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Biotechnology-Operations.pdf
Principles and Practices
BIOTECHNOLOGY OPERATIONS
S E C O N D E D I T I O N
Principles and Practices
BIOTECHNOLOGY OPERATIONS
S E C O N D E D I T I O N
John M. Centanni Michael J. Roy
CRC Press Taylor & Francis Group 6000 Broken Sound Parkway NW, Suite 300 Boca Raton, FL 33487-2742
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Library of Congress Cataloging‑in‑Publication Data
Names: Roy, Michael Joseph, author. | Centanni, John M., author. Title: Biotechnology operations : principles and practices / John M. Centanni and Michael J. Roy. Description: Second edition. | Boca Raton : CRC Press/Taylor & Francis, 2017. | Michael J. Roy’s name appears first in the previous edition. | Includes bibliographical references and index. Identifiers: LCCN 2016034538 | ISBN 9781498758796 (hardback : alk. paper) Subjects: | MESH: Biotechnology--organization & administration | Biomedical Technology--organization & administration | Program Development--methods | Total Quality Management--methods | Planning Techniques Classification: LCC TP248.2 | NLM W 82 | DDC 660.6--dc23 LC record available at https://lccn.loc.gov/2016034538
Visit the Taylor & Francis Web site at http://www.taylorandfrancis.com
and the CRC Press Web site at http://www.crcpress.com
Although our passion is in the expeditious development of biomedical products, it
is also important to recognize the selflessness of research volunteers and patients
who have the compassion and strength to participate in human clinical research
studies for the betterment of others and in the hope of advancing medicine;
without this, the development of new treatments would not be possible.
vii
Contents
Preface .....................................................................................................................xv Acknowledgments ............................................................................................. xvii Authors ................................................................................................................. xix
1. Introduction to Biotechnology Operations: Planning for Success ....... 1 Biotechnology Operations .............................................................................. 1 Marketing, Financial, and Business Considerations for Development .....4 Product Development Planning .................................................................... 7
Rationale for Product Development Planning ........................................ 7 The Targeted Product Profile .................................................................. 10 The Product Development Plan .............................................................. 16
Clinical Development Planning ......................................................... 18 Project Management Planning ........................................................... 19 Regulatory Planning ............................................................................ 20 Nonclinical Planning ........................................................................... 22 Biomanufacturing Planning ............................................................... 23 Quality Control Planning ................................................................... 24 Quality Systems and Quality Assurance Planning ........................ 26 Additional Elements of Product Planning ....................................... 26
Summary of Planning for Success .............................................................. 28
2. Project Management .................................................................................... 29 Biotechnology and Project Management.................................................... 29 Background of Project Management ........................................................... 31 Project Management Plan ............................................................................. 32
The Project Management Environment ................................................. 34 Project Objectives and Schedules ........................................................... 36 Sociotechnical Considerations ................................................................ 37 Participants in Project Management ...................................................... 37
Project Management in Biotechnology Operations .................................. 41 Establishing Project Management .......................................................... 41 The Work Breakdown Structure ............................................................. 42 Forming a Project Team and Hands-on Project Management ........... 46 Team Dynamics ......................................................................................... 46 Communication and Feedback ............................................................... 49 Project Risk Assessment and Management .......................................... 51 Metrics and Tracking Progress ............................................................... 53 Resources: Planning and Usage .............................................................. 54 Human Factors in Project Management ................................................ 55 Project Completion.................................................................................... 57
viii Contents
Project Management with Contracts and Collaborations ........................ 59 Virtual Teams ................................................................................................. 60 Tools for Effective Project Management ..................................................... 61 Summary of Project Management in Biotechnology Development ....... 64
3. Regulatory Affairs ....................................................................................... 65 The U.S. Food and Drug Administration: Law and Regulations for Biopharmaceuticals ....................................................................................... 65
Historical Basis for FDA Regulation ...................................................... 65 Regulatory Organization of the FDA ..................................................... 66 Food and Drug Law, Regulation, and Guidance .................................. 71
FDA-Regulated Products .............................................................................. 71 Biologics ...................................................................................................... 72 Drugs .......................................................................................................... 75 Medical Devices ........................................................................................ 76 Combination Products.............................................................................. 77 Other Classes of Biotechnology Products and Their Review at the FDA ....................................................................................................... 79
Products for Veterinary Use ............................................................... 79 Cosmetics, Food, Dietary Supplements, Homeopathic, or Nutritional Products ............................................................................ 79
FDA Regulatory Information and Resources: Regulatory Intelligence ..... 81 Regulatory Operations for FDA Applications ........................................... 84
Regulatory Planning and the Regulatory Environment ..................... 84 Risk Versus Benefit.................................................................................... 84 Applications Seeking FDA Investigational Use or Marketing Approval ..................................................................................................... 87 Investigational Use Applications. The Investigational New Drug Application ...................................................................................... 89
Common Technical Document ........................................................... 91 Electronic Submission of a CTD ......................................................... 92
Marketing Applications: BLA and NDA ............................................... 96 Medical Device Applications. 510(k) and PMA ......................................... 99
Special Documents, Pathways, or Exemptions ................................... 101 Generic Drugs and Biosimiliar or Follow-on Biologics ..................... 104
Other Regulatory Activities ....................................................................... 105 Public Meetings and Advisory Committees ....................................... 105 Postmarketing Requirements and Activities ...................................... 107 Advertising and Promotion ................................................................... 108
Summary of Regulatory Affair Activities in Biotechnology Operations ..................................................................................................... 109 References ..................................................................................................... 111
ixContents
4. Regulatory Compliance ............................................................................. 113 Regulatory Compliance .............................................................................. 113 Quality Systems to Meet Regulatory Compliance .................................. 113
Compliance and Quality Systems ........................................................ 113 Current Good Manufacturing Practices for Manufacture and Quality Control ....................................................................................... 114 Current Good Laboratory Practices for Nonclinical Laboratory Studies ...................................................................................................... 117 Current Good Clinical Practices for Clinical Studies ........................ 117
Compliance for Biopharmaceuticals: Other Regulations of Importance .................................................................................................... 119
Compliance for Import of Biopharmaceuticals into the United States ............................................................................................ 119 Compliance for Medical Devices .......................................................... 120
Inspection and Enforcement ...................................................................... 120 Inspections ............................................................................................... 121 Enforcement Actions .............................................................................. 123 Product Liability...................................................................................... 125
Compliance with Non-FDA Regulations: International, National, State, and Local ............................................................................................ 126
International and Foreign National Regulatory Authorities for Medical Biotechnology Products .......................................................... 126 Transporting Infectious or Otherwise Hazardous Materials........... 132 Importing, Possessing, or Transferring Controlled Biotechnology Materials ........................................................................ 134 The Public Health Security and Bioterrorism Preparedness and Response Act of 2002 .............................................................................. 136 Importation or Exportation of Biotechnology Products for the Purpose of Treatment of Diseases in Humans ................................... 137 Occupational Health and Safety ........................................................... 140 Environmental Regulations in Biotechnology .................................... 141 Genetically Modified Organisms or Molecules .................................. 142 International Diligence in Biotechnology Operations ....................... 144
Summary of Regulatory Compliance ....................................................... 146 Summary of Non-FDA Compliance ..................................................... 147
References ..................................................................................................... 148
5. Quality Systems .......................................................................................... 149 Overview of Quality in Biotechnology..................................................... 149 History: Evolution of Quality Concepts and Practices ........................... 150 Quality Systems Approach to Product Development ............................ 153 Planning a Quality System ......................................................................... 156
Defining Objectives and Ensuring Management Support................ 156 The Quality Manual ............................................................................... 156 The Quality Plan ..................................................................................... 157
x Contents
Hallmarks of Quality: Fundamental Criteria for Building Effective Quality Systems ........................................................................................... 159
Management Responsibility .................................................................. 160 Defined Quality System ......................................................................... 162 QbD and Design Control ....................................................................... 163
Quality by Design .............................................................................. 163 Design Control .................................................................................... 164 Design Change ................................................................................... 169
Contractor, Vendor, and Consultant Control ...................................... 169 Product Identification and Traceability ............................................... 171 Process Control ........................................................................................ 172 Environmental Controls ......................................................................... 173 Inspection or Testing (Quality Control) ............................................... 173 Release of Material, Service, or Product .............................................. 174 Change Control and Corrective or Preventive Actions ..................... 175 Packaging and Labeling ......................................................................... 176 Preservation, Storage, and Handling ................................................... 176 Servicing ................................................................................................... 178 Customer Concerns and Adverse Event Reports ............................... 178 Document Control .................................................................................. 178 Training .................................................................................................... 178 Auditing ................................................................................................... 179
The Quality Assurance Unit ...................................................................... 180 Manage the Quality Assurance Function............................................ 181 Control Documents and Manage the Documentation System ......... 182 Investigate Situations: Manage and Control Change ........................ 184 Ensure Qualified and Trained Staff ..................................................... 184 Perform Audits ........................................................................................ 185
Initiate a Quality System for a Biotechnology Operation ...................... 188 Unique and Effective Approaches to Quality Management ................. 190
Risk-Based Approaches to Quality Systems ....................................... 190 Total Quality Management ................................................................... 190 Six Sigma .................................................................................................. 191 Statistics in Quality Assurance ............................................................. 191 Quality Systems for Research ............................................................... 191 Resolving Quality Issues or Problems ................................................. 192
Summary of Quality Systems .................................................................... 193 References ..................................................................................................... 194
6. Biomanufacture ........................................................................................... 195 Overview of Biomanufacturing Requirements ....................................... 195 Design in Biomanufacture .......................................................................... 196 Technical Considerations for Biomanufacture ........................................ 200 Phases and Scale-up: The Biomanufacturing Life Cycle ....................... 201 Raw Material Considerations ..................................................................... 205
xiContents
Compliance and Quality in Biomanufacture: Current Good Manufacturing Practices ............................................................................. 207 Biomanufacturing Processes for Biotechnology Products ..................... 209
Expression of Recombinant Proteins and Nucleic Acids .................. 209 Production of Recombinant Molecules from Expression Vectors .................................................................................................. 209 Genes, Vectors, and Host Cells ......................................................... 210 Bacterial Cell Expression Systems ................................................... 212 Yeast Cell Expression Systems ......................................................... 213 Mammalian or Insect Cell Expression Systems ............................ 213 Production of Master Cell Banks and Working Cell Banks ......... 216
Biomanufacture of Recombinant Proteins .......................................... 217 Planning Production of a Recombinant Protein ............................ 217 Upstream Process: Production by Bacterial or Yeast Cell Fermentation ....................................................................................... 218 Upstream Process: Production by Mammalian or Insect Cell Culture ......................................................................................... 220 Upstream Process: Recovery ............................................................ 221 Downstream Process: Purification .................................................. 222
In-Process Testing and Analysis of Bulk Substance .......................... 230 Production of Bacterial Plasmid DNA ................................................. 231 Production of Live Recombinant Organisms: Bacteria and Virus ... 232 Production of Products Composed of Mammalian Somatic Cells or Tissues ........................................................................................ 234 Production of Cellular Products Derived from Pluripotent (Stem) Cells............................................................................................... 236 Production of Biological Molecules by Transgenic Animals or Plants ......................................................................................................... 238 Production of Biologically Active Lipids, Glycolipids, and Complex Carbohydrates ......................................................................... 245 Production of Biologically Active Peptides ......................................... 245 Production of Combination Products: Biopharmaceutical with a Drug or Medical Device ...................................................................... 247
FP: Formulation, Fill, Finish, and Labeling .............................................. 248 Biomanufacturing Facilities, Utilities, and Equipment .......................... 253
Facility Design Considerations ............................................................. 253 The Facility and Utilities: A Controlled Environment ...................... 254 Operation of Clean Work Areas for Biomanufacture ........................ 255 Biomanufacturing Equipment .............................................................. 257
Contract Manufacturing Options .............................................................. 257 Validation of Biomanufacturing Facilities, Utilities, Equipment, and Processes ............................................................................................... 259 Summary of Biomanufacture ..................................................................... 261 References ..................................................................................................... 262
xii Contents
7. Quality Control ........................................................................................... 263 Quality Control Overview .......................................................................... 263
Definition of Product Attributes ........................................................... 265 Analytical Methods to Measure Attributes .................................... 266 Traits of Analytical Methods ............................................................ 267
Drafting a Certificate of Analysis (Bulk Substance) .......................... 267 Selection of Analytical Methods ........................................................... 270 Development of Specifications .............................................................. 277 Entering Test Results .............................................................................. 282
Certificate of Analysis for Drug Product .................................................. 282 In-Process Testing ........................................................................................ 285 Analytical Methods ..................................................................................... 286 Additional Analytical Tools and Concepts .............................................. 295 Quality Control of Cell Banks .................................................................... 297 Samples and Sampling ................................................................................ 298 Analytical Controls and Reference Standards ........................................ 299 Test Failures, Out-of-Specification Results, and Retesting ..................... 300 Testing for Product Stability ....................................................................... 302 Quality Control Testing of Raw Materials ............................................... 308 Quality Control and the Manufacturing Environment ......................... 310 Qualification, Validation, and Verification of Analytical Methods ...... 312 Application of Statistics in Assay Performance and Validation............ 317 Summary of Quality Control ..................................................................... 318 References ..................................................................................................... 319
8. Nonclinical Studies .................................................................................... 321 Nonclinical Studies and Risk Assessment ............................................... 321 Biopharmaceutical Delivery, Pharmacokinetics, and Pharmacodynamics ..................................................................................... 323
Product Delivery to the Body ................................................................ 323 Adsorption, Distribution, Elimination, and Metabolism (ADME) .. 325
Absorption........................................................................................... 325 Distribution ......................................................................................... 325 Metabolism and Biotransformation ................................................. 327 Excretion .............................................................................................. 328
Pharmacokinetics and Pharmacodynamics ........................................ 328 Application of Pharmacokinetics and Pharmacodynamics in Biopharmaceutical Development .......................................................... 333
Safety Assessment of Biopharmaceuticals ............................................... 336 Toxicology ................................................................................................ 336 Design of a Safety Assessment Program ............................................. 337 In Vitro Screens: Surrogate Measures of Toxicity .............................. 340
xiiiContents
In Vivo Safety Testing of Biopharmaceuticals .................................... 342 Animal Model Development ............................................................ 342 Test Product Formulations, Routes of Delivery, and Dosing Designs ................................................................................................ 344
Protocols and Performance of Biopharmaceutical Safety Studies in Animals ................................................................................................ 346 Elements of a Nonclinical Study Design ............................................. 347 Nonclinical Safety Testing ..................................................................... 351 Acute Toxicity Testing ............................................................................ 351 Subchronic and Chronic Toxicity Testing ........................................... 356 Reproductive, Developmental, and Teratogenicity Toxicity Testing ....................................................................................................... 359 Carcinogenicity Testing ......................................................................... 360 Immunotoxicology .................................................................................. 361 Genetic Toxicology .................................................................................. 363 Tissue Binding or Local Tissue Tolerance ........................................... 367
Quality of Nonclinical Studies: Current Good Laboratory Practices ..... 368 Summary of Nonclinical Studies .............................................................. 369 References ..................................................................................................... 370
9. Clinical Trials .............................................................................................. 371 Introduction to Clinical Trials.................................................................... 371 Background of Clinical Research .............................................................. 373
Introduction ............................................................................................. 373 Historical Information on Clinical Trials ............................................ 374
Organization of Clinical Research ............................................................ 375 Phases of Clinical Trials ......................................................................... 375 The Science of Clinical Research .......................................................... 376 Quality in Clinical Research and Current Good Clinical Practices ...... 377 Clinical Development Planning ............................................................ 377
Infrastructure for a Clinical Trial: Individuals, Documents, and Investigational Product ............................................................................... 378
Design of Clinical Trials and the Clinical Protocol ............................ 378 Human Subjects, Patients, and Volunteers .......................................... 388 The Sponsor ............................................................................................. 388 The Principal Investigator and His or Her Study Staff ..................... 391 Institutional Review Boards, the Process of IC, and IC Form .......... 392 Investigational Product .......................................................................... 394 Collection of Clinical Data: Case Report Forms and the Patient Diary ......................................................................................................... 395 Clinical Testing Laboratories ................................................................. 396 Reporting Results of Clinical Trials: Clinical Summary Reports .... 397
xiv Contents
Clinical Trial Operations ............................................................................ 397 Activities Leading to a Clinical Trial ................................................... 398 Phase 1 Clinical Trial: First-In-Human Study ..................................... 400 Clinical Pharmacology Studies of Biopharmaceuticals in Human............................................................................................404 Phase 2 Clinical Trial: Proof-of-Concept Study .................................. 405 Phase 3 Clinical Trial: Therapeutic Confirmatory ............................. 406 Phase 4 Clinical Study and Risk Evaluation and Mitigation Strategy ..................................................................................................... 407 Clinical Trials for New Populations or Indications ........................... 408 Global Clinical Trials .............................................................................. 409
Quality Systems for Clinical Trials: Current Good Clinical Practices .................................................................................................... 409
Quality and cGCP in Clinical Trial Operations ................................. 410 Integrity of Clinical Study Data and Documents ............................... 413 Monitoring and Auditing Clinical Trials ............................................ 414 Ethical Behavior and the Well-Being of Clinical Trial Subjects ....... 415
Summary on Clinical Trials ....................................................................... 416 References ..................................................................................................... 417
Additional Readings ......................................................................................... 419
Glossary ............................................................................................................... 425
Appendix ............................................................................................................. 451
Index ..................................................................................................................... 459
xv
Preface
This book resulted from the authors’ experiences gained while working in biotechnology development at industry, government, and academia, and while teaching a graduate course titled biotechnology operations. This course is offered to graduate students in the master of science (MS) in Biotechnology Program at the University of Wisconsin-Madison (http:// www.ms-biotech.wisc.edu/). In this course, we examine the undertaking of developing biotechnology products, focusing on the scientific and manage- ment skills of biomanufacturing, clinical trials, nonclinical studies, project management, quality assurance, quality control, and regulatory affairs. The course emphasizes both operational planning for success and integration of plans and efforts in these seven functional areas. The instructors real- ized from their experience in the biotechnology industry the great need to carefully plan and fully integrate biotechnology development projects. The course is taught in that manner and this book reflects that philosophy; thus, this book is a practical guide for students and for those employed or inter- ested in biotechnology.
This book is intended to meet a need and to fill a gap. Despite the wealth of experience with operations in the biotechnology industry, there was no single comprehensive and practical, yet fundamental, guide available. Many books and most individual scientific or trade publications are highly techni- cal and focused on a specific aspect of biotechnology. They do not empha- size the themes of planning and integrating the seven operational endeavors. Biotechnology Operations: Principles and Practices is written with the objec- tive of presenting a roadmap and reference for biotechnology operations, integrating these functional areas through the processes of product plan- ning and design, and the practice of project management. It applies lessons learned in the biotechnology industry over past decades as novel products have been developed from emerging scientific discoveries. The lessons high- light development principles that could help the industry to bring to market more efficiently and quickly the safe and effective biotechnology products of the future. While focused largely on biopharmaceuticals, this book also reflects development of other biotechnology products. It is anticipated that this book will provide the reader a clear understanding of basic principles and practices, and assist in reducing risks and in resolving problems as future biotechnology discoveries are developed into products.
In preparation of the 2nd edition of this book, and at the request of the readers, we have enhanced our use of examples by including additional text boxes, diagrams, and figures. This 2nd edition now includes up-to-date meth- odologies associated with current biotechnology industry practices; incor- porated are examples of tissue engineering, stem cell technologies, and the
xvi Preface
use of alternative bioreactors. Chapter 2 now includes additional schematics to better depict abstract concepts. Chapter 3 is expanded to include current thinking of the FDA on various topics, and now includes specific infor- mation on submission formats and processes such as Common Technical Document format and electronic submissions. Chapters 2, 5, and 6 contain additional illustrations and examples of design and change control, man- agement responsibilities, quality audit process, biomanufacturing facilities, whole animal bioreactors, and stem cell manufacturing processes. Chapter 7 is updated and includes depictions of testing equipment, figures outlining new concepts, and examples of trending and trend analysis. Chapter 8 now includes specific study design examples that have been used successfully to support translation of new biopharmaceutical products into human clinical trials. Finally, Chapter 9 includes an emphasis on the practical use of Good Clinical Practice (GCP) and how it directly applies to human clinical study management.
The target audience for this book is advanced undergraduates or postgrad- uate students pursuing an advanced degree in biotechnology and individu- als working in any aspect of biotechnology product development. This book should be particularly relevant to students interested in biotechnology, bio- pharmaceutical product development, and those already working in biotech- nology. The information presented in this book can be used to expand upon one’s current experience while providing an additional level of appreciation and overview of the product development process. For those in the biotech- nology industry, this book provides guidance on planning a new develop- ment program or managing an ongoing program. Noting that irrespective of the nature of the new biomedical product, the principles and practices outlined in this book are essential for the success of developing and market- ing of a new product.
xvii
Acknowledgments
The authors sincerely hope the experiences, ideas, and examples related in Biotechnology Operations: Principles and Practices will inspire the reader to plan and implement meaningful strategies and thereby expedite the devel- opment of desperately needed new medical products. Many of the examples and suggestions in this book represent challenges and successes that we’ve experienced throughout our careers. It is our passion to contribute ways that facilitate the transition of new therapies from the discovery or research envi- ronment into the clinic.
Special thanks go
• To the many students in the Master of Science in Biotechnology Program for helpful discussions and feedback on the best way to present this wealth of information.
• To Eric Schmuck and Derek J. Hei for their assistance with develop- ing ideas and materials for this 2nd edition.
• To our many colleagues (especially Anthony [Tony] Clemento, Natalie Betz, and Edmund J. Elder Jr.) who have contributed to dis- cussions and suggestions that made writing this book possible and also for their continued support and encouragement.
Our special thanks go to Kurt Zimmerman, program director of the Master of Science in Biotechnology Program at the University of Wisconsin-Madison, for his continuing support and for providing a program in which the stu- dents are trained and encouraged to become industry leaders.
xix
Authors
John M. Centanni, MS, has a faculty associate appointment in the School of Medicine and Public Health at the University of Wisconsin-Madison, in the Master of Science in Biotechnology Program. He has firsthand experience of leading functional groups in biotechnology firms as a project manager. His strong scientific background has allowed him to serve multiple scientific R&D roles in the biotech industry contributing to the development of pre- clinical safety studies, quality control assays, and animal models. Centanni has participated in the development and implementation of quality systems to meet regulatory compliance in both the industry and academic envi- ronments. He has directly overseen the regulatory and clinical operations associated with a number of early phase, multicenter, human clinical trials. Centanni has worked in the pharmaceutical and biotechnology industry since 1987 and has more than 20 years of product development experience for drugs, biologics, and devices. He has instructed and trained basic sci- entists and clinical researchers in regulatory compliance and expectations associated with clinical product development (Good Laboratory Practice (GLP), Good Manufacturing Practice (GMP), and Good Tissue Practice (GTP), and Good Clinical Practice). Centanni is experienced in preclinical research, regulatory, quality, clinical development, and project management, and has been involved in the development and registration of pharmaceuti- cal products across a number of therapeutic categories.
Centanni is the director of the Investigational New Drug (IND)/ Investigational Device Exemption (IDE) Consultation Services, where he leads a team of consultants at the University of Wisconsin-Madison, Institutes for Clinical and Translational Research. In this role, he provides campus- wide support to clinical investigators advancing their investigational prod- uct from the research environment into the clinic. This support ranges from strategic support for the selection of viable product development candidates to characterization of products and design and implementation of human clinical trials.
Centanni is also an active participant in the Stem Cell and Regenerative Medicine Center and Cardiovascular Regeneration Focus Group at the University of Wisconsin. He serves on a number of grant review panels that include California Institute for Regenerative Medicine (CIRM), Institute for Clinical and Translational Research (ICTR) Novel Therapeutics Pilot Awards, and American Burn Association (ABA) Multicenter Clinical Trials Group.
Before joining the University of Wisconsin, Centanni directed the regula- tory, quality, and clinical efforts of a small biotechnology firm in Madison, Wisconsin. As an accomplished molecular and cellular biologist, Centanni has successfully directed multimillion dollar translational and clinical
xx Authors
research projects as principal investigator. Additional professional attributes of Centanni include a notable patent portfolio as an inventor on more than a half dozen intellectual property filings and authorship of several scientific journal articles and book chapters.
Centanni is a graduate of Hood College, Frederick, Maryland, with a mas- ter’s degree in biomedical sciences supported by a thesis and defense. Prior to graduate school, Centanni earned a BS in biology at the University of Wisconsin-Oshkosh, Wisconsin. In his free time, Centanni enjoys saltwater fishing, snorkeling, traveling, and playing racquet ball.
Michael J. Roy, PhD, is an emeritus professor at the University of Wisconsin-Madison, where he previously taught in the Master of Science in Biotechnology Program in the School of Medicine and Public Health. He has successfully developed biopharmaceutical products and medical devices for private and publically held firms, nongovernmental organizations, and the federal government for more than 27 years, serving as a consultant in bio- technology development over the past decade. Many of his efforts, including product development planning, regulatory affairs, quality systems, project management, biomanufacturing, quality control, and clinical studies, focus on early development of novel biotechnology products, notably vaccines and antimicrobial agents.
He is a graduate of the University of Wisconsin-Madison with a PhD in pathology, of Louisiana State University Medical Center, New Orleans, with an MS in tropical medicine and medical parasitology, and of the University of Wisconsin-Platteville with a BS in biology. Colonel Roy is retired from the U.S. Army Reserves, where he was involved in developing in vitro diag- nostic devices and vaccines and in establishing quality systems at the U.S. Army Medical Research Institute of Infectious Diseases, Fort Detrick. He also enjoys hiking, raising hardwood trees in southwestern Wisconsin, and has archeological interests of that region.
1
1 Introduction to Biotechnology Operations: Planning for Success
Biotechnology Operations
Biotechnology encompasses a wide variety of scientific, business, and operational endeavors in life sciences. It is applied across a broad range of specific disciplines, for example, plant, animal, medical, microbiologi- cal, biopharmaceutical, agricultural, and environmental to name just a few. Biotechnology is practiced worldwide and at many institutions: small pri- vate firms, large public corporations, nonprofit organizations, universities, and research institutes. Those practicing biotechnology include individuals with diverse skills and backgrounds: entrepreneurs, scientists, business- persons, managers, product developers, and other highly educated and motivated specialists. As seen by the inexperienced and at the macro level, biotechnology appears to be a vast three-dimensional matrix, broad and oftentimes baffling in scope and operation. However, to those experienced in biotechnology, there is organization and rationale. The keys to successfully managing a biotechnology firm are a focus on carefully crafted plans and efforts to accomplish a specific objective and to integrate operational activi- ties within the operational matrix. This is especially true for biotechnology product development operations, where the objective is to increase the value of specific products by moving them through sequential phases and to the marketplace.
Virtually every aspect of biotechnology has two common themes: (1) to extend our knowledge of life sciences and (2) to produce a product or service that someday will improve the condition of humankind. In the commercial sector of biotechnology, there is also the objective to profit financially. There are subplots to every biotechnology endeavor as well. Developing a novel biotechnology product, especially a biopharmaceutical, is an extremely technical, highly regulated, complex, expensive, and long process. Biopharmaceuticals are in development for more than 5 years, and it is not unusual for schedules to extend, from research to market approval, beyond 10 years. The risks associated with biotechnology are tremendous,
2 Biotechnology Operations
since most biopharmaceuticals fail at some point in development. Yet, there are compelling reasons to undertake biotechnology product development. The profits can be substantial, and there are needs and markets for useful products. For some individuals, it is not financial incentives, but altruistic purposes or the challenge of pursuing an ambition and lifelong dream. This provides a stream of bright individuals willing to labor at bringing biotechnology products to market. So, biotechnology development contin- ues to grow in importance, size, and scope, and is highly regarded by the public.
Biotechnology has its own jargon as evidenced by terms used in this book and other references listed in the Additional Readings, and a great amount of operational information, notably regulatory, is available at websites and some of these are identified in this book.
Words, some considered jargon, have developed to describe certain aspects of the biotechnology operational trades, and these can be confusing, even counterintuitive, to the uninitiated. The reader may refer to Glossary for definitions used in this book.
This book focuses on biotechnology product development, specifically the scientific skills commonly applied worldwide to move in an ordered man- ner a product from concept at the laboratory bench to the marketplace. It emphasizes product design, development planning, project management, and elements of each major operational function applied to the development process. These combined activities we refer to as biotechnology operations. The seven major functional areas of biotechnology operations, identified in Box 1.1, are further described in individual chapters of this book. Additional functional areas, such as business development and finance, also directly impact biotechnology operations, and these are recognized because they are keys to success.
The focal point of a biotechnology operation is the product, and at the heart of product development are the user and intended use. The opera- tional team of professionals works together to add value, bring the product to market, and ultimately to the end user. Hence, a key to building a suc- cessful biotechnology operation is to maintain this focus on product and its intended use and the user. In biopharmaceutical development, the intended use is the product indication, a word that will be used repeatedly in this book. In medicine, an indication is defined as the reason a product is used to diag- nose, prevent, or treat a specific disease or condition. An indication also identifies, to a great extent, the intended user of a biotechnology product. This is especially true for biopharmaceuticals. In addition to having an indi- cation, biotechnology development is also based on an understanding of the molecular or cellular nature of a product and on the product’s safety, strength, purity, and potency.
Seven major areas of biotechnology operations are listed in Box 1.1. In addition, there is a need to integrate and coordinate each of these skills in
3Introduction to Biotechnology Operations: Planning for Success
an effective and timely manner, focusing on making operational headway, and moving the product toward market approval. Given the complex- ity of biotechnology operations, the need for careful planning is intui- tive. Planning is an activity that results in a written strategy. Together they establish the objectives and also map out a means of integrating the skills and events that lead to success. Indeed, a product development plan (PDP) allows a development program to be successful. Without a carefully crafted and functionally integrated plan, biotechnology operations typi- cally fail.
To begin our journey through biotechnology operations, this chapter intro- duces the planning process for product development. Think of the plan as a skeleton and each element of the plan a bone that gives structure to the over- all operation. Chapters 3 through 9 describe individual functional areas that execute or flesh out the plan and provide operational activities (Box 1.1). The functional areas do the heavy lifting, so to speak, in an operation, and six of them are considered the muscles of an operation. Chapter 2 describes proj- ect management, the operational function that serves as the neural system to a biotechnology operation, coordinating movement of operational elements according to the plan.
BOX 1.1 SEVEN MAJOR AREAS OF BIOTECHNOLOGY OPERATIONS
Operational Area Definition Chapters
Project Management
Lead the planning, organization, and management of the overall development project and associated resources.
2
Regulatory Affairs Advise on regulatory aspects and climate for product development, coordinate activities with regulatory agencies, and ensure regulatory compliance.
3 and 4
Quality Assurance Provide support to ensure that all efforts and the product are of highest quality through quality management, audits, documentation, and other quality functions.
5
Biomanufacture Produce the highest quality product through phased manufacturing development and final commercial production.
6
Quality Control Ensure quality product through laboratory testing. 7 Nonclinical Develop pharmacology and toxicology laboratory and
animal studies and reports to ensure the safe and proper use of the product.
8
Clinical Determine the safety and effectiveness of the product when used to treat human subjects.
9
4 Biotechnology Operations
Marketing, Financial, and Business Considerations for Development
Biotechnology products in general and biopharmaceutical products in particu- lar, with their stringent regulatory guidelines and strict need for a high benefit- to-risk ratio, are particularly expensive to develop. So expensive, in fact, that investment capital and public funding often provide insufficient resources to support the complete product development cycle. Today, the total development cycle costs for a biopharmaceutical can reach or exceed one billion U.S. dollars. Although somewhat less expensive to develop, other types of biotechnology products, such as those in the agricultural or environmental sectors, might still cost in excess of one-half billion U.S. dollars. Indeed, some biotechnology firms never even enter the development arena because of high cost and inability to raise capital to meet projected expenses.
Biotechnology firms rely on both public or private financing and partner- ships with traditional pharmaceutical firms to provide capital needed to reach their development goals. Of course, money always comes with tradeoffs and an investor or partner may hold definite ideas and opinions regarding how the biotechnology firm should develop the product. In the end, some biotech- nology firms are acquired by the partner during the development cycle and well before a product comes to market. Raising capital is not a subject of this book, but one must consider expenses and budgets during development plan- ning and again at every milestone.
Once a project has begun, financing and budgets continue to have an impact on decisions made both in planning and in executing a project. Indeed, they are often the primary consideration regarding a decision on whether or not to con- tinue a product development project. There are tradeoffs for the biotechnology firm. Development of a specific product may necessitate the sacrifice of other endeavors, such as pursuing promising lines of research. The philosophy of a company may have to be changed to pursue development, with hiring of devel- opment staff offset by the loss of research scientists. Facilities inevitably must be added or modified to suit development efforts and, as noted later, this can be extremely resource intensive. Once these resources have been committed, there is no turning back without incurring significant loss of time and money. No wonder biotechnology executives typically refer to the decision to embark on development as betting the farm or entering the valley of death.
Given all these warnings, what is the prudent way for a biotechnology firm to enter product development? The answer is simple: one step at a time, with a market analysis, a carefully defined product and indication and a well- considered PDP.
Earlier in this chapter, a metaphor—skeletal, muscular, and neural systems—was used to introduce the concepts of biotechnology development plans, operational elements, and integration by project management, respec- tively. This metaphor is further explained and developed in Box 1.2. Further
5Introduction to Biotechnology Operations: Planning for Success
BOX 1.2 A BIOLOGICAL METAPHOR FOR PLANNING BIOTECHNOLOGY OPERATIONS
A metaphor to planning a biotechnology operation is taken from the organized development of the mammalian neural, muscular, and skel- etal systems. This metaphor seems relevant, given the biological nature of our professional work.
An organism is composed of individual organs and tissues, and as they develop and function they work together in harmony and allow the animal to function and survive. The skeletal, muscular, and neu- ral tissues provide functions, respectively of support, movement, and perception of or reaction to stimuli. Each tissue arises in an exact man- ner, shaped according to a plan programmed in the genetic code. The developing skeletal system is composed of bones, logically arranged and able to provide the outline of a unique organism.
To begin the metaphor a biotechnology operation functions, or should function, in the manner of a healthy organism, with the indi- vidual organs and tissues coordinated and working in harmony. An operational plan, the PDP, is the skeleton of that operation. It provides shape to the overall project. Although the individual bones of an ani- mal form a strong framework, they must move in an integrated and coordinated manner. For this to happen in an organism, muscle is the organ system holding bones in a particular manner, yet moving them so they are useful structural elements. In a like manner, the PDP is moti- vated by the various functional areas of biotechnology development— clinical, manufacture, nonclinical, quality assurance, quality control, and regulatory affairs—that implement the plan, providing outcomes, yet allowing movement and flexibility of operation.
Returning to the organism in this metaphor, a neural system signals the bones and muscles to work together in a timely and effective man- ner. The neural system makes the bones and muscles useful to the body by coordinating endeavors, both as affecters and effectors. Thus the bones and muscles achieve specific objectives. In biotechnology prod- uct development, the neural system is represented by project manage- ment, a key function that ensures the various elements work together in harmony, sensing the operating environment, and reacting accordingly.
Perhaps the most important part of this metaphor is to imagine an organism deficient in one of these three elements: skeleton, muscle, or neural. Indeed, there are diseases for which this is the case. The result is illness and eventual death. Here the metaphor carries to the biotech- nology operation, because without each of the functional elements, a PDP to bring them all together, and a system to integrate and manage
(Continued)
6 Biotechnology Operations
to this metaphor, consider that these three organ systems would not function properly in any animal without support provided by other organs: the heart, liver, and kidneys for example. So, it is in biotechnology development, where support from research, marketing, business development, management, and other areas is essential to the life of the operation. An important element of any good development program is the need to consider the advice, exper- tise, and support of individuals with skills that do not apply directly to the technical agenda of an operation but have great impact nonetheless. We have mentioned financing and now consider input from the business and market- ing professionals. While these professionals might seem at times to perceive situations and issues differently from operational staff, their skills and judg- ment are indeed important throughout the development process and their input is especially critical to success at the planning stage.
They sit on the product development team, advising and planning from business, finance, and marketing standpoints. The product development team members, often referred to as a project team, should pose to them criti- cal questions from the outset of the planning process. Is there a market for the biotechnology product as it is currently designed and, if so, is the market large and extensive enough to generate a profit and is it open to introduc- tion of this new or improved product? Or should another product design be chosen? Is there competition and, if so, is it prohibitive to the intended market? Will it be necessary for the firm to develop or further develop the market and, if so, how long might this take? Are there advantages and dis- advantages to the market due to regulatory pressure, not just the U.S. Food and Drug Administration (FDA) but any regulatory agencies? How might we price this product in the current market? Here, the business and finance elements of the entity become especially important, and a well-considered business plan provides valuable information for development planning pur- poses. At this time, it may be difficult to exactly identify business advan- tages of a particular product, but certain elements can be considered. At a strategic level, several questions are posed. In theory, is money available to develop products in this market sector and, if so, is there precedent? What are the potential sources of funding and are partnerships with larger firms possible? Alternatively, might competitors try to impede our progress in an effort to retain their market share? At this early stage of predevelopment, it is
BOX 1.2 (Continued) A BIOLOGICAL METAPHOR FOR PLANNING BIOTECHNOLOGY OPERATIONS
their operation, the product development program does not function properly and eventually does not survive. Alternatively, if skeletal, muscle, and neural systems are healthy and carry their weight, then the organism, and by analogy the biotechnology operation, prospers.
7Introduction to Biotechnology Operations: Planning for Success
be impossible for even the most seasoned business experts to have all the answers; indeed, meetings at this time may generate more questions than answers. Yet, such discussions are critical to the planning process.
Product Development Planning
Rationale for Product Development Planning
Biotechnology operations have borrowed many concepts and operating prin- ciples from the drug industry. Indeed, both drug and biopharmaceutical devel- opment projects often focus on preventive and therapeutic biopharmaceuticals intended for use in humans. Drug development, a phased or step-wise process well established by the drug industry and regulatory authorities, is commonly applied to biopharmaceutical development. Figure 1.1 outlines functional ele- ments involved in a phased scheme for a biopharmaceutical or drug develop- ment project and the approximate schedule for each. It represents a project beginning with discovery or engineering of a novel biologic and ending with a product entering the marketplace. It is an idealized and simplified cartoon, but, in reality, the process is much more complex than depicted and may be abbre- viate or lengthened. Nonetheless, such schemes are developed and applied as planning and operational management tools, thus providing visual representa- tion of the major events, processes and milestones, and facilitating communica- tion and understanding by project teams and upper management.
In Figure 1.1 the stages are defined as: Stage I—Research, the molecular or cellular entity is discovered, isolated, engineered, and characterized. This ini- tial stage begins with the discovery of a novel product. The research labora- tory assists in the characterization and perhaps reengineering or refinement of the product to meet design criteria. In Stage II—Development, a host of activi- ties are described throughout this book that led to marketing approval in year 2011. The design of a targeted product profile (TPP) and product development plan (PDP) are prepared under the guidance of formal project management (Figure 1.2), which initiates the development pathway. Project management and quality assurance are active throughout the development life cycle, whereas formulation, analytical, pharmacology, biomanufacture, toxicology, clinical, and regulatory activities are staggered within the development life cycle.
Discovery research is the foundation on which most biotechnology products are based. Some refer to it as Phase 0 in the development process because dis- covery must happen before Phase 1 or early development may begin. It is sci- entists in laboratories who discover, sometimes serendipitously and, in other instances by plan, the information on which biotechnology product develop- ment is based. Gene cloning, propagation of stem cells, engineering a drought- resistant trait into plants, and a monoclonal antibody directed against a tumor protein are but a few of the thousands of the proven discoveries that have been
8 Biotechnology Operations
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9Introduction to Biotechnology Operations: Planning for Success
the foundations for important and useful products. In most instances, these dis- coveries are patented, which legally ensures that the discoverer, or the affiliated institution, receives proper credit for any worthwhile product that might be developed from their invention. Patents ensure that the patent holder, the dis- coverer, reaps a monetary reward if the technology is licensed or the product is marketed. As product development requires substantial resources, typically tens to hundreds of millions in U.S. dollars, only a few biologic discoveries are taken through the development life cycle to become a product. Most biotech- nology products are therefore based on a unique discovery that either has a pat- ent or is patentable. However, few discoveries or inventions in biotechnology are themselves marketable products; they must first be developed.
What then can we do with an exciting, patented biological innovation that holds potential value to humankind and in the marketplace? What must we do to develop that product? First, we carefully and exactly define the prod- uct. Although this may sound simple, reaching a definition is no easy task and, unfortunately, many discoveries enter product development without an exact definition of what the intended product is or what is expected of it. In such cases, the PDP, and hence product development operations are unfo- cused, wasteful, and far too often unsuccessful. A biotechnology product, and product develop pathway, must be planned, it cannot simply evolve.
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FIGURE 1.2 Planning backward: Targeted product development. A product development plan (PDP) is written in a reverse manner (arrow on the left) from what one might imagine. Working from the targeted product profile (TPP) (lower left), label claims are the first step in the planning process. Then, working backward, or from lower left to upper left in the figure, the planner develops a PDP, planning each part of the project, listed in the middle of the figure. The plan is then implemented (shown on the right side, reading top to bottom) in a forward manner, through each stage of development (provided in Figure 1.1) to produce a final product with approved label claims. (Courtesy of Anthony Clemento, 2008.)
10 Biotechnology Operations
There is not enough time and money to take any product development route other than that of a well-considered PDP.
How does planning work? Let us consider another metaphor to explore tar- geted product planning. Suppose we had the power to design and then develop a new species of mammal. Our first step would be to define, in various ways, the purpose of this desired mammal. Specifically, we begin by asking how it would meet needs of the user. Let’s say that we wish our new animal to pick fruit from trees in orchards. Then our design, based on this user need, would be bipedal and tall, with long arms and dexterous fingers. It should have the strength to stand for hours, and muscles that allow it both to stretch and to rapidly pluck fruits from a tree. It should have intellect: an ability to differentiate oranges from apples and to discern ripe oranges from immature fruit and for a brain to signal the muscles and skeleton to pick that fruit. Hence, we have defined a creature intended to pick fruits from trees. In planning the bone structure of this animal, would we borrow the design of a dog or a meadow vole? Certainly not, instead we would shape our plan, the bones if you will, around a bipedal creature, per- haps a primate. But we would design especially long bones in the arms and legs, a vertical or erect and strong vertebral column and lengthy arm bones with many digits. Indeed, we might include bones for four arms, one to grasp the tree branch, one to pluck the fruit, one to catch the fruit and yet another to transfer it to a basket. Hence, our bone structure forms a framework for the intended creature.
The plan for the muscular system would make these bones useful to the creature’s intended purpose. Would we link these bones with muscles that allow our creature to run fast, like a cheetah? Probably not; we would instead give it muscles that allow those bones to stand all day, to continually grasp limbs, and to carefully pick, grasp, and transfer fruits.
We would plan a neural system that coordinates these musculoskeletal func- tions, one that achieves the primary objective of picking fruits, but also allows the grasp to rapidly change as the creature reaches for a new branch, to pluck the fruit with one arm even as the grasp is changed with another arm and the fruit is transferred with a third arm, and to discriminate a ripe from unripe fruit, immediately before it is plucked.
Biotechnology product development, to be successful, follows a specific plan- ning process in much the same way as we designed the fruit-picking creature. However, in biotechnology, the long process of discovery research and economic realities do not allow us the luxury of millennia, the time needed for evolutionary processes in nature. In biotechnology operations, products are developed rapidly and efficiently from innovations. We begin with a clear understanding of what the product is and what it must do and how the product will be used. This is writ- ten in a targeted product profile (TPP).
The Targeted Product Profile
Product development planning is said to happen in a backward manner because the process begins with generation of a TPP, which in fact is a draft
11Introduction to Biotechnology Operations: Planning for Success
of the product label with product claims. The planning process is outlined in Figure 1.2. In biopharmaceutical development, the TPP has in the past been referred to as draft or concept product labeling. The FDA strongly encour- ages sponsors, defined as the entity responsible for developing a biopharma- ceutical, to prepare and use a TPP to support communication with regulatory authorities. More recently, investors in biotechnology have asked petitioners to provide them with a TPP along with the business and technical plans.
Simply stated, the TPP is a clear and detailed description of what a product should be, how it will appear, and, most importantly, what it must do. The term TPP says it all, establishing a target of or focus on the product and profil- ing or summarizing characteristics of the product. Box 1.3 lists the elements of a typical TPP for a biopharmaceutical product. Generation of a TPP is truly the first step in managed product development. A TPP is written by a lead author, someone familiar with both the product and with various aspects of biopharmaceutical development. Teamwork is essential for a successful start, and this means appointing a product development team and project man- ager (PM) and holding team meetings at regular intervals (Chapter 2). Team members review and recommend changes to the draft TPP. Members of the product development team, each representing a functional area, investors, and upper management are involved in this process with a final TPP as the
BOX 1.3 ELEMENTS OF A TPP
• Trade name and chemical name: A draft trade name or interim designation, such as a compound number, is developed for the product.
• Warnings: Warnings might be added for this product, based on its class of product and previous experience. Messages to the prescriber or user for this product are noted.
• Description: The product’s nature and classification are devel- oped. The formulation in the final container, with excipients, is included.
• Clinical pharmacology: The mechanism of action, pharmaco- dynamics, and pharmacokinetics that are known to date or should be explored are presented. Drug interactions are given.
• Clinical studies: Identified are pivotal clinical studies to include patient populations, endpoints, and outcomes.
• Indications and usage: The expected indication is given as is the intended patient or user population.
• Contraindications: Situations in which the product should not be used (e.g., pregnancy or congestive heart failure) are stated.
(Continued)
12 Biotechnology Operations
team’s first goal. These early interactions set the context and tone for later discussions (team members bond, agree, or disagree), and leadership skills become evident. The need for additional professional skills is recognized, and so teams are filled out to meet management and technical objectives and, early on, thought is given to the nature, scope, and possible general sched- ule of the development project. Thus, preparation of a TPP provides a critical guidance document and solidifies the new product development team.
Of all elements attributed to a TPP, the first, most important and, often times, the most contentious, is establishing the indication, also referred to as the label claim or simply the claim (Figure 1.2). Note similarities between information in a TPP and in an actual product label. Any differences are largely because of the fact that a TPP is an expectation in nature and tone,
BOX 1.3 (Continued) ELEMENTS OF A TPP
• Warnings and precautions: The users or physicians become aware of events or reactions to the product and the more serious or common of these are given in a warning. They are written for the physician and user, or composed as information for patients, specifically written for the user. Instructions for special situa- tions are also placed here, and specific items are highlighted as paragraph headings. Recommendations may be given to stop using a product, for example, if a disease progresses or if certain symptoms are noted. Drug interactions, use in nursing mothers or in pregnancy, pediatric and geriatric use, or use in other spe- cial populations are generally included in this section.
• Adverse reactions: Types of adverse reactions that might be acceptable, given the intended use and user profile, are identified.
• Overdosage: This describes reactions or remedies, should a patient take more than the prescribed amount.
• Dosage and administration: This provides a statement on how the product will be provided as dosage form to a patient in final format. The intended final container or delivery system is described.
• How supplied: This describes the packaging format that will be produced and marketed.
• References: A few key scientific publications regarding the product and indication are included.
• Patient information: This expands special instructions that might be required for proper handling, storage, or use by the patient.
13Introduction to Biotechnology Operations: Planning for Success
whereas the actual labeling of a marketed product is FDA approved, the real thing, based on data. For a biopharmaceutical, an indication might be defined as a treatment or prevention for a disease or condition that has a specific cause and symptom. Let’s begin by demonstrating poorly worded indications. One, for a peptide therapeutic product, is to lower blood pressure in benign hypertension. Another vague example of a vaccine composed of a recombinant protein is to prevent malaria. It is critical that an indication be as specific as possible and that it be matched with a proper biopharmaceutical. Here the biotechnology development team must set aside bias and grand or long-range projections of safety or efficacy (e.g., this biopharmaceutical is so great it will cure every type of cancer and never result in a side effect) and instead focus on the research data. The key is to settle on an indication for which the product would likely reach the market in short order. Having said this and returning to our examples, the indication for the peptide might be more rationally stated as lowering blood pressure in individuals with uncomplicated benign hypertension and between the ages of 60 and 82 years, where blood pres- sure has remained elevated above 140/90 mm Hg despite the use of other common drugs and where there are no symptoms of congestive heart failure. The example of the malaria vaccine might better be stated as indicated for use in infants and children between the ages of 6 months and 5 years for prevention of serious disease and death from falciparum malaria in endemic regions of Africa, Asia, and South America. The TPP also makes claims for safety parameters, and so it is impor- tant to consider the safety profile that would be acceptable for the product and include this in the final profile.
A claim is a contention or assertion that something will be achieved. Biotechnology products are all accompanied by printed labeling, and it is here that claims on product safety and efficacy are made. Claims are also reflected in advertisements and labeling, not just for medicines but for all types of products. Consumers read these claims (or should read them) when making purchases and before use. A TPP is draft labeling with predicted claims, and as such the TPP is used to guide the planning and actual development of all biotechnology products, not only biopharmaceuticals. Some examples follow. First, a recombinant bacterium is indicated for remediation of crude oil spills in salt water where the air and water temperatures are more than 40°F, wave action is not severe, and the spill is contained to a geographic region of area less than 100 km2. Or a genetically engineered soybean has the indication to increase yields by more than 20% in comparison with other varieties when grown in zones 3 or 4, and where rainfall averages between 12 and 20 inches per year, there is no irrigation, and the soil is slightly acidic or neutral.
Returning to biopharmaceutical development, a TPP discusses the other objectives, and these are listed in Box 1.3; note the various claims identi- fied on the TPP, such as indication and safety profile. It is worth noting that results from research or early development completed to date, market driv- ers, and perhaps the experiences and ingenuity of individual team members are the basis for deciding on each claim. The target population is identified
14 Biotechnology Operations
as part of the claim. For example, a product is to be used only in adults of more than 50 years of age and in otherwise good health. Often, experts, such as physicians highly regarded in a specialty area, are consulted before the team reaches a consensus on a target indication and population.
The next step in developing a TPP for a biopharmaceutical is to determine the target dosage and route of administration best suited to the product and the population identified in the indication. The peptide example might be best administered by the parental route, such as subcutaneous, because peptides might not be amenable to the hostile environment of the gut. The vaccine might be preferred as an intramuscular injection from a disposable syringe. In each example, the dosage may need to be 1 mL or less. Dosage forms and strength refer to the formulation of the product and how it might be presented in a marketed or final container, such as a vial or syringe. The peptide might only be stable in a buffer of low pH. In the case of the malaria vaccine, the product profile includes a preservative, so it can be used in the tropics. This, in turn, necessitates a formulation that allows it to be shipped and stored with breaks in refrigeration. For the microbes used to remediate an oil spill of more than 100 square miles, it might be necessary to consider a product that could be disbursed from large mechanical sprayers on aircraft. The seeds of a drought-resistant soybean plant might need to be planted further apart from each other, as compared to current soybeans. This type of information is agreed by the product development team and included in the TPP.
The TPP also considers dosage form and strength. For medical products, there will be limitations as to the mass of product, peptide, or recombinant protein that can be held in 1 mL of solution. The optimal formulation, one that is simplest and least expensive, may not be feasible, and the product team could decide a special formulation was necessary, for example instructions to keep a protein product from aggregating and thus preventing loss of activity. In these examples, the TPP prompts the team to consider manufacture, formu- lation, and quality control issues and highlights the need for additional steps in development and, perhaps most importantly, identifies both complexities and costs of actually developing the individual product.
Contraindication refers to those times when the team recommends that the product simply should not be used, when it might be unsafe, for exam- ple. Basic contraindications should be considered, and here again it might be helpful to consult a physician with experience of treating the disease in the indicated patient population. Warnings and precautions, on the other hand, are more difficult to define at this very early stage of development and in the absence of any safety information on the product. However, warnings and precautions from products similar in nature, treatment indication, and target patient populations may be instructive as to what may or may not be accept- able for this product. The contraindications, warnings, and precautions often narrow the indication, and this is important information to consider in prod- uct development. For the peptide used to treat hypertension, it might be con- traindicated to use the drug in patients with certain other cardiovascular
15Introduction to Biotechnology Operations: Planning for Success
diseases as known from experience in cardiovascular medicine and phar- macology of similar products. The malaria vaccine might be contraindicated when the patient was already infected with the parasite. The remedial bacte- rium might be contraindicated when other petrochemicals, such as gasoline or diesel fuel, were present. The drought-resistant soybean plant might not be used within a kilometer of other soybean fields. The main point is that a knowledgeable product development team confronts these issues during the process of developing a TPP and well before development begins. This facili- tates early planning to resolve, if possible, each potential problem or issue.
Identification of undesirable and product-related adverse reactions, risk of overdose, and interactions of the biopharmaceutical with other drugs are, to a great extent, items that must be addressed during clinical studies (Chapter 8). However, it is possible during TPP preparation to consider the limits of adverse events or precautions the team might allow for a product. With the peptide antihypertensive, serious illness or death resulting from therapeu- tic doses, no matter how infrequent, might pass the acceptable threshold for adverse events. The malaria vaccine for children should not cause local reac- tions and discomfort that are of great concern to the child or a parent. In the case of the remedial bacteria or the soybean plant, one might respectively establish limits regarding how extensively the bacteria could multiply in the environment in the absence of crude oil or how far the soybean could spread to neighboring fields. The process does identify, to the development team, certain limits that might be applied to the development program.
Use in special populations further defines when and how one might use the product; it extends the indication by considering individuals of certain age groups, such as adolescents or the elderly, or of physiologic status, such as nursing mothers or pregnant women. Drug abuse and dependence is typi- cally not an issue with biopharmaceuticals but can be important with certain types of drugs.
Adding a description of the product to the TPP would seem, on the surface, to be a simple task but product development teams often find it to be a chal- lenge, especially in regard to describing all intended physical, chemical, and biological characteristics. This is discussed in greater length in Chapter 7. Any biotechnology product that will be used in man, animals, or the envi- ronment will need to be very well characterized in all respects but, at this juncture, product characteristics are unknown. Preparation of the TPP forces the team to consider what types or classes of characteristics must be exam- ined for in the product during development. For any of the examples we have used, biological characteristics should include potential toxicity or half- life and description of any living cells. Chemical characteristics may include the molecular nature of a product and also any impurities or contaminants. Physical characteristics are size or shape or the ability to withstand adverse conditions of an acceptable molecule or organism.
Clinical pharmacology may also be unknown at this early stage of development. The term takes into consideration the distribution of the
16 Biotechnology Operations
biopharmaceutical in the body, the kinetics of distribution from the time of dosing through the time of clearance, and the dynamic properties while it is in tissues (Chapter 8). But there should be some information, from laboratory or animal studies, on which the team can develop desirable parameters or acceptable limits. The antihypertensive peptide should clear itself from the body before another dose being given, and the malaria vaccine should not remain in a subcutaneous tissue indefinitely. The remedial bacterium should be cleared from the environment and not be present long after the crude oil has been eliminated, and there may be limits on how long the soybeans can self-reproduce under field conditions.
Nonclinical toxicology testing (Chapter 8) is very important because the tox- icity profile of a product in animals is often a predictor of toxicity in humans. Clinical studies follow nonclinical toxicology and, as discussed in Chapter 9, they are designed with the results of nonclinical studies in mind. The nonclin- ical and clinical toxicology profiles are certainly unknown at this early stage, but the development team does have the opportunity and obligation to set lim- its for safety and efficacy parameters, even if they are general, for each product in its TPP. Here the history and labels of competitive products or good medical judgment come into play, along with scientific and medical experience. Would one consider developing and marketing the example peptide antihypertensive if it consistently caused rats to die of hypotensive shock at the intended human dose? Might the malaria vaccine be advanced to clinical trials in children and infants or would it even be marketable if it caused severe local reactions in both rabbits and nonhuman primates? For the remedial bacterium, would one wish to put it into a field trial in an ocean lagoon if the bacterium itself led to the death of fish or invertebrates in an aquarium setting? Would it be prudent to place the drought-tolerant soybeans into field trials if they were found, in greenhouse studies, to spread the resistance gene to other species of legumes? Limits for nonclinical and clinical toxicity can and must be established.
The preparation of a TPP not only motivates the team to discuss potential issues early on, even before a PDP is written, it also forces members of the team to consider limits to the technology well before a major investment is made in developing the product. Hence, the value of a TPP goes far beyond internal use by the sponsor. Once completed, a TPP often becomes the technological extension of a business plan and is invaluable for business development and helps to raise working capital from investors. It is a foundation for communi- cating the technology, potential benefits, and possible risks to the public and to regulatory agencies, simply because it clearly demonstrates that the sponsor has considered implications, good and bad and known and unknown, of the technology.
The Product Development Plan
A PDP, also called a product development strategy, extends the TPP, pro- viding a roadmap to reach the stated goals. Further, it defines how issues
17Introduction to Biotechnology Operations: Planning for Success
and unknowns identified in the TPP will be addressed and thus become the basis for scheduling activities and budgeting resources over that schedule. The PDP is shared by everyone on the product development team as a common narrative understanding of what has to be done and how it will be accomplished. Project managers use the PDP for exact task integration, scheduling, and tracking. The PDP may be shared, in confi- dence, with potential investors or partners and regulatory agencies so as to demonstrate that the sponsor has the will and a valid strategy to take the product to market and thus make a return on investment.
Consider again that planning is a process and a plan is a written document. Typically, the planning process is managed by a PM, although the plan itself is written by individual members of the project team. There is no established order to preparing the individual chapters, discussed later, but most organi- zations find it is quite helpful to develop a draft or at least an outline of three sections, clinical, regulatory, and project management, before beginning the others. The draft project management plan establishes the project team and provides guidelines, early on, as to the planning process itself. The planning process requires much discussion, and this comes at team meetings or tele- conferences. So having a project management approach established early on facilitates communication and preparation of each section of the PDP. Since the planning process works backward in a development scheme (Figure 1.2) and since a Phase 3 clinical study is critical to achieving market approval of a biopharmaceutical, it is very helpful to draft a clinical plan before other sec- tions are prepared. Also, having regulatory input upfront provides important guidelines, especially for biopharmaceutical products.
Once the project management, regulatory, and clinical plans are in outline or early draft format, a designate from each functional area drafts the appro- priate section of the plan. The planners each apply the method of working backward to prepare at least a solid outline. They can then expand the plan, adding detail while working forward through early, mid, and late phases. Integration of the elements is important, and the process is facilitated by effective project management, frequent meetings, and cooperation on the part of every team member.
The contents of any one PDP are difficult to predict because every product is unique. Yet experience provides suggestions to ensure any PDP is under- standable. The PDP has a clearly stated purpose and objective, focused on the product as described in the TPP. It considers each one of the seven func- tional areas. It identifies significant risks and foreseeable difficulties and makes arrangements in the plan to address them. The risk-to-benefit discus- sion is real and not overly optimistic. The plan is comprehensive by provid- ing precise technical descriptions. Important steps or stages are not avoided or omitted, and adequate resources are committed for each functional area. Finally, the PDP offers a realistic schedule.
Provided next are elements found in most PDPs, written as statements that should, along with the writings in this book and other publications, stimulate
18 Biotechnology Operations
thought and focus on planning your product. The order of sequence includes the following:
1. Clinical 2. Project Management 3. Regulatory 4. Nonclinical 5. Manufacturing 6. Quality Control 7. Quality Systems and Quality Assurance
Additional planning considerations are discussed in the various chapters in this book.
Clinical Development Planning
Overall Clinical Development Planning
• Prepares a broad overview plan for clinical development and con- firms intended label claims and intended medical outcomes follow- ing treatment.
• Lists, by phase of development, all clinical trials that are anticipated. • Defines the most challenging aspects of clinical development for
this product and indication. • Identifies safety, tolerability, or toxicity factors that are of concern for
the investigational product. • Describes what has been learned from previous clinical studies with
products of this type and for similar indications. • Defines pharmacoeconomic and marketing issues related to the
product should it be approved for the stated indication and patient population.
• Identifies key decision points in the clinical development scheme.
Clinical Development Planning by Phase
• Describes the Phase 3 clinical trial design and elements of a Phase 3 concept protocol to include hypothesis, objectives, outcomes, end- points, and measurements.
• States the regulatory guidance or precedence needed to develop the Phase 3 clinical approach.
• Identifies a study or studies to be performed in Phase 2 and given the intended design and outcomes of Phase 3.
19Introduction to Biotechnology Operations: Planning for Success
• Describes how Phase 2 studies are to be temporally staggered? • Provides a brief concept design for each Phase 2 study, indicating
the outcomes, endpoints, measurements, and number and nature of subjects tested.
• Lists Phase 1 studies to be completed before beginning Phase 2. • Provides a brief concept design for each Phase 1 study, indicating the
objectives, endpoints, measurements, number and nature of subjects tested, and most likely outcomes.
A General Clinical Development Plan at Each Phase or Study
• Identifies criteria for patients or subjects enrolled. • Describes unique designs, such as adaptive or crossover, contem-
plated for any study. • Lists the resources required to perform the study and describes
requirements for clinical study centers or sites. • Identifies multicenter studies to be performed in late stage and includes
foreign clinical trials considered at early- and mid-stage studies. • Describes logistical considerations and management of multicenter
trials. • Names the most likely opportunities for clinical study sites and iden-
tifies studies to be outsourced and the sponsor’s roles and responsi- bilities to be delegated to outside consultants or contractors.
• Identifies analytical or medical tools or procedures that will be developed to measure clinical endpoints and describes how and when each is to be developed.
• Names the internal staff requirements at each stage of development. • Provides the general statistical approach and lists requirements for
data handling, statistical analysis, and report preparation. • Outlines the clinical study’s monitoring and auditing plans and
describes how clinical quality will be ensured for each study. • Identifies clinical trial material (product) requirements at each phase
of development based on the concept protocols and number of sub- jects and doses per protocol.
Project Management Planning
• Defines the overall objectives and scope of the project. • Identifies the overall policy for project management applied to the
development project. • Defines requirements for support from upper management.
20 Biotechnology Operations
• Performs a general work breakdown structure of the major areas of effort known to date, provides an estimated schedule for the project, and illustrates this in a chart (e.g., Gantt or PERT).
• Defines roles, responsibilities, and authority of the PM. • Defines team composition in all areas over the course of the project. • Defines team communication methods along with anticipated fre-
quency of each type of communication and identifies special com- munication requirements because of distances or international participation on the project team.
• Identifies methods to involve contractors, consultants, or vendors with the team.
• Identifies responsibilities of the team and of the PM for risk assess- ment and risk management.
• Identifies methods to be used by the team to solve problems. • Identifies methods for the team’s decision-making process. • Defines responsibilities and processes for risk analysis, mitigation,
and management. • Defines tracking and metrics procedures to be applied and indicate
their frequency of use. • Discusses budget and human resource responsibilities of the PM. • Develops a project schedule. • Provides, in general, the objectives and schedule for project closure.
Regulatory Planning
Planning the regulatory approach and operational elements requires sev- eral skills. First, regulatory intelligence is conducted. Next, a draft plan is formulated. Finally, all other sections of the PDP are reviewed to ensure that each is consistent and compliant with the current regulatory environment.
Regulatory Intelligence
• Describes what is known about this product or a similar product (predicate) from the regulatory literature.
• Describes how predicate products were designed, mentions their origin and history, and identifies the methods and technologies used in their discovery and development.
• Identifies potential regulatory routes of approval, both U.S. and for- eign, used to develop similar or predicate products.
• Lists the technical (e.g., manufacture, control, nonclinical, or clinical) and regulatory successes and failures for each predicate product and explains why each succeeded or failed to gain market approval.
21Introduction to Biotechnology Operations: Planning for Success
• Discusses the impact this technical and regulatory intelligence might have on the intended PDP.
• Discusses how the national political environment may or may not be supportive of this product and lists state, local, or cultural practices or laws that might be unfavorable to such a product or indication.
• Discusses how the public might perceive the relative benefits and risks of this product during the investigational phases. Will public opinion matter, one way or another, to regulatory agencies in regard to this product and indication? It also mentions outstanding safety issues that might concern regulatory agencies and any regulatory precedent for handling these issues?
• Identifies how FDA regulations might be expected to change before approval in any given market.
Regulatory Planning
• Identifies regulatory objectives such as Investigational New Drug Applications and Biologics License Application.
• Defines any special regulatory pathways, activities, or options that will or might be considered.
• Prepares a regulatory risk-to-benefit analysis for this product. • Provides one or more possible regulatory outlines or roadmaps with
proposals to overcome perceived or real regulatory hurdles. • Identifies and proposes means to manage regulatory risks in the U.S. • Defines global, or ex-U.S., regulatory strategies, primary and alterna-
tive, considers each major market separately, and explains unique regulatory guidance and country-specific regulatory hurdles.
• Proposes responses to some possible regulatory changes that could occur before market approval.
• Identifies likely postmarketing regulatory activities and anticipated advertising and promotion guidelines and restraints for the product and the labeling claims.
• Identifies methods that would most effectively facilitate regulatory communication with each agency or office within an agency, defines each means of communication with a regulatory agency and at each phase—early, mid, and late—of development, and discusses the most challenging aspects of the regulatory communication plan.
• Provides an estimate of the number of investigational documents and market applications that must be filed and the temporal rela- tionships of each.
• Provides answers to questions, What are alternative regulatory routes to approval, such as orphan product or fast-track status that might
22 Biotechnology Operations
apply to this product and indication? Have any of these routes been tried with this class of product and, if so, what were the outcomes?
• Provides an answer to the question, How will compliance be accom- plished under current Good Manufacturing Practices, current Good Clinical Practices, and current Good Laboratory Practices and at which phase of development will they be needed?
• Provides answers to questions such as, If compliance activities are managed in house, what are the internal programs and guidelines for handling FDA inspections? Based on risks to the user associated with the product, is an FDA inspection likely during early investiga- tional phases of development?
Nonclinical Planning
• Identifies precedence and regulatory guidance for pharmacokinetic and pharmacodynamic studies performed at each phase of develop- ment for this class of biopharmaceutical and any predicate products.
• Identifies safety, tolerability, or toxicity factors that are of concern for the investigational product.
• Refers to intended human dose, dosing regimen, length of dosing, and route and method of administration in the clinical plan.
• Defines the most challenging aspects of nonclinical development for this product and indication.
• Outlines objectives, concept study design, and relative schedule for all intended studies: • Pharmacokinetic and ADME • Pharmacodynamic • In vitro toxicology • Acute toxicology • Subchronic toxicology • Chronic toxicology • Specialty toxicology in animals
• Identifies analytical or clinical evaluation tools or procedures that will be developed or used to measure endpoints in animals and describes how and when each is to be developed.
• Names the internal staff requirements at each stage of development. • Provides the general statistical approach for these nonclinical stud-
ies and give requirements for data handling, statistical analysis, and report preparation.
• Outlines the nonclinical study’s monitoring and auditing plans and describes how clinical quality will be ensured for each study.
23Introduction to Biotechnology Operations: Planning for Success
• Identifies nonclinical study materials (product) requirements based on the concept protocols and number of animals and treatment doses per protocol.
• Defines each concept study design once a clinical plan has been drafted, putting them into perspective with the overall development scheme, schedule, precedence, and guidance; examines all require- ments to achieve the objective: scientific, material, and time and monetary limitations; and proposes budget and schedule for each.
Biomanufacturing Planning
• Identifies and describes the product’s type or class and summarizes information on the biomanufacture of predicate or similar products after considering the TPP and research results on the product.
• Outlines a biomanufacturing design, including overall objectives and goals for each phase of development, early, middle, and late, after referring to the draft product design, and considers product risks and hurdles for the biomanufacturing plan.
• Drafts or outlines a biomanufacturing plan based on this design and considers product quality attributes both from the standpoint of process control and for quality control testing. In drafting the plan, it considers the ultimate objective, biomanufacture of commer- cial product upon market approval, and works backward: it begins with commercial manufacture of product and proceeds in plan each phase in reverse order.
• Provides a plan for scale-up of biomanufacture to produce required amounts of product at each phase and also considers purity and potency requirements at each phase.
• Defines plans for application of current Good Manufacturing Practices at each phase of biomanufacturing development.
• Considers each raw material or component that will be used in production and identifies potential quality criteria and the sources, and any regulatory guidelines on the quality of proposed raw materials.
• Identifies and reviews the history of any expression system or host cell line that will be used and determines if there is precedent for using the proposed production system and, if so, considers issues revealed in previous biomanufacturing efforts.
• Identifies any genetic engineering or other biological manipulations that might be required of the product or a host cell line before the product enters biomanufacture after reviewing the research back- ground on the product and its current status in research or early devel- opment; for example, the need to develop or modify an expression
24 Biotechnology Operations
vector, to evaluate a construct for a particular trait, or to characterize or do further research on a gene, a vector, or a host cell line.
• Plans the production and in-process and quality testing of any cell banks.
• Defines early, middle, and late stage development production schemes for this product, focusing on quality specification and quan- tity requirements and the chosen processes; considers upstream pro- duction and downstream purification processes for bulk substance and formulation, fill, and finish for the final product; and applies objectives and criteria for quality and quantity, yield, and scale-up at each stage.
• Identifies requirements for in-process testing after defining the processes.
• Considers once again risks associated with the chosen processes, raw material requirements, and unique aspects of production.
• Identifies special requirements for formulation, fill, and finish and labeling of the final product.
• Defines the containers or delivery devices to be used and storage conditions and requirements.
• Defines facility requirements for each stage of biomanufacture, con- siders both quantity and quality, and discusses approaches for meet- ing these facility requirements or for utilizing contract manufacturers.
• Discusses the need to provide aseptic manufacturing environments and requirements for clean work areas with classified air supply, segregation of product, potential for campaign manufacturing or shared manufacture, and flow of product within a facility.
• Identifies equipment and utility requirements at each stage of bio- manufacturing development and considers special environmental issues that are relevant to production of this product.
• Provides an overview of validation requirements and plans for the biomanufacturing facilities, utilities, equipment, and pro- cesses proposed in the manufacturing scheme and at each stage of development.
Quality Control Planning
• Understands from the TPP and draft manufacturing plan any requirements for quality control testing of product for both product release and stability.
• Identifies product, bulk substance, and final product attributes (e.g., safety, purity, and potency) as they will be considered for testing, and identifies one or more analytical requirements for each attribute.
25Introduction to Biotechnology Operations: Planning for Success
• Designs a quality control assay for each analytical requirement and considers a hypothetical specification for each.
• Designs the remainder of the assay development life cycle and har- monizes this with phases of manufacturing, nonclinical, and clini- cal development for each quality control assay.
• Provides a plan to identify how and where the assay will be performed and estimate resource requirements both for assay development and to perform the assay on expected samples, release, and stability for each assay.
• Identifies analytical methods that will be developed or used to mea- sure the quality of each cell bank that is to be tested under quality control. For each method, it describes how and where the assay will be performed and estimate resource requirements both for assay devel- opment and to perform the assay on each sample, and harmonizes them with the manufacturing plan.
• Identifies analytical controls and reference standards and describes how and when they will be developed or otherwise obtained and, in general, give qualitative and quantitative requirements for each assay.
• Outlines the initial (early phase) stability test requirements for bulk substance, final product, and cell banks or other intermediates. It describes the attributes that will be tested and identifies one or more tests for each attribute and outlines the stability test criteria that will be applied at later phases of development. It also describes any sta- bility indicating assays that must be developed beyond those con- sidered and planned for release of bulk substance and final product and outlines the frequency of testing under stability protocol.
• Describes any requirements for quality control to measure the qual- ity of the manufacturing environment or output of utilities.
• Identifies quality control tests that will be verified to ensure compli- ance with compendial methods and the phase of development for each verification.
• Identifies quality control tests that are candidates for qualification and state the requirements, expected outputs, and phase of develop- ment for each assay qualification.
• Identifies quality control tests that should be validated and give expected outputs and phase of development for each assay qualifi- cation and describes the resources that might be required for assay validation and harmonizes the analytical validation with manufac- turing development and the manufacturing process validation plan.
• Describes the program that will be developed to investigate test fail- ures or out of specification test results and investigations and har- monizes this with quality assurance plans.
26 Biotechnology Operations
• Describes efforts planned to ensure quality control is in compliance with current Good Manufacturing Practices (cGMP) and harmo- nizes them with plans of regulatory affairs and quality assurance.
Quality Systems and Quality Assurance Planning
• Considers the appropriate quality systems (e.g., current Good Manufacturing Practices, current Good Clinical Practices, current Good Laboratory Practices) that must be in place for compliance.
• Provides answers to questions such as, How and at which stage of development will each quality system be developed and instituted? Will they be performed in-house or by a contractor or partner?
• Identifies the hallmarks of quality that must be established for each quality system that will be instituted in-house or at a contract site.
• Identifies applicable U.S. and ex-U.S. regulations that drive the requirement for each quality system.
• Describes the requirement for in-house quality activities, and dis- cusses requirements for a quality assurance unit, quality manual, and quality plan.
• Outlines the elements of the quality manual. • Outlines the elements of the quality plan. • Describes the roles for quality in quality by design and at each stage
of design control. • Identifies requirements for the quality assurance unit and specifically
refers to needs for quality management, document control, auditing, and training.
• Identifies needs for quality agreements with contractors or collaborators.
• Identifies any special quality management requirements, such as, Total Quality Management, Six Sigma, or risk-based approaches.
• Identifies any requirements for quality assurance support in research activities.
Additional Elements of Product Planning
In addition to these seven functional area plans, three additional elements of project planning and implementation deserve mention. These are product design, project risk management, and the risk-to-benefit ratio of the product itself. Failure to apply these concepts and practices can result in delay or fail- ure of product development; hence, they are considered during the product development planning process and identified in the PDP.
27Introduction to Biotechnology Operations: Planning for Success
Product design immediately brings to mind an engineering endeavor, something that is applied to medical devices but not to a biological prod- uct. In fact, product design is an important aspect of the planning process and, in this book, the concepts and practices of design, design control, and quality by design are discussed at length in Chapter 5. Design is a process that focuses on the product itself and, as one might expect, design begins with product criteria and attributes listed in the TPP. Design focuses on criti- cal quality attributes that are often realized only after a certain amount of product development planning has taken place, and the product team has had an opportunity to review in detail the candidate product and the pro- posed development plan. In a practical sense, this means that design activi- ties, which are described in Chapter 5, often interrupt the planning process and require the team to revisit the nature of the product. This can even mean returning to the research laboratory bench and changing or tweaking the candidate product to improve it before it enters, or reenters, the develop- ment arena. Quality by design goes one step further, building quality into the product and hence into the product’s design process. In effect, this means that, in addition to user and performance requirements, there is a conscious effort to design quality criteria into the product. Of course, this means the quality criteria must be available as product development planning begins and suggests they be included in the TPP. Design also introduces the idea of design controls, steps in a formal design process, as discussed further in Chapter 5. Design is applied to development processes and, in Chapter 6, an example is given with the design of the manufacturing scheme.
Project risk assessment, mitigation, and management strategies involve application of procedures and practices to identify potential or actual risks and to reduce their chance of occurring or, should they do happen, their impact on a project. Risk management has a significant impact on improving product quality, safety, and effectiveness and hence is of direct consequence to the user. It should be part of every PDP, considering both the product and the development processes. Since risk management activities are often the purview of the project team, this subject is discussed in Chapter 2.
Risk-to-benefit evaluations are related to risk management and bring with them other connotations. The term and its concept were developed in the health products industries and by regulatory agencies as a means to convey a specific idea: Any product must deliver more benefit than risk to the user. Immediately one realizes this concept carries with it philosophical as well as practical and technical implications. We specifically ask how on earth do we weigh risk ver- sus benefit for any given biotechnology product. The simple answer is that somehow we do this for every biopharmaceutical before it reaches the mar- ketplace. A biopharmaceutical intended to treat cancer as a terminal disease is allowed to have significant associated risk, usually seen as side effect. A vac- cine intended to prevent a nonlife threatening infection in infants is allowed to have a low incidence of risks, and these side effects must be considered
28 Biotechnology Operations
mild. We make these choices, relying upon the judgment of experts with input from the public since as the user they are the object of risk or benefit. We are not always correct in these analyses, but overall our record is excellent. A PDP always considers risk-to-benefit of the product and, because it is driven in large part by regulatory authorities, it is discussed further in Chapter 3.
Summary of Planning for Success
The primary theme of this book, one that is ingrained into each chapter, is to carefully plan biotechnology operations. This demands that, once a product vision has been established, a long-range plan, the PDP, is produced to guide development, manage resources, and reveal to upper management and inves- tors the progress (or lack thereof) of development during this long period. However, to produce a meaningful PDP, it is first necessary to write a clear defi- nition of the product to be developed. Hence, a TPP precedes the PDP. The TPP is in fact a draft of the intended product label and states the product description, intended indication, and patient population allowable warnings or contraindi- cations, and dosage or route of administration. Hence, the TPP and PDP are key planning documents for the beginning, and they guide development through the products life cycle. This process has been referred to as planning backward, since we first identify, in the TPP, where we wish to be once the development is finished, then fill in all those tasks that must be accomplished to reach those objectives. Once the planning is finished, the actual development moves for- ward, following plans outlined in the PDP. Project teams, composed of those subsequently responsible for the product development, write a TPP and PDP.
29
2 Project Management
Biotechnology and Project Management
Market demand for biotechnology products encourages biotechnology firms and large, well-established companies to engage in product development, a process intended to move discoveries from the laboratory through the devel- opment cycle to the marketplace. Successful development adds value to a product and ultimately yields profit. Hence, the process of transitioning can- didate product out of discovery research and into and through development must be carefully managed. Yet, examples of mismanagement of biotechnol- ogy product development projects are noted regularly in trade and business publications.
What then might a biotechnology firm do to manage stress points and challenges, avoid development failures, and increase the chances of suc- cessful product development? One answer is a simple and inexpensive rec- ommendation: apply principles and practices of project management to the product development endeavor throughout the product development cycle. In other words, use project teams to apply principles of project management to integrate the six functional areas of biopharmaceutical development— manufacture, quality control, regulatory affairs, quality assurance, nonclini- cal studies, and clinical research—and coordinate these efforts under a team leader, the Project Manager (PM). When done correctly, and given a robust technology, it results in a successful outcome.
Project management is the discipline of applying tools, techniques, and skills to plan, organize, and manage resources through the various phases of a project to accomplish project goals. Project management has strategic plan- ning, operational, and closing phases as shown in Figure 2.1. In the planning phase, a group composed of functional area managers, the PM and corporate executives, consider the objectives and scope of the technical program out- lined in the targeted product profile (TPP) and Product Development Plan (PDP). This product development planning process, described in Chapter 1, is based on the biopharmaceutical product characteristics and indication, resulting in a TPP and a PDP. It is important to have the TPP and PDP com- pleted and agreed before writing a full Project Management Plan (PMP).
30 Biotechnology Operations
Indeed, these technical roadmaps are a requirement if the newly formed project management team is to draft a useful PMP. In effect, the PMP is incorporated into the PDP.
The PMP is transparent to the team and to upper management, and it applies concepts of project management such as team composition, com- munication, risk analysis and mitigation management, tracking, human ele- ments, project completion, project management tools, and resources, both human and monetary. These concepts are discussed later in this chapter. Project management planning greatly increases the chances of project suc- cess of meeting the objective of getting a product developed and to market, on budget and on schedule.
During the operational phase of project management (Figure 2.1), a project team, led by a PM, follows the project plan, always reflecting upon the tech- nical tasks, milestones, and schedule of resources and activities provided in the overall PDP. PMP provides a foundation and charter for the project team, individuals who will work together over several years to bring their product from laboratory to market.
A history of success is the reason that project management has proven so popular and effective in the biotechnology industry and why it is almost universally applied to biotechnology product development. There are other reasons. First, project management is very malleable and it allows a firm to customize project teams and a management structure for development of each product, no matter how unique the product or the project. Second, it relies upon team leadership to meet objectives, keep schedules, and move products to the marketplace. The cost of a single day’s delay in the biophar- maceutical industry can be one million U.S. dollars! Third, project man- agement is goal oriented. By its very nature and definition, a project team has clear goals, enabling the team to focus on larger product development
Initiate and
operate
Monitor
Output: Managed project
Development operation
Output: Completion
Closing
Plan
Output: Project Management Plan
Project budget Project schedule
Project objective and scope Draft targeted product profile and product development plan
Identify stakeholder and management expectations Agree to project and corporate governance
Apply project management and performance tools Technical functional areas; financial; communication and
feedback; risk and mitigation; team dynamics; resolving problem; and metrics
Clinical + nonclinical + biomanufacturing + QC + QA + regulatory
FIGURE 2.1 Stages of project management and elements of project management plan.
31Project Management
objectives. Fourth, projects are structured, and this structure is in a written PDP, described in Chapter 1. Structure provided in a PDP and PMP assists a project team to consistently achieve tangible and profitable results. Fifth, a project team applies management principles. Each project has a defined beginning, an end, a schedule for completion, and tools such as task lists and schedules that assist the PM and team. Project teams are consistent and resilient. A team stays with the program, from start to finish, pursuing the objective no matter if individuals leave or new persons join the team. Another tool is the shared budget for resources: human, fiscal, and capital. Resources are allocated per plan and according to schedule as PMs strive to maintain a balance in resources, as they are expended toward a common goal. Sixth, a team is diverse and professional. People with various skills coming from all backgrounds— contractors, employees, consultants, and clients—are intimately involved and work together on the team. Finally, there is synchronization, as phases and activities of the project are sequenced to balance resources, time, and performance against the objectives and the plan. This sounds a bit idealistic, but, for many well-led teams, it is a reality.
In this chapter, we review the field of project management as practiced in biotechnology operations, notably by biopharmaceutical firms. It discusses the history of project management, the construction of a project team, and selection of a PM, reviews operational principles and practices of leading a team, and it provides information on project management tools and tech- niques. Since project management involves social and technical elements, this chapter touches on both areas. Although not intended to be a com- plete review of the subject, it should serve as an introduction and provide some idea of how a biotechnology operation can be successfully managed. Hopefully, the reader will appreciate the value of project management and also understand how he or she could apply these skills in his or her work environment.
Background of Project Management
Project management evolved within the engineering industry. Specifically, it was first used on large, high-cost, and complex projects that applied cut- ting edge technology. Examples are projects to build the first atomic weap- ons, to construct large bridges, to put a man on the moon, or to build any major defense system or novel automobile. Advances in technology drove the need for project management. Projects became larger and more complex. Consider a feat like construction of the Panama canal, completed more than 100 years ago. It was so technically complex and grand for that time (and per- haps even for our generation) that the project begged for organized manage- ment. Further, individual workers brought to the workplace special skills,
32 Biotechnology Operations
and these individuals, and their work, had to be integrated and scheduled. Choreographed might be a better word. Two hundred years ago shipbuilding required woodworkers, blacksmiths, sail makers, and perhaps a few other skills. Today, designing and building new aircraft depend on the integration of individuals with thousands of skills and subskills. Costs of shipbuilding are managed in part by careful scheduling of parts and labor. In summary, project management appeared because it was needed in a technological soci- ety, and it has evolved to meet demands of cost, quality, and schedule.
In the 1960s and 1970s, as pharmaceutical development technologies became more complex and regulation of the drug industry further compli- cated this endeavor, the largest pharmaceutical firms began to adapt, from other industries, the principles of project management. At first, these prin- ciples were applied to pharmaceutical manufacture, as engineers, trained and practiced in project management, brought skills to increasingly more complex pharmaceutical plants. They were successful in managing teams and complex technologies, and this was noted by upper management. By the 1980s, project management was being applied to the full scope of pharma- ceutical development, from discovery to postlicensure activities. Also at this time, the new industry called biotechnology was just beginning to emerge. Not surprisingly, as scientists and engineers migrated from pharmaceutical to biotechnology firms, they transferred project management knowledge and skills to biopharmaceutical companies. Today, project management has been adopted by pharmaceutical, medical device, and biotechnology firms worldwide.
Project Management Plan
The process of planning project management itself begins once an objec- tive has been established for a development program and both TPP and PDP have been drafted (Chapter 1). A project may be a 3-month process to produce a recombinant DNA molecule for sale as a laboratory reagent, or it can be a complex, 10-year biopharmaceutical effort to develop a monoclonal antibody to treat a life-threatening disease of children. No matter the com- plexity or length of a project, both management and technical aspects must be carefully planned.
Hence, a project management plan establishes goals or objectives for the life cycle management of a project, recognizing hurdles and providing a long-range framework to minimize risks and to achieve goals and rewards. It also puts into place procedures and processes for management. Project management planning simply takes elements of good planning practices— planning for success, looking at the ultimate objective and defining goals along the way, and incorporating quality systems—and formalizes them
33Project Management
into a document or set of documents that can be shared by all team members throughout the life cycle of the product. If this is achieved and the objective is clear and shared by all team members, then transition from strategic proj- ect and management planning to operational project management is easy for the PM and the development team.
A project management plan may have five basic elements. Each reflects a phase in the life cycle of project management and indeed in the life of a technical project:
1. Initiation: Starting the project in a positive manner and formation of a team
2. Planning: The subject of much of this chapter 3. Executing: The technical and management aspects according to the
plan 4. Monitoring and controlling: Functions that ensure the project is meet-
ing objectives 5. Closing
Further to the five basic elements are a host of other considerations for a project management plan, outlined in Figure 2.2. These will be discussed individually in this and other chapters. It is worth noting that the project planning process, those meetings and discussions that seem to take forever, and the operation and execution of the development project itself include the
Team and dynamics
Contractors and collaborators
Project planning
Resources
Metrics and tracking
Objectives
Initiation and completion
Schedule Scope and complexity
Communication
Social considerations
Technical aspects
Risks and constraints
FIGURE 2.2 Inputs for project management planning.
34 Biotechnology Operations
same technical and management elements. Thus, considering each element in the project management plan is key to operational success of the project.
The Project Management Environment
The environment in which a biotechnology product is developed mat- ters to project management planning almost as much as the TPP and PDS. Biotechnology firms come in all sizes and with various types of structure or organization. These factors matter to effective project planning and manage- ment. For this discussion, firms are stratified and considered based upon size and complexity, with virtual biotechnology firms at one end of the spectrum and large, experienced companies at the other.
Virtual firms have few full-time employees. A project management team at a virtual firm might be composed of from one to a very few employees and in addition include outside partners, consultants, or contractors. A key rep- resentative, perhaps the titular CEO or a key investor, could lead this small team. Although there is little formal training or experience on the part of the PM or project team members and despite the fact that each team member may be responsible for two or more functional areas, small teams at virtual firms often outperform their counterparts at much larger biotechnology or pharmaceutical companies.
Small biotechnology firms normally have little project management infra- structure at the time their first product enters development. To establish project management at a small firm, the technology should have reached a level of maturity and the pathway forward must be clear. Specifically, five elements must be in place because they form the foundation for successful product management:
• Management decision and support: A business decision to move for- ward, made by executive management or a board of directors based upon project benefit or attributes, risk (technical and commercial), and resources. There is the intention to apply project management to product development, and there are or will be resources available.
• Planning: A written PDP and TPP or their equivalent provide ade- quate information, stating objectives and spelling out a clear route forward.
• Feasibility to begin to move forward: All elements of the plan are feasi- ble in the current financial, technical, and regulatory environments.
• Estimate of completion: A realistic schedule based on estimates of experienced professionals.
• Decision points or milestones: Milestones and decision points are evi- dent in the plan. For example, Go/No Go criteria to advance the proj- ect to the next phase are established at the beginning of the project and revisited at the start of each phase.
35Project Management
Once a decision has been reached to apply project management to a product development pathway, management intent is best demonstrated by appoint- ing the initial or core project management team and a PM and, most impor- tantly, relegating authority and responsibility for product development to this manager and team. Supporting a development project can be particu- larly difficult for the product discoverer, company founder, or executive of an entrepreneurial firm who has, for years, focused on discovery research or business development. To many founders, a seemingly easier route to suc- cess may be continuation of discovery research, where perceived risks are lower than those in the route of product development. For some executives, it is difficult to let a project team take control of functions considered essential to success of the firm. These are emotional decisions that must be made for the small biotechnology firm.
Established biotechnology firms may have it a bit easier when they begin a new development project because they have the experience and infrastruc- ture. Indeed, their existing and often mature project management programs provide experienced and highly trained staff dedicated strictly to building and managing teams. Many issues related to start-up operations—build- ing the first team and introducing employees to the principles and practices of project management—may not apply to the larger biotechnology firm. However, the established firm has other hurdles to productive project man- agement. A few are as follows:
• Complex organizational structure and rules confound efforts to complete any one project on time. For example, merger or acquisi- tion, a common occurrence in larger firms, results in changes to a major contractor, disrupting continuity of operations and schedules.
• Priorities change frequently and without clear direction from upper management. For example, the clinical indication for a product changes radically because of revised market objectives.
• Upper management is far removed from project teams. • A large organization may be slow to respond to opportunity or to
change, when these are necessary or desirable. • Projects are abandoned in midcourse and without explanation to the
development team. • Problems, incurred in one project, spill over to another project. • Elements of a project must be reworked because they are considered
unsatisfactory to someone outside the team. • Communication breaks down due to change in mode of communica-
tion. For example, a new videoconferencing system is required for all project team meetings, but it does not work properly.
• Corporate politics impact PMs. • Team membership changes during corporate reorganization.
36 Biotechnology Operations
These examples can complicate efforts toward successful product develop- ment in a larger biotechnology firm, and they have led some PMs to wish they were employed by a smaller organization.
Project Objectives and Schedules
Biotechnology firms, especially small and midsized companies, often suffer from malignant optimism and fail to recognize that few firms successfully complete projects planned under highly optimistic or unrealistic schedules. Hence, the PM is responsible for ensuring that a realistic schedule is com- posed and communicated to the team and to the management. History sug- gests this is a difficult task, since the common story in the biotechnology industry goes something like this:
• Project development team is formed. • Senior management provides the ultimate objective for product
development. • Project development team prepares a product development strategy
with schedule. • Senior management demands that work be completed in one-half
the time allotted by the team. • Project begins under the accelerated schedule. • Within 1 year, the project is offtrack, management is angry and team
members are discouraged. • Pessimism or outright failure in the face of a sometimes promising
technology.
What is the solution to artificial compression of schedules, an issue that constantly plagues small and midsized biotechnology firms? First, upper management must recognize that the product management team is com- posed of individuals who, together, have years of experience in estimating development times. These individuals have been responsible for meeting schedules in the past. Second, members of the product development team must recognize the need to expedite development, but not at great risk to delaying development. Moving quickly along the development pathway is a hallmark of the industry and provides the biotechnology firm with a competitive advantage. Although it is inappropriate for upper management to establish unrealistic schedules or to diminish resources below a certain level, PMs should take reasonable risks in establishing optimistic time lines. This is balanced against three very real project constraints: scope, time, and budget. Managers periodically weigh these constraints while advancing the project through each phase; balancing these constraints is a major challenge, while also taking into account quality and performance, in the product
37Project Management
development planning process; yet it is a key to success. Communication is key to ensuring every team member is aware of the constraints.
Project scope and complexity are important considerations to planning. Simply stated, there are simple and easy projects, and then there are complex and difficult projects. Yet others are somewhere in between these extremes. Project difficulty and complexity, if they exist, become apparent upon reviewing the PDP and have a great impact on the project management plan. Complex projects call for more involved and extensive project management.
Sociotechnical Considerations
To be effective, project management focuses on two critical areas: one tech- nical and the other social. First, it must apply project management skills to the plan with due consideration to implementation. Examples are establish- ing objectives, developing work breakdown structure (WBS), and monitor- ing resources. Project management must also influence individuals whose cooperation and help is needed to complete the project successfully, for example, establishing buy-in from the supervisor of a key team member. This need to use both technical and social skills for effective project man- agement has resulted in the realization that this trade is a sociotechnical endeavor. Executives of small biotechnology firms, though successful at influencing outsiders, such as investors or the scientific community, some- times lack the ability to influence technical aspects of product development. Hence, they call upon PMs to play this role. PMs must therefore have strong interpersonal skills, notably the ability to influence others to achieve project goals and to interact with upper management. Thus, it is critical for upper management to understand this and to follow-through by identifying and retaining experienced individuals, those with both technical background and social skills.
Participants in Project Management
A biotechnology project team is composed of many individuals, and they are led in this regard by an appointed PM. Teams vary greatly in size and scope, depending upon the complexity of the project and the size of the biotechnology firm. Each individual on a project team has a vested interest in reaching the same objective no matter what his or her technical skills, employment rank, or title. Team members may not be employees; instead they are consultants, contractors, or investors. Individuals who might be considered for inclusion on a biotechnology product development team are given in Box 2.1. Individuals who serve on teams have roles, both pro- fessional and managerial, and some have, shall we say, special status. It would be nice to think that everyone involved in a biotechnology project is equal in the eyes of the project and upper management. Unfortunately, this is seldom the case. A PM and the team recognize key participants in
38 Biotechnology Operations
a project, referred to as stakeholders. These individuals have a significant vested interest in the project, even though they are often not, from a prod- uct development standing, the most active members of the team. Indeed, some stakeholders, such as major investors or executive level management, seldom if ever participate in routine team functions. Their interests are nonetheless held above those of others on the team, and the PM pays spe- cial attention to their opinions and desires. Even though they sit apart from the team, stakeholders have great influence on team activities, and each stakeholder expects regular and often direct communication from the team, usually by way of the PM. From this, it can be inferred that the PM, in addi- tion to managing the team, is responsible for communicating with, indeed for influencing, stakeholders. This can be a stressful and time-consuming task in itself. Experience suggests that stakeholders often hold the positions described as follows:
• Project champion: This person, sometimes referred to as the project leader (as distinguished from the PM), is capable of influencing bio- technology projects based on scientific expertise, organizational power, or responsibility for a critical resource (e.g., a patent). In other firms, the project champion is an executive manager. In either case, they may be a figurehead (e.g., the historic founder of the firm or discoverer of the product or technology) and may or may not serve on the project team or even have a specific technical role. But they may also consult directly with project team members. They are often accountable only to upper management, such as a board of directors or president, and not to a PM or to the team.
BOX 2.1 SKILLS OF INDIVIDUALS ASSIGNED TO A BIOTECHNOLOGY PRODUCT DEVELOPMENT PROJECT TEAM
1. Project Manager
2. Project leader or project champion 3. Finance 4. Legal and contracts 5. Research 6. Business development 7. Marketing 8. Quality Control 9. Quality Assurance 10. Clinical studies 11. Nonclinical studies 12. Manufacture 13. Regulatory affairs
39Project Management
• Major investor: The golden rule is stated to be: He who has the gold makes the rules. This has great meaning to a biotechnology firm, where cash flow is always an issue. Today, investors are very proac- tive. Few attend project meetings, but most major investors expect to be frequently informed by the project team manager on techni- cal successes or failures, news about reaching or missing milestones, and updates on expenditures.
• Chief executive or board member: Executive officers in small biotech- nology firms are very hands-on with project teams. Most do not micromanage their teams, but instead stay in constant contact with the team leader and key team members. They are often scientifically astute, interested, and inquisitive. Keep in mind that they are an important bridge for your firm, communicating the good, bad, and ugly to analysts, investors, and the public.
Individuals actually serving on the project team may or may not be consid- ered stakeholders.
• Functional area manager or director: These individuals, and there may be many, are key architects of the project, responsible for decisions about strategy, plan, resource requirements, and determining status. Although they may not have authority to allocate resources, their influence looms large in other ways. They might direct key technical or administrative aspects for project support and frequently main- tain a commanding presence in the smaller biotechnology firm. They are accountable in two directions: to corporate executives on project matters, but to line management for functional responsibilities.
• PM: Individuals responsible for leading a team are influential as well. In some instances, they are or soon become stakeholders themselves, even though they may be subordinate to executives and directors. PMs often have great responsibility, but without direct authority. We refer to roles of a PM throughout this chapter. The way a project is managed and executed are keys to a project’s success or failure. Hence, it stands to reason that selection of the appropriate PM is an important decision. The manager should be experienced with a project of this scope and nature, although it is certainly not neces- sary for a candidate to have great technical knowledge in that area. Ideas for correctly matching a PM with a project are listed in Box 2.2. Attributes of excellent PMs is given in Box 2.3.
But, where do we find a PM with these attributes? In selecting a PM, the small biotechnology firm, with a staff of perhaps 20–100 individuals some of whom have previously served on project management teams, may have qualified applicants already on staff. Although the firm many not have a
40 Biotechnology Operations
BOX 2.3 ATTRIBUTES OF EFFECTIVE PM
General management Conflict resolution Leadership Team building Planning and scheduling Resource
allocation Anticipation of change Acceptance of change Adaption to
change Execution of change Effective communication Team building Negotiation Leading decisions Risk analysis Risk mitigation Risk management Organization Technical knowledge People and team skills Critical thinking Facilitation Begging, nagging, and playing devil’s
advocate
BOX 2.2 CONSIDERATIONS FOR SELECTION OF A PM
• What are the objectives and what is the anticipated length of the project?
• What is the scope of the project management function and hence the PM? Is it an individual project, a nested project, an integrated project, or a series of projects?
• Has the project team been previously led by someone, and if so, what was the outcome and what are the lessons learned from that leadership?
• Is strategic and operational planning involved? • Will he or she allocate resources, human or monetary, and
make priority decisions? • Is the project at more than one location, or in more than one
country? • To whom will the PM report and at what level within the
organization? • On what criteria will project management staff and team mem-
bers be selected? • How will their performance be evaluated? • What are the roles and responsibilities of project champions
and functional directors?
41Project Management
seasoned and full-time PM, there could be an employee who, through expe- rience at another firm, has basic skills and thus qualifies to lead a product development team. The midsized biotechnology firm will have experienced project management processes and, like a large firm, have project manage- ment staff with individuals willing and available to move to a new project. Selection must be rigorous no matter the situation. Individuals designated to the project team should have an opportunity to interview candidates. Final selection is influenced by those who understand and, preferably, have prac- ticed project management in the biotechnology industry.
Project Management in Biotechnology Operations
Establishing Project Management
It is important to decide exactly when to begin the formal process of project management. Some guidelines and common practices are instructive in this regard. Project management, as described in this chapter, is seldom used in discovery research, and so the concept is often foreign to the management of a small company. Certain scientists further argue that formal project man- agement inhibits good research because a highly structured environment is not conducive to discovery. Others suggest that it inhibits direct management of projects by executives. However, most would also agree that an organiza- tion developing a product through application of more than one functional operational area must institute at an early stage of product development some method to coordinate and integrate activities and participants. In the end, most biotechnology firms elect to apply project management principles and practices to their projects.
Given that projects, and hence project management, have a defined begin- ning, when should the biotechnology firm make the transition and establish formal project management? The best answer may be: whenever planning, coordination, and scheduling activities will, in some way, help the team and the stakeholders achieve a common goal.
The process is not difficult at experienced, typically large or medium sized, biotechnology or pharmaceutical firms. They begin project management at the outset of technical efforts, immediately on approving an operational concept and even before a PDP is prepared. These firms have professional project management staff to draw from or the resources to hire new staff. Larger firms have significant infrastructure in project management headed by a vice president dedicated to the task. Also, institution of a project team usually follows internal guidance, instructions from upper management, and established corporate guidelines.
42 Biotechnology Operations
The process is not often seen at the inexperienced and smaller biotech- nology firms. Despite any recommendation to make a conscious decision and begin project management at a defined point, the fact is that project management usually evolves at smaller biotechnology firms, with little conscious effort on the part of executive management. Executives may real- ize, perhaps after witnessing a failure or set back, that, to develop a product within the allotted time, a team leader is needed to manage the project, lead the team, ensure a smooth and timely sequence of events, and care for mundane items, such as setting an agenda, preparing minutes, communi- cating with stakeholders and preparing formal project management tools such as Gantt charts and reports. This process is project management by evolution, and it is a characteristic of less experienced biotechnology firms. Often in this situation, executives draw the project management designate from within the ranks of the project team, even if that person has other duties.
Another challenge to establishing a managed project in small firms is an open wariness of any managed development process and thus hesitation to appoint a PM. At the start, team members may voice many and varied ideas and opinions concerning the scope, purposes, and strategies for the project and disagree on management guidelines and styles. This is often the first sign that a new team is embarking on a sociotechnical endeavor and, in the absence of a full-time PM, early conflicts must be handled gently so as to avoid delay or disruption to the new project. Even more reason for executive management to complete project management planning, appoint a PM and organize a project team as soon as the requirement is identified.
The Work Breakdown Structure
In planning a new biotechnology project and preparing a project manage- ment plan, it is important to devise a WBS. The process that results in a WBS, itself a narrative document or chart or both, is based on an understanding of the deliverables, the materials, service, or product that the project is intended to produce. To begin, it is necessary to have completed the TPP, a PDP, and the intended scope of work. At this stage of planning, these documents are often rough draft documents. The WBS simply breaks the intended project into smaller, subcomponents that are more manageable. In a WBS for develop- ment of a biopharmaceutical product, the organization looks somewhat like an organization chart with branches representing the functional areas such as clinical and regulatory affairs. In practice, this is done first on a large sheet of paper or it can be organized using project management software, a tool that will be discussed in the section “Tools for Effective Project Management.” Work is broken down first by deliverables or milestones, then into subhead- ings referred to as tasks, then into subtasks, and so on. Examples are shown in Figures 2.3 through 2.5. Each figure shows work breakdown of the same project but using a different project management tool.
43Project Management
ID Task Name Duration Start Finish
1 Research 50 days Mon 3/14/11 Fri 5/20/11
2 Confirm Efficacy 5 wks Mon 3/14/11 Fri 4/15/11
3 Test Blood Levels 5 wks Mon 4/18/11 Fri 5/20/11
4 Quality Control 40 days Mon 4/18/11 Fri 6/10/11
5 Certify M9 Blood Assay 8 wks Mon 4/18/11 Fri 6/10/11
6 Develop M9 Tissue Assay 6 wks Mon 4/18/11 Fri 5/27/11
7 Assays Completed 0 days Fri 6/10/11 Fri 6/10/11
8 Manufacturing 135 days Mon 1/17/11 Fri 7/22/11
9 Manufacture Pilot Lot 40 days Mon 1/17/11 Fri 3/11/11
10 Manufacture Clinical Lot 40 days Mon 5/30/11 Fri 7/22/11
11 Product Available 0 days Fri 7/22/11 Fri 7/22/11
12 Quality Assurance 70 days Mon 3/14/11 Fri 6/17/11
13 Audit Preclinical Site 3 wks Mon 3/14/11 Fri 4/1/11
14 Review Assay Certification Documents 5 days Mon 6/13/11 Fri 6/17/11
15 Preclinical 156 days Mon 4/25/11 Mon 11/28/11
16 Write Protocol 8 wks Mon 4/25/11 Fri 6/17/11
17 Perform Study 86 days Mon 6/20/11 Mon 10/17/11
18 Order Animals 20 days Mon 6/20/11 Fri 7/15/11
19 Treat Animals 1 day Mon 7/25/11 Mon 7/25/11
20 Observe Animals 60 days Tue 7/26/11 Mon 10/17/11
21 Write Report 6 wks Tue 10/18/11 Mon 11/28/11
22 Preclinical Report Completed 0 days Mon 11/28/11 Mon 11/28/11
23 Clinical 84 days Mon 3/14/11 �u 7/7/11
24 Design Clinical Protocol 30 days Mon 3/14/11 Fri 4/22/11
25 Finish Clinical Protocol 14 days Mon 6/20/11 �u 7/7/11
26 Regulatory 41 days Tue 11/29/11 Tue 1/24/12
27 Draft IND Sections 4 wks Tue 11/29/11 Mon 12/26/11
28 Finish IND 21 days Tue 12/27/11 Tue 1/24/12
29 Submit IND to FDA 0 days Tue 1/24/12 Tue 1/24/12
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Task
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Project: Fig 2.1 MS Proj TU Date: Fri 5/21/10
FIGURE 2.3 Gantt chart format for biopharmaceutical development project. Schedule of events for a project shown in Gantt chart format including the work breakdown by task and task ID number. The start and finish dates and the duration of each task are given in the narrative listing. The right- hand panel depicts the project in chart format using solid bars to summarize the duration of a set of tasks (e.g., research) and shaded bars to represent individual tasks. Diamonds represent milestones and arrows interconnect tasks to reveal dependencies. Special computer software is used to compose complex Gantt charts.
44 Biotechnology Operations
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45Project Management
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46 Biotechnology Operations
Forming a Project Team and Hands-on Project Management
However, or whenever, formal project management enters the picture, proj- ect management planning will begin with a meeting of project management team members led by the designated PM. Note that this first activity is project management planning, not operational or product development planning. To be successful, upper management must wholeheartedly and visibly support the institution of formal project management whether or not there is dis- sention regarding the need for a project team. Indeed, a way to open the first meeting is to have the president or CEO review for team members the development strategy and the initial or draft project management plan. This empowers the team and its leader and clearly demonstrates the intended objectives, deliverables, and measurement criteria of the project; it gets every- one on the same page as a team. Network building, as team members form interpersonal relationships, is another important objective of the first meet- ing, as networks form links and bonds between individual team members. These first steps have been referred to as the forming stage of a project team.
Once the strategic objective has been reviewed, the PM might focus the team on critical elements of a project management plan to ensure that all team members share the same vision. This stage has been referred to as the storming phase, perhaps because such discussions can seem quite unsettled. One of the most important tasks is development of the team charter, and this is expanded in Box 2.4. If nothing else, this first meeting establishes, through the outline of a team charter, team identity both internally, among team members, and externally, to senior management or to investors. Finally, the team agrees to certain attributes that have been stressed by or seem impor- tant to upper management. Some examples (and these may be very different depending on project objectives, the firm, and the team) are that the team must meet expectations of stakeholders, be on or under budget, and keep on schedule. Note that the team, not specific individuals, now meets goals and objectives. This has been referred to as the norming stage as team members agree to norms or standards. For some members new to project management teams or the concept of teamwork, this may be a new concept. Thus it may require explanation by and patience on the part of the PM.
With this accomplished, the fourth, longest, and final stage of a project team may begin and this is referred to as performing.
Team Dynamics
Team dynamics refers to various team activities and interactions, often defined as communication. As one might guess, social interactions, especially those identified in Figure 2.6, are at the heart of team dynamics. Of greatest concern to the PM team are identification of risks, resolution of problems, and the decision process, three highly related and very dynamic responsibilities.
47Project Management
Decisions are often required when problems or issues arise and must be addressed by the team. In a high-technology industry such as biotechnol- ogy, we frequently encounter problems. These may be technical in nature, but often they are based in regulatory, quality, or management and admin- istrative areas. Our problems are also derived from the fact that this indus- try is competitive and therefore fast paced and dynamic. Recommendations
BOX 2.4 GUIDELINES AND ELEMENTS OF A CHARTER FOR PROJECT TEAMS
• Establish team work rules: • Identify means of selecting milestones and deliverables
since these are key to project development and tracking. • Agree on means to identify and track tasks. These basic
building blocks, often technical, must be pursued in sequence.
• Discuss means of team communication. • Establish milestones:
• Points in the project, actually major events, at which time progress can be measured against an objective.
• It may be necessary to initiate planning in various func- tional areas before this begins.
• Define required skills: Who must sit on the team and when in the project cycle will their input and skills be needed?
• Reveal constraints: Identify boundary conditions, notably as resources.
• Outline a team charter and include key elements: • Our team goals • The team’s members and what each represents • The roles of each team member • Our project team manager, leaders, and stakeholders • The team quantity (size) • Our team’s qualities (skills) • The team’s identity
• Set the network: Draw out an organization chart for the team. • Share important technical information: Even use the first meeting
to begin technical cross-training and discussions.
48 Biotechnology Operations
for addressing problems typically encountered by project teams are listed in Box 2.5, where problems are classified as one of two types: adaptive or technical.
Risks are related to both problems and decision making and are related to solving problems and making decisions. Risks impact product development, adding to the already complex nature of projects and their management. PMs are often called upon (or volunteer) to take the lead in risk identification and mitigation; or at least lead the team in these functions. Risk management is discussed in the section “Project Risk Assessment and Management.”
PMs cannot and should not attempt to measure risk or resolve problems by themselves. Instead their role is to motivate team members to find the best resolution in the shortest period of time and to lead the team in the problem- solving process. The PM also communicates the problem, and its resolution, to team members, upper management, and stakeholders.
The most important and challenging team dynamic for which PMs provide a leadership role is the decision process. Decisions face the team at virtually every meeting. Someone once said that the only reasons for having a team were to solve problems and to make decisions! Although this might not be completely true, it makes an important point. An agenda would seem barren without the need to make at least one decision. Experienced PMs have likened the process of leading the decision process within a biotechnology team to the activity of herding cats. To avoid unsettled outcomes, the prudent PM estab- lishes, at the first team meeting, decision-making guidelines. The process itself should be identified and recorded, so that it is followed in the future. Criteria for reaching a decision (e.g., by majority rule and by consensus) and time lim- its are established. The PM needs the support of every team member, even those who argued against the chosen course of action, once a decision has been reached. Meeting minutes reflect each decision taken by the team. Finally, the
Respect
M ak
e
Resolve Focus
Engage D
em ea
no r
IdentifyCom municate
FIGURE 2.6 Team dynamics. Positive team dynamics result from an engaged team, where members engage each other respectfully and focus upon relevant issues, all in a positive demeanor.
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team should consider how decisions will be communicated to stakeholders and how failure to reach a decision might be quickly and amicably be resolved.
Communication and Feedback
Once a team has been established, the charter written and the PM appointed to lead the team, it is necessary to consider communication issues. Communication for the project team requires interactions between team members, the project sponsor (typically senior management), functional area managers, executive management (e.g., president and chief financial officer), and business users, the likes of sales, marketing, legal, contracts, and business development. Although
BOX 2.5 MEANS OF TEAM COMMUNICATION
• Formal meetings: Even if the team is scattered in several coun- tries, there is a need to meet face to face at regular intervals. Formal meetings always have an agenda and result in formal minutes.
• Informal meetings: Yes, there are times when some, but not all, members of the team must meet. Informal meetings are best summarized by e-mail messages or memoranda that identify the purposes and outcomes for the meetings and are shared with all members of the team. The PM may be responsible for gathering these messages and providing them to the larger team. Video or telephone conferences may be used for infor- mal meetings and, for some participants, for formal meetings.
• E-mail, Telephone, and FAX: Today, e-mail is a primary means of day-to-day communication, especially when individuals are separated, even by short distances. Telephone conversations remain very important to the professional. How and when is the content or outcome of these communications to be copied to the PM or to other members of the team? Often this is done periodically, perhaps at meetings, where important elements of previous conversations or messages are revealed to members of the larger team. In other cases, important communications between two team members are transmitted directly to the PMwho then ensures that they are disseminated to other team members on a need to know basis.
• Project management tools: Today the PM has at his or her dis- posal computer-based tools, such as MicroSoft Project® and Share-Point, and document sharing tools, such as Box®. These may be used to identify and outline tasks, define resources, track goals and tasks, and support preparation of reports.
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these individuals may not attend each project management team meeting, they are by nature team members. Indeed, team activities and decisions will affect them directly. Their membership and participation are encouraged, and effective two-way communication, in whatever format, is essential to success. The PM is kept informed of all current events by each team member.
The PM outlines a communication plan with the purpose, frequency, and format for communication between team members and with executive man- agement and stakeholders. Examples of project messages are a corporate- wide or public announcement, upon reaching a milestone, or the financial information, first reviewed in a budgetary meeting with finance before the close of a fiscal quarter. Today, there are many methods of communication from which to choose, but proven methods used by the biotechnology indus- try are listed in Box 2.6.
BOX 2.6 STEPWISE APPROACHES TO RESOLVING PROBLEMS
1. Identify the problem. Be very specific. 2. Classify the problem. Is it adaptive or technical? 3. Assess the problem in relation to the team and team members.
Is it the team’s or the project’s problem or does it belong to another entity? If it does not belong to this team, then refer it to the proper authority.
4. Identify the importance of the problem to operations and objec- tives of the team.
5. A PM ensures all team members are aware of the issue and remain engaged and informed throughout the remainder of the problem-solving process. Keep the team informed, focused, and involved and maintain an urgency to find a cause and resolution.
6. Look for causes and identify the nature and source of each pos- sible cause.
7. Identify approaches to resolve the problem. a. For technical problems, consider technical solutions.
Engage technical experts and apply scientific methods. b. Adaptive problems are often more difficult to address or
resolve. They may be complex, occur in a changing envi- ronment, and lack predictability. It may be necessary for a team composed largely of individuals with technical or sci- entific backgrounds to resolve an adaptive problem. Seek help of experts.
8. Once resolved, document the problem, the process, and the resolution. Implement the solution.
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Project Risk Assessment and Management
Risk identification and management, important assets to a biotechnology project, are often project team functions, at least for projects under the team’s purview, and are led by the PM. Processes inherent to project management lend themselves to identification and control of risks. The PM is assisted in risk management efforts by others on the team, often including representa- tives of quality assurance and regulatory affairs.
Each biotechnology operation faces a number of risks, as exemplified in Figure 2.7. Team meetings provide a forum to discuss each risk, perceived or actual. Yet, it may be difficult for the project team to hold open and frank discussions of risks, even those with a high probability to occur and then confound, delay, or impede a project. Perhaps this stems from the nature of biotechnology itself, an entrepreneurial endeavor operating in an envi- ronment with many inherent financial, technical, and operational risks.
Common project risks in biotechnology
operations
Unacceptable safety profile
Low efficiency
Unacceptable quality
Regulatory disapproval
Unable to meet schedule
Excessive cost
FIGURE 2.7 Examples of project risks in biotechnology operations. Which of these are adaptive, which are technical, and which may have elements of both? Which are often considered together, proving that we seldom encounter a single risk?
52 Biotechnology Operations
Yet boards of directors and a biotechnology firm’s investors expect that significant risks be revealed, at the time of investment and throughout the operational phases of a project. Regulatory agencies increasingly demand that risk analysis be part of any product development program since such efforts result in safer and more efficacious biopharmaceutical prod- ucts. Hence, project teams in biotechnology development are increasingly becoming the clearinghouse for risk assessment and mitigation and, with this, the PM becomes a leader in risk identification, assessment, and mitiga- tion activities.
The practical aspects of project team risk management require consider- able time and effort. A project team is aware of their role in risk management and ensures defined processes are in place so each response is appropriate and all responses are consistent. The process allows for the initial assess- ment of potential risk elements and for prioritization of the risks as well as for implementation of mitigating actions and periodic reassessment of risks. A project team performs an initial risk assessment at the outset of a proj- ect and renews and revises this assessment at predetermined milestones or whenever significant changes are made on a project. For example, product and process risks are reexamined prior to clinical trials, in the case of medi- cal products, or before field studies, for agricultural products. Biotechnology operational projects establish milestones, and risk assessment is considered a milestone-related task. The PM ensures that risk assessment programs are established with milestones and scheduled for completion. Whenever a project team is involved, risk management is part of the project management plan. At an early meeting, the team agrees to the most likely possible risks proposed by individual team members. Next, each risk is subjected to one of three types of risk assessment—fault-tree analysis, informal assessment, or failure mode and effects analysis (FEMA)—by the appropriate professional or committee. Informal risk analysis is used whenever the likelihood of a risk is low, if it is a highly technical problem or if the risk has minor conse- quences. One individual or a few people may review the issue and report back to the project team.
Fault-tree analysis is a complex process often used in engineering sys- tems, but can be quite helpful with some biotechnology operational endeavors, such as biomanufacturing. FMEA is most commonly applied to projects in the biotechnology, drug, and medical device industries. A team usually performs this analysis. Every step of a process or feature of a product is listed, and possible failure modes are identified. Then each possible failure is assigned a score, and the team makes plans to measure and address risks that exceed a certain score. FMEA can be time consum- ing upfront but, under the guidance of a project team, it ultimately saves considerable time and resources. The PM must drive these processes and ensure they are brought to completion. Finally, it is often the PM, speaking on behalf of the team, who communicates risk information to executive management.
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Metrics and Tracking Progress
Once the project is underway, the PM is responsible for tracking prog- ress of each task against the plan and schedule. Metrics demand the team have a method to measure progress of each task and of the overall project. Although some tracking is done informally, using memory and communi- cation with colleagues, most is performed using written schedules and lists of interrelated tasks and milestones prepared with computer programs such as Microsoft Project®. A scheduled list of tasks forms a track or road- map for the project; this can be visualized in Figure 2.3. Such tools are introduced later in this chapter. The purposes of tracking are both to moni- tor progress against objectives (e.g., milestones) and to control the process. Tracking allows the PM to predict if any piece of the project (e.g., a task) seems to be at risk of failure or is heading offtrack.
Specific activities in tracking include collecting actual work and cost per- formance information and estimates to completion of milestones. The PM then compares actual performance with the plan and, if necessary, revises tasks, working with project team members in an effort to bring the proj- ect back ontrack. Indeed, a diligent PM uses tracking to identify problems before they delay or limit progress. Tracking methods are also used to com- municate issues to the project team. Reports to senior management and to investors are based upon data obtained by timely and accurate project tracking.
Additional guidelines for effective project tracking by the PM are to
• Review all aspects of a project at regularly scheduled meetings. • Make changes between meetings and notify team members. • Use various tracking tools, timelines, and charts, to communicate
with the project team. • Keep management and team members informed through meeting
summaries and minutes. • Consider that changes in project/schedules necessitate both reas-
sessment by the team and changes in resources.
Since biotechnology development projects frequently encounter issues that must be addressed by the team and since potential delays are not uncommon to product development operations, tracking is an intensive but important aspect of project management.
Another metric tool available to PMs and teams is earned value manage- ment (EVM), which is a means of identifying variances. This, in turn, allows teams to make more accurate forecasts, which management greatly appre- ciates, and to recommend changes whenever necessary. Its use depends upon application of proper metrics. EVM compares work performed to work planned, as shown in Figure 2.8.
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Resources: Planning and Usage
Resources are the people, facilities, equipment, raw materials, and money applied to a biotechnology development project. The process of allocating or reallocating resources is referred to as budgeting. Budgets themselves are negotiated during the project’s planning stage, and executive management provides a team with a specified amount of resources. Each team member, having outlined the pathway and schedule to the objective has, for his or her functional area, identified the resources his or her department needs to achieve objectives. This process requires great time and effort, some negotiation, and full justification. The PM is responsible for preparing an overall budget, by task, year (or quarter), and functional area manager. The PM tracks progress of the project against consumption of resources and is responsible for identifying budgetary risks and overages. This is done peri- odically, and reports are provided to team members and executive manage- ment. Project management programs, such as Microsoft Project®, are very helpful, as they provide a means of entering and tracking resource usage as compared to completion of tasks or achievement of milestones. The result- ing charts and graphs allow the PM and team members to visualize this information.
0
20
40
60
80
100
120
5 10 15 20
Ea rn
ed v
al ue
v s.
ex pe
ct at
io ns
(% )
Time (wks)
Task 1 Task 2 Task 3
FIGURE 2.8 Earned value versus time. Three tasks, each represented by a distinct line in the graph, are expected to complete by week 20, a major milestone, of our project. The PM has measured progress for each task, as a percent of anticipated progress at 5, 10, and 20 week time points in the project. Now, at week 15 and using this earned value chart, he or she projects an outcome for each task at 20 weeks, projecting how the curve might appear between weeks 15 and 20. Task 1, marked by diamonds, has progressed very well, is on schedule, and should complete by week 20. Task 2, marked by squares, is making progress and may be completed by week 20, but, then again, it might be delayed. Task 3, marked by triangles, has lagged behind schedule throughout the project and, without immediate assistance and perhaps some luck, is unlikely to complete at week 20.
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Finances are important to a project, but people are the primary resource and so a PM considers the many facets of human resource planning and use. The PM is aware of several human resource factors. First, most staff assigned to a proj- ect actually report to functional area supervisors. Ultimately, it is the respon- sibility of functional area managers or supervisors, not of the PM, to manage staff and performance issues on a day-to-day basis. Indeed, many members of a project team will be responsible to more than one project, and each of their staff has additional responsibilities, as well. Furthermore, since human resources are typically the greatest expense at any company, salaries have a tremendous impact on the overall project budget. The PM pays particular attention, through tracking, to utilization of staff and supervisors assigned to their project. Also, if not enough staff are assigned to a particular technical task, then the project is at risk of failure. Even if a staff member is assigned to a team full time, their time is defined and limited. Planning and tracking of human resources are ultimately and routinely the responsibility of functional area supervisors, but, for the overall team effort, they are always major concerns to the team’s PM.
Budgeting monetary requirements is a process referred to as costing by PMs. As noted earlier, there are people costs (internal employees), but there are also external (e.g., consultant), capital, revenue operating, raw material, energy, and other project costs. Accurate costing may be beyond the train- ing or experience of some PMs. Hence, a prudent PM will ask for assistance from the finance department to plan cost budgets and to serve on or advise the project management team. Since the financial department is ultimately responsible for the annual corporate budget and since financial staff have experience in estimating costs, they can be immensely helpful, even indis- pensable, to a PM and the team. Indeed, a financial officer is a great asset to any project team, no matter how technical the objective. Yet even with the assistance of financial staff, the PM, representing the team, has continuing responsibilities for planning and tracking a project’s resources. Project plan- ning tools provide the PM with a means of accomplishing these objectives throughout the life cycle of a project.
Human Factors in Project Management
A newly appointed PM either adopts a preexisting team, builds, or rebuilds a project team. We have already discussed administrative aspects of building teams and human resource functions, but there is another side to the issue. Even though the PM does not directly supervise team members, they still take on numerous responsibilities related to coordinating activities of team mem- bers and they do this in both the planning and performance stages of a project. A short list of a PM’s human resource duties might include the following:
• Integrating human resource planning and incorporate strategic planning into the project
• Structuring teams
56 Biotechnology Operations
• Fostering working relationships between team members • Interacting with supervisors and human resource professionals • Respecting dual roles or career paths (e.g., discovery scientist and
development leader) of team members • Leading creative and innovative people • Remaining open to new ideas from team members • Managing unique personalities • Anticipating and then managing change in teams and team members • Managing conflict within the team • Trusting team members with scientific and technical expertise
Interestingly, most of these responsibilities match skills that were listed for effective PMs (Boxes 2.2 and 2.3). Although managing a team can be a daunt- ing task for anyone, it may quickly overwhelm a new PM, the individual with little experience or training in interpersonal relationships or in human resource management. For those with no supervisory authority, human issues can be the leading cause of anxiety, frustration, and stress. This is particularly true in smaller biotechnology firms, where the PM may have limited experi- ence and little or no authority but the responsibility to manage team mem- bers, some of whom might be brilliant, opinionated, scientifically experienced, extremely busy, or hold lofty titles. Certainly this is a challenge for any PM.
The PM also ensures a balanced team, one composed of members suit- able for achieving the objective. Balance must be established at the project’s initiation and maintained through the project’s lifetime, even as the need for certain skills varies from phase to phase. The PM spells out roles and expec- tations for each team member, and, since executive management may not agree on every proposed position, filling out a team may involve explanation and negotiation.
The most effective teams have a project champion, sometimes called a project leader, in addition to the appointed PM. The PM must ensure that the roles of project champion, a proponent of the technology and often a stake- holder or influential scientist, do not conflict with theirs. It is important for executive management and everyone on the team to recognize that the role of PM differs from that of project champion or from those of each functional area director serving on the team. These roles are based, to a great degree, on each organization’s philosophy, organization, and policies. The PM must understand this and adapt his or her team leadership style accordingly.
Thus, there is a need for the PM to manage egos, the team, and the project. Not long after project teams are formed, the PM and functional area manag- ers or directors may come to view each other as different and even difficult individuals in each other’s minds. Yet, in a successful biotechnology opera- tion, each person will realize the importance and the contributions of others on the team.
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However, there are instances in which true emotional hostilities break out between team members or the PM. Dealing with difficult people or difficult situations is a critical issue in project management. Indeed, some argue that conflict is inevitable on any project team, even a well-managed one. However, the PM is well advised to never ignore conflict, but to recognize it, identify the sources, and work with the team to manage disputes, overt or hidden, and simmering. There are many sources of conflict, a few of which are listed in Box 2.7. Conflict is recognized by the PM in many ways and some, such as body language or facial expressions, are subtle. Conflict must be differ- entiated from disagreements, which can actually be a positive for team per- formance as it engages team members in healthy debates and discussions. However, a real conflict is never to be ignored because it can get out of control and disrupt progress. More often than not, executive management expects the PM to resolve conflicts that involve the team or the project and to lead the team down a pathway paved in productive behaviors. It is often necessary for the PM to mediate conflicts and negotiate resolutions. There are many approaches and tools, some of which are shown in Figure 2.9. In summary, people skills are essential for effective project management and leadership.
Project Completion
Yes, projects are actually completed, perhaps not always on time or within budget, but, like a movie, each project does have an ending and some are happy and others sad. Wrap-up is an important part of any project and
BOX 2.7 COMMON HUMAN SOURCES OF CONFLICT ON PROJECT TEAMS
• Team size or composition not suited to project • Lag in schedule or task or project priorities • Technical failures • Inherent tensions (especially true with matrix organizations) • Disagreements, often longstanding, on results or decisions • Individual background or developed styles of team members
(e.g., communication) • Difficult individual behaviors (e.g., divisive, passive, and
aggressive) • Senior management intervention or micromanagement of the
team or project • Personal work styles • Scarce resources
58 Biotechnology Operations
should be considered by the PM as a separate task. What better way to indi- cate to the team that an end is in sight than to reveal, early on and in the proj- ect schedule, a date for the final project meeting! The last meeting includes a project wrap, simply a review of what has happened: the good, the bad, and the ugly aspects of the project. The final meeting is a learning experi- ence as well as cause for celebration. Paperwork, such as reports, and other outstanding responsibilities are assigned and scheduled. From the meeting, a lessons learned document is produced for management. It need not be long or detailed, but it must be honest and reflect actual team performance and project outcomes. Whether technical aspects of the project succeed or fail, the team is recognized by the PM and executive management for a job well done, a team effort.
Lessons learned reports and meetings need not wait for total completion of a project. They can also be used whenever major milestones are reached or when a significant risk or issue has been resolved. Such a look-back exercise allows for team members to share experiences and better prepare for future challenges.
PM facilitates communication
Compromise
Problem solve
Accommodate
ConfrontDirect
Reconcile
Collaborate
FIGURE 2.9 Preventing or resolving conflicts. The project manager facilitates commu ni cation and hence pre- vents or resolves conflicts by applying various social skills.
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Project Management with Contracts and Collaborations
Outsourcing of technical efforts, such as manufacturing, quality testing, and nonclinical animal or clinical studies, is a common, indeed an important, practice in the biotechnology industry. Managers from functional areas are responsible for their contractors, and they assign one individual the respon- sibility for outsourcing a piece of work and managing the agreement. He or she has specific technical, project management, and contractual experience and can best ensure a successful outcome to partnerships and agreements. The PM seldom has direct responsibility for a contractor or consultant, but they do consider all contractual efforts as integral to the overall project. Hence, the budgets and schedule of tasks and milestones for a contractual, collaborative, or consultant’s efforts are, in all respects, part of the project and considered in the PDP, WBS, and schedules and reports.
Numerous outsourcing models are available to the PM. Some examples are as follows:
• Competing several vendors with similar and acceptable capabilities • Selecting vendors from a list of prequalified contractors • Partnering with a particular vendor or sole-source contracting with
an established vendor
Virtually any service or material may be outsourced by a firm. Vendors pro- vide functional and technical services, such as manufacture and regulatory affairs. Although not generally recommended, the virtual biotechnology firm may even use a consultant or contractor to provide project management services and to manage their other contractors, consultants, and vendors.
In addition to managing contractors or vendors, biotechnology firms often collaborate and codevelop products. These business arrangements are usu- ally between two biotechnology companies or between a biotechnology firm and a large pharmaceutical firm. Another business model is partnership between a biotechnology firm and a contract research organization (CRO). Some CROs provide services in part for equity in the product and sponsoring firm. Whatever the business arrangement, the project team at a biotechnol- ogy firm must follow progress of each and every aspect of codevelopment.
Biotechnology firms often enter into partnerships with larger biotechnol- ogy or pharmaceutical companies. Here, interfaces may be quite broad and also have depth, extending well into highly technical endeavors. In a partner- ship example, one party manufactures product, whereas both parties provide quality control testing services. Each partner must understand the nature of the product and the full scope of manufacturing and control. In addition, they both must have a clear understanding of all technical details in these areas. Teams can become large, with many functional specialists, up to 50
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residing at several locations, comprising a single team. This in itself presents a challenge to the PM. When developing partnerships in operational areas, individuals in business may not appreciate future needs for professional project management to guide the relationship. To ensure success in highly technical partnership, there is often a need to meld two different technical and business cultures (e.g., the culture of a large pharmaceutical firm with the culture of the biotechnology firm), and this requires much coordination between many individuals at both organizations. It also involves contract or legal specialists for both parties. PMs are often responsible for forming integrated project teams and ensuring effective project leadership. Business development must work with PMs before a partnership deal is consum- mated, and PMs are well advised to include business developers on their teams. Alternatively, arrangements with partners may have collaborating parties working quite independently from each other. In such cases, interface between the parties is more commonly one on one: between respective PMs and between like functional area managers from each organization.
No matter what the business or management relationship, strong proj- ect management experience, leadership, and negotiating skills are absolute musts if these arrangements are to succeed. Hence, the project team and manager are fully aware of all contracts and collaborations and the state- ments of work, roles, and responsibilities of each one.
Virtual Teams
Today many teams can be described as virtual, and this is especially true in biopharmaceutical development. Twenty-five years ago team members were typically collocated, but today individuals are often separated by distance, time zones, and organizational boundaries, and work together as a virtual team. Add to this the international nature of many teams, complicating mat- ters with very different time zones, native languages, and cultural practices. Electronic communication and digital technologies facilitate such project teams, and many PMs are experienced at working in such venues. The criti- cal success factors for virtual teams are many; indeed, books have been writ- ten on just this subject. A few issues are worth highlighting. Cross-cultural understanding is a must, and it is often the PM’s responsibility to facilitate cultural understanding and to ensure that confusion and division do not arise from such matters. It is most critical that every team member has a well-defined role, one that is understood and appreciated by all other mem- bers. Cultural differences across organizations, even when they are located in one country, must be considered. Electronic collaboration and communi- cation technology must be current, rapid, and dependable. The latest gadget may not be the best, but modern, proven technologies must be available to
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each team member. Face-to-face meetings, to include at least key members of the team, are essential at least in the beginning and periodically thereaf- ter. Stakeholders and senior management and supervisors of team members may be numerous, widely scattered, and reflect the cultural makeup of the team. Standard team procedures and guidelines are most essential with vir- tual teams, and it helps to have them written. Despite these issues, virtual teams have proven to be very effective in the biopharmaceutical industry, especially when there is excellent and experienced team leadership.
Tools for Effective Project Management
How is it possible to put together a project and then communicate and track it over several years especially given the complexity, size, and inevitable changes to many biotechnology efforts? Today, PMs have at their disposal and at reasonable cost, comprehensive and powerful project management tools to help in these efforts. Microprocessors and project management soft- ware provide four areas of project support. They are used to define plans and schedules, identify resources, track tasks and milestones, and produce reports. Software will assist the PM in establishing a WBS, listing each task and placing under it any number of subtasks. This organizes the project so that the reader discerns project structure and definition.
The process itself is rather simple. The user first prepares a list of what must be accomplished, breaking the list down in outline format. The list, a breakdown of the project by tasks and milestones and estimates of the sched- ules for each, becomes the input for entering each task and subtask into a project management computer software program. Tasks and milestones are also linked to each other as a means of identifying and demonstrating depen- dencies and interrelationships. The software then presents this information in both graphical and written format to the user; this is the draft or initial output of a WBS (Figures 2.3 and 2.5). It is then shared by the PM with other members of the team for review and comment. With a little training, each team member may now visualize both the overall project plan and WBS and they then visualize their designated role and responsibilities for the project. Task and milestone relationships and integration into the overall schedule are also clear. Potential risks and problems of the intended project become apparent to each team member when witnessed in graphical format; hope- fully these weak points can be corrected. The next step in using this tool is for the reviewer to identify specific steps that might be taken to resolve issues over the course of the project. The draft WBS, referred to as output, may now be revised based on recommendations of project team members. This review- to-revision process is repeated several times before project tasks, milestones, integration, and schedules are finally established to the team’s satisfaction.
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The computer software or program most widely used by PMs today is Microsoft Project®, but other good programs are on the market. A Gantt chart, shown in Figure 2.3, depicts the output from MS Project and pro- vides an example of how this chart demonstrates a WBS. For this example, the project was divided into functional areas and each was entered as a line into Gantt format as research, quality control, manufacturing, qual- ity assurance, preclinical, clinical, or regulatory affairs. Then tasks, each a technical or administrative step in a project, were entered under a respec- tive functional area. A task is a piece of work, clearly definable in terms of technical requirements and schedule. In the example, the tasks Write Protocol, Perform Study, Write Report, and Report Completed were listed under the functional area Preclinical. Tasks were further broken down into subtasks, and this is shown in the example (Figure 2.3) under the area of Nonclinical and task Perform Study. Here, three subtasks were entered as Order Animals, Treat Animals, and Observe Animals. As each task or sub- task was entered, it was assigned an identification number, to the left of the name, a start and finish date or duration and predecessor tasks, to the right of the name. For the example of the Preclinical task ID number 21, Write Report, it began on October 18 and took 6 weeks, until November 28. It had one predecessor task, ID #20, and successor task #22, Report Completed. A milestone, which is a major event in the project schedule, is completion of the report; this was added to indicate the date on which this and other related tasks were completed.
The software produces a visual in Gantt chart format, to the right of these entries. Horizontal clear bars identify each task against the schedule or cal- endar, whereas the overall schedule for each functional area is shown in a dark horizontal bar. Milestones are visualized in the Gantt chart as dark diamond shapes. Vertical lines in the chart outline the dependencies of tasks, shown as arrows leading from a predecessor to a successor task. Other infor- mation, such as notes and resources, may be inserted by adding columns. If resources are included, budgets are then calculated by task or subtask and by specific time period. The integrated nature of each task and milestone is readily apparent from the chart.
Once a project is underway, tracking functions of the program allow the PM to compare actual to planned progress. It is also a simple matter to enter proposed changes and determine how any given change or set of changes will impact the overall timeline or any other task. Once a change is made to one task, the hierarchy and schedule are automatically recalculated and the outcome is made immediately obvious on the revised Gantt chart. For example, let us suppose the animals do not arrive on July 15, as scheduled in Task ID #18, but instead arrive 2 weeks later on July 29. By simply changing this date, from July 15 to July 29, it is possible to learn if this will delay any other task or even delay or otherwise impact the project completion date. The project manger is able to demonstrate this to the project team and this
63Project Management
allows the team to consider alternative arrangements or risk mitigation strat- egies. Indeed, PMs often project Gantt charts at team meetings to demon- strate to the whole team exactly how a change or delay might impact the overall sequence of events or schedule. This tool is a very powerful means of communication.
Project management software also assists the PM with preparation of reports, outlining various elements and adding to the report-specific examples and illustrations. In addition to the Gantt chart, these programs are capable of presenting the project or any stage in the project in other illustrative formats, such as scheduling charts. As is the case for much of today’s software, project management software has dozens of other help- ful functions, too numerous to mention here.
PMs use other methods to visualize a project or to present this informa- tion to various individuals or groups, such as the project team, investors, or senior management. Although the Gantt chart may be familiar to members of a project team, it can be foreign to stakeholders or individuals not familiar with project management. How then does the PM present the visual repre- sentation of a complex biotechnology project to an audience that is unfamil- iar with the Gantt format and the project itself? To speak to such an audience, the PM uses simpler formats. These are found in commonly available and easy to use software programs. Examples of two formats, a timeline and a PERT chart, are shown in Figures 2.4 and 2.5, respectively. Notice how these simple formats of the information presented in the Gantt chart (Figure 2.3) allow one to tailor formats to communicate to specific audiences.
Other direct communication tools are available to the PM, and some were mentioned earlier in this chapter. Some are better for a particular purpose or situation than are others, but they all serve the PM and team by communicat- ing the project to those outside the team.
Telephones remain a frequent means of communicating one on one, and teleconference meetings are a common audio tool used for communication by the team. Face-to-face meetings are colocated at one site but, for many teams, meetings in one room are infrequent, occurring quarterly or annually. Today video conferencing is frequently used, and this technology is typically dependable and user friendly. Electronic mail (e-mail) is an excellent means of communication since it is near instantaneous (if the recipients read their messages) and can accommodate a group of any size. It allows individual or group responses. However, it is rather impersonal and is not effective when cross-interactions must be rapid and fruitful and completed rapidly, or when emotion is a critical component, such as resolving a long-standing dis- agreement among team members. Also, it is more difficult to reach decisions or clarify complex technical or scheduling issues by e-mail, and we are all aware of misunderstandings that arise using this method of communication. PMs employ a variety of communication methods, enhancing communica- tion with a mixture of techniques.
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Summary of Project Management in Biotechnology Development
Chapter 4 focuses on project management, the endeavor that pulls together the biotechnology operation, integrating functional areas into an organized whole; this is aimed at achieving a common goal, usually the successful development of a biopharmaceutical product. A project team, managed and led by a PM, is necessary to achieve such lofty and expensive objectives in our industry. A team is composed of individuals representing each tech- nical and administrative aspect of the biotechnology operation. The PM is the individual responsible for organizing, orchestrating, and monitoring the various processes, tasks, or work activities of the team. The PM and his or her authorities and responsibilities must be clearly established by upper management and identified to the team. The PM is responsible for manag- ing the project by ensuring proper planning, stimulating effective commu- nication within the team and to stakeholders outside the team, guiding risk management and problem solving, keeping metrics on progress, or lack thereof, understanding and guiding team dynamics, tracking technical and financial aspects of the project, coordinating with contractors, collaborators, and consultants, and, unfortunately, providing a place for team members and management to vent their frustrations. It is critical from the beginning of a project to follow a written project management plan based on technical plans previously drafted for the project. The PM also creates and manages lists of tasks and schedules, so important to establishing, tracking, and mea- suring a successful project. In each of these endeavors, there is reliance on credibility, trust, knowledge, experience, and, most importantly, on project objectives and plans from the standpoint of the team and of the PM.
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3 Regulatory Affairs
The U.S. Food and Drug Administration: Law and Regulations for Biopharmaceuticals
Historical Basis for FDA Regulation
Food and drug regulation evolved in the twentieth century, which was a reflection of major changes in the way in which foods and drugs were pro- cessed and sold. In the nineteenth century, these products were processed on a small scale. Grain was milled locally, community butchers slaughtered ani- mals and sold meat to neighbors, and local pharmacists and physicians formu- lated and dispensed medications. This changed late in the nineteenth century, notably in the food industry, as large mills and slaughterhouses became a part of the Industrial Revolution. But abuses, like the sale of adulterated foods in some instances, led to social revolt and the desire for governmental regula- tory controls. Upton Sinclair’s book, The Jungle, a revealing look at practices in the meat industry, is thought to have stimulated the U.S. Congress to pass the Pure Food and Drugs Act in 1906 (Sinclair, 1905). For drugs, this act focused on the need to inform the public about foods and drugs through the use of honest labeling. A label reveals the contents of a container and cannot provide false or misleading information in a fraudulent manner. The 1906 Act did not, however, establish the need for review of each product by a federal agency before it could be marketed and sold to the public.
This initiative was to come later, in 1938, when the U.S. Congress passed the Federal Food, Drug, and Cosmetic (FD&C Act or the Act) Act of 1938. This act, passed in the middle of the Great Depression, changed the Food and Drugs Act of 1906 in several key ways. First, it no longer required the government to prove fraud was committed if a drug claimed a curative or therapeutic effect. It also required a premarket drug review of a New Drug Application (NDA), in which the sponsor of a product, the company distributing and selling a drug, provides written evidence that its product was safe and effective. It authorized other government actions as well: the Federal Trade Commission (FTC) was to review drug advertising, promotional claims, and material; the Food and Drug Administration (FDA) would inspect drug manufacturing
66 Biotechnology Operations
facilities and enforce the law and levy fines and punishments and it prohib- ited false therapeutic claims; and it defined classes of regulated products— biologics, medical devices, and cosmetics—as existing under these rules. The FD&C Act of 1938 is the foundation for today’s regulation of drugs, biologics, and medical devices, and it was the basis for establishment of the FDA.
The FD&C Act and other food and drug laws are often responses by the public and government to tragic and avoidable situations. The Biologics Control Act (BCA Act) of 1902 was the result of 10 children contracting teta- nus after taking a poorly made antitoxin. The Cutter Incident of 1955, in which children were exposed to live polio virus from a poorly manufactured vac- cine, was the basis for expansion of biologics regulation. Other amendments often followed problems, abuses, or deficiencies, perceived and real, in drug, biologics, and medical device manufacture, control, evaluation, and market- ing. This trend has continued unabated for 50 years and, unfortunately, it may be horror stories that lead to additional food and drug laws or amend- ments in the future. A few of the many acts and amendments in the past 100 years are provided in Box 3.1.
Many products resulting from biotechnology are considered biologicals or biopharmaceuticals. Two acts of Congress, the BCA Act of 1902 and the Public Health Service Act (PHS Act), established special rules for biological or biopharmaceuticals. Today, many biologicals result from biotechnology endeavors. Through these acts and amendments, Congress has delegated to the FDA the responsibility of ensuring compliance of biopharmaceuticals. This, in turn, has profoundly impacted the means by which many biotech- nology products are developed, manufactured, tested, distributed, and sold in the United States.
Regulatory Organization of the FDA
The FDA is responsible for protecting the public health by assuring the safety, efficacy, and security of human and veterinary drugs, biological products, medical devices, our nation’s food supply, cosmetics, and products that emit radiation. The FDA is also responsible for advancing the public health by helping to speed innovations that make medicines and foods more effective, safer, and more affordable, and helping the public get the accurate, science- based information they need to use medicines and foods to improve health.
As such, the FDA is a regulatory agency responsible for many consumer products used in the United States today, which is a huge task, especially considering the impact these products have on our health and livelihood. Clearly, it would be impossible for the FDA to oversee or individually inspect each product item that is sent to consumers. The FDA also communicates information to various interest groups, pharmaceutical, biological, and med- ical device industries, to those who distribute or prescribe the products, such as pharmacies and physicians, and to consumers. Each interest group has responsibilities for reporting, to some degree. For industry oversight, FDA
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inspections represent but a small and selected fraction of the material that is distributed to the user.
The FDA is an agency within the U.S. Department of Health and Human Services (DHHS) and is composed of various organizational units or offices and seven centers. These are shown in the current organizational chart in Figure 3.1. The responsibilities of centers or offices are listed in Box 3.2. Specific
BOX 3.1 EXAMPLES OF FOOD AND DRUG LAWS FROM 1906 TO 2013
Name and Year Purpose
Pure Food and Drugs Act (1906) Prohibits interstate commerce of adulterated or mislabeled food or drugs.
Food Drug and Cosmetic Act (1938) Provides for safety testing prior to marketing, adequate labeling, appoints FDA responsibility.
Public Health Service Act (1944) Regulation of biological products. Kefauver–Harris Amendment (1962) Requires drugs have proven efficacy. Fair Packaging and Labeling Act (1966)
Honest and informative labeling on consumer products with FDA responsibilities.
Orphan Drug Act (1983) Encourages development of products to treat rare diseases.
Federal Anti-Tampering Act (1983) Makes it a crime to tamper with prepackaged consumer products.
Drug Price Competition and Patent Term Restoration Act (Waxman–Hatch Amendments) (1984)
Drug price and completion and patent restoration. Generic drugs.
Prescription Drug Marketing Act (1987)
Requires licensing of drug wholesalers, bans diversion of drugs.
Prescription Drug User Fee Act (1992) User fees established for FDA review of applications.
FDA Export Reform and Enhancement Act (1996)
Controls for imported and exported products.
FDA Modernization Act (1997) Regulate advertising. Pediatric Research Equity Act (2003) Require clinical research to include pediatrics. Food and Drugs Administration Amendments Act (2007)
Broaden and upgrade drug safety programs.
Biologics Price Competition and Innovation Act of 2009 (BPCI Act) (2010)
Created an abbreviated licensure pathway for biological products that are biosimilar or interchangeable with current FDA approved reference product.
FDA Safety and Innovation Act (FDASIA) (2012)
Promote patient access to new products.
Drug Quality and Service Act (DQSA) (2013)
Identify and trace prescription drugs.
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BOX 3.2 RESPONSIBILITIES OF SELECTED CENTERS AND OFFICES AT THE FDA
Office of the Commissioner
Office of Orphan Products Development (OPD)
Review applications for orphan drug designation
Office of Combination Products (OCP) Identify primary review office for combination products
Office of Regulatory Affairs (ORA) Ensure regulatory infrastructure and enforcement
Center for Biologics Evaluation and Research (CBER) Office of Biostatistics and Epidemiology (OBE)
Statistical review and support
Office of Blood Research and Review (OBRR)
Blood products and device review and research
Office of Vaccines Research and Review (OVRR)
Vaccine products review and research
Office of Compliance and Biologics Quality (OCBQ)
Inspections of biologics facilities
Office of Cellular, Tissue and Gene Therapies (OCTGT)
Review of genetic therapy and cell and tissue products
Center for Drug Evaluation and Research (CDER) Office of Compliance Surveillance, monitor, inspections Office of Surveillance and Epidemiology
Epidemiological review and support of new and approved drugs
Office of Clinical Pharmacology and Biopharmaceutics
Pharmacology review and support
Office of New Drugs Review of IND, NDA, ANDA Office of Nonprescription Drugs Review of OTC drugs Office of Hematology and Oncology Drug Products
Review of drugs to treat or prevent cancers
Office of Pharmaceutical Science Drug development and testing Office of Generic Drugs Review of ANDA applications Office of Biotechnology Products Review of biotechnology products,
therapeutic proteins, and monoclonal antibodies
Office of Testing and Research Analyze drugs, ensure product quality Office of New Drug Quality Assessment
Ensure critical pharmacological attributes, tests
Office of Translational Sciences Statistics, clinical pharmacology Division of Drug Information Public and professional information Office of Medical Policy Review advertising and promotions
(Continued)
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responsibilities for biologicals and biotechnology products are outlined in Box 3.3. Amendments to the FD&C Act typically apply to biologics as well as drugs. Some therapeutic biological products that had been reviewed and regu- lated by Center for Biologics Evaluation and Research (CBER) in the past, such as therapeutic monoclonal antibodies, are now reviewed by FDA’s Center for
BOX 3.2 (Continued) RESPONSIBILITIES OF SELECTED CENTERS AND OFFICES AT THE FDA
Center for Devices and Radiological Health (CDRH)
Office of Compliance Inspections of device manufacturers Office of Device Evaluation Review marketing applications for medical
devices Office of In Vitro Diagnostic Device Evaluation and Safety
Review marketing applications for in vitro diagnostic devices
Center for Veterinary Medicine (CVM)
Office of New Animal Drug Evaluation Review marketing applications for veterinary drugs
BOX 3.3 REVIEW OF BIOTECHNOLOGY PRODUCTS AT THE FDA. RESPONSIBILITIES BY CENTER
Center for Biologics Evaluation and Research (CBER)
Center for Drug Evaluation and Research
(CDER)
Center for Devices and Radiological Health
(CDRH)
• Vaccines • Plasma or serum products • Blood products • In vitro diagnostics for
blood • Gene therapies • Somatic human or animal
cells or tissues • Pluripotent cell-derived
products • Stem cell products • Allergenic materials • Antitoxins, toxoids, and
toxins • Antivenoms • Combination products in
which the biologic is primary mode of action
• Monoclonal antibodies • Therapeutic immune
therapies • Cytokines • Therapeutic proteins
derived by biotechnology
• Enzymes • Interferons • Growth factors • Peptides • Small molecule drugs • Combination product
in which the drug is primary mode of action
• Medical devices of biotechnology origin
• Radiation-emitting devices
• Medical imaging agents • Surgical and therapeutic • Diagnostic and
Radiopharmaceutical • In vitro diagnostics • Combination products
in which the device is primary mode of action
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Drug Evaluation and Review (CDER). Finally, certain aspects of biotechnology product development are regulated by agencies, local, state, national, and inter- national, other than the FDA. These are also reviewed in Chapter 4.
Food and Drug Law, Regulation, and Guidance
Laws enacted by Congress and signed by the President are the basis for FDA functions. Regulations are established requirements, developed by an autho- rized federal agency, to interpret the intent of laws. Regulations interpret the law, considering the intent of Congress when the law was established, and they apply technical, scientific, and administrative best practices to fulfill the law. Regulations cannot be simply mandated by the FDA but must go through a rulemaking process established by the Administrative Procedures Act (APA Act) of 1946. The rulemaking process mandates that a regulatory agency propose to the public every regulation and seriously consider the comments received in response. Hence, rulemaking is a very transparent process with significant influence by citizens and organizations. Once the discussion period is completed, the regulation is then published and goes into effect. Regulations are placed into the Code of Federal Regulations (CFR), bound in numbered volumes by functional area. Food, drug, biologic, and medical device regulations are published in Part 21 of the CFR, hence reference to 21 CFR. Regulations have the impact of law and, if violated, are enforceable by authorized law enforcement agencies (Chapter 4).
Regulations alone may fall short in their ability to fully interpret the law and provide scientific or technical guidance in highly specialized areas. Thus, regulatory agencies interpret regulations through the use of highly technical publications referred to as guidelines. A guideline, unlike a regula- tion, does not carry the weight of law but instead suggests to an interested party the best technical, scientific, or administrative practices that may be considered as a means of achieving the intent of a regulation. The FDA has dozens of guidelines written and updated by scientists working on cutting- edge technologies; they are available to the public. For individuals working in biotechnology, guidelines are extremely important resources because they facilitate targeted planning of product development by identifying scientific, technical, and regulatory processes. This can greatly increase regulatory compliance and prevent loss of time in development.
FDA-Regulated Products
FDA regulates a vast array of products, and these will be described as indi- vidual classes of products along with information about the FDA center that regulates them. An important aspect of the regulatory planning process is
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to understand exactly the nature of a product and to focus regulatory efforts to that area. Jurisdiction for product review and responsibility at the FDA would seem obvious, but, with many biotechnology products, this may not be the case. In developing a regulatory plan, it is extremely important to determine jurisdiction, and this is often based on both the type or class and the intended use of product—information that should be available in the targeted product profile (TPP) (Chapter 1).
Biologics
The PHS Act defines a biologic as any virus, therapeutic serum, toxin, antitoxin, vaccine, blood, blood component or derivative, allergenic product or analogous product…that is intended for use in the diagnosis, cure, mitigation, treatment, or prevention of disease. For many years, this was construed to mean virtually all biotechnology products. However, over time and with the advent of biotechnology, biological products became very diverse. As noted earlier, some biotechnology-derived products, such as monoclonal antibodies, enzymes, cytokines, and simple protein therapeutics, are considered well characterized in that their molecular nature is well known. These well-characterized products are reviewed by CDER. In this book, the term biopharmaceutical refers to prod- ucts under the former definition of a diverse universe composed of biologicals or biologically-derived molecules, cells, tissues, or organisms intended for use in the diagnosis, cure, mitigation, treatment, or prevention of disease. Since a biopharmaceutical may be reviewed either by CBER or CDER, depending in part on nature or indication, it is important to determine for each product the most likely route of development.
CBER is the FDA lead regulatory office for many biologicals, notably the blood and blood-derived products, cellular, tissue, and gene therapies and vaccines. Three offices within CBER (Box 3.2) manage activities and regulate these products.
Office of Blood Research and Review (OBRR) handles blood products, most of which are derived from human whole blood or plasma. Even in a high biotechnology world, human-derived blood and blood products, such as red blood cells, plasma, platelets, and clotting factors, comprise a large industry. In addition, recombinant blood products, such as clotting fac- tors, are approved, and others are in development. Human-derived blood products are highly regulated from their sources (blood and plasma donor centers) to the finished product; this is partly because of the risk that such products could contain adventitious agents. Blood establishments must be registered, and they are routinely inspected by the FDA. Blood products are rigorously tested for a wide variety of infectious agents. To ensure the safety of the blood supply, materials such as blood bags and infusion lines used in blood collection, handling, and testing are also regulated by OBRR, CBER. Blood products used for further manufacture, such as platelets or plasma,
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are subjected to rigorous methods to remove and inactivate viruses such as hepatitis or human immunodeficiency viruses.
The Office of Cellular, Tissue and Gene Therapies (OCTGT) regulates cel- lular and gene therapies and human cell and tissue products. Most of these products are derived from biotechnology, whereas others directly apply bio- technology methods. Many are live microbial products, such as viral vectors that have been engineered to carry a therapeutic gene. Others are macromol- ecules, such as plasmid DNA, intended to have a therapeutic effect upon cell entry.
Somatic tissues or cells, such as allogeneic skin grown in culture to replace that of a burn patient, or bone marrow-derived cells, selected for a trait and expanded in culture, are reviewed by OCTGT as well.
Human organs, such as kidneys, lungs, or livers, intended for whole organ transplantation, are not regulated by the FDA. However, the agency does regulate human cells or tissues, or products made from cells or tis- sues, intended for infusion, implantation, transfer, or transplantation. The agency also regulates animal whole organs such as pig skin or liver that might be transplanted into humans. The primary reason for the regulation of these xenogeneic products is that they are manipulated and are likely to have a systemic effect in terms of safety. Similarly, although whether a cel- lular product is autologous or allogeneic is important, again the level and type of manipulation and potential distribution in the human body are of primary importance and influence how these products are similarly regu- lated. The safety of minimally manipulated blood and tissue products is primarily based on registration of donor centers, careful selection of donors, and testing of the donated cells or tissues. Current Good Tissue Practices (CGTP) require special handling of cells or tissues, and any added materi- als, such as supplements, that are ultimately considered a part of that prod- uct. Human organs, such as kidneys, livers, or lungs, intended for whole organ transplantation, are not regulated by the FDA unless this activity is part of human clinical research.
The FDA has recently proffered guidelines for pluripotent (or stem cell- derived) products that are now used in a number of early clinical trials. This area of exploration is in its infancy with several pluripotent cells studies initi- ated and several hundred studies evaluating stem cells. Stem cell technolo- gies will continue to mature as early development pathways demonstrate that safe and effective treatments can be developed from pluripotent cells realizing their great potential in regenerative medicine and as therapeu- tic products. For example, the FDA has made clear that nonclinical animal studies are critical to understanding possible risks. Nonclinical testing must be designed based on the source of the cell, the intended use and route of delivery, and data from laboratory studies. The cells themselves must be fully characterized and major issues, some of which are listed next, must be defined in the laboratory and in animal studies.
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• Mechanisms of action, physiological parameters • Distribution and persistence in the body or migration to tissues
other than the target • Original function in the tissue or organ of origin versus intended
therapeutic function • Differentiation potential • Ectopic growth potential • Tumor, benign or malignant, formation
Interestingly, even though these issues may seem unique to stem cells, they are in general the same questions posed by regulatory agencies to sponsors of a wide variety of biotechnology products.
The Office of Vaccine Research and Review (OVRR) reviews biological products that are intended to protect from or cure disease via an immuno- logical affect. Hence, virtually all vaccines and allergenics, with the possible exception of cancer vaccines, are under the purview of OVRR. Vaccines, like genetic therapeutics, represent a wide variety of biotechnologies, too numer- ous to mention here. All products reviewed by OVRR have in common an intended mode of action: to elicit an immune response. The indication may be therapeutic, or it might be preventative. Product types come to OVRR in a vast array of technologies, and today most are derived from biotechnol- ogy; only a very few candidate vaccines, such as influenza, remain as natural products. Live bacterial or viral vectors, such as Escherichia coli or adenovi- rus, are used both to stimulate an immune response to that organism or as carriers intended to stimulated immune responses to other proteins geneti- cally engineered into the host. Vaccines may be given by oral, intranasal, intramuscular, epidermal, or other routes. Vaccines today are often part- nered with a delivery device, making a combination product. Most biotech- nology products engineered and intended to stimulate an immune response goes to OVRR for review. Vaccine adjuvants, molecules intended to improve immune response when given with an antigen, are also reviewed. Allergenic products, often natural substances purified and then used to treat allergies or in hypersensitivity testing, are products reviewed by OVRR as well.
CBER also reviews unique types of biological products. Exact regulatory pathways have not been developed for every type of product that might be conceived by the biotechnology industry. It is worth noting that one class of biologics, those derived from plants, are also reviewed by CBER. Drugs and biologicals may be derived from bioengineered and selectively bred plants, and these may be reviewed by CBER or by Center for Drug Evaluation and Research (CDER) or by both centers. One such drug is an antimalarial drug, which was in short supply worldwide in 2011. In this situation, the product receives review with special attention paid to the recombinant nature of the plant from which the product, not itself a recombinant molecule, is derived.
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The basis for regulation and review may be based largely on the indication and manner in which the product will be used and less on the nature, drug or biological, of the product.
All centers and offices within the FDA and the drug regulatory agencies of other developed countries play a significant role in providing support for therapeutic products and vaccines used in global health, even if those products will not be licensed in the United States. This is due in part to the FDA’s leading international reputation for scientific product review and also because many target countries cannot afford to have in their governments a food and drug authority. When a biopharmaceutical firm applies to test a product indicated for global health in the United States, it will be reviewed by the FDA and comment will be provided. Indeed, most global health prod- ucts are first tested in the United States or another developed country before being fielded in developing countries, where it is necessary to conduct field trials and it is desirable to manufacture commercial product. Hence, the FDA, along with institutes and academic centers in the United States and other Western countries, provides early product development support and advice that can be transferred to target countries.
Drugs
Drugs are broadly defined as products used to mitigate, treat, or prevent disease in man and that affect physiology or anatomy of the human body. The industry standard for a drug is a small or large molecule that has a well- defined chemical structure. Following market approval, drugs are recognized as such in reference texts, such as the United States Pharmacopoeia (USP) or the National Formulary (NF) (Chapter 7). The USP and NF are published by a gov- ernment- charted laboratory, the United States Pharmacopeia. Pharmaceutical is synonymous with the word drug. The historical definition of drug is of a small molecule, or ethical, pharmaceutical, a compound that is synthesized from nonbiological sources. Examples of drugs under this definition are acet- aminophen or aspirin, antibiotics such as penicillin, and the anesthetic ether. Drugs of this nature are under the purview of the FDA’s CDER (Box 3.2). Certain molecules of biological derivation, such as monoclonal antibodies and therapeutic proteins, are reviewed under CDER even though they do not meet the historical definition of drug. Prescription drugs or biopharmaceuticals are those products that must be prescribed by a licensed medical professional and dispensed by a licensed pharmacist. Over-the-counter (OTC) drugs are those that do not require a prescription. Most biopharmaceuticals are dispensed by prescription.
Generic drugs are drugs that no longer have marketing exclusivity due to patent protection but are still regulated under drug regulations. Generics must be tested in adequate clinical studies, albeit abbreviated, and they must receive marketing approval from the FDA under the Abbreviated New Drug
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Application (ANDA). The Waxman–Hatch Act of 1984 was instrumental in establishing regulations that guide marketing approval of generic drugs.
CDER, in addition to reviewing small molecule drugs or pharmaceuticals, also takes the lead for review of a number of biopharmaceuticals. Examples are shown in Box 3.3, but the list is growing as new biopharmaceuticals enter clinical trials or receive market approval. Recombinant therapeutic proteins are macromolecules or peptide products intended to treat disease. Responsibility for review or co-review within CDER may also be deter- mined by the indication or intended use. This is because CDER is also orga- nized by disease area. Hence, a therapeutic recombinant protein intended to treat gastrointestinal cancer might be reviewed by the oncology group, specifically by the Division of Biologic Oncology Products (DBOP) group. A monoclonal antibody protein directed against an infectious bacterium of the gastrointestinal tract could receive primary review from the anti-infec- tive group and consulting review by the gastroenterology group. Another example of a biopharmaceutical product falling under CDER is monoclonal antibodies. They represent a class of molecules that are produced in vitro as a result of genetic engineering. Monoclonal antibodies are produced to react with a variety of target proteins and to have a therapeutic effect on the patient. Examples are antibodies directed against inflammatory cytokines; these are engineered to stop undesirable inflammation due to autoimmune diseases. They are produced in genetically engineered, immortalized, and cloned cells in much the same way as other recombinant proteins. Today we understand the molecular structure of most monoclonal antibodies. Therapeutic monoclonal antibodies are reviewed by CDER with primary responsibility resting in its Office of Biotechnology Products (OBP), but may also be reviewed by experts in the disease. In the case of the mono- clonal antibody directed against an inflammatory cytokine, rheumatolo- gists employed in another division of CDER might work closely with OBP on review of regulatory documents. In conclusion, the division of review responsibility for a given biopharmaceutical may, in CDER, be more diffuse than it is for one at CBER.
Whichever division or office within CDER or CBER has primary respon- sibility for review of a given product, there might also be specialists from another center involved in the review of that product as well. There are specialists within each of the centers for functional areas such as nonclini- cal studies and toxicology, clinical trials, manufacturing, and quality con- trol. Hence, review of applications by any FDA center represents a team of experts, often from a variety of offices or centers.
Medical Devices
This major class of products includes instruments, prostheses, delivery tech- nologies, in vitro diagnostic tests, implants, apparatus, and a host of other engineered yet nondrug and nonbiologic items. Regulatory responsibility for
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medical devices rests with FDA’s Center for Devices and Radiological Health (CDRH). Examples of medical devices range from tongue depressors to cardiac pacemakers, from dental amalgams to CAT scanners, and from devices and software to transmit X-rays to HIV test kits. Unlike drugs and biologics, medi- cal devices are classified, a priori, based on their risk to the user and the level of regulatory control, review, and safety concern. Under an established hierarchy, medical devices with the greatest risk to the user are given the highest classifi- cation. A tongue depressor is Class I, General Controls, a test for the common flu virus is Class II, General and Special Controls, and a heart–lung machine is Class III, Special Controls and Premarket Clearance or Premarket Approval (PMA). In general, the FD&C Act requires safety testing for all devices, but does not require testing for efficacy studies of Class I and Class II medical devices. Most Class III devices need some assurance of adequate performance. Many Class II devices are eligible for regulatory approval under a regulation referred to as 510(k) approval if they are substantially equivalent to a predicate device. Manufacturers find this an attractive rapid means of seeking regulatory approval from CDRH, but it only applies to certain devices. There are many other differences in the regulatory requirements and review and structure of processes leading to market application and approval for devices as compared to drugs and biologics. Although few biotechnology products are considered medical devices, some are a combination of a device and a drug or biologic.
Combination Products
Many biopharmaceutical products are not simply classified as biologics, drugs, or medical devices, but instead are a mixture of these product classes. For example, a drug aimed at a blood cancer cell, historically a drug, may be attached to a protein, a biologic, that targets the cancerous cell. A recombi- nant vaccine protein, a biologic, may be delivered to the skin using a novel jet injector, a medical device. The combinations and permutations seem limit- less, with new biotechnology combination products appearing daily.
The FDA accommodates these so-called combination products through the Office of Combination Products (OCP) responsible for defining jurisdiction and ensuring a coordinated review of applications. Jurisdiction is based on the primary, or most important, mode of action for the product. It is critical to identify and then to evaluate every potential combination product early in the planning process so as to establish the most likely regulatory path forward. The first step is to determine if a product is, in fact, a combination product in the eyes of the FDA. Referring to examples cited earlier, the drug would provide the primary mode of action because it would kill the cancer cells, and CDER is likely the lead for review. The vaccine protein would be primary for that example because it elicits and thus provides the desired outcome: protec- tive immunity. Although a paradigm such as the one shown in Figure 3.2 helps a sponsor to identify the primary review office, consultation with the FDA, through a formal Request for Designation, is recommended.
78 Biotechnology Operations
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Other Classes of Biotechnology Products and Their Review at the FDA
Products for Veterinary Use
Biologicals, including biotechnology products used to treat or prevent disease in domestic animals, are largely regulated by the U.S. Department of Agriculture (USDA) and not by the FDA. However, the Center for Veterinary Medicine (CVM) of the FDA is designated for animal drugs and medicated feeds. CVM reviews marketing applications, ensuring such products are safe and effective for their intended use. The process is similar to that applied to human drugs, and includes the need to file an Investigational New Animal Drug (INAD) exemption to support investigational use or a New Animal Drug Application (NADA) to receive product-marketing approval. Animal feeds, both those that contain medications, such as antibiotics, and those without supplements, are under the purview of CVM, because they should not contain harmful sub- stances and must be properly labeled. Biotechnology-derived products could fall under both the medicated and nonmedicated classifications of animal feeds, and these products must receive marketing approval from CVM.
Cosmetics, Food, Dietary Supplements, Homeopathic, or Nutritional Products
Regulations do exist for cosmetics, products that are applied externally, but have no claim of therapeutic value. Cosmetic regulations, as compared to drug rules, are relatively simple, yet they ensure these products are safe and not adulterated. A biotechnology-derived product could, by intended use, be considered a cosmetic and, if so, cosmetics regulations would apply.
Products that are nutritional in nature and for which no claims are made of a therapeutic or medicinal effect are regarded as a food by the FDA. The FDA regulates most food products, with the exception of meat and a few other animal-derived items that fall largely under the U.S. Department of Agriculture’s purview. Additives for food and food colors do fall under the purview of the FDA. Foods and additives are not considered in this chapter.
Dietary ingredients and dietary supplements are not subject to premarket approval by the FDA, but the agency must be notified of marketing intention nonetheless. These products are regulated by the FDA based on their nature, but largely on the indication and the therapeutic or preventative claims made by the sponsor. Classification of such products requires experience as the complex guidelines are based on nutritional or medicinal claims made by the sponsor, which themselves are often difficult to parse or comprehend.
Dietary supplements and nutritional products are now generated by bio- technology, although the vast majority are still naturally derived or chemi- cally synthesized. Dietary supplements, regulated largely as food and not as drugs, are products taken by the mouth and intended to supplement the diet and not intended to treat, diagnose, cure, or alleviate the effects of disease. Again, the claim made on the product label is instrumental in determining the level
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of product regulation and the designated regulations and review process at the FDA. If the intended probiotic use is for therapeutic purposes, then the associated applicable regulation under the FDA would change accordingly. This shift in probiotic use is an example of regulatory evolution, that is how historic regulation is applied to novel or modified products that will now provide therapeutic benefit if given in accordance with its intended use. An example of this, a traditional food not marketed as a food supplement with claimed medical benefit and as a human therapeutic, is briefly described in Box 3.4. Certain homeopathic compounds may fall under similar rules and
BOX 3.4 NONTRADITIONAL USE OF FOOD SUPPLEMENTS AS A THERAPEUTIC
• Probiotics are microorganisms that have been traditionally used to fortify the gut microbiota. They are regulated by the FDA as a food supplement. As fortifiers, many probiotics have been given Generally Recognized As Safe (GRAS) status by the FDA.
• The potential health benefit and ability of probiotics to prevent or cure disease have been gaining scientific traction over the past decade; a cause–effect relationship continues to be sought that includes clues of strengthening the immune system.
• If the intended use of a probiotic being provided to people is to study, diagnose, cure, mitigate, treat, or prevent a disease, then the regulation of a probiotic is as a therapeutic and falls squarely within the purview of the medicinal branch of the FDA.
• Likewise, a study to evaluate the ability of bacterial coloniza- tion to treat or prevent disease in patients with an immune dis- order must be performed under an IND application submitted to the CBER at the FDA.
• As a therapeutic, both safety and efficacy will be carefully eval- uated by the FDA to minimize human risk and hopefully dem- onstrate human benefit outweighs potential risks of human exposure.
• As the increased use of probiotics continues to be investigated for their ability to promote a positive host response to live organisms (e.g., bacteria, fungi, or virus), the FDA issued a draft guidance to assist with the development of these prod- ucts intended for therapeutic purposes (Guidance for Industry: Complementary and Alternative Medicine Products and Their Regulation by the Food and Drug Administration, December 2006) (FDA, 2006).
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have their own set of definitions, depending on claims made on therapeutic value. Certain homeopathic compounds are formulated in pharmacies for individual patients, a process that is legal in the United States under spe- cific circumstances. As with nutritional supplements, the indication deter- mines the level of regulation, as pharmacies typically fall under the laws of individual states. With homeopathic medicines, there is a point at which the distribution or volume of sales can conflict with this definition. Then the FDA regulations may be brought to bear, and the product may be declared a drug, biological, or device. Few biotechnology products produced for thera- peutic purposes would be considered homeopathic medicines or nutritional products.
FDA Regulatory Information and Resources: Regulatory Intelligence
This section focuses on the functions of FDA regulatory intelligence and a biotechnology firm’s need to continually obtain information that allows an understanding of the regulatory environment as it applies to the develop- ment of a product. Also, a prudent biotechnology firm will have in place processes to abstract and communicate FDA regulatory information to operational staff and upper management. Sources of information on non- FDA regulatory bodies are described in Chapter 4.
Regulatory intelligence is the process of finding and analyzing publically available regulatory information. It is not necessary, nor is it possible, to know all regulations that apply to every biotechnology product; it is more important to understand the overall environment and possible sources of regulatory information and to be able to locate information on particular types of products.
Gaining regulatory intelligence is always a first step in preparing a regu- latory plan. To find pertinent regulatory information on a particular prod- uct or similar products, it is necessary to first understand the product and indication, as provided in a TPP; then a structured search is initiated. The search begins as a broad investigation to understanding the regulatory environment using general and specific sources. This provides a general background on the subject. It then focuses on areas of interest, moving from one regulatory source to another until it seems that all regulatory databases have been exhausted. Scientific databases list additional product informa- tion, and these are examined. The result is a bibliography that provides a regulatory history and scientific background of the product and product class, an exact idea of the current regulatory environment, and insights into future regulatory initiatives. Taken together, this background information
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serves as a foundation for preparing the regulatory plan and a basis for later updated information on the product and its regulation.
Today, public regulatory information, with the exception of regulatory textbooks and some international guidelines, is available on the Internet at no cost to the user. Hence, the key to finding the information is to use a library of websites for searches and to apply search engines. Some searchers use regulatory information blogs to network with colleagues, an approach that is particularly helpful for locating very specific bits of information. Also, before beginning a search, it is a good idea to develop a method to catalog the information so that it is easily retrieved for review and reference. Most regulatory libraries are electronic databases. Simple systems, such as using office support software with desktop search capabilities, are fine for smaller databases, although more complex and dedicated software is avail- able for regulatory libraries consisting of thousands of references.
National regulatory agency websites, such as http://www.fda.gov, and international or harmonization sites, such as http://www.ich.org, are a wealth of information on FDA regulations and international standards, sometimes providing far too many resources outside the intended scope. Compendia methods, standards, and monographs can be readily obtained from U.S. Pharmacopeia National Formulary (USPNF) via hard copy or is assessable online at http://www.usp.org. Scientific literature through PubMed or a gov- ernment and institution or university library search engine provide jour- nal articles of a regulatory or technical nature. Professional and industry or trade journals post articles on various regulatory topics and recent articles offer insights into recent or hot topics.
Review and careful evaluation of brochures or dossiers of similar cur- rently marketed products may prove to be extremely valuable in under- standing the precedence for this type of product. Valuable product-specific details of priority nature such as methods or manufacturing components may be obtained from vigilant searches of the public access U.S. Patent and Trademark Office (http://www.uspto.gov) dbase. A patent search is likely to provide critical information regarding freedom to operate and market- ing or potential current market exclusivity.
Regulatory intelligence does not end with an initial search but contin- ues through the product lifecycle. Information is obtained through peri- odic searches of websites, by using commercial regulatory intelligence software, and through e-mail alerts, many provided by various govern- ment agencies including the FDA, by using blogs and through diligent personal networking, by e-mail, from professional meetings, and by networking with other professionals. Some long-standing and helpful U.S. sources of intelligence on biopharmaceuticals are given in Box 3.5, although other sources, notably international guidelines, are described in Chapter 4.
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BOX 3.5 WEB-BASED SOURCES OF U.S. REGULATORY INFORMATION
U.S. Government Regulatory Trade Organizations
Food and Drug Administration
http://www.fda.gov Regulatory Affairs Professional Society (RAPS)
http://www.raps.org
National Institutes of Health
http://www.nih.gov Drug Information Association (DIA)
http://www. diahome.org
U.S. Pharmacopoeia
http://www.usp.org Food and Drug Law Institute (FDLI)
http://www.fdli.org
Clinical Trials Registry
http://www. clinicaltrials.gov
American Association of Pharmaceutical Scientists (AAPS)
http://www. aapspharmaceutica. org
PubMed (National Library of Medicine)
http://www.ncbi.nlm. nih.gov/pubmed
Pharmaceutical Education and Research Organization (PERI)
http://www.peri.org
California Institute for Regenerative Medicine (CIRM)
http://www.cirm.ca.gov International Conference on Harmonization (ICH)
http://www.ich.org
Medical Dictionary for Regulatory Activities
http://www.meddra. org
World Health Organization (WHO)
http://www.who.int
FDA News and Announcements
http://www. GXPnews.com
Regulatory Newsletters
Parenteral Drug Association (PDA)
http://www.pda.org
http://www. fdcreports.com
http://www.foi.com Biotechnology Organization (BIO)
http://www.bio.org
http://www.foi. com
http://www.bioworld. com
Pharmaceuticals for Practitioners
http://www. pharmaportal.com
http://www. fdanews.com
http://www. fdaadvisorycommittee. com
Association of Clinical Research Professionals (ACRP)
http://www.acrpnet. org
The Organization for Professionals in Regulatory Affairs
http://www.topra. org
Applied Clinical Trials
http://www. appliedclinical trialsonline.com
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Regulatory Operations for FDA Applications
Regulatory Planning and the Regulatory Environment
Biotechnology endeavors are influenced in many ways by regulations and the regulatory environment. To succeed, the biotechnology professional must understand the regulatory landscape lying ahead before embarking on product development. This is achieved by preparing a regulatory plan, an early and important part of any product development strategy (PDS). Benefits are obvious. It builds a framework for the overall operational plan and serves as a foundation on which many aspects of nonclinical, clinical, manufactur- ing, and control planning may be built. A regulatory plan identifies poten- tial regulatory hurdles through inspection of product-specific regulations and regulatory agencies. It allows a biotechnology firm to communicate two messages to outside parties, investors, or potential partners: they will be successful, in part, because they have identified regulatory issues specific to their product and they intend and plan to address those matters. A well- considered regulatory plan goes beyond reacting to the environment and also identifies regulatory opportunities such as unmet needs or accelerated pathways to enhance product value. Indeed, a well researched and reasoned regulatory plan seeds new ideas into a biotechnology firm’s business plan.
Novel biotechnology products often sail into uncharted regulatory waters. There are new diseases or unique indications for novel diseases, product lines marketed never before, and radically new technology approaches to unsolved problems. As noted earlier, the breadth of biotechnology products is enormous, meaning that regulatory requirements cut a very wide and deep swath. The regulatory plan for a given product may need to cover a number of diverse areas and also delve into each of these areas, perhaps setting precedent in one or more areas. Elements of a regulatory plan are given in Box 3.6 and discussed further in Chapter 1. In biotechnology, the regulatory planner is challenged from many directions. Even if regulatory pathways are clear and simple with much precedent, a well-conceived regu- latory plan adds product value by demonstrating how the product may be a regulatory slam-dunk. The time and effort involved in preparing a regula- tory plan are justified for any biotechnology product with reward derived from a well-planned and efficient regulatory approval process.
Risk Versus Benefit
The FDA is chartered to protect the public health. Hence, at the heart of regulatory judgment for any given product is the need to weigh risk versus benefit, as it relates to individual and public health. This can be challenging for a biotechnology firm, especially the small or virtual operation, to appre- ciate. Nonetheless, it is critical at some point to think like the regulators and
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BOX 3.6 ELEMENTS OF A REGULATORY PLAN
• The product
• Characteristics • Class • Competitor or equivalent • Indications: primary and secondary • Limitations in safety or effectiveness, stability, shipping/
handling, scaling, price competitive, and population • Special safety considerations • Product blemishes
• The regulatory environment
• Scope of applicable regulations: Global, national, state, and local
• Purposes for regulations: Medical, agricultural, environ- mental, and safety
• Regulatory agencies or guidelines: FDA, USDA, ICH, European, Japanese, and rest of world
• Laws, regulations, and guidelines • Precedence for product class or competitor products
• Global strategy
• Global market targets • Sequence or timing of applications • Exclusivity by market • Application and approval methods by market or country,
ex-U.S. • Influences on regulatory environment
• Political: U.S. and ex-U.S. • Social: U.S. and ex-U.S. • Advisory committees • Local or state authorities • Special interest groups
• Regulatory communication
• Formal applications: IND, NDA, BLA • Written communication: Letters, amendments
(Continued)
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consider the risk-to-benefit balance that a product must bring to the market. Any sponsor has surely considered the benefit that his or her product might provide to mankind. In fact, the benefits of any biotechnology product have most probably been touted to the world, in an effort to garner financial and public support, during research in early development phases. But it is not typical, certainly not in the early stages, of biotechnology development, to admit that a product also carries risks; it is counterintuitive to the entrepre- neurial environment to delve deeply into the possibility of product-associ- ated risk. Traditionally it is not good business practice to highlight potential product risks to the public. However, the FDA must, by law, consider the possibility that any product can cause harm, even that the risks might out- weigh the benefits. Regulatory agencies look at any product as possessing possible risk as well as benefit and they have an obligation, as part of their mission to public health, to evaluate and, sometimes, to publicize product risks. This is a reason some biotechnology firms are asked by the FDA to place a black box warning onto the labels of a marketed product. These dif- ferent ways of thinking create a tension between biotechnology firms and regulatory agencies.
In developing a regulatory plan throughout the development cycle, it falls on regulatory and quality professionals to carefully weigh product benefits against risks and pass the results of this analysis along to members of the product development team. This is done so that risks can be addressed before they become an issue with regulatory authorities or, worse, cause harm to public health. Regulatory risk identification and management begins early
BOX 3.6 (Continued) ELEMENTS OF A REGULATORY PLAN
• Meetings or teleconferences • Public conferences • Timing of each major communication
• Options for special designations and pathways
• Accelerated approval • Emergency use and treatment • Orphan product designation • New drug product exclusivity • Fast track designation • Breakthrough therapy • Priority review • Rolling review
• Regulatory risks
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in the development cycle, as a formal and broader process of product and project risk analysis and management, as discussed in Chapters 1 and 5. Here, we point out the simple fact that regulatory agencies perceive poten- tial risks early in a product’s life cycle. If risks are not recognized as such by the sponsor, then the FDA may demand that they be both recognized and mitigated. This process may go on through the entire cycle of product devel- opment. Failure to heed warnings by the FDA on risk can lead to regulatory action by the FDA. The regulatory professional is often the biotechnology team member to elicit support of product development team members to identify and manage technical risks so that they do not later become prod- uct or regulatory risks.
If after a careful evaluation, it is determined that a product’s risks out- weigh benefits and these risks cannot be effectively mitigated, then a biotech firm is encouraged to withdraw or fail the product early in the development process to ultimately minimize losses and avoid potential harm to people.
Applications Seeking FDA Investigational Use or Marketing Approval
The development life cycle for a biopharmaceutical is heavily influenced by regulatory requirements, hurdles some might say. These requirements are considered met only after a biotechnology firm has communicated scientific plans or results to the FDA. Indeed, the regulatory process is a dialogue between the agency and the sponsor. The sponsor, a legal entity responsible for the product, may be an individual, but, in most cases, it is a corporation or an institute. A sponsor such as a biotechnology firm has one individual responsible for signing regulatory documents, and this person is the designated sponsor’s representative. During the develop- ment life cycle, a dialog between the FDA and the sponsor’s representative is carried on through a series of meetings, teleconferences, and written documents. These steps in communication with the FDA can and should represent important milestones in meeting objectives in the overall prod- uct development cycle. The overall sponsor–FDA communication process, shown in Figure 3.3, is much the same for drugs and biologics even though the nomenclature differs in some respects. Certain aspects of medical device development, including major documents, can differ significantly from drugs or biologics.
The FDA regulatory process begins with a designation by the sponsor that a candidate product will be used in humans for the treatment or prevention of a disease or disorder. The process of developing a TPP (Chapter 1) is criti- cally important in part because the sponsor clearly identifies the intended use, indication, and nature of the product. Consideration as a medical device, a drug, a biologic, or combination product commits to a regulatory pathway that has been established by precedent and regulation or guidelines. Thus, the TPP process is a critical first step in the development of a PDS and for regulatory planning.
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Biotechnology research laboratory:
Potential product
Intended for human use?
Drug or biologic?
Nature of product and intended use (indication)
Veterinary or laboratory use
Target product profile and product development
strategy
Briefing for pre-IND meeting
Phase 1 studies
Phase 2 studies
Pre-IND meeting with FDA
Phase 3 studies
Yes
No
Complete technical requirements, prepare IND
Pre-phase 2 meeting with FDA (optional)
Pre-phase 3 meeting with FDA
Prepare BLA or NDA
File BLA or NDA
Pre-BLA meeting with FDA
Correspondence and meetings with FDA
BLA or NDA information filings and reports
File IND
Phase 1 study report, annual reports
Phase 2 study report and annual reports
Reports and amendments Phase 4 studies,
promotional review and reports
REMS
Correspondence and meetings with FDA
FDA concurrence
Marketing approval
FIGURE 3.3 Regulatory activities and milestones in biopharmaceutical development.
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Investigational Use Applications. The Investigational New Drug Application
The Investigational New Drug (IND) application is a request to the FDA to perform human safety and effectiveness studies on a biopharmaceutical or drug. The IND application often represents the first legally binding docu- ment that is submitted by a sponsor to the FDA. However, the IND filing at the FDA is often preceded by less formal means of communication, such as a pre-IND meeting or teleconference. A new drug is a construct or mol- ecule that has an ingredient or combination of ingredients that have not yet received marketing approval. A product is considered investigational, although being evaluated for safety and efficacy under an IND. Indeed, even a compound with a slight molecular variation from an approved com- pound may necessitate filing an IND with the FDA to investigate safety and efficacy. Products approved for an indication are also considered a new drug if they are to be used (investigated) outside the approved label claim or used for another indication. The investigational use of a new drug is the use of that new drug in a human clinical study or studies by or on behalf of a sponsor (Chapter 9). The IND process focuses on submission, by the sponsor to the FDA, of a clinical protocol, a written plan to test that drug in the first clinical study ( first time in man) and the scientific data and background information related to that drug. The format for an IND can be found at http://www.fda.gov. Many other countries have an investiga- tional process and associated applications, but the process and format vary across borders.
In the Unites States, the sponsor and FDA are both responsible to the public for ensuring that a new drug will not present unnecessary risks to those receiving it and that any risks are balanced with potential benefits. The contents of an IND, outlined in Box 3.7, are standard for any IND, and the sponsor is responsible for generating this application and sub- mitting it to the FDA. In doing so, the sponsor must attest to the validity of data and background information provided to the FDA by completing and signing a form (FDA Form 1571). The information provided in an IND is critically important to the FDA, as this represents the basis for their review and determination of risk versus benefit assessment. As shown in the IND format (Box 3.7), the information comes from a variety of sources, which are described in Chapters 1,4 through 6,8, and 9. Of considerable importance is Section 6 of the IND, providing the Clinical Protocol and other clinical investigation documents, which are described in Chapter 9. The nonclinical information (described in Chapter 8) and any laboratory studies related to laboratory studies, such as biodistribution, are provided in Section 8 of the IND. Previous human use of this or closely related products are communicated in Section 9 of the IND. Section 7 of the IND is reserved for chemistry, manufacturing, and control of the product (described in Chapters 6 and 7).
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BOX 3.7 CONTENTS OF AN IND
• Section A(1)*: Cover page is Form FDA 1571 • The sponsor’s statement to certain questions about the
product and responsible individuals is placed onto a stan- dard form provided by the FDA. Signature sponsor’s repre- sentative. Delegation of responsibilities.
• Section B(2): Table of contents. • Section C(3): Introductory statement and general investiga-
tional plan. • A review of the background, perhaps noting the research
basis for the product, information on the disease or condi- tion, and an explanation of why the product was chosen to treat or prevent the indication.
• Section (4): Reserved for FDA requests. • Section D(5): Investigator’s Brochure (IB).
• IB is a summary of critical information found in the IND, and is written by the sponsor to fully inform the clinical investigators and staff. Not required for single center spon- sor–investigator IND.
• Section E(6): Clinical protocols • The protocol describes in detail the nature of the study
and all procedures that will be taken during the clinical trial. Supportive documents include the Form FDA 1572, a signed statement of the principal investigator, clinical pro- tocol synopsis, CVs of clinical staff, sample consent form, and other documents related to the clinical investigation.
• Section F(7): Chemistry, manufacturing, and control information • Here the sponsor describes the manufacture of the product
and provides detailed information on the raw materials, process, and facilities. Quality control assays are identified and certificates of analysis presented. An environmental statement is required.
• Section G(8): Pharmacology and toxicology information • All nonclinical studies are described with full study reports.
All nonclinical or preclinical studies that are related to (Continued)
* The FDA has used two systems, one numbering (1–10) and the other lettering (A–I), to identify the elements of an IND.
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Common Technical Document
The IND format continues to be replaced by a more universally accepted investigational application format, the Common Technical Document (CTD), developed by the International Conference on Harmonization (ICH). The goal of this harmonized format is to reduce or avoid duplicative testing during the drug development process and eliminate unnecessary delays in the global development of regulated products. The contents of the IND and CTD are the same, but the reporting format or sectional structure differs, as presented in Figure 3.4; additional details and more specific information on CTD can be found at http://www.fda.gov. Rather than the previous nonstan- dard format of 10 sections that make up an IND application or 20 sections of a market application, the CTD format has five modules, irrespective of the type of application. Of the five modules, only four of them (Modules 2–4) are actually universal to all regions and recognized as part of the CTD. Module 1 is region specific, intended for administrative purposes. For example, a CTD submitted to the FDA includes in Section 1 a cover letter, Form FDA 1571, and a Table of Contents required by the FDA. Together with four subsequent modules—quality, safety, efficacy, and multidisciplinary—the modules facilitate ease of navigation and accommodate for regional differences in
BOX 3.7 (Continued) CONTENTS OF AN IND
safety or toxicology and key studies that demonstrate potential efficacy, such as pharmacokinetic and pharmaco- dynamic studies, are included.
• Section H(9): Previous human experience with the investiga- tional drug. • Descriptions and references to all previous uses of this or
closely related products with special attention to safety issues in humans.
• Section I(10): Additional information. • Certification of compliance (Form FDA 3674), summary of
FDA communications, letters granting permission to refer- ence Master Files or other INDs
• References • A list of scientific publications cited in the text
• Appendices • Large documents, such as nonclinical toxicology reports
or pertinent scientific publications, may be included if they have been abstracted or cited in an earlier section.
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submission requirements for several geographic areas to include the content and format of technical data to be accepted by the United States, Canada, Europe, and Japan. Again, the CTD format applies to applications for all investigational stages as well as those to support approval and marketing.
Electronic Submission of a CTD
The CTD format provides a rigid structure or platform to accommodate an electronic submission, as the electronic CTD (eCTD) format. The content of
Module 1
Regional administrative information
(Not part of the CDT)
Module 3
Quality
Module 4
Nonclinical study reports
Module 5
Clinical study reports
Module 2
Common technical document summaries
2.1 CTD table of contents 2.2 CTD introduction 2.3 Quality overall summary 2.4 Nonclinical overview 2.5 Clinical overview 2.6 Nonclinical written and tabulated summary
Pharmacology Pharmacokinetics Toxicology
2.7 Clinical summary Biopharmaceutics and associated analytical methods Clinical pharmacology studies Clinical efficacy Clinical safety Synopsis of individual studies
Administrative information and prescribing information 1.1 Table of contents of the submission including module 1 1.2 Documents specific to each region
(e.g., application forms, prescribing information)
3.1 Module 3 table of contents 3.2 Body of data 3.3 Literature references
4.1 Module 4 table of contents 4.2 Study reports 4.3 Literature references
5.1 Module 5 table of contents 5.2 Tabular listing of all clinical studies 5.3 Clinical study reports 5.4 Literature references
FIGURE 3.4 Common technical document (CTD) structure.
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an eCTD submission is designated into specific modules, sections, and docu- ments creating a backbone structure. This structure then results in each doc- ument to be contained in a subfolder of a specified section, and each section in the folder of its specified module. The eCTD format is the only acceptable elec- tronic submission format of regulatory documents to the FDA. Furthermore, for some device submissions, the FDA offers a software system (eSubmitter) for use in the electronic submission of certain types of device applications.
Overall it is easy to appreciate the level of standardization and presumed simplicity that electronic submissions offer large corporations. Unfortunately, complying with the electronic submission requirements has created great challenges for small biotechnology companies and academic institutions. There are meticulous requirements in formatting and naming conventions often requiring expensive software packages, regulatory expertise to ren- der electronic documents required to generate an acceptable electronic sub- mission, and only those submissions that are demonstrated to be error-free are accepted into the electronic submission system. Document rendering to ensure an acceptable format for FDA electronic software acceptance takes countless hours and typically requires a small group of dedicated staff for each eCTD submission requiring additional resources, which is challenging for a biotech company that may generate only a few submissions over the course of a 1-year period. One option to acquire the ability to compile and submit eCTD documents to the FDA would be to use template documents. This may be the most cost effective option but does require a significant investment in time and effort with a steep learning curve. A second option is to purchase an e-Submission software package to include publishing and validation capabilities. Although more straightforward, this option is more costly than the piecemeal approach previously described using template doc- uments and rendering them via trial and error. A final strategy to consider may be to hire a consultant or to outsource data management for a submis- sion. This may turn out to be the most expensive option but requires the least amount of training and is expedient.
The IND is a legal document, and it is important for the sponsor to provide only information that is true and accurate, and to disclose any and all infor- mation relevant to the product, notably safety data. On receipt, the FDA has 30 calendar days to review the IND and decide whether or not to allow the sponsor to proceed with the proposed clinical study. If during this 30-day review period, the FDA decides there is insufficient information to evalu- ate safety based on the information provided in the IND application, then additional information will be requested of the sponsor. If the 30-day review period is reached and the FDA review team identifies issues regarding prod- uct safety or potential undue risk to human subjects, then the application will likely be placed on Clinical Hold until these issues are resolved. A formal clinical hold is a mechanism used to ensure that the study will not proceed until the FDA is satisfied with the sponsor’s ability to address all safety con- cerns identified as a result of the review process.
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This formal iterative process is dictated by documented communications usually in the form of letters and amendments to the IND application. A sponsor has the right to appeal a clinical hold but, in doing so, must pro- vide significant data to refute or better explain the safety concerns brought forth by the FDA. Release from a disputed clinical hold or appeal typically involves a process of scientific negotiation and may include the role of an FDA appointed ombudsman but this process takes time, and thus a clinical hold or appeal process is an undesirable process for any biopharmaceutical development program.
To better ensure the chances of a successful IND, a sponsor often com- municates and discuses with the FDA his or her intentions to submit an IND, well before the IND is written. This is a formal process with several steps. The FDA provides guidelines for meetings with sponsors, classify- ing each meeting type as either A, B, or C and offering guidance on the associated timing and how to plan a successful meeting at various stages of development and for different purposes. Once a sponsor has some sci- entific information about a product and clinical trial design, the time is right to request a pre-IND meeting. To benefit the most from this meeting, it is advisable to prepare well in advance by crafting specific questions for the FDA to comment and provide input either agreeing with the develop- ment plan, clinical approach, and regulatory strategy or not. This dialogue with the FDA is an opportunity to evaluate sensitive areas or potential areas of specific safety concerns of the FDA review team. In scheduling a pre-IND meeting, the sponsor submits a formal written request to the FDA. This request includes an information packet or premeeting package, outlining information and data relevant to the product, and summaries of the clinical protocol, nonclinical safety data, and objective of the meet- ing. Also included in this information are potential meeting topics and associated questions, a list of meeting participants and their role in prod- uct development along with proposed meeting dates. This information is submitted to the FDA in advance, typically 30 days or more, prior to the proposed meeting dates. This is accompanied with a request that the FDA meet either in person or by teleconference with the sponsor to discuss any issues. The sponsor also queries FDA staff’s opinion on specific items of concern. For example, the sponsor may ask whether a particular nonclini- cal study was necessary, or if a certain manufacturing method or the con- cept clinical protocol design was acceptable. The FDA will usually provide written feedback after review of the information packet just prior to the scheduled meeting date. In the interest of being sensitive to the many time commitments of FDA review staff, the FDA will provide a statement in its written response. For example, the FDA might respond to the request: If you find that our written responses and advice are sufficiently clear and complete to obviate the need for further discussion, please inform us as soon as possible so that the meeting time may be cleared. Alternatively, the FDA may recommend the meeting as proposed.
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Timing of the request for a pre-IND meeting is crucial because the sponsor must have acquired enough information about a product to allow the FDA to make an informed response to the specific questions being asked. At the same time, it is not appropriate for a sponsor to approach the FDA requesting pre-IND feedback shortly before filing an IND. As previously mentioned, a formal process with associated deadlines and responses is required for FDA interactions to occur. The result of a well-defined, -timed, and -executed meeting with the FDA is thoughtful recommendations from the agency to the sponsor which are based on questions provided to the FDA in advance of the meeting. Pre-IND communication greatly reduces the chances of a clini- cal hold on the future IND application if, in fact, the sponsor addresses the issues raised by the FDA in their pre-IND responses.
Responsibilities for the sponsor and FDA to communicate do not end with filing an IND application and receiving clearance to begin a clinical study. Once the product is considered an Investigational Drug, the pro- cess of submissions, responses, meetings, and teleconferences has only just begun. Meetings and communications between the sponsor and FDA con- tinue throughout the life of an IND and the development cycle it supports. These are shown in Figure 3.3. A few commonly used meeting venues, in addition to pre-IND, are noted here. A meeting is sometimes held prior to initiating Phase 2 clinical studies (pre-Phase 2), and one is always held prior to performing a pivotal clinical study, Phase 3 (pre-Phase 3 meeting), and prior to submitting the marketing application, a NDA or Biologics License Application (BLA) (pre-NDA or pre-BLA meeting). These meet- ings allow the FDA and the sponsor to agree to clinical study designs that, if successful, support the next step of the approval process (e.g., market- ing application). Sponsors also request, during the course of development, a meeting with the FDA to discuss a special issue, such as the need to do an additional nonclinical study or the design of a new manufactur- ing plant. Although these subject matter-specific agendas may be com- bined with a clinical or milestone meeting, they typically require input of experts outside the clinical arena, and thus result in individual meetings or teleconferences.
Throughout the life of the IND, note that is not unusual for an IND to be active for more than 10 years, the sponsor must file written reports and data with the FDA. Annual reports outline progress on the program and changes in technology, summarize product manufacturing and disposition, and summarize clinical and nonclinical studies, and are provided on the anniversary date of the IND. In addition, and at anytime during the year, significant changes made or findings related to safety of the investigational product, notably toxicology or adverse reactions seen in clinical or nonclini- cal studies, must be immediately communicated to the FDA in writing. There is also a need to keep the FDA abreast, by written correspondence, of matters arising in development. In any given year, a sponsor is likely to submit sev- eral letters, many containing updates or modifications to protocols or data, to
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the IND. Each letter is considered an amendment under that IND and there- fore represents a legally binding statement. For example, the 10th letter or amendment to an IND would be numbered by the FDA as BB-IND32401-010, where BB stands for a biological, IND32401 is an FDA-assigned number given to that original IND (on first receipt), and each future amendment to the IND is labeled sequentially. So sequential number 10 (0010) represents the 10th amendment to the original IND submission. Ensuring the accuracy and completeness of each submission, managing meetings and premeeting information, and maintaining all submissions to an IND are important tasks and require professional regulatory support.
If undue risk is noted by the FDA, the agency can, at any time, impose a clinical hold and halt ongoing or prevent planned clinical studies. Clearly, there is a need for the sponsor to be diligent and to have a means, typically through a formal regulatory process and professional staff, to monitor all aspects of product development and to report findings and changes to the FDA in a timely manner.
Marketing Applications: BLA and NDA
A marketing application (NDA or BLA) from a sponsor is a request from the sponsor to the FDA for approval to enter a product into interstate commerce and to make claims of safety and efficacy about that product in the labeling. Hence, a marketing application is composed of all the information that is rel- evant to that product, all clinical and nonclinical study designs and results, and everything else known or discovered about the product, its research, manufacture, and control. NDAs and BLAs, submitted to CDER and CBER, respectively, are massive documents, containing narrative summaries, tabu- lated information, raw data, and explanations of how the data were gener- ated, analyzed, interpreted, and submitted. The sponsor also proposes the product labeling and provides critical documents stating the indication and making claims for the product and the product label, which is the written information affixed to the container of product. The proposed labeling must provide warnings, contraindications, directions for use, and other informa- tion. An outline of the most important (and required) product labeling, the package insert, is shown in Box 3.8. The Physician’s Desk Reference and other publications provide a collection of product labeling for FDA-approved drugs and biologicals, bound into one volume. Product labeling may also be found at the website of the FDA and those of product sponsors or manufac- turers. Biotechnology firms seldom make draft product labeling available to the public until a product has been approved.
The proposed labeling drives the application review process at the FDA. Each of the product claims in the proposed labeling must be supported by the data submitted to the FDA in the NDA or BLA. For example, if the spon- sor claims in the product labeling that a monoclonal antibody would stop the growth of prostate tumors for a period of 2 years (an endpoint) and prolong
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BOX 3.8 ELEMENTS OF PRODUCT LABELING (PACKAGE INSERT)
Trade name and chemical name. • Description: Describes the drug’s nature and classification.
Identifies how it is supplied in the final container and lists any excipients.
• Clinical Pharmacology: This gives the mechanism of action, pharmacodynamics, pharmacokinetics, and known drug interactions.
• Clinical Studies: All pivotal clinical studies and important smaller studies are described, to include endpoints and outcomes. Data are often summarized in graphic or tabu- lar format and all important results, efficacy and safety, are identified.
• Indications and Usage: The exact indication is given exactly in a very few brief sentences.
• Contraindications: Here are listed any situations in which the product should not be used.
• Warnings: Messages to the prescriber and user are noted, sometimes in bold and capitalized text to stress special safety issues. Warnings may be listed by organ system (e.g., cardiopulmonary events) or by disease (e.g., malignancies). These may be in bold and surrounded by a black box.
• Precautions: These are items that the user or physician should watch for, issues less common or important than those given in warnings. They are general, written for phy- sician and user, or they are information for patients, specifi- cally written for the user. Instructions for special situations are also placed here, and specific items are highlighted as paragraph headings. Recommendations may be given to stop using a product, for example, if a disease progresses or if certain symptoms are noted. Drug interactions, use in nursing mothers or in pregnancy, pediatric and geriat- ric use, or use in other special populations are generally included in this section.
• Adverse Reactions: General adverse reactions are first described, which are coming from reactions seen in the clin- ical studies. Here again key results of clinical trials are often presented in graphic or tabular format. A table presenting
(Continued)
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the life expectancy of the patient for 4 years (a second endpoint), then the design of the pivotal clinical trials must be focused on acquiring data to con- firm those endpoints. For example, if product labeling submitted to the FDA claims a monoclonal antibody is 98.7% pure, then the manufacturing and quality control data provided in the BLA must show both a specification for this purity level and provide data, from testing multiple lots of product, that it does in fact reach that purity level. Then BLA must further demonstrate that the monoclonal antibody will be manufactured and tested in such a way that there is high probability that future lots of product will achieve that same 98.7% purity. If the product labeling claims this monoclonal antibody did not result in autoimmune disease in nonhuman primates when it had been given more than 4 years, then that claim must be evident from the non- clinical data submitted under the BLA.
Further, for each NDA or BLA, it is typical for the FDA to request addi- tional data from the sponsor, sometimes asking that an additional study be performed, and inspections of clinical, nonclinical, and manufacturing facilities are routine during the examination product. These inspections are referred to as Pre-approval Inspections (PAI), biomonitoring (BIMO), or post-approval. Exact wording on labeling claims may be open to negotiation
BOX 3.8 (Continued) ELEMENTS OF PRODUCT LABELING (PACKAGE INSERT)
the most common adverse events is usually provided here. The most common adverse events are then described in a narrative paragraph. Finally, adverse events are listed by body system, cardiovascular through urogenital.
• Overdosage: This describes what is known if a patient should take more than the prescribed amount.
• Dosage and Administration: Information on how product is provided to a patient in final or dosage format is provided. This repeats and provides details on what has been given under description.
• How Supplied: If multiple formats exist (e.g., liquid or a tab- let), then each is described. The NDC number is given for each.
• References: A few key scientific publications are cited. • Administrative: Manufacturers’ name, address, and license
number with dates are given. • Patient Information: Instructions that are for the patient,
especially concerning proper handling, storage, or use, may be given on a separate but attached pamphlet.
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between the sponsor and FDA, but the claims must always be supported by data. Inevitably, the FDA asks a sponsor to make changes to their proposed labeling; discussions can be contentious but negotiations inevitably result in fair and balanced wording to labeling, and the FDA always makes the final decision. Note the cyclic nature of the product labeling in a well-conceived biopharmaceutical development project. In Chapter 1, we described the need for TPP (draft labeling) to drive the development process and now, at the NDA or BLA stage, we finalize that labeling. All the effort of the biopharma- ceutical development team went into generating data that would support the claims made in the draft labeling, presumably the same claims seen in the proposed and final labeling. With product labeling, the development process is truly a cycle, beginning with a visualization of what will appear in label- ing and ending with proposed and approved labeling, which is supported by many years of effort and investment in between!
On submission of the NDA or BLA, the sponsor must pay a significant sum of money, referred to as an FDA user fee. This fee supplements FDA resources for product review and is not refundable if the application is denied by the FDA. Additional fees must be paid for each manufacturing facility at appli- cation and periodically for the marketing life of the product. Review by the FDA takes months since each claim must be carefully examined in light of the data presented in the application. No wonder that a prudent biotechnol- ogy operation carefully prepares an NDA or BLA and submits it to the FDA only once it is supported by data. Failure is expensive for a sponsor in many respects!
The format for an NDA and BLA has changed to harmonize with the inter- national community. As previously mentioned in this chapter, most coun- tries have adopted the CTD format, and the electronic or eCTD is encouraged over the historic hard-copy paper submission.
Medical Device Applications. 510(k) and PMA
Sponsors of new medical devices, regulated through FDA’s CDRH, are faced with a variety of pathways to market approval. For any given device, the pathway is determined largely on the risk posed to the user by that class of device and, to a lesser extent, on the nature of the device and previous experience with that device. As noted earlier, medical devices are in one of three classes based on the risk posed by the device. Class I, generally low risk devices, must be developed under general controls, processes that include the quality systems regulation (QSR) for manufacturing and record keeping. Very few Class I devices require premarket notification, or 510(k) process. Class II devices, of moderate risk, are subject to both general controls and spe- cial controls and most are subject to premarket review and clearance under
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510(k) premarket notification process, a process described in Subsection “Medical Devices”. Class III devices carry potentially more risk and include many life-sustaining or -supporting implantable devices. These are subjected to the most rigorous controls. In addition to general and special controls, they are subject to PMA. Some Class III devices may be marketed under the 510(k) rules, but most must undergo the PMA process. The PMA pro- cess demands proof that the device is both safe and effective, which usually means clinical studies are required. For human studies of medical devices, an Investigational Device Exemption (IDE) is required of devices presenting the highest risk to the user. No matter which route is taken to market, medical device labeling is required in a device marketing application, and it cannot be false or misleading.
Few biotechnology products are considered medical devices alone, but some are combination products and so the sponsor of a combination device must consider medical device regulations in their development scheme. However, the inclusion of a medical device into a combination product does not lessen the regulatory hurdles, rather it likely increases them due to the added product complexity. As noted earlier, the final determination of which center at the FDA will be designated as the lead review for a biopharmaceuti- cal combination product is based on which component, device or biological, primarily produces the major effect. More often than not, it is the biological that is designated the primary mode of action and, in such cases, the product will follow the IND to BLA (or IND to NDA for a drug–device combination) route to approval with concurrent review by CDRH.
A device submitted to the FDA for clearance under 510(k) must demon- strate substantial equivalence to a legally marketed predicate device. The device must have performance standards, it must have an indication for use, the proposed labels must be accurate, and it must be fully described to CDRH in the 510(k) application. There must be evidence that it will be legally marketed and information on the device’s safety and efficacy profile must be provided. The PMA and its contents are equivalent to those of the BLA or NDA, because they demand a significant amount of nonclinical and clinical study data to support label claims and because the manufacturing controls are quite stringent. Clearly, most device manufacturers would pre- fer to register their devices under 510(k) approval process.
In vitro diagnostics (IVDs) are a class of medical devices that have a few special rules. Since many biotechnology products are used as key reagents with IVDs, their regulation is of great interest to biotechnology firms. IVDs encompass a host of products, from complex instruments to test kits used to diagnose life-threatening diseases to simple diagnostic laboratory reagents. Many IVD biotechnology products are marketed fol- lowing 510(k) premarket notification, and a few must enter the market through PMA route; either pathway for IVD is under review by CDRH. Also under the review of CDRH are biotechnology products, reagents, and instruments designated for Research Use Only, Investigational Use
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Only, Analyate-Specific Reagents, and Laboratory Developed Tests. Biotechnology firms may find a ready market for their products, some- times originally developed as laboratory reagents, with IVD manufac- turers who purchase reagents to include in their test kits. Before making a decision to enter the IVD field, firms should understand the possible impact of entering a regulated arena. Considerable diligence, research, and regulatory planning are advised.
Special Documents, Pathways, or Exemptions
• Master Files: The FDA provides a means to file confidential informa- tion for any type of product; the submission is called a Master File (MF), and the contents are maintained similar to other regulatory filings. There are five recognized types of MFs designated: Type I, II, III, IV, and V, with the most common MF used by the biotechnology industry to be Type I for manufacturing site, facilities, operating pro- cedures, and personnel, Type II for drug substance or drug product, intermediate product, and materials used in their preparation, and Type III for packaging material. The advantage of filing an MF is to allow the FDA to review important information without divulging confidential information to other industry partners on manufactur- ing processes or testing methods for example. The MF mechanism allows for the use of information that is already on file at the FDA to be accessed for the new application that should minimize review time. This mechanism of cross-referencing information may also be used to avoid duplicating costly and lengthy pharmacology or toxi- cology studies. The MF may be referenced by the FDA in support of an IND, NDA, or BLA, with a letter of permission from the sponsor of the MF to FDA. The FDA reviews the MF only as an informa- tional source and in no case does the FDA constitute the contents of the MF as an application for investigational use or market approval. For example, a biotechnology Firm, A, produces a synthetic mole- cule that is sold to another biotechnology Firm, B for use in Firm B’s in vitro diagnostic. Firm A prepares an MF that describes in detail specific proprietary information, for example the manufacture and control methods used to make and test this molecule and Firm A then submits or files it with the FDA. Firm A also prepares a letter of cross reference or letter of authorization (LOA) for Firm B, and in it they give the FDA permission to review their MF as part of Firm B’s IND 510(k) filing. Firm B provides Firm A’s letter of cross reference in their application. In effect, Firm B does not have access to the tech- nical details of Firm A’s proprietary information, yet the FDA can review this information in great detail and on behalf of Firm B to ensure that it is pure and potent and suitable for use in Firm B’s IVD. MFs are commonly used in a number of ways to support both the
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regulatory and business interests of companies. MFs are required to be kept current through the standard annual reporting mechanism.
• Animal Rule: This regulation applies to biopharmaceuticals for which there is no possible or ethical way to test efficacy in humans. Examples are reasonably well-understood diseases caused by bio- logical weapons of terrorism or mass destruction. It allows the sponsor to test the efficacy of the product, a countermeasure, in a well-developed and surrogate (to man) animal model. The product’s safety is then studied in human clinical studies under IND. The IND or PMA process is used throughout development with the only dif- ference being that efficacy studies are typically conducted in at least two animal species.
• Accelerated Approval: Drugs or biologicals indicated for the preven- tion or cure of serious or life-threatening diseases are eligible for a program known as Accelerated Approval. Under this regulation, the FDA may approve such products on the basis of a surrogate or clini- cal endpoints if these are likely to predict clinical benefit. Clinical studies are still required, but they may not require the stringent and long clinical processes for other drugs. There are restrictions on the approval, however, and sponsors carefully choose this route in spe- cial projects.
• Emergency Use and Treatment IND: An FDA regulation allows inves- tigational drugs to be used outside the standard clinical protocol in serious or life-threatening situations and with FDA concurrence. This regulation may speed a drug to a patient who might other- wise be ineligible to enroll in the study, such as someone living far from the study site. There are caveats to this as the sponsor must supply product at no cost, and there is risk that the drug may be misused or result in harm to the patient. Biotechnology firms devel- oping life-saving products agree to Emergency Use with caution, but consider it important as a good faith effort to speed a product to needy patients. Treatment IND is a slightly different approach for investigational biopharmaceuticals. There must be preliminary evi- dence of efficacy and an indication for a serious or life-threatening disease or it may be used if there is no alternative drug available and if death is expected from the disease within months. Examples are advanced cases of AIDS and cancers. The product is typically in Phase 3 clinical studies, and the information on its use must be reported to the FDA.
• Orphan Product Designation: This leverage option is commonly used by biotechnology firms, because many biopharmaceuticals are developed for the diagnosis or treatment of rare diseases or condi- tions. By definition, a rare disease or condition affects fewer than 200,000 people in the United States per year. It provides incentives
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for firms to develop biopharmaceuticals that are used less frequently and therefore have a smaller market value. This program has been quite successful for more than 30 years and is coordinated by the Office of Orphan Products Development (OPD), FDA. A common European and FDA application is available. The sponsor benefits by receiving both 7 years of FDA-administered market Orphan Drug Exclusivity (ODE), exclusive market rights for the indication and a tax credit of up to 50% of R&D costs, waived FDA fees, clinical trial tax incentives, and protocol assistance, and reduced statistical bur- den such as numbers of subjects required in a Phase III human clini- cal study to support approval. Also, the FDA awards development grants to sponsors of orphan products.
• New Drug Product Exclusivity: This is protection from competition, for 3 or 5 years after marketing approval, for the holder of an NDA or BLA when the drug is a new chemical entity that is a product unique both in nature and the marketplace. This exclusivity, in addi- tion to that provided by a patent, encourages biotechnology firms to develop novel products.
• Fast Track Designation: Another program to expedite products to patients with serious or life-threatening diseases or conditions, and where there are clinical data to demonstrate the potential to address an unmet medical need, is called Fast Track Designation. The sponsor must request Fast Track Designation from the FDA, and certain criteria must be met. However, a sponsor with Fast Track Designation receives special consideration from the FDA, such as additional meetings in which guidance may be provided, Priority Review of market applications, and even a program to review cer- tain sections of the NDA or BLA incrementally, thus saving time in the review process.
• Breakthrough Therapy: A breakthrough therapy designation is granted to products that are intended to treat serious condition and have preliminary clinical evidence demonstrating substantial improvement over the current standard of care therapies. Request for consideration of this product designation occurs during clini- cal development under an active IND. Benefits of this designation include intensive development guidance, organizational com- mitment to involve senior managers and experienced reviewers, Rolling Review, and other actions that would expedite the review process. If the FDA determines the product does not meet the break- through criteria, they will provide a nondesignation letter to the sponsor stating this status was not granted and explain the reasons for the decision.
• Priority Review: This expedited review process is intended for a product developed to treat a serious condition or a labeling change
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to a pediatric study. A request for Priority Review is to be submitted with the marketing application, and if successfully granted, it will provide a shorter clock for marketing application review to 6 months from the standard 10-month review period. The FDA will acknowl- edge, in writing, if a Priority Review has been granted.
• Rolling Review: Product approval for a Rolling Review entitles the sponsor of a product to submit sections of an application for FDA review prior to the assembly of a complete marketing application submission. The intent is to avoid delays associated with comple- tion and assembly of a complete application and instead allow the FDA to review individual sections of an application such as chem- istry, manufacturing and controls, toxicology, or clinical sections. The most common and beneficial review to take place as a Rolling Review is preliminary evaluation of clinical data that demonstrate effectiveness.
Generic Drugs and Biosimiliar or Follow-on Biologics
Generic drugs have the same active ingredient as brand name marketed drugs and represent look-alikes that enter the market following expira- tion of patent protection. They are often made by manufactures other than the company that originally made the brand name drug, and they must be tested in small, head-to-head studies—laboratory and clinical pharmaco- kinetics and pharmacodynamics—to demonstrate chemical identity and pharmacokinetic and pharmacodynamic similarity. The approval process for a generic drug at the FDA follows the sponsor filing an ANDA, and indeed it is just that.
It is unlikely that generic drugs will be derived from biotechnology. However, their equivalent, biosimilars, also referred to as follow-on biolog- ics, biogenerics, or generic biologics, are biologic look- and perform-alikes. The FDA approved the first biosimilar product Zarxio (filgrastim-sndz) manufactured by Sandoz, Inc., a Novartis company (Princeton, NJ) in March 2015. Zarxio is biosimilar to Neupogen (filgrastim), which is marketed by Amgen, Inc., (Thousand Oaks, CA) originally licensed in 1991. Zarxio is approved for the same clinical indications as Neupogen and can now be pre- scribed by physicians as a substitute for Neupogen. Congress has met stiff resistance on two fronts: first, from biotechnology firms that currently hold the market in the absence of patent protection and second because there are serious questions regarding whether or not biological molecules can, in fact, be reproduced to mimic exactly the purity, potency, efficacy, and safety of the predecessor molecule. Clearly, any biosimilar product will need to be thoroughly tested in adequate and well-controlled preclinical and clinical studies. The extent of testing continues to be debated, but the biogenerics have now reached the market.
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Other Regulatory Activities
Many additional regulatory activities must be completed during the product life cycle, some before and others after product registration. Several examples are as follows:
• Establishment registration: Establishments manufacturing any drug or biopharmaceutical or medical device, whether U.S. or foreign, must be registered with the FDA.
• Licensing issues: Divided, shared, or contract manufacturing. To accommodate the complex and sometimes specialized manufac- turing schemes required for biopharmaceuticals, the FDA allows manufacturing of one product at two or more sites. For example, a biopharmaceutical might be produced by fermentation at one site, then shipped to a second site for purification, formulated at a third site, and filled and labeled at a fourth site. Such divided or shared manufacturing, generally done largely by contractors for the spon- sor, is allowed if it is carefully controlled and defined and if each site is a registered establishment.
• Proprietary name: A sponsor wishes to have a unique name, apart from the often long and confusing chemical name, for their bio- pharmaceutical. To avoid duplication for confusion in labeling, the FDA is responsible for approving the proprietary name for each biopharmaceutical.
• National drug code: In addition to the unique name, the FDA issues with marketing approval a unique drug number, a National Drug Code (NDC), and this is clearly marked on all labeling.
Public Meetings and Advisory Committees
A cornerstone of good government is the right to speak in public for or against an issue, especially if that issue arises from a government or government- regulated activity. Biotechnology products and the marketing approval of biopharmaceuticals and medical devices are no strangers to the public arena, and their use and release into the environment have been a matter of debate ever since recombinant Pseudomonas syringae was sprayed on strawberry fields of California in 1983 to prevent frost damage. The photograph of sci- entists dressed in protective gear and spraying the bacterium elicited much debate worldwide. We noted earlier that a key part of the formal process to make or to change a regulation is the public rulemaking process in which the public has an opportunity to review and comment on proposed regula- tions before rule publication and codification. In addition, regulatory agency
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processes may be influenced by public petition: requests made to produce, remove, or change a regulation. Other rules allow the public to demand eco- nomic or environmental impacts for a regulation, and these processes also mandate the public be informed and allowed to influence the government’s decision. All of this applies to the FDA and other regulatory agencies. States have similar rules that result in public hearings or meetings. Just as it was in 1983, public debate is a very important aspect of rulemaking.
The FDA also uses advisory committees to its and the public’s advantage by asking expert panels to review data on safety and efficacy for products near completion of review and recommend approval. The FDA has estab- lished these committees for every class of drugs, biologicals, and medical devices under its purview. Members meet at established intervals to make recommendations on a variety of subjects. At the top of their list are product- specific recommendations, notably whether or not a product should receive market approval. This is typically done after the FDA has completed the review. For this, the committee is asked to answer a series of questions, such as, Does the committee view this product to be safe for its intended use? Panel members vote, but the recommendation is not binding to the FDA, and the agency will sometimes decide in a manner not consistent with the panel’s majority recommendation. Clearly, such meetings and the commit- tee’s voting record and recommendation are extremely important to the bio- technology firm sponsoring the product put before a committee. Another function of advisory committees is to make recommendations on groups or classes of products. For example, if a class of monoclonal antibody, repre- sented by several similar products, appears to cause an unexpectedly high number of allergic reactions, the FDA might ask a committee to meet and discuss the situation and perhaps make a recommendation, such as posting a warning on the label. Advisory committees also perform more mundane tasks such as reviewing research laboratories at the FDA. It is very important that members of these committees be experts yet have no conflict of interest such as working in a commercial environment with the products on which they make recommendations or receive money from the sponsor. Indeed, a member should have no strong personal bias for or against a technology. As one might imagine, it can be a challenge for the FDA to find the right experts to serve on advisory committees. Committee meetings are open and announced to the public, comments are solicited, and minutes and votes are a matter of the public record.
Having read this chapter, one might ask, What is there about a biopharma- ceutical operation that is not made public and that can be kept confidential and proprietary? The answer is quite a lot; much, perhaps most, of the tech- nical information that the firm considers proprietary, and all of the financial facts are kept from public view. The FDA does not delve into a firm’s finances or the public or private nature of a company, which is a territory for the Securities and Exchange Commission or the Internal Revenue Service. The FDA does not consider marketing other than whether or not promotion is
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in line with approved labeling and in a few other areas related to market approval. Proprietary information that must be disclosed to the FDA by the biotechnology firm in applications or correspondences is maintained as con- fidential and remains hidden from public view. Regulatory agencies, unlike many organizations in political capitals, do not leak confidential information to the public or to the media.
Although a limited amount of information is accessible to the public through Freedom of Information Act (FOIA), this information is nonproprietary or reportable observations from inspections and provided in warning letters or Form FDA 483. Indeed, proprietary information is redacted before public dis- closure through FOIA.
Postmarketing Requirements and Activities
Interaction with regulatory authorities does not stop once market approval is given. The FDA can withdraw marketing approval any time after it has been granted. This has been demonstrated in cases where the Agency inves- tigates a proposed noncompliance issue which results in the ability to prove cause (i.e., demonstrating noncompliance with regulation). This happens when a sponsor (holder of the market approval letter) fails to meet report- ing requirements or if a product proves unsafe. What are postmarketing requirements of the sponsor? Some of them are as follows:
• Post-approval maintenance of the approved NDA or BLA: Sponsors must file annual reports with the FDA as long as they are marketing an approved drug. Elements of an annual report include but are not lim- ited to labeling, chemistry, manufacturing and control, nonclinical testing, and clinical data. Reporting of changes from the original mar- ket approval is especially important to regulatory agencies. It may be necessary for the sponsor to report significant changes immediately and not wait for the annual report. At the time of market approval, it is normal for the FDA and sponsor to agree to certain postmarket- ing clinical studies, such as Phase IV or monitoring of special patient populations. These commitments by the sponsor also include adver- tising and labeling changes, product complaint reporting schemes, or events that trigger product recalls. The FDA and the public take these commitments quite seriously.
• Reports of adverse drug events (experiences): ADEs are explained in Chapter 9. Briefly they involve reactions in patients using a biophar- maceutical, and they must be reviewed by the sponsor if they fall under certain guidelines for severity or frequency and might be related to the product. Physicians and users may report these expe- riences to the sponsor and sometimes directly to the FDA. Direct communication is referred to as MedWatch for many medications,
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whereas specific products such as vaccines have a unique report- ing system. Certain types of products, such as vaccines, have unique reporting requirements. The sponsor, in turn, must report serious experience situations to the FDA in an Alert Report within 15 days. The rules are not complex, but they are considered extremely impor- tant to maintaining public health and a safe source of biopharma- ceuticals, as well as keeping a positive image for the firm and the biopharmaceutical industry as a whole.
• Risk evaluation and mitigation strategy: The FDA has instituted this program, referred to as REMS, to improve postmarket approval safety of medical products. REMS includes guides for medications, patient-friendly labeling, and improved communication from the FDA or sponsor with health care providers to better ensure proper use of products. The document is a plan that is submitted by the sponsor to FDA for approval as part of the marketing application.
• Dear doctor letters: The FDA believes that an effective means of communicating new information, especially risks, for prescrip- tion biopharmaceuticals is by ensuring well-informed prescribing physicians. Letters to doctors often fill that objective, along with announcements in medical journals, and through the public media.
• FDA letters to manufacturers: Letters to manufacturers are another matter, since these are targeted directly to the sponsor and are often issued in response to compliance issues. Communication between sponsors and the FDA are therefore discussed in Chapter 4.
Advertising and Promotion
Drugs, biologics, and medical devices are heavily marketed to various target populations—physicians, nurses, pharmacists and patients, or end users— and we accept this in our society. As compared to many other countries, biopharmaceutical marketing and promotion are lightly regulated in the United States. As discussed elsewhere, marketing and advertising activities are regulated through claims made on the label. A biopharmaceutical label is defined as, A display of written, printed, or graphic material on the immedi- ate container of a drug. Labeling, as defined by the FDA and used as a noun is “any written, printed or graphic material on the drug, on any of its con- tainers or wrappers, or on any material accompanying it” (Federal Register, 2006). Hence, the package insert, that lengthy document that contains pre- scribing, safety, and dosage information, and is stuffed into boxes of OTC or prescription medications, is labeling. Promotional labeling is any labeling used in advertising or marketing activities. Promotional labeling is at the heart of biopharmaceutical sales, and so it becomes a point of contention whenever it does not reflect the approved labeling. For example, one of the most egregious violations of FDA marketing rules is to promote the use of a
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biopharmaceutical for an indication or use that is not given in the approved label. This is known as promotion for off-label or unapproved use. Although the FDA does not restrict licensed medical practitioners from prescribing medications for or advising patients to take medications outside the labeled information, the FDA does not allow sponsors in anyway to promote this practice. Hence, it is only legal to promote biopharmaceuticals, or other drugs and biologicals, in accordance with the approved label. Advertisements must be balanced and complete, again as driven by information included in the FDA-reviewed and -approved label.
A product’s label will almost certainly change during the postmarketing period, and this results in refined definitions for what can and cannot be included in promotional materials. To ensure that biopharmaceutical pro- motional information is in line with current labeling, the FDA insists that it be provided for review by the agency at certain times during product devel- opment. Investigational products may never be promoted. Most or all pro- motional materials for approved products must be submitted to the FDA as they will be used immediately after market approval. This first advertising campaign is referred to as a launch. Also, most other promotional materials generated postlaunch must also be submitted for review. Dissemination of scientific and medical information is also closely monitored by the FDA and direct-to-consumer advertisements, such as television and newspaper ads, is controlled. An important subject of ongoing debate is whether or not the consumer or the health practitioner is given adequate information about safety issues that are known to be or could even possibly be related to a product.
Summary of Regulatory Affair Activities in Biotechnology Operations
This discussion completes this chapter on regulatory affair operational activities, and it reiterates themes that were introduced in Chapter 1. Having a historical perspective of FDA regulation and understanding organization of the FDA provides a foundation to better understand the regulatory con- siderations, regulation, and guidelines throughout the product development process. Early in this chapter, the different types of regulated medicinal products were highlighted under the broad categories of drugs, biologics, devices, and combination products. Further mention is made of biotechnol- ogy products that are considered cosmetics, dietary supplements, or veter- inary products but are not regulated as medical products. The regulation of medical devices is complex since they include a wide variety of product types and today involve many combination products. Developing regula- tory strategies to enable efficient biopharmaceutical development based on
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understanding how each biopharmaceutical product is regulated by the FDA is a process collectively termed regulatory intelligence. Use of regula- tory intelligence allows us to strategize and plan for biotechnology opera- tional adjustments at every stage of product development.
It is also important to continuously assess the risk-to-benefit balance during product development with an eye toward understanding how the regulator will interpret our analysis of biopharmaceutical development information. In most cases, the benefits must outweigh the associated or anticipated potential risks. The IND is a formal application to and for review by the FDA, and the process used to obtain their permission to use an investigational product in a human clinical study. Another application, the BLA or NDA, is used to request FDA authorization for approval to mar- ket a medical product, biologic or drug, respectively, in the United States. Each application has specific format requirements with different formats for paper versus electronic submissions. It is important to recognize that user fees may be associated with the submission of a regulatory application. An important and essential part of the successful development of a biotechnol- ogy product is managing an active relationship with the FDA and being responsive to any FDA request.
An introduction and various examples of special documents, pathways, or exemptions established specifically or expediting product development for particular underserved clinical indications have been provided. Also pre- sented in this chapter are a brief discussion on generic drugs or biosimilars that have now reached the market, a summary of the approval mechanism, and the testing requirements that continue to cause much debate. Adequate testing of generic or biosimilars remains controversial since demonstration and testing of comparability are usually required only for the active ingre- dient, and overall the product must pass the brand product specifications with regard to potency and percent impurities. It is well documented that the impurity profile of medical products is the primary contributor of safety issues.
The FDA has many regulatory responsibilities and oversight that are briefly mentioned in this chapter which include review of establishment registra- tion information and of priority naming conventions, licensing issues, and issuing product codes. As a public service, the FDA organizes public forums inviting a subset of experts but is open to the public and often attended by special interest groups—patients, lobbyists, or other stakeholders who are interested to hear firsthand on the potential issues and current thinking on specific topics. It is also very common for the FDA to rely on outside expert opinion that is solicited by establishing advisory committees and meetings on specific topics.
Information has been provided in this chapter on the various types of formal FDA applications including those for investigational product use, labeling, and commercialization. Also, recall that compliance with detailed postmarketing requirements is essential to lawfully market medical products
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throughout the product lifecycle. These marketing requirements include advertisements, promotions, and medical claims.
Regulatory planning starts early and involves successful product develop- ment through the very end. Planning must consider all regulatory aspects of the product and its development; each step in the cycle. The product labeling is central to the development lifecycle, beginning with a draft labeling or TPP and ending with approved labeling for the marketed product.
References
FDA. 2006. Guidance for Industry: Complementary and Alternative Medicine Products and Their Regulation by the Food and Drug Administration. US Food and Drug Administration. http://www.fda.gov/downloads/regulatoryinformation/ guidances/ucm145405.pdf (accessed May 31, 2016).
Federal Register. 2006. Department of Health and Human Services. US Food and Drug Administration. http://www.fda.gov/OHRMS/DOCKETS/98fr/06-545. pdf (accessed May 31, 2016).
Sinclair U. 1905. The Jungle: The Uncensored Original Edition. Sharp Press, Tucson, AZ.
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4 Regulatory Compliance
Regulatory Compliance
Information in this chapter builds upon an understanding of regulatory operations (Chapter 3) by examining the broad world of regulatory com- pliance, and discussing the FDA requirements to integrate quality into all aspects of biopharmaceutical development programs and by review- ing many regulations outside of the FDA that impact most biotechnology operations.
Quality Systems to Meet Regulatory Compliance
Compliance and Quality Systems
The Oxford English Dictionary (1997) defines compliance as “the act or instance of complying; obedience to a request or command.” Further defined for bio- technology product development, compliance is the act of meeting a plethora of rules, regulations, and directives. Compliance impacts each biotechnology development function every day. It involves constant vigilance to identify and understand each applicable regulation and, most importantly, it drives the bio- technology firm to institute and integrate programs that ensure obedience to these regulations and directives. Compliance is achieved largely by ensuring quality in all aspects of development and at every step in the development cycle. This is best done by instituting quality systems, described in Chapter 5. Indeed, results of FDA inspections repeatedly demonstrate that firms with mature and effective quality systems consistently have, in the eyes of regu- latory agencies, fewer deficiencies than do operations with deficient quality systems. With this in mind, it is easy to consider the need to integrate, into a biotechnology operation, scientific and technical skills, regulatory guidance, and quality systems.
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Quality systems are composed of quality hallmarks, features of a well- established, compliant, and smooth operation (Chapter 5). In this chapter, we discuss the intersection of compliance with quality system. Three examples of quality systems—current Good Manufacturing Practices (cGMP), cur- rent Good Laboratory Practices (cGLP), and current Good Clinical Practices (cGCP)—are outlined in the following and each will be further described in Chapters 6 and 7 (manufacture; quality control), in Chapter 8 (nonclini- cal), and in Chapter 9 (clinical), respectively. These are excellent examples of systems applied by most nations to protect public health by ensuring safety and efficacy of biopharmaceuticals. They are presented here as U.S. FDA regulations, but similar good practices of many nations are currently being harmonized into international compliance guidelines and so these elements now reach worldwide.
Current Good Manufacturing Practices for Manufacture and Quality Control
cGMPs were established to prevent drug and medical device manufacturers from producing and selling adulterated products to the public. In brief, adul- terated products are those that contain harmful ingredients, misrepresent strength, are mixed with other substances than are approved, do not con- form to performance standards, packaging, labeling, and storage conditions, in which strength/quality/purity do not match the label, or lack adequate manufacturing controls. This, in turn, was desired by the public because adulteration had occurred in medical product manufacture, and the practice was not tolerated by the public. cGMP is an established quality system that has been shown to have a positive effect on the quality of biopharmaceuti- cals and hence cGMPs have been adopted as the manufacturing standard, worldwide.
In the United States, biopharmaceutical manufacturing compliance is based upon regulatory requirements for manufacturing processes and utili- ties, codified for human and animal drugs and in 21 CFR 210 and 211 as well as in other sections of FDA regulations. Important elements of cGMP are listed in Box 4.1. cGMPs are now quite well harmonized worldwide and any differences in cGMPs largely reflect the nature of the product or differ- ences in its usage. It is repeatedly stated that “…quality cannot be tested into a product but that the sum total of what constitutes that product must be of the highest quality” (Federal Register Online, 1996). Biopharmaceutical manufacture and control are, under cGMP, based upon the idea that a prod- uct, the process to make it, and the laboratory control tests must be designed in a manner that meets the intended use. cGMPs strive to meet that stan- dard. These quality systems concepts are discussed further in Chapter 5 and examples or application of cGMP to actual manufacturing and control pro- cesses are demonstrated in Chapters 1 through 4, 6 and 9.
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BOX 4.1 HIGHLIGHTS OF CURRENT GOOD MANUFACTURING PRACTICES (cGMP)
• 21 CFR 210—Current Good Manufacturing Practice in manu- facturing, processing, packaging, or holding of drugs Status, applicability, and definitions
• 21 CFR 211—Current Good Manufacturing Practice for fin- ished pharmaceuticals
A. General provisions B. Organization and personnel
– Responsibilities of quality control (assurance) unit, per- sonnel qualifications and responsibilities
C. Buildings and facilities – Design, construction, lighting, ventilation, plumbing,
sewage, washing and toilet, and sanitation D. Equipment
– Equipment design size, location, construction, cleaning, calibration, maintenance; automatic, mechanical and electronic equipment, filters
E. Control of components and drug product containers and closures – Receipt and storage, quarantine and release, testing and
use of components, containers and closures; retesting; rejection; drug product containers and closures, waste removal and flow
F. Production and process controls – Written procedures, vendor qualification, deviations;
yield; equipment identification; sampling and testing of in-process materials and drug products; time limita- tions; control of microbial contamination; reprocessing.
G. Packaging and labeling control – Materials examination; issuance of labels; tamper-
evident packaging; inspection; expiration H. Holding and distribution. Warehousing and distribution.
(Continued)
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BOX 4.1 (Continued) HIGHLIGHTS OF CURRENT GOOD MANUFACTURING PRACTICES (cGMP)
I. Laboratory controls – Testing and release for distribution; stability testing;
special testing; sampling plans, reserve samples; labo- ratory animals
J. Records and reports – Cleaning and use logs, equipment calibration records,
component, container, closure and labeling records; master production and control, laboratory, distribution and com- plaint records and review;
K. Returned and salvaged drug product • 21 CFR 600—Biological products
A. General provisions B. Establishment standards
– Personnel; establishment, equipment, animals; records, retention samples, product deviations; temperatures during shipment
C. Establishment inspection D. Reporting adverse experiences
• 21 CFR 610—General biological products standards A. Release requirements B. General provisions
– Methods and processes: General safety, inactivation, sterility, purity, identity, constituent materials, combi- nations, cultures
F. Dating period limitations G. Labeling standards
– Container and package labels, name of product, manu- facturer and distributor, export
• 21 CFR 630 and 640—Standards for human blood and blood products
• 21 CFR 660—Standards for diagnostic substances for labora- tory tests (blood products)
• 21 CFR 680—Additional standards for miscellaneous (biological) products
(Continued)
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Current Good Laboratory Practices for Nonclinical Laboratory Studies
cGLP regulations were established because certain individuals were perform- ing nonclinical laboratory studies in an unscientific or uncontrolled manner and the results of these studies could not be trusted. Important toxicology data were found to be questionable. In response, a regulation, 21 CFR 58, was established in 1979 with the purpose of ensuring the quality of nonclinical safety studies for medical substances. The key elements of FDA cGLP regula- tions are outlined in Box 4.2. Taken together, cGLPs ensure that testing has been performed in a sound scientific manner and with an established qual- ity system. Notable are requirements for study protocols, appropriate data capture, accurate reports, internal quality audits, and acceptance of results by both the scientists and a quality assurance professional.
Current Good Clinical Practices for Clinical Studies
cGCPs represent a quality system that ensures the highest quality science and ethical treatment of human subjects for clinical studies of all types and at all phases of development. With cGCP the burden for quality is shared between the principal parties conducting a clinical trial: sponsor, investiga- tor, and, if one is used, contract research organization (CRO). Unlike cGLPs and cGMPs, which are to be found in one or a few sections of 21 CFR, cGCPs are codified in a number of chapters and sections of the regulations. This is due in large part to the broad scope of clinical trials overall, the fact that they involve FDA functions, and generally recognized and codified rules for the conduct of research that involves human subjects no matter what the reason for their enrollment. The key components of cGCP are outlined in Box 4.3. In the biopharmaceutical industry, an important foundation of cGCP is that the regulation both protects users of biopharmaceutical prod- ucts and also safeguards the well-being of human subjects, those individu- als taking personal risk by volunteering to test new products. Indeed, the protection of human subjects is paramount in cGCPs, as it should be.
BOX 4.1 (Continued) HIGHLIGHTS OF CURRENT GOOD MANUFACTURING PRACTICES (cGMP)
• 21 CFR 820—Quality system regulation (for design and manu- facture of medical devices)
• 21 CFR 1270—Good Tissue Practices • 21 CFR Part 11—Electronic records; Electronic signatures
• Controls for closed and open systems, signature manifesta- tions and record-linking, electronic signature components and controls, identification and passwords.
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BOX 4.2 ELEMENTS OF CURRENT GOOD LABORATORY PRACTICES (cGLP)
• 21 CFR 58—Current Good Laboratory Practices for nonclinical laboratory studies
A. General provisions – Definitions, applicability, and inspections
B. Organization and personnel – Personnel, management, study director, quality assur-
ance unit C. Facilities
– Animal care and supply; handling test and control arti- cles, laboratory areas, specimen and data storage
D. Equipment – Equipment design, maintenance, and calibration
E. Testing facilities operations – Standard operating procedures, reagents and solutions,
animal care F. Test and control articles
– Test and control article characterization, handling and mixtures
G. Protocol for and conduct of a nonclinical laboratory study J. Records and reports
– Reporting study results, storage of records and data, retention of records
K. Disqualification of testing facilities – Grounds for disqualification, notices, final orders,
actions, public disclosure, and suspension • 21 CFR Part 11—Electronic records; Electronic signatures
• Controls or closed and open systems, signature manifesta- tions and record-linking, electronic signature components and controls, identification and passwords.
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Compliance for Biopharmaceuticals: Other Regulations of Importance
Compliance for Import of Biopharmaceuticals into the United States
Importation of biopharmaceuticals is regulated by a number of agencies in every country of the world. For the United States, Center for Biologics Evaluation and Research or Center for Drug Evaluation and Research oversee importation and
BOX 4.3 REGULATIONS FOR CURRENT GOOD CLINICAL PRACTICE (cGCP) AND CLINICAL TRIALS
• 21 CFR Part 11—Electronic records; Electronic signatures • Controls or closed and open systems, signature manifesta-
tions and record-linking, electronic signature components and controls, identification and passwords
• 21 CFR 50—Protection of human subjects. Informed consent • General requirements, elements and exception for informed
consent, additional safeguards for children • 21 CFR Part 54—Financial disclosure by clinical investigators • 21 CFR Part 56—Institutional review boards
• Organization, personnel, functions, operations records and reports, administrative action for noncompliance
• 21 CFR 312—Investigational new drug application and foreign clinical trials • Responsibilities of sponsors and investigators: responsibili-
ties of sponsors, transfer of obligations to a contract research organization, selection of investigators and monitors, informing investigators, review of investigations, record- keeping and retention, inspection of records and reports, disposition of investigational drug, assurance of IRB review, disqualification of clinical investigator.
• Drugs intended to treat life-threatening and severely- debilitating illnesses, emergency use.
• Foreign clinical studies not conducted under an IND, pub- lic disclosure of data and information.
• 21 CFR 314—Applications for FDA approval to market a new drug
• 21 CFR 320—Bioavailability and bioequivalence requirements.
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exportation of biologics or drugs, respectively, to ensure they comply with all U.S. laws and regulations. The FDA works closely with Customs and Border Protection (CBP). Inbound shipments in violation are detained by CBP on behalf of FDA or the United States Department of Agriculture (USDA). FDA must be advised if a final biological or drug product is manufactured overseas. A foreign manufacturer must have a U.S. FDA license to manufacture and dis- tribute that product. This means the product must have an approved marketing application and, before this is granted, the foreign manufacturer must usually pass FDA inspection. A product approved and manufactured in the United States may, however, be exported from the United States to another country without additional FDA authorization to export. In such cases, FDA provides, on behalf of a biopharmaceutical sponsor and to a foreign regulatory agency, a Certificate to Foreign Government to substantiate marketing approval in the United States. Investigational biopharmaceuticals are another matter. The man- ufacturer need not have a U.S. FDA license but they must declare a valid and active IND by number and name. CBP screens such shipments carefully, notify- ing FDA and USDA if paperwork is in any way out of order or incomplete.
Compliance for Medical Devices
There are aspects of medical device compliance that differ from other FDA products, and the biotechnology firm developing a combination product with device components is well advised to understand these nuances. As noted earlier, registration and listing of U.S. establishments developing or manufacturing devices is critical to understanding medical device compli- ance. Also, devices are classified according to the level of risk to the user. Quality systems regulations (QSRs) and guidelines demand a strict quality system for development and production of medical devices. While some aspects of device QSR, identified in Chapter 5, now apply to drugs and bio- logics, they are quite detailed for devices. Certain reporting requirements are also unique to medical devices, as identified under the FDA’s medical device reporting (MDR) regulations. Additionally, there are stringent rules on track- ing of certain medical devices. Medical device import and export compliance has many similarities to drugs and biologicals, but some processes do differ and are important to firms engaged in international transport, manufacture or marketing of medical devices or combination products.
Inspection and Enforcement
We are certainly all aware that in any society it is necessary to enforce laws and regulations. Yet, skirting or blatantly disobeying regulations just seems to come naturally to certain individuals, typically those motivated by
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personal gain, and so societies have established means of ensuring, or try- ing to ensure, compliance by everyone. These are (1) enforcements, a means of imposing on individuals the observance of law and (2) inspection, the official and careful examination of an item or an activity. Biological, drug, and medical device activities have, in the past, been found to be deficient and in some cases there has been proved a serious intent to produce adulterated product, to falsify nonclinical or clinical study data or to avoid providing human subjects with their legal rights. Such behavior does, unfortunately, exist. In an effort to ensure that all biopharmaceutical products are both safe and effective and to increase public confidence in the biotechnology industry, the FDA inspects virtually all aspects of regulated development and enforces regulations intending to keep biopharmaceutical products safe and effective.
Inspections
Inspections provide one means of ensuring compliance and most coun- tries have enacted laws to allow regulatory inspectors to review facili- ties, records, and operations that produce or distribute investigational or approved products. U.S. FDA inspections are typically conducted for the following reasons:
• Periodic review of an operation to ensure continuing compliance • Supplier of products to the government • Directed review due to issues related to a product • Revisit, following finding of deficiencies on an earlier inspection • Following a recall or complaint • Preapproval visit based upon a market application or amendment or,
more rarely (but in the case of a new technology, an investigational new drug [IND] Application).
The Food, Drug, and Cosmetic Act of 1938 gives the FDA broad author- ity in what may be inspected as long as the items—facilities, records, even vehicles—bear on whether a product (e.g., a biopharmaceutical or active ingredient) or service (e.g., a nonclinical or clinical study) is in compliance with the Act. Personal, financial, or business information is not a target of inspections and technical information is kept confidential for inspectional reports. Individuals are not required to sign affidavits but information they disclose may be used in a case against the firm.
Let us examine the FDA inspection process as it might happen at a bio- pharmaceutical firm. Primary reason or types of FDA inspections include preapproval, routine, and directed or for-cause. An inspection typically begins in the morning of a weekday as FDA inspectors present their credentials and state the reason for their visit. If a firm refuses entry, then the FDA
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will seek an administrative inspection warrant or, if serious breaches of the law are suspected, a criminal search warrant. The inspection itself involves a review of the plant, facilities, and records. FDA inspectors are highly trained and inspection teams, varying in size from one to a dozen FDA employees, include individuals with various expertise. For example, the team sent to a biomanufacturing plant might include individuals expe- rienced in record review, others with expertise in technologies used at the plant, and specialists with a deep understanding of general manufactur- ing processes and regulations. Inspections may be brief, lasting less than a day and conducted by one individual, or they may take weeks and involve teams of inspectors, visiting continuously or sporadically. Inspectors care- fully research the history of a product and the facility before they visit, and they are guided by FDA’s Inspection Operations Manual. Further, inspec- tors now use a systems approach when visiting an operation. There are compliance trends that lead to investigational emphasis, and these issues should be evident to the biopharmaceutical community through meetings and from press releases by the FDA. Today, for example, this is corrective and preventive action (CAPA), sources of active pharmaceutical ingredi- ents (APIs), and production and process or facilities and equipment con- trols, but in the future it may be other topics that the FDA and the public believe require immediate attention to ensure a supply of safe and effica- cious products. A thorough investigation begins at the top, looking at man- agement responsibility and involvement, moving to design control and always touching on the hot topics. During an FDA inspection, typically at the end of each day, a debriefing session takes place to ensure aware- ness and status of the inspection and also provides an opportunity for any clarification or misunderstanding. Upon completion of the inspection, the inspector conducts an exit interview with management and provides a list of any notable observations.
A variety of documents may be prepared by the FDA as the result of an inspection.
• Form FDA 483, Inspectional Results, lists notable observations made by the inspectors. It is issued to the firm before the inspector leaves on the final day.
• Upon returning to their FDA office, inspectors file details of their findings and present evidence or exhibits of deficiencies, uncovered in the Establishment Inspection Report (EIR).
Inspections result in one of three courses of action, as recommended by the FDA. For the biopharmaceutical firm, the preferred outcome is no action indi- cated (NAI); a clean bill of compliance health, if you will. Another possible outcome is voluntary action indicated (VAI) and the third is official action indicated (OAI). A prudent biotechnology firm will take any inspectional findings of VAI or OAI very seriously. The report on their firm becomes a
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matter of public record, and competitors and customers may file a Freedom of Information Act (FOIA) request to obtain Form FDA 483 or the EIR, redacted of confidential information. Management and quality staff of the firm is involved in all reviews of and responses to inspectional findings. In the ideal situation, a team of supervisors carefully examines the inspectional findings and compares them to regulations cited and to company records or proce- dures identified and described. Indeed, a systems approach is applied and the firm typically generates a voluntary plan to correct each deficiency cited by inspectors; this plan is submitted by the sponsor to the FDA for review. Negotiations between the agency and management of the firm may follow and there is usually a final resolution and agreement, satisfactory to regula- tory authorities. Such is the outcome for most VAI situations. Enforcement action is, however, indicated for OAI determinations. Time and again it has been shown that FDA has the upper hand in these matters and rarely does a firm avoid the need to admit to and correct OAI deficiencies found on an FDA inspection.
Overseas biopharmaceutical manufacturing facilities produce prod- ucts that are imported into the United States by national and international firms. There are strong economic drivers: lower cost of goods and operating expenses, ease of sourcing materials, and relatively low wages to prepare bulk substances and even final product. This shift in international manufac- turing has increased the demand for the U.S. FDA inspections of overseas operations. Bulk API or final product cannot be imported into the United States without an appropriate FDA approval, to include approval of the overseas manufacturing facility and hence the products made in that facil- ity. FDA inspectors perform routine inspections of overseas biopharmaceu- tical manufacturing and quality control operations and, in larger countries, FDA employees may even reside in that nation. Further, the FDA performs 100% screening of API and product that enters into the United States, to include surveillance inspections of imported goods to ensure compliance with the U.S. requirements. Ultimately, the U.S. firms, or the U.S. represen- tative of a foreign firm, the importer, is responsible and held accountable for safety and efficacy of foreign-manufactured ingredients used in their finished product.
Enforcement Actions
The Food, Drug, and Cosmetic Act of 1938 went beyond inspectional author- ity and action and delegated certain authority for enforcement to the FDA. This gave the agency authority for seizure, injunction, civil penalties, and criminal prosecution, or import and export restrictions for certain prod- ucts. FDA enforcement actions may, however, only be applied to certain acts, with the most common being production or delivery of adulterated or misbranded product into interstate commerce or of adulterating or misbranding the product once it is in commerce, refusing to permit an
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inspection, failing to register a manufacturing facility, and adulterating or removing labeling. Adulteration and misbranding require further defini- tion as these acts apply to biopharmaceuticals, but not every nuance can be listed here. Adjectives used in specific definitions of adulterated include: putrid, filthy or decomposed, lacking indicated strength, quality or purity, out of cGMP compliance, or having a deficient container. Phrases used to define misbranded labeling are: false or misleading, failure to list essential elements such as name of drug or manufacturer and directions for use, and directions that result in a dangerous situation when followed. The point is clear and the public agrees that products meeting definitions of adulter- ated or misbranded should be pulled from the market. Consider now how this discussion on FDA inspection and enforcement directly relates to dis- cussions regarding regulatory operations, quality systems, manufacturing and control, and nonclinical or clinical studies.
The FDA may only bring to bear enforcement actions if the product is introduced into interstate commerce. Courts have interpreted the terms adulterated, misbranded, and interstate commerce quite broadly and it is virtually impossible for a firm or even a university or institution to avoid compliance with FDA regulations. So, what might the FDA do if the sponsor fails to correct deficiencies uncovered by the FDA? There are many possi- bilities but those most commonly used are an enforcement letter or warning letter to the sponsor, forced recall of the product, and judicial enforcement. Debarment or disqualification from participating in the FDA regulated activities is an option when responsible individuals are identified. It is not unusual for the FDA to take two or more of these actions before resolving a case.
• Debarment: It is imposed when action is sought against individuals. For example, an officer of a biotechnology firm may be debarred from working in the regulated industry for a period of time. The investigator of a clinical study or the director of a nonclinical may be debarred from conducting further studies to support the develop- ment of a regulated product after proven egregious behavior.
• Letters: For OAIs or when VAIs are not resolved to the satisfaction of the FDA, the sponsor is sent a strongly worded warning letter in which the FDA states the case against the biopharmaceutical firm. The letter is addressed to an individual, usually an executive, at the firm. The agency then posts this letter at http://www.fda.gov for public access. Letters also list additional enforcement action, including possible criminal action that could be taken against the firm or high-level individuals at that firm unless the matter is resolved to the satisfaction of the FDA. Not surprisingly, several issues are resolved to FDA’s satisfaction shortly after a warning letter is issued, the ultimate step in ensuring compliance through administrative means.
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The FDA always has at its disposal judicial actions. The FDA, like many other regulatory agencies, is not alone authorized to bring enforcement action but must use judicial tools in conjunction with the Department of Justice. Judicial Enforcement is reserved for situations that cannot be resolved or those in which public health or safety of individuals is at risk. FDA may apply the following judicial actions:
• Seizures or Recalls: The FDA may send federal marshals to a plant with instructions to seize all product and the FDA may then order the company to announce and to pay for a complete recall of all product: sold, on the shelf or in distribution.
• Injunctions: Injunctions—temporary, preliminary, or permanent—are legal tools used to keep a party from doing something or to proactively make them do something. For example, an injunction on a manufactur- ing facility may prevent, by law and legal enforcement any employee and/or management to manufacture, process, pack, label, hold, or dis- tribute product in a facility that produces product that has been called into question by the FDA authorities. Unlike an administrative action, injunctions carry the force of criminal penalties. Consent decrees of permanent injunctions may result and they can remain in effect forever or they can expire on a particular date.
• Criminal prosecution: It really happens, firms and individuals will go so far as to face criminal prosecution over a disagreement with the FDA. Others flee the country before they can be prosecuted. The Department of Justice is always involved and there is coordination with the FDA’s Office of Criminal Investigations. The capstone to this process is that criminal prosecution involves strict liability, which means that a corporate officer need not commit the act or even know that a specific act was committed. Prosecution can rest upon failure of a responsible individual to seek out and remedy when situations occur or have occurred. In most cases of criminal prosecution, the FDA involves officers of a firm and may or may not involve technical operators or supervisors. Food for thought.
Product Liability
The biopharmaceutical firm must also be concerned about another legal issue involved in possible adulteration or misbranding issues. Product liability, or other civil actions related to poorly designed products, incorrect manufac- ture or control, or inadequate or misleading clinical or nonclinical studies that result in harm to a private party, such as the user of a biopharmaceutical, can result in civil actions. Civil suits are commonly pursued in the United States and it does not require FDA action for a biotechnology firm or for the officers of that firm to end up in a court of law accused of selling bad product or of putting a human at risk.
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Compliance with Non-FDA Regulations: International, National, State, and Local
When we think of compliance in biotechnology operations, most people have a mental image of meeting regulations of the Food and Drug Administration. While U.S. FDA compliance is important to most biotechnology firms, some will never need to consider 21 CFR. Yet every company will face non-FDA compliance issues. These issues can arrive with little warning and they can have a tremendous and, unfortunately, negative impact on operations. For example, virtually every biotechnology firm ships biologicals and chemi- cals across state lines and international borders. Shipping such materials is highly regulated by several agencies at the national level and perhaps also at the state level. Another example is disposal of waste generated in labora- tories and during nonclinical and clinical studies.
This chapter provides an overview of regulatory compliance situations that are frequently encountered by biotechnology firms in the United States and yet do not fall under the purview of U.S. FDA. The subject matter has been organized under headings related to a particular activity, but the reader will find that a single activity may be regulated by two or more agen- cies at the local, state, and federal levels.
International and Foreign National Regulatory Authorities for Medical Biotechnology Products
National interests and international political differences can be major hurdles to multinational regulatory approval of biotechnology products. Attempts are underway by both regulatory agencies and biopharmaceutical firms to eliminate these differences through transnational harmonization and, for much of the world, by strengthening national regulatory authorities (NRAs) in some countries. While these efforts may not bring every national agency into agreement, they are making a difference in many international biotech- nology markets, notably for countries consuming the greatest amounts of biopharmaceuticals. Despite the lack of movement by some nations, NRAs are generally moving in the direction of harmonization. Today, it is pru- dent to assume the fastest route to multinational approval for biotechnology products in a multinational marketplace is through the early application of harmonized documents.
Organizations encourage and provide guidelines for harmonization. For example, the International Federation of Pharmaceutical Manufacturers and Associations (IFPMA; http://www.ifpma.org), a trade organization, and several NRAs are working to harmonize international regulations through the International Conference on Harmonization (ICH). ICH, a nonprofit group mentioned throughout this book, promulgates harmonized guidelines on
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various subjects applied to biopharmaceutical development (http://www. ich.org). ICH topics are divided into four major categories and ICH topic codes are assigned according to these categories: (1) Q, or quality topics, are those relating to chemical and pharmaceutical quality assurance (e.g., qual- ity control test validation and stability testing); (2) S, or safety topics, are those relating to in vitro and in vivo preclinical studies (e.g., carcinogenicity testing); (3) E, efficacy topics, are those relating to clinical studies in human subject (e.g., dose response studies, Good Clinical Practices); and (4) M, multidisciplinary topics, are cross-cutting topics, which do not fit uniquely into one of the aforementioned categories (e.g., medical terminology, or elec- tronic standards, and the common technical document, or CTD). A partial list of ICH documents relevant to biotechnology product development is in Box 4.4. Most ICH guidelines are accepted by the FDA and so they are a particularly helpful guidance for ensuring safe and effective biopharmaceu- ticals enter the U.S. marketplace.
The FDA also provides at its website hundreds of additional guidance documents for virtually every aspect of biotechnology product develop- ment (http://www.fda.gov/regulatoryinformation/guidances/default.htm). Examples of FDA guidance documents are listed in Box 4.5. It is worth not- ing that the U.S. Federal government regulations, those for all agencies, can be searched online by keyword or numeric citation at the Government Printing Office’s electronic Code of Federal Regulations website, http:// www.ecfr.gov/.
The WHO also certifies the quality of products in international com- merce and provides international standards for nonclinical and clini- cal testing of drugs and biologicals (http://www.who.int). However, the WHO website is complex and it can be difficult to identify specific guide- lines. As a general rule, WHO guidelines and standards are neither as detailed nor as stringent as those of developed countries. The WHO guid- ance on pharmaceutical development is an example (http://www.who. int/topics/pharmaceutical_products/en/). WHO also provides guidelines for clinical trials, including an international registry of clinical research and templates for study documents, and biological product development, including those for nonclinical safety testing, formulation, distribution, and purchase.
European and Japanese regulations continue to have, in these countries and much of the rest-of-the-world, a direct impact on development of biopharma- ceuticals. While each member of the European Economic Community (EEC) has a national regulatory authority and national regulations, harmoniza- tion within the EEC is being led by the European Medicines Agency (EMA), responsible for scientific evaluation of medicines for use in the EEC, and the Committee for Human Medical Products (CHMP), responsible for prepar- ing opinions on questions concerning medicines for human use and cen- tralized marketing assessment and authorizations. Additional information
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BOX 4.4 EXAMPLES OF ICH GUIDELINES USED IN BIOTECHNOLOGY DEVELOPMENT
• Quality • Q1A—Stability testing of new drug substances and products • Q2(R1)—Validation of analytical procedures • Q3A(R2)—Impurities in new drug substances • Q5A(R)— Viral safety of biotechnological products derived
from cell lines of human and animal origin • Q5D— Derivation and characterization of cell substrates used
for production of biotechnological/biological products • Q6B— Specifications: Test procedures and acceptance crite-
ria for biotechnological/biological products • Q7— Good manufacturing practice guide for active phar-
maceutical ingredients • Q8(R2)—Pharmaceutical development • Q9—Quality risk management • Q10—Pharmaceutical quality system
• Safety • S1B—Testing for carcinogenicity of pharmaceuticals • S2—Guidance on genotoxicity testing and data interpretation • S4—Duration of chronic toxicity testing in animals • S6— Preclinical safety evaluation of biotechnology-derived
pharmaceuticals • S9—Nonclinical evaluation for anticancer pharmaceuticals
• Efficacy • E2A—Clinical safety data management • E3—Structure and content of clinical study reports • E6(R1)—Good clinical practice • E8—General considerations for clinical trials • E9—Statistical principles for clinical trials
• Multidisciplinary • M1—Medical dictionary for regulatory activities (MedDRA) • M2—Electronic standards for the transfer of regulatory
information • M3—Nonclinical safety studies • M4—Common technical document
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BOX 4.5 EXAMPLES OF U.S. FDA GUIDELINES FOR DEVELOPMENT OF BIOPHARMACEUTICAL PRODUCTS
• Chemistry, Manufacturing, and Controls (CMC) • Points to consider in the characterization of cell lines used
to produce biologics. FDA. August 1993. • Points to consider in the manufacture and testing of
monoclonal antibody products for human use. February 1997.
• Content and format of chemistry, manufacturing, and controls information and establishment description infor- mation for a vaccine or related product. FDA. January 1999.
• Drugs, biologics, and medical devices derived from bio- engineered plants for use in humans and animals. FDA. September 2002.
• INDs for Phase 3 and Phase 3 studies—Chemistry, manu- facturing, and controls information. FDA. May 2003.
• Comparability protocols—Protein drug products and bio- logical products—Chemistry, manufacturing, and controls information. FDA. September 2003.
• Sterile drug products produced by aseptic processing— Current Good Manufacturing Practice. FDA. September 2004.
• CGMP for Phase 1 investigational drugs. FDA. July 2008. • Process validation. General principles and practices.
Guidance to industry. November 2008. • Assay development for immunogenicity testing of thera-
peutic proteins. December 2009. • Characterization and qualification of cell substrates and
other biological materials used in the production of viral vaccines for infectious disease indications. FDA. February 2010.
• Analytical procedures and methods validation for drugs and biologics. FDA. July 2015.
• Recommendations for microbial vectors used for gene ther- apy. FDA. October 2015.
(Continued)
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BOX 4.5 (Continued) EXAMPLES OF U.S. FDA GUIDELINES FOR DEVELOPMENT OF BIOPHARMACEUTICAL PRODUCTS
• Quality • Medical device quality systems manual. December 1996. • Labeling for human prescription drug and biological
products—Implementing the new content and format requirements. January 2006.
• Quality systems approach to pharmaceutical CGMP regu- lations. FDA. September 2006.
• Q10—Pharmaceutical quality system. April 2009.*
• Request for quality metrics. FDA. July 2015. • Clinical
• Structure and content of clinical study reports. July 1996.*
• Protocol development guideline for clinical effectiveness and target safety trials. July 2001.
• Guidance for clinical investigators, sponsors, and IRBs: Adverse event reporting to IRBs—Improving human sub- ject protection. FDA. January 2009.
• Adaptive design clinical trials for drugs and biologics. FDA. February 2010.
• Early clinical trials with live biotherapeutic products: Chemistry, manufacturing, and control information. FDA. September 2010.
• Guidance for clinical investigators, sponsors, and IRBs: Investigational new drug applications (INDs)— Determining whether human research studies can be con- ducted without an IND. FDA. September 2013.
• Guidance for clinical investigators, industry, and FDA staff: Financial disclosure by clinical investigators. FDA. February 2013.
• Oversight of clinical investigations—A risk-based approach to monitoring. FDA. August 2013.
• Considerations for the design of early phase clinical trials of cellular and gene therapy products. FDA. June 2015.
(Continued)
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regarding exact responsibilities of various agencies within the EEC is given at the EMA website at http://www.emea.europa.eu. The Japanese Ministry of Health, Labor and Welfare, website (http://www.mhlw.go.jp/english/; Pharmaceuticals and Medical Devices tab), provides regulations and guide- lines for biopharmaceutical products marketed in Japan and this regulatory guidance also influences regulatory agencies of countries of East Asia and the Pacific region. Australia (Therapeutic Goods Administration, http:// www.tga.gov.au) and Canada (Health Canada, http://www.hc-sc.gc.ca) have strong regulatory infrastructure for biomedical products, as well. In addi- tion, many trade organizations, such as IFPMA, the Biotechnology Industry Organization, BIO, (http://www.bio.org), and the Pharmaceutical Research and Manufacturers of America (PhRMA), (http://www.phrma.org) actively support national regulatory authorities, ICH and the process of harmoniza- tion, and international guidance, since it is good business and good regula- tory practice for member firms.
Finally, marketing approval is important if biotechnology products are to be used in developing countries, many burdened with much of the world’s infectious diseases. This is especially important for drugs and biologicals
BOX 4.5 (Continued) EXAMPLES OF U.S. FDA GUIDELINES FOR DEVELOPMENT OF BIOPHARMACEUTICAL PRODUCTS
• Preclinical • Formal meetings between the FDA and sponsors or appli-
cants. FDA. May 2009. • Nonclinical safety studies for the conduct of human clini-
cal trials and marketing authorization for pharmaceuti- cals. FDA. January 2010.*
• Process validation: General principles and practices. FDA. January 2011.
• Rare diseases: Common issues in drug development. FDA. August 2015.
• Product development under the animal rule. FDA. October 2015.
* FDA Guidelines based upon ICH Guidelines.
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developed to treat diseases such as AIDS and malaria. As noted earlier, EMEA, FDA, and the drug regulatory agencies of other developed coun- tries are influential in the development and regulation of products for global health. Also, the WHO and several nongovernmental organizations/ agencies (NGOs), are actively involved in searching for means to gain regu- latory approval in countries that do not currently have national regulatory authorities or lack a science-based review system for biopharmaceuticals.
Transporting Infectious or Otherwise Hazardous Materials
Successful transportation, national and international, of materials and products is important to any biotechnology firm. The transportation com- munity, such as national and international shippers, and transportation regulatory agencies, like the U.S. Department of Transportation (DOT) are diligent about the materials acceptable for shipment (http://www.dot. gov). Proper shipping procedures for hazardous and infectious materials are enforced because they protect employees of the shipping firms, ensure public health, and allow all compliant firms to transport a variety of mate- rials, some considered hazardous. While most biologicals and chemicals can be shipped, many require special precautions in packaging, labeling, and handling.
The shipper bears virtually all responsibility for ensuring safe shipping of infectious or otherwise hazardous materials. Therefore, the shipper must be aware of the various and sometimes complex regulations and then properly classify, identify, package, mark, label, and document the substance being shipped. References to common shipping regulations or guidelines for the United States are listed in Box 4.6. The International Air Transportation Association (IATA) is a clearinghouse for international air transport and for- eign, national regulations (http://www.iata.org). By regulation, the shipper of dangerous goods must be a trained person and they must comply with regulations and also certify that materials will arrive at their destination in good condition and not present any hazards to humans and animals dur- ing shipment. Commercial carriers refuse to accept any package that fails to comply with international, national, and the shipper’s regulations or guidelines. Failure to comply with shipping regulations often means that a material does not reach its destination and, for spills or human exposure to infectious, chemical, or radiological substances, can result in substantial fines for noncompliance.
Biological materials are especially important to the biotechnology indus- try. They are considered as (1) infectious (etiologic) agents; (2) diagnostic (clinical) specimens; or (3) biological products. Infectious substances are those known or reasonably expected to contain pathogens. Pathogens are microorganisms (including bacteria, viruses, rickettsia, parasites, and fungi) or recombinant microorganisms (hybrid or mutant) that cause infectious disease in humans or animals. This includes (1) all cultures containing or
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suspected of containing an agent which may cause infection; (2) human or animal samples that contain such an agent in quantities sufficient to cause infection should an exposure to them occur due to a transport mishap; (3) sample from a patient with a serious disease of unknown cause; and (4) other specimens not included earlier but designated as infectious by a qualified person, for example, a physician.
Diagnostic specimens are any human or animal material including, but not limited to, excreta, secreta, blood and its components, tissue or tissue fluid, being transported for diagnostic or investigational purposes, but excluding live infected animals. Diagnostic specimens resulting from medical prac- tice and research are not considered a threat to public health. An example is a serum sample not suspected of containing an infectious agent that is shipped to a laboratory for routine testing.
BOX 4.6 SOURCES OF INFORMATION ON TRANSPORTATION OF BIOTECHNOLOGY
MATERIALS OR PRODUCTS
• Transportation within the United States • Hazardous materials regulations, 49 CFR Parts 171–178, U.S.
department of transportation • Interstate shipment of etiologic agents, toxins, radiologic agents,
42 CFR Part 70–75, U.S. public health service, centers for disease control and prevention (CDC)
• Occupational exposure to bloodborne pathogens, 29 CFR Part 1910.1030, The department of labor, occupational safety, and health administration
• Visit the website of the intended shipping firm, FEDEX, USPS, or UPS, for guidance. • International transportation
– Recommendations of infectious substances and diagnostic speci- mens, United Nations. http://www.who.int/csr/emc97_3m. pdf
– Technical instructions for the safe transport of dangerous goods by air, International civil aviation organization (ICAO)
– International air transportation association (IATA). http:// www.iata.org
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Biological products may have special licensing requirements. These speci- mens are further defined as those products derived from living organisms that are manufactured and distributed in accordance with the requirements of national governmental authorities. They are used either for prevention, treatment, or diagnosis of disease in humans or animals, or for development, experimental or investigational purposes and include, but are not limited to, finished or unfinished products such as vaccines and diagnostic products. This general definition would include many biotechnology products. Hence, diligence and thorough research of any and all guidelines is required by any biologics manufacturer prior to shipment. As noted earlier, import or export of infectious agents is highly controlled. For infectious materials and vectors imported from foreign countries, there are requirements for importa- tion permits and shipping labels issued by the U.S. Public Health Service and posted on the website of Centers for Disease Control and Prevention (CDC), U.S. Department of Health and Human Services (http://www.cdc.gov).
Biotechnology firms also ship a variety of chemical substances and dan- gerous goods, defined as a substance capable of posing an unreasonable risk to health, safety, or property when transported by commercial carrier or by air, or identified as explosive, corrosive, flammable liquid, oxidizer, or com- pressed gas. Indeed, one of the most often overlooked chemical dangerous substances transported by biotechnology firms is solid carbon dioxide, or dry ice. Several steps must be taken before a chemical substance is transported. First, the shipper must determine if the good can be shipped by commer- cial carrier. If so, then written guidelines are followed for packaging, mark- ing, and declaring the good. A Dangerous Goods Declaration is an essential part of every shipment and it appears outside, on the carton, along with clear labeling as to contents, risk and response in case of spill or carton damage.
Importing, Possessing, or Transferring Controlled Biotechnology Materials
The Animal and Plant Health Inspection Service (APHIS), USDA, regulates the importation, possession, and transfer of many controlled plant and ani- mal materials (http://www.aphis.usda.gov). Specifically, under the regula- tion defining the scope of regulated materials (7 CFR 340), the Biotechnology Regulatory Services (BRS) division of APHIS is responsible for importation, interstate movement, and field release of genetically-engineered plants. The BRS website (http://www.aphis.usda.gov/biotechnology) is the most com- plete and up-to-date source of information for those engaged in agricultural biotechnology. While many agricultural pests, actual or potential, are obvi- ous, we must also consider that even if a material is not to be used for agricul- ture it could, in the eyes of USDA, be considered a pest to agriculture. Hence, through APHIS and BRS, importation is controlled for virtually all animal- and plant-origin materials and animal products and byproducts as well as biological materials that contain or have been in contact with materials of
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animal origin. The USDA also controls selected microbial agents that could pose a risk to animals or plants in this country. This includes in vitro mate- rials, such as cell lines. Requests for permits authorizing the importation of such controlled materials must be submitted to APHIS and importation can- not commence until the application is approved.
APHIS regulates genetically engineered plants by administering the Federal Plant Protection Act and the Lacey Act. This legislation authorizes APHIS to control interstate movement, imports to the United States, and release (for field testing) of “organisms and products altered or produced through genetic engineering, which are plant pests or for which there is rea- son to believe are plant pests” (CFR, 2011). A plant pest is a risk to other plants and ecosystems. The term is generally applied to weeds, insects, diseases, or untested genetically modified organisms (GMOs). GMO release in the United States is discussed later. Applying the term plant pest to a genetically engi- neered plant means only that the nonpest nature of the plant has yet to be demonstrated. APHIS requires a permit and concurrence of individual state departments of agriculture for movements across state lines. For field test- ing of a new plant, referred to as environmental release, a permit may also be required from APHIS. For selected plants, one of two other processes, the notification process or the petition process, may be used in place of a permit.
Today, many firms are developing genetically engineered plants that pro- duce drugs or biological compounds intended for medical or veterinary treatments. The FDA has responsibility for regulating the active ingredients produced by these plants. APHIS ensures engineered plants do not pose a significant plant pest risk, a risk to threatened and endangered species, or a risk to people working with them. An APHIS permit is required to take such plants to the field so as to reduce the risk of harm to other organisms, to evaluate any special considerations for containment, and to prevent such plants from entering the food supply.
APHIS is also responsible for protecting animals important to agriculture. This agency facilitates international trade, monitors health of animals before they enter the United States, and regulates the import and export of animals, animal products, and biologics. It is in the import and export area that the National Center for Import and Export (NCIE) has the greatest interaction with the biotechnology industry. Generally, a USDA veterinary permit is needed for import of nonhuman materials derived from animals or exposed to animal-source materials. A wide range of materials, for example, animal tissues, RNA/DNA extracts, hormones, anti-sera, and monoclonal antibodies for in vivo use, are regulated.
USDA also regulates the care and use of laboratory animals (http://www. usda.gov). Any biotechnology firm that does business with a research animal breeder or vendor or itself houses or uses animals is familiar with this exten- sive set of regulations.
The U.S. Fish and Wildlife Service, part of the U.S. Department of Interior, enforces possession or transfer of certain species (e.g., endangered birds or
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primates) or any part of those species (e.g., feathers, eggs, blood, or tissue) under the Endangered Species Program for the Endangered Species Act (http://fws.gov/endangered). Permits are required to transport or hold spec- imens and the law is enforced at U.S. borders by CBP, part of the Department of Homeland Security.
The Bureau of Industry and Security (BIS), of the U.S. Department of Commerce, formerly the Bureau of Export Administration, oversees U.S. exports of dual-use commodities, technology, and software (http://www.bis. doc.gov). The Bureau has the lead role in both export licensing process and enforcement operations. Their mission, based upon national security, is to control exports of sensitive products to entities that could misuse U.S. technol- ogies and products. BIS licenses exporters of certain products, including bio- technology-related products such as fermentation equipment. In doing so, it requires exporters to notify other parties of the sale and the conditions of sale and to obtain written acknowledgment from the end-user of the intended use. Licensing conditions are sometimes necessary to make certain that approved items are in the correct location and being used in an appropriate manner. This Bureau does not regulate all goods, services, and technologies but it does control the export of certain microorganisms, toxins, and equipment used to make these items. The items are provided in the Commerce Controlled List (see the Regulations tab at http://www.bis.doc.gov). To further complicate matters, other U.S. government agencies regulate more specialized exports. For example, the U.S. Department of State has authority over defense articles and defense services. A list of other agencies involved in export controls can be found on the website for BIS.
The Public Health Security and Bioterrorism Preparedness and Response Act of 2002
This law has certainly complicated movement or use of many biotechnology products within the United States and across its borders but it has also enhanced security while allowing legitimate biotechnology efforts to continue. It includes sections on “Enhancing Controls on Dangerous Biological Agents and Toxins,” providing for the regulation of biologicals specified by the Department of Health and Human Services and the Department of Agriculture. It recom- mends interagency coordination between the two departments regarding control of overlapping agents and toxins and provides for criminal penalties regarding certain biological agents and toxins. The CDC has primary responsi- bility for implementing the provisions of this Act (http://www.cdc.gov). APHIS is the agency fulfilling roles designated to USDA.
The USDA Regulations are within 7 CFR and 9 CFR while the CDC regula- tions are 42 CFR 73. In general, the regulations are aimed at animal and plant agricultural and human health threats, respectively, but there is some over- lap. The regulations establish and enforce safety procedures for listed agents and toxins, including
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• Measures to ensure proper training and appropriate skills to handle agents and toxins, and proper laboratory facilities to contain and dispose of agents and toxins
• Safeguards and security measures to prevent access to listed agents and toxins for use in domestic or international terrorism or for any other criminal purpose
• Procedures to protect animal and plant health, and animal and plant products, in the event of a transfer or potential transfer of a listed agent or toxin in violation of the safety procedures, as well as safe- guards and security measures
• Appropriate availability of biological agents and toxins for research, education, and other legitimate purposes
The regulations themselves cover requirements for registration, security safety and emergency response plans, training, transfer, record keeping, inspections, and notifications. They regulate molecular parts of organisms, since particular genes or proteins from these organisms might also con- stitute a risk to public health. Even a small amount of nucleic acid from a select agent may be regulated. A permit system allows a research investiga- tor or biopharmaceutical product developer to import, keep, transfer, or test (e.g., field test a genetically engineered plant) an agent. Since the penalties for improper or illegal possession, use, or transfer of the agents are severe, those biotechnology firms using even seemingly safe and innocuous agents or molecules should become familiar with the select agent list and the regu- lations well before they consider transferring the material to their labora- tory. Noteworthy is the fact that this and other Acts often contain unusual or unexpected and sometimes confusing or conflicting clauses. Examples of issues from the Act of 2002 are provided in Box 4.7.
Importation or Exportation of Biotechnology Products for the Purpose of Treatment of Diseases in Humans
FDA regulations, as discussed in the section “Compliance for Import of Biopharmaceuticals into the United States” define requirements for import or export of any “virus, therapeutic serum, toxin, antitoxin, or analogous product” for the “prevention, treatment, or cure of diseases or injuries of man” (CFR, 2015). It is important to note the roles customs and other Federal agencies play in the importation and exportation of these products, since the scope of regulations encompasses many biotech- nology products. Labeling requirements are absolute since they inform customs and other government officials. If a product is intended for human use, then it must be labeled and may be inspected and sampled by CBP (http://www.cbp.gov). FDA may be contacted and may even inspect the shipment. If there is no evidence that the product is licensed
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BOX 4.7 A DISCUSSION ON THE SELECT AGENTS AND PUBLIC HEALTH SECURITY AND BIOTERRORISM
ACT OF 2002 (PUBLIC LAW PL107-188) AND THEIR IMPACT ON BIOTECHNOLOGY OPERATIONS
• In 2002, Congress passed the public health security and bioter- rorism act, identifying the need to keep a list of select agents and toxins. Select agents (biological agents or toxins) were specifi- cally identified, or declared by USDA and Department of Health and Human Services. The U.S. Center for Disease Control and Prevention maintains and changes the list, approves laborato- ries qualified to transfer, use, or control select agents and regu- lates, to a certain extent, practices in those laboratories. A full and current list of select agents is at http://www. selectagents. gov. Some examples are botulinum neurotoxins, SARS- associated coronavirus, certain hemorrhagic fever viruses, Shiga toxin, Rift Valley fever virus, Xylella, a bacterial pathogen of citrus crops, swine flu virus, and avian influenza virus. Most would agree that we do not want just anyone, and certainly not our residential neighbors, holding these agents.
• Congress also mandated that certain individuals, including among others those adjudicated as mentally defective or (hav- ing) been committed to any mental institution, and/or those hav- ing been discharged from the Armed Forces of the United States under dishonorable conditions, and/or those determined to be unlawful users of the Controlled Substance Act (21 USC 802). In the United States, there are many citizens who have at one time been in a mental institution, discharged dishonorably or used a controlled substance, to include LSD, codeine, cocaine or mari- juana, as defined by the Department of Justice, Drug Enforcement Administration, Officer of Diversion Control. Current informa- tion can be identified at http://www.deadiversion.usdoj.gov
• The purpose of this discussion is not to argue or in any way malign the Act of 2002; it was well-meant and passed by knowledgeable legislators and the president. The purpose is to stress how very important it is for a professional working in biotechnology to be familiar with a host of detailed public laws and regulations before embarking upon a laboratory or devel- opment project. Further, the professional should consult with regulatory authorities or legal counsel whenever perceived issues or conflicts arise.
(Continued)
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by FDA, then it is held by CBP. If the product is of animal origin (e.g., a horse antiserum against snake venom), it may require a USDA permit, as well. If any part of the product is from an endangered species, it will also need a permit from Fish and Wildlife Service under the Endangered Species Program. In effect, CBP serves as a gatekeeper at the U.S. borders, acting on behalf of several federal agencies. In summary, importation and exportation of all biotechnology materials and products must be carefully researched by the shipper with the expectation that the regulations of multiple federal agencies could complicate, delay, or stop the movement of these goods.
BOX 4.7 (Continued) A DISCUSSION ON THE SELECT AGENTS AND PUBLIC HEALTH SECURITY AND
BIOTERRORISM ACT OF 2002 (PUBLIC LAW PL107-188) AND THEIR IMPACT ON BIOTECHNOLOGY OPERATIONS
• Now, in light of terms of the Act of 2002 identified earlier, consider in three examples, provided in the following, YOUR responsibility as a manager or supervisor at a biotechnol- ogy firm when a select agent is handled by your employees.
1. My operation handles, under proper conditions, recom- binant Corona virus to infect animals for testing Corona virus vaccines. I know some employees are veterans of the armed forces; must I determine if each veteran was honorably discharged?
2. We will use small amounts of botulinum toxin in a mouse model to develop QC assays to test our monoclonal antibody, intended to neutralize the toxin. Although Botox® is currently a product licensed for medical use and widely available in doc- tor’s offices, must I query each employee to ensure that no per- son was at any time committed to a hospital mental ward, if even for one day and decades ago?
3. Our laboratory is studying protein expression of Xyella in an effort to develop antibiotics and combat this disease of citrus crops. Must I query all employees to ensure that no one has ever used marijuana or possessed or taken a cough remedy or another medicine containing codeine?
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Occupational Health and Safety
A biotechnology firm must have effective health and safety policies and practices for one simple reason: it protects employees, their most valuable asset. We all know that work can affect our health and, when queried, people state that good health is a leading factor in quality of life. If a workplace is safe, people enjoy their jobs and are more interested and involved in their employment.
The biotechnology laboratory work environment includes hazards but it need not be unsafe. We work with harsh chemicals, acids, corrosives, radio- chemicals, and biological agents such as viruses and toxins. In research labo- ratories, individuals are often in close contact with these materials and, in biopharmaceutical manufacturing, there may be large volumes of potentially hazardous materials in the workplace. The work environment may also have carcinogens, flammable gases or liquids, steam and hot fluids.
As with just about everything in biotechnology, the key to providing a safe and comfortable work environment is good planning. Every biotechnology firm should have a health and safety policy and plan and procedural docu- ments, all receiving the full support of upper management and line supervi- sors. Good policies emphasize prevention rather than reaction to incidents or accidents. Standards for health and safety are based upon risk assess- ments and regulatory requirements. Health and safety plans state objectives or goals and standards or specifications. The results, based upon measur- able outcomes, are compared at regular intervals against health and safety objectives.
Biotechnology firms should have a visible organization or structure to sup- port a health and safety plan. An environmental health and safety specialist is the individual responsible for developing, implementing, and monitor- ing industrial safety programs within the biotechnology company. While a smaller firm may not require a full-time health and environmental safety offi- cer, a consultant, such as an occupational safety specialist, is an important member of the corporate team. These professionals inspect laboratories and product development, manufacturing and testing areas to ensure compliance with federal Occupational Safety and Health Administration, state, and local regulations and corporate policies. They evaluate new equipment and raw materials for safety, and monitor employee exposure to chemicals and other toxic substances. A safety specialist also conducts training programs in haz- ardous waste collection, disposal, and radiation safety.
Finally, management must make every effort to encourage a safe and healthy culture within the biotechnology firm. Communication is an important part of the process, with periodic seminars and, most impor- tantly, an effective means for employees to express their health and safety concerns to management. A safety training program gives employees an opportunity to learn more about safety as it relates to their particular job assignments.
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Local, state, and federal agencies, notably OSHA of the Department of Labor, regulate health and safety in the workplace (http://www.osha.gov). As early as 1985, OSHA began to examine the health and safety issues related to biotechnology. OSHA originally felt that no additional regulations were needed for such workplaces since other standards, such as those for general laboratory safety, provided an adequate basis for protection and safety. These OSHA regulations and standards are at 29 CFR 17. In addition, blood-borne pathogen guidelines, which apply to all occupational exposure to blood or other potentially infectious materials, and exposure to other infectious organisms are responsibilities of both OSHA (http://www.osha.gov/SLTC/ biologicalagents/index.html) and the National Institute for Occupational Safety and Health (NIOSH), Centers for Disease Control (http://www.cdc. gov/niosh/homepage.html).
In summary, one of the most important aspects of safety, from the point of view of the employee in a biotechnology firm, is to have a clear under- standing of hazards in their workplace. Simple acts, such as participation in training, wearing safety glasses, proper disposal of waste, and review of material data safety information are operational keys to health and safety in the biotechnology work environment. A health and safety plan and effective training can go a long way to reach these objectives.
Environmental Regulations in Biotechnology
There are many environmental hazards in biotechnology operations and there also are numerous federal, state, and local regulations as well as agen- cies to enforce them. Failure to heed environmental guidelines by a firm puts both the company and community at possible risk. These issues, or even perceived problems, are often highly publicized within a state or com- munity and, when a violation becomes known, it creates a negative image of the biotechnology industry as a whole. To make matters worse, there already are concerns, worldwide, about the release of genetically modified molecules or organisms into the environment. And there are the less publi- cized, but still very real issues, related to environmental release of materials from biotechnology laboratories or operations.
A relevant example is the receipt, handling, and disposal of radioiso- topes, functions regulated by the Nuclear Regulatory Commission (NRC), Department of Energy (http://www.nrc.gov/) and by state and sometimes local agencies or other federal agencies. For example, OSH handles certain aspects of exposure to ionizing radiation (29 CFR 1910.1096). NRC manages radioactive materials by controlling the production, shipment, use, and dis- posal of these materials. It does so through licensing responsible entities, such as universities or biotechnology firms. NRC also allows individual states to regulate certain activities through the Agreement State Program. A biotechnology firm wishing to purchase, receive, use or dispose of a radio- isotope must apply for and receive a license and agree to keep careful records,
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train employees, follow detailed rules, accept unannounced inspections, and pay fines for noncompliance.
Sponsors of INDs, BLAs, or NDAs are required to file an environmental impact statement or seek categorical exclusion for each product and under each regulatory filing with FDA.
Biotechnology firms also face complex regulations, many local, deal- ing with the disposal of chemicals and biological substances. With a few exceptions, it is not difficult to dispose of small amounts of nonhazardous chemicals or biological materials. However, in biotechnology operations, notably manufacturing, larger amounts of biological and chemical materials may need to be released into the environment. Examples include disposal of a large mass of recombinant bacteria following fermentation or of large volumes of organic solvents following molecule purification. In most cases, these cannot simply be sent to the local landfill or flushed down the com- munity sewer. Regulations for waste disposal often fall under the purview of the Environmental Protection Agency (EPA) (http://www.epa.gov) but one can expect to find complex and extensive state and local regulations, as well. Indeed, local officials often understand all regulations—federal, state, and local—that apply to their community.
Major regulatory applications, such as IND, NDA, or BLA, must consider environmental issues related to the manufacture and use of each product. National environmental law and policy, notably the National Environmental Policy Act of 1969, drive this requirement. Each of these documents must contain either an Environmental Assessment or the sponsor must state and show that the actions described in the document are categorically excluded from an Assessment. Environmental Assessments can be large and complex documents and they are not required for most biopharma- ceuticals, at least not in early development. However, with some products in the IND stage and with many products in large-scale manufacture, the sponsor must complete an assessment prior to submitting a regulatory application to FDA. The agency provides guidelines for those wishing to choose between claiming exclusion or preparing an assessment.
Genetically Modified Organisms or Molecules
Several federal agencies are responsible for regulating the release of GMOs or chemicals into the environment. The role of USDA has been mentioned. The EPA is also involved because of the Microbial Products of Biotechnology section of the Toxic Substances Control Act, or TSCA. TSCA authorizes EPA to, among other things, review new chemicals before they are introduced into commerce (http://www.epa.gov/opptintr/biotech and www2.epa. gov/regulation-biotechnology-under-tsca-and-fifra). FIFRA, the Federal Insecticide, Fungicide and Rodenticide Act mandates EPA registration of all pesticides, including those derived from biotechnology (e.g., genetically- modified microbial pesticides and herbicide-tolerant crops). Intergeneric
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microorganisms, that is, microorganisms created to contain genetic mate- rial from organisms in more than one taxonomic genus, are considered new chemicals under TSCA and therefore EPA reviews and regulates the use of intergeneric microorganisms in commerce or for commercial research.
Plants and domestic farm animals are genetically engineered in the United States and such endeavors, especially their field testing and com- mercialization, are highly regulated. The USDA is the lead regulatory agency for genetically engineered crop, providing regulatory guidance and licensing of crops, for planting, commercial and investigational, in the United States. BRS at APHIS takes the lead on such matters at USDA and the primary regulation is 7 CFR 340. Environmental regulations, such as those handled by EPA, may also apply. And if a food or a biopharmaceuti- cal is derived, in part or in whole, from a genetically engineered crop, then FDA also provides guidance and may need to give final approval prior to commercialization. Regulations for genetically modified domestic farm (i.e., nonlaboratory) animals, including those used for food or production of biopharmaceuticals, are complex and any given situation must be care- fully researched with special considerations given to guidelines provided by USDA, FDA, and EPA.
The Office of Biotechnology Activities (OBA), Office of the Director, National Institutes of Health (NIH) is involved in activities that affect many biotechnology firms (http://www4.od.nih.gov/oba). These activities include biomedical technology assessment, biosafety, and biosecurity. Examples in genetic research and biotechnology are that OBA: (1) monitors human gene transfer and aspects of stem cell development; (2) manages several review committees for novel technologies; (3) advises other government agencies or departments; (4) develops policies and procedures, reviews established Institutional Biosafety Committees (IBC); (5) provides information to the public; and (6) develops registries of activities.
Although OBA is not chartered by Congress as a regulatory agency, as is the FDA, it has significant influence in several areas of biotechnology and can have an immediate and significant impact on fields of biotechnology, notably emerging technologies, and on biotechnology firms themselves. OBA is influential largely by providing guidelines to the public. A typical action is a decision as to whether an institution, such as a university, may receive NIH funding. For example, if OBA decided that a type of human gene cloning experiment was not appropriate, then any university or bio- technology firm that did such experimentation could lose all NIH funding for any research purpose. Since many biomedical entities, such as insti- tutes, universities, and even some biotechnology firms in the United States, receive NIH funding, OBA guidelines have significant impact. Furthermore, unlike many FDA activities in which the information is held in confidence, OBA activities are largely public and both the press and interest groups often monitor and publicize issues and the biotechnology firms that are involved with OBA.
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OBA promulgates the NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules (NIH Guidelines), which is the premier guidance document for nucleic acid research. It specifies all aspects of genetic engineering and production of GMOs, outlining respon- sible research practices in basic, animal and clinical nucleic acid research. OBA sponsors the Recombinant Advisory Committee (RAC), experts appointed to monitor scientific progress in basic and clinical research involving recombinant nucleic acid and human gene transfer. RAC rec- ommends changes to the NIH Guidelines and its members review human gene therapy protocols.
OBA also manages compliance with the requirements for Institutional Biosafety Committees or IBCs. An IBC must be established by any entity that receives NIH funding and performs genetic engineering. Many biotechnol- ogy firms that do not receive NIH funding also use an IBC for the purpose of reviewing experiments involving genetic engineering or transfer of GMOs to the environment. The primary role of an IBC is to ensure that all recom- binant DNA research conducted at or sponsored by that institution is con- ducted in compliance with the NIH Guidelines, but the roles of IBC have been expanded at many institutions to include other aspects of laboratory research with genetically modified materials or organisms.
OBA maintains databases on technologies under their purview and thus fosters transparency on novel technologies that may be of concern to scien- tists and to the general public.
Somewhat apart from these government agencies, but important to certain biotechnology firms, is the National Science Advisory Board for Biosecurity (NSABB), now hosted by the National Security Agency. NSABB advises U.S. federal agencies on security issues related to life sciences, notably on ways to minimize the possibility that knowledge and technologies emanating from vitally important biological research will be misused in a manner that threatens public health or national security.
Taken together, these advisory committees, while not themselves regu- latory agencies, make recommendations that are very influential to gov- ernmental regulatory agencies and to the biotechnology industry overall. Their impact is particularly aimed at cutting-edge technologies. The pru- dent biotechnology operation, public or privately held, will carefully moni- tor the activities of each committee and be constantly and fully aware of recommendations or guidelines they produce.
International Diligence in Biotechnology Operations
Biotechnology is an international endeavor. Most biotechnology firms expect to sell their products or services overseas as well as in the United States. Sometimes biotechnology research can be completed in a national
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environment, heeding only U.S. requirements but this is a rare situation and most biotechnology firms or endeavors are or will become transnational or intercontinental businesses. A biotechnology firm must diligently plan to incorporate their operation and, of course, their services and products, into international environments and markets. International awareness and com- pliance is therefore especially important to the success of biotechnology firms.
International regulations and guidelines are far too numerous to cover in this chapter. However, a single example, genetic engineering of plants from which food is derived, is given to emphasize the value of understand- ing international, as well as country-specific, information prior to embark- ing on biotechnology product development. Two entities, the Food and Agricultural Organization (FAO) of the United Nations (http://fao.org) and the Biotechnology Organization, BIO (http://BIO.org) are actively involved in efforts to harmonize international guidelines for genetically modified foods or GMOs that provide foods. The biotechnology firm proposing to export recombinant organisms, food produced by recombinant plants or animals, or equipment, supplies and raw materials or services for the production of such products, should consider the guidance provided by and experiences of these organizations or their members.
An example of an international guideline intended to harmonize the movement of genetically engineered foods is the Codex Alimentarius (http://www.codexalimentarius.org/), a collection of internationally adopted food standards presented in a uniform manner. Codex standards are meant to ensure that consumers receive products that meet internation- ally accepted and minimally acceptable quality levels, are safe, and do not present a health hazard in accordance with FAO guidelines. The Codex is written by an international Committee and is approved by a WHO body, the FAO. The priority of the Codex Commission is to protect the health of consumers and ensure fair practices in food trade. One would therefore assume that by following the Codex, a biotechnology firm could trade their product or service worldwide. Unfortunately, this is not the case, since national laws and regulations for genetically modified foods still differ considerably on many points. As with most international guidelines, the Codex guidelines are binding, in a national sense, only when fully rati- fied by all parties. However, most international guidelines dealing with biotechnology products and services are not yet, and some may never be, accepted by all nations.
There are many regulations impacting biotechnology operations and their reach is worldwide. Indeed, regulations outside FDA likely dominate the FDA aspects. It is impossible to imagine a product that is not touched by one or more of the regulatory bodies mentioned in this chapter. A sample problem is provided in Box 4.8 as a means of testing the reader’s knowledge in this area.
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Summary of Regulatory Compliance
Biotechnology activities of all types are highly regulated. FDA regula- tions apply to all aspects of biopharmaceutical research, development, and commercialization. Regulatory compliance, which simply means meet- ing FDA regulations, is a necessary and important aspect to developing a
BOX 4.8 APPLICATION OF NON-FDA REGULATIONS. AN EXERCISE
• This brief exercise is offered to provide an appreciation for the complex analysis that must be performed in an effort to deter- mine which non-FDA regulations and appropriate regulatory bodies, state, local, national, and international, apply to your biotechnology operation.
• Suppose your biotechnology firm wishes to ship from London to Maryland a live, attenuated, recombinant bacterium, derived from an infectious agent. You would then grow, in small amounts, that bacterium in the laboratory and pilot manufac- turing area. This would then be studied as a candidate vaccine in the laboratory, in animals, and then, perhaps, in a Phase 1 study in humans.
• A few situations you must consider are listed below. Please add to the list, to include federal, state, and local governments, expand on the regulatory role of each agency, and cite the regu- lations themselves.
• Export from the United Kingdom: Agencies in the United Kingdom
• Transport within the United Kingdom: Agencies in the United Kingdom
• International transport by common carrier • Import and entry into the United States • Transportation within the United States • Classification as an agent within the United States • Research in the laboratory in the United States • Development in the biomanufacturing facility, the animal test-
ing facility and the clinical facility in the United States • Environmental concerns and disposal in the United States
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biopharmaceutical throughout the entire product development pathway. Regulatory compliance is achieved by operating within systems that are established and followed to ensure product quality and adherence to appli- cable regulations. Quality systems include implementation of best practices such as GMP for manufacturing activities, GLP for preclinical safety evalu- ation, and GCP for clinical activities. FDA takes a systems approach when evaluating an organizational level of compliance, beginning with one sys- tem then expanding to other systems, thereby assessing the overall com- prehensiveness of the quality system. In this way, FDA develops a level of confidence that systems instituted by a sponsor are sufficient to support product safety. Inspections are utilized to evaluate staff, training, documen- tation, material flow, manufacturing and testing procedures, data integrity and interpretation, each of which contributes to achieving and maintaining regulatory compliance. Similar compliance requirements exist for medical devices. There is an increased level of regulatory control and review for those devices posing greater potential risk to the end user. The FDAs’ QSR for medical devices establishes a rigorous system, a powerful tool used to achieve and maintain regulatory compliance. QSR is now generally accepted and applied to the regulation of drugs and biologics. A noteworthy and pow- erful component of regulatory compliance is inspection and enforcement. FDA directed inspections are usually triggered for one of three reasons: (1) required preapproval inspection, (2) routine inspection of a regulated facility, or (3) for cause (FDA, 2016) precipitated by a serious adverse event report, a complaint or follow- up to a previous inspectional deficiency. At the conclusion of an inspection, if a serious noncompliance issue is identified by the FDA, then a list of inspectional observations, commonly referred to as a FDA 483, is issued. An unacceptable response to a 483 can result in enforce- ment action by FDA, such as issuance of a warning letter. Noncompliance or suspected noncompliance weighs heavily on a biotechnology company plac- ing the company at great risk for product liability concerns, negative indus- try perception, and decreased product sales.
The biotechnology is highly regulated industry; therefore any entity devel- oping biopharmaceuticals must be compliant with FDA regulations. This is essential to the viability and success of any biotechnology firm. The most successful approach is to demonstrate quality compliance through a compre- hensive quality systems approach; this is resource intense, and requires sig- nificant resources, regulatory expertise, and extensive checks and balances.
Summary of Non-FDA Compliance
In any given country, several federal agencies, in addition to a food and drug agency, may regulate a single activity or function performed by a bio- technology firm. Hence, international, national, state, and local regulations must be considered, and there can be considerable overlap in regulatory authority. Due diligence and careful planning regarding all regulatory
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compliance is essential to the success of any biotechnology firm. Congress continues to change the laws regarding various aspects of biotechnology operations and at the same time executive agencies change or add regula- tions, while courts interpret application of laws. Adding to the complex- ity of regulatory compliance, any given government agency may change its mission or focus or become overwhelmed with regulatory submissions. As biotechnology becomes more global, the U.S. biotechnology firm must con- sider international guidelines and the regulations of countries other than the United States. To be successful, the biotechnology operation must be aware and diligent of all U.S. and global compliance issues as well as FDA regulations and guidelines.
References
CFR. 2011. Introduction of Organisms and Products Altered or Produced Through Genetic Engineering Which are plant pests or which there is reason to believe are plant pests. CFR Part-340. National Archives and Records Administration.
CFR. 2015. Food and Drugs Chapter I- Food and Drug Administration Department of Health and Human services. CFR Part-600. US Food and Drug Administration.
Federal Register Online. 1996. Medical Devices; Current Good Manufacturing Practice (CGMP) Final Rule; Quality System Regulation. 61(195): 52601–62. http://www.wais.access.gpo.gov (accessed May 31, 2016).
Oxford English Dictionary. Oxford University Press. 1997. University of Oxford, Oxford, UK.
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5 Quality Systems
Overview of Quality in Biotechnology
Quality impacts every aspect of a biotechnology operation. While this might seem like a bold statement, those involved in biotechnology would certainly agree it is true. The requirement for quality in biopharmaceuti- cal development is backed by a host of regulations (Chapters 3 and 4). As applied to biotechnology operations, a state of quality is necessary in all endeavors and quality increases the value of services and products. We refer to quality systems, quality by design (QbD), quality control, and the roles of quality in compliance to name but a few quality terms. Indeed, the word quality has various meanings to different individuals and for each situation in which the word is used, so it is perhaps best defined in the context of each usage. This book tries to do just that. Yet, we need to begin this chapter on quality systems by establishing a basic definition for qual- ity. The reader may take their choice from any one of five definitions given below.
• “Quality. The degree of excellence of a thing; general excellence.” (Oxford English Dictionary, 1997)
• “Quality is the totality of features and characteristics of a product or service that bear on its ability to satisfy stated or implied needs.” (British Standards Institute, 1991)
• “…specified requirements for a product can be stated in terms of an established design…and [where]confidence in product conformance can be attained by demonstration of…capabilities in production.” (International Standardization Organization, 1994)
• “High quality is freedom from defects.” (source unknown) • “Continuous improvement and waste reduction.” (Henry Ford)
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Practical man that Henry Ford he goes; right to the point of saving money to produce a quality automobile at low cost.
Even though 100 people might provide 100 different definitions for qual- ity, a common theme in each definition would be that a product is fit or fit for use, that it performs as intended and, hence, the user is satisfied. But, how do quality and these definitions affect the biotechnology indus- try? First, everyone in the business of biotechnology is either producing a product, such as a patent, a vial of therapeutic medicine or a recombi- nant crop plant, or they are producing a service, such as testing products in animals or humans, manufacturing active ingredients or testing those ingredients. Everyone engaged in biotechnology wants their product to be fit for the intended use and thus to have satisfied customers, it is as simple as that.
In this chapter, the reader is introduced to the concepts and practices of quality, and the terminology associated with this field. A brief history of quality is used by way of introduction. This is followed by a review of the hallmarks of quality, that is, criteria that are common to most of the quality systems used in biotechnology. To finish, we review modern ways of incor- porating quality into biotechnology endeavors, identify how quality systems are generally applied in our industry and provide guidelines for establishing and managing a quality system in a biotechnology firm.
History: Evolution of Quality Concepts and Practices
Although it is not obvious, quality has a long background, outlined in Figure 5.1. Quality is rooted in the ancient history, beginning when indi- viduals made goods for themselves or, through barter, for their neighbors. Since buyer and seller usually lived in the same village, the act of provid- ing a bad product led to a bad reputation for both product and producer, or seller. As it does today, a negative image resulted in loss of business for the producer and it also led to social and economic pressures to make only quality products. With the industrial revolution, manufacturers were at a distance to the buyer and thus there was little face-to-face contact between the parties. New forces ensured quality under this rapidly evolving system. Oral, then written, warrantees were developed. For example, manufacturers began to list themselves as the source of a product and, in some cases such as processed foods, they also provided consumers with the ingredients. If the product was of high quality, this served as a form of advertisement. However, poor quality products led to a negative image and even to con- flicts between the buyer and seller. This led to various quality initiatives
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and, over the past 100 years, a number of ideas for ensuring quality of products and services have been adopted to ease those conflicts. Some of the most familiar are
• Standards for measuring instruments, for example, kilogram weight standards
• Marks or seals, for example, UL Seal of Approval on electrical appliances
Quality
Outside groups certify quality
Late twentieth century
Code of Federal Regulations 21
Part 820 Quality System
Regulation
Work groups focus on quality Today
Quality Assurance Guaranteed to customer
Today
Tradesmen compete on quality of their products
Prior to nineteenth century
Industrial revolution demands group efforts toward quality products
twentieth century
Quality systems identified in written documents
Today
Standard International Quality Systems
Today
ISO 9001 Certified
Quality Management System
FIGURE 5.1 Evolution of quality systems.
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• Supervisory responsibility for quality production on the assembly line, for example, Henry Ford ordering his foremen to be responsible for quality in each of their assembly line areas
• Regulatory quality requirements were codified, such as the Food and Drug Act, to mandate quality in drugs and medical device development
• Worker responsibility for quality production on the assembly line, for example, as touted by automobile manufacturers in the late 1980s
• Statistical trend analysis, for example, medical chart reviews by health insurance firms to ensure that patients received the best health care for the price
• Fresh ideas, such as ISO 9001, Total Quality Management and Six Sigma, as introduced over the last three decades
Therefore, when and how did the concepts and practices of quality come to be applied to biotechnology? Quality was applied to virtually all aspects of biotechnology operations very early, as soon as biopharmaceutical develop- ment began in the 1980s. For what reasons was quality applied to biotech- nology? For business reasons—good quality results in an excellent product or service and this, in turn, leads to healthy sales. For compliance reasons— various regulatory agencies said quality assurance was necessary for most biotechnology products, certainly for those with a potential impact on the health or safety of the user or on public health, in general.
Long before biotechnology endeavors moved from laboratory benches and into development, the pharmaceutical industry had, for these reasons, applied quality concepts to all aspects of drug development. The medical device field was not far behind, adopting slightly different but no less stringent quality systems. Since many biotechnology firms must compete within those industry segments and because they are often regulated by the likes of the U.S. Food and Drug Administration (U.S. FDA) or the U.S. Department of Agriculture, the evolving biotechnology industry had no choice but to be compliant with modern quality standards. Does this mean that most, if not all, biotechnology product devel- opment or services must consider quality? Yes, today quality is a must. How then can a biotechnology firm begin to develop a product or service without first instituting a quality system and how could they continue to develop products or services with a weak quality system? They cannot. Pretty blunt language but it is a fact, you cannot provide services or develop products in biotechnology unless you follow quality systems that are accepted by the consumer and regulatory agencies for that product or service. Most biopharmaceutical firms must follow several quality systems. In summary, compliance with regulations is a major rea- son for using quality systems in biotechnology operations.
Another compelling reason is that quality constitutes good business prac- tice. When a biotechnology firm provides a quality product or a service, cus- tomer satisfaction increases and with it increases sale of the product or service.
153Quality Systems
Under a quality system, the number of complaints is significantly reduced. Within a firm, employee pride and satisfaction are enhanced in a quality envi- ronment and significant cost savings are realized from increased productivity and reduced waste of materials. Quality also means speeding products to the market and keeping them on the market with result as excellent profit.
Quality Systems Approach to Product Development
Biotechnology firms develop and market products. To reach the market place firms adapt or invent various development systems—manufacturing, qual- ity control, or nonclinical studies to name a few—for the purpose of product development. A system that cannot be neglected is a quality system, an orga- nized body of immaterial things, if you will, aimed at ensuring the utmost qual- ity of the product or service. A quality system then takes into consideration the many facets or hallmarks of quality that have been adopted by our industry. Hallmarks of quality are listed in Box 5.1 and discussed further in a later section
BOX 5.1 HALLMARKS OF QUALITY SYSTEMS
Management responsibility
Defined quality system(s)
Quality by design and design control
Contractor, vendor, and consultant control
Product identification and traceability
Process control
Environmental control
Inspection and testing (quality control)
Control and release of material, services, or product
Change control and corrective or preventive actions
Packaging and labeling
Preservation, storage, and handling
Servicing
Customer concerns and adverse event reports
Risk and risk management
Documentation
Training
Auditing
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of this chapter. Hallmarks are immaterial things, tools really, that are applied to ensure a quality product and, hence, user satisfaction.
Several quality systems have been defined, for example, ISO 9001 and current Good Manufacturing Practices (cGMP). Each falls under a guiding authority (e.g., U.S. FDA), each has a specific objective, and each is applied to a particular functional area. For example, cGMP is a quality system applied to the manufacture of biopharmaceutical products while current Good Laboratory Practices (cGLP) are applied to nonclinical safety testing of prod- ucts. Specific quality systems are described throughout this book. Quality systems are sometimes defined by regulatory bodies while others represent consensus within an industry. In the United States, cGMP and cGLP are rec- ognized and enforced by the U.S. FDA while elsewhere international com- mittees (ICH, International Conference on Harmonization) often define or redefine quality systems.
One or more of the following six quality systems is often incorporated into biotechnology operations. Each is briefly described in the following and they will be mentioned in greater detail elsewhere in this book.
• Current Good Manufacturing Practices: cGMPs are regulations used worldwide to ensure the quality manufacture and control of drugs (Chapters 6 and 7), biopharmaceuticals and medical devices, world- wide. Despite attempts to harmonize cGMPs between nations, there are differences in national cGMPs and an international version as well. In addition, cGMPs also have unique guidelines pertaining to manu- facture of special classes or even of unique types of products. For exam- ple, the cGMPs for medical devices considers engineered products and, in some cases, the software to operate those products. Each class of biotechnology product may also have unique aspects of cGMPs; for example, vaccines have several unique manufacturing guidelines and monoclonal antibodies also have special quality features.
• Current Good Laboratory Practices: cGLPs are applied worldwide for evaluating the safety of medical products in nonclinical (in vitro or animal) studies. As is the case with cGMPs, cGLPs are not fully har- monized across countries and there are differences in how they are applied to various product classes. Additional information on cGLPs is provided in Chapters 4 and 8.
• Current Good Clinical Practices (cGCP): cGCPs are regulations used worldwide for evaluating the safety of medical products in clini- cal (human) studies at all phases of development. As compared to cGMPs and cGLPs, cGCP regulations are harmonized across coun- tries, although there are differences in some aspects of this quality system due to various types of products and cultural or political features of national regulations. Additional information on cGCPs is given in Chapters 4 and 9.
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• ISO 9001: ISO, an acronym for International Standards Organization, is an internationally recognized standard for quality of virtually any manufactured product and many classes of services. While it is not often applied for the development of biopharmaceutical prod- ucts, it is a standard quality system for medical device development, manufacture, and control in much of the world. Additionally, many service providers to the biotechnology industry are ISO 9001 certi- fied. ISO 9001 emphasizes quality processes, making the case that a desired result is achieved more efficiently if activities and resources are managed as a process, rather than as isolated events. ISO 9001, like other quality systems, is based upon the hallmarks of quality. Indeed, the hallmarks of quality were originally promulgated by the ISO organization. A firm seeking ISO 9001 certification institutes a quality system under ISO 9001 guidelines and then requests a pre- certification inspection. If they pass, certification is used to demon- strate the quality nature of their products or services. ISO 9001 is used throughout the biotechnology industry but is less commonly applied to biopharmaceutical development. ISO provides many other quality systems, each designated by a unique ISO number and each designed for a specific operation. For example, ISO 17025 certi- fication is for testing and calibration laboratories.
• Quality system regulation (QSR): QSR was developed specifically for the regulation of medical devices but, because QSR is also highly effective at maintaining quality in operations, it has been more broadly adopted by other centers of the FDA as a quality standard for a variety of medicinal products. QSR, defined in 21 CFR 820, sim- ply provides a framework, considerations, and general processes to ensure establishment of the essential elements of a quality system.
• Pharmaceutical quality system (ICH Q10): The International Conference on Harmonization provides the pharmaceutical quality system, a comprehensive system based upon quality concepts. These con- cepts include cGMP, pharmaceutical development, and quality risk management, applicable throughout the product lifecycle (e.g., prod- uct development, technology transfer, commercial manufacture, and product discontinuation). Implementation of this quality system is intended to foster innovation and continuous improvement and strengthen the connection between pharmaceutical development and downstream manufacturing activities.
Each of these distinct quality systems serves a purpose and has a unique scope. It should not be surprising that the institution of any quality sys- tem by a biotechnology firm should be preceded by a conscious decision to institute the system and careful planning to ensure that it is implemented correctly.
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Planning a Quality System
Defining Objectives and Ensuring Management Support
Like any endeavor in the biotechnology industry, quality is planned before a system is instituted. To initiate quality planning there are decisions on the exact nature, scope, and objectives of the quality system to be instituted. Most firms begin with a single quality system and then grow into additional systems as they are required. If product is to be manufactured in-house, the first quality system would be cGMP. Alternatively, if cGMP manufacture and control were delegated to a qualified contractor then the firm’s first qual- ity system might be cGLP or cGCP. A biotechnology company that does not have FDA-regulated products, such as an agricultural biotechnology firm, might wish to become ISO 9001 certified or it could elect to first establish ISO 17025 for laboratory functions. And a firm producing medical devices would likely consider the quality systems regulation (21 CFR 820).
Once a quality system has been chosen, then formal, written documents, the quality manual and the quality plan, are considered as early elements of quality development. To begin, management empowers an individual or group of individuals to prepare a quality manual and begin to write the quality plan. These might be drafted by consultants but more commonly a quality professional is employed by the firm at this juncture. The result is that everyone at the firm understands the purpose and objectives of the proposed quality system and what will be done to incorporate it into daily operations. But, most importantly, everyone is now aware that management currently supports and will continue to support quality initiatives. This ensures that employees, contractors, and consultants of the biotechnology firm buy into the development of a quality system.
Experience suggests that these first steps are the most critical to success, and yet they are often the most difficult to achieve in a biotechnology firm. This is because managers and executives, especially entrepreneurs, are typically unaware of the importance of quality systems as good business practice and as a means to achieve regulatory compliance. Far too often, a firm will hastily develop a quality system without management support and absent either a clear objective or a quality plan. In such cases, there is little planning and the quality system is, more often than not, inappropriate for the firm’s business objectives, hastily constructed and wasteful of resources. Getting started on a quality sys- tem therefore involves significant thought, management support, and planning.
The Quality Manual
Once management has made the decision to establish a quality system at a bio- technology firm, the next step is to create a quality manual. A quality profes- sional is appointed the champion and lead author of this document. The manual
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is a short, usually under 10 pages, document that provides elements, listed in Box 5.2, of the intended quality system. Each section must be tailored to the particular biotechnology operation. Once written, the quality manual is signed by the senior management. It states, for all employees, contractors, and consul- tants of the biotechnology firm, the corporate expectations with regards to the quality system and employee performance under that system. Just as a corpo- rate employee manual spells out human resource policies, the quality manual provides quality policies as guidance to all employees, explaining to each why quality is considered important to them and to their customers.
For example, if the chosen quality system is cGMP, then the manual would discuss the firm’s commitment to manufacture quality product, choose competent subcontractors, perform internal audits, apply specifications, and follow other hallmarks of a cGMP system. The first chapter of a quality manual, the statement by management, is critical and must be signed by the highest levels of management. The second chapter spells out responsibili- ties for everyone in the firm, providing an organization chart and authority and responsibilities for quality assurance and other functional departments (e.g., manufacturing). It clearly defines for everyone the areas involved in the planned quality system. The remainder of the manual spells out quality sys- tem functions in a general sense, leaving details for the quality plan.
The Quality Plan
Having stated objectives and elements of the quality system in the qual- ity manual, the firm now identifies a route to best implement the chosen quality system. An initial step is to review the business plan, the targeted product profile (TPP) and product development plan (PDP) (Chapter 1) and then, under the objectives stated in the quality manual, write a supporting
BOX 5.2 ELEMENTS OF A QUALITY MANUAL TO SUPPORT A QUALITY SYSTEM
Chapter 1: Introduction and statement of management Chapter 2: Responsibilities and organization Chapter 3: Overview of studies performed under the quality system Chapter 4: Type of quality system(s) Chapter 5: Employee management Chapter 6: Safety Chapter 7: Facilities, equipment, and reagents Chapter 8: Special operations Chapter 9: Records and documents Chapter 10: Other elements of the quality system
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quality plan. This means the quality system is appropriate for and supports the intended operations and biotechnology products or services that will be developed by the firm. This may sound easy, but far too often previous and inappropriate experiences become the sole basis for preparing the new qual- ity plan. Each quality plan is unique and must focus on the new biotechnol- ogy product and development objectives. In other words, the quality shoe must fit the business and operational foot.
To begin, the quality planner studies the objectives and processes of all functional and operational areas, current and intended, for that firm. For example, biotechnology firm A may be focused upon genetically engineered plants, agricultural products, and with the intention of doing all product development in-house. Firm B may develop medical biopharmaceutical prod- ucts but, as a virtual company, intend to have most manufacturing, nonclini- cal and clinical efforts completed by contractors. In contrast, firm C may plan to produce recombinant biological molecules for use in research laboratories and in the manufacture of in vitro diagnostic medical devices. Each of these three examples requires a unique quality plan since each firm faces a differ- ent set of regulatory requirements, operational processes, and business objec- tives. Operational research is critical to establishing the correct quality plan.
It is also very helpful to appoint a quality steering committee and to work with project management committees. Members of these committees instruct the quality planner on the technology and resources in their departments and, at the same time, work closely with quality professionals to establish the corporate quality plan. Typically, there is a bit of attitude adjustment required of any product development team embarking on the quality planning process. This is especially true at a firm that has no past experienced working with a quality system. A quality committee with regular meetings ensures that cooperation between the functional area managers and the quality planners begins at the outset and continues through the process of quality planning.
Project committee meetings are another means of disseminating quality planning information and for communicating objectives and quality proce- dures between quality professionals and other functional area supervisors.
Why is there a great need to sell the concept of quality to staff? Managers of other departments may view quality as a threat, since it represents someone from outside their department reviewing critical functions and even suggest- ing operational changes. The journey of establishing a quality system reaches a first milestone the moment managers and supervisors from throughout the firm visibly and fiscally support the draft quality plan. A positive attitude from man- agers goes far because it instills a team spirit and positively influences everyone on the product development team. Using tact and convincing arguments, qual- ity professionals strive to instill this attitude on fellow team members.
Support of peers and managers cannot stop with the quality plan, and qual- ity support must continue beyond establishment of the quality system. This includes continual sharing of information, full integration of the quality staff into product development teams, provision for additional quality resources
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as the scope and complexity of efforts expand, and continual dialogue between quality and functional departments. As will be discussed in later chapters, such processes are not unique to the new biotechnology firm but apply equally to the established firm with a broken or deficient quality system.
Once comments of the team members and upper management have been incorporated, the overall scope and purpose of the quality plan should be finalized. Regulations are perhaps the most important consideration, since a quality system exists, in part, to maintain compliance with the regulations. In Chapter 4, the elements of cGMP, cGLP, and cGCP are outlined and their practical applications are discussed in Chapters 6, 8, and 9, respectively. Next, the quality sections, each focused on a hallmark of quality (Box 5.1) are written. Not each hallmark applies to every product or quality system, so certain hallmarks may be omitted from any quality plan. But omission of a hallmark must be a conscious decision and based upon the fact it is not applicable in a given case. Again consider that each quality system is unique.
Another important element of a quality plan is the quality organization the biotechnology firm wishes to use in daily quality operations. Guidelines for building a quality assurance unit (QAU) (referred to by the U.S. FDA as the quality control unit) or department within a biotechnology firm will be dis- cussed in a later section of this chapter but a few points are relevant to quality planning. The QAU must reflect the size and complexity of the overall quality plan. Minimal considerations are: quality management, documentation, train- ing, and auditing. In a virtual biotechnology firm, these tasks may be completed by a single individual and that person might even be retained as a consultant or contractor, not a full time employee. However, use of consultants is not always possible once a functioning quality system is in place and operational. In an established biotechnology company, a general rule of thumb is the need for one dedicated quality assurance individual for each 25 employees. Of course this ratio, 1:25 may change greatly, for example, if the majority of technical employ- ees are working in development and not basic research. Once again, it is very important to tailor the quality plan and project quality resource needs to busi- ness and development plans. A final note is to consider the quality manual as a living document, one that can be changed if it is not working effectively to support corporate, quality, regulatory, or technical objectives and functions.
Hallmarks of Quality: Fundamental Criteria for Building Effective Quality Systems
The term hallmark is derived from the ancient practice of marking precious metals with a stamp that identified the source and quality of these sought- after materials. Over time, hallmark has come to mean a distinctive feature of an item, especially a feature that makes the item stand out as excellent.
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In the discussion on quality planning and in Box 5.1 are noted features of a quality system that make a quality operation stand out: the hallmarks of a quality system. Apparently, the term quality hallmarks was first introduced to describe features of the quality system ISO 9001 but today hallmarks are characteristic of any quality system. Indeed, a novel quality system is easily built around the most common hallmarks to be described in this section.
Management Responsibility
A quality system cannot exist without the involvement of management, including executives at the highest levels. Quality principles (Figure 5.2), promulgated in ISO 9001, provide eight quality management principles: leadership, involvement of people, process approach, systems approach to management, customer focus, continual improvement, factual approach to decision making, and beneficial supplier relationships. These principles, at the heart of any quality system, are woven throughout an effective biophar- maceutical development program and each will appear again in later discus- sions. It is from these principles that a quality system owes its existence.
Quality management
principles
Leadership
Process approach
Factual approach to
decision making
Mutually beneficial supplier
relationships Continued
improvement
Consumer focus
Involvement of people
System approach to management
FIGURE 5.2 Eight principles of quality management.
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In a practical sense, how then is management involved in quality? Figure 5.3 identifies the primary responsibilities. Leadership is critical. Indeed, management, both executive management and functional depart- ment managers, must support quality efforts of their biotechnology opera- tions. This requirement is so basic to each quality system that regulatory agencies insist management support be clearly identified, and in writing. We noted earlier that management involvement in quality must be specified in the quality plan and other quality documents. The head of the QAU reports to a high-level executive and this reporting structure is reflected in corpo- rate organization charts. Operational documents also reflect that the qual- ity management-to-executive management relationship exists on a daily or certainly a weekly basis and is functional or even dynamic. This organiza- tion of quality resources is driven by regulatory compliance and good quality management practices. Today, inspectors from regulatory agencies such as U.S. FDA typically inspect, for quality compliance, biotechnology firms in a top-down manner, reviewing operational management records in the first hours of an inspection in an effort to learn if management is actually involved in and committed to quality efforts. If this is not the case, then the firm may fail the inspection right from the outset. Indeed, when a firm fails an inspec- tion of a quality system and receives a warning letter, this unwelcome corre- spondence from the FDA is inevitably addressed directly to the president or chief executive officer of that firm since this is the person with direct respon- sibility for all aspects of operations and quality activities (Chapter 4).
Upper management commitment
Involvement and leadership
Customer
Management reviews
Quality policy
Quality plan
Delegate responsibility and authority
Effective quality
assurance unit
FIGURE 5.3 Upper management responsibilities in a quality system.
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Management responsibility continues largely through management reviews, a process in which designated upper management considers all aspects of their quality system to ensure it meets stated objectives and remains suitable and adequate to meet current operations and future plans. Management review is especially important for an organization that is evolving, changing, or growing and whenever new regulatory guidelines, new products or services, or new markets arise. Thus, management review identifies opportunities for changes to the quality system.
Guidelines for management involvement are established in writing and the results are reflected in corporate documents. Executive management under- stands the biotechnology, regulatory and quality processes to the extent that they can make intelligent, high-level decisions concerning the overall qual- ity, safety, and efficacy of their products. Management has set appropriate goals and objectives, has established standards, and is clearly receiving feed- backs from their functional area managers. And they are aware of signifi- cant changes in all areas of operations and how these changes are executed. Upper management not only empowers quality leaders but also maintains the authority to make changes when quality issues are brought to their atten- tion and the record should show this to be the case at their firm. An absentee or otherwise disengaged executive is not appreciated in the biotechnology industry, especially by regulatory agencies, since executives often do not have his or her finger on the pulse of their firm.
Defined Quality System
This hallmark simply means that one or more quality systems have been established at the firm; this was mentioned earlier in this chapter as an early step in quality planning. Further, each hallmark identified as part of that chosen system must be important to operations and to the completion of the product development life cycle at that firm and for that product. From a standpoint of compliance, regulators expect each quality system, such as cGMP, cGLP, cGCP, or ISO 9001, to be identified in the quality plan, any quality manual and in other quality documents. Also, the chosen quality system must fit the product accordingly. For example, cGLP regulations are, by the U.S. FDA definition, a quality system used for nonclinical safety studies. Yet, cGLP is sometimes applied to systems far removed from this scope. This leads to organizations touting their compliance with cGLP when, in fact, they do not even meet the U.S. FDA’s intended scope of the regulation. This inevitably leads to inappropriate use of the system and con- flicts between the firm and regulatory agencies or customers. Actual quality practices must meet procedural definitions as well. Simply stated, if you tell customers or regulatory authorities your operation is compliant with a par- ticular quality system then it must indeed meet both the scope and practices for this system.
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QbD and Design Control
QbD has, in just a few years, become a key component of biotechnology operations and most quality systems. Regulatory agencies now recommend QbD for most medical products just as they demand management involve- ment in the quality system. It is no coincidence that design control has come to represent the heart of QbD, an important aspect of development for any biotechnology product and a requirement for biopharmaceuticals.
Quality by Design
QbD applies quality concepts to the design and development of biopharma- ceutical products of all types and for every indication. It is based upon cus- tomer needs for quality products, excellent science, design control, and risk analysis and management. In the first step and as described in Chapter 1, QbD identifies product requirements to meet the needs of the user, a patient, and the health care provider. The next step, designing a product such as a molecule or a cell, seems counterintuitive. Isn’t biotechnology itself based upon discov- ery, the concept of developing a product from a new finding? In the real world, a scientist makes a discovery in the laboratory and then finds investors or another sponsor to take this new molecule or cell forward to the marketplace.
But QbD does not fly in the face of reality. Instead, it suggests taking this discovery, new molecule or cell, whatever, and matching it with a require- ment or a need and a patient population. Sometimes, however, the match is not good and the molecule must be rediscovered or at least modified to meet the exact need. For example, a new monoclonal antibody may be dis- covered using an animal model system such as a mouse. The intended use is to develop this monoclonal antibody as a therapeutic for human malaria in man. In mice, the antibody works as a therapeutic, clearing an infection with mouse malaria parasites. The discoverer wishes to take this finding forward but, in planning the project, discovers that the mouse monoclonal will not work against human malaria parasites and that the mouse antibody might elicit undesirable reactions if it were given to humans. There is no market for a therapeutic for mouse malaria. This is where QbD might come into play. In this example, the scientific concept is solid but the initial discovery is not useful to the intended user. By redesigning the molecule, and this can actu- ally be done both by generating new monoclonal antibodies or by genetically engineering the mouse antibody to be a humanized molecule, it is possible to rediscover the concept and to design a new molecule that will meet require- ments of the human user.
Indeed, QbD is not the traditional or twentieth century concept of drug dis- covery, in which thousands of molecules were first generated by an organic chemist, then screened for any attribute in laboratory and animal studies. This older concept of pharmaceutical discovery has been compared to a fun- nel with a fine screen and very narrow opening that often led to nowhere for
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most molecules. Very few molecules made it into the development pipeline, yet many of these were a bad fit for their intended purpose and were consid- ered great molecules, but needing a useful and marketable indication.
Therefore, QbD was introduced and since 2004 the U.S. FDA has strongly recommended it as an operational method for modernizing both drug dis- covery and drug development. It has quickly spread to the biotechnology industry in part because it works quite well and also because the twentieth century drug development process, the funnel concept of screening many compounds to find just one candidate that suited an indication, simply does not suit discovery and development of large molecule and cellular products. The funnel concept just did not work for biopharmaceuticals and they often had to be redesigned to suit a specific need. Another element of QbD is a need to consider the TPP, discussed in Chapter 1. QbD goes even further than early stages of discovery and development and continues throughout the life cycle of the product to ensure continuous improvement and innovation as critical quality attributes are applied to product development. And QbD is used with functional areas of development, notably manufacture, qual- ity control, nonclinical studies, and clinical trials. An example is given in Chapter 6, where QbD is applied to the design of a manufacturing system for a biopharmaceutical product.
The concept of QbD was really an idea whose time had come in the early twenty-first century and both industry and the International Conference on Harmonization (ICH) deserve credit for encouraging its use. ICH describes, through their quality or Q series of documents (e.g., Q8, Q9, Q10, and Q12), QbD concepts and practices for biopharmaceutical products. These arguably are the leading reference documents on QbD today and should be consid- ered by any biopharmaceutical firm entering development and establishing a quality system.
Design Control
QbD applies a formal means and format of designing a biopharmaceutical product. Design control, at the heart of design, is best designated as the gen- eral arrangement or layout of a product. It demands both an active process and results from that process, notably written design documents. Historically, the process of design control evolved from engineering projects, notably in the medical device industry, where physical design of a product was seen as good business practice and, more recently, as a regulatory requirement. For example, a new heart pacemaker would only function correctly if it was designed to meet certain specifications. To be effective and safe for the user, and thus marketable, an implantable pacemaker is a maximum size, very reli- able, for example, in the accuracy of beats-per-minute, and useable, for exam- ple, have a battery life that supports many years of use by a patient. If the management asked for a new pacemaker and did not insist on a user-friendly certain design, they could find themselves with an implantable device that
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was 10 cm in diameter, produced 30 ± 20 beats/min, and had a battery life of 30 min, hardly desirable to the customer or a regulatory agency and quite difficult to market. Design control takes user needs into consideration before the product is produced. It is a good business practice.
Today, design control is applied to biopharmaceutical development with the designer following certain steps, these having been adopted by indus- try and regulatory authorities. Elements of design control are shown in Figure 5.4 and further defined in Box 5.3.
Target product profile (TPP)
Design input
A plan of product design
Review
Output: A product design
Review
Product output and verification
Review
Design validation
Final development
and production
Changes to design and specifications
(Figure 5.5)
Management guidance
Design specifications
Engineers Scientists
experimentation
Design team
Development team
FIGURE 5.4 Elements of design control.
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• Design control is product specific. The product and indication for the biopharmaceutical are specified and each design control process is for one product with a unique indication and no other. Further, design is repeated each time a change is proposed to that prod- uct or its indication. As an example, consider again the monoclo- nal antibody against a malaria parasite. Having a firm grasp that the concept is feasible from studies in mice, it is now designed as a humanized monoclonal antibody to bind and kill the human malaria parasite. The concept of QbD and specifically the process of design control now impacts future development of this monoclonal antibody. Further, quality criteria now take on much greater quality implications, for planning is the first step of development.
• A product design plan is drafted. Design planning is a process much like product development planning but is much more specific in scope, focusing on how the design process itself will take place. The design plan identifies the product and indication and then it outlines vari- ous elements of design, notably input, output, review, and decision, serving as an agenda for product design. A product design plan might have an outline as shown in Box 5.3. But, in addition, it would describe who is responsible for each step of design, when it would occur, how it would be completed, and what the expectations might be.
• Design documents and records reflect each step of the design control process. Requirements for written records are established. These
BOX 5.3 ELEMENTS OR STEPS IN DESIGN CONTROL
• Product is specified: The product and indication are clearly described (e.g., using TPP).
• Product design plan is drafted: This plan then drives the process. • Design process is fully documented: Detailed records are kept
throughout the process. • The process involves the full product development team. • The design control process begins and these steps may be repeated
until the team is satisfied with the final design.
Input Review Output Verification Validation Change
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include process documents such as agenda and minutes to meetings and product documents, for example, the draft meeting minutes and the product’s written draft labeling, user’s instructions, techni- cal descriptions, specifications, and drawings that have been gen- erated during the process. Plans are made to keep product- specific historical files and this is accomplished by the following quality procedures.
• Design control involves professionals, technical and management, serving on a design team with responsibility for the product’s development. Research, manufacturing, sales, marketing, qual- ity assurance, quality control, senior executives, regulatory, project management, finance, and personnel must somehow be involved in the processes referred to as design input, output, review, and change. If this design review group seems similar to a project team described in Chapter 2, then you are correct. Very often the members of a proj- ect design team for a biotechnology company will be assigned to the development team as well.
• Design input is the next step. The purpose is to enlist opinions of various individuals regarding how the product should be designed and this, of course, takes into consideration the operational plans we have discussed throughout this book. For QbD to be effective, the quality attributes of the product must be a prime consideration in design input. Quality professionals should be actively involved early in the process. Leadership is key to success of design input and here an effective project manager may lead the team effort and ensure constructive communication. In a practical sense, design input involves sitting around a table as a team, speaking, listening, and learning from each team member. Everyone will, at first, be sur- prised at differences in individual perceptions for a single product and indication. Returning to the example of the humanized mono- clonal antibody to treat malaria, marketing may suggest that it be manufactured in final form for $50 per dose and be used to treat three species of human malaria parasites. Manufacturing personnel may disagree, suggesting that it could only be produced and for- mulated for over $200 per dose. And clinical affairs might suggest that initially it can be marketed to treat only one species of malaria parasite, as initial studies to treat three parasite species would be prohibitively expensive and time consuming. Consensus must be achieved in these meetings, otherwise everyone is advocating a unique design and efforts will be disjointed, at best. In effect, design input involves soliciting everyone’s opinion and clearly describing a single route forward, that is, reaching a firm decision.
• This all leads to design output, which, as the name suggests, is nothing more than producing, in a written report, consensus of
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opinions, a feasible product development objective, a consensus on the design, and an idea of how all these efforts might be applied to achieve a common objective. Design output provides important key product specifications, agreed to by the team members and by upper management. For example, the team might agree the monoclonal antibody against human malaria infection must target one malaria parasite species, have a therapeutic effect in 80% of patients, give an excellent safety profile in infants and children, and be manufactured for $100 per dose. Of course, the actual output document is much more detailed but nonetheless the most important attribute is a clear design for the intended product.
• Design reviews are performed throughout the design and opera- tional phases of biopharmaceutical development. Once an output document is established, it should be critically reviewed by a much larger audience. Using partners, contractors, and consultants is recommended as is review by the upper management and techni- cal staff. Reviews often lead to new ideas and improvements and these then become additional input, thus starting the design pro- cess over again or leading to changes in design. This might precipi- tate another round of design review and revised outputs. Hence, QbD never involves a single meeting, in which everyone agrees and from which design output is completed days later. Instead, it requires many meetings, interspersed by additional laboratory and marketing research, and further input from management. During this period, the evolving design is reviewed by the team. Once a design has been agreed, design change is inevitable. Problems are encountered in development and they must be reviewed at frequent intervals by the team. Quality issues continually arise and must be addressed in a revised design.
• Design verification follows. Economics suggests that sooner or later the design cycle must end and the product must be produced, at least on a limited scale, and then tested. In biopharmaceutical develop- ment, this involves manufacture, quality control and studies, both nonclinical and clinical. The testing aspects are considered design verification. The product, and hence the design, is tested in labo- ratory animals and man. It determines where and how the design, which is really a model on paper, will be developed and produced. For the monoclonal antibody against human malaria, this might mean that extensive laboratory testing reveals, for example, the mol- ecule is stable at a particular temperature, that it can be produced in a 10 L bioreactor and is not toxic to a small animal. Design verifica- tion, analogous to Phase I and early Phase II development, involves laboratory and early clinical experimentation and testing. In the end, design verification leads to confirmation or rejection of individual
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components in the design and, if things are not perfect, it can trig- ger another round of design input, output, review, and verification. It is at this stage in development that the design team begins to also function as a product development team.
• Once a product has withstood initial testing and is verified, design validation may be used to further substantiate the adequacy of the product’s design. Validation typically has a more stringent defini- tion than verification. It requires additional testing and might even involve changes to variables that had not been previously tested. In biopharmaceutical development we often consider design validation to be middle or late stage (Phase II or Phase III) development encom- passing advanced nonclinical and clinical trials.
Design Change
Change is expected during a product’s life cycle. Biotechnology products do not speed through the full course of development activities without several changes in TPP, PDP, or product design. While change is expected and even good in many cases, change must not be a random or uncontrolled event. Changes in product development must be controlled throughout the design control process and the development life cycle.
In summary, a hallmark of quality is the concept of QbD incorporating the process of design control. We think of this in much the same way as plan- ning development but it has much broader and deeper implications, taking into account the ideas of design and great technical depth and detail.
Contractor, Vendor, and Consultant Control
Every biotechnology development program depends upon acquisition of goods and services. A quality system takes great care to source only the best raw materials, advice, and contract support. It does so by incorporating corporate policies and procedures for obtaining, by purchase, collaboration or contract, materials or services. This, along with good business practice and common sense prevent the purchase of materials or services that might foul part of the product or development scheme. Unlike large pharmaceuti- cal firms with in-house resources and procedures available to closely man- age and control vendors, small- to medium-sized biotechnology firms often do not have this expertise and capacity. In fact, a virtual biopharmaceutical firm may rely upon very few employees to manage a significant amount of the operational effort that is actually performed or supplied by contractors and vendors. Also, consultants are often retained by biotechnology firms to provide critical advice or prepare important documents, such as regulatory submissions or quality plans. The quality of these materials and services is important to the success of any product development effort but the buyers
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must themselves ensure compliance with established quality criteria. How can this be accomplished in a fast paced biotechnology environment? First, a buyer establishes specifications for every material or service considered for purchase or hire. Second, they consider more than one offeror, when- ever possible, and carefully evaluate each one by reviewing the corporate history, experience, and references. Vendors or providers passing these ini- tial screens might then be evaluated in greater depth through audit of their facilities.
Further to the example of a monoclonal antibody to treat malaria infec- tions, the manufacturing and quality group considers the need to purchase saline for formulation of the product. Specifications, such as USP grade nor- mal saline for injection in 1-L sealed glass bottles, are established. A request for proposal is sought from three or more contractors. Once the vendor’s pro- posals have been reviewed, further information, such as copies of Certificates of Analysis, for the last three lots of saline, might be requested. Additionally, staff might determine if a supplier complies with a quality system such as cGMP or ISO or they may request inspectional results from previous ISO 9001 or FDA inspections. Given that the quality of this saline is important to the overall quality of the monoclonal antibody product, it would be pru- dent to schedule an audit of the saline manufacturing facility once a likely supplier has been identified from the list of three candidates. Audit proce- dures are explained later in this chapter. Once the saline arrives, samples from each lot might be retested for critical parameters (e.g., sterility, pH, or concentration of sodium chloride) by the firm’s quality control laboratory to determine if it does indeed meet the specifications cited in the vendor’s certificate of analysis. Quality control testing of raw materials is described in Chapter 7. Upon receipt, saline is kept at the recommended conditions and it may be tested for stability prior to reaching the expiration date.
Ensuring the quality of consultants or advisors and service providers is also important to a biotechnology firm. Technical requirements and the intended scope of work are clearly stated, to include the amount of control, review and supervision that will be provided by the firm to the consultant. Resumes of candidates are reviewed and references are checked to ensure that each consultant applicant is qualified by experience and training.
Service providers, such as contract research organizations (CROs) per- forming manufacturing, quality control, clinical and nonclinical studies, are thoroughly examined and references reviewed before contracts and agree- ments are signed. These are high-cost and high-profile contracts involving months and years of effort. Potential failure or delay by a CRO has great negative impact upon a biotechnology development program. The steps involved in selecting a CRO are no different from those used to select a ven- dor for raw materials. The purchaser establishes exact specifications and fully describes the intended scope of work. At small biotechnology firms, it may be necessary to hire an experienced consultant to write this critical document and then participate in the selection process. The scope includes a significant
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amount of detail on technical and quality aspects and a request for proposals is advertised. Proposals include elements of technical performance, cost, and quality. Once proposals are received, the candidate list is narrowed to only qualified proposals. Again, it may be necessary for the small biotechnology firm to retain consultants to assist in review and to use the expertise of the project team, to include those with finance and contract responsibilities, dur- ing the review and selection process. Audits and a check of references is an absolute must when considering CROs for a major contract. Once a vendor is selected, they sign both technical and quality agreements. The firm may assign one or more employees on staff to oversee and manage a major devel- opment contract.
Quality agreements are based upon the quality expectations and specifi- cations for the product or service. They define the quality system and pro- cesses to be used by the CRO or vendor to manage quality aspects of the material or service contract. The scope of the quality agreement covers the full scope of quality efforts. The nature of each quality system that applies to the service or material is clearly stated. The vendor-client relationship, with responsibilities, procedures for changes to the deliverables and procedures to resolve disputes, is described in some detail. A responsibility matrix may be quite helpful. Since ongoing monitoring and auditing are likely to be part of the contract, the exact nature of these activities is stated in this agreement. And terms are clearly defined in an effort to prevent misunderstandings. For example, quality terms that may be confused are substantial deviation or minor error and these are either not used or they are clearly defined under the scope of the quality agreement.
Biotechnology firms rely heavily on outsourcing and success or failure of a service or material provided by a vendor can have a huge impact on an oper- ation. It is critical to ensure that quality criteria and functions are considered in all contract, consultant, and vendor agreements and activities.
Product Identification and Traceability
The sponsor must have in place a system to identify all materials and prod- uct as it moves through the development life cycle. This control applies in-house and to deliverables from services, such as quality control test- ing and clinical or nonclinical studies. Product identification and trace- ability are considered quality responsibilities at a biotechnology firm. For manufacture of a product, this may be the application of a unique num- bering system to identify lots of manufactured product and a method to trace numbered lots through the distribution system. With such systems, a manufacturer may trace forward to learn where and how the product was manufactured or to trace backward to identify the origin and handling or possession of each raw material, facility, and piece of equipment used in its manufacture. Clearly, product identification and traceability necessitate a mature and infallible documentation system, a quality function that will
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be discussed later. For a service provider, the identification procedures may be more involved, requiring multiple levels of identification. For example, a contractor’s quality control testing laboratory has numbering or labeling systems to track each sample and results as they pass through the test- ing and reporting process. A clinical or nonclinical study is based upon a specified and numbered protocol and each segment of that study—animal or human subject, test product, treatment or procedure—is uniquely iden- tified to ensure the protocol was executed flawlessly and that each compo- nent was performed completely and correctly.
Today, the quality function of ensuring identification and traceability increasingly relies upon barcoding and microprocessor-based systems. Quality electronic records and signatures are critical to performance require- ments. Hence, the software and microprocessors themselves must be of the highest quality and suited to the tasks of identification and traceability. There are regulatory requirements for microprocessor-based systems (Chapter 3).
Process Control
A phrase common to the pharmaceutical industry is you cannot test quality into a product. This means quality must be built into the product at every phase of development and throughout the process, that is at each step in the production process. Biotechnology products are no exception to this rule and therefore process controls are used to ensure quality throughout the pro- cess. Process control relies on clear and concise written documents to guide operators at each step of biopharmaceutical production. These are standard operating procedures (SOPs) and batch production records (BPRs) or work instructions. For nonclinical studies or clinical trials, protocols and SOPs are used to guide processes.
Protocols, defined by the Oxford English Dictionary as “official formality and etiquette” (Oxford English Dictionary, 1997), are formal written and approved instructive documents that describe, step-by-step, how studies and trials are performed. The BPR, a prospectively generated, formal written and approved document, guides manufacturing processes. BPRs are also used to collect information or data as it is generated. Typically, a protocol or BPR gives the reader broad step-wise guidance and references to SOPs, documents provid- ing more exact technical instructions and describing exactly how the process is performed. In a quality environment, such as those mandated by cGMP, cGLP or cGCP, SOPs are also used to provide a controlled work environment, thus ensuring products are made in the appropriate atmosphere, to instruct the use of equipment and utilities, and to mandate exactly how other activi- ties are undertaken in the manufacturing environment.
The technical staff of a biotechnology firm prepares BPRs, SOPs, Protocols, and related documents but each one must be reviewed and approved by both supervisors and quality assurance staff. As approved documents, BPRs, SOPs, and protocols, are highly controlled and may be changed only using formal
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processes and with approval of all responsible individuals. Documentation control and change control responsibilities, a quality assurance function, are described later in this chapter.
Environmental Controls
A product is only as good as the environment in which it is produced and biotechnology products are manufactured or tested in a wide range of environments. Genetically, engineered plants are grown in green houses or open fields under controlled conditions. Sterile, recombinant proteins are manufactured and then formulated in highly controlled, indoor, envi- ronments (Chapter 6). Clearly there is a significant difference between the controlled environment in a corn or tobacco field and the environment within a biopharmaceutical manufacturing facility; yet each environment meets specifications suited for their product’s specifications and intended use. We think of biomanufacturing endeavors as happening in clean rooms and in fact this is by far the most common practice. Environmental controls bring into play the issue of the quality of the facility, the air, the water, and the personnel. For genetically engineered plants grown outdoors, the quality of the soil may also be considered. What goes on in the facility is also important. The flow of raw materials, product and waste, and person- nel in a biopharmaceutical manufacturing plant can have a major impact on the quality of the product. The quality of the people is no exception since their performance is part of the overall environment in a biotechnol- ogy operation. We discuss in Chapter 7 the laboratory testing efforts that go into ensuring the quality of all aspects of biotechnology operational environments. Quality is intricately involved in these efforts and ensures, through inspection, audit, validation, review, approval, and documenta- tion, that the environment meets preset specifications and is suitable for the intended operation.
Inspection or Testing (Quality Control)
Inspection and testing are integral to manufacture of biopharmaceutical products. As described in Chapter 7, quality control efforts are technical, performed primarily in laboratories. Today, quality control testing is often administratively separate from the QAU. Nonetheless, quality assurance has significant responsibilities for ensuring that testing is fully qualified or validated for the intended purpose, that testing was performed accord- ing to procedures and that test results conform to specifications. Unlike discovery research, quality control testing is a formal process, performed under SOPs and using highly developed qualified or validated proce- dures. A distinguishing feature of quality control is the use of specifica- tions, normally a range of acceptable test values, against which sample test results are compared (Chapter 7). Quality control testing is performed
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on raw materials upon arrival, to ensure that they are, in fact, identical to claims made on the label or certificate of analysis. In-process testing is performed on samples taken during the manufacture of biotechnology products. While quality cannot be tested into a product, in-process testing in part ensures the quality of products along the production process. Drug substance and drug product, defined further in Chapter 6, are tested in the quality control laboratory. Then, once finished and filled into a con- tainer, final biopharmaceutical product is further inspected and tested. Stability testing demonstrates that a product remains pure and potent once it has been stored for a designated time, hence the need to pro- vide a stated shelf life for each biopharmaceutical. Quality control also involves the calibration or certification of test and measuring equipment to ensure that it meets standards or specifications. Just as manufacturing processes must be validated, so too must analytical tests and measuring devices (Chapter 7). Quality assurance professionals work closely with quality control to ensure that adequate test methods are established, that test results are compared to specifications, and that all testing is fully and precisely documented.
Release of Material, Service, or Product
Before it is released for use, a manufactured product must conform to all specifications. This means that raw materials, processes, environments, identification labels, and results of inspections and testing must meet prede- termined specifications. Variance from any one specification can, in theory, lead to the rejection of that product so that it cannot be released for public use. The jargon used in the biotechnology industry to describe an accept- able product is conform or pass while an unacceptable product, one that does not meet specifications, is nonconform or fail. Even though it is the other operational groups— manufacturing and quality control—that produce and test product, it is the quality assurance function that reviews the data and decides whether a product conforms or does not conform to specifications, that is, passes or fails. Product that does not conform is usually placed into quarantine while the data is reviewed and the situation investigated. But if failure is the product’s eventual fate, the product is destroyed or completely reprocessed if appropriate. Quality assurance has the task of reviewing all records and, ultimately, deciding whether or not a service or product is released to the market.
Release applies to services as well as product. For example, written reports of clinical and nonclinical studies are released to the client only after they are reviewed, approved, and released by quality assurance. Not only is the report reviewed but also the performance and compliance of the complete study, from protocol through data collection, are closely scrutinized to ensure integrity and correct translation of results to the report.
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Change Control and Corrective or Preventive Actions
As one might expect, not everything goes perfectly or as planned in a biopharmaceutical operation. Despite the best intentions or planning, some- times stuff just happens. If a lot or batch of product or a report does not pass scrutiny by quality professionals, it is unacceptable for release. Unacceptable products or reports are considered nonconforming since they do not meet specifications or were not made according to written procedures; it does not pass. And quality audits may uncover hidden defects in a process, raw mate- rial or test program. Nonconforming product may also be identified by cus- tomers who complain about a product even after it passes and reaches the market. In cases of nonconformance, corrective and preventive actions must be taken immediately. These decisions, and the investigations that typically precede them, are the purview of quality assurance.
A quality system includes procedures to collect and, when appropriate, review nonconformance issues as they arise. Quality assurance is tasked with ensuring that timely collection of information, including customer com- plaints, identifies problems with a product, study, or other service. Quality professionals then make management aware of the issues and ensure that appropriate corrective action is taken to resolve the problem and prevent it from recurring. Corrective plans, instructions for investigating failures or deviations and initiating corrective action, are written and executed for this purpose. Investigations are prioritized based upon the risk a problem poses to the user. A serious problem with a biopharmaceutical, one that might put patient’s lives or health at risk, is addressed immediately and decisively, per- haps with a product recall and halt to production. Perceived or unproven problems are investigated over time as quality assurance staff follow trends and discuss the situation with professionals such as functional area manag- ers and consultants or vendors. Corrective actions follow identification of the root cause and lead to application of controls and additional monitoring to confirm effective resolution. Quality professionals pass information on to senior management since, as noted in Chapter 4, executives are ultimately held responsible for resolving issues. Another result of an investigation and identification of root cause of a problem is preventive action, a process also guided by quality assurance. Preventive actions address the root cause of a problem and are intended to prevent the problem from reoccurring.
Change is normal, often good, for all aspects of a biotechnology operation; earlier in this chapter, change was discussed in relation to design control. Corrective and preventive actions may lead to change in a manufacturing process or clinical or nonclinical study. As manufacturing progresses through the development life cycle, changes are made to improve product yield and quality. Clinical or nonclinical protocols require change, mid-study, to correct omissions or errors that threaten the integrity of the study itself.
Change, a carefully managed process, is referred to as change control and should always be under the control of employees and change must
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be reviewed and approved by quality assurance. A scheme for controlled change is shown in Figure 5.5. For biopharmaceutical development activi- ties, the controlled change process is mandated by regulatory agencies. Change is typically planned and executed by a functional area manager (e.g., manufacturing or clinical) but the QAU may recommend change and in any case QAU must approve plans to change and provide oversight and approvals. Hence, it is always a team effort. Change in a biotechnology operation, no matter how seemingly insignificant, requires forethought, extensive discussion, focused decision, and follow through in action and documentation.
Packaging and Labeling
Each biotechnology product has a container and, adhered to the outside, a label that exactly identifies its contents. Containers must be appropriate to hold a given product and to maintain its identity, purity, and potency. A product label contains very important product information, such as lot number, exact name, strength or dose, formulation, expiration date, and warnings or critical instructions to users. Labeling, as described in Chapter 6, provides additional information on a product as printed matter that is inserted into the packaging (hence the term package insert). If any of this information is incorrect, then the product itself is compromised, misbranded in parlance of the biopharmaceutical industry, because it is not correctly packaged or labeled. Production and use of packaging and labeling are highly controlled processes and their quality must be perfect to prevent omission or error, otherwise the product is considered mis- branded. Therefore, quality criteria for these processes are every bit as stringent as they are for making the product itself. Whenever a package or label is generated, quality assurance approves the printed material before it is used and they also ensure that the packaging and labeling processes are performed correctly.
Preservation, Storage, and Handling
A biotechnology product may be perfectly manufactured and labeled but if it is improperly preserved, stored, or delivered, then it is of no use. In fact, improper storage renders a product adulterated. Therefore, a quality system ensures that product is properly handled postmanufacture. For example, if a protein solution is, by specification, to be kept frozen but inadvertently warms to room temperature on the loading dock, then it is no longer a qual- ity product. Biopharmaceutical manufacturers make every effort to ensure adequate controls are instituted and followed so that only pure and potent product reaches the customer. Quality procedures and records are used for all aspects of transportation, storage, and delivery of a biotechnology product.
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Servicing
This aspect of a quality system is seen largely in the medical device indus- try, where it is essential to provide service, calibration, and technical support to customers. However, in other areas of biotechnology, such as production and sales of research reagents, manufacturing equipment or analytical instru- ments, servicing is also important. While the QAU does not itself provide ser- vicing to a customer, it ensures that servicing programs are instituted and they monitor the quality of those efforts.
Customer Concerns and Adverse Event Reports
Every biotechnology firm with an investigational or marketed product collects and reviews comments from customers and, in the case of medi- cal products, collects safety data. Trend analysis is often used to prioritize complaints and identify problem areas. Management and quality assurance review complaints and establish and maintain programs to address cus- tomer concerns.
Document Control
Also referred to as record control, documentation is a major element of any quality system. Keeping quality records is a major endeavor in a biotechnol- ogy firm and even a small operation generates thousands of critical docu- ments each month. These records, identified by document type in Figure 5.6, are used to support regulatory applications, to provide data used in busi- ness development, to verify compliance with regulations, to demonstrate the application of appropriate quality systems, to record technical proficiency, to document and track changes, trends and issues. Records, written, printed, or electronic, are often legal documents, meaning they can be requested by a court of law. Each record is reviewed for accuracy and completeness and, in most cases, signed and dated. Then it is archived, where it is available on short notice. Document management, as performed by the QAU, is discussed later in this chapter.
Training
Employees must obtain the appropriate training before they begin work and that training is kept current during the course of their employment at a bio- technology firm. Training is directly relevant to an employee’s duties and it is performed to assigned procedures. For example, it is acceptable to employ an individual in a biopharmaceutical manufacturing operation if they have training and experience in a given skill area but it is not acceptable to employ a laboratory technician in manufacturing if they do not have train- ing and experience in that skill area or are not properly supervised during
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a training period. A quality system ensures that employees are fully trained and also have on file current job descriptions and documentation of training and appropriate education before they are qualified to perform a particular job function.
Auditing
The audit function is key to maintaining a quality system in biotechnology. The International Standards Organization defines a quality audit as, “A systematic and independent examination to determine whether quality activities and related results comply with planned arrangements and whether these arrange- ments are implemented effectively and are suitable to achieve objectives” (International Standardization Organization, 1994). There are several impor- tant phrases included in this definition. First, auditing is systematic because it is a carefully planned and executed activity. Second, the audit is independent of the entity being audited to avoid any potential for conflict of interest. Third, audits focus upon quality aspects and not on highly technical activities. In other words, an auditor examines whether or not the appropriate quality system is
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Procedures (SOPs)
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Release, approval, change, and validation documents
Quality record files and archives
FIGURE 5.6 Pyramid of quality documents.
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in place to support a particular technology and functional area and has limited regard for scientific or technological details. Compliance, really the demonstra- tion thereof (Chapter 4), is of utmost importance in a quality audit. This may seem counterintuitive for a high-technology industry, but scientists are respon- sible for technical issues while quality professionals work with technical staff to ensure the quality aspects of that same operation. Fourth, an audit compares what was actually performed against planned arrangements. It determines if per- formance matches instructions. Finally, an audit examines whether or not these planned arrangements were really appropriate to achieve the stated objective.
For internal audits, a biotechnology firm’s quality assurance auditors inspect the records of a functional department, such as manufacturing, within that firm. External audits are performed on contractors, vendors, or collaborators, operations external to the firm performing the audit. It would be impossible to audit every function, internal or external, of a biotech- nology firm or that firm’s contractors and vendors. Restraints of time and resources allow only for the more critical functions of vendors to be audited and therefore priorities and audit plans are established. Also, there are many ways to perform an audit; one method does not fit each entity or situation. Quality assurance procedures for conducting audits are explained later in this chapter.
The Quality Assurance Unit
Under the hallmarks of quality section, aforementioned, we discussed the attributes of a quality system. In most cases, these attributes are managed, at least to some degree, by a group of professionals within a biotechnology firm referred to as quality assurance or the QAU or more informally, QA. U.S. FDA regulations (21 CFR) refer to this entity as the quality control unit but within the industry quality assurance is the term most widely used. The QAU serves several important roles at the biotechnology firm. First, it maintains a compli- ance posture to help the firm meet regulations. This means that QAU works closely with functional area supervisors and coordinates frequently with regulatory affairs staff. Second, QAU serves the users of product or clients of services by ensuring they receive products or services of the very highest quality. Third, the QAU is the gatekeeper of the quality plan, quality manual and the various hallmarks of quality for the firm.
By now it has become clear that five aspects of quality operations— management, documents, training, auditing and change control, and inves- tigations—occupy much of each quality professional’s time and effort. These are shown in Figure 5.7. They are also quite important to the success of a biotechnology firm, especially for those involved in biopharmaceutical development. These five aspects also garner quite a lot of attention during
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inspections performed during due diligence or by regulatory agencies. This section provides more detail on those five important functions, expanding on descriptions provided in the hallmarks of quality section and describing how each is handled by a QAU.
Manage the Quality Assurance Function
A QAU is designated under the quality policy and the unit’s responsibilities and authority are described in the quality plan. A primary function of the QAU is management of the quality system or systems instituted by the bio- technology firm under the policy and plan. The QAU is managed by a trained and experienced quality professional. Indeed, and as noted earlier, this indi- vidual at a small biotechnology firm may write the Policy and Plan and initi- ate the QAU. In addition to possessing quality assurance skills, the head of the QAU understands the technology being developed by the firm and techni- cal aspects of operational areas falling under the proscribed quality systems. They also have knowledge of regulatory guidelines, especially as they apply to the quality systems. In some firms, the quality manger will be responsible for implementing one or more quality systems. To work effectively with the product development team and interact with upper management, the head of the QAU possesses leadership and negotiating skills, as well.
As with other operational units in the biotechnology firm, the QAU is managed in all respects. Preparing budgets, managing personnel, commu- nicating, serving on teams, and establishing priorities are but a few of the routine management tasks. Two quality requirements, communication and coordination, stand out and, if they are performed effectively, distinguish the excellent QAU from the mediocre department. These requirements are
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FIGURE 5.7 Responsibilities of the quality assurance unit.
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important because quality functions, and therefore quality professionals, are highly integrated into the daily operations of a biotechnology firm. Quality professionals provide information, notably advice, to many individuals and they coordinate their activities with many others at the biotechnology firm. Functional area managers depend upon communication and coordination with quality assurance professionals if they are to achieve their objectives. This means that quality assurance leaders must themselves understand com- plex technological and regulatory systems and then integrate their quality processes in a timely and effective manner to meet the objectives of the team. This makes quality assurance management a unique endeavor and provides many internal and external challenges to biotechnology development teams.
Control Documents and Manage the Documentation System
Earlier in this chapter, we briefly mentioned documentation as a hallmark of quality and stated that everything, no matter how seemingly insignificant, that happens in a biopharmaceutical operation is recorded. Tiers of docu- ment types were identified in Figure 5.6. The QAU manages and controls each of these entries as an official record. In this section, we summarize doc- uments and review quality assurance responsibilities and procedures used to ensure an effective and compliant documentation system.
Documentation, a process, provides a means to generate, review, use, and store documents. There is a saying in the pharmaceutical industry, if it was not written down, then it was not done. Every aspect of biotechnology opera- tions, including plans and processes generate piles of records, a variety of documents. While written records are the norm in many firms, the trend is to move to electronic records and signatures. Most small biotechnology operations begin by using paper document systems but, with growth of a firm, electronic systems are adapted and they do provide certain advantages. Yes, transitioning from paper to electronic records does not in itself simplify the documentation challenges. Capture, review, audit, and storage of docu- ments must also be done exactly and these tasks fall to the QAU. Regulatory agencies expect nothing less. Further challenging documentation efforts for growing biotechnology firms is operational growth beyond the capacity of the existing documentation system. Finally, senior management never sees all these documents and therefore seldom appreciates their volume or the complexity of properly reviewing, approving then organizing and maintain- ing all these files.
Who then is responsible for writing, reviewing, and approving each of these documents? Plans and strategies are written by high-level manage- ment with the assistance of technical and administrative staff. Protocols are prepared by investigators or study directors, individuals responsible for designing and completing a nonclinical or clinical investigation or valida- tion. Procedures, manufacturing records, and work instructions are written by technical staff, those who know exactly how a technical procedure must
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be performed. Finally, each document has considerable detail. Many instruct what is to be done, by whom, and when. Other documents, so-called forms, prompt for data entry and thus record what was actually done, who did it, and when it was performed. Corrections, reviews, and approvals are exactly recorded on each.
Documents are prepared by operational staff and reviewed by their super- visors and managers and by quality assurance specialists. Approval, that is formal signing of each document, is the responsibility of three or more indi- viduals: the writer, the reviewer, and a representative of quality assurance. Therefore, in a biotechnology firm, QAU has the major task of reviewing and approving each document generated by the development team and, in many cases, by each contractor, consultant, and service provider. This is further complicated by the fact that documents are often amended, and each change or amendment must be reviewed and approved by QAU.
To guide documentation activities, the QAU has, you guessed it, their own SOPs to guide quality activities. Notable are procedures for writing, review- ing, approving, and changing each type of document. QAU establishes and maintains an archive in which to place, in an organized manner, those important documents that customers, regulatory agencies, and investors might wish to review at a later date. Regulations clearly spell out the length of time a firm must keep operational documents.
Research activities are generally not under the prevue of a QAU. However, QAU may be asked to review and maintain laboratory notebooks prepared in research laboratories, despite the fact they do not come from development operations. Well kept, accurate and detailed research notebooks are impor- tant to any biotechnology firm since they are used for intellectual property applications and because they are valuable to a product development team as the basis for planning early development activities. For example, and as noted in Chapter 7, many quality control assays used to support product manufacture begin as research tools in the laboratory and these records are helpful if not essential to establishing early specifications. And the exact his- tory of genetic constructs, as produced in a firm’s research laboratory, are important documents to support safety claims of products derived from that research. A prudent biotechnology firm gives research documents the same care provided to documents from development.
Plans, protocols, procedures, and records instructing biotechnology oper- ations, defined in an earlier section, are used in each functional area, ensure consistency of operations, and are required by regulatory agencies. Each of these documents must be reviewed, approved, and distributed to users. Most are revised periodically, meaning the process is repeated at least annu- ally. Data captured on forms or in BPRs are also reviewed, approved, and archived. Even a small operation generates a large amount of data, further complicating the documentation task.
The list of operational documents does not end here. Many other docu- ments are identified in chapters throughout this book. But there is some relief
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for the QAU as they do not manage every record generated by the biotech- nology firm. Notably absent from their responsibilities are financial, human resources (excluding training), business and marketing records.
Investigate Situations: Manage and Control Change
Change is normal in biotechnology; changes are made to plans, proto- cols, and procedures. Any document or process or study can be changed but change is in a controlled, proscribed manner; hence the term change control. Earlier in this chapter, changes to product designs were discussed. However, downstream from the design phase, and in all functional areas, change also occurs and indeed is to be expected. Hence, all change pro- cesses within a biotechnology operation are managed and ultimately approved by QAU to ensure the integrity of each study, process or pro- cedure and to communicate a proposed or completed change to all par- ties involved. The QAU maintains procedures for change. If a document, study, or process is to be changed, then the individual with responsibility for that functional area recommends the exact proposed change to the product development team. The various departments involved review the proposal and they then discuss the risks and benefits as well as the technical issues or challenges associated with the proposed change. Discussion and agreement to an intended change improves the likeli- hood of making the correct decisions and thus preventing subsequent problems with a product or study, as might be the case if change was made without proper consideration of all factors. A proposal for change may go through several iterations and extensive discussion before it is finally approved by each member of the team and by QAU. The change control process is complex, often lengthy and thus involves careful docu- mentation and, finally, modification of a document, such as a protocol or SOP. As implied, a documentation system incorporates levels of interde- pendencies between documents. Therefore, in many cases a change to one document may warrant changes to another associated document that may be affected by this change.
The QAU also manages documents and approves activities such as devia- tions, the retrospective discovery that a procedure had not been correctly performed. Variance, change that must be made without proper discussion, review or approval, is another type of change that is handled by QAU.
Ensure Qualified and Trained Staff
Individuals working in biotechnology must have the training, education, and experience commensurate with their assigned duties. Operational area supervisors are responsible for ensuring this is the case for their employ- ees. However, because adequate training of personnel is critical to the safety of employees and the quality of biotechnology products, certain training
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responsibilities are given to QAU. In addition to maintaining training records for each employee, the QAU provides training on subjects related to quality, compliance and documentation, and record-keeping. The QAU also ensures that supervisors are qualified to provide technical training and confirms that each trainer maintains and follows a training plan and schedule. QAU staff members also coordinate training activities with senior management and evaluate training programs to ensure they are effective and compliant.
Perform Audits
Earlier, we noted auditing as a hallmark of quality and its importance to a quality system. Here, we discuss the performance of quality audits, internal and external, as managed and conducted by the QAU.
Quality audits support quality operations by applying two types of audits to compare actual performance and conditions to stated requirements (e.g., in SOPs). The external audit is conducted by a company engaged in an agree- ment or wishing to do business with another company. The vendor audit is an example. The internal audit is a firm’s own audit of its internal opera- tions. Virtually everything in the operational arena of a biotechnology firm is audited internally on a periodic basis or when problems are identified in a particular area. An example of an internal audit includes the review of study protocols, study records, and study reports at a nonclinical or clinical study site to ensure compliance with cGMP or cGCP, respectively, with the firm’s internal operation. In a manufacturing plant, an internal audit reviews SOPs, BPs, equipment and validation records, quality control testing, and records related to raw materials.
External audits may, for a reputable vendor, be performed by checking credentials, reviewing certificates provided by the vendor or performing telephone interviews and reference checks. However, for key materials and services, a quality auditor visits a vendor’s manufacturing facility, nonclini- cal laboratory or clinical site and carefully inspects to ensure compliance with expected quality criteria. Reputable product and service providers to the biotechnology industry are frequently audited by their clients. Therefore, a vendor or contractor who denies or evades a quality audit without good reason is suspect.
Several rules apply to the auditing process. The auditor must be independent of the entity being audited. For internal audits, the auditor and the audited department typically have parallel but independent reporting schemes in the corporate structure. In a practical sense, this means the QAU performs the audit and reports directly to a senior executive in the biotechnology firm. Even small firms may have a person trained to perform audits. Hence, audits and audit reports have become a major means of ensuring quality of a prod- uct or service. Careful preparation is important for success of any audit and usually outlined in a prospectively developed audit plan to include the pur- pose and scope of the audit. The format, length, and organization of the audit
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report should also be considered before the audit is initiated. The auditing firm must address several questions in the audit plan. For example, what is the purpose of the audit? Are there specific issues or problems with the system being audited? What is being audited and who are the individuals involved? The auditor should be carefully selected by QAU. It would not be correct to send a very thorough and detail-oriented individual to perform an audit that was intended as a superficial overview of a vendor’s quality system. Alternatively, if there was a need to examine in detail any aspect of that quality system, for example, a biopharmaceutical aseptic fill operation, then an expert in this area should perform the audit.
Audits performed by QAU typically focus upon whether procedures exist and if they do, whether or not a technical operation is following that pro- cedure. However, during the audit it is not uncommon for the auditor to observe a technical aspect of the actual operation that the auditor, in their opinion, believes to be erroneous. For example, an auditor observes that a manufacturing procedure is being performed according to SOP, that the operator is properly trained and keeps accurate and proper documentation. Yet, the auditor disagrees that this procedure, as written and performed, is not technically incorrect or unclear. Such issues arise in many audits and they beg the question of whether or not the auditor is justified or even quali- fied to critique the technical approach being used. If the auditor was from the U.S. FDA, we might say yes, the auditor may critique technical aspects, since U.S. FDA inspectors evaluate both technical and quality aspects of an operation. But what is the answer if the audit is internal or if the audit is per- formed by a biopharmaceutical firm on their contractor? One key to avoid- ing such issues is for QAU to be very specific about the scope and purposes of each audit, internal or external. Nonetheless, such issues frequently arise and they often lead to conflicts, which can be troubling, time consuming, and often difficult to fully resolve.
Performance of the audit is important to maintaining validity of the outcome. QAU is responsible for ensuring audits are planned, performed to standards and procedures and fully reported with proper action taken to resolve findings. The audit process is outlined in Figure 5.8. An audit begins with extensive planning. For an external audit, QAU seeks input from management and functional area supervisors. An audit plan identi- fies the purpose and scope, standards and procedures and, in the case of external audit, the plan is typically reviewed with internal managers before the audit is conducted. Most audits begin with an introductory meeting at which agenda, participants, purpose, and scope are confirmed. It is com- mon for such audits to adopt a systems approach to obtaining information in an audit. Most audits focus upon documents, both prospective and instruc- tional documents such as SOPs and performance or data records. The facil- ity, such as laboratories or a manufacturing area, may then be inspected by the auditor. This is critical because a quality audit is performed largely to demonstrate that procedures were in fact performed according to written
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procedures or requirements. Notes are taken by the auditor. At the conclu- sion of the audit, a closing meeting is usually held so both parties have an opportunity to discuss the findings of the audit and perhaps resolve any apparent discrepancies or misunderstandings.
Every audit results in a written report, prepared by the QAU’s auditor, to relate important and relevant findings or discoveries. The report states the facts and cites regulations or guidelines, but without being judgmental or finely interpreting regulations. The audit report may also make recommen- dations when deficiencies are found, but it should not mandate exact pro- cedures to resolve these issues. Functional area supervisors at the audited
Plan Identify scope, standards,
and procedures
Conduct
Meet Ask questions
Inspect
Review documents
Examine situation
and facts
Prepare listing of findings
Report
Findings
Communicate
Write Meet
Concur Complete report
Actions
Input
QAU procedures and management
recommendations
FIGURE 5.8 The quality audit processes.
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entity are left to take corrective action but the auditor follows up to ensure that corrective action has adequately addressed the original issues.
There is another side to viewing audits, since every biotechnology firm will, at some time, be audited by an outside party. This may happen as due diligence, the result of pending business arrangements, as an inspec- tion by an interested party, such as a regulatory agency, a client or a col- laborator, or it may be a potential or active customer, someone wishing to purchase materials or services. When audited, employees should make full disclosure of any records requested by the auditor but they are not responsible for volunteering additional materials or information that is not specifically requested. It is in everyone’s interest to be ready with complete, unambiguous, and well-organized materials for the auditor to examine. The entity being audited typically sets the tone for the audit and a positive, cooperative tone is especially important since, like a dental appointment, getting it completed quickly and painlessly is in the best interest of all parties.
Initiate a Quality System for a Biotechnology Operation
Once a biotechnology firm decides to develop a product and enters the regu- lated or quality compliance arena, it finds an effective means to build qual- ity systems into the proposed development operation. As described earlier, quality planning, specifically the quality plan and quality manual are key to initiating an appropriate and effective quality system, one that will grow with the operation. Quality manuals and quality plans are living documents and may be changed as operations increase in development phase or scope. Thus, while quality assurance is seldom the first operational element at any biotechnology firm, it must be adopted early and then it soon becomes an integral part of the development effort. The excitement and expense of enter- ing into product development sometimes obscures this immediate need for quality function; if this happens, quality efforts lag and adequate systems will likely suffer as a result. Nonetheless, senior management support of the QAU enables the establishment of an effective quality system, which sup- ports the other operational areas.
Therefore, where does one begin in this quality assurance process? The need for a quality assurance role at the biotechnology firm is often spelled out by a consultant or by a new employee who has worked in a regulated biotechnology environment and is familiar with quality and compliance. Or, it may be stimulated by recurring problems in operations. However, it begins, the growing biotechnology firm may retain an experienced quality consultant, someone who has built a quality department within a growing
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operation, or they may elect to hire a quality professional with the same experience. Few succeed in establishing an effective quality assurance func- tion by relying solely on tasking an inexperienced or untrained staff member with such responsibilities.
The next step is to generate management support and then quality poli- cies and plans that focus on the mission and operations of the organization. Budget must be considered, of course. The quality function is tailored to fit the PDP and it is written in parallel to, or as part of, that overall plan. Establishing a QAU is especially challenging for the virtual biotechnology company, the firm with less than 10 full-time employees managing a full product development program. Here, it may be necessary to delegate, over a long period, quality functions or oversight to a contractor or consultant.
Many quality issues face the maturing or fully mature biotechnology firm, defined here as 100 or more employees working in a fully opera- tional environment. First is the inability to sustain growth of the qual- ity function. Unlike the virtual or start-up operation, the maturing firm already has a QAU but it may be woefully understaffed, with manage- ment emphasizing rapid product development over quality programs. Indeed, mid-sized firms can, for various reasons, have less concern for quality functions than do smaller, younger companies. Functional man- agers may ignore quality; or quality staff may experience burn-out and lose interest. A weak QAU is easily detected during business due dili- gence or inspections by regulatory authorities and this lowers the value of a biotechnology firm in the eyes of potential business investment, part- nership, or purchase.
Operational growth requires additional resources for quality efforts and so growth in quality requirements are planned and budgeted. Growth requires addition of specific quality skill sets. A documentation specialist may be needed or an experienced individual is required to perform audits. Mundane issues, such as space to work and secure files in which to archive all those documents, face the growing quality operation. The key to success- ful growth of the QAU in a mid-sized biotechnology firm is effective and timely quality planning through revision of the quality plan.
Unfortunately, some biotechnology firms have, for whatever reason, a largely dysfunctional QAU. This situation often results from inadequate planning, poor management support and, unfortunately, ineffective lead- ership. Such QAUs first require immediate management involvement and support. This does not mean simply throwing money at the problem but more often requires investigation followed by organizational change or res- olution of issues, for example, resolution of interdepartmental squabbles, by upper management. Inability for any one department to operate effectively is often a reflection of an ineffective project team. Once the root cause has been identified, then senior management begins to repair the quality sys- tem and QAU.
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Unique and Effective Approaches to Quality Management
Risk-Based Approaches to Quality Systems
Risk-based approaches are a popular and effective means of ensuring quality in development. The U.S. FDA recommends risk assessment and management as a means to enhance and modernize pharmaceutical and biotechnology manufacturing and product quality. This initiative uses a scientific framework to find ways to mitigate risk posed by medical devices, drugs and biotechnology products.
Risk-based approaches in quality assurance examine a biopharmaceutical operation as a process and then identify those areas within that process that pose the greatest risk to the product. They also examine issues or problems associated with a product or particular type of study, such as number of fail- ures. It focuses efforts early in the life cycle of product development. This naturally fits with quality development efforts for any product. Risk man- agement then uses the scientific method to examine the risks and to address and lessen those risks using appropriate quality systems. Continuous, real time quality is a hallmark of this approach.
Risk analysis and management procedures are described further in Chapter 2, since they typically involve multiple operational areas and often fall largely within the purview of project managers. Nonetheless, the QAU has critical functions and plays an important in this area, often identifying risks and recommending that risk approaches be initiated or completed.
Total Quality Management
TQM aims at customer satisfaction. It has been adopted by many firms, including large biotechnology companies, and is especially popular with sales and marketing groups. It is a structured system for satisfying internal and external customers and suppliers by integrating the business environ- ment, stressing continuous improvement, refining development processes, encouraging maintenance cycles, and changing, for the better, organizational culture. The term structured system relates to the fact that TQM relies upon principles of quality systems and an environment that fosters such systems. TQM has three cornerstones:
• Everyone and Everything: total quality involves every individual and all activities in the company.
• Quality: conformance to Requirements (meeting Customer Require ments).
• Management: quality can and must be managed. As one might imag- ine, TQM must be driven from the top.
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Six Sigma
This program has been adopted by many firms worldwide as an avenue to produce quality products and reduce customer complaints. A major objec- tive is to incorporate a quality system that is so effective that less than 5% of a firm’s revenues are used to address and repair quality issues. Specifically, it aims to reduce product or service failure rates. The six sigma process encom- passes all aspects of a business, including management, service delivery, design, production, and customer satisfaction. As compared to an operational quality system such as cGMP, in which only the manufacturing and control departments are directly affected, six sigma involves every aspect of a firm.
Statistics in Quality Assurance
Quality Assurance collects a significant amount of data and uses this data to map trends and plan future endeavors or prevent recurrent problems. Statistical analysis of that data is critical to making decisions and taking proper actions. One outcome of statistical analysis is a control chart, which graphi- cally represents one or more aspect of the quality process and identifies varia- tions beyond limits. As such, statistics is a tool that allows the QAU to monitor processes and provide meaningful information to functional area managers and upper management. The quality programs described earlier, such as six sigma or TQM are dependent upon statistical analysis of quality data.
Quality Systems for Research
What quality considerations should be given to (or imposed upon) biotech- nology research laboratories? Is there a compelling business reason to estab- lish a quality system for research efforts in any context? Application of a full quality system may be helpful for research quality, but in many cases it may actually hinder research endeavors. Research laboratories are for dis- covery and not for structured development. Research results must be of high quality but discovery research does not directly lead to products or to users. Particularly frustrating are attempts to impose on a discovery research labo- ratory a formal quality system, such as cGLP, when it is neither needed, by regulation, nor helpful to achieving objectives. The quality standards applied to research and development or commercial applications are different.
However, some hallmarks of quality, mentioned earlier in this chapter, can be very effective at improving productivity and reproducibility in the research environment, at least if they are applied correctly. For example, vendor control is an excellent way to save time and money. An effective documentation system can be very supportive of patent applications and improve records upon which future product development efforts depend. Hence, the small firm, engaged exclusively in research, is encouraged to institute quality hallmarks that help their laboratories achieve success
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without burdening research efforts through establishment of a fully com- pliant quality system. Other hallmarks of quality, notably training, man- agement responsibility, design of experiments, or vendor control to name a few, can greatly improve the productivity of a research laboratory.
Resolving Quality Issues or Problems
The quality assurance professional takes on huge responsibilities and has great authority within a firm. He or she reports to upper management and may be more influential than many other functional area managers. One example is in a growing biotechnology firm, where the QAU might ques- tion whether a clinical study site is fully qualified, under cGCP, while the clinical manager strives to meet an already challenging schedule to begin a study. Another example, this in manufacture and control, is whether or not to release product for a clinical study because a specification was not fully met by analytical results. Specifications for product in early develop- ment can be ambiguous in certain respects and individuals on the product development team may disagree on whether it passes or fails. In another example, the QAU may question the validity of an important aspect of an expensive nonclinical study and suggest that portions be repeated. Note that repeat testing has a set of regulatory guidelines around the nature and frequencies of appropriate retests to be conducted. These quality opinions have a tremendous effect on day-to-day operations, expenditures of time and money and, in the end, the success or failure of a biotechnology firm. Such situations often leave the quality assurance professional in the hot seat. Since disagreements are not infrequent and because operational depart- ment directors disagree on important matters, tempers may flare or dis- agreements linger and fester.
Well-led project teams are perhaps the best means of resolving differences while ensuring correct decisions. Several guidelines must be considered. In the biopharmaceutical industry, FDA regulations make it clear the QAU has, under cGMP, cGLP, and cGCP, the final word on matters relating to quality, even though a high-level manager can and has been known to reverse QAU decisions. Second, individuals with different backgrounds often perceive the same situation or interpret the same data quite differently. We see this poli- tics, in the mass media and at scientific meetings; science is not immune to disagreements. These differences can lead to animosity and disregard for the other’s opinion. Great quality professionals are therefore good at nego- tiation, which is based upon understanding the other person’s point of view and then trying to work within that opinion to reach a solution or common understanding. This, in turn requires them to listen carefully and to be patient. They also must clearly explain the reason for a judgment and they are well advised to seek the opinion of regulatory professionals. Again, a good project management team with a strong project manager is wonderful at facilitating negotiations, if only by setting a positive environment.
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Upper management plays a major role in preventing hostilities and resolving disagreements between the QAU and other operational areas. Misunderstandings in biotechnology operations are often the result of poorly-established or undercommunicated corporate and product devel- opment objectives. In these all-too-frequent instances, upper management bears responsibility. Management is responsible for defining quality policy and responsibilities and authority in the quality manual. Further, upper man- agement must recognize when communication has broken down between quality assurance and another department manager and then make every effort to resolve the differences and have each faction work together toward a common objective. To perceive developing issues in a timely manner, upper management must always be involved and alert. Project managers and team members ensure that management is engaged in development activities, including professional roles and disagreements.
Why is it so vitally important to identify quality issues in a biotechnology firm? First, these small firms are so fragile, very susceptible to failure for a number of reasons. Second, they have little depth—fiscal resources, product line, facilities—to rely upon in times of trouble. Third, the team has worked together for a brief period, as most firms are relatively new and develop- ment may have just begun. Differences related to quality aspects of the oper- ation could spell the difference between success and failure, especially in biopharmaceutical endeavors. Often, quality issues are for a variety of rea- sons invisible to insiders but most obvious to outsiders, such as consultants and auditors. Indeed, seasoned professionals in biotechnology have said that by examining the roles, authorities, and responsibilities of a QAU, one can quickly surmise a key indicator or success versus failure at a young biotech- nology firm.
This section on resolving quality issues has provided no magic solutions for problems one might encounter in a biotechnology operation. It aimed instead to summarize but a few of the situations one might encounter in the operational environment of a biotechnology firm. By understanding quality systems and through careful planning and effective management, quality functions and the quality professional who manage them, the qual- ity endeavors can be a valuable asset to any biotechnology development team and the firm they represent.
Summary of Quality Systems
Quality assurance is a planned and structured function designed to ensure each product or service provided by a biopharmaceutical firm will meet established requirements and user expectations. Quality planning and insti- tution of a quality system early in biopharmaceutical product development
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is critical to the success of any biotechnology operation. Hallmarks of quality are distinctive features of excellence comprising any quality system and include features such as management responsibility, definition of the qual- ity system, design and design control, contractor control, product identifica- tion and traceability, process control, environmental control, quality control or testing and release, change control and corrective or preventive actions, packaging and labeling, preservation, storage and handling, servicing, and customer concerns. The quality assurance function focuses on the quality attributes of quality management, documentation, investigation and change management, training, and auditing. Effective quality systems are devel- oped specifically for each process and product or service and established quality systems, such as cGCP, cGMP, ICH Q-10, cGMP, or ISO 9001, are often required of biotechnology firms, especially for biopharmaceutical develop- ment. Today, several unique and effective approaches can be adopted to bet- ter manage a quality system but their benefit must at least equal their risks. The QAU is the functional area that manages a quality system within the overall biotechnology operation. The QAU controls documents and manages the documentation system, investigates situations, manages and controls change, ensures all staff are qualified and trained to perform their duties, and performs audits, internal and external. Initiating or maintaining an effective and respected quality system in a biopharmaceutical operation is a challenging task and relies in part upon both technical and social skills of quality professionals. Further, the quality system must be matched to requirements and be always balanced with the operation, solving and not creating problems or issues.
References
British Standards Institute. 1991. Standards of BSI, BSI, Herndon, VA. International Standardization Organization. 1994. ISO 9001, Geneva, Switzerland. Oxford English Dictionary. Oxford University Press. 1997. University of Oxford,
Oxford, UK.
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6 Biomanufacture
Overview of Biomanufacturing Requirements
The biotechnology operation focuses on the development of a specific prod- uct. This concept carries with it the need to plan and then develop a bioman- ufacturing process to produce a biological substance of high quality and in amounts required for testing and marketing. Further, the biomanufacturing materials, processes, and the resulting product must be compliant, that is, they must satisfy regulatory agencies through application of good science and a quality system: current Good Manufacturing Practices (cGMP). To achieve these objectives, the biotechnology operation must develop a bio- manufacturing plan.
Even for the simplest product in the hands of experienced bioprocess engi- neers, biomanufacturing is a demanding endeavor and requires consider- able planning, time, and financial and human resources. False starts in the biomanufacturing pathway, which is usually the result of inadequate plan- ning, often lead to project failure and termination.
Hence, biomanufacturing planning begins early in the product develop- ment cycle and is based on the exact understanding of the product’s nature and intended use or indication. The overall product development plan (PDP) (Chapter 1) coordinates the manufacturing plan with plans for the quality assurance, quality control, and regulatory, clinical, and nonclinical aspects of product development. To ensure this integration, the biomanufacturing planning process requires leadership from biomanufacturing experts, con- siderable time and effort, and frequent interactions between individuals from various departments.
This chapter on biomanufacturing considers design and planning, pro- duction technologies, compliance and quality, major stages and steps of manufacturing for various types of biotechnology products, and the manu- facturing facility.
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Design in Biomanufacture
At the heart of a biomanufacturing plan is the manufacturing design or scheme—pictured from beginning to end—with the various control testing, quality, and regulatory elements that impact product production. The objective of biomanufacture is to produce a product that has the attributes, for example, strength, identity, purity, potency, and safety, commensurate with the intended use. Product attributes are further defined in Chapter 7. Each biotechnology product is unique and is, or will be, produced using both well-characterized and well-known commercial processes and special methods developed for that particular product or class of products. The flow diagram shown in Figure 6.1 is a general format or template used to design a product-specific biomanufac- turing scheme. Three stages of biomanufacture—(1) upstream processing; (2) downstream processing; and (3) formulation, fill, and finish—are the back- bone of a biomanufacturing design. In the first stage, upstream processing, the product is produced from raw materials by using process technologies such as cell culture, fermentation, and synthesis. The second stage, downstream processing, involves purification of the desired product by its separation from impurities and contaminants. In biopharmaceutical processing, the output is referred to as the bulk (drug) substance (BS). For a biopharmaceutical, the BS is also the active pharmaceutical ingredient, which means that it has the ther- apeutic activity. Stage 3 processing ensures that the product is fit for use, by applying the processes of formulation, filling into a container, packaging, and labeling. The result is a final product (FP), which is ready for use.
Quality by design (QbD) is a concept applied to all product development endeavors (Chapter 5) in biotechnology and is a critical and early aspect of biomanufacture. It evolved in part from regulatory and quality initiatives in the medical device industry late in the twentieth century. More recently, the principles and practices of QbD have been adapted to biopharmaceutical development. Product development or manufacturing QbD is driven in part because regulatory agencies have provided evidence that, when followed, QbD consistently leads to high-quality products. This represents a para- digm shift from the previous practice of simply testing samples of the FP for quality in the hope of demonstrating quality. Indeed, this past practice was referred to as testing quality into a product. This practice has now changed, and although quality control testing (Chapter 7) remains an important aspect of product quality, it is now recognized that production design and process practices play equally important roles in ensuring a pure, potent, consistent, and predictable quality product every time. In other words, quality should be built into the product by design rather than by solely relying on the FP testing. The biotechnology industry also recognizes that QbD makes good business sense. Janet Woodcock of the FDA defined QbD in 2005 as “a maxi- mally efficient, agile, flexible pharmaceutical manufacturing sector that reliably produces high quality drug products without extensive regulatory
197 B
iom an
ufacture
Stage 1 Upstream processing
Raw materials
Target product profile
Stage 2 Downstream
processing
Stage 3 Formulation, fill,
finish, and labeling
Genetic construct, expression vector
Package label
Aseptic fill
Formulation
Design of manufacture in a product
development strategy
Initial product capture
Purification 1
Purification 3
Testing
Staff, quality assurance, and quality control
Purification 2
and andandand
Research seed
Cell banks
Cell culture fermentation
Initial recovery
Testing
Testing
Testing
Cell paste
Stages:
Processes:
Outputs: Bulk
substance (BS)
Final product
(FP)
Testing
Testing
Testing
Testing
Testing
Testing
Inputs: Facility,
utilities, and equipment
FIGURE 6.1 General outline of biomanufacturing activities by stages or steps of biomanufacture. This flowchart traces the biomanufacturing scheme applied to many biotechnology products. Boxes in the upper row define inputs, that is, the resources required to begin biomanufacture of a product. The flow- chart below is divided into three stages typical of a complete manufacturing process and describes outputs (or results) from each of the three stages. Process elements, shown in the shaded flags, are typical for production of a recombinant protein product.
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oversight.” Yet again in 2014, elements of QbD were highlighted by Margaret A. Hamburg, Commissioner of Food and Drugs, in her outline of FDA stra- tegic priorities for the next 4 years. She stated that the continued use of risk- based approaches would help to ensure product quality in pharmaceutical development. (Hamburg 2015)
In its simplest form and as provided in the International Council for Harmo nization of Technical Requirements for Pharmaceuticals for Human Use quality guidelines (Chapter 4), the concept of QbD instructs the devel- oper to design a product, so it consistently meets the desired performance criteria and always meets expected quality attributes. This definition demands much of a product’s sponsor. First, it identifies the need to inte- grate a manufacturing plan into the overall PDP, described in Chapter 1. It also directs the use of formal manufacturing design process, in which the product designer considers and documents both the expected performance and quality attributes of the product.
QbD, as it applies to biomanufacturing and the biomanufacturing porting of the PDP, is shown in Figure 6.2. The nature of a biotechnology product,
Processes
Design space
QC plan:
TPP
User needs Regulatory requirements
Available manufacturing: –Raw materialsAvailable tests:–Assays
Input
Specifications
Limits
Specifications
–Processes –Facility and equipment–Laboratories
Quality by design Input risk analysis
Review output Document
Manufacturing plan
Quality requirements
Management Financial Personnel
LimitsLimits Specifications
–Tests –Specifications
–Release –Stability
–Validation
Manufacturing process plan: –Stages –Steps
Facilities, equipment, and validation
Output and PDS
FIGURE 6.2 Quality by design in biomanufacturing. Manufacturing design begins with inputs, notably user needs and regulatory requirements, that are synthesized into a targeted product profile (TPP). A design space has specifications and limits as boundaries and quality requirements, available manufacturing resources, tests and management, and resources as inputs. This allows the manufacturing process to be designed within a design space, represented in the center. Outputs of the design process include a quality control plan; the manufacturing process plan; and a strategy for facilities, validation, and other requirements.
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as provided in a targeted product profile, must be carefully considered in the manufacturing design. QbD requires that a manufacturing process be designed using scientific approaches, quality criteria risk management, and design space. QbD prompts the need for application of design con- cepts: input, output, reviews, design space and specifications, and ranges of acceptable values or limits. These are discussed in Chapter 1. Design space, as it refers to biomanufacturing and control activities, is the requirement to design within limits or boundaries. For planning biomanufacture, the limits are imposed by constraints of manufacturing technology and by product specifications. Specifications, further defined in Chapter 7, are measurable quality criteria for a product. Although these boundaries on design space restrict initial manufacturing design, they allow later changes in the manu- facturing processes, with minimal justification, as long as the changes are implemented within the design space and consider the specifications. This allowance is based on the premise that changes to a biomanufacturing pro- cess made within a reasoned design space will most likely not change the quality of the biopharmaceutical product. Planning under QbD helps to ensure that a biomanufacturing process will be robust and the product will predictably be consistently of high quality throughout the life cycle.
As shown in Figure 6.2, QbD also applies the concepts of input and out- put to biomanufacture. Input, notably user needs, the nature and the profile of the product, and manufacturing and control technologies are considered within the confines of design space. Within the design space, active manu- facturing design and review lead to the output, notably plans for the appro- priate processes, facility, and quality control tests.
In manufacturing design, the concept of risk analysis, elaborated upon in Chapter 1, is considered in light of how the manufacturing process and the design space might impact the user of the biopharmaceutical product. Any biotechnology product carries some inherent risk to the user or to the public. Other risks are imparted through the product’s manufacture. Both design and manufacturing planning activities identify and attempt to mitigate all risks. QbD considers these risks in a logical manner and demands that the design should take into account any possible risk.
Biomanufacturing design and planning are greatly impacted by affordable biomanufacturing technologies available for a given type of product and process. For a given product, the biomanufacturing planner has a variety of process methods from which to choose. The process technologies chosen during design are added to the plan and referred to as input. The results of these technological applications are known as output, which also become embodied in the manufacturing plan. Consider, throughout the remaining chapter, the need to design a biomanufacturing scheme, applying product limits and specifications, the inputs and outputs of a design, and the need for understanding process and product risk at every point. These concepts help us better understand why products are manufactured in a certain manner. Finally, some biomanufacturing process change is inevitable for even the
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best conceived biomanufacturing plan. Consideration is given for making changes in a manufacturing plan as long as the changes are kept within the boundaries of design space and the risk is carefully considered.
Technical Considerations for Biomanufacture
Biomanufacturing is a relatively new field, which has expanded, by quan- titative and qualitative measures, rapidly. Before 1970, the manufacture of biological products was accomplished largely by purification of biologically active molecules from various natural sources. For example, albumin was precipitated from human plasma and then separated from other blood pro- teins. Some years ago, vaccines were strictly natural products, such as sub- units of viruses or protein toxins, derived from microbes grown in culture. Although these methodologies are still considered biological technologies, the advent of genetic engineering led to the endeavor we now refer to as biotechnology. Genetic engineering made possible the transfer of genes from one organism to another, and this science allowed us to genetically modify bacteria or mammalian cells, which in turn led to biomanufacture, the pro- duction of small amounts of recombinant proteins or nucleic acids. To make large amounts of these recombinant products, first for further evaluation and then for commercial use, biomanufacturing protocols and technologies or methods were expanded. Biomanufacturers soon discovered that product quality was important to ensure proper function. If a molecule were of poor quality, it would not perform as intended, when used in critical test proto- cols, such as in animal or clinical studies. Notably, when a manufactured bio- technology product failed to perform consistently, the sponsor was left with little product value and doubts about the product’s utility and marketability.
It was also recognized that quality control testing of a manufactured biotech- nology product, alone, did not ensure product quality. Indeed, product qual- ity reflected both the processes and the technology applied to biomanufacture, such as facilities, utilities, and equipment used in biomanufacture. Consistency of manufacture was also critical to achieving the desired attribute; the biophar- maceutical product had to be the same every time it was manufactured. Hence, commercial biomanufacture demanded attention to product consistency.
Rapid advances in biotechnology have challenged the young field of bio- manufacture in other ways. Many organisms capable of expressing recom- binant proteins expanded greatly in just three decades, and the types and classes of biotechnology products that must be manufactured by our indus- try continues to both expand and diversify. More advanced expression systems that utilize somatic or stem cell engineering are the examples of rapidly growing scientific methodologies that have brought about the need to develop and apply new biomanufacturing technologies. Transgenic plants
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and animals have become commonplace, and biologically active molecules are now regularly processed from these sources. Synthesis of biologically active molecules is a field that continues to expand. Other challenges include developing novel products, refining old processes to produce currently mar- keted products in a more economic way, improving the quality or consis- tency of investigational and marketed products, and engineering production of generic or follow-on biopharmaceuticals—new products that are safe and effective, exactly like a predicate product.
To meet these challenges, careful planning and development of new manu- facturing technologies, analytical tools, and processes continues unabated. Biomanufacturing scientists continue to invent, apply, adopt or adapt processes, procedures, and skills to meet this increase in demand. Facilities and equip- ment have been designed or redesigned, built, validated, and commissioned to house and support these more sophisticated processes. In summary, there is much activity in the field of biomanufacture, which is leading to excellent mar- keted products, and each success is based on proper manufacturing planning and design and the ingenious application of existing and novel technologies.
For the remaining chapter, we present an overview of the stages and steps used in biomanufacture, identify technical considerations for vari- ous processes, and integrate quality and compliance into biomanufacturing schemes. We then apply biomanufacturing criteria and technologies to sev- eral classes of biotechnology products, highlighting differences and similar- ities of various products. At the end of this chapter, we describe the design, use and validation of biomanufacturing facilities, utilities, and equipment. Quality control and quality assurance activities are closely associated with biomanufacture, and these are discussed in Chapters 7 and 5, respectively.
Phases and Scale-up: The Biomanufacturing Life Cycle
Biomanufacturing is performed throughout the life cycle of a product. We iden- tify phases in the life cycle and further ask the biomanufacturer to ensure that a product possesses particular qualitative and quantitative attributes or traits in each phase of development, with process and product specifications becoming increasingly more stringent as the cycle progresses. For biopharmaceuticals, manufacturing phases of development follow those applied to clinical studies (Chapter 9): Phase 1 (early phase), Phase 2 (mid phase), Phase 3 (late phase), and Phase 4. This approach makes sense because we use product in greater amounts as the number of human subjects increases at each clinical phase. At Phase 1, requirements are in the hundred of doses, but as product approaches the mar- ketplace, product might be needed for millions of users. Compliance issues, specifically adherence to cGMP, also increase in intensity and importance, as biomanufacturing development increases through the phases. Both total
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amount of product and quality criteria undergo change, as the product is used in a greater number of individuals. These relationships between clinical phase, biomanufacturing phase, product quality, and product quantity are shown in Box 6.1, and these relationships must each be considered in a manufacturing plan. A greater amount of product is produced in each subsequent phase, and along with this comes the need to better characterize the product and to meet ever greater compliance standards through improved quality and production systems. Phased product biomanufacturing development is a dynamic process, and change is desirable and normal. How this change is anticipated, planned, controlled, and executed in the manufacturing plan is critically important to the overall product success.
The quality of product required at Phase 1 clinical studies is, to some extent, mandated by cGMP, but it is also a function of the indication, intended use, and proposed manufacturing process. In planning and devel- oping the process, it is important to consider that greater amounts of prod- uct will be required later in the development; therefore, a biomanufacturing process must be amenable to change to accommodate scale-up and to meet more stringent quality and compliance criteria. Process control, quality con- trol testing, and consistency of manufacture are important measurements that can demonstrate product quality at Phase 1. Hence, the process is well- defined and some quality control assays are established before initiation of biomanufacturing. In early phase development, it is best to produce multiple batches of BS and multiple lots of FP to understand and control variation. This information can be used to ensure consistency of manufacture— amount and quality—from batch to batch and from lot to lot.
At mid-phase development, the quality objectives are to confirm and extend the findings of early phase biomanufacture. Biomanufacturing scale-up at mid-stage further tests the application of quality criteria to the process and to the end product. Product manufactured at mid stage is used in those clini- cal studies, and it is also applied to refinement or qualification of analytical tests. Process improvements are often implemented and tested to ensure that changes yield a product with the same or better quality attributes than those seen at early-stage development. The ranges of acceptable values for product specifications, both process and quality control, are often narrowed at Phase 2. Mid-phase biomanufacture provides a product that is used for the qualifica- tion or validation of analytical tests and also for additional stability studies. It is not uncommon for a biomanufacturer to miss critical mid-phase manu- facturing objectives. Failure in achieving consistent manufacture may result in the need to abort the ongoing processes. In such situations, it is important to review the manufacturing plan, make changes, and then repeat mid-phase biomanufacturing before progressing to scale-up or late-stage manufacture. Unfortunately, such advice is too often ignored, leading to biomanufacturing failures at Phase 3, which result in the need to repeat manufacturing devel- opment, often including Phase 2 and Phase 3 biomanufacturing and clinical studies, both very expensive propositions.
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BOX 6.1 BIOMANUFACTURING ACTIVITIES BY PHASES OF BIOPHARMACEUTICAL DEVELOPMENT
Phase Design and
Plan Manufacturing
Processes
Quality Control
Laboratory Quality and Compliance
Planning (0) Targeted product profile
Product development strategy
Technology Documentation
system
Develop constructs Technology
transfer from R&D laboratory
Cell bank development
Research laboratory development of critical analytical tools
QC constructs and cell banks
Identify regulatory guidance
Establish quality plan and basis for quality system
Ensure quality assurance activities
Early phase (1) Implement design and process development schedule
Accept constraints Produce clones
and cell banks Perform the
process two times or more
Produce product for nonclinical and Phase 1 studies
Establish certificate of analysis with product attributes, tests, and specifications
Test Phase 1 products
Evaluate final product stability
Institute Phase 1 cGMP compliance
Formalize training system
Manage documentation system by using version- controlled documents
Mid-phase (2) Refine plan based on findings
Scale-up for multiple batches and lots
Phase 3 requirements
Adjust process and refine steps
Further develop assays, qualify critical tests, refine specifications and add new tests
Test Phase 2 products
Expand viability testing
Increase scope and depth of cGMP compliance
Implement more qualification activities
Perform internal and external audits
Late phase (3) Plan commercial process and validation activities
Execute multiple lots at or near commercial scale
Validate process and facility
Validate or verify each assay
Test product at scale-up and for Phase 3
Expand viability testing
Come to full cGMP compliance, as applied to commercial production
Approve validation
Postlicense/ commercial (4)
Plan and document all change
Manufacture for commercial market
QC for commercial product
Approve change Maintain full
cGMP
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Mid-phase is the best time to make significant and necessary process changes. Process changes may result in changes to the purity or potency of the product, and these may negate the validity of nonclinical and clinical data generated during Phase 1 and Phase 2 studies. For these reasons, every effort is made at this time to improve the process without changing the molecular or cellular nature or the quality profile of the biotechnology product.
Late-phase biomanufacturing development focuses on preparing material for Phase 3 clinical studies. Production at this stage also ensures a robust process at greater scale, and the late-phase product is used for further assay development or validation and stability studies. Biomanufacturing process validation, described later in this chapter, is another objective of the late- stage biomanufacturing program. Manufacturing changes, in addition to scale-up, can be instituted at late stage, but they must be thoroughly ana- lyzed to ensure that the product remains consistent in quality with the mate- rial made in the earlier phases and used in clinical and nonclinical studies.
To this point, we have discussed changes to biomanufacturing processes as a qualitative perspective. We now cover the subject of quantitative manu- facturing changes or scale-up. Early phases of biomanufacture yield limited amounts of product, certainly not a sustainable quantities required to address market demand. Biomanufacturing scale-up, depicted in Figure 6.3, is used to gradually increase the total amount of the product that is available from each production run. A production run is a distinct series of processes, and each run results in one batch of BS or one lot of FP. Scale-up must be considered within the confines of design space. One should, at the outset, have an idea of commercial requirements and then base a scale-up plan on these require- ments. Scale-up may follow several pathways. Scale-up may be achieved by increasing the yield (1) by increasing the amount of product produced within
Nonclinical Phase 1
Appl icabil
ity of good
man ufact
uring prac
tice r egula
tion
Prod uct c
harac teriza
tion
Phase 2
Phase 3
Quality systems
FIGURE 6.3 Manufacturing by phase of development. Simultaneous increases in product quantity, quality, characterization, and regulatory compliance through phases of biomanufacture. (Courtesy of Anthony Clemento, 2008.)
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a batch or a lot; (2) by upping the scale of manufacture for each batch or lot; and (3) by increasing the number of manufacturing runs. Often, a scale-up plan considers two or even all three methods to increase the total amount of BS and FP. This is particularly important with therapeutic biopharmaceuti- cals, as small changes in molecular structure may have significant implica- tions on function, thus translating to safety or efficacy outcomes for patients.
Scale-up is an expensive process, which is not typically initiated until suc- cess has been demonstrated in clinical or field studies. Scale-up has a greater impact on the production of BS than on the production of FP. Scale-up of final drug product manufacture often involves increasing the size of the formula- tion batch by using larger vessels, by building another production facility, or by outsourcing to a qualified contract manufacturing organization (CMO) that has the capacity to perform a greater volume of fill, finish, and labeling. Of importance, scale-up to obtain greater amounts of BS most often cannot be economically accomplished by simply multiplying the number of bioreac- tors or fermenters, or by installing several rows of chromatography columns. Instead, scale-up typically involves developing new or modifying existing technologies to produce much greater volumes in a single batch (i.e., a single large fermentation vessel). Biomanufacturing systems are often finicky when it comes to scale-up; hence, there is a need for imagination and extensive experimentation. At large scale, the experimental processes require a sig- nificant investment in equipment, are costly to perform, and require large amounts of raw materials. This scaling process forces the operator to devise BS production systems that appear to be quite different from the smaller sys- tems used in early phase biomanufacture. Process and laboratory controls are continually applied during the scale-up process, as changes to yield are likely to impact product quality. It is an important consideration to limit all changes to a process within the designated design space.
Raw Material Considerations
Like any other manufacturing endeavor, biomanufacturing requires raw materials, which are sometimes referred to as components. These provide the structural building blocks, dynamic metabolic energy source, and bio- manufacturing environment for every product and process. Raw materials such as water, gases, salts, and nutrients are critical elements employed at every phase of the biomanufacturing process. The quality of each raw mate- rial should remain unchanged throughout the manufacturing cycle, but amounts increase with scale-up. Requirements and specifications of raw material are included in a manufacturing plan.
Box 6.2 presents a list, albeit incomplete, of raw materials that may be used in upstream production, such as fermentation of yeast, to produce a recombinant
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protein. Box 6.3 provides a list of raw materials that may be used in downstream production, that is, in the purification process of that product. Since the quality product output is, in part, a reflection of the quality of the input raw materi- als, biomanufacturers, and regulatory authorities take the source and quality of each raw material very seriously, no matter how insignificant it may appear to the process or application. Special consideration is given to the raw materi- als that contact or are incorporated into a FP. Raw material specifications and acceptance criteria are critical to consistently meeting the standards set forth in manufacturing plans and procedures. The possibility of raw materials contain- ing toxins or adventitious agents is especially noteworthy, because these impu- rities present risks to the user, and because once introduced into the product stream, these may be difficult to detect and remove.
A raw material for biomanufacturing may be purchased from a ven- dor or it could be produced in-house, by the product manufacturer. For example, sodium chloride is typically purchased, whereas highly purified water, water for injection (WFI), is often produced in the sponsor’s facility. As you might expect, raw materials are a highly controlled commodity in the biomanufacturing arena. To prevent misidentification or contamination,
BOX 6.2 EXAMPLE OF A MATERIAL LIST: UPSTREAM FERMENTATION
Material Number
Description and Attribute Source Specification Comment
H2–115 Working cell bank TA Biotechnical
CoA Manufactured 01/11/2010
145621 Yeast nitrogen base without amino acids
DB/Fidco CoA No animal product
RX001 Glycerol Spectarm USP No animal product
LC1121 8 N Ammonium hydroxide
TJ Booker USP Not applicable
31772 Glucose/dextrose TJ Booker USP Not applicable 32274 Tryptic soy agar
plates Remel SOP QC-1181 Passed
32371 Yeast peptone dextrose plates
Remel SOP QC-1181 Passed
16–2010 Water for injection TA Biotechnical
SOPs MF-1141 and QC-1832
Passed
4–115 Biotin Spectrum CoA Not applicable
Note: Manufacturer’s material number: CoA, manufacturer-provided certificate of anal- ysis; SOP, internal testing by QC laboratory standard operating procedure with specification and passed by QC and QA; USP, U.S. pharmacopeia-grade material; animal product, manufacturer- provided certificate ensuring that no animal prod- uct was used in this material.
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vendor-supplied raw materials are inspected, clearly labeled, sometimes retested, and then kept in controlled storage areas of the manufacturing facility until used in the manufacturing process. Quality of raw materials is further discussed in Chapters 5 and 7.
Compliance and Quality in Biomanufacture: Current Good Manufacturing Practices
Quality considerations for biomanufacturing begin with design and plan- ning and continue throughout the life cycle of a product. In the United States, biopharmaceutical manufacturing quality is promulgated in a set of regula- tions known as cGMP. Other countries also have manufacturing guidelines,
BOX 6.3 EXAMPLE OF A MATERIAL LIST: DOWNSTREAM PURIFICATION
Material Number
Description and Attribute Source Specification Comment
1–110 Clarified fermentation supernatant
TA technology Batch production record-661–00
Manufactured 01/10/2011
040721 Water for injection TA technology SOPs master formulation-1141 and QC-1832
Passed
SF-1418 Sodium phosphate (monohydrate)
TJ Booker USP Not applicable
TM0012 Sodium hydroxide, pellets (NaOH)
Spectarm USP NF Not applicable
SF-1416 Sodium phosphate (dibasic) heptahydrate
TJ Booker USP Not applicable
C3HN5– 9990
Millipak-20 filter units (0.22 µm)
Milepour CoA Meet specifications
65 SD105 Superdex 200 chromatography gel
EG healthcare CoA cGMP grade
30 SO672 Sepharose high performance chromatography gel
ED healthcare CoA cGMP grade
Note: Manufacturer’s material number: CoA, manufacturer-provided certificate of anal- ysis; SOP, internal testing by QC laboratory standard operating procedure with specification and passed by QC and QA; USP, U.S. pharmacopeia-grade material; and NF, national formulary.
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and for the biotechnology firm intending to export a biopharmaceutical, attention must be paid to directives from European, Japanese, Canadian, World Health Organization, and other national and international agencies or organizations. In addition, the International Conference on Harmonization of Technical Requirements for Pharmaceuticals for Human Use has guide- lines on manufacturing quality. These references are further identified and discussed in Chapter 4.
Biotechnology products and raw materials, those other than biophar- maceuticals, also have manufacturing and product quality criteria, either known as an industry standard or established by industry trade organizations, national or international bodies, and regulatory authorities. For example, the International Standards Organization guides activities and establishes stan- dards for biomanufacturing and thousands of other industrial endeavors.
Good manufacturing practice guidelines, no matter what the standard or guideline source, are followed by biomanufacturers, first because they are regulatory requirements and second because they are logical for business development, product marketing, financial stability, and product liability reasons. Product recalls are expensive for a biotechnology firm, and adverse events due to production and release of substandard product can devastate the reputation and lead to financial ruin, even for a large company.
The objective of cGMP is to consistently produce and deliver the highest- quality product to the user. Today, cGMPs apply beyond production activities in a biomanufacturing facility. They encompass the concepts of biopharma- ceutical design, risk analysis, and manufacturing planning, the functions that begin well before the product even enters the facility and extend to warehous- ing, an activity found at the far end of the biomanufacturing development pathway. The full embracement of cGMP, the U.S. FDA regulation, is phased into the manufacturing plan, as shown in Figure 6.3. Phase 1 manufacturing, under cGMP, ensures that raw material and process hazards are identified. Steps are also taken to ensure that these hazards do not endanger human subjects during any phase of clinical investigation. However, FDA recognizes that not all aspects of cGMPs apply to a given product, especially in early development, and FDA offers additional guidance for Phase 1 manufacture of biopharmaceutical products. Application of cGMP requirements is consid- ered in a manufacturing plan by focusing on product-specific attributes and quality issues that might affect biopharmaceutical manufacture for nonclini- cal and Phase 1 clinical studies. Risk analysis of the manufacturing plan is one way to identify quality issues, and apply cGMPs to production processes in a rational manner.
The plan also considers manufacturing compliance as product moves through subsequent phases of manufacture and greater numbers of subjects are exposed to a product. Now, cGMP application has broadened and has become increasingly stringent for each stage of biomanufacture. There is a heightened level of importance and need for cGMP in certain processes such as aseptic technique or sterile fill, because these processes are critical to
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ensure product safety. With respect to sterility and several other manufac- turing controls, there is a single interpretation of cGMP and it applies from Phase 1, or early phase biomanufacture, to commercial manufacture. Yet, less risky processes have little impact on safety and thus are of lesser con- cern. Therefore, the concept of cGMP application is considered a gradient, beginning with cGMP at Phase 1 biomanufacturing and increasing through commercial production and weighing the risk of a practice or material at every phase. The cGMP regulations are outlined in Chapter 4. Specific exam- ples of quality criteria and application of cGMP are provided in subsequent discussions of biotechnology products and biomanufacturing technologies in this chapter.
Biomanufacturing Processes for Biotechnology Products
The discussion on biomanufacture now shifts from general subject matter to focus on various biotechnology products and the technologies used to manufacture those products. We begin by reviewing standard production methods used to biomanufacture recombinant proteins in living cell-based systems and follow with a discussion on the use of transgenic organisms. The information then moves to the field of stem cell or somatic cell and tis- sue production, delves into technologies such as the synthesis of biologically active molecules, and introduces the growing field of combination products, where biopharmaceuticals are merged with medical devices or pharmaceuti- cals (drugs). There is a great diversity of biological products, so it is impossi- ble to mention each one or even to discuss each class. However, the examples should provide the reader with an idea of what has been achieved and, in a few instances, what could be done in the future to manufacture biotechnol- ogy products.
Expression of Recombinant Proteins and Nucleic Acids
Production of Recombinant Molecules from Expression Vectors
Laboratory methods to manipulate living organisms and the biological mol- ecules they produce are at the heart of biotechnology. Operational endeavors, including biomanufacturing, flow from discoveries made in basic research laboratories where tools or methods are devised and first applied in discov- ery research. Hence, it is no wonder that the first step in biomanufacturing is the discovery or invention of an organism or molecule that expresses a desirable trait or otherwise serves a useful function. To date, thousands of discoveries or inventions have enabled genetic engineering of nucleic acids and living organisms. Research laboratories have capitalized on a wealth of
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information regarding recombinant DNA, cell metabolism, and the basis for life itself. But there is a caveat in all this. Although it is these scientific find- ings that constitute the foundations on which we base product development, it is the biomanufacture, the production of large amounts of high-quality biotechnology product, that brings the product to the market and the user.
Today, the production of recombinant molecules, notably proteins and nucleic acids, represents, by volume, the bulk of biomanufacturing capacity. A variety of active recombinant molecules; proteins such as insulin, human or bovine growth factor, monoclonal antibodies, and vaccine antigens; and nucleic acids for genetic therapy and diagnostic purposes have entered the marketplace. Some are sold in large quantities and represent blockbuster products in the marketplace. Today, biomanufacture of recombinant proteins and nucleic acids meets the growing demand and represents an important economic sector of the biotechnology industry.
Genes, Vectors, and Host Cells
The first stage in biomanufacture of a recombinant product involves three processes: gene isolation, cloning and development of an expression vec- tor, and production of cell banks. The process and controls of this first stage are outlined in Figure 6.4. First, a gene of interest is identified and isolated through molecular cloning, most often by using polymerase chain reaction and other technologies. Alternatively, the gene may be selected from an established library of cloned DNA. The gene is characterized by molecular weight determination and DNA sequencing. A vector, available from public or private vector libraries, is selected, based on suitability for biomanufac- ture of the designated product. The attributes for selection include ability to adapt and function in a suitable host, replication, promotion of protein expression, protein chain termination, and absence of undesirable character- istics, among others. The gene is then inserted into a selected vector by using methods such as recombinase-based cloning or restriction-ligase cloning.
Next, the vector is transformed into a host cell: bacterial, yeast, insect, or mammalian cell. The host must also be carefully chosen so as to be compat- ible with the vector. Each type of cell has particular attributes, and no cell is universally well suited for expression of every recombinant DNA or pro- tein molecule. After transformation, the vector must be stable, that is, held within the host, and be maintained as one or more copies of the vector over many generations, as the host divides. Methods are applied to increase the chances of successful transformation, but many attempts may be required before a stable and fruitful match between the host and the vector is achieved. Ultimately, transformed hosts are produced and one single isolate is selected. This selection is done only after performing extensive testing to ensure that all qualities have been achieved. Once the transformed host is deemed acceptable, it is cloned by limiting dilution to ensure that all future transformed cells are derived from a single cell. The progeny of this host
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Procedure Information and testing
Step 1. Isolate and characterize gene
Step 2. Clone gene into expression vector
Step 3. Transform host cell with vector
Step 4. Clone host cell
Step 5. Select and expand cell to research seed
Transfer to development
Step 6. Produce master cell bank
Step 7. Produce working cell bank
DNA and gene Source and derivation
Homology of sequence (genome database) Molecular weight
Vector Derivation, history, and map
Sequence Selectable markers Signal sequences
Cloning site
Cloned vector Sequence
Map Alignment of gene in vector
Host cell Derivation and history Purity and morphology
Raw materials (medium, supplements, etc.)
Transformed host cell Copy number
Stability Expression gene
Purity and morphology
Cloned host cell Purity and morphology
Copy number Gene expression
Research cell bank Purity and morphology
Copy number Gene expression
Viability Sterility
Master and working cell banks Identity of host, vector, and gene insert Viability, sterility, purity, and morphology
Vector copy number Molecular markers
Annual testing of cell banks Sterility and copy number
Expression and stability
Vector
Box II
Box I
Box III
Box VI
Box V
Box IV
Box III
Box II
Box I
Box I
FIGURE 6.4 Production and testing of a recombinant molecule in an expression system and production of cell banks. This flowchart serves as an example of the steps that are taken in the early develop- ment and biomanufacture of a recombinant molecule in an expression vector. The expression system is constructed by genetic engineering and then produced as cell banks. Quality testing is performed throughout the process.
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cell are considered a research seed, and this seed is characterized for purity of cell line, retention of vector, and other traits or attributes (Chapter 7).
Selection of the host cell is worth additional mention. There are many species to choose from, and within a species, there are several strains, each of which is well characterized. Bacteria, notably Escherichia coli, and yeasts, such as Pichia pastoris, are common choices of prokaryotic host cells.
Host cell lines are purchased from a reputable source, such as the American Type Culture Collection, or a biological supply house. These vendors main- tain several strains or lines of cells, each with a full genetic history, and the buyer expects and should receive only top-quality, highly characterized cell lines. Much like pedigreed horses, cell lines and strains are noted for various attributes, such as a posttranslational capacity or large yields of recombinant protein, under specified conditions. In addition, they may also have known limitations or deficiencies, such as slow growth or stringent nutrient require- ments. Although several species and strains of host may be able to express a given recombinant protein, there are caveats; thus, great care is taken in selection of any expression system.
Bacterial Cell Expression Systems
Bacteria are often chosen as host cells because they express large quantities of a wide variety of proteins very economically. Escherichia coli has been used for decades because its genome has been sequenced, its laboratory strains are plentiful and very safe, and this bacterium is receptive to accepting, hold- ing, and expressing recombinant genes from vector plasmids. Escherichia coli is often the first choice when biomanufacturing is considered. Within the species of E. coli, there are many strains to choose from, and each strain has particular attributes and advantages as well as disadvantages. For example, some strains are best suited to secrete the desired recombinant protein into the culture media during fermentation, which can simplify downstream processing. However, production by E. coli can also have drawbacks. With certain proteins, E. coli does not secrete but instead harbor protein internally, within inclusion bodies. To obtain recombinant product from inclusion bod- ies, cells must be split open, that is, lysed, which adds extra steps, compli- cates purification, and possibly adds unwanted impurities to the product stream. Protein from inclusion bodies may not be properly folded, neces- sitating refolding steps. However, purification from inclusion bodies may be easier and more productive than purification from cytoplasm.
Another disadvantage to bacterial expression systems is the inability to make or correctly complete certain posttranslational modifications to a recombinant protein. Bacteria do not add carbohydrates to proteins, as do eukaryotic cells. Hence, if glycosylation is required for bioactivity, a bacte- rial host cell may not be the best choice. Another issue with bacteria is the production of undesirable contaminants, which are released into the process stream. Gram-negative bacteria have certain molecules such as the cell wall
213Biomanufacture
component called endotoxin. If molecules like endotoxin cannot be readily separated from the desired protein, then such organisms are not good can- didate hosts. These examples demonstrate the importance of identification of the proper host before beginning experimentation. Alone, this aspect of planning may save considerable time and resources.
Yeast Cell Expression Systems
Yeasts are eukaryotic, unicellular organisms that offer both advantages and disadvantages as host cells. Two species are commonly employed, but other species are available. Saccharomyces cerevisiae, brewer’s yeast, is well charac- terized as an expression host, as is P. pastoris, which has the advantage of secreting recombinant proteins into the culture medium. Yeast cells grow rapidly and economically in commonly defined medium, even in large ves- sels up to 10,000 L or more, which can enhance protein expression scale-up.
Yeasts are very efficient in producing some recombinant proteins, and the fermentation of yeast cells is usually inexpensive. Both yeast and bacteria can be grown in the same types of fermentation vessel, and the equipment is standard, reusable, and comparatively inexpensive. Yeast host cells are available in many strains, allowing selection based on attributes. As with bacterial host cells, and unlike mammalian cells, yeast growth medium is very well defined, so there is little concern about the introduction of host cell adventitious agents, such as human or animal retroviruses, with yeast cells. In contrast to bacterial cells, yeasts have the capability to correctly add and process many posttranslational modifications. Yeast strains commonly used in fermentation are genetically engineered to be inducible. This highly desir- able trait means that the yeast cells can begin to produce greater amounts of a recombinant protein when a simple chemical, such as glycerol, is added to the fermentation chamber or when an exact environmental condition is established in the chamber. It allows for greater control of fermentation. Hence, production in a yeast cell system provides an opportunity for pro- duction of recombinant molecules.
Mammalian or Insect Cell Expression Systems
Mammalian or insect cell expression systems are increasingly selected by sponsors for biomanufacture, especially for production of high-value human recombinant proteins, such as monoclonal antibodies. Although transforma- tion of a mammalian or insect cell line with a genetic construct can prove more difficult, as compared with bacteria and yeast cells, these cell systems have the advantages of accepting and expressing a large gene and completing most posttranslational modifications. Because large proteins with glycosylation (such as monoclonal antibodies) are common to the world of biopharmaceuti- cals, mammalian cells are frequently chosen as an expression and production system. However, as compared with yeast or bacterial cells, mammalian or
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insect cells are often less robust and more fragile, grow more slowly and may be more fastidious, thus requiring more stringent environmental controls. They may require a continuous flow of complex medium to deliver nutrients and may demand continuous waste removal. Concerns regarding the pres- ence of latent virus in mammalian cell lines, specifically cells from a new and poorly characterized cell clone that might harbor and then shed viral parti- cles into the product stream, have slowed the introduction of new cell lines. However, advances in mammalian and insect culture techniques and extraor- dinary characterization efforts have overcome some of these difficulties, and today, there are several effective cell bioprocessing systems in the market.
Several factors enter into the choice of a mammalian cell line intended for protein expression. The ease of transfection with a particular gene or transfec- tion technology, cell growth and protein secretion profile, and environmental requirements, all enter into the decision. Hence, the key to selecting the cor- rect cell line for the expression of a given gene is experimentation with several highly regarded lines, which allows comparison with a particular construct. Cells are named by their derivation. Cells of the epithelial origin are the most used for biomanufacture. The mammalian Chinese hamster ovary (CHO) cell, a cell line in use for more than 50 years, is high on the list of choices. This cell line was widely used first in virology and cancer research laborato- ries and later in biomanufacture. The CHO cells are very well characterized and certified free of adventitious agents (with the exception of endogenous retrovirus-like particles). Products derived from CHO cell production have been used for decades without safety problems. Other cell lines chosen for biomanufacturing are African green monkey kidney (Vero), Madin–Darby Canine Kidney (MDCK), human embryonic kidney (HEK-293), baby hamster kidney (BHK), human retinoblast (Per C6), and murine myeloma (NSO).
The process of establishing an expression vector, referred to as transfec- tion, is outlined in Figure 6.5. The expression gene is isolated and cloned into an expression vector by using methods described earlier in this chapter and technically in the manner described for yeast and bacterial hosts. The method of delivering that gene to mammalian cells, transfection, differs noticeably from the methods applied to bacteria or yeast cell. One of the several transfec- tion methods, most commercially available, may be used to transfect a gene to a mammalian cell. Notably, the method must deliver the intended gene directly into the nucleus and integrate it into the chromosome of the target cell. Most transfection methods rely on chance, that is, the probability that a gene will enter into the nucleus, integrate into the genome of a mammalian cell, and result in a stable and productive transformed cell. In practice, this means treating a large number of cells and using cloning and selection meth- ods to determine which cells are stably transformed. Additional experimen- tation is performed to characterize cell lines, and the best cell line is chosen for expansion to become the transformed cell research seed (Figure 6.5).
Transfected insect cells are also used to produce recombinant proteins, often times large quantities of proteins that could not be well expressed in
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Choose transfection method
Mechanical: Gene gun, electroporation, and optical laser
Chemical: Calcium phosphate, liposomes, and cationic polymers
Infection: Adenovirus and lentivirus Others: Nucleofection and impalefection
Construct expression gene (selection and
marker)
Transfection
Stable transfection With matched cell line+transfection
method+genetic construct
Genetic construct
Select stable transformed
clones
Expand clonal cells
Master cell bank
Working cell bank
Evaluation
Establish gene Establish cell line:
Mammalian or insect
Mammalian or insect cell line
Expand cells
Established cell line
Test: Transient transfection Evaluate transfection
Efficiency Test expression level
Test: Stable
transfection
Test: Stable cell
line
Test: Purity and
viability
FIGURE 6.5 Process of gene transfection for mammalian cellular expression. An established cell line is selected and tested in the left panel, and a gene is engineered in the right panel. By using a carefully chosen transfection method, the cells are transfected with the gene, and after evalu- ation, these cells are used to produce the expression product.
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other systems. The process of gene transfer to insect cells is quite differ- ent from that applied to mammalian cells. The gene of interest is inserted into the genome of baculovirus, a virus that normally infects insects, which then acts as the delivery vector. The cell lines used as targets are derived from insects and are thus free of potentially harmful human viruses but are capable of hosting baculovirus. Insect cells are also desirable because, like the mammalian cells, insect cells also perform complex yet accurate posttranslational modifications of the expressed proteins. On infecting an immortalized insect cell of a well-characterized cell line, genetic information is transferred from the virus to the cell nucleus; some insect cells are stably transfected. The transformed cells are then identified, selected, character- ized, cloned, and expanded to produce a research cell seed. As the trans- fected insect cell grows and multiplies, it expresses the recombinant gene of interest and the gene product, a recombinant protein, is produced, which can then be harvested.
Production of Master Cell Banks and Working Cell Banks
Research seed, described earlier, is transferred from the research laboratory, where it is produced, to a development laboratory, where it undergoes addi- tional examination and characterization. The complete history of the con- struct, to include descriptions and sources of all raw materials and detailed summaries of the procedures used to derive the seed, is archived as the research seed history. Once development scientists are satisfied that the seed is adequate for production of the intended product, master and working (or production) cell banks (WCB) are manufactured.
Cell banks provide a uniform stable stock of cells with genetic inserts. These stocks are available to production for future use. Cell banks include the master cell bank (MCB), which is derived from the research seed, and the WCB, which is derived from the MCB. To produce an MCB for bacterial or yeast cells, a single clone of research seed is expanded in the culture and then transferred for further growth in a shake flask or small fermenter, as outlined in Figure 6.4. A limiting dilution step may be employed before transfer to ensure that a single cell is indeed the ancestor of the MCB. The cells are har- vested and counted, and specified numbers are aliquoted into several hun- dred vials; these are labeled, cryopreserved, and placed into secure storage, usually divided between two or more storage sites. The vials of an MCB are the ultimate source from which the product is derived for decades to come. Since an MCB is limited in the number of vials produced while demand for MCB could be great, a WCB is produced from a vial of MCB. Procedures used to produce and control WCB are very similar to those used to produce and control MCB. Biomanufacturing uses stock from WCB, until it is exhausted, and then another WBC is produced from a vial of MCB.
To ensure identity, purity, and safety of MCB and WCB, samples taken immediately after production from both MCB and WCB are extensively
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tested, as outlined in Figure 6.4 and described in Chapter 7. Tests of MCB and WCB samples are repeated at specified intervals (e.g., annually). This strin- gent test regimen once again emphasizes the need to develop analytical tools early in development, even before biopharmaceutical production begins. At this stage, a significant resources have gone into the development of the con- structs, MCB and WCB, as such they represent a precious commodity and should be treated as such. Secure storage of all cell banks is critical. The pro- cesses of producing research seed, MCB and WCB, are often both rewarding and instructive to a new biotechnology operation and often represent their first introduction to biomanufacture under cGMP. There is great satisfaction in having completed the first stage of biomanufacturing by having produced the foundation for later production efforts.
Biomanufacture of Recombinant Proteins
Planning Production of a Recombinant Protein
In a product-specific manufacturing plan, most processes have at least three stages, and each stage is further divided into several technical steps, as out- lined in Figure 6.1. We previously covered the steps of Stage 1. Now, we will consider Stage 2, upstream processing, which involves the production of recombinant protein in the expression system, and Stage 3, downstream processing, which involves the purification of recombinant protein product as BS. A good biomanufacturing plan goes beyond the initial process outline and also considers facility, utilities, equipment, raw materials, quality con- trol testing, staff requirements and compliance, or cGMP for both upstream and downstream processing.
The initial attempt at biomanufacturing using a new process or for a new product is referred to as pilot production. Pilot production involves per- forming defined, sequential runs in an attempt to develop the process and to eventually get it right; that is, to make a safe, pure, and potent product. Indeed, pilot production is much like research experimentation because it involves trial and error, tweaking various systems, and even making sig- nificant changes in process protocols and procedures. Pilot production may precede Phase 1 production, described above, or it may overlap or be syn- onymous with Phase 1 production. The term run is used in biomanufacturing to describe the performance of one defined process, such as all steps in the upstream fermentation stage or a full set of process steps and fermentation followed by purification, from beginning to end. It also demands repeat- ability to confirm that the system is performing properly and consistently. It is not unusual for a biomanufacturing operation to attempt a new pro- cess in 5 or even 10 runs before it is considered reproducible and robust. Hence, no matter what the biotechnology product is, pilot production can be a long, arduous, and expensive endeavor, stretching over several phases of development.
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Upstream Process: Production by Bacterial or Yeast Cell Fermentation
Fermen tation is an ancient process, best exemplified by brewing of beer in the presence of yeast. It is a skill that has developed over the ages. Substrates for fermentation of biomolecules remain simple and include well-characterized materials such as water, salts, and sugars. In some instances of biomanufac- ture, more complex nutrients, such as soy extracts or vitamins, may be added to the fermentation vessel. Animal materials such as liver powder or serum supplement are used in the fermentation of a biopharmaceutical product only in special circumstances where they are essential to the success of a process. Such materials can harbor adventitious agents that can contaminate FP, and thus, their use is discouraged.
Fermentation to produce biotechnology products is performed in a fer- menter, a closed and sealed glass or stainless steel vessel with a series of por- tals, stirring devices, and tubes entering the chamber (Figure 6.6). To begin the fermentation process, one must have raw materials of the highest quality, including a growth medium, gasses, a seed of recombinant bacterial or fungal
FIGURE 6.6 Equipment for microbial fermentation. This picture shows fermentation equipment in a bioman- ufacturing suite. The operator in the center is programming the microprocessor controller in the square unit. On either side of the controller stand two fermentation vessels, a small one in the background on the bench top and a medium one behind the controller. In the foreground is a large, cylindrical storage vessel made of stainless steel. (Courtesy of Waisman Biomanufacturing, University of Wisconsin, Madison, Wisconsin. www.gmpbiomanufacturing.org.)
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organisms, and a means to control the process. Seed material, formed of bil- lions of organisms that are capable of active division, is derived from a vial of WCB that has been expanded in a flask containing the defined medium. This is called the inoculum. The environment inside the vessel is controlled by human intervention or, when on auto-pilot, by a microprocessor. The fermen- tation process is initiated once all ingredients and the seed have been added together in the vessel and the fermenter has been closed and sealed. Once the environment inside the chamber is optimal, the cells replicate and are active manufacturers of the recombinant protein product. The chamber is typically stirred or otherwise agitated in an effort to evenly distribute gasses (particularly oxygen), nutrients, organisms, and, if secreted, the recombinant protein product. As stirring and movement of gasses may cause foaming, addition of antifoaming agent, a chemical designed to reduce microbubble formation, is often helpful. High shear stress and foaming contribute to pro- tein denaturation and as such is avoided whenever possible. Cell growth and product production are monitored by taking samples from the chamber, and critical measurements, such as pH, gas tension, and osmolality are measured by probes placed directly in the chamber. This in-process testing allows the operator to follow the progress and correct variables if deviation from the specified limits is required. For example, if pH drops out of range, sodium hydroxide may be added to raise the pH, thus getting the fermentation sys- tem back into an optimal pH range.
Division and growth are separated into several phases, as shown in Figure 6.7. First is the brief lag phase, during which organisms adjust to the culture environment. Next is the exponential growth phase, during which organisms divide rapidly and hopefully produce large amounts of the intended recombinant protein product. The deceleration phase represents a slowing in growth, and the fourth phase is stationary, with little growth or even increasing amounts of death. The final phase, decline, represents reduction in metabolism and is indicative of large amounts of organism death. Once measurements determine that the organisms have grown to the required optical density, or that the death is extensive, or that sufficient product has been produced, the fermentation is halted by radically chang- ing the pH, by rapidly cooling the chamber, or by some other interven- tion that is conducive for being gentle on the recombinant protein product and facilitating high yields. It is important to identify the optimal growth and harvest phases of the formation process. Termination of the growth phase is important in an effort to maximize the production of the recom- binant protein while reducing the number of unhealthy or dying organ- isms. Dying organisms release destructive enzymes (e.g., proteases) and debris or impurities into the medium; continuing a controlled fermentation beyond that point can interfere with product purity or complicate down- stream purification. The result of a successful microbial fermentation is a vessel filled with slurry of organisms, debris, expended medium, and large amounts of the intended recombinant protein product.
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Certain cells, notably yeast used in some fermentation systems, may be cued or induced to begin the production of recombinant protein. To engineer an induction system, a gene or genes are inserted into the vector construct for the purpose of controlling protein production by the expression system. These inducible genes are active in the presence of certain environmental cues or products. An example is selective induction of recombinant protein production by P. pastoris on addition of glycerol to the fermentation vessel.
Upstream Process: Production by Mammalian or Insect Cell Culture
Mammalian or insect cells are cultured in a sealed chamber referred to as a bioreactor. Although mammalian or insect cell culture has superficial resem- blance to fermentation, the process is, overall, quite distinct. The objective in both systems is to produce a recombinant protein product that is either stored within the cells or secreted into the medium. Both bioreactors and fer- menters are closed and sealed systems with a high level of monitoring and environmental control. Mammalian or insect cells typically demand more complex substrates than bacteria and yeast. Hence, cell culture media used in a bioreactor contains a complex mixture of nutrients and vitamins. When animal products are used for biopharmaceutical production, in fermentation or cell culture, they must be carefully tested and controlled, so as to ensure
Stationary
Lag
Growth (exponential)
Decline or death
Deceleration
Time
C el
l n um
be r/
m L
FIGURE 6.7 Phases of microbial growth in fermentation.
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that potential microbes and other impurities do not contaminate the cells or the product. Typically, mammalian or insect cells must be grown as adherent cells. This is because, unlike bacteria or yeast, these cells in nature exist in a tissue or an organ, where cells are interconnected and held firmly to a base- ment membrane or other connective tissue protein. Alternatively, hollow fibers, convoluted vessel surfaces, and microcarrier beads, sometimes coated with collagen or other connective tissue matrices, may be used to create opti- mal microenvironments, facilitating cell anchoring or adaption to culture conditions, yet confining cells to a small area in the absence of mechanical stresses. Mammalian cells engineered to grow without anchorage are grown in suspension, but agitation or stirring is exceptionally gentle, because insect and mammalian cell membranes are fragile. Gentle air movement or wave action is used in some systems to maintain the requisite movement of the cell medium for suspension cultures. Mammalian cells are particularly suscep- tible to reduced growth because of low oxygen tension, high carbon dioxide tension, buildup of waste, changes in pH, and other metabolic-environmen- tal influences. Cell bioreactors are closely monitored for temperature and the addition of gases, buffers, and nutrients is highly controlled, both by the operator and by microprocessors. The vessel environment of a cell bioreac- tor is monitored using specialized probes and microprocessors by operators; adjustments in gasses, buffers, and nutrients are efficiently achieved using aseptic ports in the vessel.
Mammalian and insect cells demonstrate growth curves, which represent cell growth as a logarithmic or semilogarithmic phase, followed by a pla- teau phase, and finally a decline phase. Although the growth and protein secretion of mammalian cells are typically slower than that seen in bacte- rial or fungal cell fermentation, under well-controlled operating conditions, the growth and protein secretion of mammalian cells may be sustained for much longer periods. A cell culture is terminated at an exact point in the growth and protein production cycle, so as to maximize protein production and minimize contaminants. The result of successful cell culture is a bio- reactor vessel filled with a slurry composed of cells, cell debris, expended medium, and the intended recombinant protein product.
Upstream Process: Recovery
Immediately on stopping cell growth in a fermenter or bioreactor, the mate- rial is harvested, chilled, and the cells and other large solids separated from the liquid phase. This is done by moving, with pumps, the contents of the chamber into a capture vessel. In the case of cells anchored to the substrate, or when product is contained within the cells, it may be necessary to dis- lodge the cells by mechanical or enzymatic means. If most of the product is harbored in the cells, as would be the case with protein that is not secreted, the cell paste is retained and the supernatant is discarded. Whole cells con- taining product are then lysed using a single or a combination of methods,
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which may include mechanical methods (e.g., sonication and homogeniza- tion), chemical methods (e.g., lysozyme and detergent), and/or shock (e.g., freeze-thaw cycles and water). Soluble proteins are then harvested, again traditionally, by using a combination of centrifugation and filtration meth- ods. Primary clarification, to remove any remaining cells, cell debris, and other large solids, is performed by centrifugation or tangential flow filtration (Figure 6.8). The resultant filtrate or supernatant containing the recombinant protein is then kept in a storage tank under controlled conditions, until it is purified. The storage step is referred to as a hold.
Downstream Process: Purification
No matter what the source—fermentation, cell bioreactor, transgenic ani- mals, or plants—recombinant proteins and other biological molecules must be purified from typically a complex milieu of cellular debris, impurities, and contaminants. By way of definition, impurities are undesirable materi- als, both particulate solids and soluble molecules, that remain with product after production. Common impurities derived from biological processing are host cell proteins or DNA, endotoxin, or other microbial toxins; cellular debris and organelles; and materials from the culture media. Contaminants are the substances that enter the product stream, often during purification, and are frequently shed or leached from the process materials or equipment. Small particles from tubing, glass, or metal containers, chromatography gels, and heavy metal ions, leached from metal containers, are the examples of contam- inants. Impurities or contaminants may be debris, suspended particulate, or soluble in nature. Both of them are considered necessary evils, because their presence reflects the contents and environment of the culture that yielded the product. However, levels of impurities and contaminants are greatly reduced
Pump Membrane
cassette
PP
Pf
Pr
Permeate
Retentate (product)
Feed
Permeate tank
Batch feed
FIGURE 6.8 Purification scheme for a recombinant molecule. This is a classic purification scheme, or down- stream process, for a recombinant protein and includes precipitation, centrifugation, filtration, and multiple chromatography steps, yielding bulk substance.
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during purification, and, in the end, biotechnology products are tested for common impurities and contaminants (Chapter 7) to ensure that the levels meet predefined specifications and the product is deemed to be safe.
Purification steps are designed to remove one or more impurities or con- taminants, at least to the greatest extent possible, and yet retain the desired recombinant protein, or other biologically active molecule, thus maximizing the yield of product. Yield is especially important because a low yield of a very pure product is as unacceptable as a high yield of the product with sig- nificant levels of impurities. Hence, in-process testing (Chapter 7) is applied throughout purification to ensure improvements in purity and maintenance of yield at every step. The field of biomolecular purification has progressed rapidly in recent decades and dozens of methods, some simple and others quite complex, have been developed and have entered the market. We will mention a few of the most commonly used methods.
To plan downstream processing, a purification scheme is produced (Figure 6.9). To do this, it is first necessary to understand the biophysical and biochemical properties of the recombinant molecule, because purifica- tion methods take the advantage of those properties. This understanding is based on experimental data, derived from the research laboratory, about the product and the nature of that product as it enters purification. Knowledge of the possible contaminants and impurities and their properties is also needed. For example, it is critical to know the isoelectric point of the desired molecule under given conditions, the pH or the salt concentration at which the molecule precipitates from solution, the size and shape of the molecule, the glycosylation profile, or any propensity to bind to other molecules or inert substrates. Purification schemes, as shown in Figure 6.9, take advan- tages of these characteristics or attributes.
Many technical methods, or purification tools, are available for down- stream processing. Choice and application of a method are based on the nature of the molecule and the knowledge that certain tools have been suc- cessfully used in the past to purify similar molecules. Some purification tools are quite simple and inexpensive, whereas others require significant investment. The sequence in which methods are applied is as important as the choice of the tools themselves, and the downstream plan must design their use very carefully. It is often necessary to test each purification method alone and the full sequence of methods, at small scale in the laboratory, in an effort to derive the optimal sequence of events, before beginning opera- tional purification. In-process tests are another critical component of a puri- fication plan. These are developed to ensure that materials such as solutions are of the correct composition, pH, or strength and can be effectively used to measure the levels of the desired molecule and impurities throughout the purification process. These assays, which can require expensive instrumen- tation and extensive development efforts, must be available from the outset of purification process development, because they are essential to the under- standing of the purification outcomes.
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Cell supernatant or cell paste
Precipitation
Centrifugation (clarification)
Tangential flow filtration Depth filtration
Size exclusion chromatography
Ion exchange chromatography
Affinity chromatography
Ultrafiltration
Bulk substance
Process
SDS-page Total protein
SDS-page
SDS-page activity
Total protein HPLC
SDS-page activity
Total protein HPLC
SDS-page activity
Total protein HPLC
SDS-page activity
Total protein HPLC
BS panel of tests
Control testing
FIGURE 6.9 Tangential flow filtration. Material enters the filtration scheme from the batch feed, usually a storage vessel (holding tank), and it is pumped under pressure (feed) across a membrane cassette, where some material of the correct molecular weight passes through the membrane cassette as permeate and is then held in a tank. Material that does not pass through the cas- sette re-enters the batch feed tank and is again pumped across, and in some cases through, the membrane cassette. Continuous flow across the membrane cassette deters clogging the cassette selective filter.
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Early in downstream processing, and often at other stages in biomanu- facture, the liquid fraction must be clarified, without significant loss of the desired protein, using combinations of precipitation, centrifugation, and fil- tration. Precipitation is a simple and inexpensive application; however, many methods for precipitation, such as changing the pH or adding simple salts, may either degrade the protein of interest or add contaminants to the prod- uct. Precipitation is based on the knowledge that a desirable protein or an undesirable impurity becomes insoluble under certain conditions such as low pH or high salt concentrations. Once a precipitate is formed, it is sepa- rated from the undesirable materials by centrifugation or filtration. Desirable molecules in the precipitate are recovered by diluting the precipitate with physiologic buffer. Undesirable proteins in the precipitate can then be dis- carded. An example of precipitation is purification of an immunoglobulin on addition of buffer with a high salt concentration. Immunoglobulin pre- cipitates in this environment, leaving impurities in the supernatant. This is centrifuged, and the pellet is recovered and diluted with normal saline to again solubilize the immunoglobulin protein. Another example is the appli- cation of a polycationic agent, such as polyethyleneimine, which precipitates undesirable nucleic acids; however, the desired recombinant protein remains in the solution. After centrifugation, the pellet with impurities is discarded and the supernatant is retained or vice versa, depending on which fraction holds the desired product.
Centrifugation, a relatively simple and often effective method, is employed whenever possible and often constitutes the first step in a purification scheme. It separates materials based on density, shape, and other physical properties that impact their gravitational movement in a fluid. It can be used without other treatments, such as in effective separation of large impurities (e.g., cell walls and nuclei), from smaller particles and soluble proteins. Centrifugation is also used in a step-wise manner to sequentially remove matter of different density. It is often applied in conjunction with other applications such as pre- cipitation. Centrifugation equipment is available in many designs and range from small instruments to large, continuous-flow machines that can process large volumes of product.
In addition, flow filtration or tangential flow filtration are the methods used in purification schemes. Both methods remove debris and clarify a solution in which the recombinant protein is suspended. Flow filtration involves pass- ing the material through a selective membrane filter, a synthetic sheet that has holes of a specific size. As solution is pushed against the filter, solids of that size or less move through the membrane and larger particles are trapped atop the membrane. However, a disadvantage of flow filtration is the buildup of material on the membrane surface, which can clog and foul the filter. Some filtration protocols, therefore, use a series of flow filtration filters. The fluid stream is first fed through filters with larger membrane holes. Then, in a series, it is fed through filters with smaller holes, thus distributing particles over many filters and avoiding fouling and clogging of a single filter.
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Tangential flow filtration (TFF) is a more expensive, but often more effec- tive, method and is also a choice for processing larger volumes. In TFF, solute passes over the filter in a horizontal stream, even as filtration is happening in a vertical plane (Figure 6.10). This horizontal movement constantly sweeps debris off the filter surface and prevents clogging. As it can be performed rap- idly, TFF is often used to exchange solutions, such as one buffer for another and to selectively remove low-molecular-weight impurities, all in a single step.
Although extremely useful, filtration must be applied judiciously, because under some circumstances it may also destroy molecular integrity. Filtration causes shearing forces as the fluids move under pressure across or against a membrane and shear can destroy cells or molecules. (Each product has a unique tolerance for shear.) The only way to fully realize the effects of shear on a given molecule is through experimentation followed by characteriza- tion of the desired molecule. Movement of fluids rich in proteins may create foaming, an indicator of protein denaturation, which is an undesirable out- come of any biomanufacturing process and must be avoided or countered. A third cautionary note is avoidance of adsorption of the desired molecule to surfaces of equipment, transfer tubes filtration membranes, and even to
Fraction collector
Filter/bubble trap
Pump A
Pump B
Load
Buffer A
Buffer B
Regen
Detector
Chromatography column
Controller
FIGURE 6.10 Flow diagram for preparative chromatography. This scheme depicts the equipment and flow for a chromatography system used as one step in the purification of a recombinant protein. The product (load) and Buffer A are pumped into the column via pump A, where the gel matrix of the column binds or otherwise slows the progress of the molecule of interest (e.g., through affinity binding, size exclusion, and ionic interaction). Other molecules pass through the column and are detected and collected into fractions. Once this has been completed, Buffer B is pumped onto the column, with the intention of releasing the bound molecule. Thus, the desired product is detected as it comes off the column, and it flows into later fractions, where it is collected.
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impurities. Again, any purification step must provide consistent and useful yield, and adsorption can greatly reduce the amount of desirable cells or proteins left in the product stream.
Chromatographic methods are used in most biomanufacturing purifica- tion schemes that involve molecular purification. Chromatography is based on various properties of proteins and other macromolecules: charge, size, shape, or affinity to a substrate. Preparative chromatography is used to purify significant amounts of materials, whereas analytical chromatography, described in Chapter 7, is used to characterize macromolecules. Preparative chromatography, the subject of this discussion, is performed using aque- ous suspension of resins or gels packed into a vertical column (Figure 6.10). Preparative chromatography columns come in various sizes and shapes (Figure 6.11) to suite the intended purpose, with some exceeding the vol- ume of household refrigerators. They are controlled with pumps, valves, and microprocessors. Each column-and-resin chromatography system has a unique set of properties that allow for the differential separation of mol- ecules, based on the physical or chemical properties of the molecules to be
FIGURE 6.11 Equipment for preparative chromatography. Two preparative chromatography columns rest on tripod supports. On the table top are controller units and pumps with tubing that lead to the glass columns. This is performed within a chromatography suite of the biomanu- facturing facility. (Courtesy of Waisman Clinical Biomanufacturing facility, http://www. gmpbiomanufacturing.org.)
228 Biotechnology Operations
separated. In the simplest chromatography protocol, clarified supernatant containing both product of interest and impurities is placed at the top of the column. Then, using gravity or a pumped stream of buffer, the super- natant is passed through the column. As this fluid passes over the column, the molecules contact the resin and may or may not bind to the column. As it passes out of the column, it is collected in a series of tubes, each fraction rep- resenting a specific volume and time of collection. By eluting with various buffers, a gradient is established on the column and desirable proteins leave the column in one fraction, whereas undesirable proteins exit the column in another fraction. This process is shown by simplified format in Figure 6.10.
Chromatography uses distinct molecular properties to separate one mol- ecule from another. For example, ion-exchange chromatography uses the charge properties of various molecules to separate desirable from undesir- able proteins. Here, the resin has a known electrical charge at a given pH and ionic strength (salt concentration). Elution buffers added to the chromatog- raphy column may be changed by the operator over time. Each buffer has a given salt concentration and pH. Taking advantage of the ionic properties of both the gel matrix and the desired protein, the buffer strength and the pH of the elution buffer are adjusted to ensure that the target protein binds, by ionic interaction, to the ion-exchange resin. For example, at pH 7.0 and ionic strength of 100 mM (Figure 6.10, Buffer and Pump A), a recombinant pro- tein might bind to the resin; however, impurities pass through the column, to be collected in the early factions. Next, a second buffer and Pump B in Figure 6.10, of pH 6.9 and ionic strength of 150 mM, is added to the column to release the recombinant protein, and so on. Eluate is collected in later fractions, and some of these contain product, largely free of impurities.
Many other types of chromatography are available to the biomanufactur- ing operator. Hydrophobic interaction chromatography takes advantage of a molecule’s affinity for or, alternatively, rejection of water. To purify hydro- phobic proteins, a gradient with high-to-low gradient of salt concentrations are established in a column containing hydrophobic interaction resins. Size exclusion chromatography takes advantage of the size and/or the shape of a molecule. It is particularly useful to purify proteins of interest if they are particularly large or small or have an unusual shape. Affinity chromatog- raphy methods employ ligands, attached to chromatography resins, to cap- ture the desired protein as it passes through the column in the presence of physiological buffer. For example, Protein A resins are commonly used to retain monoclonal antibodies to a column. The protein A molecule, derived from bacteria, naturally sticks to antibody molecules. When Protein A is immobilized on a resin and placed into a column, any antibody passing over resin will, in physiological buffer, adhere to the Protein A while impurities pass through the column. In the second step, a buffer solution, known to force Protein A to release the antibody by molecular or ionic competition, is passed over the resin. Now, monoclonal antibody, without impurities, elutes into the subsequent fractions.
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Another purification method is the use of tags in affinity chromatography, with polyhistidine tag being quite popular. Here, the protein of interest must, in research or early development, be genetically engineered to have at the C- or N-terminus a series of nucleotides that repeatedly code for histidine. Polyhistidine tag chromatography is an affinity method in which the chroma- tography resin immobilizes nickel ions. As the protein harboring the polyhi- stidine tag passes over the column, it binds to nickel, while other impurities, without the tag, pass through the column. The bound his-tagged protein is then conveniently rinsed with several volumes of buffer. Subsequently, a selective elution buffer is passed through the column, triggering the release of protein from nickel. More sophisticated tags may include a cleavage sequence to facilitate the release of the tag from a therapeutic protein. Although provid- ing an efficient purification method, the use of tags is discouraged by regu- latory agencies, as they introduce an added impurity with the capability of influencing the protein structure and function or elicit an immune response if used in the production of a therapeutic protein.
The pharmaceutical industry commonly employs the use of two indepen- dent chromatographic methodologies in tandem (e.g., size exclusion followed by ion exchange) as a common purification technique, which result in a rela- tively pure protein product with minimal impurities. More chromatographic methods are available and still others have been developed to purify spe- cific biomolecules. For some molecules and even for living cells, in situations where readily available purification methods are not useful, a new and very product-specific chromatographic method such as affinity chromatography is often developed out of necessity.
Purification is a lengthy process, as the tools are applied over days or even weeks. Pauses, referred to as holds, in a series of events are commonly incor- porated into process schemes to allow for in-process testing and operating staff breaks in schedule. However, pauses require product storage, and stor- age can result in product degradation. Therefore, pauses must be carefully planned, controlled, and monitored. During a hold, the product may be susceptible to degradation as a result of impurities such as proteases in the material, to other influences of the hold environment, such as oxygen or pH, or even to the container surface, which acts as a catalytic agent. In general, greater lengths of storage time and higher storage temperatures accelerate product degradation. To prevent degradation, it is important to understand the contributing factors along with remedies to allow for planned and con- trolled holds, in order to prevent degradation of the product. For example, protease degradation is reduced by storage in various buffers or addition of protease inhibitors, substances that are inherently safe and can later be sepa- rated from the product. Oxygen tensions can be adjusted, antioxidants can be added, or containers may be lined with inert materials to prevent product breakdown. Planning each process and hold step is based on the knowledge of the product, possible impurities and contaminants, and the product’s sta- bility profile.
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The end result of purification efforts is BS, a pure, potent, and stable prod- uct within the proper bulk container. For a biopharmaceutical, this is referred to as bulk (drug) substance.
In summary, purification processes are planned on the basis of the prop- erties of the product and possible impurities or contaminants. Success at purification frequently involves technical planning based on the product’s characteristics and then experimentation and trial and error in the research laboratory. Purification is first attempted at small or model scale, to better understand the attributes of each application. The biomanufacturing opera- tor does not expect to get it right the first time. Indeed, it may be shown that a purification tool or a series of methods, intuitively good choices, negatively impact the molecule of interest; or it may be discovered that product yields are unacceptably low. If the negative impact is irreversible and the protein cannot be recovered to the native form, then the operator might drop that step and try another. Alternatively, the method can be modified. A third possibility is the application of a recovery step that intends to return the mol- ecule to its native or desired state. In reality, it may be necessary to apply sev- eral tools and determine, experimentally and by trial and error, the impact of each tool, before the correct process or formula is discovered. Again, success in this endeavor is based on the knowledge of the protein and on the fact that various tools are available to the operator.
In-Process Testing and Analysis of Bulk Substance
In-process testing is a hallmark of product purification. The operators need to know, at each step, whether their purification scheme is achieving the intended objectives of removing impurities and contaminants while enrich- ing the desired product, without significant product loss. Hence, quality con- trol (in-process testing) is applied to samples taken at the completion of each step. More information is provided in Chapter 7 about the individual ana- lytical tools commonly applied for in-process testing of biopharmaceuticals. Examples are measurements of product, particles, contaminants, or impuri- ties. At each step, the operator is interested to learn whether the product remains in the stream and, if so, to identify its molecular integrity. Relatively rapid methods, for example, examination and measurement of protein bands after sodium dodecyl sulfate polyacrylamide gel electrophoresis of a sample quickly provide information, both qualitative and quantitative, about the yield of both the intended protein and the contaminants and impurities at each step in the process. These tests must be readily available to a biomanu- facturing operator.
Both quantity and quality of a BS matter greatly to the manufacturer. Quality control test results of BS must demonstrate that product, at this stage, possesses all intended attributes. In Chapter 7, we will discuss the tests used to measure those attributes. Each test is classified under the attribute it mea- sures: identity, safety, purity, and potency. Purity is of particular importance,
231Biomanufacture
because it is a key objective of downstream processing. However, in a general sense, how do we define purity of a molecule such as a recombinant protein? One guideline often applied to biopharmaceuticals is that more than 95% of the BS is the intended and intact ingredient and less than 5% of the BS are the known or unknown impurities and contaminants. Most biomanufacturing operations strive for more than 98% or greater purity, certainly for commercial manufacture. However, there are caveats to this purity guideline. First, the balance of material in BS, the remaining 2% or 5% if you will, must be known, indeed be characterized, for it cannot be toxic or allergenic or potentially toxic to the user and it must consist of various materials without a predominant molecular entity: impurity or contaminant. Second, it is not always possible to reach the 95% purity level, and in such instances, it may be acceptable to identify impurities and show that they cannot be harmful to the product or the user.
Knowing what could be or what should not be in BS is helpful in making these determinations. For example, a recombinant protein product derived from bacterial host cells might be expected to have very small or trace amounts of bacterial chromosomal or plasmid DNA, and it would be tested for such impurities and measured against a specification for host cell DNA. In addition, in-process testing focuses on the materials that are there and those that could be there but not on those that are highly unlikely to be there. For example, the bacterial product would not be expected to have mamma- lian or yeast cell DNA, and so, the operator would not test for DNA from eukaryotes. In-process testing will, in part, help the operator to understand the makeup of material and to pinpoint the step at which it entered, or was not fully eliminated from, the product stream. Understanding the potency of product is another critical step in characterizing BS, and at this stage of manufacture, there must be either an indirect or a direct measure of potency.
Recombinant proteins are not the only biotechnology products produced by biomanufacturing technologies. The following paragraphs provide by way of example an overview of possible manufacturing approaches for a few of the many other biotechnology products.
Production of Bacterial Plasmid DNA
Bacterial plasmid DNA, used in DNA vaccines, genetic (DNA) therapeutics, diagnostic tests, or as research laboratory chemical, is produced by bacterial fermentation. Once purified, biomanufacturing processes may yield up to 1 g of plasmid DNA per 10 L fermentation. RNA can also be produced, albeit in milligram quantities, by in vivo transcription in E. coli. To produce DNA, a plasmid vector is constructed in the research laboratory and tested for the intended biological effect. An appropriate cell line of E. coli is transfected with the DNA plasmid and cell banks are produced from research seed. Beginning with WCB, cells are grown in a fermentation vessel and then har- vested and lysed to release supercoiled plasmid DNA, the intended product
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for most purposes. The plasmid DNA is then purified using physical separa- tion and chromatographic methods. Impurities, such as chromosomal DNA, RNA, and bacterial host cell proteins must be considered during purifica- tion. During all processing, the plasmid product must remain supercoiled, that is, in the closed circular form that coils about itself. Quality control test- ing focuses on identity, purity, and identification of minor impurities and potency in a relevant biological assay for all DNA products. The form of the plasmid (linear, circular, relaxed, or supercoiled) is also determined by analytical testing.
Production of Live Recombinant Organisms: Bacteria and Virus
Live virus, bacteria, and even protozoa are used as biopharmaceuticals or as diagnostic or laboratory reagents. Virus, for example, retrovirus, may be used in gene therapies to transfer therapeutic genes to patients. Owing to their efficient ability to infect cells, live viruses such as vaccinia, adenovirus, and alphavirus are often employed as delivery vehicles for vaccines. Live bacteria are used as investigational biopharmaceuticals, both therapeutically and as vaccines. Even protozoa, such as attenuated malarial sporozoites, serve as live vaccines.
There is considerable experience in bacterial and viral culture and puri- fication, largely for vaccine production, yet there are potential issues with some viral or bacterial constructs. One concern is the possible release of live recombinant virus or bacteria into the environment or spread to close contacts. Another concern is whether these microbes retain the capacity to infect or cause diseases in humans, plants, or animals. To address these issues, live organisms are carefully designed and endowed with redundant systems engineered into their genome. Some systems ensure they cannot survive outside the environment provided by cell culture or a living host organism. Other systems delete the genes responsible for replication, infec- tion, or pathogenicity, rendering them incapable of causing disease. Further, safety issues related to exposure and release are considered in manufac- turing plans and process controls. Extensive safety testing is performed to measure the attributes of organism in research seeds, and stringent specifi- cations are applied.
These recombinant organisms are constructed in the research laboratory by using well-characterized, attenuated strains of virus or bacteria. Live bacteria or viruses (Figure 6.12) are grown by fermentation and cell culture, respec- tively. Since live virus is propagated in cultured cell lines, it is necessary to identify a virus host cell, often of human origin, which yields large quanti- ties of viral particles and is free of adventitious agents or other undesirable traits. This requires significant resources, because the number of satisfactory choices is limited, and the processes of adapting virus to the host, cell bank- ing (both MCB and WCB), and testing for identity and purity are lengthy and expensive. Cells are first grown to optimal cell density in a bioreactor
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Viral activity Number particles Surface protein
Certificate of analysis for BDS
Certificate of analysis for BS
Cell substrate (Diploid cell and
culture collection)
Viral product Virus research seed
Cell control Characterization
Viability Identity Purity
Virus clonal selection and expansion
Viral MCB and WCB (virus seed)
Grow virus on cell Substrate harvest cell/
supernatant
Cell lysis
Virus separation (centrifugation and
filtration)
Whole virus
Purification (centrifugation and chromatography)
Killed or inactivated whole virus product (BS)
Master cell and working cell banks (MCB and WCB)
Live purified virus (BS)
Viral subunit
Process
Viral activity Number particles Surface protein
Purity
Viral activity Number particles Surface protein
Viral activity Number particles Surface protein
Viral activity Number particles Surface protein
Chemical inactivation
or kill
Viral disruption
Viral activity Number particles Surface protein
Viral activity Number particles Surface protein
Certificate of analysis for BS
Control testing
FIGURE 6.12 Production and preparation of virus (live, killed, or subparticle).
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or in large flasks. Cell growth medium is both well defined and well char- acterized to prevent contamination or cross-contamination of cultures with adventitious agents. Raw materials from undefined or animal sources, such as serum, are generally unacceptable as media supplements. Virus from WCB is inoculated onto cells and, after a period of incubation and viral replication, the cells are harvested, lysed, and then virus is separated from large debris by centrifugation or filtration of the medium. It is then purified using density gradient centrifugation or selective filtrations, to separate viral particles from impurities such as cell debris and culture medium. Virus of high purity, BS, results from the process, and this is extensively tested for identity, safety, ste- rility, purity, and potency. Potency tests are developed to reflect the intended use of each product. Virus may be examined for desirable traits, those that enhance the intended mechanism of action. For example, if a vaccine must first attach to epithelial cells to be immunogenic, testing might examine the virus for the ability to bind that receptor. Hence, both potency and safety tests are often complex assays, many immunological or molecular and others performed in tissue culture or using live animals.
In contrast to virus, live bacteria are propagated from bacterial WCB as recombinant biotechnology products. These are grown in defined medium, preferably without animal-derived supplements and in the manner described earlier for fermentation of recombinant proteins. Bacterial cell growth and expression are monitored for amount and for desirable traits or attributes, often identified in the research laboratory. The cells are harvested in a man- ner that retains viability and are processed further to remove impurities, such as dead cells, cell debris, and components of the medium. Purification of live bacteria is based largely on physical separation with the help of cen- trifugations, washes, and filtrations. Bulk substance consists of live bacterial cells held in a physiological medium and perhaps with a cryopreservative, because they will likely be stored in the frozen form. Testing protocols are developed to ensure that traits are retained throughout the processes. Identity, safety, purity, and potency testing are also completed. Potency test- ing may include the ability to express an antigen or attach to a cell substrate.
Production of Products Composed of Mammalian Somatic Cells or Tissues
With the advent of tissue and cell replacement therapies and the new era of regenerative medicine, biomanufacturing operations developed methods to expand somatic tissues and cells. While selected individual cell types, such as those used in laboratory tissue culture, had been produced for decades, the growth of somatic cells and tissues intended as replacement therapies in human subjects present new manufacturing challenges. Today, autologous tissue regeneration and replacement is a growing biotechnology industry and a quality system suited for this technology, Good Tissue Practice, has been developed as regulatory guidance.
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Replacement of knee joint cartilage provides one example. The objective is to grow, in vitro, healthy autologous cartilage that can be used to replace dis- eased cartilage in a joint. To begin, a piece of cartilage is surgically removed from a healthy joint of the patient and the cells from this healthy tissue are transferred to a biopharmaceutical production facility. Here, the cartilage cells are expanded on an inert biological matrix to confluence and form a sheet of cultured cartilage cells. This sheet of cells is returned to the surgeon, who then implants it into the damaged joint. While this product does not utilize recom- binant technology, it does apply the biotechnology practices of cell selection or isolation, purification, cell culture expansion or growth production, and quality control testing that accompanies a complex biomanufacturing process. Many technical and quality hurdles have been overcome, largely through planning, proper application of the existing technologies, and invention of new methods.
This approach has also been used to illustrate another example, growth of autologous epidermis, new skin for patients subjected to severe burns. A flow diagram for skin production is shown in Figure 6.13, in which major
Donor (patient) epidermal cells
Dermal substrate
Adventitious agents Viability
Bioburden
Quality of collagen and fibroblasts
Mature epidermal tissue (BS)
Certificate of analysis for BS
Process
Viability Purity
Identity
Biological function: Viability
Histological analysis Barrier function
Animal collagen matrix
Downstream control testing
Upstream control testing
Epidermal cell purification
Seed epidermal cells on dermal substrate
Propagate keratinocytes on dermal substrate
Stratified epidermal tissue
Fibroblasts
FIGURE 6.13 Production of epidermal somatic cells and skin tissue.
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attributes and production steps are highlighted. The seed tissue is taken from an unaffected area of the skin of a burn patient. This critical raw mate- rial must be of high quality. Culture methods encourage rapid and consis- tent growth of skin tissues on a matrix or artificial membrane, to the stage of confluence, thus achieving a sheet of tissue two to three cell layers thick. Scrupulous aseptic technique is applied at every step to ensure a safe and sterile product. Unique quality control methods focus on attributes of this product. For example, measurements of skin tissue tensile strength ensure that the tissue can be transported and then surgically sutured without tear- ing. Identity testing confirms that donor skin sample matches exactly the skin tissue yielded at the end of manufacturing. The outcome is a biomanu- facturing system producing a high-quality skin or a tissue product able to close horrific burn wounds, achieve homeostasis, and alleviate patient suf- fering and provide new market opportunities.
Production of Cellular Products Derived from Pluripotent (Stem) Cells
The biomanufacturing community has rapidly evolved to produce prod- ucts from pluripotent cells (i.e., stem cells); however, there remain many unknowns regarding the clinical feasibility of this technology. The following overview of technologies available for production and testing of differenti- ated cells from pluripotent cells gives one an idea of how biomanufacturing might be performed in the future. More times than not, groundbreaking sci- ence and dire clinical need are what drive the rapid evolution of technology to accommodate the applications of new cellular therapies from benchtop to bedside. One fairly recent example of rapidly evolving technology is the development of induced pluripotent stem cell (iPSC) methodology, which was initially discovered in 2006. Induced pluripotent stem cells are spe- cialized adult stem cells reprogrammed either chemically or genetically to a more undifferentiated and stable stem-cell-like state. These specialized cell populations are essentially coaxed into dedifferentiating from what appeared to be a committed state into a less-defined developmental state, which is better equipped to promote tissue repair or organ-specific regenera- tion. For example, the seemingly most appropriate cellular therapy product for a heart indication would be to use progenitor heart cells (iPSC-derived) destined to become a cell type specific to the heart (e.g., cardiomyocytes) rather than using stem cells isolated from bone marrow, which rely on micro- environmental cues when delivered to the injured heart to facilitate heart muscle regeneration.
Although iPSC technology is nearing evaluation in the clinic, many issues have yet to be better understood or resolved to ensure that a reason- ably safe product will be used in human clinical studies. One such issue is the challenge of controlling and characterizing the differentiation state of the cellular product; the desire is to have a homogenous cell popula- tion, which has been difficult to achieve. Although much improvement
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has been made, this inefficient programmed cell differentiation is much less than 100%, which raises concerns about the multiple cell populations and the potential variability associated with every differentiation process. The differentiation inefficiency results in a less characterized heteroge- neous cell population, with undesirable carryover of cells that represent an impure cell population with variable differentiation potential. This differentiation inefficiency is apparent in vitro and as such raises safety concerns in the regenerative medicine community of unpredictable cell fate and unwanted differentiation potential after in vivo delivery of an iPSC product. Technology developments will continue to strive to main- tain control and confidence of the differentiation state regardless of the manufacturing process scale and will be able to demonstrate that the cell population can be safely administered to patients (e.g., maintain nontu- morigenic potential). In vivo distribution and persistence in the human body will remain an important safety consideration, until control of the differentiation process is achieved, thus resulting in a homogenous cell product that is predictable.
A biomanufacturing plan for products derived from stem cells is based on a cell therapy indication and a product composed of differentiated, tissue- specific, but viable growing cells of a particular lineage. As a new technology, regulatory guidance—even general information from FDA—is important to developing a compliant product. Fortunately, FDA has announced general guidelines for development from pluripotent cell sources and established helpful information on related products under Good Tissue Practices and other regulatory guidelines. These recommendations, while still quite gen- eral and largely unproven for pluripotent cell-derived products, provide a foundation for production schemes.
However, many questions regarding application of these types of new regenerative technologies to ensure production of a safe and effective prod- uct have yet to be answered. First, do the proposed manufacture and control methods result in a safe product and how exactly do we demonstrate the prod- uct safety by using currently available scientific methods? Second, can any biomanufacturing scheme actually generate differentiated cells and tissues from pluripotent cells, and, as BS, FP, and after growing in the patient, will these cells demonstrate the attributes of identity, purity, and potency? Third, is it possible to apply to pluripotent cell-derived biomanufacturing protocols those methods and quality criteria that are used for somatic cell and tissue production, or will it be necessary to begin anew and develop unique schemes for these cell types. In addition, when produced in great numbers, will dif- ferentiated cells, derived from pluripotent cells, remain differentiated or will they revert to an undifferentiated status or even to a malignant state of differ- entiation? In addition, how do we ensure that pluripotent stem cells, derived from an unknown source, do not carry adventitious agents? Do traditional methods applied to somatic cells provide adequate safeguards for pluripotent cell-derived products?
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The biomanufacturing design and subsequent plan must address these issues and answer questions by using novel manufacturing methods that go hand in hand with appropriate quality control tests. A hypothetical scheme for biomanufacture and control of a pluripotent cell-derived prod- uct is proposed in Figure 6.14. Despite the novel technology and source of the product, issues that confront the manufacturing team are very similar to those experienced several decades ago by teams of biomanufacturing scientists who intended to produce a recombinant protein in E. coli. Then and now, application of precedent, good planning and careful and thorough experimentation are keys to preparing a biomanufacturing plan, moving a novel product through the manufacturing cycle, and bringing it to market. Indeed, by application of good scientific and manufacturing practices, it is conceivable that any biotechnology concept can be taken from the research laboratory and be successfully produced and marketed for the benefit of mankind.
Production of Biological Molecules by Transgenic Animals or Plants
A host of alternative production systems for recombinant biopharmaceutical molecules are currently in development. Most purport to make a product that is of equal or greater quality when compared to biomanufacture pro- duced by traditional methods such as fermentation or cell bioreactor pro- duction. Most animal and plant systems promise expediency, higher quality, and lower cost. Although increasingly more popular, today only a few plant or animal biomanufacturing concepts are technically mature or proven and many have already fallen by the wayside, as they have proven difficult to manage, give small yields, or produce an unacceptably impure or impotent product. Nonetheless and based on some successes, notably production of recombinant protein in transgenic goats, there are a host of new plant- and animal-based biomanufacturing technologies in development. Biopharming is a casual name given to the application of transgenic plants or animals to produce biopharmaceuticals.
A transgenic plant or animal is a plant or animal that has been genetically altered using recombinant DNA techniques to create a genetically unique organism. The transgenic organism contains an exogenous gene or genes that have been intentionally inserted into their genome. Once inserted, the expression of the exogenous gene can express the protein of interest, often a glycoprotein, and, in some cases, secrete this protein with tissue fluid. In the case of a transgenic plant, the protein can then be extracted from bio- mass, that is, stems or leaves, or from seed. In case of transgenic animals, the protein is available from secretions, notably milk. Hence, the plant or ani- mal functions as a bioreactor, producing appreciable amounts of the desired protein as BS. As one would anticipate, production of a transgenic organism capable of producing and secreting the perfect protein is highly technical and requires significant experience and skill. As with any biopharmaceutical, the
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Process Control testing
Human stem cell (Pluri/multipotent)
Clone: Derive cell line
Master and working cell banks
Expand culture from cell bank
Differentiation in culture flask to smooth muscle
Harvest and pool
Purification: Centrifugation selection methods
Selective adherence to substrate
Differentiation protocol for smooth muscle
Collagen substrate (Urinary Bladder Fibroblast)
Degree heterogeneity Adventitious agents
Microbial burden Phenotype and genotype
Differentiation
Heterogeneity Phenotype Genotype
Differentiation
Heterogeneity Phenotype
Growth characteristics Degree differentiation
Safety testing
Quality of raw materials
Heterogeneity Phenotype
Growth characteristics Degree differentiation
Safety testing
Heterogeneity Phenotype
Growth characteristics Degree differentiation
Safety testing Expand smooth muscle to tissue
Mature smooth muscle to tissue
Induction factors (Cytokine and Growth)
Certificate of analysis for BDS
FIGURE 6.14 Hypothetical scheme for biomanufacture and control of human cells and tissues (bladder smooth muscle) derived from pluripotent cells.
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protein product must be isolated and purified from other molecules and it must possess posttranslational modifications and structure that allow full biological function and should be without modifications that could make the molecule allergenic or nonfunctional. This process, using a transgenic goat secreting in milk, is provided in Figure 6.15.
Biomanufacture of proteins that demand complex glycosylation and post- translational carboxylation has been successful using genetically modified animals where it has failed using genetically engineered bacteria or yeast. Recombinant whole animal systems, such as transgenic goats, have been used to produce effective molecules of human blood proteins such as anti- thrombin, fibrinogen, or alpha-1 antitrypsin. Further, to allow for convenient collection and purification of the protein, the transgenic gene product must be expressed and secreted into a harvestable body fluid. To achieve such design objectives, human proteins have been transferred to dairy animals, such as goats, for the purpose of gene expression in the mammary tissues and protein secretion into the animal’s milk. Protein is then purified from milk by using protocols that consider separation from cellular debris, other milk proteins, and fat. This is accomplished using downstream purification methods such as centrifugation, filtration, and chromatography, but under the design plans that consider the unique impurities in mammalian milk. Issues related to adventitious agents have been addressed. Additional infor- mation on downstream protein purification methods is provided later in this chapter. A specific example of how transgenic goat emphasizes the economic advantages and the scale of production this technology can offer are pre- sented in Box 6.4.
Rabbits have also been used as a reliable and efficient source of a clotting factor for a rare blood disorder. In this case, a transgenic rabbit is created as a bioreactor, and a C1 esterase inhibitor, produced in the milk of transgenic rabbits, is licensed for the treatment of hereditary angioedema. A lactating rabbit can produce 10–12 g of protein per liter of milk, whereas traditional methods of protein expression from cell culture systems are less efficient, with yields ranging from 0.2 to 1.0 g of protein per liter of culture media. These are just a couple of examples where cutting-edge biotechnology has demonstrated the proof of concept and then has been successfully developed to reach the marketplace.
A second method, producing biopharmaceuticals in plants, has been tested in a variety of plant species for over two decades. A scheme for production of a recombinant protein by a transgenic higher plant species is depicted in Figure 6.16. The ability to transform commercially useful plants, first with genes of other plant species and then with genes of animal origin, has pro- vided the foundation for this technology. Cultured plant cells of both higher- and lower-order plants, for example, maize or tobacco and algae or mosses, have been tested as biomanufacturing systems. Higher-order plants are usu- ally used; however, systems using lower-order species are under develop- ment. The first step is the production of a transgenic plant, a process that is
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Vector containing transgene
Injection of transgene into goat embryos
Transgenic goats
Offspring
Goat milk containing biopharmaceutical protein
Embryo implantation
Vector
FIGURE 6.15 Transgenic goat producing biopharmaceutical protein in milk. The recombinant process begins with constructing a vector containing the protein of interest, designed to target pro- tein expression in the milk. The transgene is injected into goat embryos by using pronuclear microinjection methods and then embryos are reimplanted into a host female goat. To iden- tify germline transmission of the transgene, offspring are screened for transgene expression in the milk. Milk is collected from the transgenic offspring and milk proteins are isolated. Recombinant protein is easily purified from endogenous milk proteins and other impurities that reside in goat milk.
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facilitated by several novel gene delivery methods well suited for transfec- tion. Once mature, samples of various plant tissues are tested for the desired trait, such as expression of a mammalian protein. Next, these plants must be cross-bred, by using methods applied to the development of hybrid plants; this is followed by another round of selection. The ultimate source of the recombinant molecule may be any plant tissue expressing the product, but seed is a preferred choice, given its ability to store large amounts of pro- tein in an environment with a low microbial bioburden. The plant tissue must then be processed to release the desired recombinant protein from cells and tissue matrix and bring it into solution. The solution is clarified of plant debris and processed using purification methods described above for other proteins in order to derive the recombinant molecule as BS.
BOX 6.4 THE TRANSGENIC GOAT AS A BIOREACTOR
• In 2009, FDA approved ATryn, a recombinant antithrombin indicated for the prevention of thromboembolic events in a rare human clotting disorder.
• Antithrombin can be isolated from human plasma; however, to fulfill market requirements, 100 kg would take one year’s sup- ply of plasma donations.
• However, the same amount of recombinant antithrombin can be collected more efficiently and economically from 150 trans- genic goats, animals expressing and secreting this protein in milk. A lactating female goat produces up to 800 L of milk in a year.
• The transgenic goat model as a bioreactor can in theory and in practice yield, after protein purification from milk, approxi- mately 4 kg of a recombinant protein.
• The cost comparison to produce the same amount of recom- binant pharmaceutical-grade protein is quite staggering when comparing traditional methods with biopharming.
• Purification is achieved by utilizing isoelectric precipitation, affinity chromatography, and size exclusion chromatography. These methodologies result in a 90% yield with greater than 99% purity.
• The production cost from a bioengineered domestic animal is estimated to be approximately one-tenth the cost of manufac- turing the same recombinant therapeutic protein in a commer- cial cell culture facility by using a bioreactor.
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Choose transfection method Molecular or Biological: DNA, agrobacterium Mechanical: Gene gun, electroporation, and
optical laser Other: Nucleofection and impalefection
Construct expression gene
(selection and marker)
Donor seed Process
Transfection Seedling+transfection
method+genetic construct
Genetic construct
Test in seedlings Ensure high-rate
transfection
Select stable Transformed plant
seed
Grow and cross-breed
Second and third Generation plant and
seed
Centrifugation and filtration
Evaluate recombinant Protein trait in
plant tissue
Discard Unproductive Transfection
Test tissue for Protein trait
First generation plants
Field growth
Harvest seed
Crush and solubilize seed
Purifications and chromatography
Protein BS
BS panel tests and certificate analysis
Activity Total protein SDS-PAGE
HPLC
Test tissue for Protein trait
Test seed for protein trait
Activity Total protein SDS-PAGE
HPLC
Test tissue for Protein trait
Control testing
Establish gene
Hybrid seed
Plant seedling
FIGURE 6.16 Production and control of a recombinant protein by a transgenic higher plant species.
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Several hurdles had confounded or currently confound the application of biomanufacture with transgenic plants. Glycosylation is one example. Plants have evolved unique methods to make posttranslational modifica- tions to proteins. Glycosylation modifications of plants are unlike those in animals, yet they are encoded in the plant genome. Sometimes, expres- sion of a mammalian gene by a transgenic plant unpredictably results in a molecule that has the expected protein backbone but possesses a totally different and undesirable carbohydrate moiety. This is the result of glyco- sylation by plant enzymes, a process that would not happen if the protein molecule were manufactured by an animal cell. A case in point is a mono- clonal antibody produced in maize cells and glycosylated with a series of carbohydrates unique to plants. Such modifications change the proper- ties, often the potency, of the recombinant molecule. Hence, careful plan- ning, based on an understanding of the product and the transgenic host system, is essential to successfully using transgenic plants or animals for biomanufacture.
Another issue is the need to redefine cell banks to meet transgenic plant technology. For example, corn, unlike bacteria or immortalized cells, relies on cell banks composed of actual monocotyledon seeds. Owing to sexual reproduction and other traits, higher plant seeds are quite heterogeneous in nature and genetic makeup. A corn seed bank is not derived from a clone and is not genetically pure or homogenous. This can cause problems with variable field growth, sexual reproductive capacity, or protein expression. In addition, the environment of field-grown plants is quite difficult to con- trol. Weather in a corn field is highly variable, and higher plants grow, quite literally, in dirty environments, certainly as compared to that of the fermen- tation vessel in an aseptic biomanufacturing facility. Thus, plant-derived recombinant molecules begin as septic entities, adding challenges to aseptic purification processes. Finally, purification of biopharmaceuticals has also presented new challenges, many unexpected, because extracts of plants have impurities and contaminants that are not found in bacteria, yeast, or mammalian cell systems. Novel approaches, such as use of unicellular and asexual plant species (e.g., algal and moss species), growth of plants in con- trolled environments, and derivation of recombinant proteins from selected plant tissues, have overcome some, but certainly not all, of the issues facing biopharming.
Yet, there are potential advantages, notably production of large quantities of product at low cost, that are derived by biomanufacture from transgenic plants or animals. Whether produced in domestic animals or plants, human recombinant proteins intended for therapeutic use need to consider the real potential of eliciting an unwanted immune response due to the xenogenic nature of the protein. The FDA has provided guidance regarding the produc- tion of therapeutic proteins by both plants and animal bioreactors. Further recognizing the potential immunogenicity and associated safety con- cerns surrounding xenogenic protein technology, FDA guidance discusses
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requirements to minimize the risks of immunogenicity and the likelihood of generating an undesirable immune response.
Production of Biologically Active Lipids, Glycolipids, and Complex Carbohydrates
Peptides, lipids, complex carbohydrates, and glycolipids are potentially important biomolecules of economic and therapeutic value. This is perhaps best demonstrated by complex carbohydrates and glycolipids, which have been successfully used as vaccine antigens, and by lipid molecules, which are then considered as vaccine adjuvants. Each has a biological derivation and function in nature and can be manufactured by man in the laboratory. Biomanufacture of these molecules in larger quantities is done through one of the following two routes: (1) production by a live organism that is not recombinant but naturally expresses the molecule in nature, followed by isolation and purification of the macromolecule and (2) chemical syn- thesis, using processes analogous to, but often more complex than, those applied to production of small-molecule drugs. An example of a natural product (platelet-rich plasma [PRP]) derived from bacterial cultures and then purified using biomanufacturing methods and conjugated to a toxin (T) to yield Haemophilus b conjugate (PRP-T) is outlined in Figure 6.17. This complex carbohydrate is used as a vaccine against Haemophilus influ- enzae type b.
The chemical synthesis of biologically active, nonproteinaceous macromol- ecules is becoming more elegant and widely adopted. For example, instru- ments are used to synthesize complex carbohydrates on lipid-like backbones, a method that is dependable but still gives low yield.
The downstream or purification steps used for lipid, carbohydrate, or gly- colipid biomolecules vary in some respects from those used to purify pro- teins. This is understandable because lipid or carbohydrate molecules are quite unique in chemical structure and composition and physical properties. Manufacturing planning takes into consideration the molecular attributes of these candidate products and matches them with technologies available for purification. Many of the methods applied to a manufacturing step may be borrowed from the research laboratory and adapted to biomanufacturing.
Production of Biologically Active Peptides
A peptide is a string of amino acids, 40 or less, by most definitions, in a given sequence. Biologically active peptides are used or tested as biopharmaceu- ticals, enzyme inhibitors, laboratory chemicals, and for other purposes. As they are shorter than proteins, peptides typically have no posttranslational modification and lack complex secondary structure. Various technologies are available to manufacture peptides, and the choice of method is based on the intended use and specifications, notably purity, the number and nature
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Purified protein carrier
(Diphtheria toxoid)
Conjugation (Reductive amination)
SDS-page HPLC Safety
Identity
Molecular weight purity
Hib, master and working cell
banks (Bacterial)
Growth in culture
Collection by centrifugation
Centrifugation and tangential flow
filtration
Kill bacteria
Remove polysaccharide
Purify polysaccharide
Chemical modification
(activation) of polysaccharide
Purification
Bulk conjugate Certificate of analysis
(BDS)
Certificate of analysis
Cell banks Hib identity
Contamination impurities
Purity Identity
Molecular size Moisture
Polysaccharides Impurities
Molecular size Degradation
purity
Growth Rate yield
Polysaccharide pH
Control testing
Process Controltesting
PRP: Protein ratio Residual conjugation
agents Unbound PRP
Molecular sizes
FIGURE 6.17 Production of a polysaccharide-protein conjugate product (Haemophilus influenzae type b vaccine).
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of amino acids in the string, the amount required, and the cost. Peptides are often manufactured using automated equipment, the peptide synthesizer, to build the string, one amino acid at a time, beginning at a solid matrix such as a plastic bead and continuing to the end. Hence, most peptides are made using tools common to synthesis of other organic molecules.
Impurities—truncated peptides, fragments, and free amino acids— remain in solution with the peptide product after synthesis, so purification is required. Further, peptides in their unprotected state, perhaps because they represent incomplete or fragmented protein sequences, can be unstable. Formulation, fill, and finish procedures must carefully consider postmanu- facture hold and storage environments to ensure a potent and stable product.
Certain other biomolecules are peptide-based biomolecules, in that they are peptides but with another large molecule bound to them. Examples include glycopeptide and protein–peptide combinations. The production of such molecules may require considerable planning and technical effort, as there are no established manufacturing methods available for these biopharmaceuticals.
Production of Combination Products: Biopharmaceutical with a Drug or Medical Device
Biomolecules are frequently used with material from another source to produce a combined effect. For example, a recombinant bacterium used to clean oil spills may be applied to the spill along with a short-chain organic molecule that disburses the oil and facilitates the metabolic activity of the bacterium. In biopharmaceuticals, the pairing of a biological product with a medical device, a drug, or both is considered a combination product by reg- ulatory agencies (Chapter 3). A recombinant DNA vaccine that is delivered exclusively with a special injector device is one example. Another example is an engineered retrovirus that must be given with a specific drug to enhance the potential therapeutic activity of the retrovirus. A case in point is the use of a drug to facilitate insertion of a therapeutic gene into the genome of the recipient. For the full intended therapeutic effect, both substances, drug and biologic, must be pure and potent under a defined schedule of usage.
Biomanufacturing plans identify combination products and present a strat- egy for producing both products to specifications that will yield the desired, combined effect. Manufacturing specifications for combination products are often challenging for the operator, because individual roles must be assigned to attributes of each product and synergistic effect also needs to be consid- ered. For the example of a recombinant DNA vaccine (a biopharmaceutical composed of plasmid DNA), delivered by a needleless jet injector, a medical device, the vaccine plays the dominant role, because it imparts the therapeu- tic effect by stimulating the immune response. However, the device must perform per established specifications, otherwise the DNA vaccine will not be delivered correctly and might not then exert the intended pharmacological
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effect, that is, vaccination. Early considerations for a manufacturing plan takes into account this needed synergy in combination products. Specific concerns in DNA-related examples might be as follows:
• DNA formulation must be compatible and stable with materials of the device.
• DNA must be concentrated so that it can fit into the device chamber during storage, but it must also be capable of rapidly exiting via the device needle on actuation.
• Device must consistently deliver an exact amount of DNA. • Device must be easily and correctly used by medical staff.
From this example, it is clear that design, actually a codesign of a biological product and a medical device, is critical to success whenever a biotechnology product is partnered with a drug, a device, or even another biopharmaceuti- cal. Manufacturing plans and technologies consider complex performance issues and ensure that these issues have been addressed early in develop- ment. Experimentation is performed to test the effects of additional vari- ables, as each product brings into play a new set of issues.
FP: Formulation, Fill, Finish, and Labeling
A product is used in clinical investigations or commercially only after it has been properly formulated, tested, placed into a protective container or a delivery device, and then labeled. Having reviewed the various types of biotechnology products presented earlier in this chapter, one might well imagine the need for a host of formulations, containers, and labels, as well the procedures, to complete their FP. In this section, we review the technolo- gies used to bring a biopharmaceutical from BS to the format intended for the user, that is, the FP.
The biomanufacturing plan for an FP focuses on the indication and the user. Biomanufacturing is a market- and user-driven process, and therefore, the formulation, container, and label are designed for a specific purpose, that is, the needs of the user. For biopharmaceuticals, the user is the patient or a subject enrolled in a clinical investigation, but for many products, a full definition of the user includes the medical professional prescribing or administering the product. For a tissue product, such as the skin or cartilage, mentioned in an earlier example, the surgeon is also a user of that product. Indeed, the patient may never see the cartilage tissue product as provided by the manufacturer, because the cartilage is transferred by the surgeon from the package into the knee joint. In contrast, for the combination product of
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recombinant monoclonal antibody in an autoinjector, the patients, not the medical professional, use the product, receiving it directly from the phar- macy and then injecting it themselves. The medial professional is, however, familiar with the product and its attributes before a prescription is written and may train the patient on the proper use of the product. Most biophar- maceutical FPs are used by more than one person and all of their needs are considered in the FP design.
The FP design considers various product attributes. Formulation decisions rest on the intended shelf life of a product, possible and desirable storage conditions, and the proposed dose. The target dose also allows planning for the amount of product that will be included in the final container and the packaging and labeling that are provided with the container.
A scheme for FP production of a typical biotechnology product is shown Figure 6.18. First step is to select or devise a formulation that suits the nature of the product, intended use, route of delivery, and container. There are many formulation choices available, and others can be developed for a unique product. Common formulations for biotechnology products con- sist of salt solutions, buffers, and a variety of excipients. Once a promising formulation is identified, experimentation and trial and error are the bases for selecting the one that is just right for that product; this means signifi- cant testing must be performed for product attributes and for product sta- bility. Since a product may need to be matched with several formulations before the correct combination of ingredients is identified, the process can be extensive.
Once a formulation has been selected, the actual formulation process for a biopharmaceutical begins with final clarification and, for most products, sterile filtration of the BS. After purification and until formulation, bio- pharmaceuticals are held as BS under refrigerated or frozen conditions. If the active ingredient in BS is stable, as a frozen recombinant protein might be, it may be stored for several weeks or for even months. In contrast, a tissue product, such as skin tissue, might withstand refrigerated storage only for a few days or weeks. Hence, the time allowed between comple- tion of bulk manufacturing and formulation, fill, finish, and labeling var- ies greatly, depending on the product type. Product stability testing is discussed in Chapter 7. No matter what the product, once the decision is made to begin formulation, the process is completed as rapidly as pos- sible to reduce hold time and thus minimize product exposure to the environment. Since most biopharmaceuticals are sterile products, formu- lation must be a strictly aseptic process applying stringent techniques and environmental controls.
An example of formulation, fill, and finish is instructive and a recombinant protein, such as a monoclonal antibody. After sterile filtration, the storage buffer for a monoclonal antibody, an active ingredient in BS, is exchanged for the FP buffer. In this example, 0.9% sodium chloride in water (normal saline) for injection is exchanged with a small amount of detergent. After mixing,
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Formulation buffer
Excipients
Test final product certificate of
analysis
Inspect label
Initial formulation
Fill into vials, syringes, injectors
final product
Label vials
Package and package insert
In-process testing
Inspect Package, carton, and lablesOuter carton
Storage and distribution
Test formulation buffer
Hold and transfer
Process
Test excipients
Control testing
Formulated product
Product as bulk substance (BS)
FIGURE 6.18 Manufacture of final product: Formulation, fill, and finish process.
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the formulated product is again sterile filtered and then transferred to the fill area, where it is dispensed into vials.
In contrast to a recombinant protein, a more complex biopharmaceutical product, such as skin tissue, may require more formulation effort. In this case, it may be necessary to rinse the skin tissue with various buffers and then place it into a nutrient solution enriched with oxygen and chemicals that maintain tissue viability. Terminal sterilization, even by filtration, is not possible with skin and other cellular (or many other biotechnology) prod- ucts, and so, strict aseptic technique is practiced throughout the formulation process. To meet stability profiles of some fastidious biotechnology products, such as live bacterial or viral vaccines, some products have unique additional steps in formulation. For example, dry powders may be prepared by spray drying the product, once it is in the final salt solution. Freeze drying, referred to as lyophilization, is also used to prepare powder from formulated liquid product, after it has been aliquoted into vials.
Fill is the next step in the process. In this step, product is placed into a final container with fill procedures tailored for each product-and-container combination. For the example of skin tissue, product might be placed into a pouch of some type. For the living tissue, the outer container might be a special transportation apparatus replete with systems to provide sterile nutrients and oxygen and an external heat source to maintain a temperature suitable for this living product.
Returning to the example of recombinant protein, the monoclonal anti- body product formulated in saline with a detergent, container and handling requirements present a less complicated fill and finish procedure. Standard pharmaceutical glass vials are chosen as the primary container. These vials have butyl rubber caps, and aluminum crimps are used to seal the cap to each vial. Vials and caps of the highest quality are scrupulously cleaned and heated to remove any contaminants before use. Vials either are filled and then capped and crimped (force a seal over the cap) manually by operators (Figure 6.19) or are filled, capped, and crimped using a machine. Prefilled syringes and injectors are becoming popular containers for biopharmaceuti- cal products, and these are always filled using automated pieces of equip- ment, which are fast, accurate, and less likely to break the aseptic nature of this operation.
Once containers have been filled, capped, and crimped, a permanent label is placed onto the container to provide the user with an abstracted descrip- tion of the contents. Containers include a description of the product, dose, volume, total number of doses per container, source (manufacturer), expira- tion date, lot number, and special instructions or warnings. Imagine fitting this much information onto a label, 2.5 × 1 cm, for a small vial!
Packaging is the next step in producing the FP. Containers are placed into a protective inner package, for example, a light cardboard. A package insert is added to this box before an adhesive label is attached to identify the contents and then it is closed with a tamper-proof seal. The package insert, sometimes
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referred to as labeling, provides product information to both the medical pro- fessional and the patient or user. Multiple product containers in their outer package are then placed into a larger carton, the outer container, and this too is labeled.
By way of a final example of formulate, fill, and finish, we consider a biotech- nology product that is not a biopharmaceutical but instead is a genetically engineered plant. It is a strain of disease-resistant corn, grown in the field, harvested, cleaned, and then placed into storage in 50-pound bags as BS. The product will be planted in fields by farmers. Before packaging, the seed is formulated by various treatments, perhaps by drying to an established moisture level, followed by addition of powdered fungicide to prevent decay during storage. The seed is then filled, placed in moisture-proof containers, such as 10-gallon plastic buckets, and the buckets are sealed. These buck- ets are then labeled on the outside, and several buckets, along with product information (the package insert), are placed into an outer carton made of cardboard, and this too labeled. The package may then be shipped to a dis- tributor or the user. Although technically quite different from formulation– fill–finish production methods used for a biopharmaceutical, the processes for this and many other biotechnology products follow the same general steps in production, protecting them from harmful environments and mak- ing them ready for the user.
FIGURE 6.19 Fill and finish of final product in a clean room. Two operators, working under a Class 100 hood, manually fill and finish vials of a biopharmaceutical. They are carefully gowned and covered to prevent any possibility of contaminating the FP. The operator on the left is filling the vials, held in a box at the center of the hood, with a dispensing pipette, whereas the operator on the right adds a cap to each vial. In the next step, an aluminum seal will be crimped to tightly close each vial and a label will be added. (Courtesy of Waisman Clinical Biomanufacturing facility, http://www.gmpbiomanufacturing.org.)
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Biomanufacturing Facilities, Utilities, and Equipment
Facility Design Considerations
Biotechnology products, notably biopharmaceuticals, are manufactured in very special facilities. Biomanufacturing facilities are not only unique to our industry but are also custom designed for each class of product and even for a specific product. All the activities we have mentioned in this chapter, with the possible exception of upstream production in biopharming, must be per- formed under a roof, above a floor, and between four walls. Biomanufacturing facilities are complex and expensive to both build and operate. Staff are pro- fessional specialists and perform everything, from emptying the trash and cleaning surfaces and equipment to aseptically filling the final containers follows and writing procedures and results in permanent records. As each biotechnology product and every biomanufacturing process for a product are unique, facilities are custom designed and have been built and equipped in every imaginable way. Still, there are similarities in facility designs for dif- ferent products, and our discussion of biomanufacturing facilities, utilities, and equipment will provide basic principles of facility design and operation.
Biotechnology firms skilled and lucky enough to find themselves in devel- opment are faced with the need to manufacture their product. After this realization, the first question to be posed by management to the product development team is as follows: When, where, and how will this be accom- plished and at what cost? The answer must be in the manufacturing plan; therefore, options must be considered and choices must be made. A team composed of individuals with business, financial, management, biomanu- facturing, and facility and process engineering experience is chartered to reach a decision on manufacturing options. There are at least three possible ways to meet a biomanufacturing requirement, and these are as follows: (1) do not manufacture the product (and hence, do not develop it further); (2) manufacture the product in-house; and (3) manufacture the product at a CMO. A fourth possibility is to split manufacture, that is, do some pro- cesses in-house and have others completed at a CMO. Not surprisingly, few biotechnology firms select the first option and the rest seem evenly divided between the choices numbered two, three, and four. Many factors beyond the technical need or a desire to have an in-house operation often influence the final decision; these are resources, location, and business and exit plans.
A biomanufacturing facility is designed to produce a specific class or type of product. One would not expect to see a recombinant seed corn operation co-located with a formulation, –fill, and finish operation, and it would be unusual to find, in the same biomanufacturing building, both epithelial cell culture and bacterial cell fermentation. However, some facilities can process quite a number of different products, that is, manufacture on a campaign basis, but even then, they are limited in scope. When considering the building,
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one must choose between erecting a new building or remodeling an exist- ing structure. Many excellent biomanufacturing operations have been built within an empty warehouse, albeit one of high quality in a good location and with adequate services. Actual facility design is best left to professional engineers and architects with experience in biomanufacturing process engi- neering and building, equipping, and maintaining facilities. Early drawings are produced by engineers and architects. These should be carefully exam- ined by process engineers, the individuals with an understanding of the pro- cess intended for the facility. General descriptions of utility and equipment requirements are added to the facility plan. After discussions and one or two more rounds of review, a final facility plan is established. Cost estimates are made by professional biomanufacturing facility engineers.
The Facility and Utilities: A Controlled Environment
Most biopharmaceuticals are sterile products of exceptionally high qual- ity and reliability. Biopharmaceutical production requires special controls, because products are intended for use as medicines by large numbers of people and many of these medicines are given by the parenteral route. Biopharmaceutical production and the facility producing the biopharmaceu- tical are highly regulated, and for marketed product, the facilities must have a government license, so as to ensure that only safe, pure, and potent prod- ucts reach users. Regulations and guidelines make clear the exact standards and specifications for biopharmaceutical- manufacturing facility design and operation. Further, good business practices also demand a quality manufac- turing facility for biotechnology products. Not a month goes by without a news story covering an inadequate and substandard drug, biologic, or medi- cal device facility that produced and continued to provide substandard, even unsafe, product to consumers. These incidents result in expensive product recalls, consumer illness or even death, and regulatory actions against the sponsor (Chapter 4). Such cases lead to negative publicity for the firm and loss of product sales, even for the product not made in that facility. Indeed, a single incident at a manufacturing facility has many times led to the loss of a product line or even to the financial demise of a biopharmaceutical firm. The combined lesson is that a manufacturing facility must be first-rate from the ground up and in all respects.
A biomanufacturing facility is, from the outside, a building that looks like any other commercial structure; however, from the inside, it is established and equipped exclusively for the production of biopharmaceutical product. How then is the proper environment for biomanufacture established under the general facility plan? First, that building is designed to house a particular process or several similar processes. Second, the design considers the need for production by aseptic processes, so as to reduce the incidence and spread of microbes and other potential contaminants through the use of segregated, clean work areas. The facility is also planned to house adequate utilities and
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equipment, which are again properly designed. Biomanufacturing requires sufficient space for operators, various processes, utilities, and equipment; further, regulations require and common sense suggests separation of vari- ous activities to prevent mix-ups, contamination, or cross-contamination of product or spread of microbes. The design must consider utilities, reliable sources of water, electricity, natural gas, and heat, ventilation, and air condi- tioning (HVAC). Finally, a facility must be well managed by highly trained and experienced professionals, and, just as with the manufacturing process, it must be run by strict and compliant written procedures and records. Does this sound rather expensive? Indeed, it is.
Once a decision is reached to build a biomanufacturing facility according to a design, additional planning is in order. Detailed product-manufactur- ing plans allow the facility planner to impose on the facility design a pro- cess map, a schematic in which the manufacturing process is drawn, stage by stage and step by step onto a general facility design. This allows one to determine whether or not all the pieces—processes, equipment, utilities, and workflow—will work in harmony. Consulting engineers and architects and biopharmaceutical process engineers who have experience in pharma- ceutical manufacturing are retained to examine rough plans and refine the drawings to ensure compliance with local and state regulations. Plans and drawings are revised, discussed, changed, and finally finished. The facility plan now supports business functions such as accepting bids from building contractors and utility and equipment manufacturers. Now, upper manage- ment can be given a firm estimate of the cost.
Operation of Clean Work Areas for Biomanufacture
Controlled processes and aseptic processing are crucial to biomanufacture. At the heart of aseptic processing is the need to keep viable particulate mat- ter, specifically bacteria and yeast, from contaminating a product. The pri- mary source of microbes is humans (e.g., skin and hair) and materials that enter into a clean area. Introduction and spread of microbes are controlled by facility, utility and equipment design, environmental awareness regarding microbial burdens (Chapter 7), and sound aseptic operational procedures. The facility floor plan is critical to maintaining a clean environment, as it allows for segregation and the logical flow of activities and products. Entry of people and raw materials and exit of waste are carefully controlled to reduce the entry of contaminants into a clean work area. Before entering, individu- als don special gowns, footwear, and masks that cover much of the body, thus reducing the microbial burden shed by them into the clean room envi- ronment. Materials are sterilized or disinfected immediately before these are brought in a clean area. Before entry, water that is to be used in a clean area is filtered to remove any viable particles. As microbes and contaminated dirt particles move freely in air, the HVAC system is designed to constantly filter air through high-efficiency particulate air filters. Moreover, the air is
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circulated rapidly and in great volume to ensure that any particles generated by process activities are swept from the room and filtered to prevent product contamination. Doors and pass-through openings control airflow between rooms, moving air by an established pattern from the cleanest to less clean rooms, thereby further limiting particulate and microbial movement. Since microbes adhere to and multiply on surfaces, all ceilings, walls, and floors are finished with highly resistant epoxy surfaces to discourage microbial col- onization and to withstand repeated scrubbing and disinfection. All equip- ment surfaces are designed to tolerate harsh antimicrobial treatments.
Movement of everything—people, raw materials, trash, and product— within a facility must flow in a predetermined direction. Highly controlled aseptic operations, such as fill and finish, are performed in highly classi- fied areas segregated from other manufacturing and nonproduction areas, so as to prevent contamination of the FP. During and after biomanufacturing operations, measurements of air quality are taken, and each room must meet an air quality standard or classification. There are two manners of classify- ing air within a clean area; both of them are shown in Box 6.5. Everything in a clean area, including air, water, surfaces, and the gloved hands of every staff member, are sampled or swabbed to test for microbial contamination.
Operations not requiring a clean environment are kept apart from clean areas dedicated to aseptic processing. For example, packaging and labeling are generally relegated to an area that is not highly controlled or classified. In addition, quality control laboratories, offices, and meeting rooms are estab- lished outside the clean area. In summary, a facility design facilitates the
BOX 6.5 MEASUREMENT OF PARTICLES IN A BIOMANUFACTURING CLEAN ROOM OR AREA
Room Classifications
Published Specifications
ISO 14644–1 FED STD 209E (U.S.)
ISO 3 Class 1 ISO 4 Class 10 ISO 5 Class 100 ISO 6 Class 1000 ISO 7 Class 10,000 ISO 8 Class 100,000
Note: The particle counter instrument is used to measure particles in air. An air sample is taken into the instrument; particles of a specific size (e.g., >0.5 µm) are counted and total air volume is measured. Particle counts are then given as number of particles per cubic foot or particles per cubic meter (ISO 14644–1; International and European). A manufacturing facility might have air classified as 100,000/ISO 8 in general preparation or laboratory areas, as 1,000/ISO 6 in clean work areas, or as 100/ISO 5 in areas for performing aseptic operations, such as filling vials.
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prevention of mix-ups, provides a flow from clean to dirty, allows segrega- tion, and isolates critical steps in an effort to ensure pure, potent, and safe product.
A biopharmaceutical operation also considers the utilities in support of manufacturing operations. Temperature is always controlled by heat- ing air-conditioning and ventilation equipment. The production of some products also requires humidity control. Gases to incubators, steam to sterilization equipment, and water to make solutions must enter the clean area. Water for injection is the purest grade and may be produced by the manufacturer in-house. However, towing to the complexity and expense of producing WFI, many biotechnology firms simply purchase it in bottles as a United States Pharmacopeia (USP) reagent (Chapter 7). Steam is used in most biomanufacturing facilities to sterilize raw materials or equipment and to clean and disinfect surfaces. In such cases, the steam is produced as clean steam, generated with special equipment from WFI. Mechanical equipment, such as HVAC and water purification systems, are always monitored and alarmed.
Biomanufacturing Equipment
Many pieces of equipment, including biological safety cabinets, centrifuges, filtration apparatus, fermenters, bioreactors, chromatography apparatus, controllers, microprocessors, and incubators, to name a few, are operated in a clean area. Special or unique pieces of manufacturing equipment—and there are many to manufacture biopharmaceuticals—must be of the highest qual- ity and some are specially designed for the biomanufacture of one product. Equipment specifications ensure proper performance, and all equipment are periodically calibrated, controlled, and monitored during operation. Rigorous cleaning is necessary. Cleaning protocols are often complex pro- cedures, because all residual product and cleaning agent and microbes must be removed after use. In addition, many pieces of equipment are sterilized before use or reuse. Once the facility has been built, facility, utilities, and equipment are commissioned and validated, along with the process itself. In addition to product-specific requirements, there are also basic quality requirements for cGMP operations.
Contract Manufacturing Options
It is no wonder that many biotechnology firms select a CMO to produce their product. At first, many resist this option because they believe that the firm will lose some control of product manufacture, especially if the CMO is some distance away. However, renting capacity can be economically attractive and
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there are many ways to establish a partnership in which the sponsor retains adequate control of the biomanufacturing program after it is placed at a CMO. Indeed, procedures, reflected in Box 6.6, are recommended for selec- tion of a CMO.
Even if a CMO is retained to perform critical biomanufacturing stages, many firms elect to perform some processes in-house. Using outside ser- vices to formulate, fill, finish, and label is quite common in the biotech- nology industry. An added benefit of contracting these types of services is that this approach necessitates the technology to be mature enough and
BOX 6.6 CONSIDERATIONS FOR RETAINING A CONTRACT MANUFACTURING OPERATION
Plans List your needs from a contract manufacturing operation (CMO), regarding type of product, phase of development, and biomanufacturing by stage. Is this a clinical Phase 1, 2, or 3 study? What exactly must be done by the CMO: early development; cell banking; upstream processing; downstream processing; formulation, fill, and finish; quality control testing; or more than one of these functions? What are the deliverables: BS; final product; amount of product; or number of containers, reports, or records?
Competencies Identify core competencies of CMO and match to your requirements: history; size; management philosophy; experience and willingness to work with small, medium, or large biotechnology firms; experience by type of product, phase, or capacity; dedicated or shared facility; references; location (region and country); profile in the CMO community; and possibility of strategic partnership.
Equipment Will it be necessary to purchase or lease specialized utilities or pieces of equipment, or is everything already in-house at this CMO? What is the cleaning and change-over process in case of dedicated equipment or shared equipment?
Microbiology If necessary, can the CMO work with live organisms or material that requires stringent or unusual aseptic technique or environment? Are antibiotics used or allowed in the facility?
Design Consider responsibilities for manufacturing design, planning, and risk analysis and mitigation.
Quality Note quality responsibilities and willingness to enter into a Quality Agreement and Technology Transfer Agreement. If it is a shared facility, evaluate change-over procedures; staffing of multiple projects in facility; and systems to prevent potential for mix-ups, contamination, and cross-contamination.
Schedule Examine scheduling possibilities, typical slack and busy periods, opportunities on calendar, and flexibility going forward. Plan for onsite observations and audits as necessary.
Cost What are the projected costs and unusual expenses associated with the CMO, and will it be cost-effective, resource-effective, and time- effective to retain this CMO?
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be amenable to technology transfer from in-house to an outside contrac- tor for this to be successful. At some stage in development, technology transfer is likely going to be imminent, either to support later stages of clinical development or to meet commercial demands requiring large- scale production. Therefore, an early lesson in technology transfer, iden- tifying subtle but important vulnerable processes in the manufacturing process, may add value during the early stages of product development and be less costly in the end. Recall that it is much easier and generally more acceptable to tweak the production process in the early stages of development than in the later, more codified stages. Preferably, there are multiple CMOs capable of doing the chosen biomanufacture. Selection of a CMO requires considerable diligence by the sponsor. A history of previ- ous projects provides assurance that a CMO can work with this product and is capable of performing the processes properly. Precontract site vis- its, audits, frequent communications, and a detailed contract are the best means of choosing the right CMO. However, a sponsor’s efforts do not end with the award of a contract, because there must be frequent visits, communication, and coordination between the sponsor and the contrac- tor throughout the life of the contract. Including contractor representa- tives in the project management team is an especially effective method of managing a contract.
Validation of Biomanufacturing Facilities, Utilities, Equipment, and Processes
Validation is an expensive and time-consuming, but very necessary, process that is completed during late-phase development and only after a manu- facturing facility, with all equipment, utilities, and staff, and the exact pro- cess have been established and commissioned. It is considered a regulatory requirement and a good business practice. Marketing approval is not possi- ble unless a manufacturing plant and process are fully validated. Validation, which is defined in a dictionary as to make sound, defensible in common prac- tice, carries a more complex definition in biopharmaceutical development. For biotechnology operations of any type, validation is a formal process of establishing documented evidence that a specific process (or test, equip- ment, facility, or utility) consistently performs and will, with a high degree of assurance, continue to perform within determined specifications and qual- ity criteria. Quite a mouthful, but once broken down, this definition makes sense in light of the complexities of biomanufacturing and the importance of quality in the endeavor.
All manufacturing systems have inherent variations, much of it being accept- able and some being undesirable. Validation is based on an understanding of
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the nature of that variation, its impact on the process, and the ability of pro- cess controls to keep that variation within manageable levels. Hence, valida- tion is an experimental endeavor based on deep knowledge of the process and under which evidence is generated under a protocol. Perhaps, this is one reason that validation is undertaken only in the later stages of manufactur- ing development. Careful planning is an absolute requirement, and valida- tion efforts are always included in a manufacturing plan.
A validation master plan is prepared, usually in late-phase development, for any major validation effort. The scope of the plan is broad and the level of detail is great. The overall philosophy and approach taken to validate the facility and process are provided in the validation plan. The master plan outlines the validation activities related to a facility, including the physi- cal plant, environment and utilities, equipment—both installed and mov- able—computer systems, software and hardware, critical raw materials, biomanufacturing processes, with all stages and key steps, including aseptic, cleaning, and monitoring processes or procedures. Many analytical tests are also validated at this time (Chapter 7).
From the validation master plan, validation protocols are written to exper- imentally examine each critical step of a manufacturing process. Validation of a facility might involve dozens or even hundreds of protocols. A protocol breaks down a system into simple parts and describes each critical parameter, quality attribute, and operational specification. It ensures that attributes are measurable and testable and that each measurement is scientifically sound. Specifications are developed for results derived from each test or measure- ment. Validation activities are spelled out in detail, usually with standard operating procedures, production records, and testing instructions, and these are identified in the protocol. The work is done by scientists and engi- neers, working closely with quality assurance professionals, and it is normal to employ consultants and contractors to assist employees with these her- culean efforts. Good statistical practices are also utilized in most protocols. Validation is fully documented, from the master plan to the final validation reports. Validation has a pass-versus-fail outcome; either one meets all the predefined specifications outlined in the validation or the validation fails. Validations are labor intensive, time consuming, which make them costly. With this in mind, the prudent biomanufacturer ensures that all systems are performing as expected before executing validation protocols.
Technical steps are involved in the validation process. First, each piece of equipment, utility, and facility component must undergo installation quali- fication, a process to confirm that each item was correctly designed, built, and installed. Limits for usage are also confirmed in installation qualifica- tion. These are simply operational limits that could not be exceeded during normal operations. For example, instructions and specifications limit the use of a 10 L fermentation vessel to 5–8 L media volume. The next step is opera- tional qualification, a verification process in which the operating ranges of each item are confirmed under operational conditions. Items may be stressed
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to the farthest ranges of operational performance. For the fermentation ves- sel, it might be tested, against specifications, three times, with 5, 6.5, and 8 L media volume. In operational qualification, calibration is completed on mechanical or electrical systems, and control systems are shown to work as designed. Process qualification is the third step, in which the manufacturing process is performed, typically three or more times, so as to demonstrate control, consistency, and achievement of specifications. In process qualifica- tion, critical systems are stressed to ensure that they function properly at the limits of operating ranges. Two or three successful repetitions are the norms under validation protocols, and each one must meet specifications. Validation is not a one-time endeavor, and critical systems must undergo the process of revalidation at periodic intervals, after market approval. In addi- tion, any significant manufacturing change must be validated.
A primary outcome from validation efforts is the confidence that the product will be successfully manufactured within specifications over a long period of time. Validation also ensures that product quality, safety, and effi- cacy are designed into the product as confirmed in early manufacturing endeavors and that ongoing process monitoring will be a part of the product manufacturing life cycle.
Summary of Biomanufacture
Biomanufacturing activities typically entail three major components: pro- duction, purification and final fill/finish. It is a phased process, improved throughout the life cycle of product development, aiming to ultimately yield a sustainable process of high quality and adequate scale of production and purification. The amount and quality of the biological substance must meet clinical study and marketing requirements. Further, there is a diversity of biotechnology products, and so, the manufacture of a given product is based on a custom design, which is based on a carefully crafted biomanufacturing plan. There exist precedence for manufacturing most types of biotechnol- ogy products, but for any new product, no matter the amount of precedence, actual production always requires careful planning and some trial and error. Therefore, biomanufacturing requires a skillful and experienced team, and patience and resources from the management. The result is a process that consistently yields a pure and potent biopharmaceutical product.
Successful biomanufacture requires a well-designed product with product specifications, because the design will change during the course of develop- ment, it also requires design control and carefully stated technical consid- erations. Further, the development process requires the use of appropriate raw materials, a step-wise effort to increase the amount produced, scale-up, quality control, and consideration of GMP. Early stages of biomanufacturing
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include production of the construct, that is, the active ingredient, a molecule, or a cell line. This is followed by production of cell banks and reference stan- dards. Quality control assays and process controls are developed along with the biomanufacturing process. Upstream processing, that is, the production of the active ingredient in bulk, often impure form; downstream process- ing, that is, purification; formulation; and filling of the FP require multiple steps, each composed of a particular subprocess. There are as many subpro- cesses as there are products. Product yield, activity, stability, and purity can be drastically different based on the nature of the product and the selection of the processes. Some examples of product classes are bacteria, viruses, pro- teins, glycoproteins, DNA, RNA, cells or stem cells, transgenic plants or ani- mals, and peptides, and they may be derived from biomanufacture involving prokaryotic or eukaryotic cell lines, biological fermentation or culture, and chemical synthesis.
Each manufacturing process occurs within an environment and environ- mental controls that surround the manufacturing process. Most early phase manufacturing is performed in small facilities, following the basic guide- lines for aseptic manufacture and a proper environment. However, as the scale of manufacture increases, the quality criteria also become stringent, until at the later stages, where both the manufacturing process and the facil- ity are validated. Appropriate facilities, utilities, and equipment are selected to provide an acceptable environment, one that meets regulatory require- ments. Production of the FP involves formulation, fill, and finish, a process that is scrupulously aseptic.
References
Hamburg MA. U.S. Food and Drug Administration/FDA Strategic Priorities 2014–2018, September 2015.
Woodcock J. 2005. American Association of Pharmaceutical Sciences-FDA-ISPE Workshop, October 5.
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7 Quality Control
Quality Control Overview
Quality Control (QC) is a laboratory endeavor aimed at ensuring that the highest-quality biotechnology products are manufactured and released to users. QC tests do not just happen; they are designed to meet certain objec- tives and quality criteria. Indeed, their design follows the principles of Quality by design (Chapter 5), which promulgates the concepts of superior quality testing programs and is based on an understanding of the product and the analytical tools used to test it. This chapter reviews the principles of QC planning, describes the life cycle of QC test and product specification development, identifies analytical methods most often applied to biophar- maceutical development, discusses the qualification and validation of these methods, and mentions the application of QC tests for product release and stability.
In contrast to the management and administrative nature of Quality Assurance (QA) (Chapter 5), QC is a technical or laboratory endeavor, which uses analytical methods to achieve specific objectives in a biotechnology operation. In the past, the terms Quality Assurance and Quality Control were used interchangeably, particularly by regulatory agencies, and this was con- fusing. Today, FDA still defines the term Quality Control as a largely adminis- trative function, which is not the same as that applied in most biotechnology operations. In this book, we define Quality Control as a laboratory or testing function and Quality Assurance as a quality management and administrative function (Chapter 5). Further, and as with any endeavor in biotechnology, QC has developed a trade or technical jargon, so commonly used terms are defined in this chapter and in the glossary.
The technical objective of QC is to apply laboratory testing to measure the quality of materials, be they raw or in-process materials and bulk substance (BS) or final product (FP), and whether the purpose of testing is at release or over time for stability evaluation. Quality control planning involves several steps, outlined in Chapter 1 and described in detail throughout this chapter. Quality control planning leads to important documents, the Certificate of Analysis (CoA) and the stability protocol. These are data-reporting formats
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that identify product attributes, test methods, specifications, and, after the completion of testing, the results.
There is a second dimension to QC planning, that is, consideration of the biopharmaceutical QC development cycle, as shown in Figure 7.1. The cycle identifies various QC functions in which processes are progressed, sequen- tially, in many small steps, or generations, and in close coordination with bio- manufacturing, clinical studies, and nonclinical testing. Each step in this cycle is considered one generation in the life cycle of a given test method and involves the test, the specification, and the product that is tested. Presumably, each gen- eration of test and specification is a slight improvement over the past, and with each step, there is a greater understanding of how test, specification, and over- all product quality relate to each other for a given biotechnology product.
Quality control planning is complex in part because a biopharmaceutical product requires many QC tests, each measuring an attribute. It is typical for an investigational product to have 10 or more QC tests and for a marketed product to have more than 20 tests. Together, the tests developed to support one product are complementary, that is, one test adds to knowledge gained from the others. Second, ongoing nonclinical and clinical studies often validate the usefulness
Begin: product attribute
Analytical requirement
Test design
Hypothetical specification (S-1)
Final test and specifications
Product (P-1)
Final specification
Refine specification (S-2)
Refine specification (S-3)
Develop test (T-1)
Product (P-3)
Redevelop test (T-2)
Redevelop test (T-3)
Product (P-2)
Validate tests against specifications
Test results (R-2)
Test results (R-3)
Test results (R-1)
Final test procedure
Phase 2 clinical
Nonclinical and phase 1 clinical
Phase 3 clinical
Phase 3 Clinical
FIGURE 7.1 Quality control cycle. The quality control test life cycle begins with an understanding of a product’s attributes and the analytical requirement. An appropriate test is then chosen or designed, and the development cycle then applies specifications (S), test development (T), input of manufactured product (P), bulk substance or final product on which to perform the assay, and results (R) from tests at that development step. It is a multistep process (e.g., S-1, S-2, S-3, and final specification).
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or validity of a test by demonstrating biological activity of that product in ani- mals or man. At some point in QC development, perhaps in the mid phase, some tests are qualified, and later in development, all of them are validated. These processes add considerably to confidence in using the assays.
Finally, QC testing is required for samples of BS and FP at release, for prod- uct on stability protocol, and at several points during manufacture (in-pro- cess testing). Multiply the number of QC tests with the number of times each QC test is performed and again with the number of samples per test, and it is clear why QC is a critical yet time-consuming function in biopharmaceutical product development.
Quality control test development begins very early in biopharmaceutical development, because all operations, especially biomanufacturing, require excellent analytical support. Failure to have available analytical methods often delays other efforts such as clinical and nonclinical studies and bio- manufacturing process development. Time and again, this point is proven, as biotechnology firms ignore the need to plan and develop QC technologies and soon find that they are unable to evaluate product quality, and subse- quently, the development program stalls.
We now describe each step in developing a panel of tests, using QC of BS as the first example and then reviewing this process as it is applies to FP and to stability testing.
Definition of Product Attributes
The QC development cycle begins with a Quality Control Plan, drafted only after there is an understanding of the intended product, a treatment indica- tion, and at least an early or research version of the candidate biopharma- ceutical (Chapter 1). In addition, some experimental work must precede QC planning, as it is not possible to develop a test for a product that has not been at least slightly characterized in the research laboratory. For example, if it is a protein, the primary, secondary, and tertiary structure, as well as the molecular, cellular, or immunological basis for its activity, should be known. Quality control planning begins once one understands how the product will eventually be used, the intended treatment indication, and the mechanism of action the product must have to achieve a desired endpoint.
An example product will be applied throughout this chapter to illus- trate the principles of biotechnology QC. Consider a recombinant protein (r-protein) product, a biopharmaceutical that functions in man by neutral- izing an undesirable molecule, perhaps a complex carbohydrate, located within a diseased cell. The therapeutic r-protein has the potential to ame- liorate a disease. However, this r-protein must first enter the cell via a receptor on the cell surface. Further, it is known that r-protein binding to the cell receptor must be of that magnitude that triggers the cell to inter- nalize the r-protein. In addition, the r-protein must survive within the cell and bind to the target molecule, an undesirable carbohydrate associated
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with cell death and disease. Binding leads to functional elimination of the carbohydrate molecules from the cell. Further, to enter the cell, the r-protein must be an intact molecule with proper primary and secondary structure, that is, it should not be degraded or unfolded.
With such knowledge of a product and it’s mechanism of action, the QC planner begins by developing a list of attributes for the product. An attribute is simply a desirable or necessary characteristic that a must possess to be safe or effective. In biopharmaceutical development, the most commonly applied attributes are appearance, identity, strength, purity, and potency. Returning to our example, several attributes of the r-protein—binding to a cell recep- tor, internalization into the cell, survival within the cell, and binding to the target molecule—are easily identified. Each of these attributes is listed in a typical CoA, and some appear several times, as they are measured with dif- ferent tests. The CoA, to be described later, is a formal document that lists each attribute, QC test, specification, and the test result.
Further to our example, the active ingredient in the example product is the r- protein. It is also referred to in testing parlance as the analyte or test sub- strate, the material we wish to measure in whatever way. The r-protein also possesses attributes. When in solution, it has a distinct appearance. Our r-protein has an identity, just like every human has a distinct fingerprint. In a given configuration, such as a vial of FP, the r-protein has a particular strength or level of active ingredient (e.g., a concentration). It also has purity, and, as with most products, impurities accompany the r-protein, even if pres- ent in trace amounts. Finally, like all biopharmaceuticals, the r-protein has a biological potency, in that it affects a biological system. A biopharmaceutical may possess other attributes, but this list is adequate to begin planning QC tests for most biotechnology products.
Analytical Methods to Measure Attributes
The heart of QC testing is developing and applying tests that measure a product’s attributes. Quality control scientists map a strategy, matching tests to particular attributes and sometimes applying multiple tests for each attri- bute. Further, they determine whether an attribute requires, or deserves, a qualitative or quantitative measurement. This necessitates an understanding of what makes an assay a good means of measuring an attribute.
Nonetheless, a perfect match between a method and an attribute is often not possible. In such cases, QC scientists adapt a given analytical method to suit the exact nature and intended use or indication of their product. Fortunately, most analytical tools are quite adaptable. Even then, some attributes for a given product cannot be measured with off-the-shelf analytical methods or even with available but slightly modified tests. Here, QC scientists must be creative and design a new or unique analytical tool, right from the begin- ning… This is often the case for potency tests, which measure complex bio- logical functions such as an immune or cellular physiological response.
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Traits of Analytical Methods
Each assay has one or more unique traits, somewhat like a facial feature, that distinguish the assay from other analytical tests and make it attractive for a particular application. Some tests have traits of qualitative analysis, others of quantitative analysis, and a few possess both. Tests are selected for their traits, a description of what they can really do for us. The QC scientist must have a pool of analytical methods available, at least in theory, once he or she begins to match a test to measurement of each attribute. Fortunately, a num- ber of analytical tools, each with its own peculiar trait or traits, are available for testing common attributes of many biopharmaceuticals.
One trait of any test is system suitability, which means that the chosen ana- lytical method must be appropriate in all ways for the intended purpose and measurement. Specificity is the second trait, and it means that a test has the ability to measure the intended product and nothing else that might be in the test material. Precision is the closeness of agreement between several mea- surements, much like precision in shooting an arrow means coming close to one point on the target with several arrows. The trait of linearity is applied when the assay must generate a linear curve. Linearity means that the results are directly proportional to the concentration of the analyte. Range, closely related to linearity, is the interval between the upper and lower concentration of analyte in the linear part of the curve. Limit of detection (LOD) is to under- stand how little of the analyte can be reliably detected in a sample. Limit of quantitation (LOQ) defines the lowest amount of analyte that can be quanti- tatively measured and not just simply detected. The trait of robustness car- ries many related meanings, but overall, it means that a test is reliable with respect to normal or expected variations in the analytical or testing environ- ment. For example, if three operators obtain the same result after each of them performs the test on three different dates, then an assay is robust and, one would think, reliable. Each trait will be discussed further in this chap- ter. Analytical tools are chosen to measure an attribute only if they possess traits that allow them to complete a stated measurement. Hence, possession of proper traits is a criterion for choosing the proper analytical tool. To para- phrase a saying of automobile mechanics, you must select the right analytical tool for each job.
Drafting a Certificate of Analysis (Bulk Substance)
As attributes and methods are identified, they are listed in tabular form on a draft document referred to as the CoA, as shown in Table 7.1 for BS and in Table 7.2 for FP. Each attribute, further defined below, are listed in the first column of the CoA. More than one test may be applied to an attribute, as each test measures a different parameter of that attribute. This discus- sion and the accompanying tables use examples of commonly applied QC or test methods, described later in this chapter, and reflect the analysis of our r-protein example product.
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TABLE 7.1
Certificate of Analysis for a Biopharmaceutical Bulk Substance (r-Protein Example)
Attribute Analytical Method Reference to
Method Specification Result
Appearance Visual inspection of BDS in clear glass tube
SOP# QC01 Liquid, opaque, off-white to straw color, no particulates or aggregates
Safety Microbial enumeration test
USP <61> <1 cfu/mL
Identity N-terminal sequence SOP# H411B Confirm known sequence Identity SDS-PAGE SOP# QC-02 Single band at 41 kDa Safety Bacterial endotoxin (gel-
clot LAL test) USP <85> <1.0 EU/mg protein
Purity SDS-PAGE SOP# QC-03 Single band, comparable to reference standard
Purity HPLC SOP# QC-04 Single peak integrated ≥98% of material in sample
Purity Aggregates by size exclusion chromatography
SOP# QC-05 ≤2% of material is aggregate
Purity Peptide map SOP# QC-06 Map equivalent to reference standard
Purity Host cell protein, ELISA SOP# QC-118 <0.1 mg host cell protein/ mg total protein
Purity Host cell DNA, fluorescence probe
SOP# QC-120 <10 µg host cell DNA/1 mg total protein
Strength Total protein by BCA SOP# QC-07 1.0 ± 0.1 mL containing 2.0 ± 0.1 mg/mL protein
Purity Silicon lubricant by atomic absorption
Contract laboratory SOP# X147–1
<10 ng silicon/mg total protein
Purity Aggregated protein
Aggregated protein by light scatter
SOP# QC-08 <7 µg aggregated protein/1 mg total protein
Potency Receptor binding SOP# QC-111 0.6–1.05 µg r-protein/1.0 µg receptor
Potency Viability of cultured cells at 1, 2, 4, 6, and 12 h
SOP# QC-09 and SOP# QC-10
>70% viability versus time 0 at each time point
Potency Accumulation of molecule in cultured cells at 24 h
SOP QC# 11 and SOP# QC-12
<10% accumulation over baseline, time 0
Prepared by/Date:___________ Approved by Quality Control/Date:__________ Approved by Quality Assurance /Date:______________
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TABLE 7.2
Certificate of Analysis for a Biopharmaceutical Final Product (r-Protein Example)
Attribute Analytical Method Reference to
Method Specification Result
Appearance Visual inspection of FDP in final container
SOP# QC-21 Clear, colorless liquid without particulates or aggregates
Safety Endotoxin gel-clot LAL
USP <85> ≤10 EU/1 mL dose
Safety Sterility, compendial USP <71> 21 CFR 610.12
Sterile
Safety Osmolality by osmometer
SOP# QC-26 200 ± 10 mOs/kg
Safety pH by pH meter microprobe
SOP# QC-27 7.1 ± 0.2
Identity N-terminal sequence SOP# H411 Confirm known sequence
Identity SDS-PAGE SOP# QC-22 Single band at 41 kDa Purity SDS-PAGE SOP# QC-23 Single band at
30 kDa, comparable to reference standard
Purity HPLC SOP# QC-24 Single peak integrated >98% material in sample
Purity Excipient glycerol atomic absorption
SOP# 11–4422C 1 ± 0.1 μg/mL
Purity Excipient, human serum albumin by ELISA
SOP# 11–2244C contractor
200 ± 20 μg/mL
Strength Total protein by BCA SOP# QC-25 1.0 ± 0.1 mg/1 mL dose and per vial
Purity Aggregated protein by light scatter
SOP# QC-28 <1 µg aggregated protein/1 mg total protein
Potency Viability of cultured cells at 1, 2, 4, 6, and 12 h
SOP# QC-09 and SOP# QC-10
>70% receptors versus time 0 at each time point
Potency Accumulation of molecule in cultured cells at 24 h
SOP# QC-11 and SOP# QC-12
≤10% accumulation over baseline time 0
Potency Reduction of disease; transgenic mouse model
SOP# QC-23, SOP# QC-24, and SOP# QC-25
>50% reduction as compared to control
Prepared by/Date:___________ Approved by Quality Control/Date__________ Approved by Quality Assurance /Date:______________
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• Appearance: Most products have a distinct appearance to the eye. Bad product often looks bad. Under appearance, traits may be further defined as color, clarity or opaqueness, or presence or absence of particulates or aggregates.
• Identity: This trait simply ensures that the product is what we believe it to be, and what we have labeled it as, and not something else. Each biotechnology product is unique and possesses fingerprints that can be analyzed.
• Safety: A safety test cannot by itself tell us whether a product is safe or not, but it can provide some assurance that it is not overtly toxic or otherwise lethal or overtly harmful to the user. In addition, multiple safety tests, each examining a specific aspect of the attribute, can additively increase the chances that a product is, in fact, safe.
• Purity and impurities: All biotechnology products are purported to be pure, that is, to have only those molecules or cells or other active ingredients they are supposed to contain. Purity is a measure of that product claim, and impurity testing informs us as to the nature and amount of anything else in the product vial.
• Strength: It is important that each product have enough of the active ingredient, so that it has the potential to cause the intended effect. If we say that there is 25 mg/mL of product in 1 mL per vial, then there should be 25 mg/mL and 1 mL in each vial.
• Potency: This is the most challenging trait to measure. It tells us that the product, in fact, has the intended biological effect.
This is the first step in drafting a CoA. The next step, as shown in column 2 of Tables 7.1 and 7.2, is to identify analytical methods to measure each of these attributes.
Selection of Analytical Methods
This section provides information on selecting a test to measure each attri- bute; by way of example, it identifies a few assays commonly applied to recombinant protein biopharmaceuticals in BS (column 2, Table 7.1). Selection of tests for FP is discussed in a later section, and a list of assays, with techni- cal descriptions, is presented there and in Table 7.2.
Laboratory tests used in QC, also referred to as analytical methods, or just methods, can be classified in several ways. Tests described in a compen- dium (e.g., a pharmacopeia) are nationally or internationally recognized and are performed in a very standard manner, no matter what the product. Tests are matched to attributes in the example CoA, and in column 3 of Tables 7.1 and 7.2, reference is made to the specific manner in which each test is performed on BS.
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A compendial test, such as that for sterility, microbial limits, or endo- toxin, is applied to a wide range of biological and pharmaceutical products. Examples of compendia, technical handbooks, or encyclopedia are given in Box 7.1. An outline and contents of the United States Pharmacopeia (USP 2015), a commonly used compendium and QC reference, are given in Box 7.2,
BOX 7.1 EXAMPLES OF COMPENDIA AND REFERENCE TEXTS FOR QUALITY CONTROL
• United States Pharmacopeia (USP): The official pharmacopeia for the United States, is published by the U.S. Pharmacopeial Convention (http://www.usp.org). The USP, along with a sister publication The National Formulary, or USP-NF provides test methods, standards, and references for analysis of medicines, reagents, and other materials. If an analytical method and a standard are applicable to a biopharmaceutical and they are approved and published in the USP, then it is very likely that FDA will demand that product testing meets this standard. Specifications may or may not be recommended by USP.
• European Pharmacopoeia (EP): The official pharmacopeia for the European Union, developed by the European Directorate for the Quality of Medicines (http://www.edqm.eu).
• British Pharmacopoeia (2016): The official pharmacopeia for Great Britain (http://www.pharmacopoeia.com).
• Merck Index (2011): This encyclopedia provides precise and comprehensive information on chemicals, drugs, and biologi- cals written as monographs and carefully indexed. It is used in biopharmaceutical development for materials and products that are well characterized and may be predicate or compara- tors for a novel compound in early development.
• Merck Manual (of Diagnosis and Therapy) (2011): A medical ency- clopedia, organized by disorders or diseases of various sys- tems or organs. It explains the symptoms or diseases, their diagnosis, and their treatment. It is a leading medical reference.
• Martindale’s: The Complete Drug Reference (2015): A very com- plete reference book that provides monographs, albeit brief, on thousands of drugs and biologicals with reference citations and manufacturers. It is carefully indexed.
• Physician’s Desk Reference (2015): A collection of the product labeling of the most commonly used drugs and biologicals in the United States. It has information on drug indications, dos- ages, side effects, and detailed instructions for use.
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BOX 7.2 AN OUTLINE OF THE UNITED STATES PHARMACOPEIA AND EXAMPLES OF ITS SECTIONS
RELATED TO BIOPHARMACEUTICALS OR DRUGS
• General Information: Provides guidance on a variety of product classes spanning the spectrum of drug and biopharmaceuti- cal development. Examples are general guidelines for oph- thalmic preparations and water for pharmaceutical purposes. Section <1231> identifies classes (qualities) of waters and their standards.
• Reagents: All types of reagents and their quality standards are described in this section. Examples are acetic acid (diluted), used in various test procedures, and pancreatic digest of casein, used in culture media.
• National Formulary (NF): Here are the recipes for making a host of products that are used in or as drugs, including many over- the- counter preparations. An example is the procedure for pre- paring the peppermint solution added to certain oral drugs.
• Official Monographs: In this section of the USP are monographs on commonly used drugs; many drugs described here are long- standing and generic. An example is aspirin delayed-release tablets. Others are products used largely in medical treatment facilities, such as lactated ringers and dextrose solution.
• General Tests: Many tests that are used in drug and biopharma- ceutical quality control are found under this heading. Certain tests used for biotechnology products are described in great detail. Examples are as follows: • <621> Chromatography: Gas, Paper and Column • <85> Bacterial Endotoxin Tests • <71> Sterility Tests • <61> Microbial Limits Tests
• General Tests also covers information that provides guidance and procedures for biopharmaceutical testing and assay devel- opment, in general. Examples are as follows: • <111> Design and Analysis of Biological Assays • <1041> Biologics • <1045> Biotechnology-Derived Articles • <1046> Cell and Gene Therapy Products
(Continued)
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along with the titles of some compendial tests commonly used to measure attributes of biopharmaceuticals. Methods of compendial tests cannot be easily modified, but sometimes they are adapted for novel applications.
Another group of analytical methods, often not found in a pharmaco- peia and referred to here as generic tests, is used to measure the attributes of identity, purity, strength, and, sometimes, potency of biotechnology products. Although they are not compendial, there may be industry or regulatory precedence, procedures, or even quality standards for their per- formance. This depends on the nature and history of the product. These QC tests can be established in most laboratories, or if they require expen- sive instrumentation, they can be performed by a contract research orga- nization (CRO). Most are readily adaptable to a variety of products, and in many cases, their methods can be changed to suit specific purposes. A biologics sterility test, described in FDA regulations (21 Code of Federal Regulations [CFR] 610.12), or visual appearance of a product would be con- sidered generic tests. It is worth noting that even some commonly used tests (e.g., high-pressure liquid chromatography [HPLC] or polyacrylamide gel electrophoresis [PAGE]) are performed in a very specific manner for each analyte, and, at most, published guidelines, such as in USP or by FDA, are general in nature.
A third group of tests includes those developed for one product or a few closely related products. These methods (some examples are given later in this chapter) often originate in a research laboratory and are further developed, adapted, and refined by the QC laboratory for use as a QC test to measure an attribute of a specific biological product. Biological potency assays often fall under this group of tests.
Quality control assays are also classified in yet another way, that is, by their intended use or application. These include tests for raw materials, for
BOX 7.2 (Continued) AN OUTLINE OF THE UNITED STATES PHARMACOPEIA AND EXAMPLES OF ITS SECTIONS
RELATED TO BIOPHARMACEUTICALS OR DRUGS
• <1047> Biotechnology-Derived Articles—Tests • <1048> Quality of Biotechnological Products: Analysis of
the Expression Construct in Cells Used for Production of r-DNA-Derived Protein Products
• <1049> Quality of Biotechnological Products: Stability Testing of Biotechnological/Biological Products
• <1050> Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin.
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in-process testing, for drug substance, for drug product, or for stability test- ing of any material, substance, or product. Hence, a given method might be applied at one or more points in the product manufacturing cycle.
As noted earlier, another means of classifying QC tests is by their intended outcome or application. Examples of test applications are appearance or description, identity, purity, impurities, potency, quality, and special tests. It is not unusual for a very adaptable test method to be used to measure two attributes. For example, one test may be used to measure both identity and purity. A test is also classified according to the method’s enabling technol- ogy, such as identification of bacteria and yeast, pH measurement, HPLC chromatography, peptide mapping, or receptor binding.
The information entered into the third column of the CoA (Figure 7.1), references by name the test, or procedure used. For compendial tests, ref- erence is made to a section of a monograph to describe the sterility test (e.g., USP <71>). For noncompendial tests, standard operating procedures (SOPs) are identified in this column, citing the number of the SOP. If a CRO performs a test, then that laboratory and the SOP used by it are identified under the test method column of the CoA.
To summarize, a key element of QC planning and performance is match- ing the proper test to an attribute, and this requires knowing which tests are available to the analyst and understanding how the attribute relates to each available test. Only then can a meaningful panel of tests be selected for the product. We return to our example in an attempt to better explain the matching process of attribute-to-test method. First is the identity test. Identity tests reveal whether or not a product is, in fact, the intended mate- rial, that is, the fingerprint of a biopharmaceutical. For the r-protein exam- ple, we know from research that it is a globular protein of known molecular weight and a defined sequence. How should we ensure that the material we manufactured is, in fact, that r-protein? A common approach is to sequence, by amino acid determination, from the N-terminus of the r-protein until about the tenth amino acid. It is highly unlikely that another protein would have the same 10 amino acids in that order at the N-terminus. One might also identify the isoelectric point of a protein by performing electropho- resis at various conditions, for example, various pH and ionic strengths. Another approach to demonstrating identity is to measure the molecular weight of the molecule under reducing or nonreducing conditions by gel electrophoresis. Although not as definitive as N-terminus sequencing, this test differentiates the analyte from many other proteins. Gel electrophore- sis is made more powerful as an identity test if, after the electrophoresis step, the protein is blotted to an inert but absorbent membrane and then probed with an antibody specific for the product. This is known as Western blot test.
A sample of reference standard, that is, a protein known to be the desired protein, is always tested in parallel with a test sample. If results of the test sample match theoretical or expected values and the results obtained from
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testing match the reference standard, then there is a high probability that the test sample is the intended molecule.
The strength of a preparation is a general measure of how much of the desired active ingredient is in the product. For the r-protein, strength might be reflected in the total amount of protein, as long as the vast major- ity of protein in the sample is, in fact, the r-protein. How do we ensure this is the case? First, we perform identity testing on that sample to know whether the molecule in the sample is the intended r-protein. Second, and as described below, we measure the purity of the molecule in a sample of product. Back to the concept of strength, for a recombinant protein, this may be measured by a total protein assay, such as bicinchoninic acid (BCA) assay, or by ultraviolet absorbance at a specific wavelength in a spectrophotometer. Then, this measurement, given in milligrams per mil- liliter, is multiplied by the percentage purity and total volume in millili- ters to give the total amount, in milligrams, of the desired protein. This paradigm demonstrates the interdependence of various QC tests and the need to interpret the result any one assay may give in relation to the result from another assay.
Several test methods may be applied to measure the purity of any product. Further, major impurities are characterized. Methods are developed to mea- sure the level of product purity, that is, the percentage or the actual amount of the stated product and the percentage or the amount of all impurities and contaminants. Other methods are then used to identify specific major impurities or contaminants to include even tiny amounts of potentially toxic or otherwise undesirable substances that might have entered into the production stream and remained in the product. The development of test- ing schemes and the selection of tests is based on an in-depth understand- ing of the raw materials, equipment, and processes used in manufacturing, as well as of the scope of possible impurities or contaminants. Impurities and contaminants are further discussed in relation to biomanufacturing (Chapter 6).
Returning to our example of r-protein, we consider the purity of that mol- ecule as well as impurities that might exist with the r-protein as BS. Here, r-protein purity is measured by sodium dodecyl sulfate PAGE (SDS-PAGE), an assay also used as an identity test. However, this assay is, at best, only semiquantitative in that it cannot accurately measure the amount of the r-pro- tein or the amount of impurities in the sample of BS. It may, however, give a reasonable estimate of purity. The SDS-PAGE test is often used in-process to follow the progress as molecules are purified across multiple bioprocess- ing steps (Chapter 6). A more exact method is applied to the measurement of r-protein purity by using an analytical chromatographic technique; HPLC is a common choice. Here, the example r-protein should appear on the chro- matogram as an independent, major peak, whereas impurities might appear as smaller side peaks or shoulders of the major peak. Further, we might use size exclusion chromatography (SEC) to show that the molecule in our
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preparations is not aggregated. This test demonstrates that the r-protein has maintained a native form and that it has not otherwise distorted through clumping. Yet another purity test, peptide mapping, may reveal that molecu- lar integrity is still present. Protein aggregates are measured by light disper- sion, if their presence is suspected. Special tests such as mass spectroscopy, nuclear magnetic resonance, and capillary electrophoresis may be consid- ered. Carbohydrate analysis may be applied if there is a need to examine cer- tain posttranslational modifications. Today, many new methods are applied to molecular characterization of certain products and some are noted later in this chapter. Since purity is critically important to molecular integrity and function and because impurities must be characterized and measured to prevent them from causing undesirable reactions in the consumer and from increasing in amount with later production, several purity and impurity tests are typically applied to a product, both as BS and as FP.
Detection of contaminants presents a different challenge because these can enter the stream from so many sources, especially if a failure goes undetected during processing. For example, most proteins are filtered at some point during purification and filters may fracture and release fibers, particles, or fragments into the product stream, thus contaminating the product. Contaminants, particulate and soluble, may enter the product stream from raw materials or virtually any substance that contacts the product stream. Given that one cannot test for every possible material that might contaminate a biotechnology manufacturing process, what should the QC scientist consider as possible contaminants for any given product? Anything that might be toxic or otherwise dangerous to the user in small amounts and is a part of the process comes first to mind. For example, if there is a possibility of bacterial growth, then endotoxin testing, described below, is critical. If there is a piece of biomanufacturing equipment that is essential but is known to sometimes shed particulates of silicon lubricant into the product stream, then it might be wise to test for silicon lubricant. Both endotoxin and silicon lubricant are considered in the example CoA for BS, as shown in Table 7.1. Clearly, an effective yet affordable contami- nant testing program involves discussion between manufacturing and control staff, with decisions based on full understanding of the production processes and the intended use.
Potency assays are critical to a QC testing scheme because they are used to predict whether the product will function as it was designed to func- tion. It would be futile to produce any biopharmaceutical product and test it for purity, identity, and safety and still not know if it could function as intended. Unfortunately, this is far too often the case with biopharma- ceutical development programs. For testing BS, potency assays are often a surrogate assay, meaning that it does not directly measures the biological function in a complex system such as a whole animal but instead measures a physiological attribute of the product in an in vitro or a cell-based assay. Surrogates are used in all aspects of biotechnology development, but any
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surrogate measure must be appropriate, well designed, and, eventually, validated against the intended use. The QC scientist developing a surro- gate assay must be knowledgeable about both the product and the thera- peutic indication, particularly the mechanism of action and the biology and molecular biology involved in the product’s therapeutic effect. Using this knowledge, scientists become inventive, even crafty, in finding analyti- cal methods that predict potency (or lack thereof) while keeping those tests as simple, inexpensive, and practical as possible. The best source of new potency tests is the research laboratory.
Returning to the example of the r-protein and examining Table 7.1 further, we see that two potency assays were developed for this BS. One measures specific binding of the r-protein to its cell-surface receptor. The rationale is that receptor binding is a critical step in the pathway to molecular activity and a biologically active r-protein must bind to that receptor. For this example, the receptor was identified and the gene was cloned in a research laboratory, so that it is now produced in small amounts (enough for testing purposes). Using analytical instruments, QC scientists next develop an in vitro assay that measures the amount of r-protein that binds to a given amount of the receptor. For example, between 0.6 µg and 1.0 µg of the recombinant prod- uct binds to 1.0 µg of the receptor. On repeatedly testing three batches of r-protein with the newly developed assay, scientists determine that between 0.75 µg and 0.92 µg of the product binds to the receptor in this test. This test is chosen as one of the two potency assays for r-protein in the BS.
For an attribute as important as potency, one always considers two or more complementary assays. This is because a single assay measures only one aspect of a product’s potency attribute. In the r-protein example, the QC scientist chooses a second assay, also developed in the research laboratory, to measure the degree to which r-protein inhibits the buildup of the unde- sirable molecule within the target cell. This is a relevant surrogate to the intended biological response because it measures the ultimate activity asso- ciated with therapeutic value, at least at the molecular and cellular levels. Shown in Table 7.1 as the last assay on the CoA, the measurement is an in vitro assay, which is likely rapid and inexpensive but hopefully very precise and sensitive to product activity.
This section of provided an overview of the early test selection, basing each assay on a product attribute. After this process has completed, the next step in the planning process is consideration of specifications. Later in this chapter, we discuss in greater detail analytical tests and their application to the QC of biopharmaceutical products.
Development of Specifications
Identifying a test to measure each attribute is important, but it is also critical to know whether each batch or lot passed or failed. This decision is based on the results obtained when the test is applied to a particular batch of product.
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The word specification carries great meaning to both QC scientists and bio- pharmaceutical development projects. A specification is a descriptor, numer- ical or verbal, that a product must achieve to be considered suitable for use. It also serves as a requirement or condition, the basis upon which a product is accepted or rejected. Specifications may be established by regulation, by precedence and proven value and capability, by outside guidance, or by a product development team. Often, a specification is quantitative, stated as a range of values (e.g., ≤12.5 units or 1.0–3.0 mg/mL or 2.0 ± 1.0 mg/mL), but it may be qualitative, a term that compares the specification to a reference, such as comparable to values of reference standard #0017, or it can be purely a descriptor such as clear, colorless solution free of particulate matter. Examples of specifications for BS are shown in the fourth column of Table 7.1.
During the development process, that is, before submitting a marketing application, many specifications may be considered interim or temporary. However, specifications codified (e.g., sterility test in 21 CFR) or established by precedence or compendium (e.g., sterility test in USP) are less flexible and are typically inviolable during the development life cycle. Specifications are taken quite seriously by both regulatory authorities and the sponsor; indeed, final or ultimate specifications established through the validation process in Phase 3 guide the release of a marketed biotechnology product in the years to come. Specifications for marketed products may be changed, but this is done only with scientific evidence to support the adjustment and follows strict change control rules. Hence, there is a great need to carefully choose and then fully develop a specification during the development process, bas- ing interim and final decisions on experimental data generated by testing multiple lots of the manufactured product.
Advances in analytical technologies for biopharmaceutical products have increased the number of tests used on any product. Regulatory authorities are quick to suggest yet another test that might ensure safety or better pre- dict efficacy of a product. In addition, specifications themselves have become more complex, quantitative, and sensitive. Indeed, the role and importance of the QC function to biotechnology development has grown considerably over the past 30 years.
Establishment of specifications for purity or impurities is challenging for biopharmaceutical product development teams, as the questions raised have no simple answers, especially because the data are limited or do not exist at the time. What quantitative limits are acceptable for purity and impurities, and what is allowed and how much? The correct answer must be based on highly regarded existing scientific data, and, for a unique product, it must be experimentally determined for each product. Thus, a final purity specifica- tion is not finally established until late-phase development, after much data have been generated. For many products, there are at the outset of testing no existing guidelines for purity specifications or impurity characteristics. At first, the sponsor must assume that a product will not be 100% pure and that impurities will exist in both the BS and the FP. Upstream manufacturing
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generates, and downstream processing concentrates, certain impurities and contaminants; no product is expected to reach 100% purity by using current purification technologies. Upstream production is a dirty process, whereas downstream processing, notably chromatography, may introduce and con- centrate novel, yet undesirable, impurities and contaminants, even while removing other impurities or contaminants and concentrating the product. Contaminants enter the stream out of necessity, because they are inherent to a required process, a necessary evil. Examples of contaminants are endo- toxin, in some systems shed by the very recombinant bacteria making the product; chromatography gels or matrices; particles from vessels and tubing; and chemicals leached into the product stream from various contacts and surfaces. Impurities are the molecules that are derived from the ingredients used to make the product but are not wanted in the BS or FP. These include cellular debris and molecules derived from the cells in which the recom- binant protein or tissue was produced and the components of the nutrient medium that fed those cells. Impurities are also seen as breakdown prod- ucts of the desirable biomolecules and include improperly folded protein, shortened versions of the protein, posttranslational product variants, and fragments or aggregates of the protein.
As noted before, a rule of thumb for establishing purity and impurity specifications in biotechnology is that the BS or FP specification should be that the product is at least 95% pure. However, this rule does not apply to every biopharmaceutical, and specifications for purity must be developed based on the intended use and attributes of the product and the nature of the impurities. For example, a protein that will be used to enrich cattle feed might be fine at 75% purity, as long as none of the impurities were toxic to cattle or man and consisted largely of protein fragments and aggregates. A biopharmaceutical intended for injection at large doses in human patients with serious disease might need to be well more than 99% pure and com- pletely free of any toxin or foreign or aggregated protein. Here again, careful planning is required to ensure that analytical methods and speci- fications developed for purity and impurities match exactly the intended use and other attributes, for example, safety, of a product. Impurities such as a virus or lethal toxin are simply not allowed in a biopharmaceutical. However, if such materials could possibly have been introduced, meth- ods must be designed to ensure their removal and highly sensitive and specific tests should be introduced to ensure that product is free of such substances. Here specifications are very stringent. The guidelines for set- ting specifications for other impurities or contaminants are established based on prior experience, on the probability that they do in fact exist, on the availability of tests to identify or measure them, and, mostly, on common sense and good scientific practice. As noted in Chapter 6, speci- fications for levels of impurities or contaminants are, in the end, often negotiated between the sponsor and the regulatory agency. This brings up a final point on the subject of setting specifications for impurities, both
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qualitative and quantitative. The views of national populations and their regulatory agencies vary greatly on the perceived risk of certain impuri- ties to the user, and the biotechnology firm should consider all market- places and national or international guidance, not just the United States, when establishing specifications for a product.
There are other outcomes to the processes of establishing and applying test methods and specifications in this cycle. Sometimes, a well-considered ana- lytical method fails miserably and is not predictive of the attribute or is oth- erwise unable to predict product quality. There is no need to consider further refinement of the test or of the specification. In other instances, the hypotheti- cal value established by QC scientists is not at all close to the experimental values. In such cases, the assay may be further studied or it may be reworked to either explain the differences or to optimize the method. More often than not, however, a well-considered analytical method is meaningful to measur- ing product quality, with only the need to adjust the specification, and thus ensures consistent quality for future batches or lots of product.
Some QC tests have, in the eyes of regulatory authorities, absolute specifi- cation requirements. As noted before, sterility tests are performed by com- pendial methods and they must meet standards published in a compendium or by regulatory agencies. There is, in the eyes of regulators and the market- place, but one definition of sterility, and adjustment of the sterility specifica- tion is simply not acceptable for a biopharmaceutical such as the r-protein mentioned in the earlier example. Other historical tests, even certain com- pendial methods, may allow specification variance, but this depends on the nature of the product and the risk to the user associated with a change in specification. Such changes must be negotiated with regulatory authorities. For example, the appearance and description for a parenteral biopharmaceu- tical, formulated as protein in a buffer, is expected to read clear solution with- out particulates. However, for some proteins at high concentrations in a buffer solution, it may be normal for the product to be cloudy or opaque. Hence, a highly concentrated protein solution may be allowed to deviate from the standard specification for most protein solutions and be considered accept- able if it has a specification for appearance of cloudy colorless solution without particulates or foreign matter.
Establishing specifications for contaminants and impurities is a chal- lenging task, because it is impossible to know in early development how an impurity might impact the safety or efficacy of a product. How does one evaluate the impact of minute amounts of a given contaminant, unless it is a known toxin? Establishing purity and impurity guidelines has led to long discussions within the international biopharmaceutical community. These discussions are based on the risk posed by certain impurities found in some products, formulation, dose, or patient population. An example relevant to biopharmaceutical protein preparations is the impact that protein aggregates might have on parental products. Years ago, it was felt that they had little impact on product safety or potency, unless they were present in significant
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amounts. Even then, a definition of significance was elusive. Recent evi- dence suggests but does not prove that protein aggregates, even in small amounts, may be immunogenic and potentially elicit an antibody response in a patient to a recombinant protein. If this is the case, the impact is prob- ably quite variable, depending on the route of delivery, amount given over the lifetime of the patient, exact nature of the protein and the aggregate, and the patient. Then how does one establish a specification for maximum allow- able amounts of aggregate for any given protein product? Panels of experts may be called to address such questions, but even then, recommendations tend to be fuzzy. Hence, challenges to planning product attributes, tests, and specifications continue through the life cycle of product development and also into marketing phases.
As with other attributes, specifications for potency of product in BS are established early in development, even when little experimental data are available. Potency tests, identified in the planning process, are created in a laboratory and then used to analyze some early batches or lots of BS or FP, respectively. Referring back to the development of the r-protein receptor- binding potency assay (Table 7.1), we know that in theory, 1.0 µg of the recom- binant product should bind to 1.0 µg receptor and also that laboratory testing revealed a range of binding activity, that is between 0.75 µg and 0.92 µg of pure r-protein binds to 1.0 µg receptor. At this time, the QC scientist must establish a specification for this potency assay based on both theoretical and derived experimental data but with the knowledge that this data might, by chance, reflect an incorrectly high or low estimate of the actual binding activity. The early or hypothetical specification, referred to as S-1 in the QC cycle drawing (Figure 7.1), is the following specification in this example: Range of protein binding, 0.60–1.05 µg of product per 1.0 µg of receptor (Table 7.1). After testing additional and subsequent batches of BS, the sponsor might discover that the range of values is too broad and, based on the data, narrow the range of acceptable values in the specification, perhaps to 0.80–0.90 µg protein per 1.0 µg of receptor (e.g., the refined specification, S-2, in Figure 7.1). Alternatively, with additional data, the original or S-1 specification could be a firm estimate, holding up to experimental results and remaining the same throughout the product development life cycle. This then is the accepted process, experimentally intensive but proven effective to establishing mean- ingful tests, test results, and specifications.
The other potency assay for BS used in our example (Table 7.1) measures another trait important for measuring the activity of the r-protein, the ability to halt buildup of a molecule inside cells. The specification was established in the same manner.
In summary, the early establishment and then later adjustment of a specifi- cation is normal part of QC testing in the overall product cycle. In early devel- opment, the process is a mix of scientific, iterative, and intuitive approaches and later, it becomes heavily scientific, driven by data from clinical, nonclini- cal, and laboratory studies.
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Entering Test Results
Results are added to the fifth and final column of the CoA (Table 7.1), after test- ing has been completed. Results are given in the same format as specifications to allow for comparison. For example, if the specification for range of protein binding is 0.60–1.05 μg recombinant protein bound per 1.0 µg of receptor, then the result should only be given as milligrams of protein bound per 1.0 µg of receptor. If the result was 0.73 μg, then this batch of BS would Pass by this standard. However, if the result was 0.55 μg, then it would Fail. The subject of handling a failure to meet a specification is discussed later in this chapter.
Certificate of Analysis for Drug Product
The process of planning QC test methods and specifications is also applied to developing testing plans for FP. Indeed, the process of potency test devel- opment is often more challenging with the final formulated product than it is with the BS.
Once BS has been tested and released, it is formulated, filled into a final container, such as a vial or syringe, and finally labeled and packaged (Chapter 6). Once finished, testing begins and results are included in a CoA for FP, a sample of which is shown in Table 7.2. This certificate, once signed and reviewed, becomes one of the several documents that support the release of FP to the user or, if product fails, the destruction of this material. The contents of the CoA for a lot of FP are important, and so, great care is taken during QC planning to choose the correct analytical tools and specifications. Again, the choices are based on the nature and attributes of, and the indica- tion for, the FP.
Quality control tests for FP may be more stringent than those for BS, and they are always focused on an attribute that is important to the intended use and the well-being of the user. In designing the tests that apply to a biophar- maceutical FP, we consider many aspects of quality. The appearance test is performed on a representative sample of FP when it is in the final container, filled and finished. The specification for appearance is designed to ensure that the QC examiner inspects a representative number of vials for certain attributes and to note the absence of undesirable and visible impurities or contaminants. For the r-protein example, we expect the FP to appear color- less and to not contain any aggregates or particulate matter. The appearance test illustrates an important point of QC testing: the SOP must be written in such a way that the operator or inspector examines for these attributes and properly identifies substandard FP. This general procedure, use of trained operators or technicians, adequate equipment and SOPs, and exact report- ing are applied to every test that is performed in a QC laboratory and to the results reported in a CoA.
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FP is subjected to several safety tests, because this is the most important attribute of any biopharmaceutical. Most biopharmaceuticals are given parentally, and hence, they must be sterile. They must also be free of or have very low levels of endotoxin or other toxic substances. Tests for specific types of undesirable contaminants or impurities are defined in a compendium or in regulations and are described elsewhere in this chapter. In addition, the general safety test is performed in the United States on most biopharmaceu- tical FPs to detect any highly toxic properties that the product might have. The CoA in Table 7.2 identifies one test for appearance and four methods for safety.
The active ingredient in FP must be exactly what it purports to be on the label and nothing else. Hence, identity testing is performed on FP. A variety of generic tests, such as SDS-PAGE, perhaps in conjunction with Western blot using a specific monoclonal antibody, HPLC, N-terminal sequencing, tryptic digest mapping, and cell karyotyping or phenotyping, may be employed in a panel of identity tests. In Table 7.2, the chosen identity tests for an FP, our example r-protein, are N-terminal sequencing and SDS-PAGE.
Despite the fact that purity has already been demonstrated and impurities identified at the BS stage, it is important to test FP for purity and impuri- ties. This is because impurities or contaminants might have been generated or introduced during formulation and fill, the processing steps from BS to FP. For example, some r-protein might degrade during processing or micro- scopic contaminants, such as endotoxin, might enter the FP if, for example, the containers were not scrupulously clean. In the CoA for FP (Table 7.2), two purity tests, SDS-PAGE and HPLC, are used to detect and identify (or measure) macromolecular impurities in FP.
Formation of aggregates in FP is a problem with formulations of certain biopharmaceuticals. They are considered impurities but can also impact the safety and potency of FP. Hence, an additional purity step for measuring protein aggregates is added to the CoA for the r-protein (Table 7.2).
The FP is also tested for concentrations of any excipients, materials that are added during processing and should exist in the FP. In the example (Table 7.2), both glycerol and human serum albumin were added to the for- mulation and the amounts of each are measured to ensure that they meet the specified concentrations.
Strength of FP is determined by an assay that measures the total amount of active substance. In the case of a recombinant protein, this might be a BCA assay to measure total protein (Table 7.2). A variety of analytical methods are available to measure all macromolecules in FP or count and measure the amount of cells or tissues in FP.
Other tests performed on FP measure attributes of the formulation that are important to product purity, potency, and stability. In the example (Table 7.2), osmolality and pH were measured to ensure that the salt concentration was correct in the formulation buffer. Maintaining pH is also important to main- tain the stability of most biopharmaceutical products.
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Development of relevant potency tests for FP challenges the design and subsequent execution of any QC plan; this requires considerable abstract thinking, laboratory testing, and interaction with scientific colleagues. FP potency tests must be meaningful and practical. A potency test that mea- sures a noncritical potency criterion is not very helpful, and any test that takes more than 60 days to complete and report is impractical. Consider also that a potency assay must stand as surrogate for the ultimate potency test: performance in many human users. There, probably, never was, and never will be, a single perfect potency assay, one that stands alone to pre- dict the biological efficacy of a FP. Therefore, sponsors seldom rely on a single potency assay but instead apply three or more potency assays, each of which may be imperfect. This does not always happen from the start of product development, but it should begin in the early phase, so that mul- tiple FP potency assays are available in mid- and late-phase development.
Another objective of potency testing is to learn whether FP possess attri- butes that result in optimal performance for the end user. This is difficult to achieve, because we often do not understand every biological factor that leads to optimal and consistent efficacy in all users. For biopharmaceutical development, this means, in theory, if not always in practice, that the potency test applied to FP mirrors exactly the potency when it is used in man. A single potency test seldom, if ever, achieves this objective, but application of multiple potency assays may support such conclusions.
Biological responses and biological molecules or cellular systems are com- plex. Application of a biopharmaceutical to a biological system-cultured cells, animal or human, is an attempt to disrupt or bring back a biological system to equilibrium. Indeed, biopharmaceutical treatment may further disrupt or complicate a biological system already out of control. As compared to small drug molecules, many biopharmaceuticals are complex biological entities. Given this information, consider how difficult it is to measure a product’s potency in a complex system. Hence, multiple potency assays provide a greater chance of ensuring product efficacy than does a single potency test, because several potency tests evaluate the impact of the product at multiple points in complex biological pathways. Although the use of a complete liv- ing organism (e.g., a whole animal) for FP potency testing brings into play all biological influences on the product and allows measurement of product potency, it is often difficult to develop and validate an appropriate animal model that mimics the human situation. Often, however, it is worth consider- ing animal models for potency testing over in vitro models.
Again, consider the example of our r-protein, indicated to treat a disease, based on buildup of an undesirable molecule inside certain cells. To reach the desired biological endpoint, the r-protein must function properly at sev- eral cellular locations and in a number of ways. Unlike the situation of test- ing for potency in a simple and highly defined in vitro laboratory model, there are other, extracellular influences that impact this molecule, as it exists in a human. Hence, a well-designed panel of potency assays for this
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biopharmaceutical takes into consideration the functions at the cellular level. For the r-protein, the initial QC plan considers three potency assays: one that examines potency in a living animal; another that focuses on the r-protein entering the target cell; and a third that involves the measurement of a spe- cific desired activity within the cell. This plan applies a commonly used and practical approach to ensure a potent biotechnology product in every lot of FP and performance of product under multiple potency tests, each of which measures a different aspect of the potency attribute. Could an animal model possibly be used to measure the potency of r-protein? Perhaps. As noted ear- lier, the best potency tests are developed in or adapted from the sponsor’s research laboratory, where the technology might have already been applied for investigational purposes.
In-Process Testing
The concept of in-process testing during product manufacture was intro- duced in Chapter 6, and several examples for various products were provided therein. Analysis of any product is important to establish effective manufac- turing processes and to maintain quality. First, timely feedback regarding product, impurities, or contaminants in the product flow allows production scientists and staff to make adjustments and resolve the issues. Stopping a process midstream and then reworking a particular step is usually a much less costly solution than uncovering a problem at the end of manufacture and then having to repeat the entire process. In-process testing also gives a level of assurance that product will be pure and potent once it is tested at the end of production. This too can save time and resources.
Although in-process testing is included in manufacturing protocols and samples are taken by manufacturing staff, typically the QC scientist will develop in-process assays and test samples supplied to the laboratory by manufacturing staff. In-process test results appear on manufacturing docu- ments such as batch production records (Chapters 5 and 6). Most in-process tests characterize products by using chemical or simple biological measure- ments, examining strength, measuring the amount of product, or testing either for product purity or for specific impurities and contaminants. Many in-process tests are developed for release testing of BS or FP and others are modifications of tests found on those CoAs. Whatever the test, it must be designed to generate results in a short period of time, usually within hours or a few days. This means that many tests, such as complex biologi- cal potency assays and those methods performed by CRO laboratories dis- tant from the manufacturing facility, are unlikely candidates for in-process use. In addition, in-process tests are simple to perform and do not require expensive, dedicated instrumentation or staff with special analytical skills.
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Despite these disclaimers, a number of tests mentioned in our discussions on release of BS and FP can be adapted, and many more are available from the methods listed later in this chapter. Some excellent in-process tests are commercially available and used to rapidly measure protein concentration, counting cells, or assess their viability. Most are adaptable to measure var- ious products for two or more attributes; examples are simple analytical chromatography methods, such as HPLC, and various types of electropho- resis, notably rapid procedures like SDS-PAGE.
Analytical Methods
Having introduced the QC test development and specification development cycle (Figure 7.1), we now examine the technical aspects and advantages and limitations of several analytical tests commonly applied to biotechnology prod- uct’s QC. Analytical methods, such as specifications, are adapted from many sources by QC scientists. Some methods, such as sterility testing, are com- pendia and are performed only using very specific recipes and reagents and according to industry standard specifications. Other methods are traditional or generic in basic design but adopted for use on a specific product or group of related biotechnology products. Generic methods provide some flexibility in the method of performance, and the specifications are product specific. There are also novel tests and analytical methods developed for a special measure- ment of one product. Methods are classified in other ways: by analytical instru- ment, degree of difficulty, foundation technology, type of product, or level of product manufacture or development. Presented below, but not classified or listed in any special manner, are certain tests commonly applied to biophar- maceutical development. Potency assays are given little attention, because, as mentioned before, they are often home-made and relevant to one or a few prod- ucts. Potency assays for a variety of biopharmaceuticals are, however, listed in the manufacturing descriptions and figures (flowcharts) of Chapter 6.
Quality control tests are performed in a dedicated laboratory (Figure 7.2), or samples may be submitted to contract laboratories for analysis. The small- or medium-sized biopharmaceutical firm is well advised to use contract labo- ratories for specialized tests (e.g., sterility), for methods that require complex or expensive pieces of equipment (e.g., mass spectroscopy), and whenever special scientific expertise is required (e.g., tests for posttranslational modi- fications). Since regulatory requirements for QC testing are extensive, such testing is rarely performed in a research laboratory.
• Sterility test: Sterility testing is required by regulatory agencies under guidelines (e.g., FDA 21 CFR 610.12). Further, sterility test- ing methods described in great detail in pharmacopeias sterility test,
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if passed, ensure that a biopharmaceutical is at the sterility assur- ance level required for a parenteral product. The USP sterility test provides 95% assurance that no more than one vial of product in one million vials will have a bacterium or fungus. Considering it is impossible to test 1 million vials in each lot of a product, the assur- ance level is a statistical relationship to the actual number tested, which can be surprisingly low. The test is performed with great care, to exact procedures and with many controls. Sampling protocols are carefully designed to ensure that representative product is selected for testing. A specification for sterility test, USP <71>, might read: Sterile or no growth, as growth of organisms is the measurement made on a sample.
• Microbial limits test (MLT) or microbial enumeration test: For in- process materials and often for BS, another compendial method, the MLT, is used in place of the sterility test to determine the microbial load or bioburden. The MLT, USP <61>, is designed to enumerate the total bioburden in a sample and to identify a few select and highly unde- sirable bacteria and fungi, should they grow from that sample. The specification for the MLT is expressed as colony-forming units per sample (dose or milliliter). A result might read: <2 Colony-forming units per milliliter of sample and no pathogens detected. Both MLT and sterility testing are very specialized and highly regulated endeav- ors; hence, biopharmaceutical firms may contract this work to spe- cialty laboratories.
FIGURE 7.2 Quality control laboratory. This area of the quality control laboratory is dedicated to microbiol- ogy testing.
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• Endotoxin test: This test measures the amount of a toxic mol- ecule, endotoxin, that is produced and shed by many species of gram- negative bacteria. A gel clot, lymph amebocyte lysis (LAL) assay, is commonly used, but there are other accepted methods. It is an important test performed on most biotechnology products because the endotoxin molecule can result in adverse events, such as inflammation and even shock and death, in humans. Thus, endotoxin serves as a sentinel for past or present bacterial con- tamination of a product. Some expression vectors themselves pro- duce endotoxin, shedding it into the product stream. Endotoxin is sticky, adhering to surfaces or other molecules, and it persists, so absence or low levels of endotoxin signals good production and purification techniques for a biopharmaceutical. As is the case for MLT and sterility testing, national regulatory agencies or interna- tional advisory groups have established specifications for endo- toxin, that is, maximum acceptable levels that cannot be exceeded in certain classes of products intended for a specific use or route of administration. A result might read: 4.3 EU/mL endotoxin by gel- clot LAL.
• Appearance: Appearance tests measure attributes such as color, pres- ence or absence of visible particulates or aggregates, and clarity. The appearance test is often performed visually by trained operators, who inspect a representative number of containers, selected at ran- dom at various times during the fill operation. Inspection of syringes or vials is typically done before an indirect bright light and a dark/ light background. Through training and carefully written SOPs, the examiner becomes proficient at identifying certain undesirable traits such as coloration, opaqueness, aggregates, or particulates. Instructions for reporting results are critical to success of appear- ance testing. Instrumentation is also used by some biopharmaceuti- cal QC laboratories to scan vials of FP or samples of BS to measure appearance. A specification might read Clear, colorless solution, free of visible particulates or aggregates.
• General safety test: A general safety test, historically required by the Center for Biologics Evaluation and Research, USFDA, for release of most biopharmaceutical FP, is exactly described in 21 CFR 610.11. It can detect general toxicity of a biopharmaceutical. However, it is no longer routinely required by FDA for most products.
• Osmolality: Virtually, all biopharmaceuticals are formulated in buffered salt solutions and kept at a particular ionic strength and pH. The ionic strength of FP, and sometimes of BS, is determined using an osmometer. Ionic strength is important to the stability of many products and reflects proper formulation. A result might read: 200 ± 10 mOs/kg.
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• pH: A basic but important measurement is pH, because biopharma- ceuticals are often unstable and macromolecules can degrade at low or high pH values. Formulations are often designed to maintain a narrow range of pH for each product. pH measurements are made using a pH meter with a microprobe. A result might read: pH 7.25.
• N-terminal sequencing: Used as an identity assay and described ear- lier, this inexpensive method establishes a unique identifier for a protein. It is performed by sequencing, from the N-terminus, the first 10 amino acids in a recombinant protein. Example of a result is: KQENMEVRLL versus known and reference standard KQENMEVRLL.
• Polyacrylamide gel electrophoresis, native or reduced molecule (PAGE or SDS-PAGE): This assay determines the molecular weight of a mole- cule and it can disclose impurities in a preparation. It is typically used with proteins and glycoproteins. The sample is subjected to an elec- tric field, electrophoresed in a polyacrylamide gel matrix, and the gel is stained with a vital dye to disclose bands of proteins, distributed by molecular weight. An example of PAGE is shown in Figure 7.3.
41 kDa
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kDa M
ark er
La ne
1 La
ne 2
La ne
3 La
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FIGURE 7.3 Polyacrylamide gel electrophoretogram. A polyacrylamide gel was stained for protein with Coomassie Brilliant Blue as a quality control test for identity of r-protein (molecular weight of 41 kDa). The gel is divided into five vertical lanes, with one standard or test sample applied to the top of each lane, followed by electrophoresis to separate proteins by molecular weight. Lane Marker receives standard protein sample containing six proteins of known molecular weight, 25, 30, 46, 58, 80, and 175 kDa. A negative control sample of r-protein, first digested with proteolytic enzyme, is applied to Lane #1. Lane #2 is a reference standard of r-protein, a positive control. Samples in Lanes #3 and #4 are from BS and FP, respectively, of a lot of r-protein manufactured by fermentation of recombinant bacteria with purification. Results of the samples from BS and FP show a single band at 41 kDa, identical to the reference standard. This electrophoretogram suggests that within the sensitivity of this assay, there is little or no other protein in the samples.
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It is not generally considered a quantitative test, but an estimate of purity can be calculated if comparisons are made on the same gel of test material versus qualified reference standards of known purity, for example, 100%, 95%, 90%, 80%, and 60%. Polyacrylamide gel elec- trophoresis, when used under various conditions, such as reducing or nonreducing, rapidly provide semiquantitative information and other valuable insights into product identity, structure (secondary, tertiary, and quaternary), and purity. When testing unreduced or native proteins, shape or change isoforms can be found, even in small amounts, and can be compared to reference standard. A high load of sample can reveal oligomeric or aggregate species at the top of the loaded lane. A typical test result might read: Native (nonreduced): Single dominant band at MW 41 kDa and two faint bands at approximately 20 and 10 kDa. Minimal amount of material at the top of the load lane. Reduced (SDS): Dominant bands at MW 20 and 36 kDa and faint bands at MW 10, 15, and 26 kDa.
• Electrophoretic methods: Other electrophoretic methods may be used. Each method, aimed at identifying a unique attribute, uses a different matrix or format to retain macromolecules while they are subjected to an electric field, two or even three dimensional. Immunoelectrophoresis is one example. Most electrophoretic meth- ods are commercially available and easily adaptable to suitable QC testing protocols. Further, some electrophoretic methods can be immediately followed by immunological assays on the sample to further identify each type of molecule (e.g., PAGE and Western blot testing). The adaptive possibilities are many and varied.
• Western blot (of PAGE or SDS-PAGE): Sometimes applied as an iden- tity test, but adapted to also detect certain impurities, a molecular profile by Western blot analysis is performed by transferring mol- ecules from a PAGE gel to a membrane and then treating that mem- brane with polyvalent antiserum, or with a monoclonal antibody, specifically reactive against the test protein product or an epitope of that test protein. The reaction is developed by immunohistochemi- cal methods to demonstrate colored band(s), which should fall at the molecular weight location of the protein and be comparable to the immunoblot bands of reference material. Oligomeric or aggregated species may also be detected at the top of the lanes. Using antisera specific for known impurities, Western blot may also identify those materials. Protein reference standards and negative control antisera or monoclonal antibodies are used. Western blots are not quantita- tive. A result might read: r-protein, major band at 41 kDa recognized as major band at 41 kDa by polyclonal rabbit serum to Protein-r and by mono- clonal antibodies 3D7, 8F8, and 4D2, specific for epitopes Ala3, Leu29, and Try54 of r-protein. No other bands detected.
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• Host cell protein: After molecules are produced in a cell-based sys- tem, they are associated with impurities and contaminants, that is, a variety of host cell substances and process materials (Chapter 6). These are often proteins: enzymes or structural molecules. Host cell proteins can be identified by a variety of methods. Popular ones are antibody-based assays, such as an enzyme-linked immunoas- say. This measurement uses a polyvalent animal antiserum raised against proteins of the host cell to measure impurities or contami- nants. As it is not possible to identify or measure every host cell contaminant, specific or marker proteins may be measured. A result might read: <0.10 mg host cell protein per 100 mg recombinant protein in the solution.
• Host cell DNA: DNA is an impurity, unless the product itself is com- posed of DNA (e.g., bacterial plasmid), in which case anything other than the desired DNA molecule is an impurity (e.g., chromosomal DNA). DNA is measured by a variety of methods, many commer- cially available, with great accuracy and specificity. Assays include threshold measurement with DNA-binding proteins, hybridization assays for specific DNA of defined origin, and quantitative poly- merase chain reaction probe methods, which are highly specific. A result might read: <10 µg DNA/1.0 mg recombinant protein.
• Host cell RNA: Some host cell production systems can yield a consid- erable amount of undesirable host cell RNA. Commercial test kits are used to measure this molecular impurity. A result might read: <10 µg RNA/1.0 mg recombinant protein.
• Carbohydrate: Some biopharmaceuticals must be posttranslationally modified to show activity. Indeed, glycosylation and a unique pat- tern (e.g., location on protein, composition, and structure or pattern of glycosylation of each carbohydrate side chain) can be important to potency. A variety of methods, adapted from classical carbohydrate chemistry and some now semiautomated, are applied as surrogate measures of potency to demonstrate identity of some biomolecules.
• Light scatter for aggregates: Subvisible molecular mini-particles and aggregates can be disclosed and measured using instruments that measure the scatter of light as it passes through a solution of prod- uct. A result might read: <0.1% scatter at wavelength 300 nm.
• Protein measurement: A variety of tests, some commercial and others developed in or adopted by the QC laboratory, are on the market to measure the amount of protein in solution. Each has advantages and limitations, so the QC scientist picks a test carefully to meet a partic- ular need and qualifies it for use with a given protein. The Bradford test or a BCA reagent-based test is commonly used in biotechnology. A result might read: Total protein, 1.06 ± 0.1 mg/mL.
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• Peptide mapping: This is an identity test. A protein in solution is digested with an enzyme, for example, trypsin, and the fragments are subjected to an electric current (electrophoresis) in a matrix and then stained. The pattern of fragments is characteristic to a given protein. The result would be obtained from the peptide map and might be given as: Matches predicted and reference maps.
• Size exclusion chromatography: The SEC method examines a sample of protein for purity and impurities by using a chromatography gel that distributes proteins on the basis of size. Aggregates of protein are detected by SEC. The results provide a measure of purity and identify impurities based on molecular size. A result might read: Dominant peaks at MW 20 and 36 kDa and faint peaks at MW 10, 15, and 26 kDa.
• Isoform characterization: The isoelectric focus assay, a high-resolution method that allows the separation of proteins based on their iso- electric point, evaluates the charge characteristics of a protein and can demonstrate isoforms, major variants, of the protein. Isoelectric focus gels are scanned and bands can be measured and identified by pI. Results are not quantitative, but estimates may be made from scans. Example of an isoelectric focus result: Major sample band lies between pI markers of 5.20 and 5.85 and is comparable to reference stan- dard. Minor variants constitute under 10% of total protein.
• Amino acid composition: This method may be used as an identity test. An analytical instrument determines the amount of each amino acid and then calculates the ratio. This ratio is compared to the expected ratio and to that measured for a reference standard. The ratio rela- tive to a reference amino acid, say L-leucine, may also be determined and compared to theoretical and reference standard. To perform the test, a sample of protein is hydrolyzed with acid and the amino acid composition is determined by an automated method. Typical result might read: Correct amino acid composition ±10%.
• Chromatography: Many analytical variations and instruments are applied to this methodology, which separates chemicals in a com- plex sample. Molecular characteristics, such as charge and molecular weight, are the basis for their separation. Sample may be entrained in a semisolid matrix (thin-layer chromatography) or in a gas stream (gas chromatography), held within a narrow column. A detector, scanner or in-line, measures molecules as they exit from the long tube. A chromatogram, to include quantitative data when reference standards are applied, is produced from the detector. Gas chroma- tography is especially useful for detecting residual solvents in a product. A result might read: Residual isobutane <10 ng/mL.
• High-pressure liquid chromatography: This has been applied to mea- sure several attributes of many biological molecules. As the name
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suggests, HPLC is distinguished by applying very high pressures to the enclosed column. It is especially useful with macromole- cules and shows excellent resolving power, with clear separations on the chromatographs. In addition, it is relatively fast and inex- pensive and quite adaptable. Sample is separated into components that appear on the output as distinct peaks or even bumps or shoul- ders on a major peak. An HPLC apparatus and sample chromato- gram are shown in Figure 7.4. It also has the ability to measure the amount of material under each of those peaks and, for reference purposes, to spike known molecules, such as impurities or contami- nants, into a sample of highly purified reference product. Indeed, this is how experiments are designed and initial results are seen
(a)
FIGURE 7.4 High-pressure liquid chromatography. Panel (a) shows a high-pressure liquid chromatography instrument, stacked modules on left, and dedicated computer containing analytical software on the lower right. (Continued)
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using several other modern chromatography and spectroscopy methods. Typical result might read: Major peak at 15.8″ comprising 97.68%. Two minor peaks at 22″ (0.88%) and at 18″ (0.27%), with slight shoulders at leading and trailing edge of major peak.
• Electrospray ionization-mass spectrometry: This instrument-based test is used to measure mass of a molecule and may be applied to study protein folding. The results are compared to both reference standard and the theoretical mass of the molecule of interest. Controls might include proteins of known masses, especially those in the molecular weight range of the test material. Results are reported in kilodal- ton (kDa) unit of protein mass. Typical result might read: 41.111 kDa (versus theoretical 41.005 kDa).
• N-terminal and C-terminal analysis by liquid chromatography-mass spectrom- etry: This method uses physical separation by liquid chromatography and mass analysis by mass spectrometry. It is highly sensitive and spe- cific and can be applied to characterize a variety of proteins. It confirms both the N-terminal and the C-terminal sequences. Typical result might read: N-terminal, KQEN; C-terminal, EIGGY; comparable to the reference standard.
−3 .7
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FIGURE 7.4 (Continued) High-pressure liquid chromatography. Panel (b) shows a chromatograph of r-protein bulk substance. Here, the large single peak to the right, at 4.85 min, is r-protein (determined by comparison to a reference standard), whereas the peak on the left, at 4.46 min, represents a significant amount of a contaminant protein, which has a shoulder peak at 4.33 min, perhaps a second but closely related contaminant protein.
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• Protein folding and unfolding by intrinsic fluorescence: Protein folding, often important to biological activity of a macromolecule, may be compromised because of incorrect posttranslational folding or mod- ification. Properly folded protein may denature during processing or storage. Hence, it is sometimes important to demonstrate correct folding in a protein product. The amino acids tryptophan and tyro- sine in proteins fluoresce under specific wavelengths of light. As proteins in the native or correctly folded state demonstrate a unique fluorescence signal and because denaturation of a protein results in a shift (e.g., red-shift) of the fluorescence emission barycentric mean value (which can be derived from an analytical instrument), protein folding can be measured. A specification for batch-to-batch variance can be established. An example is barycentric mean Lambdanm < 358 for fluorescence measured between 300 nm and 400 nm. A result might read: Lambdanm 340 for fluorescence measured at 380 nm.
• Mass spectrometry-time of flight: This method relies on a complex piece of equipment and represents one of the several new and promising analytical tools that may be used to characterize proteins and other biological molecules. It is useful in describing protein folding.
Additional Analytical Tools and Concepts
The tests identified above are used to test the attributes of biotechnology products that exist as molecular entities, but what about testing for appear- ance, safety, identity, purity, and strength of living biopharmaceutical prod- ucts, such as attenuated organisms used in vaccines, retroviral vectors, and somatic or pluripotent-derived cell and tissue products, as introduced in Chapter 6? Here, we review, with very general descriptions, a few of the many methods used in the QC of various biopharmaceuticals, notably live materials.
• Cell karyotyping: A karyotype represents the appearance and num- ber of chromosomes in the nucleus of a eukaryotic cell. Cytogenetic analysis of a cell’s karyotype is used as an identity or a purity test to demonstrate quality of a cell line, especially when that line was used as a somatic or pluripotent cell-derived therapeutic or to pro- duce macromolecules. Karyotyping is performed in specialty labo- ratories, and reference cell lines are required. The number of cells with an abnormal karyotype is measured. The species origin of the cells is also confirmed. The result would be presented to identify the cell line, as compared to a reference cell line, and might also attest to its purity.
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• Cell phenotyping: Any cell trait or characteristic distinctive of that cell line is considered phenotypic. Cell phenotyping, used when a cell line or tissue is derived from somatic or pluripotent cell sources, measures one or more molecular parameters to demonstrate that cells are identical to those intended and that the cells have neither differentiated or dedifferentiated and they are not contaminated. The result would be presented to identify the cell line and attest to its purity.
• Microbial identification: Bacteria, fungi, yeast, and viruses, includ- ing retrovirus and bacteriophage, are biotechnology products. They are derived from many sources and manufactured in various ways, both with and without cell-based systems. An identity test demon- strates, in various ways, that the microbe is as purported. Bacteria and yeast are identified by traditional methods such as culture on selective media and metabolic properties. Viruses are identified by growth characteristics on selective cell lines. All microbes may be further identified using species-specific antibodies to agglutinate or label with fluorescent dyes, to kill them in the presence of comple- ment, or to neutralize their activity. Describing their morphology or ultrastructure is also an effective means. Polymerase chain reaction is increasingly used to identity live organisms or DNA molecular products.
• Monoxenic nature of microbial product: The purity of a microbial prod- uct may be demonstrated to show that all organisms in a product have the same trait. Cell phenotyping or karyotyping or microbial identification methods, described above, can be applied for this purpose. Other methods use growth characteristics or a panel of chemical or immunological reagents to selectively identify possible contaminants. An example is the use of selective media that support the growth of most bacteria but not of the strain or species compris- ing the product. Polymerase chain reaction, using probes against DNA from a variety of possible contaminants, is another test used to reveal the purity of a culture.
• Attenuation of microbial product: Many products, notably those intended for genetic therapy or vaccines, are attenuated, so that they do not produce disease. Quality control tests for safety focus on ensuring markers of attenuation, such as inability to grow on cer- tain substrates or in specific cell lines. These traits, or lack thereof, may also be evident by using antibody or molecular probes, such as immunofluorescence assays. Polymerase chain reaction and other genetic probes identify a gene of interest in any host.
• Expression of a molecule by a vector or host cell: Other biotechnology products are engineered to express a molecule, which, in turn, exerts an immunological or therapeutic effect on the user. Quality
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control tests are based on methods mentioned above, but instead of searching for the absence of a trait, they focus on confirming that an attribute is present and active or functioning. Immunological and molecular probes are used to ensure that an expression product is expressed and exists at the expected location, such as on the cell surface.
• Adventitious agent testing: Adventitious agents may be found in a variety of cell products and cell banks. Considerable effort is put in examining samples for microbial agents, for example, mycoplasma and retrovirus. Some testing is performed to identify, through cul- ture or immunological methods, the agents themselves. More often, indirect measures of adventitious agents, such as electron micros- copy to detect viral or viral-like particles or polymerase chain reac- tion, are used to locate and, sometimes, identify undesirable microbes or the DNA fingerprint they leave behind. If infectious agents are not obvious at the outset, they may be induced by applying a vari- ety of chemical or biological stimulants to the cell line. For example, endogenous retrovirus can be induced with nucleotide analogues and then detected by electron microscopy as viral particles. Viruses can be detected by injecting cell lysate into newborn animals and then examining the animals months later. For some biotechnology and blood-derived or supplemented products, such tests are critical to ensuring safety.
Quality Control of Cell Banks
As noted in Chapter 6, the identity, purity, and viability of cell banks (micro- bial, mammalian, or insect cells) are very important attributes to the overall manufacturing quality and success. Every cell bank is tested both at the time of release and at specific intervals. The attributes considered in most test protocols are identity, sterility, purity, and viability (potency). Following are the tests applied to the QC of cell banks, both master and working cell banks, as well as the progeny of cell banks that are used in extended manufacturing campaigns.
• Identity of cells: The identity of cells in a bank is established by applying tests to ensure that they are the intended species and strain. Particular methods are described above, under the headings Microbial identification, Cell karyotyping, and Cell phenotyping.
• Identity of insert: If a cell line has been genetically engineered to express a product, then the process of expression and the expression product
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are tested to ensure that banked cells possess these intended capacities. Methods described in the earlier sections of this chapter are applied, as appropriate, to detect the nature and function of the molecule or other attribute.
• Potency: This is defined as the capacity for cells in a cell bank to divide after removal from storage. This function is clearly basic to the intended use, and so, quantitative potency testing is an annual QC physical examination for any cell line.
• Purity: To ensure that a cell line has not been contaminated with another product, and this does happen, then the monoxenic nature of the cell line must be established using the methods described above, notably adventitious agent testing.
Samples and Sampling
Consideration of sampling methods, assay controls, and reference standards is an important aspect of QC planning because it is critical for each test sam- ple to represent the whole of that batch or lot of BS or FP. Thus, sampling must follow standard procedures, with methods tailored to the intended scope and nature of each test and heed statistical considerations such as sample size or random selection. Indeed, papers and books are written about sampling methods for QC of various consumer products, including pharmaceuticals. Beyond the number of samples taken, there must be a plan for sampling per- formance. For example, if one wished to sample 100 glass vials containing a biopharmaceutical from a total lot of 10,000 vials, it would be important to take vials periodically, perhaps 10 vials at each of 10 time points, throughout the fill, as opposed to grabbing the first or last 100 vials in the fill line.
Also, when seeking a representative sample from a single container it is necessary to take the sample in the proper manner. For example, if one is taking sample for aggregate testing from a small vial containing a recombi- nant protein, the container is first gently stirred and then the sample is taken, ensuring aggregates, that tend to settle to the bottom of a vial over time, are fairly represented throughout the sample. Sampling raw materials provided by a vendor in large containers drives the need to ensure that containers are selected in a representative yet random manner and, for each selected container, to randomly take material from within that container. The gen- eral concepts of sampling are included in a QC plan, and specific sampling methods for each assay are written into the test protocol or SOP. If not every sample is subsequently assayed, then it is important to ensure that tested samples are indeed representative of all samples taken. Consultation with a statistician is often helpful throughout the sampling process.
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Analytical Controls and Reference Standards
To ensure that each QC test is reliable and to maintain consistency in test- ing, control samples, reagents with known analytic values, are always tested in parallel with unknown samples. While every assay requires one or more controls, a full set of control materials is too often missing from an otherwise adequate test procedure. In establishing a QC test, a panel of control samples is carefully chosen and then applied correctly within the test scheme. Each con- trol reagent is certified for a particular purpose, and during assay qualification or validation, it is shown to produce the intended outcome. Positive and nega- tive control materials are generated through biomanufacturing or purchase from vendors, before beginning the analytical work. Controls are stored in a manner that retain all desired attributes. Controls for quantitative assays, such as those requiring generation of a standard curve, require particular attention.
In contrast to control reagents, a reference standard is a test sample with known analytical values. It has a pedigree. A reference standard consistently produces, by repeated testing with a standard assay and over a long period of time, the same result. Reference standards are by definition used with every assay or panel of assays. In some instances, more than one reference standard is required for an assay, such as when a standard curve is produced by limit- ing dilution of the standard.
A reference standard also is the same or nearly the same as the actual test material, the product or analyte. For example, if actual samples are substance or product that contains excipients and buffers, the reference standard is for- mulated in the same way. Hence, the best source of each reference standard is a batch of substance or a lot of product manufactured and controlled in the same manner as is currently used. However, in those instances when a recom- binant protein cannot serve as a reference standard due to issues such as insta- bility, it may become necessary to purify and store small amounts of the native molecules to avoid the issue, that is are stable formulations. Producing a refer- ence standard can be a very resource-intensive project. How does one obtain a reference standard? One option is to store BS or FP in aliquots and then use one aliquot, taken from the first batch or lot of product, as the first reference standard. Over time and as manufacture and control expertise improves, this first reference standard is replaced with material from new batches of BS or lots of FP. This is also the case when tests are redeveloped, as shown in Figure 7.1 as T-1, T-2, and T-3. It is not unusual for five or more individual, reference standard lots to be used over the development cycle of a single assay. Cross- over studies are performed to compare, in great detail, an old to a new refer- ence standard, that is, to ensure that the pedigree is maintained. To further complicate the matter, a reference standard, like a control, is used and relied upon over a long period of time and thus each is kept in extended storage. Early in development, and often for years into the development cycle, there is limited information on stability profiles of controls and reference standards.
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This creates unknowns regarding their reliability when used to test product over that same period or into the future. A plan to meet each of these chal- lenges becomes a part of the QC plan and this is typically done for each assay.
Test Failures, Out-of-Specification Results, and Retesting
As one might expect, failures are experienced in the QC laboratory just as they are in other aspects of biotechnology operations. When the QC test result on a given batch or lot does not meet the specification it deviates or is out of specification. In some biotechnology operations, such as biomanufacturing, a failed process can be repeated, at least if there is an explanation and time and money allow. Failure in testing, specifically the failure for a test to meet a specification, can be difficult to resolve. There are strict controls on managing deviations in a regulated environment, and biopharmaceutical QC follows guidelines and regulations promulgated by FDA. Further, some highly pub- licized judicial actions taken by FDA originated from improper retesting and reporting of QC test results.
Deviations in QC testing demand, by regulation, an internal investigation. Investigations involve a complex process, having four major components that are described in greater detail in Chapter 5. A case study of an out of specification for a potency test is reviewed in Box 7.3. An investigation uses established methods, a root cause analysis, a corrective and preventive action plan, and approval of the outcomes and recommendations by supervisors, QC, and QA. Further, findings of the investigation may lead to a recommen- dation of significant actions, such as qualification or validation even replac- ing an assay before it can be used to further test a product. Other resolutions, closely monitoring the performance of the assay, replacing key components such as a reference standard, and establishing formal audits or documenta- tion systems, may be recommended by the QA unit. Since management is ultimately responsible, the failure may be raised to that level.
It is important to prevent test failures and this is achieved in several ways. First, full QC planning prevents many failures and sticking to the accepted plan avoids other. Another approach is to follow a QC development cycle (Figure 7.1), that is by developing each assay one step at a time and not skip- ping a step or a critical experiment. The most frequently cited reason for fail- ures in QC testing is, in this author’s experience, setting unrealistic (meaning too stringent) specifications in early and middle phases of the development cycle. In effect this means that the development team establishes a speci- fication before adequate data was available to support that specification. Following the hallmarks of quality and abiding by a quality system, current Good Manufacturing Practices for most biopharmaceutical QC operations is another way to ensure success with QC endeavors.
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BOX 7.3 CASE STUDY: QUALITY CONTROL TEST FAILURE AND INVESTIGATION
The r-protein was formulated and filled into single-dose vials for use in a Phase 3 clinical trial. Three final lots, #1, #2, and #3, each having 8000 vials at a cost of $25 per vial, were manufactured and then tested by QC. For Lot #3, a FP potency test (Table 7.2), Accumulation of Molecule in Cultured Cells at 24 h, failed to meet the specification of lesser than 10% accumulation over baseline, time 0 (of the test assay). The result for Lot #3 was 12%.
QC and Quality Assurance (QA) would not release the lot, despite the fact that it was needed for patients in the trial and represented an investment of more than $20,000.
QA called for an investigation by an ad hoc committee composed of supervisors from research, manufacture, QC, and QA. Careful review of all documents failed to reveal evidence of errors to properly per- form the test or maintain all records. It did reveal that the standard operating procedure (SOP) for preparing the cultured cells, a critical biological component of this potency assay, allowed a wide range in the age of the cells that could be used as cellular substrate in the assay. Specifically, cells could be used from 2 to 8 days. Research pointed out that older cells, those older than 5 days, could, at times, be in asynchronous cycles; indeed, some cells even experienced death at 7–8 days.
Further investigation showed that Lot #3 was tested using 7-day-old cells, within limits stated by the current SOP but perhaps not advisable. Lots #1 and #2 and other lots tested previously used cells of 3–5 days of age.
Laboratory investigation in a well-designed and -controlled study tested Lots #1, #2, and #3 using cells of various ages, 2–8 days old. Findings consistently demonstrated that older cells yielded higher val- ues, often greater than 10, of accumulation of molecule in cells.
A second laboratory investigation tested Lot #3 repeatedly with cells of 3–5 days of age and all results were lesser than 10. There was a sci- entific basis to this finding as well, given the mechanism of action for r-protein.
The committee recommended release of Lot #3, changes to the SOP, allowing only cells of 3–5 days of age, and careful and complete docu- mentation of the investigation and all laboratory results.
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Testing for Product Stability
All BS and FP are placed in storage and transported for defined periods and at established environmental conditions. BS is usually stored for shorter periods, days or weeks, as compared to FP, for months or years. There are significant advantages, both economic and operational, to establishing a long shelf life for BS or FP. Substance and product stability profiles are established over the product development cycle by using many storage conditions, each exact, and by testing with multiple assays at many time points. It must be demonstrated that BS and FP remain pure, potent and safe during storage and transport. Because some products are likely to be subjected to shaking, inversion, humidity, and temperature incursions, cool and hot, during stor- age stability testing is performed. Stability protocols include subjecting both BS and FP to undesirable but possible conditions. Establishing a stability profile is a long, tedious and expensive endeavor but it is necessary and usu- ally well worth the effort.
Experimental stability protocols are designed by QC and manufacturing staff to evaluate the attributes and desired traits of BS and FP, as might be expected under conditions of storage, handling and shipment. Elements of a stability plan are outlined in Figure 7.5. Most biotechnology products today are kept refrigerated or frozen, but all products are subject to fluctuations in temperature and humidity, or they face exposure to light. Environmental conditions certainly change as the product is moved: from manufacturer to truck; then to wholesale warehouse; again to truck or aircraft; then to phar- macy, mailbox, or automobile; and finally to the consumer. Also, if a prod- uct is to be shipped, stored, and used in regions with a warm moist climate where it is difficult to always maintain a cold chain, or in very cold regions where it might be exposed to freezing temperatures, if just for 1 h, then stabil- ity studies need to be especially rigorous. Stability data provides a sponsor with very useful information because it aids in developing proper product formulation and approaches to marketing, transport, and storage. Regulatory authorities demand that storage conditions be explicit and highly visible in labeling. In addition, because one cannot sell degraded product, stability test results impact both business and marketing plans. There are significant eco- nomic advantages for a product with a long shelf life at ambient temperature, and there are marketing hurdles for the sponsor of a product that must be kept frozen, especially if the product must be kept at −80°C or cannot reach room temperature even for 1 h. Consider also the difficulties of manufactur- ing, stocking, and rotating a supply of a product with a shelf life of just 1 year and one can imagine the advantages of a 3-year shelf-life. While experimental in nature, the design of stability protocols is driven by quality, business, and market interests, because the information derived from stability studies in part ensures the sales of that product. Hence, stability testing plans are an important aspect of the overall QC plan.
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Stability testing, typically a QC function, is a formal process under which the actual shelf life of a biopharmaceutical is identified through experimentation. A key milestone is early development of a written sta- bility plan, developed by a multifunctional team before stability testing begins. The team begins by reviewing what is known about the product
Elements of a stability study plan
Study protocol
Pull samples at predefined intervals for
analysis
Results Interim and final
study reports
Analytical testing
Stability chambers
Refrigerate Humidity Incubate Light/dark
Cycle samples
Warm → Cool Freeze → �aw Light → Dark
Sample prep
Degas or gas overlay Adjust volume Mix, shake, and invert
FIGURE 7.5 Elements of a stability plan. The stability plan considers the physical environment (test cham- bers) for samples, a sampling and preparation scheme, cycling of some samples, selection of (pulling) samples to test, analytical testing, and interpretation of results.
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and similar products or molecules. First, it is important to know if a prod- uct could be stored at room temperature or in the refrigerator, as opposed to in a freezer, as simpler or ambient storage reduces cost and complica- tions of shipping and storing the product. Second, the shelf life is deter- mined empirically for each proposed storage condition. How can this be approached? Third, we often attempt to improve the shelf life or simplify the storage conditions for a product by experimenting with different for- mulations. For a given product, we know in general that certain chemicals preserve cells or proteins better than do others. What might these be and which might be experimentally evaluated? Finally, we seek an understand- ing of how long product can withstand excursions, such as high or low temperatures, high humidity, shaking, inversion, and so on, before it loses potency. These conditions must be carefully chosen, since resources limit the number of variables that may be tested.
Stability testing is an experimental endeavor, designed as a matrix experi- ment and guided by a written protocol. It applies not one but a panel of assays, each capable of measuring a stability-indicating attribute for the product. Further, to provide enough material for testing, many samples of product, both BS and FP, are stored in a variety of configurations. For example, storage at three or four temperatures and, for FP, in two positions (upright and inverted). With testing by three to four assays required at each of the seven different time points, sample requirements may reach hundreds or thousands of vials or syringes of FP or samples of BS for every lot and batch, respectively. During stability testing, storage conditions must be care- fully controlled and documented. Thus, there is a need for many samples and a laboratory infrastructure able to accommodate various environmental conditions and perform many tests. Stability testing is very labor intensive and expensive, but it is absolutely necessary from both regulatory and busi- ness standpoints.
Stability-indicating assays, if not available as QC release assays, are modi- fied from existing research or development assays or they are developed by QC scientists to fit this purpose. They are selected by first identifying those product attributes providing meaningful information about the shelf life of that product. Since any assay used to measure a product attribute is, in the- ory, a stability-indicating assay, the CoAs for the release of BS (Table 7.1) and FP (Table 7.2) provide a foundation for developing stability-testing protocols, as shown in Table 7.3 for BS and Table 7.4 for FP. However, a CoA intended for product release lists far more assays than are necessary or could ever be performed on the variety of stability test samples and at every time point. To downselect the choices to perhaps three to six assays per stability protocol, the QC scientist must first determine which attributes of the product are most likely to be the stability-indicating attributes. Next, they choose those attributes and experimentally determine if in fact they really are stability indicating.
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What are the reasons for choosing an attribute and assay for inclusion in a stability protocol? First, certain types of assays are, by tradition, stability indicating for specific classes of biopharmaceuticals. Examples include peri- odic sterility testing and appearance of all products and tests to detect the degradation of protein in storage or cell viability for products with living cells. Second, it is good to have a balanced assay portfolio, choosing one or two methods for the attribute of purity or impurities, one for potency, and one or two for safety. The nature of the product gives a clue to what con- stitutes a good stability-indicating assay, as does the understanding of the mechanism of action. For example, if a protein product tends to aggregate at the formulated concentration, then a quantitative test for aggregates is in order. Further, each test must be sensitive, so that loss of product integrity or activity is, in fact, detected early and long before the product is completely unsafe, degraded, impure, or impotent. For some products and tests, this selection is challenging. For the r-protein in our example, instability might be reflected in denaturation or breakdown of the molecule. This would result
TABLE 7.3
Tabular Synopsis of a Stability Protocol for Biopharmaceutical Bulk Substance (Example of Formulated r-Protein in Vial)
Test/Specification
Post Manufacture (Months)
1 5 12 36 60
Appearance Clear, straw-colored liquid without particulates or aggregates
X X X X X
Safety Microprobe pH 7.1 ± 0.2 X X X X X Purity SDS PAGE: Single band at 41 kDa,
comparable to reference standard X X X X X
HPLC: Single peak integrated >98% material in sample
X X X X X
Potency Blocking assay: cultured cells: >60% inhibition of secretion as compared to reference standard
X X X X X
Safety Bioburden: USP <61> <5 cfu/mL and no evidence of pathogenic organisms
X X X
Note: X = This test will be performed at this time point. Accelerated stability testing only to 12-month time point.
This protocol is performed on samples of BS that have been stored at static conditions in the upright position at the following temperatures (one set per temperature in static conditions): at all time points at -70 ± 10°C (recommended storage temperature); at Day 1 and Day 5 when kept at 21 ± 2°C (room temperature); and at Day 1 and Day 5 when kept at 37 ± 1°C (accelerated temperature).
Another set of BS samples is tested after static storage at –70 ± 10°C but after having been subjected to three freeze/4-h thaw cycles before testing.
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in an inability to perform the key biological functions and reach the desired endpoint. Referring again to the r-protein, and focusing only on FP, initial consideration is given to several key assays for early stability experiments. The tests such as SDS-PAGE and HPLC are selected for purity and impuri- ties, and the potency assay is chosen for blocking accumulated molecules by cultured cells. These are chosen in part because both are meaningful to this protein and its biological activity and because each test is simple, accurate, well controlled, and can be performed on large numbers of samples, in a relatively easy and inexpensive manner. Since vials are opened to test the product anyway, we might add tests to record the appearance and measure
TABLE 7.4
Tabular Synopsis of a Stability Protocol for Biopharmaceutical Final Product (Example of Formulated r-Protein in Vial)
Test/Specification
Recommended Storage Temperature 4 ± 2°C Post Manufacture (Months)
1 2 3 6 12 24 36 48 60
Appearance Clear, colorless liquid without particulates or aggregates
X X X X X X X X X
pH: 7.1 ± 0.2 X X X X X X X X X Purity SDS PAGE: Single band
at 41 kDa, comparable to reference standard
X X X X X X X X X
HPLC: Single peak integrated >98% material in sample
X X X X X X X X X
Potency Blocking assay: Cultured cells: >60% inhibition of secretion as compared to reference standard
X X X X X X X X X
Safety Sterility: USP <71> Sterile
X X X X X X
Note: X = This test will be performed at this time point. Accelerated stability testing only to 12-month time point.
This protocol is performed on FP that has been stored at static conditions in the upright position at the following temperatures (one set per temperature in static condi- tions): 6 ± 2°C (recommended storage temperature); 21 ± 2°C (room temperature); and during the first 12 months, 37 ± 1°C (accelerated temperature).
In addition, one set is tested after storage at 6 ± 2°C, with vials kept in both upright and inverted positions.
Another set of vials with FP is tested after static storage at 6 ± 2°C, with vials in upright position, but only after sample vial has been subjected to five freeze-thaw cycles before testing.
Yet another set is tested at all time points and at the 6 ± 2°C temperature storage con- dition in an upright position but only after sample vial has been subjected to shaking at 30 oscillations per minute for 6 h immediately before testing.
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the pH at every time point, because changes in pH are often related to pro- tein instability and because degradation often discolors or adds precipitate to a vial of product. In addition, using a single vial of product, both tests, that is, the appearance in the unopened vial and the pH, can be performed on a single vial, conserving the expensive product. Regulatory authorities ask that sterility be examined once each year, because sterility test is a test of container (vial) integrity. Returning to the example, SDS-PAGE provides an indication of whether or not the r-protein was breaking down under stressful conditions. In addition, HPLC, a much more sensitive test, confirms and extends any observations of protein breakdown, perhaps detecting the changes earlier than SDS-PAGE. Besides, HPLC analysis of r-protein might provide, on the chromatogram trace, a clue as to the breakdown products and impurities that accumulate over time. A stability-indicating assay is also chosen for the attribute of potency, based on the requirement that r-protein inhibits the buildup of intracellular carbohydrate molecules. The assay uses cultured cells, providing a well-characterized and much used method that should reflect biological activity, or loss thereof, in a relatively simple and reproducible format. Now, this stability-testing concept is designed into a written FP stability testing protocol, and the test scheme is summarized in tabular outline, as shown in Table 7.4.
Consider also that controls are included with each assay to demonstrate that a stability assay detects unstable product. These controls, partially and fully degraded or inactivated product, are tailored for use with a par- ticular assay. To develop a control, it is critical to first understand the most likely routes of degradation and then mimic this degradation in a mean- ingful way. For the r-protein example, it is not adequate to simply boil the r-protein or completely digest it with trypsin; this would not mimic possible environments in actual storage or transport. Instead, it is necessary to care- fully devise accelerated degradation by using conditions that mimic possibly real environments. These might include using room temperature found in temperate and tropical environments or one or more cycles of freeze-thaw. Producing controls is itself challenging and requires time and knowledge of possible degradation pathways.
The same process is followed for developing a stability protocol for BS, but some different assays may be chosen. It is seldom necessary to store BS for long periods, that is, months or years, because BS is formulated and filled to FP within days or weeks of manufacture. In addition, it is much easier to faithfully keep bulk containers of molecules at extreme conditions, for example, −80°C, as compared to FP. An example of a BS stability protocol is shown in Table 7.3.
Stability testing is performed on FP and BS kept not only at recommended (labeled) conditions, say refrigerated or frozen, but also at several subopti- mal conditions that might be experienced in a real situation. For the r-protein example, the intended storage conditions are refrigerated, but for stability testing, the QC scientist also tests the samples kept at room temperature
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and at a temperature above room temperature. Such protocols are referred to as accelerated stability, when they often represent accelerated decline, degradation, or decay. As noted earlier, vial handling is an important vari- able. Initially, the vials are kept upright during stability test storage, but in later stages of development, vials are placed in other configurations of stor- age, such as inverted or horizontal. Shaking and exposure to humidity or light might also be included in last-phase stability studies. Finally, several time points must be tested at each of the chosen conditions, because it is not known exactly how long the product will remain stable in any given environ- ment. The end result is a set of protocols that provide a stability test matrix. Although stability testing adds considerable effort and expense to a prod- uct’s QC program, it results in extremely valuable information and becomes the basis for ensuring proper handling, shipment, and storage of the FP. In addition, of course, it ensures that a high-quality product—pure, potent, and safe—is sold to the user. This, in turn, enhances marketing opportunities and prevents future recalls or complaints.
Quality Control Testing of Raw Materials
The quality of BS or FP reflects not only the manufacturing process and release testing, but also the quality of each raw material that goes into making that product. Therefore, raw materials are carefully selected and controlled, with involvement of staff from manufacturing, QC, and QA. Manufacturing staff have primary responsibility for selecting the finest materials to use in each process. Quality Assurance professionals approve those selections and are further involved by later signing-off on use of each lot of raw material, once it has arrived at the manufacturing facility. Quality control staff consider the technical quality of raw materials, either by reviewing the results of the tests performed by the vendor or by testing or retesting the material at the sponsor’s laboratory.
Almost any conceivable material, live or nonliving and chemical or bio- logical, is represented in the brief history of biotechnology. Microscopic particles of pure gold, live invertebrates, radioisotopes with short half-life, toxic or oncogenic chemicals, liable cells, and recombinant microbes are a few examples, and for every manufacturing protocol, there are salts, organic chemicals, and just plain water. Quality attributes of raw materials differ based on their intended use and integration into the product. Items that do not contact the product, such as a detergent used to clean the floors in a manufacturing facility, receive the least attention. For example, QC at a biopharmaceutical firm would review the CoA provided by the detergent’s supplier to ensure that this material is generally recognized as safe, that it has been tested by the manufacturer or distributor, and that it meets the
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specifications; the biopharmaceutical firm might not retest this detergent to verify this CoA. In this manner, the manufacturer’s CoA attests to the safe, pure, and potent nature of that raw material. Certain other raw materials, such as culture media or salts provided to the biopharmaceutical manufac- turer by an established and reputable vendor and carrying both a USP cer- tification on the label and a CoA, might not be routinely retested. The QA department might audit these vendors periodically (Chapter 5), but unless there are potential issues with the vendor or the raw material, laboratory testing might not be repeated, at least not often. The third and the greatest level of attention is given to the raw materials that become part of the prod- uct or directly contact the product and do not carry a recognized certification (e.g., USP), those that are notoriously difficult to control, or those that have even a remote possibility of harboring adventitious agents or toxic materials. These raw materials, examples of which are given below, demand retesting in the biomanufacturer’s QC laboratory, or they should receive other special consideration to ensure that they are, in fact, safe, pure, and potent. General testing requirements for a few classes of raw materials commonly used in biomanufacturing are given below.
• Solid containers and process equipment: Containers, such as hold ves- sels, and process equipment, such as plastic tubing and filters, are known to shed particles or to allow chemicals to dissolve (leachates) into the product stream. However, biomanufacturers take great care in choosing the process materials that release particles or chemicals. In steps such as product holds, in which this might happen even with the highest-quality materials, the QC laboratory may be called upon to test for those possible contaminants in the raw material, in- process samples, or the product. Standard assays are available to the biopharmaceutical industry for detecting particles and many leach- able chemicals. Adventitious agent testing was described earlier in this chapter.
• Water: As noted in Chapter 6, large amounts of water are used in bio- manufacturing, and this must be of the highest purity and without any microbial contamination or undesirable dissolved chemicals. Water is purchased as purified or as water for injection by some oper- ations, but biomanufacturers often purify water themselves, begin- ning with an excellent source of tap or well water. Since it is used in large volumes, impurities or microbes that enter at upstream steps in biomanufacturing may be carried through or even concentrated during the process and thus end up in the product. Hence, water of all grades must be tested for traces of chemicals, such as total dis- solved carbon; particles; endotoxin; yeast; and bacteria. Compendia (e.g., USP <1231> Water for Pharmaceutical Purposes) describe the various levels of water quality and the tests used by QC laboratories to ensure that this critical reagent remains pure and safe.
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• Inorganic and organic chemicals: Large amounts of salts or saline solutions are purchased or prepared during biomanufactur- ing. Whenever possible, compendial grade or otherwise certified reagents are purchased from a reputable vendor and the CoA is carefully reviewed before acceptance. Retesting may or may not be called for, depending on the material source, the previous test pro- tocols or certifications or lack thereof, and the risk profile. When a USP-grade material is not available, the QC laboratory may test a raw material to ensure that it meets the established specifications.
• Culture media and supplements: These raw materials, most of which have a CoA but no USP designation, are critical to most biotechnol- ogy operations, because recombinant molecules, cells, and tissues must be grown in basal media, typically enriched with a variety of natural or synthetic supplements (e.g., animal or human albumin and vitamins). Although manufacturing operations attempt to use only well- characterized or chemically simple materials, this is not always possible. In addition, it is sometimes necessary to use plant- or animal-derived materials to produce a biopharmaceutical. As with inorganic and organic chemicals, it is important to understand the nature and origin of each product used to grow and maintain cells or tissues. Quality control plays an important role in analyzing the nature of each raw material and ensuring that it is purchased and inspected carefully. Again, vendor-supplied CoA are scrutinized and, for some items, are tested again. Although synthetic and plant- derived natural supplements, such as vitamins or growth factors, are of moderate concern, animal-derived products are of great concern to both sponsors and regulators. This is because animal products can carry microbial toxins, animal viruses, or prions, and if present in a supplement, these agents could be transferred via the product to humans. Certificates of analysis are scrutinized, vendors are asked to certify the origin and microbial purity of such products, and vendor audits are commonly performed. In rare instances, such as with a special animal serum, it may be necessary to further process and test the product at the sponsor’s laboratory, or if a risk of disease trans- mission even exists, it may be necessary to simply find an alternative source or another supplement.
Quality Control and the Manufacturing Environment
As noted in Chapter 6, there is a great need to ensure a consistent and high- quality environment in the areas of a facility dedicated to aseptic manu- facture of a sterile product. One means of demonstrating compliance with
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environmental standards is to test swabs taken from personnel, equipment, or facility surfaces and also from samples of air and water. The QC laboratory is often responsible for this environmental sampling and testing. Samples are taken during periods without manufacturing activity (static environ- ments) and also during actual manufacturing operations (active environ- ments). Instruments are used to sample air or water, to count the number of nonviable particles, and to culture and count the colonies representing viable particles, that is, bacteria and yeast. Samples are also taken by swab- bing the uniforms and gloves of operators, as well as work surfaces, walls, and floors. This information is used to better maintain a clean environment and to alert or alarm the manufacturer whenever the aseptic or low-particle nature is detected or a work area has been compromised. Data are then plot- ted as a trend analysis, so that the staff may visualize the level of bioburden as it changes over time. This facilitates early action in the face of a developing problem. A trend analysis curve for environmental monitoring is shown in Figure 7.6. The QC laboratory may also be responsible for additional testing of environments in biomanufacturing, such as testing for residual product or cleaning agents, and using analytical methods to identify trace amounts of product or undesirable chemicals on work areas and process equipment.
10
9/2 4/2
00 2
10 /24
/20 02
11 /24
/20 02
12 /24
/20 02
1/2 4/2
00 3
2/2 4/2
00 3
3/2 4/2
00 3
4/2 4/2
00 3
5/2 4/2
00 3
6/2 4/2
00 3
7/2 4/2
00 3
8/2 4/2
00 3
9/2 4/2
00 3
100
1,000
10,000
1,00,000
Mean 0.5 Mean 5
FIGURE 7.6 Trend analysis. Plot of particles sampled in biomanufacturing room 121A, on specified dates in the years 2002–2003. Each data point in this chart represents mean value of actual particle counts (Chapter 6). A particle counter was used to measure the number of particles in a given volume of air and the data from multiple mean values (particle counts on x axis) were entered into a spreadsheet and then graphed with standard deviations (vertical lines). The upper line presents counts for particles of 0.5 µm or less diameter, and the lower line represents particles of 5 µm or less diameter.
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Qualification, Validation, and Verification of Analytical Methods
Validation was mentioned in Chapter 6 as an important aspect of bio- manufacturing. Validation or the related process of qualification are also performed on individual analytical tests. Typically, an assay is validated during the late phase of the development cycle. Assays are often quali- fied in mid phase, but some critical assays may be qualified even earlier in development, and these tests may also be validated in early phases. Critical assays are the ones that are important to the safety of the product or to ensuring potency before use in clinical studies.
Regulatory agencies demand that analytical tests be validated before licen- sure. The USP defines validation of an analytical procedure as “the process by which it is established, by laboratory studies, that the performance char- acteristics of the procedure meet the requirements for the intended ana- lytical applications” (United States Pharmacopeia–National Formulary 2016). The International Council on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use adds that validation must “ demonstrate that the (test) procedure is suitable for its intended purpose” (United States Pharmacopeia–National Formulary 2016). Validation of an assay also ensures the established specification is appropriate for a particular use and product. Although tentative or working specifications may be estab- lished long before assay validation begins, the validation process is a means of confirming or adjusting those specifications. Hence, assay validation and establishment of final specifications are highly integrated processes.
Verification applies to ensuring proper application and use of compendial assays. Specifically, verification documents that a laboratory is, indeed, per- forming an assay in the correct manner.
Assay validation is an experimental endeavor, and the process is always performed under a written and approved protocol, one designed to achieve specific purposes and perform exact experiments for each assay-product combination. The protocol states a purpose and the scope and provides pass- versus-fail rules for method validation outcomes. In general, the purpose is to prove that the analytical method can perform adequately as a written SOP for the intended purpose. The scope of use is also stated, along with the specification that is currently under consideration. A series of experimental procedures then challenge that assay to demonstrate suitability by a number of criteria or traits that are established before testing. The most commonly applied traits were briefly mentioned earlier, as they are important to con- sider well before validation begins. These are described below as:
• System suitability: This is the ability for an analytical system to achieve the objectives of the assay and is defined in several ways. First is the need to ensure that equipment is suitable for the intended purpose
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and is installed and operational (see Installation Qualification and Operational Qualification, Chapter 6). For some analytical tests using highly sensitive instruments, the conditions and settings are carefully defined. Reagents are shown to be suitable and reference standards and controls adequate and well characterized. Interfering substances in any reagent, including the sample matrix, are considered because any substance could artificially enhance or inhibit an analytical sys- tem. Sampling and sample preparation studies are completed to ensure matrix compatibility with the test method and other reagents. A system suitability report is prepared to demonstrate that the com- plete analytical system, including instrumentation, is suitable for the intended use and for further validation of the test method.
• Limit of detection and limit of quantitation: An early step in valida- tion of quantitative assays is determination of the lowest amount of analyte that can be detected, that is, the LOD. The amount that can be accurately measured in a quantitative assay is the LOQ, usually two values that bracket. For the r-protein CoA example of total protein, for BCA assay (Table 7.2) in which the specification is 1.0 ± 0.1 mg/mL, an LOD might be 0.05 mg/mL, whereas the LOQ might be 0.1–2.0 mg/mL of protein in phosphate-buffered saline. These values of LOD and LOQ would be experimentally deter- mined and would further ensure that actual test measurements are taken within the proper range of values.
• Linearity: For quantitative measurements, it is necessary to demon- strate linearity, the ability, within a given range of analyte in sample, to obtain test results that are directly proportional to the concentra- tion of that analyte. In other words, a linear curve must be generated within the dilution work area. Standards, shown to be of the high- est quality by other methods, are prepared at concentrations in a range and with a matrix that match that of the intended test sample. For example, if one expects the r-protein to exist in a sample of BS in phosphate-buffered saline at concentrations of 15 ± 5 mg/mL, then one might establish test dilutions of standard BS in phosphate- buffered saline from 5 to 30 mg/mL and then test them using the standard assay. Duplicate assays might be performed on each of five days and the results plotted as a linear regression. Acceptance cri- teria for this example is based on statistical analysis and might be a coefficient of determination, r2 > 0.98, and y-intercept of the lin- ear regression. Linearity demonstrates that the assay results may be extrapolated from the linear curve, within this range of values, and that this can be achieved repeatedly.
• Precision: Analytical precision refers to the ability of an assay to repeatedly produce the same or very similar measurements on repeated testing when variables are held constant. Precision and
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accuracy, described below, are visualized in Figure 7.7. Precision is referred to as the degree of scatter. Critical variables, such as mea- surement of sample volume or weight by a standard procedure, may need to be evaluated first to ensure that pre-analytical procedures are precise. Then the test is performed repeatedly in the same man- ner. A test is precise if the results have little scatter. Acceptance cri- teria for precision of an assay are usually given as the percentage relative standard deviation for a given number of sequential tests of the same sample. Along with robustness experiments, preci- sion validation ensures repeatability, intermediate precision, and reproducibility.
• Accuracy and range: Accuracy is a measure of how well an assay agrees with a known true value, as visualized in Figure 7.7. Put another way, it identifies the total error of the method, consider- ing both the systemic or inherent technical errors and the random errors for that test. Thus, range is the interval allowed between the upper and lower concentrations of analyte in the sample. In a practi- cal sense and for many assays, the acceptable range is demonstrated as an outcome of linearity testing. To validate accuracy and range by using the example of the r-protein, one might make several dilu- tions, within, and just below and above, the limits used to produce
Precision and accuracy
Precise Imprecise
Accurate
Inaccurate
FIGURE 7.7 Precision and accuracy. This figure depicts, as holes in targets, the concepts of precision and accuracy.
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the linearity curve. Each of these dilution points is repeatedly tested several times to produce multiple results at each dilution point. The linearity of the response is evaluated to determine whether, indeed, the linearity testing results are confirmed and to learn the actual range of acceptable accuracy values. The percentage relative stan- dard deviation would be calculated for each dilution point and com- pared to acceptance criteria, perhaps 95% to 105% of the theoretical value. Typically, in such plots of multiple dilutions, the accuracy falls outside these acceptance criteria, at dilutions above or below those determined for the linear curve. This further determines a range of values, calculated as baseline values, and here, as milligrams per milliliter of total r-protein, which then are used to accept samples for the assay. It also provides a percentage relative standard devia- tion for each dilution within this range, a value that can be applied to reference standards in the future.
• Specificity: Specificity is the degree to which the measurement is due to the analyte of interest and not due to other substances that could interfere with the assay or confound analytical results. Such substances might include components of the matrix, such as mac- romolecules or buffer salts, impurities or degradation products, or similar but undesired molecules. The assay is shown to exactly identify an analyte, to differentiate analyte from impurities, and, when desired, to provide an accurate or exact result related to other product attributes. Specificity validation requires input of these substances by spiking a known pure sample. Purity of a known sample, often the reference standard, is achieved by adding sub- stances that are expected to interfere with the assay and might be present in a sample. After multiple analyses of the various samples, reference and reference plus substance, or other substance alone, one can determine the degree to which the assay is specific for the intended analyte. Acceptance criteria might be given as a percent- age of the reference standard, such as ±10% of reference standard. Any values obtained for a sample outside the reference standard range or value would indicate undesirable interference.
• Robustness or ruggedness: This refers to the overall reproducibility of the test results obtained when aliquots from a homogenous lot of sample are analyzed under normal, expected operational condi- tions, given that even the most consistent conditions introduce small variations. Hence, variations in instruments, reagents, or test condi- tions are introduced during the experiments used to validate the assay. For example, a given reference standard might be tested under the same procedure by different operators using one instrument on the same day, might be tested using three different instruments but by the same operator on the same day, or might be tested using the
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same instrument and the same operator but on different days. An experimental matrix is developed and parameters are carefully cho- sen and then varied to ensure meaningful and affordable robustness testing in multiple experiments.
Some assays require limited time or effort to validate, whereas other assay validation protocols demand months of planning and experimentation and consume significant resources. For example, an HPLC assay of a well-char- acterized vaccine recombinant protein might be simple to validate, but an immunopotency assay of the same protein performed in rabbits to deter- mine the immunogenicity might require 12 months of effort and 10 times the resources. Assay validation must be considered for each assay and be carefully planned well in advance. Indeed, it takes much time and requires the input of many experts to develop a good validation protocol even for one test method. Validation applies to the assays used to test BDS and FDP, both for release and stability purposes. A proper assay validation tests multiple batches or lots of product, because consistency of results is important. In addition, fully quali- fied controls and reference standards are always used. The product, BS or FP, that is used in assay validation protocols is made by product manufacturing processes that are or that exactly mimic commercial procedures. Clearly, QC assay validation and manufacturing scale-up validation require a tremendous effort, concise coordination, and a significant investment of time and money. Owing to this, the QC plan must be carefully devised and the assays them- selves must be scientifically sound before the sponsor begins assay validation in late stages of the product development cycle.
Selected assays may be qualified before they are validated. Qualification is in many respects a mini-validation, as it focuses only on important aspects of assay validation and is performed under abbreviated protocols. In contrast to valida- tion, qualification is completed earlier in development with only those assays that are considered critical to demonstrating purity and potency or with those in which confidence is lacking because of their newness, uniqueness, or com- plexity. Qualification may also serve to establish product release specifications for critical attributes, as multiple lots of product are tested using qualified assay procedures. Another purpose for assay qualification is to give the sponsor confidence that an assay is predictive of product quality for use in early clini- cal trials. Results of qualification are predictive of validation. If an assay fails qualification, then it is a bad candidate for full validation; the consequences of assay validation failure can be great. Finally, qualification is sometimes recom- mended by a regulatory agency to a product in early development, so as to alleviate fears of using impure or subpotent product in clinical trials.
Assay verification refers to a process applied to commonly used assays, notably those published as a standard method in a pharmacopeia or other authoritative reference or regulation. Verification ensures that a method has been established correctly when adapted into a new laboratory. A compen- dial assay may be established in a biotechnology laboratory that has little or
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no experience with that test. In such cases, verification is a formal process, similar to qualification, in which the sponsor ensures proper performance and outcomes in the hands of less- experienced operators or at a new labora- tory. Assays such as sterility test (USP <71>) and endotoxin test (USP <85>) are candidates for verification, because they are critical to product safety yet already well characterized and have highly detailed standard procedures.
Application of Statistics in Assay Performance and Validation
Utilizing good statistical practices throughout the assay development and validation life cycle is important to ensuring correct performance while minimizing bias. Perhaps the greatest threat to proper test performance and interpretation of results is bias, a systemic distortion of results. Bias often appears unbeknownst to the QC scientist; indeed, this is inherent in the defi- nition. Factors generating or influencing bias must be identified; statistical analysis is an important means of detecting bias. In addition, statistics is key to correctly analyzing measurements, especially those considered a quanti- tative measurement of an important attribute.
Statistical analysis is important to the QC scientist, because quantitative or semiquantitative assays, and most potency assays and many purity tests fall into these categories and require comparison of results to a standard curve. This, in turn, requires constant calibration and ensuring linearity of these tests. Demonstrating a linear response ensures that results are meaningful and statistical analyses are applied to experimental results. For tests based on linearity analysis, the statistical methods chosen have a great impact on assay performance metrics such as accuracy or reportable range of values. Statistical tests are also applied to assay qualification and validation, and appropriate data analysis methods have been established for these endeavors.
Indeed, two metrics are considered for any assay, the measurements them- selves and the variability of those measurements. The measurement must be specific and accurate. Quantitative tests are either demonstrated to be linear or, in some assays (e.g., dose response), they are nonlinear but require curve fitting with a specific equation. Variability takes into consideration precision, range, LOD, LOQ, and robustness. For these, statistical rules are applied to interpretation of actual results. For example, assay precision is determined by calculating the mean, the standard deviation, and the variance or coef- ficient of variance. Certain statistical rules argue for focusing on the standard deviation and its corresponding 95% confidence interval and considering coefficient of variance of lesser importance. In the case of a potency assay for a biopharmaceutical product, in which there may be considerable inherent variance, such statistical rules must be considered in design, performance, and validation of the assay. Acceptance criteria are established only after
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careful statistical analysis of data generated by extensive use of the assay. Good statistical practices are seriously considered from the outset of QC planning and then throughout the QC cycle, because proper application of statistical methods to analytical endeavors leads to reduced development times, ensures that testing meets intended use, and prevents bias from enter- ing into any analytical test.
Trend analysis is an important management tool, which is applied to most test results in an effort to identify movement of values for controls, reference standards, and test samples. A trend analysis for environmental monitoring is shown in Figure 7.6. Data from a similar trend analysis of environmental monitoring are analyzed with statistics, first to establish real-time data alert and action limits, and second (Figure 7.8) to determine whether or not any sample exceeds these limits.
Summary of Quality Control
Quality control is a technical or laboratory function to ensure, in part, the purity, potency, and safety of biopharmaceuticals. Planning for and devel- opment of QC for a given product is based on the attributes of that product, because each test focuses on a particular attribute. Attributes are appear- ance, safety, identity, strength, purity, and potency. Certain analytical tools are available to sponsors at contract laboratories, but meaningful tests must also be developed specifically for each product. This is especially true for tests to measure purity and potency. A specification is established for each test, beginning with early manufacture, and each specification serves as a boundary to establish whether a product passes or fails testing. However, as data are generated, a specification may change during the development
Alert limit: 95% UCL = 10.860
Center line = 10.058
LCL = 9.256Q ua
lit y
ch ar
ac te
ri st
ic
9.0
10.0
11.0
Sample 3 6 9 12 15
UCL-action limit: 99.7%
LCL-action limit: 99.7% Alert limit: 95%
FIGURE 7.8 Application of statistics to trend analysis. Trend analysis data for an assay were statistically analyzed and then graphed for upper (99.7%) and lower (95%) confidence levels (UCL and LCL), on which action and alert levels, respectively, were established.
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cycle, as greater experience is gained on each test and with each batch or lot of product. This information—attribute, test, and specification—is written into a batch- or lot-specific document, referred to as a CoA, along with the specific test results. This certificate compares specification to actual result and is thus used to decide whether or not a batch or lot of product meets, by test results, specifications and hence whether it may or may not be released (pass or fail) for use.
A large number of analytical tools are available to QC scientists and many more are being developed each year. In QC planning, it is incumbent on the scientists to choose the correct tests to determine the quality of a product. Other considerations early in QC development are use of reference standards and test controls, samples and sampling, and the need to establish in-house, special tests that are not available elsewhere. Quality control tests are used not only to release product but also to measure stability of product after it has been transported or stored under various conditions. Hence, stability protocols are also developed for each product and for analytical methods included in those protocols. The QC laboratory is also responsible for moni- toring the environment of a manufacturing facility and operation and for testing raw materials to ensure that whatever goes into a product is of appro- priate quality. Assay qualification, verification, and validation are performed during the development life cycle to ensure that analytical tools perform as intended. Statistical analysis plays an important role in evaluating analytical data, both the performance of tests and the test results.
Reference
British Pharmacopoeia. 2016. Medicines and Health Products Agency, London, UK. Merck Index, 15th Edition. 2015. Royal Society for Chemistry. Cambridge, UK. Merck Manual (of Diagnosis and Therapy), 19th Edition, 2011. Merck and Company,
Kenilworth, NJ. Martindale: The Complete Drug Reference 38th Edition. 2015. Pharmaceutical Press,
Royal Pharmaceutical Society. London, UK. Physician’s Desk Reference, 69th Edition, 2015. Medical Economics Company, Inc.
Montvale, NJ. United States Pharmacopeia. 2015. United States Pharmacopeia. Rockville, MD. United States Pharmacopeia–National Formulary. 2016. General Information/<1225>
Validation of Compendial Procedures. In: USP39-NF 34 Page 1640. Pharmacopeial Forum: Volume No. 35(2) P. 444.
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8 Nonclinical Studies
Nonclinical Studies and Risk Assessment
The assessments of risk and benefit for any candidate biotechnology prod- uct are experimentally and progressively evaluated first in the laboratory, then in animal models, and finely in people. Specifically, this involves under- standing the nature of the biological construct or molecule, its purity, and its potency after manufacture, as well as the safety and efficacy profile. Nonclinical studies, performed in vitro and in animals, are primary means of measuring the potential product risk, and much of this testing precedes clinical trials. Results of nonclinical studies serve to better ensure that prod- uct benefit will indeed outweigh risk once it reaches clinical studies and the marketplace. Nonclinical study activities precede clinical research for good reason. It is the user, often times the human subject enrolled in a clini- cal trial, who bears the burden of risks associated with evaluating product safety. Thus, the sponsor of a novel biopharmaceutical provides clear experi- mental evidence that risks are tolerable and the product itself is unlikely to result in disease or death to the human subjects or, on marketing approval, to the public.
Information demonstrating safety and tolerability of a candidate biophar- maceutical is presented in the Investigational New Drug Application (IND), specifically in the Pharmacology and Toxicology section. Here, test results, that is, in vitro laboratory and animal test data, demonstrate in various ways both how the biopharmaceutical behaves in biological systems (pharmacol- ogy) and whether or not it is toxic (toxicology). Some pharmacology and toxicology test systems are simple and are applied to samples of the biophar- maceutical in a laboratory setting, using tests focused on answering a single question, for example, the mutagenic potential of a compound. Other tests are performed in appropriate animal models and these are supplemented with additional laboratory testing. An adequate and well-controlled panel of nonclinical studies, an example of which is shown in a general scheme in Figure 8.1, can demonstrate, beyond reasonable doubt, that the biophar- maceutical possesses desired pharmacological attributes and the levels of exposure at which it is safe, not toxic, and well tolerated in animals.
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Humans are exposed to many biotechnology products that are not bio- pharmaceuticals. Products used for environmental, industrial, or agricultural (including food) purposes are studied in formal toxicology tests, both labora- tory and animal. However, biopharmaceuticals are given the greatest safety scrutiny and testing because they are directly given to large numbers of humans, sometimes over long periods, and many are injected into the body.
Nonclinical testing of biopharmaceuticals has its foundation in the drug industry, where a general understanding and appreciation for the value of pharmacology and toxicology have led to successful development and
Pharmacology
Consider possible animal models
Consider possible in vitro and animal models
Select animal models
(1 or 2) Select in vitro
models
Select animal models
(1 or 2)
ADME and pilot studies 1. Formulation, dose, and route delivery 2. Metabolism or catabolism 3. Biotransformation 4. Tissue distribution
Acute toxicology Short-term dosing
In vitro toxicology
Assay development Measure product in tissue and blood
Special toxicology 1. Tissue binding 2. DNA integration
Definitive pharmacokinetic and pharmacodynamic studies
1. Cmax 2. Tmax 3. Concentration effect
Subchronic toxicology
Chronic toxicology 1. Long-term dosing 2. Reproductive 3. Immunotoxicology 4. Neonatal toxicology
TPP Toxicology
1. Medium-term dosing 2. Local tissue 3. Route delivery
FIGURE 8.1 Scheme of nonclinical activities in biopharmaceutical development. The targeted product pro- file (TPP) is instrumental in developing a nonclinical plan. Both pharmacology and toxicology studies are performed to identify a safe and effective dose. Assays are developed to measure product and metabolites in blood and tissue, and appropriate animal models are used in vari- ous types of studies.
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marketing of small molecule drugs. Studies of dose-response, pharmacoki- netics, and pharmacodynamic relationships, and toxicology, as well as the development and application of in vitro laboratory tests and animal models are the scientific tools that are routinely used by scientists and described in this chapter. However, because drugs and biopharmaceuticals differ in many respects, the panel of tests required for biopharmaceuticals is often unique, even if the basic principles are the same. Consider that biotechnol- ogy products are typically large molecules, living cells, or microbial products and have unique patterns of biodistribution and distinctive toxicities. Drugs are usually small organic molecules. Biopharmaceuticals do not always lend themselves to testing in traditional in vitro tests or animal models that have been developed to assess the safety of drugs. Nonclinical biopharmaceutical scientists apply knowledge and experience borrowed from the small mol- ecule drug industry, but they have also developed unique methods to effec- tively study the pharmacology and toxicology of biological molecules and cells. Further, as compared to testing drugs, pharmacology and toxicology testing of biopharmaceuticals often requires unique and expensive tests and development of applicable animal models.
Biopharmaceutical Delivery, Pharmacokinetics, and Pharmacodynamics
Product Delivery to the Body
Biopharmaceuticals pose unique formulation and delivery challenges because of their large size, complex structure, and vulnerability to degra- dation. Many injectable formulations are difficult to administer because of high viscosities associated with concentrated formulations that are often encountered when doses reach hundreds of milligrams required in the final formulated product. Most biopharmaceuticals and drugs are transferred from the final container, such as a vial or a syringe, to an initial target tis- sue, and only then, it is distributed to the target organ or tissue, where it has the intended therapeutic effect. There are many ways to achieve this objective, some of which are listed in Box 8.1. Many drugs are given orally because they are taken up in the digestive tract without first being metabolized. Oral presentation is rarely the case with biopharmaceuticals today and most are given parenterally. Products given intravenously are designed and intended to be distributed throughout the body very rap- idly. Other parental routes of delivery are intravenous, subcutaneous, and intramuscular. Monoclonal antibodies and therapeutic proteins are often given by one of these routes. Vaccines are usually given subcutaneously or intramuscularly but some are given intranasally and others intradermally.
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Certain cellular therapies are delivered parenterally, often by direct injec- tion into a target organ or tissue. Oral ingestion is by mouth, but in this case, the biopharmaceutical must be specially formulated, so that gastric and intestinal acids and enzymes do not degrade the product before it crosses the intestinal or gastric mucosa. In addition, special consideration is given to the size of a molecule taken orally, as large molecules such as an anti- body would not be readily absorbed in appreciable amounts. Pulmonary delivery for lung absorption is sometimes applied to smaller biomolecules such as insulin. In addition, topical application of biopharmaceuticals is another route of delivery and is exemplified by transcutaneous delivery
BOX 8.1 ROUTES OF ADMINISTRATION TO ANIMALS OR MAN FOR A BIOPHARMACEUTICAL
1. Parenteral (injected) a. Intravenous b. Intra-arterial c. Intramuscular d. Subcutaneous e. Intradermal f. Intracardial g. Intraocular h. Intraperitoneal i. Epidural 2. Oral 3. Inhalation
4. Body cavity
a. Intranasal b. Sublingual c. Rectal d. Intravaginal e. Intrauterine f. Intraurethral g. Intra-auricular 5. Topical a. Percutaneous (transdermal) b. Cutaneous c. Ophthalmic
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of vaccines or therapeutic peptides. In development are a host of special delivery methods for biopharmaceuticals such as patches, microneedles, and special injection devices (e.g., pumps). Additional delivery methods that require adjustments in formulation are being developed to enhance in vivo accessibility; these include the use of controlled-release preparations (e.g., microspheres and microparticles), protein modifications (e.g., albu- min fusion), and genetic manipulations (e.g., site-directed mutagenesis).
Adsorption, Distribution, Elimination, and Metabolism (ADME)
Once a biopharmaceutical has crossed all barriers and is in the blood, it must reach a target organ or tissue, an exact location to produce its thera- peutic effect. For some biopharmaceuticals, this step can be challenging. It must be absorbed, usually into the blood stream and remain stable if it is to be distributed. A method of delivery may fail to achieve this objective. For example, many therapeutic monoclonal antibodies are injected into the subcutaneous tissue or muscle, even though the target organ, for example, rheumatic joints, is some distance away. The biopharmaceutical must be distributed and absorbed in adequate amounts, before it results in local reactivity or is metabolized or otherwise eliminated. Biopharmaceutical products are, therefore, designed to be absorbed, distributed, and then metabolized and eliminated (but not too rapidly). These functions, known as ADME, are studied and reported for each biopharmaceutical prod- uct because these are of critical importance to success in clinical trials. Figure 8.2 outlines the possible tissue relationships between each of these functions.
Absorption
How is absorption defined for a biopharmaceutical? Oral-gastrointestinal absorption is unlikely, because most biopharmaceuticals, composed of pro- tein, cells, RNA, or DNA, are recognized as just another food substance, and thus, the gastrointestinal juices and enzymes digest them. In addition, few biopharmaceuticals might be absorbed across the skin (topical) or mucosal surfaces (transmucosal), because they are simply too large to diffuse intact across such barriers. Hence, most biopharmaceuticals are given by the paren- teral route, which means that they are directly injected into either the blood stream or a tissue. From here, biopharmaceuticals either have a local effect or enter the blood or lymph systems, facilitating distribution to other tissues.
Distribution
After application of a product, it is very important for the biopharmaceuti- cal to be distributed to the tissue or organ where it will have the greatest therapeutic effect. It should not reach tissues, or build up, creating reservoirs,
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where it may be toxic. It is also critical to sustain a certain level of biophar- maceutical in the blood and tissue. Many biopharmaceuticals reach a state of equilibrium on reaching the blood stream; whereas for other products, there is a rapid drop in circulating levels after injection. The pharmacologi- cal and toxicological implications of parenteral delivery may be considerable, because a biopharmaceutical in the blood is rapidly distributed, allowing exposure to many organs and tissues, and not only to the target tissue or organ. Alternatively, a biopharmaceutical may be injected into a firm or semisolid tissue, such as subcutaneous, where it resides in a depot and is slowly released into and distributed by the blood stream. For some products,
Animal model development
Biopharmaceutical injection of product
(e.g., parenteral)
Heart Lungs Kidneys Brain
Available, free product
Metabolism
Tissue bound Circulation Biotransformed
FIGURE 8.2 Outline of adsorption, distribution, metabolism, and excretion (ADME) studies. Using animal models and sensitive and specific assays, product is measured in various tissues and blood. Additional assays are used to measure metabolites.
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this can result in sustained levels of biopharmaceutical, with blood absorp- tion and then distribution occurring over a longer period, as compared to an intravenous injection approach.
However, for other biopharmaceuticals, it is not desirable to distribute the product to certain tissues, as the molecule might be toxic to certain tissues or organs. Some molecules tend to accumulate in one or another tissue or cellu- lar compartment, a reservoir; this may be desirable or it may lead to toxicities. For example, a product could rapidly accumulate in the liver, where it may be hundreds of times more concentrated than in other tissues. If the product is therapeutic in the liver, then it may be best to have that biopharmaceutical largely concentrated right there and unequally distributed in the body. In contrast, if the product is toxic to liver cells at high concentrations, then it is not good for the biopharmaceutical to accumulate there. Sometimes, it is best to avoid the blood stream, when possible, and deliver the biopharmaceutical directly to the target organ. However, this can be challenging with certain types of products that target hard to reach organs such as pancreas or brain. Biodistribution studies to determine these pharmacological parameters are critical to understanding pharmacology of a product. In addition, informa- tion gleaned from distribution studies is used to support the development of new formulations and delivery methods aimed at improving the therapeutic value and reducing the toxicity of a biopharmaceutical.
Metabolism and Biotransformation
Biopharmaceuticals eventually change in the body to another form and become metabolically inactive through normal processes such as enzymatic degradation. A few biomolecules, such as the DNA in a genetic therapy or a pluripotent-cell-derived product, may not follow this rule, because they are developed for the purpose of longevity in the body. Yet, biotransforma- tion, a term used to describe any biological process that converts the original product to another molecular format, is the rule that applies to biopharma- ceuticals. In some cases, biotransformation enhances the therapeutic activity, whereas in others, it decreases, limits, or terminates the biological activity. Physiological, genetic, and environmental factors may be, and often are, involved in biotransformation. Although we can establish the average time of biological activity in a given population, it has been nearly impossible to reliably predict, for a single individual or animal, how long a particular biopharmaceutical will remain active. Living organisms are quite diverse, when it comes to processing biopharmaceuticals. Further complicating the picture, the coadministration of two compounds can have unexpected effects, because metabolic drug interactions are possible. Drug interactions can impact absorption, distribution, pharmacokinetics, metabolism, or excretion, and many patients take two or more drugs or biopharmaceuticals. Metabolism and biotransformation studies can assist in understanding the overall pharmacological profile of any product.
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Excretion
Clearance is a process in which a biopharmaceutical is eliminated from fluid phases, tissues, or organs. With most biopharmaceutical products, clearance is expected to take place through the processes of metabolism and excretion, but first, the molecule must remain in the target tissue or organ long enough for it to have a therapeutic effect. Excretion cannot be too rapid. With many small molecule drugs, the absolute rate of clearance is a linear function of the concentration in blood. However, with biological molecules, this is not always the case, and the rate of clearance is not simply the rate of elimination divided by blood concentration. In addition, while small molecule drugs are often cleared by liver and kidney, larger biological molecules are not often metabolized in the liver and are retained, not excreted, as they pass through the kidney. For many biopharmaceuticals, the sites of metabolism and excre- tion are unknown, and it is assumed that components of degraded biophar- maceuticals, such as polypeptides, amino acids, and nucleic acids, are simply catabolized to a certain degree and then used by the body to produce energy and to build other macromolecules.
Pharmacokinetics and Pharmacodynamics
The science of pharmacokinetics attempts, for a given biopharmaceutical, to understand ADME and to explain the outcomes that follow dosage of that product. Pharmacokinetics is the study of complex interactions that fate between an active compound and the cells, tissues, and organs of the body. Only certain aspects of pharmacokinetics are discussed here. Pharmacokinetic studies for pharmaceuticals and drugs differ in many respects, because small molecules and the large molecule biopharmaceuticals are often quite differ- ent in biological properties and mechanisms of action.
However, some rules of pharmacokinetics do explain the behavior of many biopharmaceuticals. For example, with specific and sensitive analytical tools, we can measure the maximum concentration of a biopharmaceutical that is reached after a certain dose is given by the route of injection. This value is called the maximum concentration or Cmax and is shown in Figure 8.3. From pharmacokinetic studies, we also determine the amount of time it takes from injection of a biopharmaceutical, until Cmax has been reached. This is referred to as Tmax, also shown in Figure 8.3. From the same experiment, it is possible to measure the half-life of the biopharmaceutical in the blood, and this is referred to as t1/2. Half-life is a derived value based on both clearance and volume of distribution. The period from injection to Cmax is called the absorption phase. Although this may be a very short time for biopharma- ceuticals given intravenously, it is an important parameter for products that are injected into subcutaneous or other tissues. The period beginning at Cmax and lasting until all product has been eliminated from the blood is called the elimination phase. If a product is given in multiple injections, then the blood level rises, until it reaches a plateau, referred to as the steady state for that
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dose and dosing regimen. These rules do not apply to some biopharmaceuti- cals. For example, vaccines seldom reach the blood in appreciable quantities, and it is difficult or impossible to measure small amounts of recombinant proteins or live cells in a solid tissue.
Clearance, discussed above, is an important aspect of any pharmacoki- netic profile. The apparent volume of distribution is another parameter; it is abbreviated as V (volume of distribution) and is equal to the amount of bio- pharmaceutical administered, divided by C, the concentration of product in drug or plasma. This value varies widely, depending on the amount of tissue binding and the degree to which the product is hidden by or binds to other materials. Using real-time models in animal or human studies and assuming that assays are available to measure a biopharmaceutical in blood, it is pos- sible to measure plasma concentration time curves for a product.
Bioavailability measures the amount of biopharmaceutical that is avail- able for use by a tissue at any given time. For most products, bioavailability is maintained through multiple doses; this keeps biopharmaceutical levels at a reasonably constant, albeit fluctuating (within a range of values), level in blood or tissue. Controlled bioavailability means that the product is con- sistently available to the patient’s tissue and ensures therapeutic effect at all times. Each of the factors—absorption, distribution, metabolism, and excretion—has a great impact on bioavailability, as do calculated values such as Tmax, Cmax, and t1/2. For successful therapy with many biopharmaceuticals,
2
5B lo
od c
on ce
nt ra
tio n
of p
ro du
ct (μ
g/ m
l)
50
100
4 6
Tmax
Cmax
t1/2
8 Time (hours)
10 12
FIGURE 8.3 Example of biopharmaceutical concentrations in blood over time. After injection of a biophar- maceutical at Time 0, the concentration rises in blood, until it reaches a maximum concentra- tion (Cmax) at the fourth hour (Tmax). The concentration then falls because of metabolism and excretion, until one-half the maximum concentration is reached at the tenth hour, resulting in value of t1/2 for the period.
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pharmacokinetic experiments develop information that, in turn, allows one to design and optimize delivery methods and dosing regimens, based on the desired effect and the amount of product available to produce that effect.
This might sounds logical, even simple, but, in fact, deriving these val- ues is a very complicated process, made more difficult by the lack of ade- quate animal models in which to study most biopharmaceutical products. Nonetheless, the product development program must take this information, generated in animals, into consideration as the target level, maintenance dose, loading dose, and individualized dose are calculated for man. Often times, the human target dose calculated from pharmacokinetic studies in animals differs significantly from the target dose estimates made, in the absence of experimental data, in the targeted product profile (TPP). For large differences, it is wise to ask why this happened and perhaps do further phar- macokinetic experimentation. Each biopharmaceutical molecule is unique, in molecular characteristics, how the product is dosed, and the indication. For example, the loading dose of a gene therapy might be high and without maintenance dose, whereas a monoclonal antibody to treat a chronic disease might require a specific dose through years of treatment, without the need for a higher loading dose at the onset of therapy. The possibilities are end- less and must be experimentally determined for each product and intended clinical indication.
In summary, the experimentally derived target level is simply the amount of biopharmaceutical that, hypothetically, will produce the desired effect when given in a particular formulation and route of delivery. The loading dose is the amount of product that will be given at the initiation of therapy to rapidly achieve the target level. The maintenance dose is the amount that must be given at set intervals after the loading dose, to maintain the target level.
In addition, speaking of the future, many traditional methods used to measure pharmacokinetic parameters in small molecule drug studies have not always been successfully applied to experimental pharmacokinetics of biopharmaceuticals. Given that necessity is the mother of invention, there are new methods that seem quite relevant for measuring various parameters such as biodistribution and clearance of biological products. Optical imag- ing using bioluminescence and dyes and variations of computer-assisted tomography are promising in this respect. Future studies of biopharmaceu- ticals will certainly be more informative; however, the measured endpoints might seem quite different from those of small molecule drugs.
The science of pharmacodynamics studies biochemical and physiological effects and the mechanism of action of biopharmaceuticals. The concept of drug- receptor interactions underlies most pharmacodynamics for small mol- ecules. Since most biological products exert their effects by interactions with molecular or cellular components of an organism, pharmacodynamics is also important for the development of biopharmaceuticals. To properly progress a product to clinical development, a scientist should have some idea of how
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a therapeutic effect is generated within a complex organism. For some bio- pharmaceuticals, such as a monoclonal antibody targeted to the receptor of a particular cell type, therapeutic effect might be well known from discovery research. Indeed, designer molecule biopharmaceuticals are developed for a specific purpose, such as binding to a receptor having a known physiologi- cal function. For other products, such as most recombinant protein vaccines, the effectors’ mechanism remains unknown at the time it is first tested in man. Some classes of molecules, often times given in one or a few doses, are never completely understood, because their exact mechanism remains in a mysterious and seemingly complex black box, even after market approval. Therefore, many biopharmaceuticals are an exception to the recommen- dation of clearly understanding the mechanism of action before using the product in man. However, attempts are still made to understand pharmaco- dynamics of a biopharmaceutical, and the information derived from these experiments is applied to designing pharmacokinetic and nonclinical safety studies and estimating clinical doses and dose regimens.
What types of pharmacodynamic information can be derived for a biophar- maceutical during nonclinical development? It is important to understand the relationship between the concentration of a product and the magnitude of the response to that biopharmaceutical. However, the response may be complex and even unpredictable in some individuals, animals, or man. A concentration-effect curve can be constructed. As shown in Figure 8.4, the experimentally derived information provides a wealth of information regarding pharmacodynamic properties of a biopharmaceutical. Potency is that part of the curve where an effect can be measured. Potency is clearly
Maximal effect
Potency range
Concentration of product
In te
ns ity
o f e
ffe ct
No effect
FIGURE 8.4 Concentration effect curve for a biopharmaceutical. The intensity of effect is proportional to the blood (or tissue) concentration of the biopharmaceutical. By measuring physiological effect, the potency range is determined, as are concentrations, with no or maximal effect.
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based on the concentration of the biopharmaceutical but can be quite variable within a given population. Maximal efficacy is that amount of biopharma- ceutical that produces maximal effect in that individual or in a population. There is also a slope to the concentration-effect relationship, and this reflects the mechanism of action of the biopharmaceutical and is seen in data from a population of animals or humans. Finally, there is biological variability, seen as standard deviation from the line traced by the population value. Much individual variability is due to genetic and other factors. At any given point on the curve, this can be significant for some products and is referred to as the individual effective concentration.
However, there are caveats to pharmacodynamic study results when bio- pharmaceuticals are tested. The standard concentration-effect relationship for a given biopharmaceutical considers a normal population, matched by age, sex, disease status, and so on. As organisms (humans) age, our response to a given biopharmaceutical changes and the concentration-effect relation- ship of an aged population may look quite different from that of a young adult population. The same could be said for populations composed of infants, children, adolescents, and those with certain underlying diseases. Pharmacodynamic variability demands that a biopharmaceutical developer understand the kinetics and toxicity of each product and carefully consider every population it might intend to treat. However, this is not an easy task. Pharmacodynamic variability, the individual variation in response to a bio- pharmaceutical, based on the mechanism of action, is an important issue in product development. This variability is a factor even after extensive phar- macokinetic and pharmacodynamic information is experimentally derived from a population of animals or human volunteers.
Bioequivalence is a term that, in its purest definition, suggests that two dif- ferent biopharmaceuticals have an equivalent effect. Stated in another way, the concentration-effect relationship of two molecules is very much alike. Consider two superimposed concentration effect lines, as pictured for the single line in Figure 8.4. However, bioequivalence also has other, sometimes more practical, meanings. It can mean that an active ingredient has the same effect even when formulated in two different ways or that one biopharma- ceutical has the same effect when given by two different routes of injection. If a reliable model with minimal variability can be developed and applied, bioequivalence testing can be an important aspect of nonclinical develop- ment, as optimal formulations, routes of delivery, and other variables may be tested, first in animals and then in man.
Bioavailability, first introduced under the pharmacokinetic discussion, above, is also relevant to pharmacodynamics. Defined in pharmacology as the fraction of the total amount of biopharmaceutical given (available to systemic circulation), bioavailability has additional meaning for certain bio- logical products that are given to produce a local effect and that rely little on blood concentration. For example, a monoclonal antibody given intra- venously has, on injection, 100% bioavailability to the circulatory system.
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However, if the same product must reach a tumor mass to produce an effect, then it most likely has a much lower percentage of bioavailability where it counts, that is, within the tumor. Experimental measurements of pharmaco- dynamic properties of this monoclonal antibody are not meaningful, unless pharmacokinetic information is available and considered. Thus, the effect of bioavailability on bioequivalence is carefully considered with every new biopharmaceutical and for each new indication for existing products.
For this and other reasons, biodynamic experimentation is challenging for many biopharmaceuticals. Consider a few examples. A gene therapy should replace a receptor, missing from birth, on a particular type of cell. This might be achieved by inserting into host cells the gene for a molecular analogue or by adding a (pluripotent-derived) cell that expresses the receptor. In either case, the therapy replaces the missing activity. But how does one measure, in a manner that is meaningful to the human situation, the pharmacodynamics of either therapeutic approach? How does the biopharmaceutical scientist determine which of the two approaches might be most successful at replac- ing the receptor, and how is this tested in a whole body situation? Some would take an experimental approach in an animal model, whereas others would argue that animal studies are not relevant and the product should forgo animal studies, and instead, pharmacodynamics should be consid- ered first in human studies (and, of course, after safety studies have been completed).
In a second example, the pharmacodynamics of a monoclonal antibody are unknown. The antibody could directly bind and neutralize a molecule excreted by a cell, or it could bind to a cell receptor and reduce the excretion of the same molecule. How does one approach pharmacokinetic studies in this case? Are the studies best done in animals, or should they be performed in Phase 2 human clinical studies?
Application of Pharmacokinetics and Pharmacodynamics in Biopharmaceutical Development
The pharmacokinetic and pharmacodynamic properties of each biopharma- ceutical present important information, because this information is applied to various decisions, including the selection of an efficacious dose and route of delivery, to ensure that the new product is tested for safety at a correct dosage. Indeed, nonclinical studies should demonstrate that new products have little chance of causing unexpected and undesirable effect when given to humans in subsequent clinical studies. If this is so, then it is necessary to understand the pharmacokinetics and pharmacodynamics of the product before embarking on an extensive program of safety testing in animals and certainly before introducing the biopharmaceutical into man.
As suggested earlier, certain tools are required for pharmacokinetic and pharmacodynamic development. One important evaluation is to directly measure pharmacokinetics and pharmacodynamics in an applicable animal
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model. A representative animal model may be available from studies of other products for a similar clinical indication. However, no matter how well proven the animal model is for another particular disease or class of product, it is impossible to know if that model will be applicable to a particular bio- pharmaceutical or its indication. For some biopharmaceuticals, it is difficult to even begin the process, because of inherent biological complexity of the product, animal physiology, potential immunogenicity issues associated with a xenogenic environment, and the ability to mimic the human clinical disease or underlying pathophysiology. For example, some of the most common bio- pharmaceuticals, such as vaccines and genetic therapies, are very difficult to classify, in part, because of the black box or great unknowns concerning the exact mechanism of action. In addition, we often lack a full understanding of pathogenesis of the disease that is to be prevented or treated. In the example of a recombinant protein used as a vaccine to prevent an infectious disease, we commonly lack knowledge on how the infectious organism is pathogenic and how exactly the immune system of our body fights that disease.
Still, it is incumbent on the product developer to use available scientific information and attempt to understand the pharmacokinetics and pharma- codynamics of each new product. With proper methods to detect active bio- pharmaceutical cells or cell receptors in blood and tissues, pharmacokinetic and pharmacodynamic measurements can be made in animals. Using more than one animal species, single-dose experiments can yield information on ADME, preferred route of delivery, optimal dose, dose linearity (Figure 8.3), concentration-effect (Figure 8.4) interspecies differences, metabolism, and excretion. Multiple-dose pharmacokinetic and pharmacodynamic studies can then be initiated or can be performed in conjunction with toxicology studies.
In addition to studies in animals, in vitro studies are often helpful. For example, it is informative to determine whether a candidate therapeutic monoclonal antibody binds specific cell types, and this might be achieved by using human cells, derived from various tissues, in culture. In addition, tests to study intracellular metabolism of compounds by using cell and organ cultures are available. In vitro methods to screen for induction of immune responses, again using cell or organ cultures, are also available.
To begin nonclinical experimentation, the biotechnology firm must have a formulation (Chapter 6) for each candidate product, and there must be enough material to allow extensive testing in the laboratory and in animals. This is sometimes referred to as the optimized clinical formulation, which means that it is the formulation of the product intended for Phase 1 clini- cal studies. Clinical quality or comparable quality product is essential for these studies to demonstrate the nonclinical safety that represents the same product intended for use in human clinical studies. A decision on how the product will eventually be administered to humans must also be made. This brings to mind the need to have an animal model to evaluate alter- native delivery methods, thus generating scientific data that will be used later to support and justify the delivery method selected for human clinical
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studies. A pharmacokinetic and pharmacodynamic program should also be designed to meet current regulatory requirements. Finally, there must be a precise means of measuring the product, as it exists in a matrix such as ani- mal blood or tissue. This means extensive analytical support (Chapter 7) to detect and measure exactly the biopharmaceutical of interest.
Results of pharmacokinetic and pharmacodynamic studies are carefully examined. Of particular importance are findings with safety implications, such as undesirable localization, notably vital organs or tissues (e.g., the central nervous system, the heart, and kidneys), of therapeutic molecules or cells. Such information is then applied to the design of toxicology studies and to moni- tor safety of subjects in human clinical trials. Nonclinical data also provide a foundation on which to establish rational toxicological studies to support clinical trials. It gives clues to development or application of the best animal models or future pharmacokinetic, pharmacodynamic, or toxicology testing. Specifically, study data may point toward an animal model that mimics the sit- uation expected in man. For example, if a monoclonal antibody was expected to have a therapeutic effect only if it remained in the human body for at least 2 weeks, it would be unwise to use, for toxicology testing, an animal in which the same molecule was undetectable 1 day after it was injected. The same can be said for the route of delivery. If a vaccine is to be given intramuscularly to man as 200 µg in a volume of 1.0 mL, then one would not choose a mouse model, because it is impossible to put this volume into a single mouse muscle.
Pharmacodynamic and pharmacokinetic studies often lead a developer to change formulations according to the anticipated or most readily available format. If a therapeutic DNA molecule did not remain, as required, at the subcutaneous site of injection for 24 h in an animal model, then it might need a new formulation, one that resulted in a depot effect to enhance longevity in that tissue. Improvements in dosing, based on data from well-designed phar- macokinetic studies, can be another positive outcome. Hence, well-designed nonclinical studies of biopharmaceutical absorption, distribution, metabo- lism, and excretion typically provide valuable information that allows for improved and more efficient safety assessment studies and may also provide a basis for the mechanism of action.
In conclusion, pharmacokinetic and pharmacodynamic studies are per- formed, with careful planning, as a series of experiments and with close coordination with other functional area experts, notably individuals from man- ufacturing, regulatory affairs, quality assurance, and clinical studies. Primary pharmacokinetic and pharmacodynamic effects are studied in animal models by using a variety of in vitro (or laboratory) methods. Experimental design focuses on the specificity of biopharmaceutical activity. If at all possible, levels of biopharmaceutical should be tested in humans, based on the information derived from these studies. For many types of cells and molecules, it is impor- tant to determine where the molecule goes within the animal, to define tissue or cellular interactions, and to identify how long it remains in any given loca- tion and how and where it is metabolized or excreted.
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Safety Assessment of Biopharmaceuticals
Toxicology
Toxicology is a science, specifically the study of adverse effects of agents— physical, chemical, and biological—on living organisms. Since any molecule can produce adverse effects, toxicology is important to all biotechnology prod- ucts, not just to biopharmaceuticals. This science covers acute, chronic, and long-term risks and uses a variety of established methods, many of which are biological. Toxicology assesses risks, that is, the probability of adverse events, caused by such effects. Toxicological studies go beyond measuring risks. These studies provide data that determine the possible causal relationships and help to establish limits of safety and design rationale and safe clinical studies. This discussion covers general approaches to toxicology while using examples derived from the safety testing of biopharmaceutical products.
The term toxicology immediately brings to mind chemical toxins, acids, bases, or organic solvents created by man for the purpose of producing other chemicals or lifestyle products. It also conjures images of physical agents such as ionizing and nonionizing radiation and ultraviolet light. Further, we consider the target of these agents to be a biological system, such as plants, animals and, most notably, humans. Chemical and physical agents can cause damage to living organisms. Drugs and medical devices are consid- ered chemical agents and physical agents, respectively, and for decades, they have been studied for toxic effects; many prove to be nontoxic but a surpris- ing number are toxic, reflecting a range of toxicities from mild to severe. Toxicology studies are routinely performed on drugs and medical devices by using both laboratory (in vitro) and animal (in vivo) methods.
In the recent past, a third type of agent, that is biological, has been added to the list of potential toxic agents. We have been aware of natural toxins, for example, snake venoms and poisons from plants, but have not had, until recently, the ability to manipulate the structure and function of biological molecules or cells and then use them to prevent or treat disease. With the advent of biotechnology, scientists began to develop biopharmaceuticals. These compounds, produced by man (or at least designed by man) and some being unique in nature, are intended for human exposure, sometimes repeat- edly and over long dosing periods. Further, biopharmaceuticals are designed to change the physiology or biological status quo of the user. However, changing the physiological balance for the better in some ways can also have undesirable effects in other ways. In addition, a recombinant molecule or cell that produces a desirable effect in one tissue might cause an undesir- able, or even toxic, effect in another tissue or organ of the same individual. Hence, with the advent of biopharmaceuticals, it became clear that each mol- ecule would be subjected to toxicology testing in the same manner as drugs and medical devices. Clearly, biological substances could cause undesirable
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effects by interacting with cells or tissues, as much as chemical or physi- cal agents. Biopharmaceuticals, if used properly or improperly, in excessive dose, or at extreme exposure, can harm body structures or processes and some might even pass these effects on to subsequent offspring. Thus, each biopharmaceutical must be studied to evaluate toxicity, or the potential to be toxic, when used in a particular manner for a given indication.
Design of a Safety Assessment Program
An effective safety assessment program must be carefully planned. Elements of planning for biopharmaceutical safety studies are outlined in Chapter 1. In this chapter, we delve into some factors that influence biopharmaceutical toxicity, discuss the tools used in these studies, and consider common study designs. The toxicologists have, or should have, four assets at their disposal. These are as follows:
1. Scientific and design precedence established, over decades, for a host of biological, chemical, and physical agents
2. In vitro methods to serve as rapid screening tests 3. Animal models and the ability to test agents in these complex
organisms 4. Established testing procedures to include acute, subchronic, chronic,
reproductive, carcinogenic, local tissue, immunological, and respira- tory toxicological protocols
The key to completing a meaningful safety program for a new biopharma- ceutical is to use these tools wisely under a product-specific and indication- driven experimental strategy.
The nonclinical plan is thus based on elements of TPP, notably the intended indication or disease, and intended product safety profile, as developed for the candidate biopharmaceutical. Consider that some biopharmaceutical products such as a therapeutic for a terminal illness, for example, metastatic cancer, have a very different profile from a product such as a vaccine that is intended to prevent a nonlife-threatening disease in infants. The plan is also based on an estimate of the clinical dose and dosing schedule, as provided in the draft clinical plan. From this information, and relying on experience with similar products, on regulatory guidelines for the class of biopharmaceutical, and on any available research data, the nonclinical professional can outline the intended approach for safety testing. Using a recombinant therapeutic protein as an example, Figure 8.5 presents a general scheme for nonclini- cal testing of a biopharmaceutical by phase of development. It demonstrates how the flow of events in a safety testing program structures a tiered test- ing approach. With each tier or ascending phase of clinical development, the product is used in both greater numbers and a more diverse population of
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individuals. This, in turn, can demand more detailed and stringent safety testing before each clinical phase.
The earliest nonclinical testing focuses on understanding the pharmacoki- netic and pharmacodynamic properties of the biopharmaceutical, as noted above. With this information, the intended human dose or doses can be esti- mated for Phase 1 clinical study. Toxicity testing applies the intended clinical doses and dosing regimen to the design of nonclinical studies that are com- pleted and reported before filing an IND or initiating the first clinical study. These early nonclinical studies include acute, subchronic, or other types of studies that may be considered by a sponsor or might be required by regula- tory agencies before initiating Phase 1 study with this class of product.
Subchronic and even some chronic testing may be required before enter- ing Phase 2 human clinical studies. The route of delivery or the formulation may be adjusted, based on Phase 1 study results, and therefore, additional acute testing may be necessary or local tolerance testing may be advised. In addition, because mid-stage clinical trials may expand into previously untested human populations (e.g., women of childbearing potential or indi- viduals with tumors or an underdeveloped immune response), it is wise to consider specialized toxicity testing before initiating these clinical studies. Since Phase 3 studies result in testing in a more diverse and much larger population and involve doses given over longer periods of time, it may be advisable to complete chronic toxicity studies at this stage of development. Besides, at Phase 3, the dose and dosing regimen would have been estab- lished, thus reducing the risk of having to repeat long, costly chronic toxicol- ogy studies. On the basis of the intended population and use, it is also wise to consider and plan for applicable specialty studies, such as those directed toward an organ system (e.g., reproductive toxicology, neurological toxicol- ogy, and immunotoxicology studies) and those focused on a product-related issue (e.g., tissue-binding or DNA integration studies).
The planning process considers current regulatory guidelines, both national (FDA) and international. It is important to consider that regulatory agencies have responsibilities toward the safety and welfare of human sub- jects who take investigational products and toward public health regarding marketed products. Nonclinical safety testing plays a major role in meeting these responsibilities. The International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) (Chapter 4) provides excellent guidance with regard to nonclinical testing strategies, and this has the advantage of worldwide harmonization. In addition, regulatory agencies ensure that adequate and well-controlled nonclinical studies have been completed according to performance stan- dards (e.g., current Good Laboratory Practices [cGLP]). Studies always meet generally accepted scientific guidelines as well. This means, by way of exam- ple, using experimental designs that test well-considered hypotheses, testing adequate numbers of the correct animal species, dosing only with mate- rial that matches the quality of product to be used in clinical studies, and
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applying proper statistical tests when analyzing results. Conclusions drawn in nonclinical study reports must be supported by data.
In Vitro Screens: Surrogate Measures of Toxicity
A relatively simple and inexpensive means of beginning a series of nonclini- cal studies is to rely on screening tests. Unlike toxicology testing in whole animals, in vitro screens are individual tests, each with a specific purpose and activity criterion. In addition, in some cases, in vitro screening tests are performed in a series to provide information on a single subject from a vari- ety of tests. Many screens are available for chemical and drug compounds, because of the large number of candidate products tested and a long history of nonclinical development. Some in vitro tests are performed by contract testing laboratories, whereas others are amenable to use in a sponsor’s labo- ratory. Although plentiful and popular for drug and chemical development, these screens may not be useful to measure activity criterion on biophar- maceuticals because test criteria might not match assay requirements and because biological molecules are often incompatible with the test matrix or design. In addition, because of the limited experience, there may be ques- tions regarding relevancy of test results when applied to biotechnology products. Thus, specific concerns can focus on the sensitivity, specificity, accuracy, and reproducibility of in vitro screening tests, when used with a biopharmaceutical.
Nonetheless, when properly applied to a nonclinical safety testing pro- gram, in vitro screening tests can provide valuable information that can be used to design more complex in vivo studies. Examples of in vitro tests are discussed here, and a longer list is provided in Box 8.2. Mutagenicity test- ing, exemplified by the Ames tests, screens compounds for mutagenic poten- tial. The Ames test relies on Salmonella bacteria as a substrate and measures alteration in structure of a gene, after application of a test compound. Other eukaryotic or prokaryotic cells may be used in the same manner, as long as there is a reliable read-out for demonstrating mutagenicity. Carcinogenicity testing takes mutagenicity one step further by asking whether the mutagenic or genotoxic potential of a compound also results in the development of car- cinogenic potential. Since not all mutagens are carcinogens and because not all carcinogens are mutagens, the mutagenicity and carcinogenicity tests can give distinct answers about product. Although screening tests can be helpful to making early decisions, they are not definitive, and carcinogenicity test- ing is considered in animals for compounds that might have carcinogenic potential.
In its simplest format, lethality testing places test material in contact with living, cultured cells and determines whether the material kills the cells. Various cell types can be used in these studies. Variations of the in vitro test measure biochemical or physiological parameters of cell health, which would indicate that a cell might die, or at least not thrive, in the presence of
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BOX 8.2 EXAMPLES OF IN VITRO SAFETY TESTS USED FOR DRUG OR BIOPHARMACEUTICAL SCREENING
In Vitro Safety Test Purpose of Test
Mutagenesis Ames screening test Genetic toxicology for mutagenicity in bacteria Mammalian cell mutation tests Genetic toxicology by mutagenicity of various
mammalian cell types
DNA Damage and Repair Chromosome aberration test Chromosome aberrations and mitotic indices in
cells Cytotoxicity tests Cytotoxic activity by using a variety of cell
types Karyotype analysis Gross chromosomal analysis Micronucleus tests Potential to induce genetic damage, measured
as induced micronuclei in a variety of cell types DNA repair tests Potential to damage DNA Sister chromatid exchange test Genetic damage, as manifested by sister
chromatid exchange in various cell types Cell transformation tests Potential to cause genetic damage, as
manifested by induced morphological cell transformation
Aneuploidy tests Chemically induced aneuploidy in cells
Other Metabolism tests Evaluation of metabolic stability of various cells Cellular anabolism Ability to affect protein anabolism in cells
Cellular respiration Ability to affect cellular respiration, measured as ATP/ADP
In vitro drug metabolism Metabolism of products by cells Human skin permeation test Prediction of dermal and ungula permeation
Mitochondrial toxicity test Damage to mitochondrial function
Hemolysis tests Damage to red blood cells
Drug interaction tests Ability of product to be metabolized in presence of other drugs
Hepatotoxicity tests Measures drug-induced liver injury
Cytokine/chemokine secretion test Potential for triggering release of cytokines or chemokines
Apoptosis assay Induction of cell death
Cellular proliferation Induction of cellular division and proliferation
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the test product. Special types of cells—cardiomyocytes, neural, epidermal, and gastrointestinal epithelial—can be used in such studies, giving rise to test protocols with specific purposes, such as neurotoxicity testing. A num- ber of screening assays, each providing a specific outcome, are currently in development or validation with focused application. However, owing to the unique nature of most biopharmaceuticals, their utility as a toxicology screen is somewhat limited.
Developmental toxicology measures toxicity of a compound as it relates to development of a fetus and, for biopharmaceuticals, is especially important for molecules that might be used in women of childbearing age. In vitro developmental toxicity screens would be considered insufficient by them- selves for risk assessment in this area. However, some in vivo models that use pregnant rodents can also be applied to screen compounds and are more rapid but perhaps less sensitive, as compared with chronic developmental toxicology studies.
In Vivo Safety Testing of Biopharmaceuticals
Although it is possible to perform some safety testing in various nonanimal models, it is also necessary to test most biopharmaceuticals in live animals. Toxicologists classify safety tests in three manners: (1) by the length of time for which an animal is dosed, that is, acute, subchronic, and chronic studies; (2) by an organ system of interest, such as neurotoxicology and immunotoxi- cology; and (3) by a particular outcome, for example, carcinogenicity testing. Since consumers demand thorough safety testing of each biopharmaceuti- cal they use and because there are ethical questions regarding the use of vertebrates in such tests, toxicology testing in animals is a serious scientific endeavor, professionally performed and regulated by government agencies.
Animal Model Development
An animal model is a nonhuman, living vertebrate used in nonclinical research to ensure that a product is reasonably safe and, in some cases, to demonstrate efficacy or benefit before use in man. Consider that every model is imperfect in some way, certainly as compared with the human situation; this is why, it is referred to as a model. Nonetheless, an animal model allows scientists to gain an understanding of a broad range of toxicological pro- cesses and outcomes, and this means that the sponsor can make informed decisions regarding the intended use of a biopharmaceutical in man. Thus, animal study results help the sponsor to avoid the risk of causing harm to humans, while at the same time, allowing the biopharmaceutical to provide intended benefit at a particular dosing regimen. For example, if a biopharma- ceutical dose of 1 mg/kg per week is toxic but a dose of 0.5 mg/kg per week is not toxic in an animal model, then the sponsor can design a clinical trial to test the lower dose and avoid the higher dose. Normally, animal models
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are chosen or developed for a specific study design, and not vice versa. As discussed in more detail later, the sequence of planning events in safety assessment is first a hypothesis or a question and then a well-considered concept study design to drive the selection of the proper model. Selection of the correct animal model or models is challenging, and in the end, compro- mises are made and the best model is identified. Because a toxicology study is designed to answer a specific question or a set of questions regarding the safety of a particular biopharmaceutical, it is often necessary to use a differ- ent animal species to answer each question posed.
For most drugs, safety testing requires the use of two or more animal spe- cies over the development life cycle, whereas for other products, notably many biopharmaceuticals, testing in one animal might meet all regulatory requirements. Normal or healthy animals are usually tested first to dem- onstrate safety. In addition, it may also be necessary to use a second model, an animal with a similar or identical disease, because the disease process may greatly modify the toxicology profile of the product. General guidelines for the selection of animal model species are listed in Box 8.3. Overall, it can be very challenging to identify two excellent animal models for acute and chronic safety testing of a biopharmaceutical. Typically, there are many trade-offs in the selection process and the perfect model may never be found. Identifying an animal model with the disease is especially challenging, because the animal species selected must be appropriate and also the dis- ease must be relevant to the human condition and must have a similar etiol- ogy, pathogenesis, and clinical outcome. For example, to examine the safety of a therapeutic vaccine to treat Alzheimer’s disease, one must identify or develop a model of the disease that arises and progresses in the same man- ner as the human disease and the animal must exhibit the same immune response as would be expected in a human (having this condition) treated with that vaccine. Advances in development, notably the use of transgenic or knock-out animals, offer appropriate models, but these animals can be very expensive. Having chosen an animal model, it is then important to carefully design the nonclinical study to take advantage of any attributes the animal may possess, while at the same time, using the correct number of animals for each question posed.
Animal models should be validated, or at least qualified, for a particu- lar application. Unless an animal model has been widely used for a par- ticular class of biopharmaceuticals or disease, it is advisable to perform pilot studies and determine whether or not the chosen species and strain is, in fact, suitable for the intended purpose. Experience with an animal model will also help to better understand and more accurately plan for animal numbers needed in a pivotal preclinical study, especially if the ani- mal attrition rates associated with the creation of the animal model are unknown. Studies performed in the research laboratory environment help to ensure that when later used in an expensive and lengthy toxicology study, there will be no technical issues and the results will be meaningful.
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Use of animal models proves very useful in identifying any delivery issues or specific challenge with measuring a proposed clinical end point before evaluation in the clinical setting. Indeed, small pilot studies of a compound in a given animal model often lead to improvement in both the application of the model and the ultimate toxicology study design.
Test Product Formulations, Routes of Delivery, and Dosing Designs
Development and selection of animal models are not the only things that need to be considered before performing a nonclinical toxicology study. There is also the need to have a final product (Chapters 5 and 6) that matches the formulation and quality of the biopharmaceutical intended for use in humans. Far too often, nonclinical studies are considered invalid because they apply to a product that differs in strength or quality from the intended
BOX 8.3 CONSIDERATIONS FOR SELECTION OF ANIMAL MODELS FOR TOXICOLOGY STUDIES
• Observe taxonomic, anatomical, and physiological similarities to humans • Consider overall anatomy and physiology of animal • Anatomy and physiology of target organ, tissue, and cells
• Demonstrates pharmacology, pharmacokinetics, and pharma- codynamics, similar to the intended human population • Metabolizes drug in similar manner • Has same receptors or mechanism of action • Expresses same target organ, tissue, or cell responses • When possible, provides a model of the human disease
• Dosing parameters match human condition • Ability to give full human dose and dose regimen to animal • Consider route of delivery
• Economics • Use enough animals of this species to fully answer the
question • Consider cost of maintaining the animals over the period
of study design • Ethics
• Is it necessary to use an animal to answer the question(s) or an in vitro system would suffice?
• Is it possible to use a species lower on the taxonomic chart?
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clinical material. Formulation, strength, and quality can greatly impact the biological activity, including parameters related to safety, of a molecule or cell. Hence, it makes sense to use the same or a comparable formulation in nonclinical studies as will be used in clinical studies. This can have a signifi- cant impact on scheduling a nonclinical study, since formulation, manufac- turing, and quality control timelines impact the nonclinical plan. In addition, there may be trade-offs. For example, if it is absolutely necessary to use a mouse to test a human dose of a therapeutic protein, it may not be possible to give a full human dose because of volume constraints in the animal. The result could be to split doses or to concentrate the formulation. These issues arise constantly in design of safety studies and must be addressed scientifi- cally in consultation with other development scientists.
Route of delivery presents another hurdle in design of nonclinical studies. For the example given above, it might be necessary to give the product to a mouse by the intraperitonal route, because it is impossible to give a full dose to this animal by the intended human route, that is. intravenously. Indeed, sometimes an animal species is chosen based on matching the intended route of delivery for man. Because pig skin is very similar in microanatomy and function to human skin, the pig is used in many nonclinical studies of biopharmaceuticals that are to be delivered to human epidermis.
Nonclinical animal studies reflect a vocabulary that is unique to this endeavor. These terms are explained here because they are commonly used in study protocols. First are the trade terms. Test article is the product or final product, as described in Chapters 6 and 7. It is the final formulation of the material that is being tested. Neat means an article is used in full strength or undiluted. Placebo represents inactive ingredient and might be referred to as control or control article. A diluent is a defined solution, buffer, or formu- lation, such as physiological saline, used to titrate the test article or control. An excipient is a nonactive ingredient included in a formulation. Common excipients for biopharmaceuticals include detergents that prevent protein precipitation and sugars that preserve the integrity of cells in a solution. A particular type of excipient, the vehicle, is a chemical that serves to enhance transfer, absorption, or distribution of the biopharmaceutical. Tween 80, a detergent-like molecule is used to prevent aggregation and as such consid- ered a vehicle in formulations of certain protein biopharmaceuticals.
A second consideration is the route by which a biopharmaceutical is given to an animal or human; the most commonly used routes are listed in Box 8.1. Attention is also given to physiological variables that can have an impact on dosing of a product to an animal or to man. Local effects are the physiologi- cal responses of the recipient to the test article when it first reaches the recipi- ent tissue or organ. Absorption and distribution are the processes by which the test material moves away from the site of delivery and establishes itself in various tissues and organs. Formulation can have a major impact on how efficiently absorption and distribution occur and where a biopharmaceuti- cal goes in the body. Next, there is the issue of metabolism, the process by
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which the biopharmaceutical is chemically changed, broken down, or other- wise used by the body. Absorption and distribution, and hence also the local effects, influence metabolism of many products and were discussed earlier.
Calculating dose is an underappreciated skill but can have a major impact on the outcome of a nonclinical study. At the outset, the design of a nonclinical study has, from the TPP, a target dose or a range of doses that clinical experts suggest might be used in human volunteers. For proposed nonclinical studies, the study design brackets the target human dose, based on an understanding of the product and the chosen animal model, as well as the dose equivalents with respect to understanding the relative body weight or surface area of an animal versus a human. If body weight is chosen as a comparator, then dosing calculations are made as milligram of biopharmaceutical per kilogram of body mass. Alternatively, dose estimates based on body surface area are becoming more common, perhaps because of obesity in society, disease being treated, or factors related to pharmacokinetics or pharmacodynamics of biopharmaceuti- cal products. The chosen dose can impact the calculated relative dose for some biopharmaceuticals and where small animals, for example, mice, are employed. Selecting a close approximation of the intended dose, dosing schedule, route of administration, and rate of administration to the future human clinical study design is important to consider in the design of animal studies. Calculation and rationale of dose and approximation of human dose equivalents also need to be carefully planned. Finally, it is important to calculate from a study design the total test article, control article, and vehicle requirements, as well as speci- fications and tests for quality, After this, the outcomes and decisions on study design should be discussed with colleagues responsible for biomanufacture and quality control, focusing on the total quantity and number of individual lots of product, diluents, and placebo.
Protocols and Performance of Biopharmaceutical Safety Studies in Animals
A concept nonclinical study design, once found acceptable to a product development team, is then written further in a nonclinical study design doc- ument, that is, the protocol. The purpose of the nonclinical study protocol is to guide the investigative team in performance of a study. Elements of a non- clinical study protocol are given in Box 8.4. Responsibility for preparing the protocol is given to an individual, the study director, who is responsible for ensuring the performance of the study according to the study protocol. This individual is also responsible for completing a study report at the end of the investigation. Nonclinical protocols are also reviewed by other scientists who serve on the nonclinical study team and are approved by the quality assurance unit. Further, any use of animals in research requires review by an Institutional Animal Care and Use Committee (Chapter 4), which is respon- sible for the ethical and proper use of laboratory animals. A nonclinical study always results in a report, a prospectively written scientific document
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that identifies the design and justification of the study, as mandated under the protocol. The study report contains the experimental results, statisti- cal analysis, list of deviations, and conclusions made by the study director and provides, in appendices, all tabulated raw data derived from the study. A supplemental report may be incorporated in the final study report; the supplemental report includes expert opinions about the clinical relevance of results by a physician or clinical pathologist. Nonclinical study reports are typically large documents, hundreds of pages, even for a small and simple studies it is common for these reports to be submitted to the FDA as part of an IND application. Finally, it is worth noting that a nonclinical study in animals costs a significant amount of money and takes considerable time (6–12 months for acute and subacute studies and much longer for chronic studies) from concept protocol design to the final study report; this is the reason that much care is taken for study design and execution.
Elements of a Nonclinical Study Design
Adequate and well-controlled nonclinical study designs have common ele- ments, no matter how the study is classified (e.g., acute, subchronic, and chronic), and each element is considered in the study protocol. Content of the protocol is, to some extent, mandated by cGLP regulations, and this alone harmonizes the format (Box 8.4). To begin, there is a facility and it is staffed with qualified individuals capable of performing functions called
BOX 8.4 ELEMENTS OF A NONCLINICAL STUDY PROTOCOL FOR SAFETY TESTING OF BIOPHARMACEUTICALS
• Title, purpose, sponsor, and testing facility • Detailed identification of test and control articles and animals • Methods of identification • Description of materials used in the study, including the
animal diet • Dosing levels of test material • Experimental design and methods used to control bias • Type and frequency of tests and measurements • Records to be maintained • Statistical methods • Dated approval of protocol by sponsor, study director, and
quality assurance • Any changes made • Approvals
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for under the protocol. The facility is properly designed and equipped for the types of studies performed. As noted above, each study is defined under a protocol and has a responsible study director. A quality assurance unit is also an integral component of nonclinical programs by conducting audits, reviews, and approvals among other functions (Chapter 5). If the study calls for laboratory analysis, which most studies do, then there are adequate laboratory facilities identified to perform the work either at the study site or, for specialized tasks, at qualified contractor sites. Animals are involved in most studies and all their support is met; this involves hous- ing, feeding, treating, and inspecting or analyzing each animal, as well as ensuring quality care and well-being. In addition to a protocol, routine procedures—everything from handling animals to archiving records—are fully described in instructional documents such as standard operating procedures (SOPs). In addition, every bit of information and all data are captured on source documents and may be transferred to study data cap- ture forms, most designed uniquely for that study. All of this information is reviewed by qualified professionals, condensed into summaries, written into a final study report, and properly reviewed and approved. Quality assurance involvement is essential to this process.
However, each protocol is scientifically unique in purpose, scope, and design and is written by the study director, with much scientific and tech- nical input and based on a hypothesis. One reason is that each nonclini- cal study is related to a unique biopharmaceutical. To test the hypothesis and answer every question posed in the study objectives, a study design includes a unique mixture of animal and laboratory treatments, tests, and procedures. The design is given at three or more levels of variables. One level defines the dose and respective controls, the dosing scheme and sched- ule, and the length of time for which animals are on study. This leads to the designation of animal groups with each group receiving a different treatment.
The second level of design provides further instruction for carrying out the essential elements of the study. An example is animal care and treat- ment, baseline pathogen screening, observations, shaving or clipping, anesthesia, food, weight, and physical environment are written into the protocol in an effort to support the scientific design. Another example is laboratory testing. Clinical laboratory testing—hematology, clinical chem- istry, urinalysis, and immunology—measures the health of the animals. Other laboratory testing may measure the level of product or metabolites from that product.
The third level of design enters into the example of laboratory testing, because the analytical laboratory must develop or have on hand an assay or set of assays that are robust and both selective and sensitive for the compound being studied, when it exists in animal blood or tissues. Assay qualification (Chapter 7) is a common practice to support nonclinical stud- ies. Thus, laboratories must be capable of performing a variety of analytical
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methods. Very special laboratory studies may be performed at another site, at either a contract laboratory or a sponsor’s laboratory. For example, it might be necessary to send animal blood or tissue to a contract laboratory to measure levels of the excipient Tween 80 if there is a concern that the excipient might be concentrated in the body of an animal. A special assay to measure the effect of a product on animals might be performed by the sponsor of a study if this test requires special expertise or laboratory equip- ment. For example, a cellular assay to measure proliferation of lymphocytes in response to a recombinant protein might have to be performed by the sponsor’s laboratory on specimens taken from animals immunized with a new vaccine.
Returning to the second level of design, the planner must consider other procedures in the in-life phase of the study. The in-life phase of an ani- mal study is defined as the time beginning with the initial assignment of animals to the study, including dosing of the animal, until the time when it is euthanized or released from the protocol. Most toxicology protocols demand frequent examination of animals during the in-life phase of the study, and this involves close observation and measurements of general animal health, such as weight and amount of food consumed per day. It may be helpful to use or develop qualitative assessment scales to better define and capture subjective data. For example, animal appearance is a very good indicator of overall animal health, but it is hard to quantify and usually not standardized; therefore, provision of a general appearance scale or of predetermined defined criteria are meaningful and provide a valid way to capture this assessment. Some commonly used measures of animal health are given, as in-life and postlife measures, in Box 8.5. After the in-life phase, defined by the protocol, most animals, perhaps with the exception of some large species and nonhuman primates, are euthanized. This begins the postlife portion of the study. For most safety studies, a formal necropsy is performed, and rigorous gross inspection is followed, after tissue preparation, by histopathological examination of every organ. Parameters to measure animal health or toxicity are listed in a protocol, and any abnormality in each of these parameters is recorded and the results are analyzed by a certified veterinary pathologist. Plans are also included for data handling, analysis by a statistician, and report- ing. Records are exact and complete, and all procedures fall under cGLP guidelines. Data and reports follow stringent criteria for analysis and review by the laboratory and sponsor and for approval and reporting to regulatory authorities.
Hence, the design and procedures provided in nonclinical and clinical studies are carefully considered, are rigorous, and are provided in detail. Nonclinical designs and protocols are instructive and directive, because both types of protocols are important to human subjects and users of the biophar- maceutical. These studies are time consuming and very costly, so there is little room and no justification for error.
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BOX 8.5 COMMON MEASURES OF IN-LIFE ANIMAL HEALTH AND POSTMORTEM CHANGES IN NONCLINICAL
TOXICITY TESTING
• In-life measures of animal health and well-being • Clinical signs
– Weight loss and weight gain – Appearance or behavior (ruffled fur and hunching) – Agonal events – Neuromuscular, ophthalmic, and other special tests
• Clinical laboratory pathology – Hematological abnormalities – Clinical chemistry abnormalities – Abnormalities in urine, secretions, or other samples
• Postmortem measures of animal health and well-being • Death (mortality)
– LD50 – Time to death – Cause of death
• Gross observations at necropsy – Edema and fluid in tissues and body cavities – Color, size, and appearance of organs – Whole body weight – Whole organ weights – Obvious signs of systemic disease – Tumors – Developmental abnormalities
• Histopathological examination of tissues – Tissue or cellular changes – Signs of infection or inflammation – Malignancies or nonmalignant tumors – Developmental abnormalities – Signs of systemic disease or stress to a system or the
body as a whole (Continued)
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Nonclinical Safety Testing
The extent and level of nonclinical safety testing are largely based on the type of product being developed, in addition to what is known about the predicted safety profile of that product. There are a number of traditional nonclinical safety tests that have been utilized for decades that the USFDA embraces as being validated and dependable and that yield predictive results that are likely to correlate with human clinical safety. Examples of these common nonclinical tests are provided in Box 8.6. Although this list of common tests is provided, they are not all required for every biotechnol- ogy product, but rather, a subset of the most relevant tests is selected to be performed. The selection of testing and extent of testing are based on prod- uct profile and other characteristic attributes of the product and include the product’s biological function. Brief descriptions and examples of the most common nonclinical safety studies are given in the following sections.
Acute Toxicity Testing
Acute toxicity testing has been defined as the short-term evaluation of toxic- ity in animals after a single dose of a biopharmaceutical. Today, the defini- tion would, by most accounts, include study designs with multiple doses of the product but over a brief period and again with short-term evaluation. If the devil is in the study details, then the devils here are the definitions of the terms brief period and short-term evaluation. For a therapeutic protein, an acute study might involve three doses over three days, with completion of in-life studies on the sixth day. In contrast, an acute study of a vaccine might be three doses over ten days, with completion of in-life studies on the eighteenth day. Hence, the definition may be adjusted, depending on the type of compound, expected dose and dosing schedule, possible toxic effects, indication, and animal model. However, there are established com- mon elements and guidelines for acute toxicity tests. These guidelines are as follows: (1) Findings are suggestive and never definitive as to the overall toxicity of the biopharmaceutical; (2) It is a screening toxicity and may be
BOX 8.5 (Continued) COMMON MEASURES OF IN-LIFE ANIMAL HEALTH AND POSTMORTEM CHANGES IN NONCLINICAL TOXICITY TESTING
• In-life or postlife observations and tests • Infection demonstrated by microbiological examination or
culture • Immune response to various stimuli • Pharmacokinetic or pharmacodynamic measurements
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BOX 8.6 COMMON NONCLINICAL SAFETY TESTS USED FOR DRUG OR BIOPHARMACEUTICAL EVALUATION
Safety Test Purpose of Test
General toxicology Maximum tolerated dose (MTD)
Dose-limiting toxicity (DLT)
No observed adverse effect level (NOAEL)
No observable effect level (NOEL)
• Establish administration schedule and approximate clinical exposure.
• Identify reversible or irreversible side effects. • Identify extent and severity of pathologic lesions. • Evaluate regenerative capacity of organs and organ
systems. • Typically one rodent and one nonrodent species. • Minimum of three animals/sex/group. • Early stage of clinical development.
Acute toxicity testing • Establish short-term exposure toxicity, based on intended human clinical exposure.
• Confirm administration and delivery schedule. • Rodent and nonrodent species. • Single-dose toxicology study or repeat-dose-range
finding study (MTD). • Early stages of clinical development.
Subchronic toxicity testing
• Establish dose and dose regimen. • Increased exposure relative to acute toxicity. • Repeat dose with study duration of 1–3 months. • Definitive toxicity profile to be obtained. • Early stage of clinical development.
Chronic toxicity testing • Establish long-term exposure toxicity and correlate with intended human exposure.
• Use larger number of animals to allow statistical evaluation and accommodate for animal attrition.
• Early to mid stage of clinical development.
Reproductive toxicology testing
• Evaluate male spermatogenesis, female follicular development, fertilization, implantation, and early fetal development.
• One pharmacologically relevant species is usually sufficient.
• Biopharmaceutical product intended for adults of childbearing potential or children with developing reproductive systems.
• Animal evaluation to include both sexes. • Focus on reproductive system and/or organs. • Fertility of both sexes, early embryonic development to
implantation. • Early to mid-stage of clinical development.
(Continued)
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BOX 8.6 (Continued) COMMON NONCLINICAL SAFETY TESTS USED FOR DRUG OR BIOPHARMACEUTICAL EVALUATION
Safety Test Purpose of Test
Developmental toxicology testing
• Establish mortality, structural abnormalities, functional impairment, and growth alterations in fetus or neonate.
• Typically use rats or rabbits. • Identify effect on embryo-fetal development. • Pre- and postnatal developmental effects. • Examine female parturition and lactation. • Used for selected biopharmaceutical products. • Mid- to late stage of clinical development.
Carcinogenicity testing • Establish carcinogenic potential. • Prolonged exposure or extraordinarily high
physiological doses. • Determine if the biopharmaceutical product resembles
similar chemical class or structure of known carcinogen.
• Use two different rodent species. • Route, dose, and dose regimen used. • Biopharmaceutical product specific. • Later stages of clinical development.
Immunotoxicity testing • Establish potential to cause immunopathology. • For immunomodulatory pharmaceuticals, consider
immunophenotyping via flow cytometry. • Used for selected biopharmaceutical products. • Evaluate anaphylactic response, delayed
hypersensitivity, acute immune response to product, potential to elicit autoimmunity, and potential of immune suppression.
• Choice of animal species and immune status is critical for validity of these studies.
• Early stages of clinical development.
Genetic toxicity testing • Evaluate potential to negatively affect genetic material. • Examine for specific mutations or more general
chromosomal breaks or rearrangements. • Utilize both in vitro and in vivo assay formats. • Early stages of clinical development.
Tissue binding or local tissue tolerance
• Evaluate local reactions to multiple doses. • Long- or short-term and based on clinical intent. • Used for selected biopharmaceutical products. • May be combined with acute, subchronic, or chronic
toxicity study. • Surrogate in vitro studies may be useful. • Animal studies are likely most appropriate. • Early stages of clinical development.
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useful to rank toxicity, as variables are adjusted (e.g., dose, number of doses, timing of each dose, and route of delivery); and (3) It represents only an assessment of potential toxicity. To further confuse the definition, acute tox- icity is defined in one manner for drugs, in another manner for compounds intended for environmental release, and in yet another manner for certain types of biopharmaceuticals.
An acute toxicity study design does, however, contain elements that deserve mention. First, some are range-finding studies, meaning they are designed to define a dose level that is suitable for the proposed clinical dose and on which to base more definitive range-finding studies. Second, end points and measurements are as important in acute toxicity testing as they are in any other study. One end point may be finding the dose level that results in significant or measurable harm or disease (requires careful testing and definition) or the dose level that might be lethal (easy to measure and define). Here, the term LD50, the dose of a product that causes the death of 50% of animals receiving the product, may be used. Although this end point is seldom measured for biopharmaceuticals, a similar concept, here referred to as XD50 can be applied, where X is a particular measurement of declining health, such as 50 g of weight loss in a rat. Toxicologist have identified and used numerous acute toxicology range-finding study designs, end points, and measurements, and now, some of these are applied to nonclinical stud- ies for biopharmaceuticals, notably to therapeutic products. An example of a typical acute toxicity study design for a biopharmaceutical product is pro- vided in Box 8.7.
How are the results of acute toxicity testing of a biopharmaceutical consid- ered and acted upon with regard to further product development of the bio- pharmaceutical? Some outcomes are easily interpreted, and at other times, they confound, more than clarifying, the interpretation of results. A lethal dose (e.g., 100 mg/kg) of any biopharmaceutical in a relevant animal species would not be considered for a Phase 1 clinical study. In addition, if that lethal dose were near or in the range that had been chosen for clinical therapy, then this product would probably not be progressed to clinical trials unless it were first reformulated or otherwise changed to significantly reduce the tox- icity, while retaining the therapeutic effect. Yet, there are caveats even with this example. Should the product be indicated for a life-threatening disease for which there is no other possible therapeutic intervention, then it might be progressed to more definitive toxicology studies. Every result must be considered in context.
Although not definitive, the results of acute toxicity testing certainly aid the selection of dosages and perhaps dose regimens, or, at least, they should, if the study was designed properly, the animal model was care- fully chosen, and the dosages bracketed the proposed human dose. Acute studies that give multiple doses are also instructive on additive effects of the biopharmaceutical in an animal, an information that can be applied to later studies. Acute studies may also provide information on proper
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timing of doses and, if properly designed, may yield meaningful phar- macokinetic and pharmacodynamic data. Again, these objectives must be considered in the acute study design and completed in the in-life and laboratory phases of the study. It is worth noting that acute toxicity stud- ies of biopharmaceuticals are occasionally performed in research labora- tories and without regard for GLP. They are considered pilot studies on which to base dose selection for an adequate and well-controlled (cGLP) subchronic and definitive toxicology study, and most often, they are not be acceptable to regulatory authorities as definitive acute toxicity studies. In such cases, results that demonstrate lack of toxicity are not compelling, but when toxicity is noted, it must be reported to regulatory authorities and considered in the overall picture of product safety. Significant benefit is derived in savings of time and money, but there is some regulatory and scientific risk involved in performing such pilot studies. Often times, non-GLP studies that demonstrate good documentation practices through the generation of a prospectively defined study protocol, quality control, accurate data capture, and generation of a final study report associated with these pilot studies may go a long way in gaining the acceptance of these study results by FDA.
BOX 8.7 ACUTE TOXICITY STUDY DESIGN FOR A BIOPHARMACEUTICAL PRODUCT—EXAMPLE
Acute Toxicity
Rationale • Determine maximum tolerated dose or no observable effect level
• Identify potential organ toxicity, determine reversible or irreversible damage, and determine clinical safety end points
• Determine dose level (basis for repeat dose toxicity study) In-life duration • 1–2 days up to 15 days Test system • One rodent animal species
• One nonrodent animal species Administration • Closely mimic that intended for human clinical
administration End points • Weight change
• Physical appearance • Gross necropsy • Clinical laboratories • Clinical pathology • Other indicators of toxicity • Mortality
Cost estimate Low to medium*
*Depending on study elements such as animal species, in-life duration, controls, dose range, and end point analysis.
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Subchronic and Chronic Toxicity Testing
There are several different definitions for chronic, subchronic, and sub- acute toxicity studies that cover any number of nonclinical safety testing when applied to biopharmaceuticals. The terms subacute and subchronic are subjective, but progression from subacute through chronic studies reflects an ever-increasing exposure of animals to the biopharmaceutical, beyond those applied in acute studies. The subacute study, a term used more often with drugs than with biopharmaceuticals, refers to the studies that are done as repeat dose and at dose levels between those of acute and subchronic studies, with durations of 1–3 months. Subchronic studies are, perhaps, 3–6 months in duration and typically involve multiple doses, if indicated. They look for cumulative biological or health changes in ani- mals. They can, however, be broad explorations, examining animals for a wide range of symptoms or diseases. They should, in the end, define toxic- ity as well as add to the pharmacologic body of information. Subchronic study results are both qualitative and quantitative and the studies attempt to, and should in fact, meet statistical end points to clearly demonstrate toxicity if it exists. The anticipated result of a subchronic study, the one supported by unequivocal data, is safety of the product at a dose and dos- age regimen that are desirable and feasible in humans. This is referred to as a clean dose level of the biopharmaceutical. Finally, a well-designed sub- chronic study forms the foundation for designing required for follow-on chronic and specialty toxicity studies.
The study design of subacute and subchronic studies are driven by many variables, not to mention the indication and nature of the product, and no single design should be considered authoritative. The selection of an ani- mal model is very important to the success of a subchronic study, and the data from acute studies and the research laboratory are most supportive. Regulatory guidelines for drugs and often pharmaceuticals specify test- ing in two animal species, including a rodent (rat) and a larger nonrodent animal (dog or nonhuman primate). Today, therapeutic biopharmaceuticals reviewed by the Center for Drug Evaluation and Research (Chapter 3) often follow this guideline. However, other biopharmaceuticals, such as vaccines and genetic therapies, have followed precedent established at the Center for Biologics Evaluation and Research and used a single species, shown to be suitable for the product, notably for the intended indication, dose, and dosing regimen. For subchronic studies with these biopharmaceuticals, it is important to perform pilot studies to test variables and the animal model before the definitive subchronic study is performed.
Design may be single dose or, more commonly, repeat dose. It is important to ensure that the chosen design is statistically valid, so that enough animals of each sex are assigned to each dosing group. Single-sex studies are acceptable but only if a good rationale and justification for why sex-specific differences are not anticipated. Different doses are tested for most biopharmaceuticals to
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ensure that the dose and dose regimen taken to human studies are safe and well tolerated and that the next higher dose is not unsafe. An example of a typical subchronic and chronic toxicity study design for a biopharmaceutical product is provided in Box 8.8.
Statistical planning is an important consideration in determining adequate animal numbers and dose escalation in prospective study design; this is especially important to be able to rely on statistical power in the design and data analysis for any unexpected results. In other words, upfront involve- ment of a biostatistician in the design of a lengthy, expensive, and labor- intensive study may allow valid statistical interpretation of unclear results via statistical analysis. Likewise, building a margin of safety into a toxicol- ogy study not only represents good science but also provides the opportu- nity to demonstrate failure (e.g., toxicological findings at high dose levels). A wide safety margin also serves as a control that demonstrates that the test system is, in fact, capable of identifying toxicities at dose levels higher than those anticipated to be used in human clinical studies. Demonstration of a broad safety range or plateau provides an added level of confidence in safety, which supports the scientific integrity of the study, rather than potentially unknowingly approaching a sharp cutoff, which represents a narrow win- dow of safety. A graphical representation of this is presented in Figure 8.6, where toxicities or incidence of safety are plotted relative to dose escalation. Additional plateaus may be identified, especially if small dose increments
BOX 8.8 SUBCHRONIC AND CHRONIC TOXICITY STUDY DESIGN FOR A BIOPHARMACEUTICAL PRODUCT—EXAMPLE
Rationale • To determine no observable effect level (NOEL) • To evaluate dose response of multiple subsequent dosing
regimens • To identify and characterize specific organ toxicities • To predict optimal human clinical dose
In-life duration 2 weeks (subchronic), up to 6 months (chronic)
Test system • One rodent animal species, the rat • One nonrodent species, the dog
Administration Three dose levels, to include levels likely to produce no toxicity and high toxicity (but <10% mortality)
End points and measurements
• Weight change • Physical appearance • Gross necropsy • Clinical laboratories • Clinical pathology • Other indicators of toxicity such as appetence or lethargy • Mortality
Cost estimate Medium to high
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are used. Note that other areas along the dose escalation scheme, for exam- ple, between points 1 and 2 or points 3 and 4 of the x-axis carry the risk of more variable incidence of adverse effects and do not represent good areas to select a starting dose.
Controls are always included in subchronic studies, and for many bio- pharmaceuticals, this means at least two additional groups of animals, one dosed with the formulation minus the active ingredient and another, the null control, dosed with normal saline or nothing at all. Details regarding per- formance of a subchronic study, such as dosing animals in all groups at the same time, can be important. Defining study termination and animal eutha- nasia is also critical to study design. It is important to leave enough time after dosing to allow toxicity to occur, but then again, this is not a chronic toxic- ity design. For some biopharmaceuticals, guidelines recommend sacrificing one group of animals after the last dose is given and euthanizing another group, treated and controlled, in a similar manner, weeks after the last dose is given. These nuances in study design demand that the sponsor is very familiar with precedent within a class of biopharmaceuticals, has read all the regulatory guidelines, and intends to meet with FDA (Chapter 3) soon after the concept design is drafted and well before the subchronic study begins.
One definition of a chronic study is long study, which takes months to even years and examines animals repeatedly and closely for death, changes in health, or signs of chronic disease. Collectively, long-term or prolonged peri- ods are defined in guidance documents as continuous dosing for 6 months, with intermittent dosing. For many biopharmaceuticals, chronic studies are not worthwhile because the product is not given over a long period of
1 0
200
400
800
1000
600
2 3 4
T ox
ic iti
es (s
af et
y)
Dose (efficacy)
Dose selection
Plateau
A
C
B
FIGURE 8.6 Using safety and efficacy in selecting a dose. Determining the most appropriate starting dose that is likely to demonstrate an acceptable range of safety and efficacy can be achieved by identifying a plateau. In an animal toxicity study, as the dose increases, so do the associated toxicities (A); in many cases, the incidence of adverse effects may level off between dose levels, which is referred to as a plateau (B). After reaching a plateau, subsequently higher doses are usually associated with increased toxicities (C).
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time to patients. A recombinant therapeutic protein, intended to be dosed just three times, on Days 0, 30, and 60, is a fine example of a product that might not require a long, chronic study. In contrast, a therapeutic monoclo- nal antibody intended to treat patients with chronic inflammation by dos- ing biweekly and for many years deserves chronic toxicity testing, as does a genetic therapy that is designed to incorporate foreign DNA into the human genome and produce a lasting effect, even though it is given one time as a single dose. Another important criterion to consider while determining the requirements of performing a chronic toxicity study may be whether long- term tissue retention of the biopharmaceutical product is anticipated when administered in humans. To know when, in the product life cycle, the chronic studies are designed and performed is not as challenging as it is to know how they are designed and performed.
In chronic toxicity testing, the biopharmaceutical is administered over much of the animal’s lifetime. Animals are kept on protocol, and are housed, fed, and observed daily, for a substantial portion of the animals expected life. It is no surprise then that small animals, especially mice, which has a lifespan of less than 2 years, are selected for chronic studies. Chronic toxicol- ogy studies are often confounded by findings that are a normal part of aging. For example, spontaneous events such as sudden and unexplained deaths of individual animals are a reality in all species and cancers—common find- ings of aging—inbred animals. Well-controlled studies are the key to dis- tinguishing product-related adverse events and disease from those simply associated with aging or a common environment. Therefore, statistically valid designs require large numbers of animals in each group to distinguish an incidence of disease or events that occur by chance from the incidents related to product toxicity. This concern drives the design of large stud- ies with many animals to account for attrition and significant numbers of samples and tests, applicable to both in-life and postlife evaluations. Hence, chronic studies tend to be large and expensive, because of which it becomes crucial to ensure proper design, to focus on the toxicity that really matters, and to ensure study protocol reviews by regulatory agencies.
Reproductive, Developmental, and Teratogenicity Toxicity Testing
Biopharmaceuticals intended for use in individuals of childbearing age or in children with developing reproductive systems are further tested to ensure that the product will have no undesirable effect on reproductive tissues or on a developing fetus. Consumers are extremely sensitive about develop- mental and reproductive toxicology for good reason. Guidelines (e.g., ICH and FDA) suggest that a sponsor consider at least three types of studies if the product can reach the gonads or fetus. The first type of study, Segment I, is toxicity to male and female fertility and of early embryonic develop- ment to implantation. End points measured in such animal studies are mat- uration of sperm or eggs, gonadal integrity, and, in females, normalcy of
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gestation until the time of implantation. This calls for using an animal model of both sexes and recently mated female animals of an appropriate model species. Measurements focus on the reproductive cells and tissue. The sec- ond type of toxicology test, Segment II, is for embryo-fetal development. In these studies and after treatment with the biopharmaceutical, organogenesis is studied in pregnant animals from the time of implantation till the second gestational period. Here, the study is designed to measure abnormalities that might develop in the fetus and the associated organs, such as placenta. The third type of study, Segment III, focuses on pre- and postnatal development. Here, dosing of animals begins in the earliest phase of gestation and contin- ues through birth of the animal. Examinations are performed at various times during the development of the fetus and include examination of neonates. Other study designs may be used, especially if the class of biopharmaceutical is suspected to cause reproductive or development abnormalities. The need for product to actually reach the reproductive system or the developing fetus is a consideration when selecting an animal model.
Since there is no typical biopharmaceutical, each product must be consid- ered on a case-by-case basis. It is worthwhile to examine precedence and regulatory guidelines before designing a study and to consider a study laboratory and animal facility with experience in reproductive and devel- opmental toxicology. Two examples on how and when to perform a study are instructive. For the first example, the biopharmaceutical is a therapeu- tic protein, intended for long-term, monthly, intravenous dosing at 100 mg/ dose. Since the molecule binds receptors of white blood cells, is able to cross the placenta, and is indicated for use by women during childbearing years, reproductive and developmental toxicology testing is considered advisable, and probably even necessary, before Phase 1 clinical studies and certainly before Phase 2. The second example is a recombinant protein of a virus, a vaccine intended for the general population, including women of childbear- ing age and children older than 2 years. It is given intramuscularly at 5 µg per dose and in three total doses. The sponsor intends to add a label warning stating that the vaccine should not be taken if a woman might conceive in the near future or is already pregnant. This vaccine may be tested for devel- opmental toxicology in young animals before clinical studies in children; however, for the adult population, and given the warning, it may never need reproductive or developmental toxicology. For either product, ADME studies would be helpful in making a decision, because they would demonstrate the distribution of the product after injection. These examples point out the need to consider all aspects of a product, notably pharmacology and the intended treatment population, before designing a toxicology protocol.
Carcinogenicity Testing
Biopharmaceuticals are not commonly thought of as carcinogens; however, in theory and sometimes in practice, they might be found to be associated with
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cancer. Carcinogenicity testing is a long and expensive process, much like chronic toxicity testing. Guidelines concerning when and how to perform carcinogenicity testing on a particular biopharmaceutical product are avail- able, and there is precedence for most classes of product. Mice and rats are used almost exclusively, and strains of each species must be selected on the basis of many factors: longevity, spontaneous tumors, capacity to develop tumors in response to known carcinogens, and tolerability of the biopharmaceutical to be tested. Design issues such as route of administra- tion, doses, dosing regimens, and termination are complex, and the sponsor considering carcinogenicity testing is well advised to seek expert guidance and regulatory guidance before embarking on study design. The field is rife with pitfalls, complications, uncertainties, controversies, and changes in recommended practices. Interpretation of results presents another oppor- tunity to seek expert opinion, especially if controls were limited in scope or number and if criteria for carcinogenicity were not well considered in the design and protocol. Although complex and difficult, carcinogenicity studies are simply necessary for some products, sometimes before mid- or late-phase clinical studies.
Immunotoxicology
This relatively new field evolved out of observations and studies on the toxic effects of chemicals on the immune system. With a large number of biopharmaceuticals targeted, directly or indirectly, to the immune system and with other biologicals likely to interface with immune cells and tis- sues at some point during their distribution throughout the body, immu- notoxicological studies deserve consideration for many classes of product. Adding to the situation is the complexity of the immune system, notably the fact that scientists do not yet understanding the intricacies and control mechanisms of this system. Major immunotoxicological concerns are as fol- lows: (1) Adverse allergic responses to the biopharmaceutical because the product or an excipient is perceived as foreign: This is manifested as imme- diate or delayed hypersensitivity reactions, some of which can be immedi- ately life threatening. (2) Immune responses to the biopharmaceutical that neutralize the molecule and make it ineffective: This is not commonly seen with products that are taken over a long period of time. Further, when bio- pharmaceuticals in solution change format, such as going from soluble to microparticulate while in storage, the propensity to elicit both allergic and neutralizing immune responses may increase significantly. (3) Upregulation of the immune response or an inappropriate immune response resulting in immunity to self and thus leading to autoimmune disease: Immediate upregulation caused by biopharmaceuticals that act to release, immedi- ately and in large amounts, cytokines or other mediators of inflammation is of special concern. Use of recombinant cytokines is especially suspected in this regard. (4) Downregulation or suppression of the recipient’s immune
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response: This is common in patients with pre-existing conditions such as cancer or immunodeficiency.
Immunotoxicology testing is highly recommended for certain biophar- maceuticals or for any product derived from and possibly containing mol- ecules from certain sources. A few examples are cytokines or cytokine-like molecules, vaccines and vaccine adjuvants, monoclonal antibodies or immu- noglobulin-like molecules, allergens, products mimicking or derived from microbes that themselves stimulate untoward responses, products derived from certain plants or those mimicking plant allergens (latex, peanut, etc.), and molecules that bind to cells or receptors on cells comprising the reticu- loendothelial system. Given the complete list, it becomes clear that immuno- toxicology is increasingly considered in design of a safety testing program. In addition, there are no simple templates for routine testing of a molecule as there are for acute toxicology studies of certain products. An example of a typical immune toxicity study design for a biopharmaceutical product is provided in Box 8.9. However, note that studies are best designed based on an understanding of the immunological properties, or potential, of the molecule. From this knowledge, it is possible to consider how relevant tests may be performed, in vitro or in vivo. An efficient approach is to piggy-back immunotoxicology studies with acute and subchronic toxicology studies, whenever possible, by adding immunological measurements to the protocol.
BOX 8.9 IMMUNOLOGICAL TOXICITY STUDY DESIGN FOR A BIOPHARMACEUTICAL PRODUCT—EXAMPLE
Immune Toxicity
Rationale To demonstrate that product is neither toxic to the immune response nor elicits an untoward immune response or reaction
In-life duration 2 weeks (acute/innate response) or 3 months (subchronic/ adaptive response)
Test system One rodent species, mouse or rat
Administration Repeat dose, as determined in acute or subchronic toxicity study
End points • Clinical laboratories (e.g., hematology) • Measure immune cells, numbers, and functional
parameters • Cytokine profiles • Mixed lymphocyte reaction • Macrophage functional assays • Histology of lymphoid organs • Immunohistological examination of cell types and
molecules, in situ
Cost estimate High
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However, these measurements always require the application of immuno- logical or cytochemical assays, some of which are expensive, time consum- ing, and technically challenging. In addition, given the species specificity of cells and molecules involved in the immune response, it may be difficult to draw valid conclusions, no matter how well designed the study or how compelling the data. For example, a humanized monoclonal antibody tested in an otherwise appropriate rat model might be expected to be highly immu- nogenic in that species. In addition, a recombinant vaccine antigen that is immunogenic and could lead to hypersensitivity reactions in man might not be immunogenic or allergenic in rabbit, rat, or mouse. The possibilities are endless and suggest that immunotoxicology testing must be carefully considered in concept and experimental design, that pilot studies are desir- able once a model has been selected, and that results be expertly interpreted before conclusions are made. In addition, even when all appropriate pre- cautions are taken, there is always a chance that the animal model cannot accurately predict immune reactions in humans. One notable example is the tragic results of a first-in-human clinical study performed by TeGenero, a pharmaceutical company in Würzburg, Germany. Severe unanticipated side effects occurred in March 2006, which resulted in what has been termed a cytokine storm, which threatened the lives of six health volunteers and ulti- mately resulted in the bankruptcy of TeGenero. The investigational product was a humanized monoclonal antibody developed to activate T regulatory cells of the immune system. In summary, preclinical safety assessments included in vitro studies, as well as a number of animal studies. Efficacy was demonstrated in rodent models and further supported by nonhuman primate studies. Immune safety was of primary importance, as the investi- gations continued toward conducting human clinical studies. A randomized first-in-human safety study was initiated with an intravenous infusion of TGN1412 at a conservatively low dose, which represented a very generous safety margin. The issues, challenges, and lessons learned from this horrific incident are summarized in Box 8.10. A reference for additional reading on this case study is provided in the additional reading section of this book.
Genetic Toxicology
Another relatively new subspecialty, genetic toxicology, studies the effects of chemical, biological, or physical agents on nucleic acids, genes, and chromo- somes. Biopharmaceuticals can profoundly affect genetic material, but the mode of action is usually quite different from that of small molecule drugs or ionizing radiation, insults that result in chemical changes to nucleic acids, producing mutations, chromosomal breakage, or abnormalities in controls. Cancer chemotherapeutic agents are examples. In contrast, certain biologi- cal products, notably genetic therapies and products containing DNA as the active ingredient, are designed to alter the genome through biological pro- cesses. They may deliver therapeutic DNA to the nucleus and even insert
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BOX 8.10 ISSUES, CHALLENGES, AND LESSONS LEARNED FROM THE TEGENERO INCIDENT
• Preclinical testing performed • In vitro safety profile: Flow cytometry, binding affinity,
and antigen-specificity studies • Safety and efficacy in rodents—normal healthy, rheuma-
toid arthritis, and autoimmune models • Safety and efficacy in nonhuman primates—demonstrate
that TGN1412 binds to primate CD28, NOAEL • Preclinical study conclusions
• No issues noted in a repeat-dose toxicity study at 4-week intervals
• No issues noted in a dose escalation study at weekly intervals
• No issues noted as a result of conducting numerous phar- macology studies in rats
• No observable adverse effect level (NOAEL) determined in cynomolgus monkey studies
• No sign of a first-dose cytokine-release syndrome (e.g., no immune system activation)
• No increase in cytokines (e.g., TNF) • Clinical study design
• Blinded, controlled, single center safety, and tolerance of single ascending dose
• Assess pharmacokinetics of the single dose at each of the four dose cohorts
• Determine effects on immune system (e.g., lymphocytes, cytokine profile, and immune response)
• Eight healthy, normal volunteers—two placebo and six investigational product (TGN1412)
• Intravenous infusion of TGN1412; eight subjects in the first cohort dosed 10 min apart
• Clinical adverse events • Adverse events began within 90 min post dose and esca-
lated in severity • Stage 1—Immune cell activation and cytokine release
(Continued)
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BOX 8.10 (Continued) ISSUES, CHALLENGES, AND LESSONS LEARNED FROM THE TEGENERO INCIDENT
• Stage 2—Shock, perfusion failure, hypoxia, and cytokine- mediated injury
• Stage 3—Multiple organ failure • Symptomatic subjects transferred to hospital critical care
unit 12–16 h post dose • Drug-related adverse events included mechanical ventila-
tion, recurrent fever, increased peripheral vascular perme- ability, and peripheral ischemia with necrotic fingers and toes
• Clinical laboratory findings • Significant and rapid increase in all cytokines on days
1 and 2 • Up to 1000-fold increase in TNF and IFN-γ at 2 h and on
day 2 • TNF spiked within 1 h of TGN1412 infusion • Cytokines decreased by day 3 • Significant decrease in CD3+, CD4+, and CD8+
• Lessons learned • Classification and sequence of adverse events: Cytokine
storm (1–24 h), reactive or transient (5–8 days), recovery (3–20 days), and long-term (15–30 days)
• Perform preclinical studies in accordance with Good Laboratory Practices (GLPs) with accurate data collection, reporting, and interpretation
• Highlight the future need for testing of biologics to be per- formed in a pharmacologically relevant species
• Reliance of dose calculation from preclinical safety studies using NOEL or NOAEL may not be sufficient and one needs to consider a broader approach, using all relevant informa- tion (e.g., product characteristics, biological potency, mech- anism of action, species specificity, and dose-response data)
• Clinical studies should be initiated with caution and be conducted in accordance with good clinical practice
(Continued)
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that DNA into the genome, making them suspect of causing genetic toxic- ity but by unique mechanisms. One example of a biopharmaceutical with the potential to cause genetic toxicity is a DNA molecule intended to repair or replace a gene within a cell of the target tissue, such as a white blood cell in the bone marrow. The molecular delivery system enhances the chance that foreign DNA in the product will enter a host cell nucleus and integrate into the host’s genome, thus enhancing its therapeutic potential. However, with this product, it is possible that the DNA would be delivered to the wrong tissue or cell, perhaps a gonad, enter the nucleus, and insert into cellular DNA of gonadal cells, perhaps even sperm or eggs. It might then be expressed in an uncontrolled or inappropriate manner or might even be passed to the next generation. Much of the genetic toxicity testing of biopharmaceuticals focuses on such possibilities, mainly on errors in well- intended gene delivery.
Genetic toxicity of a biopharmaceutical is typically studied both in vitro and in vivo, if a suitable animal model is available. An in vitro protocol might use mammalian cells in culture to determine the frequency at which inappropriate insertion or expression occurs. More definitive animal stud- ies are designed to inject biopharmaceutical into an animal and, using
BOX 8.10 (Continued) ISSUES, CHALLENGES, AND LESSONS LEARNED FROM THE TEGENERO INCIDENT
• Hospitalized subjects continued to improve. Five sub- jects were released approximately 30 days post dose; one remaining patient was hospitalized for almost two addi- tional months
• Clinical design for first-in-human study should be science based and carefully justified based on the compound tested
• Care must be taken in determining the route and rate of investigational product administration
• Timing of subject dosing is especially important to high-risk studies; in this study, a staggered dose administration by a couple of hours would have minimized the risk to perhaps only a couple of subjects
• The clinic must be prepared to provide appropriate and adequate emergency support under current practices, and this must be documented. In this study, the clinical research unit was located on the premises, and perhaps, this provided a false sense of security, which contributed to the lack of urgency to get the subjects to the hospital critical care unit
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sensitive nucleic acid probes, measure in various types of tissues or cells the nucleic acid that has been introduced. Additional studies of cells or tissues can determine whether the therapeutic nucleic acid is actually inserted into the genome of living cells. Returning to the example, a therapeutic plasmid DNA, intended to deliver a missing gene to myeloid cells in the bone marrow, could be injected into mouse bone marrow and then located and identified with nucleic acid probes. Locating the gene in bone marrow or blood cells 3 days after injection would be an expected finding and considered a desir- able event. However, discovery of this DNA in testicular or ovarian tissue of the same mouse would be a cause for concern, because the injected product might have entered the nucleus and even inserted into the genomic DNA of the germ cells. Although most biopharmaceuticals have little potential for influencing the genome or altering DNA or RNA, a few products have significant potential to do this. Here again, it is advisable for the sponsor to review regulatory guidance, identify and study precedence for their class of product, and seek expert advice, because genetic toxicology studies are long, arduous, and expensive, and failure to conduct them when required can lead to significant delays in development.
Tissue Binding or Local Tissue Tolerance
Biopharmaceuticals are sometimes given in large amounts to a single site on the body; most of them are injected. For example, a monoclonal antibody may be periodically injected subcutaneously in doses, each over 100 mg. A number of untoward reactions can result, and nonclinical study designs consider how local reactions are detected at the site of injection or deposition in an animal model. The mechanism of action can be quite different for each biopharmaceu- tical and tissue. Local immune and inflammatory reactions can result, espe- cially after multiple doses, and these may be chronic or acute. Cells or vaccine antigens can, by product design, remain at the site of injection and cause prob- lems that are not anticipated and are not immunologic, such as proliferation of fibrous or adipose tissue. A common method for studying tissue binding or local tissue tolerance is to add measurements of local reactivity to already des- ignated acute, subchronic, and chronic toxicology protocols. In one example, tissue samples are taken periodically by biopsy and again at the time of sacri- fice and studied for signs of local toxicity.
Inappropriate cell or tissue binding by a product could result in damage to tissue or even lead to untoward reactions or disease. What can be done to ensure that a biopharmaceutical, designed to bind a particular receptor or cellular molecule, will bind only to the intended target and not to the inno- cent bystander cells or tissues? Evaluation of nonspecific binding needs to be closely evaluated since many therapeutic biopharmaceuticals are developed for the purpose of binding to a certain cell surface molecule, and monoclo- nal antibodies directed against a number of proteins which places emphasis on this concern. Toxicology studies to determine tissue-binding patterns are
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called for whenever a product could inappropriately bind to normal cells. These studies are performed using immunohistochemistry and other meth- ods that clearly demonstrate tissue or cell binding or the lack thereof. Various human tissues (cadaver material) are used as substrates in these studies. Other approaches may be applied for certain types of molecules. As these studies are somewhat artificial, that is, they are performed in vitro and not in a living organism, the significance of cross-reactive binding study results may be unclear. In this case, it may become necessary to perform additional experimentation in live animals. As noted earlier, methods are now available to track biopharmaceuticals in a living animal.
Quality of Nonclinical Studies: Current Good Laboratory Practices
Consumers and the government recognized in the 1970s a need for quality in preclinical testing of drugs. The response was institution, by FDA, of a qual- ity system known as cGLP. This set of regulations, outlined in Chapter 4, is applied to all safety testing of biopharmaceuticals. For non-FDA-regulated products, such testing may be required by other government agencies respon- sible for licensing (Chapter 4) products released into the environment and simply contacting humans. It is important to consider the scope of the GLP regulation, stated in 21 CFR 58 “for conducting nonclinical laboratory stud- ies that support or are intended to support applications for research (clini- cal investigation) or marketing permits for products regulated by FDA” (CFR 1978). The scope further defines nonclinical laboratory studies as “in vitro or in vivo experiments in which test articles are studied prospectively in test systems under laboratory conditions to determine their safety” (CFR 1978). The definition goes on to exclude clinical studies and laboratory studies that are designed as exploratory or to determine potential utility or product char- acteristics. The scope of GLPs is important in three important respects. First, it does not apply to research, early development, or clinical trials. Second, it excludes quality control of product (which falls under cGMP). Third, it does apply to all laboratory studies, including animal studies, in which a claim is made for product safety. Thus, GLPs apply to most of the work we discussed in this chapter, with the possible exception (if safety claims are not made on the results) of pharmacology studies.
At the heart of GLP regulations are requirements for (1) a quality assur- ance unit (defined in Chapter 5) with broad authority to review and approve or disapprove of just anything and everything; (2) a study protocol; and (3) a study director. In addition, cGLPs are comprehensive, with every other aspect of laboratory operations and procedures, from facility standards to
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elements of animal feed, found in these regulations. Elements of cGLP are listed in Chapter 4. Finally, many of the terms used throughout this chapter to guide the scientific, management, and administrative aspects of a nonclin- ical study, including this term itself, were either introduced or institutional- ized by cGLPs on their introduction in 1976.
Summary of Nonclinical Studies
This chapter reviews methods used to assess the risk and benefit of a candi- date biotechnology product, with emphasis on biopharmaceuticals, as per- formed in nonclinical laboratory and animal studies. These studies begin once the nature of the biological construct or molecule, its purity, and its potency after early production have been characterized and defined. Initial safety studies are performed to determine product risk in animals before being evaluated in human clinical studies. Nonclinical study results are used to assess benefit versus risk and provide valuable information for designing the first-in-human clinical study. For pharmaceuticals, nonclini- cal testing provides the foundation for understanding the pharmacological and toxicological properties of the product in animals and in vitro assays. Of importance is to understand the different testing requirements associated with product-specific attributes. For example, in vitro laboratory tests are used to characterize the investigational product, both during the early stages of product development and throughout commercialization. The laboratory testing is then typically used to evaluate each lot of product that is intended to be used, to support additional preclinical safety testing platforms such as animal studies. Animal testing is conducted to support preclinical safety; these studies typically include absorption, distribution, elimination, and metabolism (ADME) studies used to determine the pharmacokinetics of a product and pharmacodynamic studies used to demonstrate how the prod- uct interacts with cells and tissues. In addition, a number of safety tests are performed to measure the toxicity of the product at predetermined doses and dosing regimens intended to match or exceed those to be used in the pro- posed future human clinical study. Toxicity studies are designed to measure acute, subchronic, or chronic effects of the product, as well as to screen for specific types of toxicities. Nonclinical studies are designed and performed to predict human risk. This very important safety testing is conducted under a quality system, cGLP. In a nutshell, studies conducted in compliance with cGLP are prospectively designed, written, approved, and well documented. A final study report is typically generated to include results, summaries, conclusions, tabulated data, figures, and tables. The data in the final study report are verified by an independent reviewer, usually representing quality
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assurance, and the report is finalized with a required signature approval from both the study investigator, the study director, and a quality assurance representative.
Reference
CFR. 1978. Good Laboratory Practice for nonclinical laboratory studies. Title 21, CFR Part 58. US Government Printing Office, Washington, DC, Source: 43 FR 60013.
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9 Clinical Trials
Introduction to Clinical Trials
A clinical trial, also referred to as clinical study or clinical research, is the overall process of evaluating the safety and efficacy of a medical product or an intervention in humans. Importantly, it is investigational and the purpose of a clinical trial is to learn about a product and how it impact humans. The intention is not to treat patients but rather to provide data regarding safety and efficacy in humans. Successful completion of several clinical trials is required for market approval of drugs, biologics, and some medical devices by FDA, and hence, clinical studies are used in the biotechnology industry to support market approval of biopharmaceuticals. However, the concepts developed for clinical studies incorporate scientific and design elements shared with field trials of other biotechnology products for which there is no testing in humans, such as field studies of environmental or agricultural products.
In the scheme of biopharmaceutical development, clinical research fol- lows animal studies and other preclinical testing and evaluation, because a product is always sufficiently tested for safety and performance in labora- tory studies before it can be evaluated in human clinical studies. Human clinical research, the first research that uses healthy volunteers, is divided into several phases of clinical development (Figure 9.1). Each subsequent phase becomes increasingly large or complex and shows a continual shift in focus from measuring product safety in small studies—often in healthy individuals—to measuring both efficacy and safety in a larger patient popu- lation. Collectively, clinical development is a long, complex, and expensive process based on scientific methods, data integrity, and a high degree of reg- ulatory oversight.
Clinical trials are an important aspect of biotechnology development, because a large number of firms develop medical products—drugs, bio- logicals, or medical devices—and because before commercialization, each medical product must be extensively tested in humans. Some biotechnol- ogy firms plan to take their product through all phases of clinical develop- ment to market approval. Others have a different business strategy and plan
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Phase 1 regulatory review
and approval (initial IND)
IRB review and approval
Protocol and supporting documents
Phase 1: Concept protocol and design
Phase 2 protocol regulatory
submission and review
Phase 2 supportive clinical studies Pharmacokinetics
Food effects QT/QTc
Special populations Mass balance
Clinical Study Report (CSR)
Phase 1/2 supportive clinical studies Pharmacokinetic
Pharmacodynamic Dosing and dose schedule
IRB review and approval
Phase 2: Concept protocol and design
Phase 1 clinical trial
Phase 3: Concept protocol and design
Phase 2 clinical trial
Phase 3 protocol regulatory
submission and review
Phase 3 clinical trial
License application and
approval
Postmarket approval/ Phase 4 clinical trials
Regulatory requirements
IRB review and approval
Elective studies
Follow-on safety New indications New populations: children, elderly,
and so on
New formulations, delivery systems,
and so on
CSR
CSR
Protocol and supporting documents
Protocol and supporting documents
FIGURE 9.1 A typical scheme of clinical trials in biopharmaceutical development. In an electrocardiogram trace the QT interval is a measure of time between the start of Q wave and the end of the T wave in the heart’s electrical cycle (QT). The QTc represents a heart rate “corrected” QT measure.
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to evaluate the product only in early phases of clinical development, until there is added value through proof of product safety in man. Most often, a biotechnology firm with candidate biopharmaceutical product will strive to add further value to that product by initiating human clinical research studies. Despite this strong desire to increase product value by performing a clinical study, small biotechnology firms often have little or no experience in this endeavor. In addition to lack of experience, many biotechnology firms have a culture of intellectual liveliness, in contrast to rather somber tone of clinical development, where there is an emphasis on scientific proof beyond reasonable doubt and thorough and sometimes detailed or rigid quality pro- cedures based on government regulations and guidelines. The result is that to be successful in clinical research, many biotechnology firms find that they must change, to some degree, their culture and means of doing business. Another option for a biotechnology firm would be to form a strategic alliance with a large industry partner who has experience and resources to conduct the necessary clinical studies. This strategy would surely provide the most expeditious route to biopharmaceutical product development and associated human clinical studies. However, this mechanism comes with a price, usu- ally in the form of the partner obtaining an equity share in the biotechnology firm and gaining some level of control over decision making and the devel- opment path forward.
This chapter attempts to explain the basic elements of clinical research, as they apply to the development of medical products. It begins with a brief his- tory of the field, provides an overview to introduce concepts and terms used in clinical research, and progresses to a section on clinical planning. Further information is provided on the design and conduct of clinical studies, with extensive discussions concerning the people and institutions involved in a typical trial. A section is dedicated to clinical trial operations, which dis- cusses each phase of clinical development. Another section covers quality systems for clinical trials, that is, current Good Clinical Practices (cGCPs). This chapter ends with a discussion of ethical behavior and the importance of ensuring the well-being of human subjects enrolled in clinical research.
Background of Clinical Research
Introduction
Historically, clinical research has been considered as either observa- tional or experimental. In observational studies, the investigator has no control over the study conditions, and investigational drugs and placebo are not given to the subjects. These studies may be referred to as epide- miological, because they measure current conditions absent novel medi- cal interventions. Although observational studies may form a foundation
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for conducting an experimental clinical study or for following safety of a biopharmaceutical post licensure, they are not used to study the safety and efficacy of a particular biopharmaceutical during the development process. For experimental clinical studies, also referred to as controlled studies, the investigator designs the conditions for the study and follows that design in an exact manner. Clinical studies of biotechnology products performed for the purpose of market application, that is, premarketing studies, are always controlled clinical studies. Most controlled clinical trials compare two or more treatment modalities. The specific treatment schedule with the test product is predetermined, and all other treatments to or medical con- ditions of the subjects are managed as similarly as possible. The design of a clinical study for a biopharmaceutical may thus compare the test product with a marketed product used to treat the same indication (a comparator) or, if no appropriate comparator is available, to a placebo (a sugar pill) used to minimize or control study bias.
Historical Information on Clinical Trials
Comparative drug studies were first reported in the eighteenth century but used infrequently until the twentieth century when, as noted in Chapter 3, laws and regulations were developed to ensure that drugs and medical devices were adequately tested for safety and effectiveness. Indeed, these regulations have, in part, influenced clinical trial designs and are based on good scientific research practices that had been used in laboratories, nota- bly the need to test a hypothesis in a formal manner. Another process called randomization or the blinding process, that is, assigning a patient to receive one or another drug, was first used in laboratory and agricultural field research and then was adopted as good scientific practice for clinical trials. Monitoring and auditing, now standard practices for clinical trials, were earlier used in nonclinical studies and were found to be an effective means of ensuring the quality of data. Indeed, as drug development became more complex and clinical studies grew larger, many scientific and quality prac- tices were applied to clinical trials, which have now become the norm for biopharmaceutical clinical research.
In the 1970s, the quality and ethical aspects of clinical trials came to be known as cGCPs, and by the 1990s, cGCPs had gained international accep- tance, as the quality system for clinical trials. During this period, the num- ber of clinical trials grew dramatically because more drugs, medical devices, and biotechnology products entered clinical development, more countries required studies to be performed on their own soil, and regulatory agencies demanded greater numbers of ever-larger studies for each product. Further, clinical investigators and statisticians devised better clinical study designs, more effective means of conducting studies, and improved means of col- lecting and analyzing data. Today, many thousands of clinical studies are underway each day in dozens of nations and throughout the world.
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Clinical research plays an increasingly important role in the development of biopharmaceutical products and also in their postmarket approval evalu- ation. Each clinical trial is also becoming increasingly complicated, expen- sive, and publically announced. Indeed, results of clinical trials are relayed almost daily in newspapers and on television. The increased complexity and demands of clinical research have led to more regulatory requirements and an increased need for documentation. Indeed, some would argue that expan- sion of clinical research could someday outgrow the ability of the clinical research community to provide infrastructure for all ongoing trials. Others disagree and feel that careful planning and exact execution of clinical devel- opment makes for a successful clinical study, allowing a sponsor to deter- mine whether or not a product is safe and efficacious in man and, therefore, fit to be marketed for the intended purpose. Generalizations in either case are not correct. Well-designed and executed clinical studies are and always will be required to demonstrate the safety and effectiveness profile of any biopharmaceutical. Clinical research requires careful planning to ensure that a valid study is completed and to fully meet the rights and interests of each volunteer enrolled in any study.
To provide an overview of clinical trials, this chapter examines the structure of clinical research principles and activities, as they are applied to the devel- opment of medical products in the biotechnology industry. We focus on key issues that one must consider for successful clinical development of any prod- uct: organization, planning, personnel, operations and processes, documenta- tion, quality, ethics, and resources.
Organization of Clinical Research
Phases of Clinical Trials
Clinical development of biopharmaceutical products is divided into four distinct and sequential phases, identified in Figure 9.1. These phases pro- ceed from the most controlled conditions and environment (Phases 1 and 2) to a scenario that is closer to the real world (Phases 3 and 4) in which the product is used to treat patients. Other terms, including the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use nomenclature, are provided.
• Phase 1. Early phase. Clinical safety and toxicology. Clinical pharmacol- ogy. Human pharmacology: The first administration of a new prod- uct to man at a fixed route and schedule is considered a Phase 1 trial. Typically, the four goals of a Phase 1 study are: (1) to estimate the maximum tolerated or safe dose; (2) to determine whether any organ systems are affected by the product; (3) to identify any toxicity
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related to the product, and if toxicity is identified, to measure the extent, duration, and reversibility; and (4) to observe any unantici- pated (desirable or undesirable) activity of the product.
• Phase 2. Expanded dosing. Pharmacokinetics. Therapeutic exploratory: The concept and definition applied to Phase 2 study has been somewhat misused and broadened by the biotechnology community. However, a textbook definition suggests that this phase of clinical develop- ment explores different dosages of the product, compares the effects of doses of the product to those of a placebo and, ultimately, deter- mines which dose has the best safety and efficacy profile. Results of Phase 2 set the stage for design of the Phase 3 clinical trial.
• Phase 3. Pivotal clinical trial. Therapeutic confirmatory: In this phase of clinical development, the selected dose is given to a much larger number of individuals, each representing the target patient popula- tion that was established in the product labeling. A Phase 3 trial is referred to by regulatory agencies as a pivotal trial, because it is the clinical basis for marketing approval or disapproval decisions.
• Phase 4. Therapeutic use: Postmarketing studies aim to further study safety and efficacy or to extend the indication or use of the product.
The Science of Clinical Research
A clinical program is designed just as any scientific research project, and the design must be scientifically sound, that is, it should test a hypothesis or a series of hypotheses, have clear objectives, and identify measurable out- comes. Quality in performance of clinical research is also important, and proper collection of data is essential. Use of other scientific tools, such as blinding of procedures and application of placebo or comparator, statistical analysis of data, and objective interpretation of results, is an essential ele- ment of clinical research. Unfortunately, some in biotechnology may forget that good clinical research is based on the scientific method and ethical guidelines and is not simply a business or entrepreneurial endeavor; hence, clinical research should be highly regulated and structured.
Clinical research is the definitive step in evaluating new biotechnology products for safety and efficacy, that is, the prevention, diagnosis, or treat- ment of disease. Clinical trials, and the systems that support them, are com- plex endeavors and require collaboration among investigators, industry, academic institutions, and government agencies. Adding to their complex- ity, clinical trials have undergone a number of changes in the past decade. Progress in biomedical sciences and biotechnology has accelerated and increased the need for clinical studies and created new opportunities to improve clinical trial processes. Advances in informatics, laboratory and clinical diagnosis, and data management have led to new ways of evaluat- ing human subjects and reporting information and data. However, a new
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biopharmaceutical product can only be brought to market by a firm if the product is first shown to be safe and effective. Stated another way, clinical development is outcome driven and the outcomes are issue focused and, ultimately, based on meaningful scientific data, which, in turn, are based on a relevant hypothesis and clinical study design.
Quality in Clinical Research and Current Good Clinical Practices
Current Good Clinical Practice is an ethical and scientific quality standard and a quality system applied to designing, conducting, recording, and reporting clinical trials. Although cGCP has taken on a definition of regula- tory compliance, it is actually more than that and represents an established means of conducting a clinical trial. There are national and international standards for cGCP but the ICH standard is the most commonly accepted version and the one followed today by most biotechnology firms and in most nations. Quality and cGCP in clinical trials are discussed later in this chap- ter, and the elements are included throughout the text.
Clinical Development Planning
The key to successful clinical research programs in this ever-changing and fast-paced environment is effective clinical development planning. Planning must focus on the clinical claim or claims and, as noted in Chapter 1, this is best stated in the targeted product profile (TPP). The primary purpose of the label claim in the TPP is to inform prescribers and patients about the documented benefits of a product. Clinical outcomes, derived from clini- cal trials, provide the basis for label claims. Further requirements for clini- cal planning are based on FDA regulations and guidelines. For example, a major requirement of an Investigational New Drug Application (IND) is the investigational plan, a section of the document that outlines the sponsor’s intended clinical research program (Chapter 3). The TPP for a biopharma- ceutical states medical objectives of the product, including the indication and therapeutic and safety profiles.
Using a well-conceived and well-written TPP with a list of desirable out- comes, a clinical development plan can be written as a predecessor to the product development plan (PDP), both described in Chapter 1. Note that the clinical development plan is not a stand-alone document; it is instead an important but integral part of and is woven into the overall PDP. In addi- tion, the clinical development plan is a living document and, like the overall PDP, can be changed at any time, as long as the changes are coordinated with other aspects of the PDP and with members of the product develop- ment team.
The clinical development plan is a written document that describes how a new biotechnology product can be progressed, in an orderly and timely manner, from first administration in man to postlicensure studies.
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It explains to colleagues, management, and regulatory agencies, the pro- posed product’s clinical development scheme, based on current knowledge, and also provides critical information to the development team, such as the rationale; time frame of Go/No Go decision points; costs, both internal and external; and an outline of the proposed clinical studies. A well-constructed clinical development plan also addresses, in a concept protocol, the important scientific, medical, and operational issues or factors, as listed in Chapter 1. Thus, the clinical development plan brings together all elements— scientific, management, and operational—into a cohesive document. As a living docu- ment, the clinical development plan provides the opportunity for adjust- ment, as new information, including preclinical, manufacturing, and clinical information, is obtained from the ongoing research. Furthermore, a complete and current clinical development plan allows the firm, investors and/or partners, and FDA the opportunity to provide valuable feedback on the future clinical milestones and regulatory path (e.g., recognize an alter- native development strategy such as qualify for orphan drug status or fast- track approval process).
Infrastructure for a Clinical Trial: Individuals, Documents, and Investigational Product
Earlier, we noted that experimental clinical studies test a hypothesis, have a written design or protocol, use scientific research methods, and are carefully controlled endeavors, employing human volunteers and professional staff. The planning aspect is referred to as clinical study design. The elements of a study design, the individuals involved in a clinical study, and the documents that support and control a study are discussed in this section. The qualifications of clinical study staff and their responsibilities are given in Box 9.1. A list of clini- cal study documents and the primary purpose of each are given in Box 9.2.
Design of Clinical Trials and the Clinical Protocol
Once a clinical plan has been completed, the clinical professionals at a bio- technology firm now focus on each clinical study identified in that plan. The basic elements of a study are outlined in a document, usually not exceeding 10 pages, referred to as the concept protocol. This is really the scientific basis for a study design and represents the initial proposal of experienced profes- sionals, such as a medical director, a clinical project manager, and various investigators. Elements of a concept clinical protocol are listed in Box 9.3. The hypothesis being tested is paramount, but other matters are also important. Objectives are keys to success. Study size, patient population, and indication are also critical to experimental design. As noted below in greater detail,
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Phase 1 studies are smaller and focus only on safety, whereas Phase 3 tri- als are typically large, multicenter endeavors that evaluate both the efficacy and the safety of a product. Once the design has been drafted, the firm must consider the management and operational elements that will support the study. This evaluation sometimes reveals the design to be overly complex and demands that the clinical protocol be revised to make it more feasible, from an operational point of view. Alternatively, it might suggest that the number of subjects in each group is too low if a meaningful conclusion is to be drawn or that the enrollment targets are too aggressive, with historic patient populations not likely to be enrolled in the designated timeline. It is especially important to ensure that the nonclinical safety study plans cover the recommended dose and dosing schedule. Noting the nonclinical product needs, it is important to ensure availability and sufficient supply of investiga- tional product that is needed to support the proposed human study design, as it relates to dose and manufacturing scale. Normally, the biotechnology firm asks experts, such as statisticians, toxicology scientists, experienced clinical investigators, and, often, FDA, to review and comment on the design
BOX 9.1 CLINICAL TRIAL: INDIVIDUALS AND RESPONSIBILITIES
• Volunteers, patients, and human subjects
• Sponsor and staff
• Medical director • Safety monitor • Auditor • Medical writer • Clinical project manager • Regulatory staff • Manufacturing and clinical trial materials
• Principal investigator and staff
• Subinvestigator(s) • Nursing staff • Recruiter
• Statistician
• Institutional review board
• Board chair and board members • Administrative staff
• Quality assurance unit
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BOX 9.2 CLINICAL TRIAL: DOCUMENTS
• Concept protocol: A brief design of a clinical study used as the basis for discussions between sponsor, investigator, and regulatory authorities. It is the foundation for preparing the full protocol.
• Clinical protocol or protocol: An instructive document that identi- fies exactly why, how, and with whom a clinical study will be performed and provides schedules of events.
• Informed consent (IC): Detailed procedures used and the time frame required to obtain informed consent.
• Informed consent form (ICF or CF): This document explains to a volunteer the potential risks and benefits of a clinical study. To enroll in a study, a volunteer must understand and sign the CF.
• Investigator’s brochure (IB): An informative document that identi- fies for each member of the investigative staff the information on the clinical study, the product being tested in the study, and the possible risks and benefits to the volunteers enrolled in the study.
• Form FDA 1572: Statement of investigator captures information on the investigative teams and is an agreement by the investi- gator to follow the protocol and regulations regarding clinical studies.
• Curricula vitae: Resumes of the principal investigator and key investigational staff.
• Clinical trials agreements: This agreement between a sponsor and an investigator or a clinical contract research organization out- lines responsibilities of each party to perform a clinical study.
• Form FDA 3674: This represents certification by a sponsor to disclose clinical trials information to clinicaltrials.gov, accord- ing to the U.S. law.
• Forms FDA 3454 and 5455: Financial Disclosure. In these doc- uments, the sponsor discloses to FDA the financial arrange- ments that exist between clinical investigators and the sponsor.
• Case report forms (CRF): These are paper or electronic forms on which the investigator enters medical information gath- ered during a clinical trial.
• Patient diary: These are the forms completed by subjects during the outpatient phase of a clinical trial to capture data on pos- sible AEs and general medical condition of the individual.
(Continued)
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contained in the concept protocol. Thus, the overall clinical objective and the objectives of each phase of development, both provided in the clinical plan, drive the full clinical study design, as outlined in the concept protocol.
The document that ultimately describes in detail the clinical study design is referred to as the clinical protocol, and this is written by clinical study staff once a concept protocol is acceptable. The most important step in any clinical study is to prepare a complete, well-organized, and scientifically sound pro- tocol. Protocols can be changed, or amended, but amendments take time and incur cost. Therefore, to avoid delays, the protocol and other clinical docu- ments consider, and provide for, every eventuality likely to occur once the study begins. Responsibility for writing a protocol may rest with the sponsor or the principal investigator (PI). The sponsor is a representative of the biotech- nology firm, whereas the (physician) investigator is the person responsible for conducting the study in accordance with the protocol. Thus, an investigator is retained by the sponsor. Today, national and international guidelines provide a standard organization, a template, for the clinical protocol.
The elements of a protocol are listed in Box 9.4. A heading sheet gives a fully descriptive title, names the investigator and their institute or employer (affiliation), and identifies the sponsor. Most institutions give a unique num- ber to each protocol. The second sheet, a signature page, provides the names and contact information of everyone responsible for the protocol and, under a statement of compliance, prompts for the signatures of both the PI and the sponsor. A summary of the protocol is typically provided in the next section, and this begins with a statement of the objectives and, in most protocols, the formal hypothesis to be tested and is followed by a brief summary of infor- mation on the product under investigation.
BOX 9.2 (Continued) CLINICAL TRIAL: DOCUMENTS
• Operations or administrative manual: This collection of adminis- trative and management information and instructions guides performance of the clinical trial and supplements the protocol.
• Form FDA 3500 MedWatch: This is a standard form used to report safety information and adverse events to FDA.
• Investigational product accountability log: It is used to document the receipt and distribution of all investigational products.
• Transfer of obligation log: It serves to document any or all obliga- tions transferred by the clinical investigator to other clinical staff members.
• Screening log: It is used to document all subject-screening events and usually includes documentation of subjects that meet study participation criteria.
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The study design is then described in some detail, because it is the sci- entific heart of clinical research. Since most studies compare the treatment under investigation to another treatment (comparator) or to no active treat- ment at all (placebo), the design describes how the comparison will be made in the study design. It includes a description of the dose or doses of product and placebo or comparator, duration and intervals for giving doses (dos- ing), and a description of dosage forms. Typically, patients are divided into groups, or cohorts, and members of each group are given one or another treatment or dose. Further, design criteria may include use of randomization, whereby patients are randomly assigned to one or another group, or blinding, the process of keeping the exact treatment for each patient hidden from the subjects, PI, study staff, and sponsor. These elements of an experimental design prevent bias from entering into a study and are absolute require- ments for late-stage studies performed in the U.S. Bias, a predisposition to
BOX 9.3 ELEMENTS OF A CONCEPT CLINICAL PROTOCOL
• Description of the biopharmaceutical investigational product • Previous use in man or animals • Stated indication • Protocol title • Potential clinical investigator(s) • Anticipated study duration • Study phase • Intervention regimens • Study objectives • Study hypothesis • Subject population and general characteristics • Major inclusion and exclusion criteria • Study design and schedule or duration, number of subjects and
groups, and • study site(s) • Study schema • Study end points: Safety and direct or surrogate efficacy • Study procedures and methods (primary, in general) • Assessments • Stopping rules • Unique scientific, ethical, or medical aspects
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a particular outcome or a prejudice, and any element of design in a protocol that might lead to bias are carefully avoided in the study. Other methods may be applied to a clinical study design to avoid bias, improve study per- formance and validity, or ensure safety and well-being of human volunteers. Stopping rules are descriptions of how a study will be halted, temporarily
BOX 9.4 ELEMENTS OF A CLINICAL PROTOCOL
• General information: Title, numbers, names of investigator, and sponsor, version-controlled.
• Background information: Description of the product and how and when to administer.
• Trial objectives, with purpose and stated hypothesis. • Trial design: Scientific design and factors that ensure or enhance
the design. • Selection and withdrawal of subjects: Inclusion and exclusion crite-
ria; withdrawal of subjects. • Treatment of subjects: Administering medications and monitor-
ing subjects. • Assessment of efficacy: Efficacy measurements and end points. • Assessment of safety: Safety measurements and end points. • Statistics: Data sets and statistical analyses. • Access to source documents or data. • Quality control or quality assurance: All aspects of compliance
and quality; monitoring procedures. • Ethics: Ethical standards for the study. • Stopping rules: Clearly defined stopping rules resulting from
safety signals or adverse events. • Safety reporting: Specific adverse event-reporting requirements
to clinical investigator, IRB, and FDA. • Data handling and record keeping: Management of data during
and after the trial. • Finance and insurance: Responsibilities for payments and liabil-
ity insurance. • Publication policy: Anticipated publication and authorship
policies. • Appendices: For example, treatment and test charts and sched-
ules, consent form, standard medical guidelines, publication policy, and references.
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or permanently, should a certain type or series of unanticipated events or adverse events (AEs) be noted during the study. These rules provide a means of enhancing study safety and well-being of human participants. Stopping rules are explicit and fully described in the protocol, and once approved by FDA, they serve as an important agreement between the PI, the sponsor, and their regulatory agency.
Other important aspects of a clinical study design are rules for selection and enrollment of patients, that is, inclusion or exclusion criteria, and for with- drawal of the subjects. It is important to carefully select subjects, enrolling only those who meet stringent criteria. In many Phase 1 studies, the investi- gator may wish to enroll only healthy or normal subjects, who are best suited for studying a product on its first introduction to man. In many other studies, the subjects represent the actual patient population, as described in the TPP, that the biopharmaceutical is intended to treat. In either situation, it is impor- tant to include in the study a specific type of individual and to exclude those who have other medical conditions or a disease that could put them at risk of undesirable reactions to the product or that could potentially confound the study results. Hence, inclusion and exclusion criteria are written into a protocol. Inclusion criteria identify attributes that the patients must have to enter, or enroll in, the study, whereas exclusion criteria identify issues that make a potential subject ineligible to enroll or participate in the study. For example, if one were to study a biotechnology product that was intended to lower blood pressure in otherwise healthy individuals, hypertension (high blood pressure) would be an inclusion criterion, whereas severe or advanced cardiac disease might be an exclusion criterion. A list of inclusion and exclu- sion criteria that might be applied to a Phase 1 clinical study in which nor- mal, healthy individuals were enrolled is given in Box 9.5.
A number of other rules, for example, how to replace, with a new sub- ject, those who withdraw from a study, may be given in the design section of a protocol. Sometimes, clinical studies must be terminated or particular subjects must withdraw, either voluntarily or on request of PI. The protocol describes in the design section how these types of enrollment decisions are to be made and carried out.
The treatment of subjects with the test product is normally described in great detail in the protocol. To prevent bias, product is administered to each subject in exactly the same manner, at a prescribed amount, and on an established schedule.
Assessment of safety and efficacy is essential to the success of a clinical study and a protocol explicitly describes how each is measured. Outcomes, broad results, or visible effects that form the basis for the study hypothesis are described in medical terms. An example of an outcome in the case of a biophar- maceutical product intended for treatment of lung cancer might be to remain free of tumor for 1 year. Second, one or more end points are clearly stated. End point is the term used to identify a measurable parameter, again exactly medically defined. End points must reflect the objectives and the disease that
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BOX 9.5 EXAMPLES OF INCLUSION AND EXCLUSION CRITERIA
• Inclusion criteria: • Age: 18–50 years • Sex: Male or nonpregnant female • Good general health, as demonstrated by medical history,
baseline laboratory tests (urinalysis, clinical chemistry, and hematology), and physical examination
• Laboratory values within 1.25 times institutional stated normal values
• Negative test results for HIV-1 and Hepatitis-A, -B, and -C • Low risk of coronary heart disease based on National
Health and Nutrition Examination Survey-1 cardiovascular risk assessment and screening electrocardiogram
• Negative tests for autoimmune diseases, rheumatoid arthri- tis, and antinuclear antibody
• Reliable access to the clinical test facility and availability to participate for the duration of the study
• Assessment of the understanding of questionnaire com- pleted before enrollment and demonstration of understand- ing of risks and benefits associated with study participation
• Ability and willingness to provide informed consent • If the participant is female and of reproductive potential, she
should: – Have a negative serum or urine beta human chorionic
gonadotropin pregnancy test performed within 3 days before study initiation
– Agree to consistently use effective contraception from 21 days before study initiation and for the duration of the study
• Exclusion criteria: • Prior receipt of similar biopharmaceutical • History of confirmed diagnosis of (disease or condition)
within the last 2 year • Use of (specific drugs) within 5 months of enrollment or
use of (specific drugs) within 2 months of enrollment (Continued)
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BOX 9.5 (Continued) EXAMPLES OF INCLUSION AND EXCLUSION CRITERIA
• Recent (within 2 weeks) use of (specific drugs) with (spe- cific effects or drug indications)
• Anticipated use of medications known to interact with (investigational class of biopharmaceutical)
• Use of any investigational or nonregistered drug or vac- cine or whole blood or blood product within 90 days of enrollment.
• Use of systemic immunosuppressive medications or cancer chemotherapeutic compounds within past 90 days
• Current or past diagnosis of Type I or Type II diabetes • History of severe allergic reactions • Screening laboratory abnormalities beyond the limits
defined in the inclusion criteria • Clinically significant medical condition, physical examina-
tion findings, other clinically significant abnormal labora- tory results, or past medic history that may have clinically significant implications for current health status in the opinion of the investigator
• Any contraindication to phlebotomy • Body mass index <19% or >30% • Acute illness at the time of enrollment • Pregnant or lactating female or female who intends to
become pregnant during the study period • Serologic positivity for Hepatitis B or C or HIV-1 • Psychiatric condition that precludes compliance with the
protocol, including ongoing risk for suicide or psychosis • Suspected or know current alcohol abuse or recreational
intravenous drug use within the last 12 months • Acute illness at the time of enrollment • Any other condition that, in the judgment of the investi-
gator, would interfere with or serve as a contraindication to protocol adherence, assessment of safety or reactogenic- ity, or a participant’s inability to give informed consent, or increase the risk of having an adverse experience to the study drug.
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is being treated, prevented, or diagnosed. In the lung cancer example, an end point might be tumor mass found in lung. Third, to adequately evaluate end points, that is, the measurements, the act of determining an amount or quan- tifiable dimension of that end point is necessary. In the lung cancer example, the measurement of tumor mass, number, location, and size of each, once each month, by a radiological method, might constitute a valid measurement. Safety end points are also measured. For example, to determine whether the patients became allergic to a biopharmaceutical product, the protocol might direct the investigator to carefully search for rashes after each treatment. The success of a clinical study rests on establishing in the protocol meaningful and exact out- come, end points, and measurements. Hence, medical experts are often asked to advise this phase of protocol development, and in later phases of clinical devel- opment, these issues lead to important discussions with regulatory authorities.
Safety, as well as efficacy, end points are described in the protocol. Should they occur in a subject, these are recorded as AEs if they are mild or limited in scope and severity and as serious AEs (SAEs) if they are severe, such as ana- phylactic shock. When faced with an AE or SAE, the investigator or another physician must determine whether the reaction is, or might be, related to the investigational product. Indeed, each protocol, no matter the phase, states a large number of safety measurements, such as clinical laboratory tests and physical examination, that must be taken for each subject to determine the safety and tolerability of the test product. Other measurements focus on the efficacy of the product being tested.
The statistical section of the protocol describes, in detail, the analyses that will be used to make comparisons of end points in the overall subject popula- tion. It also applies statistical principles to support the design of the study. For example, the statistical discussion provides rationale for the number of subjects enrolled in each group, treatment or placebo, of a Phase 3 clinical trial. Another section of the protocol deals with administrative issues, such as control of the test product review, approval of study documents, methods for collecting and recording raw data, and details such as insurance, or publication policy.
Most clinical trials enroll the total subjects numbering in dozens (Phase 1), hundreds (Phase 2 or Phase 3), or low thousands (Phase 3). Yet, other clinical studies of biotechnology products, notably pivotal Phase 3 trials, are quite large and are conducted simultaneously by many investigators at several sites. Today, Phase 3 international trials may enroll in excess of 60,000 sub- jects at more than 100 sites in more than 20 countries. Such big trials demand much administrative support and instruction. For these purposes, a clinical operations manual is used in addition to the protocol. The Ops Manual is an extension of the protocol and describes in greater detail all administrative aspects of the study and provides detailed medial and managerial instruc- tion to the staff involved in the clinical study. Using operations manuals is a good business practice, because these manuals further ensure success of a scientifically well-designed study by providing consistent procedures at each trial site.
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Human Subjects, Patients, and Volunteers
A clinical trial includes humans willing to participate and receive either the product or, perhaps, the placebo. Later in this chapter, we describe the rights of those individuals who volunteer to receive investigational prod- ucts on behalf of the sponsor and the PI and, hopefully, for the betterment of the health and well-being of all mankind. For this, we, in biotechnology, greatly appreciate their participation. Since every volunteer enrolls in a clini- cal study by his or her own free will, we refer to these individuals with the general term volunteer. For products that could provide some benefit and are used in individuals with a disease or a medical condition for which the product is indicated, the volunteers are referred to as patients. In some stud- ies, such as with preventive biopharmaceuticals (e.g., a vaccine) or where the studies enroll healthy individuals (e.g., Phase 1), the volunteers are referred to as subjects. We use the terms volunteer, subject, and patient without fur- ther definitions in this chapter.
The Sponsor
In the case of industry studies, the sponsor typically is not only the finan- cial and resource backer of a clinical study but is also the IND holder and, as such, takes ultimate responsibility for the study. Responsibilities of sponsors before, during, and after a clinical study are clearly defined in regulatory guidelines. First and foremost is the responsibility of ensuring the rights and well-being of every human subject or patient and of maintaining quality, through quality assurance and quality control, of the trial. To meet this obli- gation, the sponsor of a clinical study has policies and procedures that dem- onstrate exact intentions. A sponsor may delegate responsibilities to another party, but this must be specific and in writing. Such is the case when an individual known as PI is retained to perform the study or when a contract research organization (CRO) assumes various functions and responsibili- ties of the clinical trial. Smaller biotechnology firms often delegate most or all clinical trial functions to others, but they can never transfer the ultimate responsibilities of ensuring that a study is conducted, recorded, and reported properly or that patients are always treated according to medical and ethical standards. A biotechnology firm engaged in clinical studies always has, on staff or retainer, clinical trial experts. Indeed, most sponsoring biotechnol- ogy firms retain internally the monitoring and auditing function of clinical trial and data management, thus ensuring the integrity of data and attend- ing to financial and general administrative duties.
An important and yet often overlooked responsibility of the sponsor is the selection of a qualified PI and, along with the investigator, the institution or the CRO at which the study is to be performed. This is often a difficult task for the biotechnology firm because, with both an exciting technology and ade- quate investment, the firm’s management may be faced with several quali- fied investigators, each of whom wishes to perform the study. Some may be
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inexperienced or otherwise unqualified to head an important clinical study but appear knowledgeable about the product. Others may have years of expe- rience as investigators but be inexperienced with this type of product. The sponsor must find a person who is both qualified scientifically and has the proper experience with the product type; finding or selecting the right PI can be a challenge for the Sponsor and is further complicated with multicenter trials, where several qualified investigators must be identified.
Sponsors ensure that all the paper work is completed during the trial. In addition, as one might expect, a great amount of paper work is, in fact, gen- erated before, during, and after each clinical study, no matter how small the trial. The IND sponsor, not the PI, communicates directly with regulatory agencies. The sponsor confirms that review was completed and approval was given by the institutional review board (IRB). Through the Investigator’s Brochure (IB), the sponsor informs the investigator and his or her staff about the product to be tested. Both the IB and IRB are described later.
The sponsor also plays important roles in relating safety information in a timely manner. First, there must be a system in place to receive review from investigators and, if necessary, report any safety information that is generated during the study. The AEs and SAEs are collected by the sponsor in a timely manner, and SAEs are immediately investigated and promptly reported to those whose job it is to influence, make, or review medical decisions, that is, the investigator, the medial monitor, the IRB, and the regulatory authorities. The sponsor has in place a system of expert review for AEs and SAEs; this is typically the job of the medical (or safety) monitor, a physician who examines each event and reports his or her opinion regarding the significance of and the relationship between SAE and the product to the sponsor. If, in the eyes of the investigator or the medical monitor, AEs or SAEs are related to the test product and certainly if the safety of the patients is at risk, the sponsor is responsible for reporting the events to all investigative staff and regulatory authorities and, in some cases, for stopping the study. Termination of a study, meaning that the product can no longer be given, is driven by detailed rules or study stop criteria, which are also provided in the protocol.
To ensure that the clinical study is being conducted properly, all aspects of the study must be audited or monitored by an experienced and knowledge- able individual, on behalf of the sponsor. Clinical trial monitoring is not the same as the role ascribed to the medical (safety) monitor, described above. In contrast to the medical (safety) monitor, who reviews AEs or SAEs pro- vided to them, trial or study monitoring is a process that involves visits by a professional auditor to the clinical study site at regular intervals to inspect the clinical study documents and medical records. This auditing or moni- toring process further ensures, among other things, that the study is being conducted according to the protocol and within the guidelines established to protect the rights and safety of the subjects or patients. The trial monitor reviews study records for completeness and accuracy, and auditors inter- view the study staff to verify that everyone is qualified to perform his or her
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assigned role in the clinical study. Deviations, variance, and deficiencies are noted by a monitor and reported to the sponsor, who, in turn, is responsible for immediately correcting the issues or, alternatively, for stopping the study, until corrections are made.
Another major responsibility of a sponsor is to prepare, update, and dis- seminate a summary of clinical, nonclinical, and other pertinent product information to the PI and his or her staff. This is done with a document called the IB, written by the sponsor before the first clinical trial and updated as new information becomes available. The elements of an IB are provided Box 9.6.
An important issue that can arise with a clinical study is conflict of inter- est, real or perceived and usually financial in nature, on the part of either PI or sponsor or both. To perform an unbiased study, it is important that the PI not be beholden to the sponsor and that any investigator assigned to a study has no significant financial interest in the sponsoring biotech- nology firm. Financial interest could result in bias on the part of inves- tigational staff, and even a perception of conflict of interest or potential bias in the sponsor–investigator relationship, particularly where it involves substantial sums of money, can call into question the merit of a clinical trial. Of course, investigators are remunerated for their time, expenses, and
BOX 9.6 ELEMENTS OF AN INVESTIGATOR’S BROCHURE
• Table of contents • Summary • Introduction • Physical, chemical, and pharmaceutical properties and formu-
lation of the product • Results of nonclinical (i.e., safety, pharmacology, and toxicol-
ogy) studies • Effects (including safety, pharmacology, and pharmacokinet-
ics) in humans known from previous clinical studies • Marketing experience, if any • Summary of data and guidance for the investigator
• Anticipated risks and adverse reactions • Summary of clinical data • Assessment and treatment • Toxicity management • Additional risks associated with this or similar products
• References
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professional expertise. However, compensation must be fair and open and a clinical investigator should not have a substantial interest in corporate stock options.
The Principal Investigator and His or Her Study Staff
The principal investigator, often referred to as the PI or simply the investi- gator, is the individual responsible for conduct of a clinical study at each clinical trial site. She or he is retained by the sponsor, who delegates specific clinical responsibilities to that PI. In return, the PI receives reimbursement for expenses, including salary for the time spent in executing the study, under the agreed protocol. There may be other benefits to PIs, such as publication of sci- entific articles and ability to work at the cutting edge of their profession. A PI may be employed at an academic institution or in private practice or he or she may be associated with a CRO. Whichever the case, the agreement between sponsor and PI typically includes funding for additional study staff, such as nurses, administrators, clerical assistants, and individuals, to recruit subjects. In some cases, the PI will be asked to prepare the protocol and other clinical documents, but larger sponsoring firms frequently provide these documents and ask the PI to follow these instructional documents. A physician may, in the U.S., be both the sponsor and the investigator, or, as happens with some biopharmaceutical firms, the sponsor may directly employ an investigator.
In effect, a PI is responsible for everything that happens at the clinical trial site, including activities by his or her staff. In the United States, a PI formally accepts this responsibility in one of the two ways, under a contract with the sponsor or, in an abbreviated manner, by signing an agreement, Form 1572, with FDA. Among varied investigator responsibilities, the most important is to exercise clinical oversight and medical judgment at the site. The PI ensures that everyone on his or her investigational team conforms to the protocol and any other instructions (e.g., Operations Manual) con- cerning the study and that subjects’ rights are fully met. Principal investi- gators are responsible for submitting the protocol to the IRB for approval and then beginning the study only after receiving this approval. The inves- tigator is also responsible for enrolling and then treating patients in the proper manner; for patient compliance in taking the investigational prod- uct throughout the study; for ensuring that clinical documents, such as case report forms (CRFs), are correctly completed; and for accountability of the investigational product. Certain administrative functions are also required of the investigator; these include maintaining professional credentials, managing the research staff, communicating with the IRB, participating in study meetings or conference calls, and maintaining good relations with the sponsor and other parties involved in the study. Last but not least, the PI is the principal scientist in a clinical study. In the end, each of the PI’s responsibilities focuses on maintaining the scientific integrity of the study, safety of the patients, and integrity of the data.
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Clearly, a busy physician investigator cannot complete a clinical trial without help, and so, investigational staff is employed at each study site. Principal investigators often enlist other physicians to work on a study; these are referred to as subinvestigators. Subinvestigators are qualified to serve in this capacity by education and training, and in many cases, they are profes- sionals working closely with the PI, such as medical residents or junior staff. Although the PI may delegate certain medical responsibilities to subinves- tigators, they still accept full responsibilities with this regard. Study nurses are typically registered or licensed nurses and are employed because of their medical training and experience and because a clinical trial involves medical procedures and measurements. Study nurses originate many study records; these documents are then reviewed and co-signed by the PI. They educate the volunteers, ensure that informed consent (IC) is always properly adminis- tered, take patient histories, and consider AEs and SAEs. Patients or subjects do not just appear magically at the study site, but they must be recruited by someone adept at identifying potential volunteers and at coordinating their initial visit. This team member is the volunteer recruiter. Administrative staff manage and organize records and files and assist recruiters and nurses. Investigational product is usually maintained and distributed by a clinical study pharmacist, and for some investigational products, this individual prepares medication according to the sponsor’s instructions. Larger studies also employ data specialists, individuals who are dedicated to transferring data from paper to electronic databases and who ensure data integrity and accuracy.
Institutional Review Boards, the Process of IC, and IC Form
We, as a society, have, appropriately, given significant rights to individuals who volunteer for and participate in clinical trials. These rights derive from a very important document, the 1964 Declaration of Helsinki. The Declaration itself is based on the Nuremberg Code of 1947. The Code was drafted in response to horrific situations that occurred during the Second World War, specifically when Nazi investigators conducted biomedical experiments on prisoners without the consent of those individuals. The heart of the Code is the requirement for full understanding of the risks by written consent from any human volunteer. This means that the person who is receiving any inves- tigational treatment, no matter how minor, must have the legal capacity to give consent (or, in the case of children, have a parent or legal guardian give consent), be so situated to exercise free power of choice without coercion, and have a clear understanding of the investigation, including possible risks and benefits.
Under the 1964 Declaration of Helsinki, a guide to physicians and others involved in biomedical research involving human subjects, regulations for clinical research now state that legally effective IC (Box 9.7) must be obtained by an investigator before involving that subject in any clinical research.
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Conditions for IC have been established in most countries for usual condi- tions as well as for unusual situations, such as children, those with demen- tia, and those in emergency situations, that is, when the individual receiving the investigational product may not be capable of being fully informed. In the United States, protection of human subjects is mandated by the Code of Federal Regulations, 21 CFR Part 50, a regulation with broad application.
In practice, IC is requested by the PI from each subject immediately before enrolling that person into a clinical trial. Human subjects are asked to review a description of the clinical study, including the design, potential benefits, and possible risks. In some cases, such as novel investigational products, sub- jects are queried or quizzed by written examination, to demonstrate that they clearly understand the study and any risks to which they may be exposed during the course of the clinical trial. Subjects are always given the opportu- nity to ask questions from the PI, even if his or her staff is administering IC. Once satisfied and willing to enroll, the subject signs an IC form (ICF or CF) in the presence of a witness. However, the consent is always reversible and, should the subject change their mind, it may be negated at any time in the study. In effect, this means that a subject may leave a clinical research study at any time and for any reason or for no stated reason.
The ICF is written after the protocol has been drafted and reviewed by the PI and the sponsor and once nonclinical toxicology information or data from previous clinical studies are available. Consent forms may be written by the PI or the sponsor. Since the CF must be approved by an IRB, this board’s pre- ferred institutional format should be considered for each clinical study site. The IRBs often request changes to a CF, and so, it is not unusual to have sev- eral slightly different versions, one for each site, in a multisite clinical trial.
BOX 9.7 ELEMENTS OF IC
• Statements that the study involves an investigation and pur- poses for the research
• Description of risks or discomforts • Description of possible benefits • Disclosure of possible alternative treatments available to the
subject • Description of processes used to maintain confidentiality • Explanation of potential compensation or medical treatments • Individual to contact for answers to pertinent questions about
the research or risks and benefits • Statement that participation is voluntary and that refusal or
withdrawal will result in no penalty
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For a variety of reasons, it is sometimes necessary to obtain the approval of two or even three IRBs for some investigational sites. It is a necessary, but sometimes a challenge, for both sponsor and investigator to ensure that each form has correct content and is acceptable under current regulations.
The IRB (known as independent ethics committee in some countries) is a committee, comprising usually five to ten medical professionals, clerics or lay persons, responsible for ensuring and protecting the rights and welfare of human subjects who participate in biomedical research. The IRB reviews pro- tocols, the IC, the IB, and related materials, such as recruiting advertisements and compensation. In doing so, the committee helps to ensure the rights of subjects. The IRB must judge whether or not possible risks to the subject outweigh potential benefits or the knowledge gained through the study. The responsibilities of the IRB do not end with approval of the study and study documents, as the IRB continues to review the program as clinical research progresses and always considers reports or changes, such as SAEs and study termination. Once the study begins, the IRB must review SAEs and other significant issues that arise. Annual reviews of each study are mandatory, whether or not there are issues related to the product, the subjects, or the study itself. Of course, no member of the committee should have a conflict of interest with any study he or she reviews.
Most institutions that conduct clinical research—universities, hospitals, research centers, and CROs—have established IRBs. Independent IRBs are also available and are used by sponsors when the investigational site has no insti- tutional affiliation. Although no accreditation is required for IRBs, their records are reviewed by national regulatory agencies, and in recent years, IRBs at some notable institutions have been suspended for failure to follow regulations. In the United States, the Department of Health and Human Services is ultimately responsible for ensuring compliance with human use regulations, but this department designates agencies under its supervision, such as FDA and the National Institutes of Health, to be involved. In a practical sense, each IRB is composed of individuals from different walks of life—ethicists, clerics, scien- tists, and lay persons—so that the review is balanced in nature and considers various professional and social aspects of the proposal. The committee meets periodically; this gives each member an opportunity to review the clinical doc- uments noted earlier. After review, these documents are discussed in an IRB meeting, and it is not unusual for the committee to ask for additional informa- tion on a particular concern or recommend changes to a document. By working together, IRBs, sponsors, and PIs support each other, ensure the integrity of a clinical study, and protect the right of human subjects enrolled in that study.
Investigational Product
Clinical trial supplies or materials include the biopharmaceutical, the investi- gational product, placebo or comparator, or diluents, and any device used to apply or deliver the product or otherwise ensure correct use and safety of the
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product as it is given to the volunteer. The investigational product must meet specifications in terms of identity, purity, strength, and quality, as discussed in Chapters 6 and 7. It is very important that the investigational product be of consistent quality for all clinical trial sites and at all times throughout the duration of the study. Investigational product used for the clinical study is provided by the sponsor, that is, the biotechnology firm manufacturing the biopharmaceutical product, in a timely manner, is properly labeled, and is kept in a secure storage location, maintaining proper environmental condi- tions (e.g., temperature) throughout the study. In a blinded study, steps are taken to identify the product and placebo or comparator correctly and yet maintain the blind. For example, a label on a vial of investigational prod- uct in a blinded study may be changed by the pharmacy to a code, so that the PI or the study nurses are not aware of the treatment, active product or placebo, given to each subject. All these clinical supply operations are care- fully managed, tracked, and documented, so the disposition of all product is accounted for. As noted earlier, a pharmacist with experience in clinical trials often manages these tasks and ensures that all clinical supplies are of the highest quality, correctly labeled for the study, fully accounted for in records, and are properly stored and distributed to study staff. He or she also ensures that any unused clinical supplies are returned to the sponsor.
Collection of Clinical Data: Case Report Forms and the Patient Diary
Accurate and timely collection of all clinical data is an absolute requirement for any clinical study. The process can be divided into four major stages:
• Preparation of document formats, forms, and media to collect the data
• Collection of data during the clinical trial • Review or audit of data to ensure completeness, accuracy, and
integrity • Analysis of data
It takes considerable planning and effort to fully and properly collect clini- cal trial data. The initial or raw data are referred to as the source informa- tion or a source document, the original document on which an observation is recorded. This includes records such as laboratory reports, clinical or patient charts, memoranda, patient’s diaries, and pharmacy dispensing records. The raw data from a source document may be initially recorded on a CRF or it may be transferred from a source document to a CRF by study personnel. The CRF is a printed, optical, or electronic document designed to record all the information, no matter what the source, required by instructions pro- vided in the protocol. Case report forms, and there are many for each clinical study, are drafted after the protocol has been completed and after it is known
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what data will be collected, in what format, by whom, and how frequently. Once finalized, CRFs are printed in final format and distributed to each clin- ical trial site. A CRF is issued to each subject participating in a study.
If patients or subjects are hospitalized throughout the course of treatment, then data are easily collected in an environment conducive to keeping com- plete and accurate medical records. More often than not, the investigational product is given during brief clinic visits, and if the patient is feeling well, he or she is sent home after a few hours (or days) at the treatment site. For other products, the patient takes the product at home and only visits the clinic initially and then periodically for follow-up physical examinations or tests. When the patient is away from the clinic, the subject diary may be used to col- lect data. In a diary, each subject records any symptoms he or she has noted during the study. Although this is not a highly reliable means of collecting data, it does sometimes reveal drug-associated AEs that occur between clinic visits.
Patient diaries are reviewed and CRFs are completed by study staff, par- ticularly by study nurses and physician investigators. These documents are then subjected to final review and approval by the PI. During the course of a study, and after CRFs have been completed by the investigational staff, they are audited by an outside representative, the clinical auditor or moni- tor, a representative of the sponsor. Whether paper or electronic, CRFs are carefully reviewed against source documents to ensure accuracy of the data. Today, data on paper records are usually entered into an electronic database to facilitate statistical analysis. This is often performed using double data entry methods, in which the same source data is entered twice by two inde- pendent people and then electronically compared for consistency of data entry. Since the transfer of data from source documents to CRFs or to elec- tronic databases is prone to human error, electronic data collection, that is, directly recording information, for example, blood pressure, from a source document into an electronic database is becoming routine practice. Although this reduces errors of transcription, it requires a validated electronic system, including both hardware and software, and well-trained clinical staff.
No matter what the format, data are analyzed according to an analyti- cal or statistical plan that is prepared by a statistician before beginning the trial. Several computer programs are commonly used to analyze data and to prepare tabular and graphic presentations of the information. Statisticians, experienced in clinical trial data management and analysis, are employed by the sponsor for these tasks.
Clinical Testing Laboratories
Clinical laboratory data are important to all clinical trials. Body tissues and fluids, notably blood, are collected and sent to a laboratory, where they are tested for various parameters. For most of these tests, the laboratory is in a hospital or other medical center and is therefore certified by an accreditation
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agency such as the College of American Pathologists and regulations such as the Clinical Laboratory Improvement Amendments. However, many clinical investigations also necessitate the performance of unique laboratory tests. These may be performed in a specialty laboratory or, in some cases, in an academic laboratory or the sponsor’s laboratory. In such cases, tests must be initially qualified for accuracy and specificity, and in later stages of clinical development, they must be fully validated. Indeed, most of the quality crite- ria applied to product quality control tests (Chapter 7) are applicable to the tests used to measure clinical end points.
The sponsor is responsible for ensuring that each clinical testing laboratory meets all requirements and that laboratory testing is completely and accu- rately documented. The PI and staff ensure that samples are taken exactly as mandated by the protocol and then properly stored and shipped to the clini- cal laboratory. The PI also reviews the results, takes proper medical action, and ensures that test results reach the patient’s records as a source document.
Reporting Results of Clinical Trials: Clinical Summary Reports
Once clinical trial data have been audited, analyzed, and tabulated, these are included in a clinical summary report (CSR). This document, normally prepared by a medical writer with the help of biostatisticians, describes the clinical trial and reports all important aspects of the study. Since each report is reviewed by the sponsor and by regulatory agencies, it must be clear, com- plete, and well written. The data are tabulated and presented in an unbi- ased, yet clear and concise manner. A report relates essential elements of the protocol, clearly describing the design, treatments with investigational product, and the population of human subjects. It discusses results of the study, presenting data in tables and figures, and drawing conclusions made by the PI and statistician with concurrence of the sponsor. Safety issues are discussed in detail and statistically significant differences between treat- ment groups, with regard to safety and efficacy data, are analyzed, usually by several statistical tests. Conclusions and discussion of the data are written in a CSR, and in many cases, a manuscript describing the study, its results, and the conclusions is submitted to a scientific journal for peer review and publication.
Clinical Trial Operations
Study resources and people involved in clinical research of a biopharmaceu- tical product were reviewed in earlier sections. We now integrate this infor- mation by describing the planning and performance of clinical studies, first giving an overview of a typical clinical trial operation and then focusing,
398 Biotechnology Operations
more specifically, on each phase of development (Figure 9.1). The discus- sion on clinical trial operations focuses on tasks that are often performed in-house, by the biotechnology firm, and that are performed for the sponsor by CROs.
Activities Leading to a Clinical Trial
Early in the development life cycle of a biopharmaceutical, the clinical plan is written and it then becomes part of the overall PDP (Chapter 1). The decision of when to enter and exactly how to design the first, or Phase 1, clinical trial may not be established by the sponsor until a later date, perhaps after sev- eral preclinical and very early development milestones have been achieved. For example, before the design of the Phase 1 trial is completed, the dates and schedules for the nonclinical studies, the manufacture and control of the clinical trial product, and filing of the IND are established. Once a tentative schedule has been set and there is a high degree of confidence that investi- gational product will be available, it is possible to prepare a detailed Phase 1 clinical trial plan and the Phase 1 protocol.
Even the simplest clinical study requires quite a lot of coordination. Even if the biotechnology firms do not have a formal medical affairs department, they often have someone on staff who has experience in managing clini- cal trials, and this individual has the responsibility for clinical planning. Alternatively, a highly qualified and recommended consultant may be retained to provide early clinical guidance. As soon as Phase 1 planning process begins, the biotechnology firm decides if all elements of the study will be performed by CROs or some aspects will be kept in-house. Seldom does a biotechnology firm have the resources to hire enough professionals to directly do all aspects of clinical work themselves. Thus, early decisions in clinical planning are usually to identify what, if anything, will be done in-house, and if clinical support is to be performed by CROs, how this will be established. If all clinical work is to be contracted, the efforts should be divided and functional areas, such as trial performance and quality efforts (i.e., auditing), should go to a second contractor. This distribution of oversight ensures implementation of the checks and balances that are very important for a successful clinical study.
Now, an experienced clinician must design the study and write a concept protocol. This may be done by a consultant or in-house staff or it may wait until the investigator and investigative site have been selected. For clinical research of many biotechnology products, the sponsor can choose from doz- ens of academic sites, usually medical schools, and CROs. For other prod- ucts, for example, in the case of a cancer treatment, the sponsor might only consider sites that specialize in treating those patients. It is really important to identify a clinical site that has access to right patients, a sufficient pool of patients, experienced staff, and the infrastructure to completely perform Phase 1 clinical trial. It is not uncommon to find an excellent investigator
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who works at an unqualified site or vice versa, that is, the ideal site but with mediocre investigators. Once potential sites are selected, site visits are con- ducted by the sponsor. A site is chosen, the PI is designated and agrees to do the study, and, after negotiation over the scope of work, budget, and sched- ules, a contract is signed. This is usually followed by selection of a CRO to perform monitoring and perhaps a third group and a fourth group to pro- vide other services (e.g., central laboratory). Now, the clinical trial team has been established for that study and site.
Laboratory support is a hallmark of any clinical trial and it comes in two types: (1) standard or routine clinical laboratory support; and (2) spe- cialty laboratory analytics. Routine clinical laboratory support is offered by almost any hospital laboratory and includes analysis such as hematol- ogy, clinical chemistry, and basic immunodiagnostics. Small or early phase trials, in fact, often use hospital laboratories. However, in large clinical tri- als, a central laboratory, represented by a single contract laboratory, is used to process, in the same technical manner, samples provided by multiple clinical study sites. Most studies also require specialty diagnostics or ana- lytical techniques. For example, it is often necessary to measure the bio- pharmaceutical in samples of blood during a pharmacokinetic (PK) study. In addition, for vaccine studies, the immune response to the product must be measured with a variety of immunological assays, mostly unique and some even difficult to perform. Specialty laboratories may offer these unique testing services, but more often, these assays are adapted to or developed for clinical studies of specific products. Biotechnology firms may either do specialized assays in-house, at the firm’s internal laboratory, or they may identify a contract laboratory capable of developing the tests. There is no standard solution, and the sponsor must carefully plan exactly how it is best achieved.
Once the clinical site has been identified, the sponsor’s representative, working on behalf of the product development team, drafts the full clinical protocol. The investigator also identifies staff, for example, sub-investigators, recruiters, study nurses, and statistician, to assist in the study. Once the pro- tocol has been written, the CF and the CRFs are drafted and, along with the protocol, submitted by the investigator to the IRB. Institutional review boards typically meet once or twice each month, and it is normal for an IRB to request that changes be made in one or more documents before they are approved. Hence, the process of protocol approval can take weeks or even months to complete.
While the PI is leading these study and protocol development and review activities, the sponsor is actively recruiting a medical (safety) monitor and a clinical monitor or auditor. These individuals review the clinical trial docu- ments before submission to the IRB and before ensuring quality and compli- ance at each study site through a prestudy site visit. At the same time, the sponsor is completing manufacture, labeling, and control of the investiga- tional product and making arrangements to have it delivered to the clinical
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site. In addition, the sponsor is actively preparing and then submitting the regulatory documents, such as the IND, to FDA.
Once the regulatory agency accepts the IND and gives permission to begin the clinical study, the sponsor delivers the product to the clinical site and the investigator begins the sequential processes of screening, accepting, consenting, and enrolling subjects. The dosing phase of the clinical trial may now begin. Volunteers are given the number of doses specified in the proto- col and efficacy end points are measured per the protocol. The volunteers are closely followed throughout the study for any sign of reaction to the product. If an SAE or a series of suspicious AEs are noted by the investigative staff, then dosing and further enrollment may be halted. In such cases, the medi- cal (safety) monitor, the sponsor, and, subsequently, the regulatory authori- ties are notified. This leads to investigation and discussions; if the safety of subjects can be ensured, the study may begin once again. However, if it appears that subjects may be at undue risk or that the product is unsafe, then the study may be terminated. Fortunately, studies of most biotechnology products are not halted in early clinical studies because of safety concerns and most studies progress to completion, as specified in the protocol. Yet, patient follow-up is often a long process, and subjects may be asked to return for physical examinations or laboratory tests for months or even years after the last dose of investigational product has been given. Extensive examina- tion of subjects further ensures the safe and tolerable nature of a new prod- uct. Throughout the study, the clinical monitor visits the site to ensure that the study is being performed according to the protocol.
Once all data have been entered into CRFs, each form is screened for accu- racy and completeness and the information is transferred to an electronic database. Statisticians are normally responsible for these steps and the statis- tical data analyses that follows. In the case of biotechnology firms, the spon- sor often retains a consultant statistician to perform analyses and to prepare tables and figures that reflect the data. A CSR is then written by the investi- gator or a medical writer. This CSR is first provided to the sponsor for review and then to regulatory authorities as definitive results of the clinical study.
The above description lists only the most important tasks, and their inte- gration, involved in a typical clinical study. A host of other issues—some financial, others medical, and many administrative—must be considered in the design and execution of every clinical trial.
Phase 1 Clinical Trial: First-In-Human Study
A Phase 1 study represents the first time a biopharmaceutical is used in human. Phase 1 studies focus largely on safety and tolerability of the prod- uct but may also include measurement of efficacy end points. The num- bers of subjects enrolled in a Phase 1 study is small, usually less than 50 and often less than 25. A sponsor may elect to do several Phase 1 studies (i.e., Phase 1a, Phase 1b, etc.) in sequence, each focusing on a particular
401Clinical Trials
scientific question. This is often the case with complex and novel biotech- nology products. Phase 1 trials may be conducted on an outpatient basis or an impatient basis, or both. In the case of a Phase 1 clinical research unit, healthy volunteers may be required to remain in the onsite clinic for a few days, weeks, or, sometimes, a couple of months during an early phase dosing study, which requires a strict time-sensitive administration and sample collection schedule. In these cases, volunteers are compensated for their time and typically reasonable living accommodations are provided. An initial Phase 1 study is usually conducted with healthy individuals. Exceptions are the products that have an excellent safety profile and are intended to treat life-threatening diseases, such as a study of a gene ther- apy to treat rapid progression of a cancer or the study of an antiviral agent to treat a chronic infection such as human immunodeficiency virus infec- tion. In such cases, actual patients, having exhausted all traditional thera- pies, are enrolled into a Phase 1 study.
The design of a Phase 1 study may be open-label, meaning that both patient and investigative staff members know the nature of the treatment (i.e., inves- tigational product or placebo) when it is given or it may be blinded or double blinded, in which case a placebo (sugar pill) is given to one group of subjects, without this knowledge being disclosed. Product dose may be escalated in Phase 1 studies, but the scheme is quite conservative and only a few indi- viduals are enrolled in each dosing group. Indeed, standard dosing schemes, such as single rising dose or multiple rising doses, are selected for each new biotechnology product. These study designs are shown in Figures 9.2 and 9.3, respectively. In a single rising dose study, subjects are randomly assigned to groups, perhaps five to ten subjects per group. The lowest dose is given to subjects in the first group and the next (higher) dose is given to individuals in the second group. The process continues, until the highest dose is reached, which is determined from toxicology study results as the maximum toler- ated dose. In a multiple rising dose design, the dose is constant for any given individual but the individual returns to the clinical trial site to receive an additional dose or doses. In the interest of safety and to monitor the possibil- ity that a particular dose might result in acute reactions, only two or three subjects in a group may be dosed with the product. This is done hours or even days before the remainder of individuals assigned to this group are dosed in the same manner.
Phase 1 trial measurements focus on safety end points, but measures of efficacy are typically performed whenever possible. Subjects may be kept in a clinic for days or even weeks after the treatment, so that they can be carefully evaluated at specified intervals. For example, frequent physical examination of subjects, use of electrocardiograms to identify changes in the heartbeat, and regular clinical laboratory testing are the hallmarks of Phase 1 studies of novel biotechnology products. Criteria for ending the treatment or dosing of human subjects whenever an SAE or multiple AEs are identified, that is, stopping rules, are very important elements of Phase 1 studies.
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Clinical Pharmacology Studies of Biopharmaceuticals in Human
Additional studies are often required to fully understand a biotechnology product before it can enter Phase 3 trials. These studies are often given cre- ative and complex numbers and letters, such as Phase 1c or Phase 2a by their sponsors. Although numbered, each study is designed to specifically sup- port the overall clinical development plan and is best referred to by its pur- pose (e.g., Pharmacokinetic Study in Normal Adults).
Pharmacokinetic studies are almost always performed if a new biotechnol- ogy product is to be given repeatedly or in significant amounts. In PK studies with human subjects, the product is given in a carefully controlled manner and then blood or another body fluid is taken at regular intervals after dos- ing, and these samples are tested to determine the half-life of the product in circulation. Although PK studies may be a part of the Phase 1 or Phase 2 investigations, they are also performed as standalone studies, designed strictly for that purpose.
Pharmacokinetic studies have been best developed for drugs, but they are used extensively for studies with monoclonal antibodies and other biophar- maceuticals intended for distribution throughout the body. Pharmacokinetic and pharmacodynamic studies are further described in Chapter 8; they are typically performed in animals during nonclinical studies before they are conducted in human.
Mass balance studies are designed to determine where in the human body a new biotechnology product goes and how long it remains in each location. To perform these studies, product may be labeled with radioisotopes (hav- ing extremely short half-lives) and then given. Product metabolism, excre- tion and even localization in organs can then be followed with radiometric devices. For biotechnology products that target a particular tissue, mass balance studies may be performed in conjunction with imaging methods that allow the molecule to be identified in a particular organ. For example, it would be important to understand if a molecule aimed at cancerous cells in lung bound largely to the tumor mass and not to critical and unaffected organs, such as heart or kidney.
Food effect studies determine whether a particular type or amount of food has an effect on the uptake and effectiveness of a new biopharmaceutical. It is from food effect studies that we learn whether a patient should ingest a product on a full or an empty stomach. Although quite important for orally ingested products, such as many drugs, food effect studies may also help to explain PK observations of biopharmaceutical products, such as unexpected patterns of excretion or binding to components of serum.
Additional Phase 1 studies may focus on subpopulations, such as a racial or geographic population, the elderly, adolescents, or children. Controlled stud- ies may also be performed to determine whether a biotechnology product will have greater or lesser effect when taken with another drug. These con- comitant medication studies measure the effects of drug-drug interactions.
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Some classes of products are known to cause very unique types of reactions and these may be studied in more detail with additional pharmacology stud- ies in human subjects.
Phase 2 Clinical Trial: Proof-of-Concept Study
The second phase of clinical development includes one or more therapeutic exploratory or proof-of-concept studies, referred to as such because they are designed to provide sufficient data to suggest that a biopharmaceutical prod- uct may well have the intended effect. Phase 2 studies may be dose ranging and demonstrate the dose that is optimal to take forward into later studies. With certain other Phase 2 trial designs, the intention is to determine the minimal effective dose, or threshold effect of the biopharmaceutical. Another intention of Phase 2 study may be to determine the maximum effective toler- ated dose, at least within the dosing criteria identified in Phase 1. In addition, Phase 2 study designs may examine various end points and measurements for those end points, searching for ones that will provide the best estimate of drug safety and efficacy in subsequent studies.
Phase 2 studies are often performed at five or more clinical study centers ( multicenter study) because there is a need of more patients—50 to 500 is a typical number—with a single disease and to determine whether results vary by study site. A single center cannot often recruit these many qualified indi- viduals. Thus, Phase 2 studies are typically multiarm studies, designed with several arms or groups (cohorts) of patients, each receiving a set dose level of the product. Whenever possible, Phase 2 studies are placebo controlled and double blinded, meaning that neither the patient nor the investigator and staff knows which treatment or placebo a patient has received. Note that in cases where studies require a patient population (diseased individuals), the control group is not kept from treatment; in place of placebo, the current standard of care is provided, which offers a benchmark for the treatment arm (investiga- tional product). If the treatment involves multiple doses of a product, as is often the case, the treatment period will be much longer in Phase 2 then it was in Phase 1, so as to determine a more realistic effect of the biopharmaceutical on both safety and efficacy. Hence, Phase 2 provides several benefits beyond evaluation of product safety and efficacy. It is a means of determining whether enough patients with the condition exist, so that a larger definitive study could be conducted. Another ancillary benefit of Phase 2 studies, those performed at more than one site by several teams of investigators, is a real-world evaluation of each site. In addition, the range of subjects’ medical conditions enrolled in a Phase 2 study may be very informative for developing a Phase 3 study design.
However, results from Phase 2 studies are seldom definitive because they do not enroll enough patients to absolutely demonstrate safety, tolerability, and efficacy. Some argue that a Phase 2 clinical study is actually a mini- Phase 3, that is, a rehearsal for the pivotal study, and therefore, the results matter greatly for business development and the decision to move forward.
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Others suggest that Phase 2 is often not predictive of the outcomes in Phase 3 but is a means of determining the best dosing regimens and a valuable lesson. Whatever the case may be for a given biopharmaceutical, Phase 2 is an important step in the clinical development of any product, and therefore, each study must be carefully designed and executed, with thorough analysis and discussion of the results.
Phase 3 Clinical Trial: Therapeutic Confirmatory
After successful completion of Phase 2, the sponsor will almost certainly hold a meeting with regulatory authorities to discuss findings and to pro- pose the design of a pivotal or Phase 3 human clinical study. Discussions between sponsor and agency typically follow this meeting, and within a few weeks, both parties should agree on the design of the all-important Phase 3 or pivotal clinical trial for the biopharmaceutical. Phase 3 studies are also referred to as adequate and well controlled, as they must be just that. They are, in fact, the study based on which the product will be registered and labeling claims will be supported.
Phase 3 trials are carefully considered, with significant input from medical experts, statisticians, and those who manage and operate the study. Phase 3 studies are always large and multicenter, and today, most of these studies are multinational. Some drugs are tested in two Phase 3 trials, both using the same product and indication. The study is statistically powered, that is, it includes enough patients, so that definitive answers as to safety, tolerability, and efficacy of product can be obtained from a single study. Placebo or com- parator is typically used and double blinding and other means of preventing bias are always included in design, where possible.
The adaptive design may also be considered, with regulatory agency con- currence, for mid- and late-stage clinical trials of certain products. Adaptive design means that changes may be made in the design of a clinical study if such change is guided by examination of data, accumulated at a particular interim milestone. An adaptive design can reduce the duration of a study or decrease the total number of patients required, and because it is based on recent information and experience, it can enhance the value of data that are generated by study completion. Although the greatest interest in adap- tive design has been with adequate and well-controlled late-stage (Phase 3) studies, this approach has also worked well with ascending-dose or other mid-stage studies. However, there are caveats. Adaptive clinical study designs are prospective and must be carefully considered with regulatory authorities before initiation. As noted above, the interim analysis on which change is based is itself carefully selected and protocol revisions are previ- ously planned, and certain changes may not be acceptable under any cir- cumstances. Nonetheless, given a wide range of acceptable design changes, the adaptive design offers numerous opportunities when properly planned and applied.
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As one might imagine, no matter what the size or design, a Phase 3 study can take years to complete and generates millions of data points and huge volumes of source documents as well as IC, CRFs, and other documents. These studies are big and expensive. With studies of some rare diseases, the numbers may be low because the number of patients and geographic locations is limited. Large Phase 3 studies typically require an Operations Manual to ensure that all aspects of the study are performed exactly the same way at each clinical site. A manual also serves to resolve problems as they arise and to facilitate communication and good medical and adminis- trative practices.
An independent committee, referred to as the Data and Safety Monitoring Board (DSMB), is included in the design of most Phase 3 studies. This board of experts is unblinded to the treatment at established intervals. The DSMB may do interim statistical analyses of the data to determine if the product seems to be working and is safe. For example, the DSMB could, early in a study, discover a distressingly large number of SAEs; in this case, it may ask that the study be halted because the risks to patients outweigh the possible benefits. In other instances, the DSMB might discover early in the study that the biopharmaceutical is safe and quite efficacious, thus recommending that it should not be withheld from the subjects in the placebo or control group.
Phase 4 Clinical Study and Risk Evaluation and Mitigation Strategy
When a biopharmaceutical has been approved for marketing by regulatory authorities, it is not unusual for the agency to ask that an extended, open- label study be performed in the postmarket approval period. Such a study, sometimes referred to as therapeutic use, Phase 4, or postmarketing, is con- ducted and financed by the sponsor. Biotechnology firms usually welcome the suggestion of Phase 4 studies, because it means that their drug could be approved in an expanded patient population without the need for additional Phase 3 trials. This result is called conditional approval. The firm receives mar- keting approval and may charge patients for the product, thus generating income but with the understanding that one or more Phase 4 studies will be conducted by the sponsor and in consideration of regulatory agency guide- lines. Product safety and efficacy may be definitively demonstrated during Phase 4. If product safety and efficacy are not demonstrated during this period, the regulatory authorities have grounds to pull the market approval and, hence, the derivation of conditional approval. In addition, extended test- ing for drug-drug interactions, effects in special or high-risk populations, and additional safety surveillance are considered for Phase 4.
Phase 4 studies are intended to reduce the risk to consumers from newly marketed biopharmaceuticals. The FDA has the authority through the FDA Amendments Act of 2007 to require risk evaluation and mitigation strategy (REMS) for all newly licensed products. This is intended to ensure that benefits to patients outweigh the risks after market approval under a
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Biologics License Application or New Drug Application. Risk evaluation and mitigation strategy considers patient population, condition severity, benefit, duration of treatment, seriousness of AE, and historic safety record. Although REMS does not involve additional Phase 4 studies, it does include specific clinical guidelines that sponsors are required to generate based on the clinical use of their product. These include a communication plan to edu- cate, inform, and raise awareness of the associated risks. Some REMS require elements to ensure safe use (ETASU), which also include a medication guide for health care providers and information and instructions on safe use for prescribers, dispensers (pharmacists), and users of selected products. If an ETASU is required, it is dispensed with the product. Risk evaluation and mitigation strategy thus represents a novel approach to use clinical informa- tion to ensure safety of already licensed products by health care profession- als and the general public. Establishment of REMS is issued outside of the product label, which has helped to get and/or keep products on the market by monitoring or mitigating the known product-associated risks. A list of approved REMS is available on the FDA website. Again, the purpose and directive of the REMS program is to ensure safe use of a product, which may require follow-up testing or monitoring of the end user.
Clinical Trials for New Populations or Indications
Given the expense and complexity of performing any clinical study, it is impossible to expect a firm to test, in the initial pivotal trial, every special population that might benefit from the biopharmaceutical. Special popula- tions may include the elderly, infants, children, adolescents, pregnant or lac- tating women, and certain racial, ethnic, or geographic minorities. Yet, other populations of individuals, notably those with an underlying disease such as liver or lung disease or impaired kidney function, are also difficult to study in initial Phase 2 and Phase 3 clinical studies. Some would argue that this dis- criminates against such populations because they have no chance to benefit from the product immediately after market approval. However, it is impos- sible to study each group of individuals in the pivotal Phase 3, because of resource and time constraints.
How does a biotechnology firm go about testing individuals of any popu- lation when pursuing a new indication? The answer is to perform another Phase 3 trial in that new population or with another indication. It is often possible to begin these Phase 3 studies at Phase 2 or after having performed small Phase 1 and Phase 2 studies. Assuming that the product has market approval for at least one indication or one patient population (the first label- ing claim), these postmarketing clinical trials may help the sponsor to market a biotechnology product under a second labeling claim and thus bring benefit to patients currently without access to that product. These Phase 3 studies, focused on broadening the indication for the product by testing it in new populations, for new indications, or, for example, applying novel methods
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or routes of administration, bring certain risk to the sponsor, as a study may uncover previously unknown safety issues such as side effects of the biophar- maceutical. This can lead to undesirable regulatory action, such as addition of warnings to current labels or the need to begin more clinical trials.
Indeed, biotechnology firms are sometimes encouraged by a number of programs, sponsored by FDA or other public health agencies such as the National Institutes of Health (Chapter 4), to test a product as soon as pos- sible, typically post licensure, for as many special populations as might ben- efit from the product. Such studies are typically done post market approval, not as Phase 4 studies but as Phase 2 or small Phase 3 studies in scope and design. If they successfully demonstrate safety and efficacy in a special pop- ulation, these studies, if adequate and well designed, may be the basis for additional labeling claims for a biopharmaceutical product.
Global Clinical Trials
Many late-stage clinical trials are performed in countries distant from the sponsor’s location. Indeed, today, it is common to place multicenter clini- cal trials in numerous countries and to perform specialty studies and even Phase 1 studies in a foreign country. The sponsor often finds such efforts save significant time and money. However, there are caveats. Cultural and regulatory differences can confound even the best planned global efforts. In addition, there are the issues of different medical standards of care and genetic differences in various populations. Some, but certainly not all, global clinical trials are managed by large CROs, organizations that maintain clini- cal trial facilities in many countries, and thus understand the language, cus- toms, regulatory environment, medical practices, and population genetics in many countries where they have offices and local national employees. Foreign and multicenter global clinical trials are certainly possible and desir- able but require considerable planning and assistance.
Quality Systems for Clinical Trials: Current Good Clinical Practices
A quality system, cGCP, is applied throughout the clinical study process, from preparation of a clinical plan to completion of the clinical study report. Why must study integrity and quality be maintained at such high levels for clini- cal trials? First and foremost, it is the right of each human subject. When a volunteer enrolls in a study, he or she is subjected to a certain degree of risk or potential risk. Owing to this, and with no guarantee of benefit, the subject has the right to know that the study will, in the end, provide meaningful sci- entific results, and certainly a correct answer, regarding the safety and efficacy
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of a biopharmaceutical. Hence, maintaining high quality and integrity of the study ensures that meaningful answers are achieved and that all rights of human subjects are met. Financial responsibility, especially meeting the study budget, is another reason to complete a study properly. Studies are expensive and only few biotechnology firms have the resources to repeat a clinical trial. Indeed, a single clinical study often means the difference between success and failure of a biotechnology firm, and this alone is a compelling reason to get it right the first time in a clinical research. An outline of the important elements to achieve GCP in a Phase 1 human clinical study is provided in Box 9.8.
Under cGCP, certain systems and procedures are applied to clinical trials to ensure the well-being of subjects, data integrity, overall quality, and suc- cess of the trial. Some examples are as follows:
• Careful planning before the study and coordination during and after the study
• Selecting proven clinical sites and investigators • Training study staff to follow the protocol and other study docu-
ments and to accurately record and transfer data • Ensuring quality and integrity of data by using time-proven methods • One hundred percent internal audit of data sets and records • Clean and screen all data entries on all documents to examine quality
and consistency of data, as it is transferred to a database
These and many other practices regarding the integrity of clinical trial data are embodied in the principles and practices of cGCP. The remainder of this section will focus on four clinical trial quality practices that are very impor- tant to quality and compliance with cGCP.
Quality and cGCP in Clinical Trial Operations
Current Good Clinical Practice is further defined by ICH as “an international ethical and scientific quality standard for designing, conduction, recording, and reporting (clinical) trials that involve the participation of human sub- jects” (ICH 1996). Compliance with cGCP provides assurance that the data and reported results are credible and accurate and that the rights, integrity, and confidentiality of human subjects are protected. The extensive ICH (Chapter 4) guideline, “Good Clinical Practices,” is the international standard for quality in clinical research. It is also adopted by most countries with a developed regulatory agency, and most of these countries have supplemen- tal regulations and guidance for conduct of studies in human subjects. The cGCP in the U.S., outlined in Chapter 4, is further defined by several federal regulations, notably 21 CFR, Parts 50, 54, 56, 312, 314, 812, and 814, which col- lectively provide extensive guidance in this country. The U.S. government
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BOX 9.8 OUTLINE OF GOOD CLINICAL PRACTICE FOR A PHASE 1 HUMAN CLINICAL STUDY—EXAMPLE
1. Subject a. Well-being—should not only qualify for the study but also
be a good candidate and the study should not create undue harm if the subject decided to participate in the study
b. Rights, integrity, and confidentiality—to participate or not participate and to understand the study risks and poten- tial benefits and alternatives to participation, shared result information, compensation, consequences of withdrawing from study, any additional costs resulting from participa- tion, contact information of responsible individual in case of any questions or concerns
c. Informed consent—need time to decide; understand study purpose, duration, and procedures; ask questions; sign; and receive a copy of the completed document.
2. IRB review and/or approval 3. Clinical investigator a. Qualified to assume the responsibility of conducting clini-
cal study b. Understands the investigational product and its appropri-
ate use c. Willing to comply with GCP and applicable regulations d. Willing to participate and prepare for audits and monitoring e. Maintains a delegation of responsibilities log f. Has adequate resources to recruit subjects, time to com-
plete the study, qualified staff, and facilities to conduct the study
g. Obtains written approval from regulatory authorities (e.g., FDA and IRB)
h. Conducts the clinical research in compliance with the clinical protocol
i. Documents and explains all/any deviations j. Responsible for investigational product use, storage, and
disposal 4. Monitoring
(Continued)
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has accepted the ICH guidelines and the U.S. and ICH systems are currently harmonized.
Management responsibility, a critical hallmark of any quality system, is clearly identified in cGCP. The ICH guideline and FDA regulations identify responsibilities of the sponsor and the PI. In the United States, cGCP allows a sponsor to transfer certain responsibilities to an investigator, an institution (e.g., an university), or a commercial entity such as a CRO, but this trans- fer must be made in writing and should be clearly described. Furthermore, the guidelines state that any responsibilities not transferred in writing to an investigator, institution, or CRO are assumed by the sponsor. Thus, cGCP demands management responsibility and vendor and consultant control.
Control of the clinical trial process is clearly mandated by cGCP, and this is done in a number of ways. As described earlier, written guidance, for exam- ple, the clinical protocol or an Operations Manual, directs the clinical trial processes. In addition, standard operating procedures are commonly applied for routine tasks performed in support of a clinical trial. Procedures and data are carefully documented in source documents (e.g., medical records), CRFs, and electronic databases. These documents also contain the provision of a proper environment for both investigational product and clinical processes. For example, cGCP provides for product identification and traceability and for
BOX 9.8 (Continued) OUTLINE OF GOOD CLINICAL PRACTICE FOR A PHASE 1 HUMAN CLINICAL STUDY—EXAMPLE
5. Reporting a. Written annual reports (e.g., FDA and IRB) b. Safety reporting c. Result reporting d. Study completion or termination 6. Documentation a. Regulatory FDA/IRB/other local of federal regulatory
bodies b. Informed consent and consent process c. Clinical protocol d. Study procedures e. Product accountability log f. Source documents and case report form g. Record archival process and storage requirements h. Training records, certificates, CVs, licenses, and so on 7. Written reports
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inspection or testing, thus meeting the cGCP requirement that investigational product be clearly labeled and that the dosage form be clearly identified. The sponsor typically assumes the responsibility for delivering a quality biophar- maceutical to the clinical site and then the investigator assumes responsibility for maintaining the integrity of that material and ensuring that each patient receives the correct product (e.g., placebo or active product).
Current Good Clinical Practices apply to virtually every operational aspect of a study. As noted earlier, responsibilities are clearly defined in writing. A case in point is control of a nonconforming study. Clinical prac- tices recognize that, despite the best of intentions and controls, there is a high probability that mistakes are made over the course of the study. These situations are referred to as noncomplaince with the protocol, the standard operating procedures, cGCP, and/or applicable regulatory requirements. Current Good Clinical Practices guidelines establish the need for self- reporting, auditing, and careful review by several parties of all documents. Most importantly, noncompliance must be reported and then corrective and preventive action must be taken. With regards to preventive actions, GCPs allow for changes in processes, as described in study documents, but they also demand that change be controlled, reviewed, and approved by respon- sible individuals.
The performance of clinical trials requires that all study staff have the appropriate education and experience and are properly trained. Earlier, the sponsor’s responsibility to select only qualified investigators and institutions to perform clinical studies was mentioned. However, the quality require- ment does not stop there. The sponsor is ultimately responsible for ensuring proper education, experience, and training of individuals in the clinical labo- ratory, in statistical group, and at any CRO. All professional staff must fully understand the protocol and IB and know their respective professional roles and responsibilities under the protocol.
Customer concerns and complaints, another hallmark of quality, focus on the satisfaction of both human subjects and, in the case of contract studies, the sponsor. As noted in cGCP, “the rights, safety and well-being of trial sub- jects are the most important consideration and must prevail over interests of science or the study staff, and society” (ICH, 1996). Everyone involved in a clinical trial must, at all times, consider the rights and well-being of each subject. Such consideration does not end with signing the CF but contin- ues to the end of the study. Indeed, for some studies of novel biotechnology products, responsibility for well-being of a subject extends through the life- time of that person.
Integrity of Clinical Study Data and Documents
Data collection and control are important under cGCP. The data presented in a CSR accurately reflect the information that was recorded in source docu- ments during the study. The objective is the completeness and accuracy, that
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is, all data points should be collected for every patient enrolled. Diligence is taken by the investigator, sponsor, and others in handling, analyzing, and reporting data.
Although 100% complete/0% errors is the goal, a number of seemingly unavoidable problems can occur and data points may be corrupted, ques- tionable, or missing from even the best designed and managed clinical study. For example, patients may fail to meet appointments or they may drop out of the study all together. This can be tolerated to some degree, but if too many patients leave the study or fail to comply with follow-up visits, it may not be adequate and well controlled and the overall study results are open to question. In addition, the investigative staff will inevitably make errors when entering data into source documents or transferring information from source documents to CRFs or electronic databases. A high error rate can invalidate a study. Serious violations of a protocol may occur if subjects are enrolled in a study without meeting eligibility criteria or completing the IC process or if there is insufficient documentation of the consent process. In such cases, the data set may be considered incomplete. One patient missing a single dose of product will not invalidate a complete data set, but when several patients miss a dose or if a few patients miss several doses, the integrity of that study will, at the very least, be flawed. These are but a few examples of why cGCP stresses the importance of excellent study management and performance.
Monitoring and Auditing Clinical Trials
Clinical monitoring, not to be confused with the medical (safety) monitor, is the process of overseeing all aspects of a clinical trial and is a responsibility of the sponsor. Monitoring begins when the first subject is enrolled and ends when the last subject is discharged from the study. Monitoring ensures that the study is performed in accordance with cGCP, the study documents, nota- bly the protocol, and other regulatory requirements. It is a big job to monitor even a small clinical trial, and it is a tremendous effort to properly monitor a Phase 3 study. However, monitoring is essential for every study.
Auditing, also described in Chapter 5, is a systematic examination of study processes and documents. An important part of monitoring, it carries with it a function of determining whether particular activities are being per- formed correctly. Auditing normally involves the careful review of clinical trial documents to ensure that they are correctly completed according to the instructions in the protocol. Auditors, also referred to as clinical research associates, are the individuals who perform the audits. The task of auditing a clinical trial is very detail oriented and analytical. Auditors visit clinical trial sites, where they review documents, speak with the investigational staff, and identify issues or problems. In many cases, they assist in resolving those issues by speaking with the PI and sometimes performing staff training. Thus, auditors perform important roles in the overall monitoring process and ensure the integrity of a study and compliance with cGCP.
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Ethical Behavior and the Well-Being of Clinical Trial Subjects
The Declaration of Helsinki, as noted earlier, is the foundation for protec- tion of human volunteers in any study. It holds clinical research to an excep- tionally high ethical standard, stating that, “Compliance provides public assurance that the rights, well-being, and confidentiality of trial subjects are protected and that the clinical trial data is credible” (ICH, 1996). This is totally appropriate, and cGCP directly supports each principle in the Declaration. Over the years, a number of human rights issues have arisen in clinical studies, even as the clinical research community applied cGCP to thousands of clinical research studies worldwide. The vast majority of human research studies are, however, without breaches of ethical behavior, suggesting that the clinical research community and cGCPs are doing an outstanding job of maintaining the principles laid down in the Declaration. Unfortunately, more ethical issues may arise in the future. Without further delving into ethical behavior in clinical studies, some examples of common ethical situations faced by biotechnology firms are worth mentioning as a close to this chapter and, perhaps, as word of caution to those entering the field of biopharmaceutical development.
Some years ago, as clinical research expanded to support development of biotechnology products, clinical investigators in private practice or associated with nonprofit institutions accepted stock options in return for providing clinical investigative services for the biotechnology firm. This seemed like a useful model in the beginning, as biotechnology firms essen- tially deferred compensation, thus saving themselves considerable upfront expenses. However, it was also felt that this practice was a potential conflict of interest and regulatory agencies argued that at the least it should be fully disclosed by both investigator and sponsor. Others asked how this differed from the accepted practice of employees (of the sponsoring biotechnology firm, including those staff members responsible for clinical monitoring) accepting stock options from their employer. Although the issue has not been fully resolved to everyone’s satisfaction, the consensus is that clinical investigators must not hold significant interest in the sponsoring entity and that any interest must be disclosed to regulatory authorities. Today, most regulatory agencies demand full disclosure by outside investigators and the indirect financial remuneration (e.g., stock options) of outside or independent investigators is capped in some instances. This situation demonstrates how important it is to ensure high ethical standards when dealing with human subjects and the clinical study process.
In another example, regulatory authorities are authorized to blacklist employees of the biopharmaceutical industries, preventing them from work- ing in our industry if there is evidence that they egregiously or repeatedly failed to comply with cGCP or other regulations. Although the practice of individual sanctioning is applied by FDA to all areas of biotechnology devel- opment, it is not uncommonly used to prevent certain clinical investigators,
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those who repeatedly failed to adhere to cGCP or those who commit a major infraction, from further participating in studies. Note that even though a blacklisted investigator need not be convicted of a crime, his or her name and affiliation still become a matter of public record. Although the clinical research community may feel singled out by the practice of blacklisting, it demonstrates how seriously we as a society take the rights of human subjects and the sanctity of clinical trials as a means of ensuring a safe and effective supply of biopharmaceuticals.
Another issue with ethical aspects is the problem of distinguishing clinical research from medical treatment. The distinction between the two is often blurred, and this challenges clinical researchers, worldwide, as they seek the best treatment for patients. For example, a new biopharmaceutical product to treat AIDS is taken to market after abbreviated clinical studies. On the one hand, this is good, because it provides access to a seemingly promis- ing treatment. On the other hand, it might also put users at potential risk of using a product that has not been thoroughly tested for safety or efficacy and this risk might have been mitigated if more extensive research had been conducted.
Another example is the off-label use of a biopharmaceutical. Sometimes, this occurs when it is quietly encouraged by biotechnology firms that wish to increase sales. It also happens when well-meaning, but sometimes poorly informed, physicians treat disease in an effort to save a patient from pain, suffering, or even death. An important question is: in such cases, are the patients being enrolled and treated in a clinical research study but without full IC? If IC is the first principle of the Nuremberg Code, the Declaration of Helsinki, and cGCP, then how can we justify such off-label use by individual medical practitioners? Alternatively, does consent alone mean that a patient is being treated ethically?
There is no simple answer to any of these examples. However, these types of questions arise repeatedly in biotechnology and apply to many of today’s most exciting biotechnology advances. There is no clear answer for every type of clinical situation. Nonetheless, those in the biotechnology industry face ethical issues, as they make difficult decisions on how to proceed into clinical studies.
Summary on Clinical Trials
Clinical trials evaluate the safety and efficacy of a biopharmaceutical product by testing that product in human subjects or volunteers. A clinical research program—and every biopharmaceutical intended for use in humans must have one—is based on the indication and patient population for the product and requires careful planning and significant resources, both human and
417Clinical Trials
monetary. Importantly, clinical studies are observational or, more frequently, experimental, and in either case, they are designed to test a hypothesis and follow the scientific method. Clinical research of a biopharmaceutical is per- formed in a series of three or four phases. Phase 1 is first time in human, focusing on safety, that is, clinical toxicology and possible adverse reac- tions, but yielding pharmacology and perhaps some efficacy data. Phase 2 is designed to expand the dosing regimen, extend findings from the first phase, and to establish a foundation for a pivotal or definitive study, Phase 3 is the third phase and it confirms the safety profile and demonstrates therapeutic or preventative efficacy. Postmarketing studies are considered Phase 4. Good Clinical Practices are followed throughout clinical development; indeed, reg- ulatory authorities will not approve data generated by a clinical trial unless data are scientifically sound and in compliance with GCPs. Hence, clinical development planning is a necessity from the outset, and counterintuitively since clinical studies are in late stages of development, the clinical plan is often the first step in an overall PDP. Clinical trials involve a number of indi- viduals, each with unique skills and management to integrate their efforts, and a large number of clinical trial documents, notably the protocol, must be prepared. A PI is the key figure in a trial, but of course, there are clinical support staff, the sponsor, and human subjects. The clinical protocol is an instructive document, exactly and fully describing why and how a clinical study is to be performed using these participants. Investigational product, or a placebo, is provided by the sponsor and then given by the investigator to each subject. Ethical behavior on the part of each participant is critical, and the well-being of human subjects is the primary objective of any clini- cal trial. Volunteers are enrolled in a study only after they give IC to par- ticipate, and this process is reviewed and monitored by an IRB. Subjects are followed throughout the trial, and AEs and detailed data about the condition of each subject are recorded and, eventually, reported to the sponsor and to regulatory agencies in clinical trial reports. The clinical program, which is applied to the overall development program, is a large operation in and of itself and must be fully managed and integrated into the overall develop- ment program.
Reference
International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH). 1996. Guidance for Industry E6 Good Clinical Practice: Consolidated Guidance.
419
Additional Readings
Chapter 1: Introduction to Biotechnology Operations: Planning for Success
Billups NF. 2015. American Drug Index. Wolters Kluwer, St. Louis. Dorland WAN. 2011. Dorland’s Illustrated Medical Dictionary. 32nd ed. Elsevier, New
York. Food and Drug Administration (FDA). 2016. Approved Drug Products with Therapeutic
Equivalence Evaluation (The Orange Book). US Food and Drug Administration, Rockville, MD; http://www.accessdata.fda.gov/scripts/cder/ob/ (accessed May 31, 2016).
Walsh G. 2003. Biopharmaceuticals: Biochemistry and Biotechnology. John Wiley and Sons, Chichester, UK.
FDA. 2016. National Drug Code Directory (NDC). US Food and Drug Administration, Rockville, MD; http://www.fda.gov/Drugs/InformationOnDrugs/ucm142438. htm (accessed May 31, 2016).
Ho RJY. 2013. Biotechnology and Biopharmaceuticals: Transforming Proteins and Genes with Drugs. 2nd ed. Wiley-Blackwell, NY.
Jameel F, Hershenson S, Khan M, and Martin-Moe S. 2015. Quality by Design for Biopharmaceutical Drug Product Development. Springer, NY.
Krinsky DL et al. (eds). 2015. Handbook of Nonprescription Drugs: An Interactive Approach to Self-Care 18th ed. American Pharmacists Association, Washington DC.
The Merck Index. 2013. The Merck Index. 15th ed. Merck & Co., Whitehouse Station, NJ.
Mosby. 2015. Mosby’s Drug Reference for Health Professionals. 5th ed. Elsevier Mosby, St. Louis, MO.
Sobti RC. 2008. Essentials of Biotechnology. CRC Press, Boca Raton, FL. Thompson Healthcare. 2010. PDR Drug Interactions and Side Effects. Thompson
Healthcare, Florence, KY. Thompson Healthcare. 2015. PDR (Physician’s Desk Reference) for Nonprescription
Drugs, Dietary Supplements and Herbs. Definitions Guide to Over the Counter (OTC) Medicines. Thompson Healthcare, Florence, KY.
Thompson Healthcare. 2016. Physician’s Desk Reference (PDR). Thompson Healthcare, Florence, KY.
Thompson Healthcare. 2016. Physician’s Desk Reference (PDR) Generics. Thompson Healthcare, Florence, MO.
420 Additional Readings
Chapter 2: Project Management
Babler SD and Ekins S. 2010. Pharmaceutical and Biomedical Project Management in a Changing Global Environment. Wiley, NY.
Brown L and Grundy T. 2011. Project Management for the Pharmaceutical Industry. Gower Publishers, Aldershot, UK.
Harpum P. 2010. Portfolio, Program and Project Management in the Pharmaceutical and Biotechnology Industries. Wiley, NY.
Project Management Institute. 2013. A Guide to the Project Management Body of Knowledge (PMBoK Guide) 20. Project Management Institute, Newtown Square, PA.
Wingate LM. 2014. Project Management for Research and Development. Auerbach Publications, Boca Raton, FL.
Chapters 3 and 4: Regulatory Affairs and Regulatory Compliance
Adams DT. et al. 2008. Food and Drug Law & Regulation. FDLI, Washington, DC. Danzis SD and Flannery EJ (eds). 2010. In vitro Diagnostics: The Complete Regulatory
Guide. FDLI, Washington, DC. Humi RA. 2012. Pharmaceutical Competitive Intelligence for the Regulatory Affairs
Professional. Springer-Werlag, NY. Kahan JS. 2008. Medical Device Development: Regulation and Law. Parexel International
Corporation, Waltham, MA. Klincewicz SL et al. 2009. Global Pharmacovigilance Laws and Regulations: The Essential
Reference. FDLI, Washington, DC. Mathieu M. 2004. Biologics Development: A Regulatory Overview. 3rd ed. Parexel
International Corporation, Waltham, MA. Pines WL. 2003. How to Work with the FDA. FDLI, Washington, DC. Pisano DJ and Mantus D (eds). 2008. FDA Regulatory Affairs: A Guide for Prescription
Drugs, Medical Devices, and Biologics. CRC Press. Boca Raton, FL. RAPS. 2008. Fundamentals of EU Regulatory Affairs. 4th ed. Regulatory Affairs
Professionals Society, Rockville, MD. RAPS. 2009. Fundamentals of US Regulatory Affairs. 6th ed. Regulatory Affairs
Professionals Society (RAPS), Rockville, MD. RAPS. 2010. Fundamentals of International Regulatory Affairs. Regulatory Affairs
Professionals Society, Rockville, MD. RAPS. 2010. US Regulatory Acronyms & Definitions Pocket Guide. 4th ed. Regulatory
Affairs Professionals Society, Rockville, MD. FDA. 2016. What does FDA inspect? US Food and Drug Administration; http://www.
fda.gov/aboutfda/transparency/basics/ucm194888.htm (accessed May 31, 2016).
421Additional Readings
Chapter 5: Quality Assurance
Avis KE et al. (eds). 1998. Biotechnology: Quality Assurance and Validation. CRC Press, Boca Raton, FL.
Haider SI and Asif ES. 2012. Quality Operations Procedures for Pharmaceuticals, API and Technology. CRC Press, Boca Raton, FL.
Jameel F, Hershenson S, Khan M, and Martin-Moe S. 2015. Quality by Design for Biopharmaceutical Drug Product Development. Springer, NY.
Ogg G. 2005. A Practical Guide to Quality Management in Clinical Trial Research. CRC Press, Boca Raton, FL.
Sandle T. 2015. Pharmaceutical Microbiology: Essentials for Quality Assurance and Quality Control. Woodhead Publishing, Cambridge, UK.
World Health Organization. 2007. Quality Assurance of Pharmaceuticals: A Compendium. WHO, Geneva.
Chapter 6: Biomanufacturing
Avis KE. et al. 1996. Biotechnology and Biopharmaceutical Manufacturing, Processing and Preservation. CRC Press, Boca Raton, FL.
Berry IR (ed). 2003. Pharmaceutical Process Validation. Marcel Dekker, NY. Doble M. 2004. Biotransformations and Bioprocess. CRC Press, Boca Raton, FL. FDA. 2009. Guidance for Industry: Assay Development for Immunogenicity Testing of
Therapeutic Proteins. US Food and Drug Administration; http://www.fda.gov/ downloads/drugs/guidancecomplianceregulatoryinformation/guidances/ ucm192750.pdf (accessed May 31, 2016).
FDA. 2014. Guidance for Industry: Immunogenicity Assessment for Therapeutic Protein Products. US Food and Drug Administration; http://www.fda.gov/down- loads/drugs/guidancecomplianceregulatoryinformation/guidances/ ucm338856.pdf (accessed May 31, 2016).
FDA. 2015. Guidance for Industry: Cell-Based Products for Animal Use. US Food and Drug Administration; http://www.fda.gov/downloads/animalveterinary/guidance- complianceenforcement/guidanceforindustry/ucm405679.pdf (accessed May 31, 2016).
Fink DW Jr. 2009. FDA regulation of stem cell-based products. Science, 324(5935): 1662–3.
Haider SI. 2009. Biotechnology. A Comprehensive Training Guide for the Biotechnology Industry. CRC Press, Boca Raton, FL.
Haider SI. 2010. Cleaning Validation Manual. A Comprehensive Guide for the Pharmaceutical and Biotechnology Industries. CRC Press, Boca Raton, FL.
Jamel F and Hershenson S. 2010. Formulation and Process Development. Strategies for Manufacturing Biopharmaceuticals. Wiley, NY.
Ozturk S. 2005. Cell Culture Technology for Pharmaceutical and Cell-Based Therapies. CRC Press, Boca Raton, FL.
422 Additional Readings
Sofer G. 2005. Process Validation in Manufacturing of Biopharmaceuticals. CRC Press, Boca Raton, FL.
Zhong J-J. 2004. Biomanufacturing: Advances in Biochemical Engineering/Biotechnology. Springer-Verlag, Berlin, Germany.
Chapter 7: Quality Control
Chakraborty C and Jhingan R. 2015. Protein Based Drugs: A Techno-Commercial Approach. Hindawi Publishing Corporation, London, UK.
Haider SI and Syed EA. 2011. Quality Control Training Manual: Comprehensive Training Guide for API, Finished Pharmaceuticals and Biotechnology. CRC Press, Boca Raton, FL.
Rodriguez-Diaz R, Wehr T, and Tuck S. 2005. Analytical Techniques for Biopharmaceutical Development. Taylor & Francis Group/CRC Press, Boca Raton, FL.
Sandle T. 2015. Pharmaceutical Microbiology: Essentials for Quality Assurance and Quality Control. Woodhead Publishing Co., Cambridge, UK.
Chapter 8: Nonclinical Studies
Cavagnaro JA. 2008. Preclinical Safety Evaluation of Biopharmaceuticals. John Wiley & Sons, Hoboken, NJ.
Choy WN. 2007. Genetic Toxicology and Cancer Risk Assessment. Marcel Dekker, Inc., NY. Descotes J. 2004. Principles and Methods of Immunotoxicology. Elsevier, NY. FDA. 2006. Guidance For Industry: Considerations for Developmental Toxicity Studies
for Preventive and Therapeutic Vaccines for Infectious Disease Indications. US Food and Drug Administration; http://www.fda.gov/downloads/ Biolog icsBloodVacci nes/Gu ida nceCompl ia nceReg u lator yI n for mat ion/ Guidances/Vaccines/ucm092170.pdf (accessed May 31, 2016).
FDA. 2015. Guidance For Industry: Product Development Under the Animal Rule. US Food and Drug Administration; http://www.fda.gov/downloads/Drugs/ GuidanceComplianceRegulatoryInformation/Guidances/UCM399217.pdf (accessed May 31, 2016).
Horvanth CJ. 2009. TeGenero incident and the duff report conclusions: A series of unfortunate events or an avoidable event? Toxicologic Pathology, 37: 372–383.
Kapp RW. 2013. Reproductive Toxicity. Taylor & Francis Group, NY. Meibohm B. 2007. Pharmacokinetics and Pharmacodynamics of Biotechnology Drugs.
Principles and Case Studies on Drug Development. Wiley-VCH, NY. Timbrell JA. 2007. Introduction to Toxicology. 3rd ed. Taylor & Francis Group/CRC
Press, Boac Raton, FL. Weinberg S. 2003. Good Laboratory Practice Regulations. 4th ed. 2003. Marcel Dekker,
NY. Zhang L. 2016. Nonclinical Statistics for Pharmaceutical and Biotechnology Industries
2015. Springer International Publishing AG, Cham, Switzerland.
423Additional Readings
Chapter 9: Clinical Trials
Chin R, Yoonsik C, and Lee BY. 2008. Principles and Practices of Clinical Trials Medicine. Elsevier, NY.
Cook TD and DeMets DL. 2007. Introduction to Statistical Methods for Clinical Trials. Chapman Hall & CRC Press, Boca Raton, FL.
Hackshaw AK. 2009. A Concise Guide to Clinical Trials. Wiley-Blackwell, Oxford. Piantadosi S. 2005. Clinical Trials: A Methodologic Perspective. John Wiley & Sons,
Chicester, UK. Rozovsky FA and Adams RK. 2003. Clinical Trials & Human Research: A Practical Guide
to Regulatory Compliance. John Wiley & Sons, NY. Stone J. 2006. Conducting Clinical Research: A Practical Guide for Physicians, Nurses,
Study Coordinators and Investigators. Mountainside MD Press, Cumberland, MD.
425
Glossary
21 CFR: part 21 of the U.S. Code of Federal Regulations, the part in which most food and drug laws are located (FDA, 2016).
483: See Form 483. 510(k) premarket notification process: a regulatory route by which to seek
marketing approval from FDA for a medical device of low to moder- ate risk and substantial equivalence to another device.
abbreviated new drug application (ANDA): an application submitted to FDA for approval of marketing a generic drug.
absorption, distribution, metabolism, and excretion (ADME): measure- ments of a biopharmaceutical in pharmacokinetic studies.
absorption phase: the pharmacokinetic phase during which a biopharma- ceutical is absorbed into the body and, presumably, into the blood.
accuracy: the measure of an assay to agree with a known true value. act: legislation that begins as a bill before congress, and once passed by
congress, becomes law. active (pharmaceutical) ingredient (API): the part of a product that has the
desired biological activity, providing the primary therapeutic and biological effects.
acute toxicity: an animal safety study that examines the toxicity of a bio- pharmaceutical after a single dose with short-term follow-up.
adaptive (study design): a study design that allows changes to be made in the protocol at a milestone, if data warrants.
ADE: See adverse (drug) event. ADME: See absorption, distribution, metabolism, and excretion. adequate and well-controlled study: a scientific study that is carefully
designed to test a hypothesis and has proper controls for the intended purpose.
adulterated: a biopharmaceutical or drug that is putrid, filthy, or decom- posed; lacks strength, purity, or quality; is not of cGMP nature; or is contained in a deficient container.
adverse (drug) event (AE or ADE): a medical event in a human subject that is undesirable and symptomatic of a physiological change or disease and is due to a particular intervention or treatment, such as use of a biopharmaceutical.
AE: See adverse (drug) event. American type culture collection (ATCC): a nonprofit organization that col-
lects, stores, and distributes biological reference samples. analyte: material or product that is being tested. analytical method: laboratory procedure or test performed on a product to
measure an attribute.
426 Glossary
analytical tool: laboratory procedure or test performed on a product to mea- sure an attribute.
analyze: in project management, it means an assessment of achievement relative to the project plan. It may include evaluation of alternatives and resource requirements and usage.
ANDA: See abbreviated new drug application. APHIS: animal and plant health inspection service of the U.S. Department
of Agriculture. API: See active pharmaceutical ingredient. ascending dose study: an experimental design in which the dose of investi-
gational product is raised with each subsequent group of volunteers. aseptic: used as a noun, it means without living organisms. As a verb or
adverb, aseptic describes processes that avoid to a great degree the inclusion of or contact with microbes.
assay: laboratory procedure or test performed on a product to measure an attribute.
ATCC: See American type culture collection. attribute: a positive, desirable, or even necessary characteristic of a product
that lends itself to testing. audit: a formal review of a process, study, or product by an auditor; it exam-
ines whether actual performance was conducted in accordance with established instructions.
batch: an amount of product that is produced together as a single entity. Batch usually refers to a defined amount of biopharmaceutical bulk substance.
batch production record (BPR): a document used in manufacturing to both guide a process and record critical information regarding perfor- mance on a particular batch or lot of product.
BDS: See bulk (drug) substance. bias: a predisposition or prejudice in scientific studies or a systemic distor-
tion of a statistical result (Oxford English Dictionary, 1997). BIO: biotechnology industry organization. bioavailability: the fraction of biopharmaceutical, of the total amount given,
available in the blood (or tissue) and its intended effect. bioequivalence: assessment of the comparative activity and bioavailability
of two products after administration to animals or humans. biologic or biological: historical terms used to describe the products that are
derived from or represent biological or living sources. biologics license application (BLA): an application made to FDA for the
purpose of gaining marketing approval for a new biological (non- therapeutic and nonpharmaceutical) in the U.S. This large document provides complete information on development of the product and its safety and efficacy.
biomanufacture: manufacture or production of biological molecules, cells, tissues, or other products derived from biotechnology.
427Glossary
biopharmaceutical: a biological molecule, cell, tissue, or other material of biological origin used in the treatment or prevention of disease in humans. Biopharmaceuticals are complex in a molecular sense and most have a biological origin.
bioreactor: a closed vessel designed to support the multiplication and growth of eukaryotic cells for the purpose of expanding a cell line or producing a biopharmaceutical.
Biotechnology Regulatory Services (BRS): a division of APHIS, the US Department of Agriculture, that regulates certain genetically engi- neered organisms.
BIS: Bureau of Industry and Security, the US Department of Commerce. BLA: See biologics license application. blinded study: a clinical study design in which certain individuals, usu-
ally the volunteers and investigative staff, are unaware of the treat- ment (investigational product, placebo, or comparator) given to the volunteer.
BPR: See batch production record. BRS: See biotechnology regulatory services. BS: See bulk substance. bulk (drug) substance (BS or BDS): biopharmaceutical product that has
been produced and purified but has not yet been formulated or ali- quoted into the final container. Also referred to as bulk drug sub- stance or BDS.
campaign (manufacture): manufacture of more than one product in a facil- ity. Each manufacturing area is used only for one product at any given time, so projects are sequential.
cap: a stopper or other seal that is placed as a seal on the container once it has been filled.
CAPA: corrective and preventive action; it is the process of investigating and correcting a deficiency, deviation, or other problem or issue in the manufacturing or quality control of product.
carcinogenicity: the ability or tendency to invoke cancer. case report form (CRF): paper or electronic form on which the investigator
enters medical information gathered during a clinical trial. CBER: Center for Biologics Evaluation and Research, the U.S. Food and Drug
Administration. CBP: Customs and Border Protection of the U.S. Department of Homeland
Security. CDC: See Center for Disease Control and Prevention. CDER: Center for Drug Evaluation and Research, the U.S. Food and Drug
Administration. CDRH: Center for Devices and Radiological Health, the U.S. Food and Drug
Administration. cell: production cell that replicates and has particular traits. In biomanufac-
ture, this is often of bacterial, yeast, insect, or mammalian origin but
428 Glossary
may be derived from almost any species: plant or animal, eukaryotic or prokaryotic.
cell bank: a source of live cells, derived from a clone or small number of progenitor cells, that are kept in storage and then used as the seed or source in biomanufacture.
Center for Disease Control and Prevention (CDC): a U.S. federal public health agency under the Department of Health and Human Services.
Certificate of Analysis (CoA): a formal document used to identify attributes or traits, quality control tests, specifications, and test results of a product or raw material.
CF: See informed consent form (ICF). CFR: See code of Federal Regulations. cGCP: current Good Clinical Practices are the regulations promulgated by
FDA and international bodies and must be followed for conduct of research in human subjects.
cGLP: current Good Laboratory Practices are the regulations promulgated by FDA and international bodies and must be followed for nonclini- cal safety studies of all biopharmaceuticals.
cGMP: current Good Manufacturing Practices are the regulations promul- gated by FDA and international bodies and must be followed for the production (manufacture) and distribution of all biopharmaceuticals.
change control: an active process under which proposed changes are intro- duced, examined, and acted upon according to plan and with full knowledge of everyone involved or impacted by the change.
charter (team): a mandate and authorization to achieve, as a team, an objec- tive. A charter is bestowed by a higher authority, such as a stake- holder or executive manager.
chemistry, manufacturing, and controls (CMC): the information related to the production, testing, and distribution of a pharmaceutical prod- uct is contained in the CMC (or Pharmaceutical Quality) section of an IND application to FDA.
CHMP: committee for human medical products of the EMEA. Prepares opinions on questions concerning medications for human use.
chronic toxicity: a safety study in animals that measures the toxicity of a biopharmaceutical given in multiple doses with follow-up over a long period of time (>6 months).
classified: a formal designation regarding the level of air quality in a clean area or room.
clean room (area): an area or room in a biomanufacturing facility that is controlled to reduce the change of microbial or particulate contami- nation of product.
clinical (study) design: a brief description of a clinical study that includes the scientific approach and hypothesis, as well as ensuring quality elements.
clinical research: See clinical trial.
429Glossary
clinical research associate (CRA): also called a clinical monitor and is part of a medical research team associated with human clinical trials.
clinical research unit (CRU): a human clinical research medical facility staffed with CRAs, nurses, and physicians, all focused on conduct- ing human clinical research.
clinical study: See clinical trial. clinical summary report (CSR): a written report that fully summarizes the
performance and results of a clinical trial, including tabulated data and statistical analyses.
clinical trial: a designed scientific study in which a principal investigator evaluates an investigational biopharmaceutical product in human volunteers.
Cmax: the maximum amount of biopharmaceutical that is available in blood or tissue after delivery of a given dose.
CMC: See chemistry, manufacturing, and controls. CMO: See contract manufacturing organization. CoA: See certificate of analysis. code of Federal Regulations (CFR): compilation of all current US federal
regulations. cohort: group of human volunteers with common characteristics and treated
at the same time, although not necessarily in the same manner. combination product: a product that combines two or three of the following:
biological, drug, and medical device. commercial production: biomanufacture of a product at the final or commer-
cial scale. Typically happens just before or after marketing approval by a regulatory agency.
common technical document (CTD): a format for preparing, organizing, and writing market applications and investigational new drug applications in many countries. The eCTD is the electronic version of the CTD.
comparative clinical study: clinical research in which the investigational product is compared with another product.
comparator: a control material, typically a product that is licensed for the same indication, used in a clinical trial to compare against the use of investigational product.
compendium/compendial: a reference book that provides product, process, and test standards and specifications.
components: materials that are used in manufacture. Often include hard- ware materials.
concentration-effect relationship: the relationship between the concentra- tion of a biopharmaceutical in the blood (or tissue) and the desired physiological effect it has on an animal or human.
concept protocol: a brief design of a nonclinical or clinical study used as the basis for discussions between sponsor, investigator, and regulatory authorities. It is the foundation for preparing a full study protocol.
430 Glossary
conditional approval: regulatory approval for a biopharmaceutical, in which the regulatory agency stipulates that certain tasks, often Phase 4 clinical studies or follow-up of patients from Phase 3 studies, must be performed as a condition to that approval.
conformance: this means that a product or, in a broader sense, a study report or other document meets specifications and regulations.
consent form (CF): format used to inform a human subject and document the process of informed consent. See informed consent.
construct (genetic): a biological material that is or has been derived from genetic engineering of a molecule or a cell. It usually refers to a plasmid.
container: the vial or other vessel that directly holds a final product. contaminant: particle or chemical that is undesirable and has entered the
product stream during manufacture. contract manufacturing organization (CMO): a manufacturing facility that
performs biomanufacturing on a contract basis. contract research organization (CRO): a corporation or institute that pro-
vides contractual support to a biopharmaceutical sponsor in areas of clinical or nonclinical studies or manufacture and control.
control: in quality control, a material that is used to ensure performance of an assay. It may be a positive or negative control.
control: in project management, it means to use influence to follow the cur- rent plan or improve it.
control article: the nonactive material that is given to experimental animals as a control and in lieu of active ingredient during a nonclinical study.
controlled clinical research: a clinical trial in which both an investigational product and one or more control substances, such as a placebo or a comparator, are given to patients randomized into groups.
CPMP: committee for proprietary medicinal products of the EMEA. CRA: See clinical research associate. CRF: See case report form. crimp: process of sealing or closing a cap onto a container of final product.
This is often done with metal bands or covers. critical pathway: the pathway in a project that is critical to achieving objec-
tives and schedules. It is also called the rate-limiting pathway, as it determines the rate at which the product is going forward.
CRO: See contract research organization. CSR: See clinical study report. CTD or eCTD: See common technical document. CVM: Center for Veterinary Medicine, the US Food and Drug Administration. cycle or life cycle (project): the overall project, from beginning to end, with
all elements included. Data and Safety Monitoring Board (DSMB): committee of independent
experts to evaluate the data of an ongoing clinical trial.
431Glossary
decision points: precise or particular moments in a project schedule that require consensus on a particular management or technical matter.
Declaration of Helsinki: a series of ethical principles used to govern the rights and well-being of human subjects in human clinical research.
design control: a formal and documented system of plans and procedures that are used to ensure the quality development of products or processes.
deviation: it is a situation when a value or process does not meet established procedures, rules, or specifications. Deviations are discovered dur- ing or after the fact and were not planned.
device: See medical device. diary: the patient diary is a record kept by all clinical study volunteers to
record any medical conditions they might encounter after the treat- ment and while not under direct medical supervision.
distribution phase: the pharmacokinetic phase during which a biopharma- ceutical is distributed throughout the body, normally from blood to tissues and organs.
DLT: See dose-limiting toxicity. documentation: a formal process of a quality system in which all documents
for a product, process, or service are carefully and fully managed from beginning to end.
dose: single delivery or application of a biopharmaceutical product. dose-limiting toxicity: it is the toxicity associated with unwanted side
effects that are serious enough to prevent an increase in dose or treatment level.
DOT: U.S. Department of Transportation. downstream: the stage of manufacturing in which product, in a crude state,
is purified to bulk substance. drug: a small molecule with pharmacological effects, usually of organic ori-
gin, and with a well-characterized chemical structure. DSMB: See Data and Safety Monitoring Board. early phase development: stage of biopharmaceutical development begin-
ning with the initiation of development efforts through the end of Phase 1 clinical trials.
EEC: European Economic Community. Efficacy: It refers to producing the desired effect, so that the therapeutic indi-
cation of the product is achieved. EIR: See Establishment Inspection Report. EMA: See European Medicines Agency. end point: a measurable entity, such as weight or blood pressure, in a scien-
tific study. enroll: to allow a volunteer to participate in a clinical study after that person
has completed the screening and informed consent processes and has been found acceptable to participate.
EPA: U.S. Environmental Protection Agency.
432 Glossary
establishment inspection report: a document prepared by FDA inspectors to note the findings made during an inspection.
European Medicines Agency: European agency responsible for evaluation of medicines for use in the EEC.
excipient: a material that is added to a biopharmaceutical product and is not an active ingredient; for example, a carrier or a preservative.
exclusion criterion: a medical characteristic of a potential volunteer that requires the investigator to disallow that individual from enroll- ment in a clinical study.
excretion phase: the pharmacokinetic phase during which a biopharmaceu- tical is excreted from the body.
experimental clinical study: a clinical trial prospectively designed as an experiment with active treatments, controls, and other methods of making comparisons between treatment groups.
expression system: a biological construct that consists of a recombinant gene stably inserted into a living cell.
FAO: Food and Agricultural Organization of the World Health Organization. fast-track approval: an FDA review and approval process that is expedited
to treat serious or life-threatening disease with a current unmet need.
FDA: See Food and Drug Administration. FD&C Act: U.S. Food, Drug, and Cosmetics Act of 1937. FDP: See final product Federal Trade Commission (FTC): a US government agency regulating
commercial practices, including advertising, within the U.S., with the exception of foods, drugs, and biopharmaceuticals.
feedback: a team member relating any aspect of the project to other team members.
fermentation: process of growing bacterial or yeast cells in a closed ves- sel under defined conditions for the purpose of manufacturing a product.
FIFRA: Federal Insecticide, Fungicide, and Rodenticide Act. fill: to actively place or aliquot a biopharmaceutical into a container. final drug product (FDP): See final product. final product (FP): it is the biopharmaceutical product once it has been for-
mulated, filled into a container, and finished. Also referred to as final drug product or FDP.
finish: to place a cap onto a container, crimp or otherwise seal the container, and, in some cases, label and package the product.
FOI or FOIA: See Freedom of Information (Act). Food and Drug Administration (FDA): an administrative U.S. gov-
ernment agency under the Department of Health and Human Services (DHHS). It is responsible for regulating many products and their development, including food, drug, medical devices, and biopharmaceuticals.
433Glossary
Form 483: FDA Inspectional Report, a form given to sponsors immediately post inspection by an inspector, listing deficiencies or deviations identified during the inspection.
formulation: addition of various solutions, buffers, excipients, or stabilizing materials to a bulk product, so as to make a solution or powder that is ready for fill into a container.
FP: See final product. Freedom of Information (Act) (FOI or FOIA): a law that allows private citi-
zens to petition a government agency such as FDA to release infor- mation that is not proprietary or confidential.
FTC: See Federal Trade Commission. functional area: a particular scientific, management, or technical activity
and suborganization aimed at fulfilling an established purpose. As regards biotechnology operations, seven functional areas are com- monly listed: clinical, manufacture, nonclinical, project manage- ment, quality assurance, quality control, and regulatory affairs.
FWS: fish and wildlife service of the US Department of Interior. Gantt chart: a computer-generated rendering of a project, using narrative
and horizontal bars to show tasks, milestones, and their dependen- cies and relationships.
genetically modified organism (GMO): an organism that has been changed or modified (e.g., additions and deletions to the genetic makeup) by genetic engineering, often referred to as recombinant DNA technology.
GMO: See genetically modified organism. GMP: See current Good Manufacturing Practices. good tissue practices (GTP): a regulatory guideline from FDA for the pro-
cessing of tissues or cells for human use. guideline: a public document written and promulgated by a government
agency, such as FDA, that recommends or suggests practices, both administrative and technical, that would, if practiced, fulfill require- ments given under regulations. However, guidelines do not have the legal status of regulations.
hallmark of quality: one of the several operational quality criteria that com- prise a quality system.
HEPA: high-efficiency particle air is a special filter that removes all but the smallest particles, leaving the exiting air especially clean and >99% free of bacteria and fungi.
hold (clinical and regulatory): a step taken either by FDA or by a sponsor to stop or not begin a clinical study of an investigational product.
hold (biomanufacture): a step in biomanufacturing where product is kept in a container, awaiting further processing.
host cell: a live cell that contains a biological molecule or microbe (not nor- mally found in that cell).
HVAC: heating, ventilation, and air conditioning.
434 Glossary
IACUC: See Institutional Animal Care and Use Committee. IATA: International Air Transport Association. IB: See investigator’s brochure. IBC: See Institutional Biosafety Committee. IC: See informed consent. ICF: See informed consent form. ICH: See International Conference on Harmonization. IDE: See investigational device exemption. identity: individuality of a product and features that distinguish it from all
other products. In other words, it is the ability of a product to be of a known and unique nature.
IEC: See Independent Ethics Committee. IFPMA: International Federation of Pharmaceutical Manufacturers and
Associations. impurity: undesirable material, usually macromolecular and submicro-
scopic, but may be visible, microscopic, or soluble organic or inor- ganic, in a product or stream. It is often inherent to the process itself, such as cell debris.
inclusion criterion: a medical characteristic of a potential volunteer that is considered a positive trait for the enrollment of that individual into a clinical study.
IND: See investigational new drug application. Independent Ethics Committee (IEC): it serves the same function as an
Institutional Review Board, to review, approve, and monitor research involving human subjects.
indication: a remedy, treatment, or prevention that is suggested by the symptoms of the disease. For a biopharmaceutical, an indication is a specific medical condition that may be treated or prevented by the product.
induced pluripotent stem cells (iPSC): a pluripotent cell or cell line that has been created from an adult cell. The adult cell has been reverted back to a more stem-cell-like state using chemicals or recombinant DNA technologies.
induction: biomanufacturing step in which a chemical is added to a fermen- ter to induce or elicit the production of a product by an organism that has been genetically engineered to respond to the chemical.
informed consent (IC): the process of informing a volunteer to a clinical trial exactly on the nature of the trial and all possible risks and ben- efits that the person might derive. To enroll, the volunteer must sign the informed consent form.
informed consent form (ICF or CF): this clinical trials document explains to a volunteer the potential risks and benefits of a clinical study. To enroll in a study, a volunteer must understand and sign the CF.
in-process testing: testing that occurs on samples taken from the process stream during the manufacturing process.
435Glossary
input: a component of design review during which the needs of the user are considered and incorporated into the product or process design.
installation qualification (IQ): a step in validation of a biomanufacturing facility, utility, or equipment in which the installation is demon- strated to be according to specification.
Institutional Animal Care and Use Committee (IACUC): this institutional committee reviews animal use and experimental protocols to ensure ethical and proper use of laboratory animals.
Institutional Biosafety Committee (IBC): a committee of scientists, ethi- cists, and laypersons established by an institution (e.g., a university) to review the engineering, use, or transfer of genetically modified organisms and related research and development.
Institutional Review Board (IRB): an institutionally based peer review com- mittee that reviews all clinical research and protocols at the institu- tion to ensure the proper treatment and well-being of volunteers.
International Conference on Harmonization (ICH): a nonprofit group, supported by regulatory agencies and medical products industries, dedicated to developing and disseminating medical product devel- opment guidelines and pathways that are acceptable to regulatory authorities in most countries and to ensuring the quality of those products.
International Federation of Pharmaceutical and Manufacturers Association (IFPMA): a trade organization that promotes harmoni- zation of regulations at the international level.
International Standards Organization (ISO): an international organization dedicated to quality through establishing requirements and speci- fications for products, services, and processes. It is not a regulatory agency but develops guidelines and provides ISO certification after review and approval. A cornerstone guideline is ISO 9001.
investigational device exemption (IDE): an application submitted to FDA to allow the human clinical study of a new device.
investigational new drug application (IND): formal application to the US Food and Drug Administration to test in human volunteers a biophar- maceutical that does not have marketing approval (investigational).
investigational product (or drug): a biopharmaceutical product that is tested in human volunteers in clinical trials and under an IND and has not received market approval.
investigator: individual leading the scientific and medical portion of a clini- cal study. The principal investigator has the responsibility for the study, whereas subinvestigators assume certain responsibilities under the principal investigator.
investigator’s brochure (IB): an informative document that identifies for each member of the investigative staff the information on the clini- cal study, the product being tested in the study, and the possible risks and benefits to volunteers enrolled in the study.
436 Glossary
in vitro diagnostic (IVD): a laboratory test used to diagnose disease and that is regulated by FDA as a medical device.
iPSC: See induced pluripotent stem cells. IQ: See installation qualification. IRB: See institutional review board. ISO: See International Standards Organization. IVD: See in vitro diagnostic. label: the printed identification for a product, usually on paper and held to
the final container or package of biopharmaceutical. labeling: the sum total of printed materials, package insert, package print-
ing, and so on that accompany, or are adherent to, biopharmaceuti- cal containers and packaging. Labeling, approved by FDA, provides the approved indication, directions for use, dosage, and other critical information provided by the sponsor to the user.
late-stage development: the final investigational stage of product develop- ment and the activities associated with this stage and with Phase 3 clinical trials.
LD50: See lethal dose, 50%. leachates: chemicals that are dissolved from a surface or other solid matrix
into the product stream during biomanufacture, thus becoming contaminants.
lethal dose, 50% (LD50): the amount of an agent that causes death in 50% or half of the population of animals over a defined study period.
letter of authorization (LOA): a formal letter submitted to FDA to allow an independent investigator to reference confidential information cur- rently on file at FDA. Also referred to as a letter of cross-reference.
limit of detection (LOD): the minimal amount of analyte that can be accu- rately detected by a particular assay in a test substrate.
limit of quantitation (LOQ): the minimal amount of analyte that can be reasonably measured, in a quantitative sense, by a particular assay.
linearity: for an analytical, quantitative measurement (test), it is the ability, within a given range of analyte in a sample, to obtain test results that are directly proportional to the concentration of the analyte.
LOA: See letter of authorization. LOD: See limit of detection. LOQ: See limit of quantitation. lot: defined amount of manufactured final product that constitutes a legally
defined entity. A lot has unique character, quality, and source and is a specifically identified amount, labeled as such.
lyophilize: a process in which a biological material in a solution is sub- jected to freezing and drying simultaneously to preserve the cells or molecules.
marketing application: application to a regulatory agency to market a prod- uct in that country. See also Biologics License Application or New Drug Application.
437Glossary
marketed product: a biopharmaceutical that has been approved for sale in that country.
market approval: permission from the Food and Drug Administration to market a biopharmaceutical in the U.S. for the indication and at the dosage given in the approved labeling.
master cell bank (MCB): it is the ultimate source of any seed. A bank of cells, usually derived from a single clone or source, is kept as a unique resource for later expansion or use.
master file (MF): a regulatory document under which a sponsor may file confidential information with FDA. Investigational use of a prod- uct is not allowed under a Master File, as it is under an IND. Drug Master Files (DMF) or Biologic Master Files (BMF) are used in bio- pharmaceutical development.
matrix: the medium in which a product is disbursed or suspended to include those of natural origin, for example, serum, or of synthetic origin, for example, phosphate-buffered saline.
maximal efficacy: the greatest effect or response that is given by a biophar- maceutical (and in the absence of toxicity).
maximum tolerated dose (MTD): this dose is the highest tested dose that does not result in an unacceptable toxicity or adverse effect.
MCB: See master cell bank. MDR: See medical device reporting. measurement/measure: the act of or a system for determining, through
laboratory, clinical, or nonclinical testing and evaluation, the quantity or quality of a particular end point or process. It refers to the use of an assay or instrument or scientific and technical skills to determine an unknown parameter. It may be qualita- tive or quantitative and is usually determined in a stated unit or capacity.
medical device (device): an object that may be any one of the many classes of physical or engineered products. It achieves its intended primary action in a manner other than pharmacological, biological, or meta- bolic means. Medical devices are used to diagnose, prevent, monitor, and treat disease.
medical device reporting (MDR): an FDA regulation aimed at ensuring that manufacturers report defects in, or adverse events associated with, medical devices.
medical monitor: also referred to as the medical safety Monitor, a medical professional assigned to review AE or SAE or other matters relating to safety of volunteers.
method (analytical): a test or analytical procedure that is used in a labora- tory to measure quality.
metrics: in project management, it refers to measurement of progress against established milestones, schedules, budgets, or other resources.
MF: See master file.
438 Glossary
middle-stage development: product development that occurs before, dur- ing, and immediately after Phase 2 clinical trials.
milestone: it is a readily identified interim event or set of events and a major waypoint in a project that demonstrates the achievement of a planned outcome.
misbranding: it refers to labeling or branding falsely or in a misleading manner or without supporting scientific data and in violation of FDA regulations. It means not completely or legibly labeled or not accurately reflecting the truth.
monitor: in a clinical trial, the medial monitor is either a safety monitor, that is, a medical professional assigned to review AE or SAE or other matters relating to safety, or a volunteer. The term monitor is also used to indicate a sponsor’s monitor, an individual that reviews activities and progress of a clinical study at the study site.
MTD: See maximum tolerated dose. multiarm clinical study: a clinical study design that includes several treat-
ment groups, each group receiving a different treatment or dose. multicenter clinical study: a trial that is performed in more than one medi-
cal center, but under the same protocol and for the same purpose. Multiple centers are used because it is not possible to recruit all vol- unteers at only one center.
NAI: See no action indicated. national drug code (NDC): a unique product identifier for human drugs in
the U.S. National Regulatory Authority (NRA): a regulatory body, such as FDA,
appointed by a national government in the area of food and drug regulation.
National Science Advisory Board for Biosecurity (NSABB): consists of a federal committee that addresses the issues related to biosecurity and biological research.
NCIE: National Center for Import and Export (of animals), APHIS, the US Department Agriculture.
NDA: See new drug application. NDC: See national drug code. neat: test article used in full strength or undiluted in a nonclinical study. new drug application (NDA): an application made to FDA for the purpose
of gaining marketing approval for a new drug (pharmaceutical or biological) in the U.S. An NDA also applies to certain therapeutic biopharmaceuticals receiving review at CDER. This large document provides complete information on development of the product and its safety and efficacy.
NF: National Formulary of USP. NIH: National Institutes of Health of the US Department of Health and
Human Services. NIOSH: National Institute for Occupational Safety and Health, CDC.
439Glossary
no action indicated (NAI): it refers to findings from an inspection report or EIR in which FDA states that no findings in an inspection warrant further investigation or action.
NOAEL: See no observed adverse effect level. NOEL: See no observable effect level. nonclinical: studies, both in vitro and in vivo, that are performed outside of
man to define in the laboratory and in animals the pharmacology or toxicity of a biopharmaceutical.
nonconformance: a product or, in a broader sense, a study report or other document that fails to meet specifications after quality control test- ing or quality review.
no observed adverse effect level: the highest dose tested in an animal spe- cies that does not produce a statistically or biologically significant increase in adverse effects in comparison to a control group.
no observable effect level: the highest dose tested in an animal species with no detected effects.
NRA: See National Regulatory Authority. NRC: Nuclear Regulatory Commission, the US Department of Energy. NSABB: See National Science Advisory Board for Biosecurity. Nuremburg code: it is a series of ethics principles established for conduct-
ing human research to protect the rights and well-being of research subjects.
OAI: See official action indicated. OBA: Office of Biotechnology Activities, Office of the Director, NIH. OBP: Office of Biotechnology Products, CDER, FDA. OBRR: Office Blood Research and Review, CBER, FDA. observational clinical study: a study that observes patients for the dis-
tribution and incidence of disease, in the absence of specific treat- ments and interventions. It is also referred to as epidemiological study.
OCTGT: Office of Cellular, Tissue, and Gene Therapies, CBER, FDA. off-track: this means that a project is not on schedule or budget. official action indicated (OAI): it refers to findings on an EIR, the inspec-
tional report, in which FDA recommends that action be taken immediately by a manufacturer and followed-up by FDA regarding deficiencies or deviations noted during an inspection. It is the most serious of the three EIR finding categories.
on-track: this means that a project is on schedule or budget. open-label study: a type of clinical study in which the investigator and, in
some cases, the patient are aware of the treatment regimen (placebo or investigational product). It is not a blinded study.
operational area: as regards biotechnology operations, an operational area is one of the seven commonly listed (clinical, manufacture, nonclinical, project management, quality assurance, quality control, and regula- tory affairs) or other developmental specialty.
440 Glossary
operational management: it refers to managing technology development under a product development plan in an operational area.
operational qualification (OQ): a stage of manufacturing facility validation in which the operation of a utility or piece of equipment is shown to meet specifications. It is performed before process qualification.
OQ: See Operational Qualification. OSHA: Occupational Health and Safety Administration, the US Department
of Labor. OTC: over-the-counter drugs. outcome: it refers to broad results or visible effects that form the basis for a
study hypothesis. In clinical studies, these are often medical items. output: a component of design control in which the process, service, or prod-
uct design, based on input and review by professionals, is proposed and documented as the process and/or the product, in whole or in part.
OVRR: Office of Vaccine Research and Review, CBER, FDA. package: the inner (surrounds a primary product container) or outer (sur-
rounds multiple inner packages) material that is used to protect a product from damage. Often, cardboard or plastic packaging mate- rial is used for packaging.
package insert: the printed, extended instructions and information, approved by both the manufacturer and a regulatory agency, for a product and enclosed in the package. The package insert is an important part of labeling and states indication, directions for use, warnings, and so on. (See labeling.)
PAI: Preapproval Inspection by FDA. parenteral: a route of delivery given beneath or through the epidermal layer
and is not oral, mucosal, or topical. parenteral product: a product that is given beneath the skin or injected. particle: it is a microscopic or visible piece of material, usually an undesir-
able contaminant in a product or stream. pathway: in project management, it refers to a well-defined course of action
or sequence of events. patients: individuals with a pre-existing medical condition enrolled in a
clinical trial for the purpose of testing a therapeutic product for that condition.
PCB: production cell bank. (See working cell bank.) PCR: See polymerase chain reaction. PD: See pharmacodynamics. PDP: See product development plan. PDS: product development strategy. (See product development plan.) PERT: it refers to the program evaluation and review technique, which is
actually an illustration of a project schedule to depict interrelation- ships of various tasks and milestones in a project.
441Glossary
PhRMA: Pharmaceutical Research and Manufacturers of America. pharmaceutical: a small molecule drug. (See drug.) pharmacodynamics (PD): the study of how a biopharmaceutical interacts
with various tissues, fluids, or organs to achieve a therapeutic effect. pharmacokinetics (PK): the study of how, when, and where a biopharmaceu-
tical gains access (e.g., absorption), is distributed, is metabolized, or is excreted by the body.
pharmacology: the study of pharmacological agents (drugs or biopharmaceu- ticals) and their mechanisms of action and effects on organisms.
pharmacopeia: a reference book that provides product, process, and test standards and specifications.
phase 1: the first clinical phase and the early phase of product development. phase 2: the second clinical phase and the mid phase of product development. phase 3: the third clinical phase and the last phase of product development
before market approval. phase 4: any development activities that occur after market approval of a
product. phased manufacture: production of product over time by using phases of
development, going from simple systems to more complex, from pro- ducing small to large batches, from small to larger clinical studies, and so on. Phases numbered as 1, 2, 3, or 4 or as early, mid-, and late devel- opment phases.
PI: See principal investigator. pilot production: earliest production of a new product in the biomanufac-
turing cycle. It is usually done on a small scale and in an experimen- tal mode.
pivotal: a clinical study, usually in Phase 3, designed to demonstrate or confirm beyond reasonable doubt the safety and efficacy of a biopharmaceutical.
PK: See pharmacokinetics. Placebo: a sugar pill, containing any substance that is known to be safe and
not cause a direct physiological or therapeutic effect and is given to volunteers assigned to a controlled clinical trial.
PMA: See premarket approval. polymerase chain reaction: a powerful molecular biology technique used to
amplify DNA or RNA. portfolio: a collection of projects or programs with common technologies or
goals. potency: measurement, direct or indirect and generally quantitative, of a
product’s biological or therapeutic effect. It is established as a qual- ity control test to evaluate BS or FD.
PQ: See process qualification. precision: the ability of an assay to repeatedly produce the same or very
similar result on repeated testing when variables are held constant.
442 Glossary
preclinical: research and early development activities that occur before Phase 1 clinical studies. It is most often used in reference to research activities.
premarket approval (PMA): the regulatory process, and application docu- ment, for marketing approval for a medical device in the higher-risk classes.
preventive action: an activity that prevents a problem or issue from occur- ring or recurring.
principal investigator (PI): the medical professional, usually a physician, who is the responsible individual at a clinical study site for a par- ticular clinical trial.
process: as a verb, a process refers to actively produce a product by using defined technical skills. As a noun, a process refers to a defined por- tion of biomanufacture.
process qualification (PQ): a stage of manufacturing validation in which a process or part of a larger process is qualified by actual performance against specifications.
product: a thing, substance, or material that is manufactured or produced during biotechnology operations. It is a result of planning and labor.
product development plan (PDP) or strategy (PDS): a document developed early in the life cycle of a product’s development cycle that provides a roadmap and specifications needed to conduct rational, compliant, and resource-effective biopharmaceutical development from early to late phases.
product life cycle: the manufacturing cycle involved in product develop- ment, beginning with planning and continuing through all phase of development and stages of manufacture.
product stream: See stream. production: the act of biomanufacturing. production cell bank (PCB): a working cell bank. program: a group of related projects that are often coordinated and share a
common objective. project: a distinct and planned enterprise that is identified by a unique
objective or goal, a beginning, an end, and a schedule. project champion: individual serving as a stakeholder or on the project
team who has a strong personal and professional interest in achiev- ing the objective or product. Champions argue and strongly sup- port the objective.
project management: the function of planning, organizing, and managing resources and schedules to bear on performance of a defined project.
project management plan: a written plan that outlines how a project will be managed using modern project management processes and tools, both technical and social.
443Glossary
project manager: a professional appointed to manage and lead a project team and the processes, tasks, budget, schedule, and other activities that fall within the scope of the project.
project schedule: a calendar with dates that demonstrates a project from beginning to completion, with all major tasks and processes. It con- tains start, milestone, and finish dates.
protocol: an instructive document that identifies exactly why and how a study (e.g., clinical, nonclinical, or validation) will be performed and provides schedules of events.
purification: it is a process used in biomanufacturing to remove contami- nants and impurities while retaining the desired product.
purity: it is the amount of product in relation to impurities that might exist in a product. It also refers to freedom from chemical or biological contamination or impurities.
QAU: See quality assurance unit. QbD: See quality by design. QSR: See quality systems regulation. qualification: it means to ensure that something, such as a test or a process,
is suitable for use. Qualification is typically less stringent than vali- dation, and the process is often applied to critical laboratory tests or manufacturing processes in early to mid-phase development.
quality: the degree of excellence of a thing; general excellence (Oxford English Dictionary, 1997).
quality agreement: a contract between two parties, generally a contractor and a client, that identifies quality aspects of a relationship, espe- cially regarding compliance issues and conformance with a quality system.
quality assurance: it is a function of planning, managing, operating, and ensuring the performance of a quality system.
quality assurance unit (QAU): the functional area at a biotechnology firm responsible for the overall quality operation and all aspects of qual- ity assurance.
quality by design (QbD): a planning process in which a product and its biomanufacture, are carefully described along with attributes and specifications.
quality control: laboratory, test, or metrology function to ensure quality of a product.
quality manual: a written document that proscribes the quality policies and, in general, the criteria, operations, and organization for quality sys- tems at a biotechnology firm.
quality plan: a written document that expands on the quality manual, and describes the quality systems and plans to operate those systems at a biotechnology firm. The quality plan is referenced in or is part of each product development plan.
444 Glossary
quality system: a designated set of components, connected in a logical fash- ion and focused on quality of a product or service. Quality systems for biopharmaceutical development are codified in regulations, guiding quality operations of a particular activity, such as manufac- ture (e.g., cGMP), nonclinical studies (e.g., cGLP), or clinical studies (e.g., cGCP).
quality systems regulations (QSR): an FDA guideline, based on a regu- lation and focused on quality design and manufacture of medical devices.
RAC: See Recombinant Advisory Committee. randomize: the act of randomly placing a subject or an animal into one or
another treatment group in a controlled and blinded clinical or non- clinical study.
range: the value defined by the upper and lower limits of a specification or concentration of an analyte.
raw materials: items that are used to manufacture a product. The term gen- erally refers to materials such as solutions, reagents, chemicals, and biological substances.
Recombinant Advisory Committee (RAC): a committee established by the director, NIH, to review and approve certain studies, in vitro or in animals or man. It deals with genetic engineering or transfer or use of genetically modified organisms.
recruiter: an individual employed by a clinical investigator or a sponsor for the purpose of identifying individuals who might wish to volunteer for a clinical trial.
reference standard: a well-characterized and known product or material against which the attribute of a test material may be compared.
regulation: a rule that has the force of law. It is an interpretation of the law by an administrative government agency in the executive branch and is intended to carry out the intent of the law.
regulatory intelligence: the process of finding and analyzing publically available regulatory information.
release: the action of allowing a product or study to be provided to the user, customer, or client.
release testing: the panel of tests that are performed on a product before it can be released.
REMS: See risk evaluation and mitigation strategy. requirement: an attribute that a product possesses, usually defined in sci-
entific terms. research seed: a microbial seed (e.g., clone) or a cell line derived from a
research laboratory. risk evaluation and mitigation strategy (REMS): risk management plan-
ning and a plan to ensure that benefits of a product outweigh the risk. It is performed by evaluation after marketing approval.
445Glossary
robust: a vigorous, strong, and sturdy manufacturing process or test method in biopharmaceutical development. It is the overall reproducibility of a process or test results when operational conditions are held within established ranges.
root cause: the ultimate or original cause for an issue or a problem that occurs. It can be clearly described in technical or scientific language and hence is preventable in the future.
run: a single and clearly identifiable manufacturing process. A single run produces an individual lot or batch or product.
SAE: See serious adverse event. scale-up: increase in the amount of biomanufacturing for a product, so as to
increase the total amount of product in a single batch or lot of BS or FP.
seed: a defined cell or viral particle from which other cells or particles may be derived.
serious adverse event (SAE): an adverse event that is serious by medical diagnosis or is life-threatening or causes death of the volunteer.
Six Sigma: a quality program aimed at reducing product or service failure rates.
sociotechnical skills: practiced ability to integrate sociological or people management expertise together with a technical knowledge and capability, so as to lead a project team.
SOP: See standard operating procedure. source document: any record, data, or other piece of information that is clos-
est to the source. These data are initial, original, or raw and on a written or electronic document.
sparge: it means to move a gas into or through a liquid in a vessel such as a fermenter.
specification: a stated value or range of values that are specific to a product attribute and quality control test. Specifications are specific, strict, and fully defined criteria based on which a product is found either suitable or unsuitable for use.
specificity: the degree to which a measurement made by an analytical test is due to the actual analyte of interest and not due to other materials in the test matrix.
sponsor: the entity (institute, individual, or corporation) that is ultimately responsible, in a scientific, business, and legal sense, to regulatory agencies, the public, and the users for a product and its development.
stability: for a product, it means the trait of maintaining purity, potency, and strength over time and in a given environment. It also refers to the property of a biopharmaceutical or product to not degrade or break down.
stability-indicating: a test that is capable of identifying when a product has lost or is losing purity, potency, or strength.
446 Glossary
stability protocol: a document that designs and plans a stability testing pro- gram for a specific product and applies various assays under an estab- lished schedule.
stage: a major division of a biomanufacturing scheme, such as upstream processing.
stakeholder: an individual who, by status or dominant position, is influ- ential to a project team and, although not always serving directly on the team, has a vested interest in the success of the team’s efforts. Project teams serve, in part, to meet the expectations of a stakeholder.
standard operating procedure (SOP): an instructive document that pro- vides exact or detailed technical or administrative procedures. An SOP may include forms to capture data during the performance of that procedure.
steady state: pharmacokinetic phase during which the concentration of a biopharmaceutical is maintained at a given level.
step: a small but important part of any stage of biomanufacture. sterile fill: a process to fill final containers with product and in the absence
of microbial contamination. Aseptic methods are used throughout. stop criterion: a medical situation that arises and, by definition, leads to ces-
sation in the enrollment and treatment of volunteers in a clinical trial.
stopping rules: a prospectively defined plan, in a clinical study protocol, to stop current treatment or additional enrolment, monitor potential safety concerns, and evaluate potential risk before putting addi- tional subjects at risk.
stream: it is the product stream, or main bulk of product, in process, during biomanufacture. It is the material that is under production at a single time in biomanufacture.
strength: measure of active ingredient in a product. It is typically measured using an analytical method that does not measure biological activity but instead measures the amount of chemical or biological substance present.
study director: an individual responsible for overall design, performance, and reporting of a nonclinical safety study.
subacute toxicity: a safety study that evaluates toxicity of a biopharmaceu- tical, given in multiple doses, in animals over a brief period (e.g., 30–60 days).
subchronic toxicity: a safety study that evaluates toxicity of a biophar- maceutical, given in multiple doses, in animals over a moderate period (e.g., 3–6 months).
subject: an individual without an underlying disease who volunteers for a clinical trial.
synchronize (project): integrate and bring together the various parts under a schedule and series of events.
447Glossary
system suitability: ability of an analytical test to achieve the objectives of the assay. All components of the test are suitable for the intended purpose.
T1/2: the time elapsed from when a biopharmaceutical reaches Cmax until it reaches ½ the value of Cmax.
tags: molecular identifiers genetically engineered into a molecule. They may be used for identification or affinity purification of that molecule.
tangential flow filtration (TFF): type of biomanufacturing preparative fil- tration that allows filtration through a selective membrane as flow of liquid sweeps the membrane surface to prevent clogging.
targeted product profile (TPP): a written document that prospectively identifies the attributes and intended therapeutic indications for a biopharmaceutical product. It is written in the format of product labeling but is a planning tool and not a means of reporting results.
task: a piece of work included in a project that is exactly defined in technical terms and has a beginning and end.
team (project): a group of professionals from different functional areas work- ing together toward a common objective, each of them bringing a specific expertise.
team dynamics: the sociological and psychological energies and motions that affect the behavior and change for a project team.
team leader: an influential individual who serves on the project team but typically not as a project manager. A team leader may be a chief sci- entist or key executive or the founder or discoverer of the technology.
test: analytical method or laboratory procedure performed on a product to measure an attribute.
test article: test product, or the biopharmaceutical in formulation, as given to animals in a nonclinical study.
TFF: See tangential flow filtration. timeline: a visualization of tasks or processes and milestones set against a
schedule. Tmax: the elapsed time from when a biopharmaceutical is given until the
maximum concentration is seen in blood (or tissue). tolerability: to determine in a clinical trial how well the subjects or patients
medically or physiologically accept or tolerate the investigational product when it is given in measured dose.
tolerated: ability of an organism to be subjected to a biopharmaceutical over a period of time without experiencing adverse effects or harm due to that product.
total quality management (TQM): a management approach to quality that aims to continuous satisfaction of the customer or client.
toxicology: the study of toxic effects of chemicals, a biological, or an ionizing radiation on a living organism.
Toxic Substances Control Act (TSCA): a US law that deals with testing before use or release to the environment of chemical substances.
448 Glossary
TPP: See targeted product profile. TQM: See total quality management. track: as a noun, it is the pathway of a project. The verb, to track, means to
monitor the processes and tasks within a project to ensure that the processes and tasks are completed on schedule and budget.
tracking: the project management process of reviewing all aspects of the project to ensure that tasks are completed on schedule and budget.
trait: a distinguishing feature or characteristic of a product. It is a specific chemical or biological feature that can be measured with an analyti- cal test.
transfection: placing a foreign or recombinant gene into a mammalian or other cell derived from animals or plants.
transgenic: an organism, such as a plant or animal, that retains one or more genes of another organism.
treatment group: in a scientific study design, it refers to a group of humans or animals that receives the same treatment, such as either investiga- tional product or placebo.
TSCA: See Toxic Substances Control Act. U.S. Pharmacopeia (USP): a compendium or reference volume that pro-
vides information on biopharmaceutical raw materials, products, processes, tests, and formulations.
upstream manufacture: biopharmaceutical production that yields the prod- uct as a crude or unpurified material. The early stages of biomanu- facturing from cell bank to crude cell paste.
USDA: U.S. Department of Agriculture. USP: See U.S. Pharmacopeia. VAI: See voluntary action indicated. validate: to provide strong evidence, usually through experimentation, that
a piece of equipment, facility, utility, test, or process performs exactly as intended and within established specifications.
validation: a process in which a test or process is demonstrated to perform exactly as intended and planned and meets established specifications.
variance: a measurement, outcome, or part of a process or study that does not meet established procedures, rules, or specifications but is known or planned before it occurs in fact.
vector: a live organism or construct of DNA (e.g., plasmid) that contains DNA or RNA of another organism, usually through recombinant technology.
vehicle: a material, such as saline, in a formulation that serves to enhance transfer, absorption, or distribution of a biopharmaceutical.
vendor: an entity that provides a material or service to a client. verification: to demonstrate, with documented evidence, that something,
such as a piece of equipment, is what it is purported to be. It is the
449Glossary
specific act of verifying and documenting that a compendial test performs as intended in a quality control laboratory.
viable particle: living contaminant (typically bacterial, fungal, or yeast) found in a product or manufacturing stream. It is an undesirable occupant of a manufacturing area.
virtual team: a project team that is separated by space and time but still functions as an effective group of individuals working together toward a common objective.
volume of distribution: the distribution of a biopharmaceutical, in quantita- tive terms, between blood and other tissues of the body after dosing. It measures the volume in which drug would be uniformly distrib- uted at any point in time.
voluntary action indicated (VAI): refers to findings on an EIR in which FDA recommends that action be taken by a manufacturer to correct minor deficiencies or deviations noted during an inspection.
volunteer: any individual who requests to be enrolled in a clinical trial. water for injection (WFI): highly purified water free of microbes, contami-
nants, or impurities and of a quality that can be injected into humans. WCB: See working cell bank. well-characterized: a product or material for which there is a significant
amount of scientific information, often chemical, biological, and physical, that provides a high degree of understanding on the nature and functional properties of the product.
WFI: See water for injection. WHO: World Health Organization, United Nations. WI: See work instruction. withdrawl: the instance of a volunteer in a clinical study leaving that
study on his or her own initiative or on request of the principal investigator.
work breakdown structure: a tool used in project management to identify the various work pieces of a project, that is, the tasks, and place them in a logical sequence of events or hierarchy. It is the basis for project planning—a visualization or narrative outline of project tasks, as mapped to component parts.
working cell bank (WCB): Derived from an MCB, this is the source of cells for production, also referred to as a production cell bank (PCB).
work instruction (WI): a document used in biomanufacturing to both guide a process and record critical information regarding a partic- ular batch or lot of product. Also called a batch production record (BPR).
yield: the amount of product that results from a step or stage of biomanu- facture. Often presented as a percentage, the amount obtained at the end divided by the starting amount.
450 Glossary
References
FDA. 2016. U.S. Department of Health & Human Services: U.S. Food and Drug Administration Home Page/Regulatory Information/Code of Federal Regulations/ CRF Title 21 – Food and Drugs: Parts 1 to 1499. http://www.accessdata.fda.gov/ scripts/cdrh/cfdocs/cfcfr/cfrsearch.cfm (accessed May 31, 2016).
Oxford English Dictionary. 1997. Oxford University Press. Oxford, UK.
451
Appendix
Overview
This addendum to this book provides an opportunity for readers to con- sider situations that occur in biotechnology operations and to develop a targeted product profile (TPP) and a product development plan for a bio- pharmaceutical product directed at treating or preventing a disease. In the first section, situations are posed. In each situation, a biopharmaceutical product is to be developed, as a team project. A small amount of technical background is provided and students are encouraged to make and state assumptions regarding the research results, intended population, indica- tion, and so on for each product. In the second section of this addendum, questions are posed with regard to planning development for each prod- uct. Each question on planning pertains to any of the six products.
Products and Projects
• Problem #1: The product to be developed is a monoclonal antibody. Your firm, ABC Biologicals, Inc., (Osco, IL) is developing a propri- etary monoclonal antibody to treat cutaneous T-cell lymphoma, a cancer of white blood cells. This biopharmaceutical product is a humanized monoclonal antibody that targets and binds specifically to a cell surface molecule, CD545. The molecule, developed in your laboratory, has been characterized in research and has been shown, in a transgenic mouse model, to bind to cancer cells, leading to their death. To be effective in mice, the product is given by subcutaneous injection. Your firm now wishes to develop the monoclonal anti- body product and apply for market approval.
• Problem #2: Your firm, EbolVac, Inc., has discovered and patented a surface protein of Marburg virus, an organism that causes hemor- rhagic fever in man. Ebola virus is considered a threat agent for bio- terrorism, and so, governments are the potential customers. A gene for an Ebola virus surface protein has been cloned and expressed in
452 Appendix
a host system as a 65 kDa glycoprotein. The product can be handled safely in the laboratory. Using a model of Ebola virus in nonhu- man primates kept in a high-containment facility, your firm has demonstrated proof of principle by protecting monkeys by using a nasal spray of the vaccine. You now wish to develop the product for human use and obtain market approval.
• Problem #3: Your firm, GenTherLine, holds an exclusive license for a novel gene therapy product composed of naked plasmid DNA. The gene therapy is intended to resolve nevoid basal cell carcinoma, a cancer of the skin, by replacing the gene for PTCH. The naked plas- mid DNA is a vector that includes the PTCH gene. It must be deliv- ered by a microneedle injection apparatus that exposes the DNA to the cells of the basal epidermis. Unfortunately, there is no animal model in which this mutation exists, as the mutation exists and is expressed only in man. Once the PTCH gene enters the nucleus of a cultured cancerous basal epidermal cell, it replaces the mutant gene that causes this disease, but the concept has not been attempted in man. You now wish to develop the product and obtain market approval.
• Problem #4: Your firm, MalarTher, has discovered a 35 kDa protein, MER24, that interrupts the life cycle of malaria parasites by blocking their ability to further infect red blood cells. MER24 acts directly on the merozoite stage of the malaria parasite, the erythrocytic or asex- ual cycle of red blood cell infection. Indeed, MER24, mimics a red blood cell receptor, thus binding to the parasite before it can bind to the same molecule on the host’s erythrocyte. It is considered a poten- tial therapeutic molecule to be given intravenously to individuals infected with Plasmodium falciparum. You now wish to develop the product and obtain market approval.
• Problem #5: Your firm, StemTechnolUS, develops human therapeutic products from human pluripotent stem cells. You have discovered and cloned a stem cell-derived cell line that differentiates into neu- rons and is suitable for repair of uncomplicated spinal cord injuries from blunt trauma. Cells can be given by surgical intervention. You now wish to develop the product and obtain market approval.
• Problem #6: Your firm, TheraGentCure, has the patent for a gene therapy construct. It is a retrovirus, specifically a lentivirus, that carries the gene for a tissue inhibitor of a matrix metalloproteinase that inhibits neuroblastoma tumor cell growth. It would be directly injected into tumor mass or into blood vessels, feeding the mass in an attempt to slow the growth of the tumor and perhaps cure the cancer. You now wish to develop the product and obtain market approval.
453Appendix
Questions by Chapter and Functional Area
Chapter 1: Background and Targeted Product Profile
• Discuss the class of biopharmaceutical represented by this product. • Provide a name for the product. • What competitive products, if any, are on the market, and how have
they been developed and marketed? Discuss market advantages your product should possess.
• Describe the disease or condition (indication) and the population that is subject to the disease.
• Develop a TPP for this product, with all elements of biopharmaceuti- cal labeling.
• Define exactly the indication and population your product would diagnose, treat, or prevent, as considered for the first market approval. Discuss the rationale for choosing this indication and the intended population.
• How, in general, will this TPP affect the plans for each functional area?
• Provide a product design, to include input, design, and output, based on the elements of the TPP.
Chapter 2: Project Management
• Identify the project and its purpose and provide an overview of the project from the standpoint of the project manager. Include purpose, scope, and technical and management objectives.
• Provide the composition of the project team enlisted for this project. Identify team members, their affiliations and locations, and authori- ties and responsibilities, and the rational for these choices. Identify stakeholders to the project and define their roles, apart from the team.
• Describe the team communication plan to include types of commu- nication and frequency of meetings.
• Describe metrics that would be used to measure the progress of the project.
• Describe the methods the project manager will apply for allocating, tracking, and managing resources allocated to the project.
• Identify the 10 major risks this project is likely to encounter. Outline the risk assessment and risk management plans for the project,
454 Appendix
focusing on risk mitigation early in the project. Describe the roles of the project manager in risk management.
• Describe your plan for identifying actual problems or issues and resolving them as a team.
• Establish a work breakdown structure, both as a narrative explana- tion and as an illustration (e.g., Gantt or PERT chart).
Chapter 3 and 4: Regulatory Affairs and Regulatory Compliance
• Is there a regulatory precedent for the treatment of this indication and population, and if so, are the predicate products approved by FDA or any other regulatory agency for marketing? How might this history of predicate products affect the regulatory development of your product? Is there precedent for this type of product (e.g., molec- ular or cellular nature) having received market approval or having been tested as an investigational drug? If so, what were the outcomes and how might this precedent affect the development of and regula- tory activities for your product? What do you think is the status of your product in the eyes of USFDA?
• Each product falls under one or the other office at FDA. Describe which office or division at FDA is likely to review your product. Explain how your product will fall into the scheme of FDA. Where will you submit applications within FDA?
• Will market approval be sought, initially, in the United States alone and/or in other countries? Describe your plans to submit marketing applications in each chosen country.
• What types of investigational use and marketing applications will be submitted to USFDA over the life of the product?
• For this product, outline the elements of the Investigational New Drug application (IND), in either IND or Common Technical Document format, as intended for FDA.
• Outline briefly an FDA marketing application in an acceptable format. Highlight the key elements that must be achieved during development, so that the application is complete. Will user fees be necessary, and if so, how much are the fees in today’s dollars?
• What other types of applications might be submitted to FDA in an effort to facilitate development, increase market share or exclusivity, or speed the process of approval?
• What FDA guidance documents are most important for your firm to consider for this product and indication?
455Appendix
• What is the nature and timing of meetings or teleconferences that might be held with FDA during the development process and before receiving market approval?
• As designed, could your product be considered in any way a com- bination product? Explain the rationale for this conclusion. If it is a combination product, then describe the impact on the regulatory plan and development pathway.
• Describe the major risks and benefits for the product, as FDA might perceive. Discuss the factors that will enter into a regulatory review of the risk-to-benefit ratio for this product.
• Describe postmarketing activities that might be required by regula- tory agencies for this type of product or indication?
• What non-FDA regulatory hurdles must be considered for this type of product? Do these hurdles pose major obstacles to product devel- opment, and if so, how will they be addressed and resolved?
• How might you add value or speed the development of this product by using various opportunities provided by FDA? Describe these programs the guidelines.
Chapter 5: Quality Systems and Quality Assurance
• Outline the contents of the quality manual that will serve your prod- uct’s development operation. Cite specifically each quality system that will be included and explain why and when in development it will be applied. Include a brief quality policy and a brief statement regarding management responsibility, as it would be approved by upper management.
• Describe the need to design your product, and explain how this will be accomplished. Provide product-related specifics regarding qual- ity by design and design control.
• Review the elements of a design program specific for your prod- uct, relating user needs, product attributes, and technical elements of design. Is it possible that your product must be reworked in the research laboratory before it begins development? If so or if not so, explain the exact reason.
• Choosing from the list of hallmarks of quality, identify what you consider to be the six most important with regard to the quality of product. Do not choose hallmarks of design. Explain why each was chosen and specifically elucidate how each will be applied to your product during both development and marketing phases.
456 Appendix
• Describe the fully functional Quality Assurance Unit that will sup- port the development of your product, and in your narrative, specifi- cally identify the examples of how the unit will function in the areas of audit, investigations, and change control.
• Describe the five most likely problems you will experience as you establish quality systems and a Quality Assurance Unit for this par- ticular product and project.
Chapter 6: Biomanufacturing
• Describe the biomanufacturing schemes that have been used to pro- duce biopharmaceuticals of a similar molecular or cellular nature. Identify regulatory guidelines that apply to the manufacture of such products.
• Discuss the major risks associated with manufacturing this type of product.
• Design a biomanufacturing plan for your product to include objectives, input, process, equipment, facility considerations, out- put, and review. Provide the process flow to include premanu- facturing preparation of constructs, upstream and downstream processing, holds, formulation, fill, finish, and raw material requirements.
• Outline how and at what phase of development the biomanufactur- ing process would be increased in scale. Provide plans to increase facility size or to use contract manufacturing operations.
• Provide general plans to validate the process, facility, equipment, and utilities.
Chapter 7: Quality Control
• Describe quality control test schemes that have been used to pro- duce biopharmaceuticals of a similar molecular or cellular nature. Identify regulatory guidelines that apply to the quality control of such products.
• Define and justify the attributes of bulk substance as they are based on its known nature and manufacturing scheme for bulk substance. Once the attributes are listed, draft a Certificate of Analysis, adding analytical methods and specifications to the attributes and consider- ing testing for more than one parameter of key attributes.
• Define and justify the attributes of final product as they are based on its known nature and the intended manufacturing scheme for final product. Once the attributes are listed, draft a Certificate of
457Appendix
Analysis, adding analytical methods and specifications to the pro- posed attributes and tests.
• Describe in-process samples that will be taken during the manufac- turing process, and identify attributes, tests, and possible specifica- tions for each sample.
• Briefly describe each test chosen for testing bulk substance and final product, for both release and stability, and assays for in-process samples. Explain why each test was chosen, based on performance, intended use, and meaningful results. Describe what is known about the specificity, accuracy, precision, range, and robustness of each assay when applied to this or to other classes of product. If an assay (e.g., potency) is to be developed just to test this product, justify the need to develop the assay and present ideas on its nature. Discuss the nature or need for control reagents and reference standards for the assays.
• Develop stability protocols for drug substance (DS) and final prod- uct (FP) and explain why each was chosen for the purpose of indicat- ing stability. Describe any tests used to measure stability that were not applied to product release.
• Consider attributes, analytical tests, and specifications for critical raw materials used in the process, working closely with the manu- facturing plan. Prepare a draft Certificate of Analysis for what you consider to be the five most critical raw materials.
• Identify which analytical tests will be qualified, verified, and/or val- idated, and mention the most likely point in the development cycle of each activity. Highlight critical assays that might require special attention in development, qualification, or validation.
Chapter 8: Nonclinical Studies
• Describe the nonclinical studies that are typically performed for this class of product and for products used with this indication. Consider both regulatory guidelines and precedent.
• What additional work must be performed to complete a pharmaco- kinetic and pharmacodynamic profile for this product, if it is given by the route and doses currently suggested in the clinical studies plan. Outline the studies that must be performed, and describe how and where these studies might be performed. Consider major design criteria for these studies.
• Describe the nonclinical toxicology studies that have been per- formed with other products in this class of product. Consider both regulatory guidelines and precedent.
458 Appendix
• Identify the nonclinical toxicology studies that should be performed to ensure safety of this product. For each study, provide a brief pur- pose and design and recommend when these studies might be per- formed by phase of development.
• Describe how quality system, cGLP, will be ensured for the planned studies.
Chapter 9: Clinical Studies
• Outline, in general, the clinical program that will be performed dur- ing the course of development.
• Given the nature of the product and the indication, outline the piv- otal or Phase 3 clinical study, and describe the objectives and indi- cation and the population in which your product will be tested. Describe the patient or subject population, approximate size, and scope and critical elements of study design.
• Outline Phase 2 clinical study on which you will demonstrate proof of principle and from which you intend to derive the information on which to base Phase 3 clinical study.
• Outline Phase 1 clinical study or studies that will be the foundation for Phase 2 study design.
• Present the requirements for choice of an investigational site and a principal investigator for each phase of the clinical development you outlined and in relation to your product and indication.
• Discuss the elements of current good clinical practices that would be required for each phase of the clinical development you have outlined.
• Describe any ethical considerations that might affect any one of your clinical trials, given the nature of the product, the study design, and the intended patient population.
459
Index
Note: Page numbers followed by f and t refer to figures and tables, respectively.
21 CFR, 71, 126, 180, 412 510(k) process, 77, 99–101
A
Abbreviated New Drug Application (ANDA), 75–76, 104
Absorption phase, 328 Accelerated Approval, 102 Accelerated stability, 308 Acetaminophen, 75 Active pharmaceutical ingredients
(APIs), 122–123 Acute toxicity testing, 351–352, 354–355 Adaptive design, 406 Adenovirus, 74, 232 ADEs (adverse drug events), 107 The Administrative Procedures Act
(APA) of 1946, 71 Adsorption, Distribution, Elimination,
and Metabolism (ADME), 325–328, 325f
absorption, 325 on bioavailability, 329 distribution, 325–327 excretion, 328 metabolism and biotransformation, 327
Adulterated/misbranded product, 123–124
Adventitious agent testing, 297–298 Adverse drug events (ADEs), 107 Adverse events (AEs), 15, 288, 383–384,
387, 389 clinical, 364–365 report, 178
Advisory committees, 106 Affinity chromatography, 228–229 Agreement State Program, 141 American Type Culture Collection, 212 Ames test, 340 Analyte/test substrate, 266
Analytical chromatographic technique, 275
Analytical methods, QC, 270, 286–295, 287f, 289f
attributes measurement, 266 controls and reference standards,
299–300 off-the-shelf, 266 qualification, validation, and
verification, 312–317 selection of, 270–277 tools and concepts, 295–297 traits, 267
Analytical precision, 313 ANDA. See Abbreviated New Drug
Application (ANDA) Animal and Plant Health Inspection
Service (APHIS), 134–136, 143 Animal model, 333–334
development, 342–344 selection, 344, 356 system, 163
Antihypertensive peptide, 16 Antimalarial drug, 74 APA. See The Administrative
Procedures Act (APA) of 1946 APHIS. See Animal and Plant Health
Inspection Service (APHIS) APIs (active pharmaceutical
ingredients), 122–123 Appearance test, 282, 288 Aseptic technique, 236, 251 Assay performance and validation,
application of statistics, 317–318, 318f
Assay verification, 316 Auditing/monitoring process, 389 Auditing process, 179, 185–188
performance, 186 quality, 187f
Autoinjector, 249
460 Index
B
Bacteria, 212, 232–234, 296 Bacterial cell expression systems,
212–213 Bacterial plasmid DNA, 231–232 Baculovirus, 216 Batch production records (BPRs), 172 Bibliography, 81 Bicinchoninic acid (BCA), 275, 283, 313
reagent-based test, 291 BIMO (biomonitoring), 98 BIO. See Biotechnology Industry
Organization (BIO) Bioavailability, 329, 332–333 Bioequivalence, 332 Biological potency assays, 273, 285 Biologics, 72–75
follow-on, 104 The Biologics Control Act (BCA) of
1902, 66 Biologics License Application (BLA),
95–96, 98–99, 107, 142 Biomanufacture, 3, 309–310
compliance and quality in, 207–209 design, 196–200 facilities, utilities, and equipment,
253–257 clean work areas for, 255–257 controlled environment,
254–255 equipment, 257 facility design considerations,
253–254 validation, 259–261
for FP, 248–252, 250f, 252f hypothetical scheme, 239f life cycle, 201–205 overview, 195 planning, 23–24, 196, 248 process for biotechnology products
bacterial plasmid DNA, 231–232 biologically active peptides,
245–247 biological molecules production,
238–245 cellular products production,
236–238 combination products, 247–248
in-process testing and bulk substance analysis, 230–231
lipids, glycolipids, and complex carbohydrates (biologically active), 245, 246f
live recombinant organisms production, 232–234
mammalian somatic cell/tissue products production, 234–236
of recombinant proteins, 217–230 recombinant proteins and nucleic
acids expression, 209–217 QbD in, 198–199 raw material considerations,
205–207 scale-up, 204–205 steps of, 196, 197f technical considerations for, 200–201
Biomedical technology assessment, 143 Biomonitoring (BIMO), 98 Biopharmaceutical(s), 1, 72, 266
application, 284 bioavailability, 329 biodynamic experimentation, 333 biomanufacturing life cycle in,
201–205 early phase, 202 late-phase, 204 mid-phase, 202, 204
class of, 78f compliance for
import, 119–120, 137 medical devices, 120
concentration in blood over time, 328–329, 329f effect curve, 331–332, 331f
delivery, 323–328 ADME, 325–328, 325f product delivery to body, 323–325
designer molecule, 331 development, 11, 29, 165, 168, 266, 284,
371 biomanufacturing activities for,
203 Gantt chart format, 43f life cycle, 87, 88f PERT chart format for, 45f timeline format, 44f
461Index
excipient, 345 fingerprint of, 274 FPs, 249, 283 immediate upregulation by, 361 immunological toxicity study, 362 label, 108 nonclinical activities in, 321–323,
322f, 337–339, 338f nonclinical safety testing for
evaluation, 352–353 operation, 257 pharmacokinetics and
pharmacodynamics in, 333–335
phased scheme, 7, 8f preventive and therapeutic, 7 problems, 175 processing, 196 promotional information, 109 safety assessment
acute toxicity testing, 351–352, 354–355
carcinogenicity testing, 360–361 genetic toxicology, 363, 366–367 immunotoxicology, 361–363 nonclinical safety testing,
351–353 nonclinical study design,
elements, 347–351 program, 337–340 protocols and performance,
346–347 reproductive, developmental,
and teratogenicity toxicity testing, 359–360
subchronic and chronic toxicity testing, 352, 356–359
tissue binding/local tissue tolerance, 367–368
toxicology, 336–337 in vitro screening test, 340–342 in vivo safety testing, 342–346
treatment, 284 type of, 78f
Biopharming, 238, 244, 253 Bioreactor, 168, 220, 238
cell, 221 transgenic goat as, 242
Biotechnology, 32 development
ICH guidelines in, 127–128 program, 169–170 team, 13
environmental regulations in, 141–142
industry, biological materials, 132–133
material transportation, 133–136 microbial products, 142 operation, 1–3, 142, 259, 300
focal point, 2 functional areas, 2–3 international diligence, 144–146 metaphor, 5–6 project management in, 41–58 quality systems, 188–189 skills and backgrounds, 1–2 themes, 1
product(s), 4, 7, 13, 253 biomanufacturing processes, 209 development, 10 fermentation, 218 importation/exportation, 137, 139 with IVD, 100 medical, 126–132 nature, 198–199 and reviews FDA, 79–81
and project management, 29–31 project team, 37–38 QC, 265 quality in, 149–150
Biotechnology-derived products, 72, 79 Biotechnology Industry Organization
(BIO), 131, 145 Biotechnology Regulatory Services
(BRS), 134, 143 Biotransformation, 327 BLA. See Biologics License Application
(BLA) Black box warning, 86 Blood-borne pathogen, 141 Blood products, 72–73 BPRs (batch production records), 172 Bradford test, 291 Brand name drug, 104 Breakthrough therapy, 103
462 Index
British Pharmacopoeia (BP), 271 BRS. See Biotechnology Regulatory
Services (BRS) BS. See Bulk substance (BS) Budgeting, 54 Budgeting monetary requirements, 55 Bulk API/final product, 123 Bulk substance (BS), 196, 217, 263, 265,
267, 282, 302 analysis, 230–231 batches, 281–282 biopharmaceutical
CoA, 268t–269t stability protocol for, 305t, 307
potency assay, 281 sterile filtration, 249
Bureau of Industry and Security (BIS), 136
C
CAPA (corrective and preventive action), 122
Carbohydrate analysis, 276 Carcinogenicity testing, 127, 340, 353,
360–361 Case report forms (CRFs), 380, 391,
395–396, 400 CBER. See Center for Biologics
Evaluation and Research (CBER)
CBP. See Customs and Border Protection (CBP)
CDC. See Centers for Disease Control and Prevention (CDC)
CDER. See Center for Drug Evaluation and Research (CDER)
CDRH. See Center for Devices and Radiological Health (CDRH)
Cell banks production, 211f, 216–217 QC of, 297–298 transgenic plant, 244
Cell-based assay, 276–277 Cell growth curve, 221 Cell karyotyping, 295 Cell phenotyping, 296 Center for Biologics Evaluation and
Research (CBER), 69–70, 72, 74, 76, 356
Center for Devices and Radiological Health (CDRH), 70, 77, 100
Center for Drug Evaluation and Research (CDER), 69–72, 74, 76
Center for Veterinary Medicine (CVM), 70, 79
Centers for Disease Control and Prevention (CDC), 134, 136
Centrifugation, 222, 225, 234 Certificate of Analysis (CoA), 263,
266–267, 319 drug product, 282–285 QC, 267–270 vendor-supplied, 310
CF. See Consent form (CF) CFR. See Code of Federal Regulations
(CFR) cGCP. See Current Good Clinical
Practices (cGCP) cGLP. See Current Good Laboratory
Practices (cGLP) cGMP. See Current Good Manufacturing
Practices (cGMP) CGTP (Current Good Tissue Practices), 73 Change control, 175, 184 Chemistry, Manufacturing, and
Controls (CMC), 129 Chinese hamster ovary (CHO), 214 CHMP (Committee for Human Medical
Products), 131 Chromatogram, 292, 307 Chromatography, 292
gel, 292 high-pressure liquid, 292–294,
293f–294f preparative, 227–228
equipment, 227f flow diagram, 226f
Chronic infection, 401 Chronic toxicity testing, 352, 356–359 Clean dose level, 356 Cleaning protocols, 257 Clearance, 328–329 Clinical data, 395–396 Clinical development, 371, 405
biopharmaceutical products, 375 key issues, 375 planning, 18–19, 377–378
Clinical laboratory testing, 248
463Index
Clinical pharmacology, 11, 15, 97, 375, 404–405
Clinical program, 376, 417 Clinical protocol, 89–90, 379, 382
clinical trials and, 378, 381–387 elements, 378, 381, 383
Clinical safety and toxicology, 375 Clinical study/research, 172, 174,
371, 376 associates, 414 organization, 375–376
cGCP, qualtiy in, 377 clinical development planning,
377–378 phases, 375–376 science, 376–377
overview, 373–374 Clinical summary report (CSR), 397, 400 Clinical testing laboratories, 396–397 Clinical trial(s), 371
documents, 379–380 historical information, 374–375 individuals and responsibilities, 379 infrastructure
clinical testing laboratories, 396–397
collection, 395–396 design of, 378–387 human subjects, patients, and
volunteers, 388 investigational product, 394–395 IRB, process of IC, and IC form,
392–394 PI, 391–392 results, 397 sponsor, 388–391
monitoring, 389 operations, 397–398
activities leading, 398–400 clinical pharmacology studies,
404–405 first-in-human study, 400–403,
402f, 403f global, 409 new populations/indications,
408–409 proof-of-concept study, 405–406 REMS, 407–408 therapeutic confirmatory, 406–407
overview, 371–373, 372f performance, 413 phases, 375–376 quality system, 409–412
clinical study data and documents, 413–414
ethical behavior and well-being, 415–416
monitoring and auditing, 414 quality and cGCP, 412–413
scheme, 372f WHO guidelines, 128, 131
CMC (Chemistry, Manufacturing, and Controls), 129
CMO. See Contract Manufacturing Options (CMO)
CoA. See Certificate of Analysis (CoA) Code of Federal Regulations (CFR), 71,
128, 273, 393 Codex standards, 145 Committee for Human Medical
Products (CHMP), 131 Common Technical Document (CTD),
91–92, 92f electronic submission of, 92–96
Communication, 37, 46, 50, 140 day-to-day, 48 direct, 38, 63, 107 electronic, 60 and feedback, 49–50 regulatory, 85–86 team, 48
Compendia, 82, 309 test, 271–273
Complex carbohydrates, 245 Computer software/program, 62 Concentration effect curve, 331–332, 331f Concept protocol, 378–379 Conditional approval, 407 Consent form (CF), 379, 393 Contract Manufacturing Options
(CMO), 253, 257–259 Contractor, vendor, and consultant
control, quality system, 169–171 Contract research organization (CRO),
59, 117, 170–171, 273, 285, 388, 391
Control article, 345 Controlled bioavailability, 329
464 Index
Controlled studies, 374 Corrective and preventive action
(CAPA), 122 Corrective/preventive actions, change
control, 175–176, 177f CRFs. See Case report forms (CRFs) Criminal prosecution, FDA judicial
actions, 125 CRO. See Contract research organization
(CRO) Cross-referencing information, 101 CSR (clinical summary report), 397, 400 CTD. See Common Technical Document
(CTD) Current Good Clinical Practices (cGCP),
114, 154, 156, 373–374, 409–416 for clinical studies, 117 in clinical trial operations, 412–413 key components, 117, 119 quality in, 377
Current Good Laboratory Practices (cGLP), 114, 154, 339, 347, 355, 368–369
elements of FDA, 117–118 for nonclinical laboratory studies, 117
Current Good Manufacturing Practices (cGMP), 114, 154, 156, 170, 202, 207–209
elements, 114–117 for manufacture and quality control,
114–117 objective, 208 quality system, 157
Current Good Tissue Practices (CGTP), 73 Customs and Border Protection (CBP),
120, 136, 139 The Cutter Incident of 1955, 66 CVM. See Center for Veterinary
Medicine (CVM) Cytogenetic analysis, 295 Cytokine storm, 363
D
Data and Safety Monitoring Board (DSMB), 407
Day-to-day communication, 48 DBOP (Division of Biologic Oncology
Products), 76
Declaration of Helsinki, 392, 415–416 Degree of scatter, 314 Design change, quality system, 169 Design control, QbD, 163–169
change, 169 documents and records, 166–167 elements, 165–166, 165f product specific, 166
Designer molecule biopharmaceutical, 331 Developmental toxicity testing, 359–360 Dietary supplements, 79–81 Direct communication, 38, 107
tools, 63 Division of Biologic Oncology Products
(DBOP), 76 Documentation system, 182–184 Document control, quality, 178, 179f Downstream process, 196, 279
recombinant proteins, 222–230 Downstream purification, 222–230
material list, 207 Draft/concept product labeling, 11, 13 Drafting CoA, 267–270
BS, 268t FP, 269t
Draft labeling, 13, 99 Draft project management plan, 17, 46 Draft TPP, 11 Draft WBS, 61 Drug(s), 75–76, 89, 102
CoA, product, 282–285 development, 7 generic, 104 interaction, 12, 327 laws, 67, 71
regulation/guidance, 71 nonclinical safety tests, 352–353 in vitro safety tests, 341
Drug-drug interactions, 407 Drug-receptor interaction, 330 DSMB (Data and Safety Monitoring
Board), 407
E
Early phase biomanufacturing development, 202
Early phase dosing study, 401 Earned value management (EVM), 53, 54f
465Index
eCTD. See Electronic CTD (eCTD) EEC (European Economic
Community), 131 Effective project tracking, PM, 53 Efficacy topics (E), 127–128 Electronic CTD (eCTD), 92–96, 98 Electrophoretic method, 290 Electrospray ionization-mass
spectrometry, 294 Elements to ensure safe use (ETASU), 408 Elimination phase, 328 Elution buffer, 228–229 EMA (European Medicines Agency), 131 Embryo-fetal development, 360 Endotoxin, 212–213
test, 271, 276, 288 End point, 354, 384, 387, 401
acute toxicity study, 355 immunological toxicity study, 362 subchronic/chronic toxicity study, 357
Engineered retrovirus, 247 Environmental assessments, 142 Environmental controls, 173 Environmental Protection Agency
(EPA), 142–143 Environmental release, 135 EPA (Environmental Protection
Agency), 142–143 Epidemiological, 373 Escherichia coli, 74, 212 Establishment Inspection Report (EIR), 122 ETASU (elements to ensure safe use), 408 European Economic Community
(EEC), 131 European Medicines Agency (EMA), 131 European Pharmacopoeia (EP), 271 EVM (earned value management), 53, 54f Excretion, ADME, 328 Expression of recombinant proteins and
nucleic acids, 209–217 bacterial cell expression systems,
212–213 cell banks production, 211f, 216–217 genes, vectors, and host cells, 210–212 mammalian/insect cell expression
systems, 213–216 molecules production from
expression vectors, 209–210 yeast cell expression systems, 213
Expression vector, 209–210, 214 External audits, 180, 185
F
Face-to-face meetings, 61, 63 Failure mode and effects analysis
(FEMA), 52 Fast Track Designation, 103 Fault-tree analysis, 52 FDA. See Food and Drug
Administration (FDA) The FDA Amendments Act of 2007, 407 FDA-regulated products, 71–81
biologics, 72–75 biotechnology products/reviews,
classes of, 79–81 combination products, 77–78, 78f drugs, 75–76 medical devices, 76–77
FDA’s Inspection Operations Manual, 122 The Federal Food, Drug, and Cosmetic
(FD&C Act) Act of 1938, 65–66, 70, 77
The Federal Insecticide, Fungicide and Rodenticide Act (FIFRA), 142
The Federal Plant Protection Act, 135 Federal trade commission (FTC), 65 FEMA (failure mode and effects
analysis), 52 Fermentation
microbial, 218 upstream, 206 vessel, 261 yeast cell, 218–220
FIFRA. See The Federal Insecticide, Fungicide and Rodenticide Act (FIFRA)
Final product (FP), 123, 196, 248–252, 263 active ingredient in, 283 CoA, 269t manufacture, 250 problem, 283 QC tests, 282–283 stability protocol, 306t strength, 283
First-in-human study, 400–403, 402f, 403f FOIA. See The Freedom of Information
Act (FOIA)
466 Index
Follow-on biologics, 104 Food and Agricultural Organization
(FAO), 145 The Food and Drug Act, 152 Food and Drug Administration (FDA),
6, 11, 65–71, 120–121, 129–131, 152, 161–162, 164, 186, 190
additional regulatory activities, 105–109
advisory committees, 106 applications, regulatory operations,
84–99 BLA/NDA, 96–99 environment, 84 IND application, 89–96 investigational use/marketing
approval, 87–88 planning, 84 risk vs. benefit, 84, 86–87
biotechnology products/reviews, 70, 79–81
CDER, 75 centers/offices, 69 enforcement actions, 123–125 form, 122–123 guidance documents, 129–131
clinical, 130 CMC, 129 preclinical, 131 quality, 130
inspection process, 120–123 law/regulations for
biopharmaceuticals food and drug law, regulation,
and guidance, 71 historical basis for FDA
regulation, 65–66 regulatory organization of FDA,
66–71, 68f letters to manufacturers, 108 product liability, 125 review staff, 94 user fee, 99
The Food and Drugs Act of 1906, 65 The Food, Drug, and Cosmetic Act of
1938, 121, 123 Formal clinical hold mechanism, 93 Formal meetings, 48 Forming stage, 46
Formulation–fill–finish production methods, FP, 248–252, 250f, 252f
FP. See Final product (FP) The Freedom of Information Act (FOIA),
107, 123 Freeze drying, 251 Funnel concept, 164
G
Gantt chart, 20, 42, 43f, 62–63 Gas chromatography, 292 Gel electrophoresis, 274 Generally Recognized As Safe (GRAS),
80 General safety test, 283, 288 Generic drugs, 75–76, 104 Generic tests, 273, 283 Genes, vectors, and host cells, 210–212 Gene therapy, 333, 401 Genetically modified organisms
(GMOs), 135, 142–144 Genetic engineering, 76, 135, 144, 200 Genetic toxicology, 363, 366–367 Global clinical trials, 409 GLP regulation, 368 Glycolipids, 245 Glycosylation, 212, 244 GMOs. See Genetically modified
organisms (GMOs) Golden rule, 39 Gram-negative bacteria, 212–213, 288 GRAS (Generally Recognized As Safe), 80
H
Haemophilus influenzae type b, 245 Half-life value, 328, 404 Hallmarks, 159, 162, 230, 399
quality systems, 153, 159–180 Hamburg, Margaret A., 198 Health and safety plans, 140–141 Heat, ventilation, and air conditioning
(HVAC) system, 255, 257 High-pressure liquid chromatography
(HPLC), 292–294, 293f–294f, 307, 316
HIV test kits, 77 Homeopathic medicine, 81
467Index
Host cell, 210–212, 296–297 DNA, 291 protein, 291 RNA, 291 yeast, 213
HPLC. See High-pressure liquid chromatography (HPLC)
Human clinical study, 410–411 Humanized monoclonal antibody,
166–167, 363 Human pharmacology, 375 Human subjects, patients, and
volunteers, clinical trial, 388 Hydrophobic interaction
chromatography, 228 Hypertension (high blood pressure),
14, 384
I
IATA (International Air Transportation Association), 132
IB. See Investigator’s Brochure (IB) IBC (Institutional Biosafety
Committees), 143–144 IC. See Informed consent (IC) ICF. See Informed consent form (ICF) ICH. See International Council for
Harmonization (ICH) IDE (Investigational Device Exemption),
100 IFPMA. See International Federation of
Pharmaceutical Manufacturers and Associations (IFPMA)
Immunoelectrophoresis, 290 Immunohistochemical methods, 290 Immunotoxicology, 361–363 Impurities-truncated peptides, 247 INAD (Investigational New Animal
Drug), 79 Inclusion and exclusion criteria, 384–386 IND. See Investigational New Drug
(IND) application Individual effective concentration, 332 Induced pluripotent stem cell (iPSC),
236–237 Industry standard, 75, 208 Informal meetings, 48 Informal risk analysis, 52
Informed consent (IC), 379, 392, 410 elements, 393 process and form, 392–394
Informed consent form (ICF), 379, 393 Injunctions tools, 125 In-life
measures, 350–351 phase, 349
Inoculum, 219 Inorganic and organic chemicals, 310 In-process testing, 174, 223, 230–231,
285–286 Inspections, 98
FDA, 121–123, 147 quality systems, 173–174
Institutional Biosafety Committees (IBC), 143–144
Institutional review board (IRB), 389, 391, 393–394, 399
Intergeneric microorganisms, 142–143 Internal audits, 180, 185 International Air Transportation
Association (IATA), 132 International Council for
Harmonization (ICH), 91, 126–128, 154, 164, 377, 412
International Federation of Pharmaceutical Manufacturers and Associations (IFPMA), 126, 131
International Standards Organization (ISO) 9001, 154–156, 170
Intramuscular injection, 14 Investigational Device Exemption
(IDE), 100 Investigational New Animal Drug
(INAD), 79 Investigational New Drug (IND)
application, 89–96, 321, 377 contents, 90–91 CTD, 91–92 eCTD, 92–96 legal document, 93 regulatory operations, FDA
applications, 89–91 treatment, 102
Investigational product, 363, 392, 394–395
accountability log, 380
468 Index
Investigator’s Brochure (IB), 90, 380, 389 elements, 390
In vitro assay, 276–277 In vitro diagnostics (IVDs), 100 In vitro screening test, 340–342 In vivo safety testing, 341–346
animal model development, 342–344 test product formulations, delivery
routes, and dosing designs, 344–346
Ion-exchange chromatography, 228 Ionic strength, 288 iPSC (induced pluripotent stem cell),
236–237 IRB. See Institutional review board (IRB) Isoelectric focus gels, 292 IVDs. See In vitro diagnostics (IVDs)
J
Jargon, 2, 174 The Jungle (book), 65
K
Kilodalton (kDa), 294 Knee joint cartilage replacement, 235
L
Label claim, 12, 13, 377 Labeling, FP, 251–252 The Lacey Act, 135 Late-phase biomanufacturing
development, 202, 204, 260 Lessons learned reports and meetings, 58 Lethality testing, 340 Lethal toxin, 279 Letter of authorization (LOA), 101 Limit of detection (LOD), 267, 313 Limit of quantitation (LOQ), 267, 313 Linearity, QC, 267, 313 Liquid chromatography-mass
spectrometry, 294 Live recombinant organisms, 232–234 Loading dose, 330 Local tissue tolerance, 367–368 Lymph amebocyte lysis (LAL), 288 Lyophilization, 251
M
Maintenance dose, 330 Mammalian/insect cell expression
systems, 213–216 Mammalian somatic cell/tissue
product, 234–236 Management reviews, 162 Market- and user-driven process, 248 Marketing
application, 96–99 approval, 87–88, 132 considerations for biotechnology
development, 4–7 Mass spectrometry-time of flight, 295 Mass spectroscopy, 276 Master cell banks (MCB), 216–217 Master File (MF), 101–102 Maximal efficacy, 332 Maximum concentration value, 328 MDR (medical device reporting), 120 Medical device, FDA
applications, 99–104 generic drugs and biosimiliar/
follow-on biologics, 104 special documents, pathways, or
exemptions, 101–104 -regulated products, 76–77
Medical device reporting (MDR), 120 MedWatch, 107 Merck Index (book), 271 Merck Manual (book), 271 Metabolism, 327 Microbes, 14, 255 Microbial enumeration test, 287 Microbial fermentation
equipment, 218f phases of, 219, 220f
Microbial identification, 296 Microbial limits test (MLT), 287–288 Microbial load/bioburden, 287 Microbial product, 73, 296 Microprocessors, 61, 219 Microsoft Project®, 53–54, 62 Mid-phase biomanufacturing
development, 202, 204 Milestone-related task, 52 Misbranded labeling, 124 Monitoring and auditing clinical trials, 414
469Index
Monoclonal antibody, 76, 98, 163, 168, 170, 228, 290, 332–335, 367
Multidisciplinary topics, 127 Multiple potency assays, 284 Multiple rising dose design, 401, 403f Mundane issues, 189 Mutagenicity testing, 340
N
NADA (New Animal Drug Application), 79
National Center for Import and Export (NCIE), 135
National Drug Code (NDC), 105 The National Environmental Policy Act
of 1969, 142 National Formulary (NF), 75, 272 National Institute for Occupational
Safety and Health (NIOSH), 141
National Institutes of Health (NIH), 143–144
National regulatory authorities (NRAs), 126
National Science Advisory Board for Biosecurity (NSABB), 144
NCIE (National Center for Import and Export), 135
NDC (National Drug Code), 105 Negative control antisera, 290 Network building, 46 Neupogen (filgrastim) drug, 104 Neural system, 3–5, 10 New Animal Drug Application
(NADA), 79 New drug application (NDA), 65, 96,
98–99, 107 NF. See National Formulary (NF) NIH (National Institutes of Health),
143–144 NIOSH (National Institute for
Occupational Safety and Health), 141
Nonclinical animal study, 345 Nonclinical planning, 22–23 Nonclinical safety testing, 339–340,
351–353
Nonclinical study, 172, 321–323, 322f in biopharmaceutical development
project, 337, 338f calculating dose, 346 design, 346 elements, 347–351 protocol, 175, 346–347 quality of, 368–369 route of delivery, 345
Nonclinical testing, 73, 332, 339 Nonclinical toxicology testing, 16 Nonconforming product, 175 Non-FDA regulations, 126
application of, 146 biotechnology
environmental regulations in, 141–142
importation/exportation, 137–139 importing, possessing/
transferring, 134–136 international and foreign NRAs,
126–132 international diligence in, 144–146
GMOs, 142–144 occupational health and safety,
140–141 The Public Health Security and
Bioterrorism Preparedness and Response Act of 2002, 136–137
transporting infectious/hazardous materials, 132–134
Nongovernmental organizations/ agencies (NGOs), 132
Normal saline, USP grade, 170 Novel biotechnology product, 1, 84, 401 NRAs (national regulatory
authorities), 126 NSABB (National Science Advisory
Board for Biosecurity), 144 N-terminal sequencing, 283, 289 Nuclear magnetic resonance, 276 Nuclear Regulatory Commission
(NRC), 141 Nuremberg Code, 392, 416
O
OAI (official action indicated), 122–124 Obligation log, 380
470 Index
Office of Biotechnology Activities (OBA), 143–144
Office of Biotechnology Products (OBP), 76
Office of Blood Research and Review (OBRR), 72
Office of Cellular, Tissue and Gene Therapies (OCTGT), 73
Office of Combination Products (OCP), 77
Office of Vaccine Research and Review (OVRR), 74
Official action indicated (OAI), 122–124 Off-label, 109 Operational documents, 161, 183 Operational research, 158 Ops Manual, 387 Optimized clinical formulation, 334 Oral-gastrointestinal absorption, 325 Oral ingestion, 324 Orphan Drug Exclusivity (ODE), 103 Orphan Products Development
(OPD), 103 OSHA (Occupational Health and Safety
Administration), 141 Osmolality, 219, 288 Osmometer, 288 Outsourcing models, PM, 59 Over-the-counter (OTC) drugs, 75, 108
P
Packaging FP, 251 and labeling, 176
PAGE. See Polyacrylamide gel electrophoresis (PAGE)
Pathogens, 133 PDP. See Product development plan
(PDP) PDR. See Physician’s Desk Reference
(PDR) PDS. See Product development strategy
(PDS) Peptide, 14
antihypertensive, 15 biologically active, 245–247 mapping, 276, 292 therapeutic product, 13
Peptide-based biomolecules, 247 PERT chart, 20, 45f, 63 Pharmaceutical, 32, 75, 229
quality system (ICH Q10), 155 Pharmaceutical Research and
Manufacturers of America (PhRMA), 131
Pharmacodynamic, 328–333 in biopharmaceutical development,
333–335 variability, 332
Pharmacokinetic (PK), 328–333 in biopharmaceutical development,
333–335 clearance aspect, 329 study, 376, 399, 404
Phased product biomanufacture, 202 pH, QC regulatory requirement, 289 PhRMA (Pharmaceutical Research and
Manufacturers of America), 131 PHS Act. See The Public Health Service
Act (PHS Act) Physician’s Desk Reference (PDR), 96, 271 PI. See Principal investigator (PI) Pichia pastoris, 212–213, 220 Pilot production, 217 Pivotal clinical trial, 376, 406 Placebo, 345, 374, 382, 387, 401, 405–406 Planning backward, 9f Plant pest, 135 Plasmid DNA, 73
bacterial, 231–232 Platelet-rich plasma (PRP), 245 Pluripotent cells, 73
cellular product from, 236–238 PM. See Project manager (PM) PMA. See Premarket approval (PMA) PMP. See Project management plan (PMP) Polyacrylamide gel electrophoresis
(PAGE), 273, 289–290, 289f Polyhistidine tag chromatography, 229 Polymerase chain reaction, 210, 296 Polysaccharide-protein, 246f Polyvalent antiserum, 290 Postmarketing requirements and
activities, 107–108 Potency, 270, 298, 331–332
assays, 276, 286 tests, 234, 266, 281, 284
471Index
Pre-approval Inspections (PAI), 98 Precision, 267, 313–314, 314f Pre-IND communication, 95 Pre-IND meeting, 89, 94–95 Premarket approval (PMA), 77, 99–100 Preparative chromatography, 227–228
equipment, 227f flow diagram, 226f
Preservation, storage, and handling, 176 Principal investigator (PI), 382, 388,
391–392 Process control, 172–173 Product attributes, QC, 265–266 Product design plan, 166 Product development
planning PDP, 16–28 rationale, 7–10 TPP, 10–16, 9f
project team, skills, 38 quality systems approach to, 153–155
Product development plan (PDP), 16–29, 195
biomanufacturing, 23–24 clinical development, 18–19 nonclinical, 22–23 product planning, elements of, 26–28 project management, 19–20 quality control, 24–26 quality systems and assurance, 26 regulatory, 20–22
Product development strategy (PDS), 16, 84, 87
Product identification and traceability, 171–172
Product labeling, 96–99, 109, 176 concept, 11–12
Product liability, 125 Product stability testing, 302–308, 303f Project champion, stakeholder, 38, 56 Project committee meetings, 158 Project leader, 38, 56 Project management, 17
background, 31–32 in biotechnology, 29–31, 30f
communication and feedback, 49–50
establishing, 41–42 human factors in, 55–57, 58f
metrics and tracking progress, 53–54, 54f
project completion, 57–58 project risk assessment, 51–52, 51f project team/hands-on, 46 resources, 54–55 team dynamics, 46–49 WBS, 42–45
contracts and collaborations, 59–60
environment, 34–36 operational phase, 30 participants in, 37–41 planning, 19–20 project objectives and schedules,
36–37 sociotechnical considerations, 37 software, 42, 61, 63 stages, 30f tools, 43f, 44f, 45f, 49
effective, 61–63 virtual teams, 60–61
Project management plan (PMP), 29–30, 32–34, 33f
elements, 33–34 environment, 34–36 participants in, 37–41 project objectives and schedules,
36–37 sociotechnical considerations, 37
Project manager (PM), 11, 17, 29, 37–39 attributes of effective, 40 diligent, 53 effective project tracking, 53 full-time, 41–42 human resource duties, 55–56 selection, 40
Project risk assessment and management, 51–52, 51f
Project team, 6, 30–31, 52 biotechnology, 37–38 formation in biotechnology, 46 guidelines and elements of, 47 human sources conflict on, 57 types, 47
Project wrap, 58 Promotional labeling, 108 Proof-of-concept study, 405–406 Protein reference standards, 290
472 Index
The Public Health Security and Bioterrorism Preparedness and Response Act of 2002, 136–137
The Public Health Service Act (PHS Act), 66
The Public Health Service Act (PHS Act) defined, 72
Public law PL107-188, 138–139 Public regulatory information, 82 PubMed, 82 The Pure Food and Drugs Act in 1906, 65
Q
QA. See Quality assurance (QA) QAU. See Quality assurance unit (QAU) QbD. See Quality by design (QbD) QC. See Quality control (QC) QSR. See Quality systems regulation
(QSR) Quality
agreements, 171 audits, 185 hallmark, 160 management
principles, 160, 160f QAU/QA, 191 quality issues/problems, 192–193 quality systems for research,
191–192 risk-based approaches, 190 six sigma, 191 TQM, 190 unique and effective approaches,
190–193 manual, 156–157 plan, 157–159 professionals, 158, 167, 175, 180, 182,
192 steering committee, 158
Quality assurance (QA), 3, 175, 180, 192 function, 181–182 planning, 26 statistics, 191
Quality assurance unit (QAU), 159, 176, 180–181, 348
auditing, 185–188 control and manage documentation
system, 182–184
functions, 181–182 investigate situations, 184 major task, 183 qualified and trained staff, 184–185 report, 161 responsibilities of, 181f staff members, 185 weak, 189
Quality by design (QbD), 149, 196, 198–199 design change, 169 and design control, 163–169 overview, 163–164
Quality control (QC), 263 additional analytical tools and
concepts, 295–297 analytical controls and reference
standards, 299–300 analytical methods, 286–295
qualification, validation, and verification, 312–317
selection, 270–277 assays, 273
performance and validation, 317–318
cell banks, 297–298 CoA, 267–270
for drug product, 282–285 compendia and reference, 271 development cycle, 264–265, 264f in-process testing, 285–286 laboratory, 270, 287f, 311 manufacturing environment, 310–311 objective, 263 overview, 263–265, 264f plan, 265 planning, 24–26, 263–264 product attributes, 265–267
analytical methods, 266–267 product stability testing, 302–308 sampling methods, 298 specifications development, 277–281 test failures, out-of-specification
results, and retesting, 300–301 testing, 265, 282
of raw materials, 308–310 results, 282
Quality system(s) in biotechnology, 149–150, 188–189 cGCP, 117, 119, 412–413
473Index
clinical study data and documents, 413–414
clinical trials, 409–416 defined, 162 ethical behavior and well-being,
415–416 evolution, 150–153, 151f fundamental criteria for building
effective, 159–180 auditing, 179–180 contractor, vendor, and consultant
control, 169–171 control and corrective/preventive
actions, 175–176, 177f customer concerns and adverse
event reports, 178 document control, 178, 179f environmental control, 173 inspection/testing (quality
control), 173–174 management responsibility,
160–162, 160f, 161f material, service, or product,
release of, 174 packaging and labeling, 176 preservation, storage, and
handling, 176–177 process control, 172–173 product identification and
traceability, 171–172 QbD and design control, 163–169 servicing, 178 training, 178–179
hallmarks of, 153 monitoring and auditing, 414 planning
objectives, 156 quality manual, 156–157 quality plan, 157–159
product development, 153–155 QAU, 180–188 regulatory compliance, 113–119
cGCP, 117, 119 cGLP, 117–118 cGMP, 114–117
for research, 191–192 Quality systems regulation (QSR), 99,
120, 155 Quality topics (Q), 127
R
RAC (Recombinant Advisory Committee), 144
Randomization/blinding process, 374, 382
Recombinant Advisory Committee (RAC), 144
Recombinant biological molecules, 158 Recombinant blood products, 72 Recombinant crop plant, 150 Recombinant DNA vaccine, 247 Recombinant microorganisms, 133 Recombinant protein (r-protein), 13, 173,
231, 265–266, 274–275, 277 analysis, 268t, 269t biomanufacture of, 217–230
downstream process, 222–230 planning production, 217 upstream process, 218–222
CoA, 313 and nucleic acids expression See
Expression of recombinant proteins and nucleic acids
production and control by transgenic plant, 243f
Recombinant vaccine protein, 77 Record control, 178 Reference standards, 274–275,
299–300, 315 Regulatory compliance, 113
for biopharmaceuticals, 119–120 inspection and enforcement, 120–125 with non-FDA regulations, 126–146 quality systems to meet, 113–119
Regulatory environment, 84–85 Regulatory information and resources,
FDA, 81–83 Regulatory intelligence, 20–21, 81–82 Regulatory operations, FDA
applications, 84–99 CTD, 91–92, 92f eCTD, 92–96 IND, 89–91 investigational use/marketing
approval, 87–88, 88f marketing applications, 96–99 planning and environment, 84 risk vs. benefit, 84–87
474 Index
Regulatory planning, 20–22, 71–72, 84 elements, 85–86
Regulatory risk identification and management, 86–87
Remedial bacterium, 15–16 REMS. See Risk Evaluation and
Mitigation Strategy (REMS) Reproductive toxicity testing, 359–360 Review-to-revision process, 61 Risk assessment, 321–323, 322f
management and project, 51–52, 51f Risk-based approach, quality
systems, 190 Risk Evaluation and Mitigation Strategy
(REMS), 108, 407–408 Risk management, 27, 49, 52 Risks impact product development, 49 Risk-to-benefit
balance, 86 evaluations, 27
r-protein. See Recombinant protein (r-protein)
Rulemaking process, 71, 105 Rule of thumb, 159, 279
S
S-1, QC cycle drawing, 281 Saccharomyces cerevisiae, 213 SAEs. See Serious AEs (SAEs) Safety assessment, biopharmaceutical
acute toxicity testing, 351–352, 354–355
carcinogenicity testing, 360–361 genetic toxicology, 363, 366–367 immunotoxicology, 361–363 nonclinical safety testing, 351–353 nonclinical study design, elements,
347–351 program, 337–340 protocols and performance, 346–347 subchronic and chronic toxicity
testing, 352, 356–359 tissue binding/local tissue tolerance,
367–368 toxicology, 336–337 in vitro screening test, 340–342 in vivo safety testing, 342–346
Safety topics (S), 127–128 Safety training program, 140 Sampling protocols, 287 Scale-up biomanufacturing, 204–205 Screening log, 380 SDS-PAGE. See Sodium dodecyl sulfate
PAGE (SDS-PAGE) SEC. See Size exclusion chromatography
(SEC) Seizures, FDA judicial actions, 125 The select agents and public health
security and bioterrorism act of 2002, 138–139
Serious AEs (SAEs), 387, 389 Single rising dose study, 402f Six sigma, 191 Size exclusion chromatography (SEC),
228, 275, 292 Skin tissue production, 235–236, 235f Sociotechnical endeavor, 37 Sodium dodecyl sulfate PAGE
(SDS-PAGE), 275, 283, 289–290, 306–307
Somatic tissues/cells, 73 SOPs. See Standard operating
procedures (SOPs) Source information/source
document, 395 Sponsor–FDA communication process,
87, 88f Stability
protocol, 304, 305t, 306t testing, 174
product, 302–308 Stakeholders, 37–39, 61 Standard operating procedures (SOPs),
172, 184, 274, 301 Statistical analysis, 191, 317, 318f Statistical trend analysis, 152 Stem cell
cellular products production from, 236–238
development, 143 technology, 73
Sterile filtration, BS, 249 Sterility testing, 280, 286–287, 307
QC testing, 286–287 USP, 287
475Index
Stopping rules, 383–384, 401 Structured system, 190 Subacute toxicity testing, 356 Subchronic toxicity testing, 352,
356–359
T
Tangential flow filtration (TFF), 223, 224f, 225–226
Targeted product profile (TPP), 10–16, 9f, 29, 87, 99, 377
contraindication, 14 draft, 11 elements, 11–12 preparation, 16 warnings and precautions, 14
Team dynamics, 46–49, 48f Teamwork, 11 TeGenero, 363
issues, challenges, and lessons, 364–366
Teratogenicity toxicity testing, 359–360
Terminal sterilization, 251 Test article, 345 TFF. See Tangential flow filtration (TFF) Therapeutic
confirmatory, 376, 406–407 effect, 65, 79, 325, 330–331 exploratory, 376 food supplements as, 80 monoclonal antibodies, 76, 325, 359 protein, 229, 351, 360 recombinant protein, 76, 265, 359 use, 376
Threshold effect, 405 Tissue binding, 367–368 Tongue depressor, 77 Total protein assay, 275 Total quality management (TQM), 190 Toxicity testing, 339
acute, 351–352, 354–355 chronic, 356–359 developmental, 359–360 reproductive, 359–360 subacute, 356 subchronic, 352, 356–359
Toxicology, 336–337, 375–376 animal model selection for, 344 chronic, 359 developmental, 342 genetic, 363, 366–367 testing, 340
developmental, 353 reproductive, 352
The Toxic Substances Control Act (TSCA), 142–143
TPP. See Targeted product profile (TPP) TQM (total quality management), 190 Tracking, PM, 53, 62 Transfection method, 214, 215f Transgenic animals/plants, 238–245 Transgenic goat, 240, 241f
as bioreactor, 242 Transportation community, 132 Treatment IND, FDA, 102 Trend analysis, 178, 311, 311f, 318, 318f TSCA (the Toxic Substances Control
Act), 142–143 Tween 80, 345, 349
U
United States Pharmacopeia (USP), 75, 271–273
sterility test, 287 validation, 312
Upstream fermentation, material list, 206
Upstream manufacturing, 278–279 Upstream process, 196
recombinant proteins, 218–222 by bacterial/yeast cell
fermentation, 218–220 by mammalian/insect cell
culture, 220–221 recovery, 221–222
Upstream production, 24, 205–206, 279 U.S. Department of Agriculture
(USDA), 79, 120, 134–135, 142–143, 152
U.S. Department of Health and Human Services (DHHS), 67
U.S. Department of Transportation (DOT), 132
476 Index
U.S. FDA regulations (21 CFR), 180 U.S. Pharmacopeia National Formulary
(USPNF), 82
V
VAI (voluntary action indicated), 122–124
Validation, 259–261, 312–317 assay, 312, 317–318 master plan, 260 protocols, 260
Vendor audit, 185 Virus, 232–234, 279, 296–297
production and preparation, 233f Voluntary action indicated (VAI), 122–124
W
Water for injection (WFI), 257 Waxman–Hatch Act of 1984, 76 WBS. See Work breakdown structure
(WBS)
WCB. See Working cell banks (WCB) Western blot test, 274, 283 WFI (Water for injection), 257 WHO. See World Health Organization
(WHO) Woodcock, Janet, 196 Work breakdown structure (WBS), 37,
42–45 draft, 61 project management tool, 43f,
44f, 45f Working cell banks (WCB),
216–217, 234 World Health Organization (WHO), 128,
132, 145
Y
Yeast cell expression systems, 213
Z
Zarxio (filgrastim-sndz) drug, 104
- Cover
- Half Title
- Title Page
- Copyright Page
- Dedication
- Table of Contents
- Preface
- Acknowledgments
- Authors
- 1: Introduction to Biotechnology Operations: Planning for Success
- Biotechnology Operations
- Marketing, Financial, and Business Considerations for Development
- Product Development Planning
- Rationale for Product Development Planning
- The Targeted Product Profile
- The Product Development Plan
- Clinical Development Planning
- Project Management Planning
- Regulatory Planning
- Nonclinical Planning
- Biomanufacturing Planning
- Quality Control Planning
- Quality Systems and Quality Assurance Planning
- Additional Elements of Product Planning
- Summary of Planning for Success
- 2: Project Management
- Biotechnology and Project Management
- Background of Project Management
- Project Management Plan
- The Project Management Environment
- Project Objectives and Schedules
- Sociotechnical Considerations
- Participants in Project Management
- Project Management in Biotechnology Operations
- Establishing Project Management
- The Work Breakdown Structure
- Forming a Project Team and Hands-on Project Management
- Team Dynamics
- Communication and Feedback
- Project Risk Assessment and Management
- Metrics and Tracking Progress
- Resources: Planning and Usage
- Human Factors in Project Management
- Project Completion
- Project Management with Contracts and Collaborations
- Virtual Teams
- Tools for Effective Project Management
- Summary of Project Management in Biotechnology Development
- 3: Regulatory Affairs
- The U.S. Food and Drug Administration: Law and Regulations for Biopharmaceuticals
- Historical Basis for FDA Regulation
- Regulatory Organization of the FDA
- Food and Drug Law, Regulation, and Guidance
- FDA-Regulated Products
- Biologics
- Drugs
- Medical Devices
- Combination Products
- Other Classes of Biotechnology Products and Their Review at the FDA
- Products for Veterinary Use
- Cosmetics, Food, Dietary Supplements, Homeopathic, or Nutritional Products
- FDA Regulatory Information and Resources: Regulatory Intelligence
- Regulatory Operations for FDA Applications
- Regulatory Planning and the Regulatory Environment
- Risk Versus Benefit
- Applications Seeking FDA Investigational Use or Marketing Approval
- Investigational Use Applications. The Investigational New Drug Application
- Common Technical Document
- Electronic Submission of a CTD
- Marketing Applications: BLA and NDA
- Medical Device Applications. 510(k) and PMA
- Special Documents, Pathways, or Exemptions
- Generic Drugs and Biosimiliar or Follow-on Biologics
- Other Regulatory Activities
- Public Meetings and Advisory Committees
- Postmarketing Requirements and Activities
- Advertising and Promotion
- Summary of Regulatory Affair Activities in Biotechnology Operations
- References
- 4: Regulatory Compliance
- Regulatory Compliance
- Quality Systems to Meet Regulatory Compliance
- Compliance and Quality Systems
- Current Good Manufacturing Practices for Manufacture and Quality Control
- Current Good Laboratory Practices for Nonclinical Laboratory Studies
- Current Good Clinical Practices for Clinical Studies
- Compliance for Biopharmaceuticals: Other Regulations of Importance
- Compliance for Import of Biopharmaceuticals into the United States
- Compliance for Medical Devices
- Inspection and Enforcement
- Inspections
- Enforcement Actions
- Product Liability
- Compliance with Non-FDA Regulations: International, National, State, and Local
- International and Foreign National Regulatory Authorities for Medical Biotechnology Products
- Transporting Infectious or Otherwise Hazardous Materials
- Importing, Possessing, or Transferring Controlled Biotechnology Materials
- The Public Health Security and Bioterrorism Preparedness and Response Act of 2002
- Importation or Exportation of Biotechnology Products for the Purpose of Treatment of Diseases in Humans
- Occupational Health and Safety
- Environmental Regulations in Biotechnology
- Genetically Modified Organisms or Molecules
- International Diligence in Biotechnology Operations
- Summary of Regulatory Compliance
- Summary of Non-FDA Compliance
- References
- 5: Quality Systems
- Overview of Quality in Biotechnology
- History: Evolution of Quality Concepts and Practices
- Quality Systems Approach to Product Development
- Planning a Quality System
- Defining Objectives and Ensuring Management Support
- The Quality Manual
- The Quality Plan
- Hallmarks of Quality: Fundamental Criteria for Building Effective Quality Systems
- Management Responsibility
- Defined Quality System
- QbD and Design Control
- Quality by Design
- Design Control
- Design Change
- Contractor, Vendor, and Consultant Control
- Product Identification and Traceability
- Process Control
- Environmental Controls
- Inspection or Testing (Quality Control)
- Release of Material, Service, or Product
- Change Control and Corrective or Preventive Actions
- Packaging and Labeling
- Preservation, Storage, and Handling
- Servicing
- Customer Concerns and Adverse Event Reports
- Document Control
- Training
- Auditing
- The Quality Assurance Unit
- Manage the Quality Assurance Function
- Control Documents and Manage the Documentation System
- Investigate Situations: Manage and Control Change
- Ensure Qualified and Trained Staff
- Perform Audits
- Initiate a Quality System for a Biotechnology Operation
- Unique and Effective Approaches to Quality Management
- Risk-Based Approaches to Quality Systems
- Total Quality Management
- Six Sigma
- Statistics in Quality Assurance
- Quality Systems for Research
- Resolving Quality Issues or Problems
- Summary of Quality Systems
- References
- 6: Biomanufacture
- Overview of Biomanufacturing Requirements
- Design in Biomanufacture
- Technical Considerations for Biomanufacture
- Phases and Scale-up: The Biomanufacturing Life Cycle
- Raw Material Considerations
- Compliance and Quality in Biomanufacture: Current Good Manufacturing Practices
- Biomanufacturing Processes for Biotechnology Products
- Expression of Recombinant Proteins and Nucleic Acids
- Production of Recombinant Molecules from Expression Vectors
- Genes, Vectors, and Host Cells
- Bacterial Cell Expression Systems
- Yeast Cell Expression Systems
- Mammalian or Insect Cell Expression Systems
- Production of Master Cell Banks and Working Cell Banks
- Biomanufacture of Recombinant Proteins
- Planning Production of a Recombinant Protein
- Upstream Process: Production by Bacterial or Yeast Cell Fermentation
- Upstream Process: Production by Mammalian or Insect Cell Culture
- Upstream Process: Recovery
- Downstream Process: Purification
- In-Process Testing and Analysis of Bulk Substance
- Production of Bacterial Plasmid DNA
- Production of Live Recombinant Organisms: Bacteria and Virus
- Production of Products Composed of Mammalian Somatic Cells or Tissues
- Production of Cellular Products Derived from Pluripotent (Stem) Cells
- Production of Biological Molecules by Transgenic Animals or Plants
- Production of Biologically Active Lipids, Glycolipids, and Complex Carbohydrates
- Production of Biologically Active Peptides
- Production of Combination Products: Biopharmaceutical with a Drug or Medical Device
- FP: Formulation, Fill, Finish, and Labeling
- Biomanufacturing Facilities, Utilities, and Equipment
- Facility Design Considerations
- The Facility and Utilities: A Controlled Environment
- Operation of Clean Work Areas for Biomanufacture
- Biomanufacturing Equipment
- Contract Manufacturing Options
- Validation of Biomanufacturing Facilities, Utilities, Equipment, and Processes
- Summary of Biomanufacture
- References
- 7: Quality Control
- Quality Control Overview
- Definition of Product Attributes
- Analytical Methods to Measure Attributes
- Traits of Analytical Methods
- Drafting a Certificate of Analysis (Bulk Substance)
- Selection of Analytical Methods
- Development of Specifications
- Entering Test Results
- Certificate of Analysis for Drug Product
- In-Process Testing
- Analytical Methods
- Additional Analytical Tools and Concepts
- Quality Control of Cell Banks
- Samples and Sampling
- Analytical Controls and Reference Standards
- Test Failures, Out-of-Specification Results, and Retesting
- Testing for Product Stability
- Quality Control Testing of Raw Materials
- Quality Control and the Manufacturing Environment
- Qualification, Validation, and Verification of Analytical Methods
- Application of Statistics in Assay Performance and Validation
- Summary of Quality Control
- Reference
- 8: Nonclinical Studies
- Nonclinical Studies and Risk Assessment
- Biopharmaceutical Delivery, Pharmacokinetics, and Pharmacodynamics
- Product Delivery to the Body
- Adsorption, Distribution, Elimination, and Metabolism (ADME)
- Absorption
- Distribution
- Metabolism and Biotransformation
- Excretion
- Pharmacokinetics and Pharmacodynamics
- Application of Pharmacokinetics and Pharmacodynamics in Biopharmaceutical Development
- Safety Assessment of Biopharmaceuticals
- Toxicology
- Design of a Safety Assessment Program
- In Vitro Screens: Surrogate Measures of Toxicity
- In Vivo Safety Testing of Biopharmaceuticals
- Animal Model Development
- Test Product Formulations, Routes of Delivery, and Dosing Designs
- Protocols and Performance of Biopharmaceutical Safety Studies in Animals
- Elements of a Nonclinical Study Design
- Nonclinical Safety Testing
- Acute Toxicity Testing
- Subchronic and Chronic Toxicity Testing
- Reproductive, Developmental, and Teratogenicity Toxicity Testing
- Carcinogenicity Testing
- Immunotoxicology
- Genetic Toxicology
- Tissue Binding or Local Tissue Tolerance
- Quality of Nonclinical Studies: Current Good Laboratory Practices
- Summary of Nonclinical Studies
- Reference
- 9: Clinical Trials
- Introduction to Clinical Trials
- Background of Clinical Research
- Introduction
- Historical Information on Clinical Trials
- Organization of Clinical Research
- Phases of Clinical Trials
- The Science of Clinical Research
- Quality in Clinical Research and Current Good Clinical Practices
- Clinical Development Planning
- Infrastructure for a Clinical Trial: Individuals, Documents, and Investigational Product
- Design of Clinical Trials and the Clinical Protocol
- Human Subjects, Patients, and Volunteers
- The Sponsor
- The Principal Investigator and His or Her Study Staff
- Institutional Review Boards, the Process of IC, and IC Form
- Investigational Product
- Collection of Clinical Data: Case Report Forms and the Patient Diary
- Clinical Testing Laboratories
- Reporting Results of Clinical Trials: Clinical Summary Reports
- Clinical Trial Operations
- Activities Leading to a Clinical Trial
- Phase 1 Clinical Trial: First-In-Human Study
- Clinical Pharmacology Studies of Biopharmaceuticals in Human
- Phase 2 Clinical Trial: Proof-of-Concept Study
- Phase 3 Clinical Trial: Therapeutic Confirmatory
- Phase 4 Clinical Study and Risk Evaluation and Mitigation Strategy
- Clinical Trials for New Populations or Indications
- Global Clinical Trials
- Quality Systems for Clinical Trials: Current Good Clinical Practices
- Quality and cGCP in Clinical Trial Operations
- Integrity of Clinical Study Data and Documents
- Monitoring and Auditing Clinical Trials
- Ethical Behavior and the Well-Being of Clinical Trial Subjects
- Summary on Clinical Trials
- Reference
- Additional Readings
- Glossary
- Appendix
- Index
