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gene_therapy_mine.pptx
GENE THERAPY
RAJITHA ROY
WHAT IS GENE THERAPY ?
Definiton: an experimental technique for correcting defective genes that are responsible for disease development
The most common form of gene therapy involves inserting a normal gene to replace an abnormal gene
Other approaches used:
Replacing a mutated gene that causes disease with a healthy copy of the gene.
Inactivating, or “knocking out,” a mutated gene that is functioning improperly.
Introducing a new gene into the body to help fight a disease.
Researchers are studying gene therapy for a number of diseases, such as
Severe combined immuno-deficiencies (SCID)
Hemophilia
Parkinson's disease
Cancer
HIV
HISTORY AND DEVELOPMENT OF GENE THERAPY
1960: The concepts of Gene Therapy was introduced
1970: Friedmann and Roblin author of a paper in Science titled "Gene therapy for human genetic disease?” cite the first attempt to perform gene therapy
1990:
The first approved gene therapy case at the National Institute of Health, U.K. It was performed on a four year old girl named Ashanti DaSilva. It was a treatment for a genetic defect that left her with an immune system deficiency
New gene therapy approach repairs errors in messenger RNA derived from defective genes. This technique has the potential to treat the blood disorder Thalassaemia, Cystic fibrosis, and some cancers
Sickle cell disease is successfully treated in mice
1992: Doctor Claudio Bordignon working at the Vita-Salute San Raffaele University, Milan, Italy performed the first procedure of gene therapy using hematopoietic stem cells as vectors to deliver genes intended to correct hereditary diseases
1999: Death of Jesse Gelsinger in a gene-therapy experiment resulted in a significant setback to gene therapy research in the United States
2006: Scientists at the National Institutes of Health (Bethesda, Maryland) have successfully treated metastatic melanoma in two patients. This study constitutes one of the first demonstrations that gene therapy can be effective in treating cancer.
2007- 2011: Research is still ongoing and the number of diseases that has been treated successfully by gene therapy increases.
Retinal disease
Colour blindness
Adrenoleukodystrophy
2011: Medical community accepted that it can cure HIV as in 2008, Gero Hutter has cured a man from HIV using gene therapy
TYPES OF GENE THERAPY
GERM LINE GENE THERAPY
SOMATIC GENE THERAPY
GERM LINE GENE THERAPY
Result in permanent changes.
Potential for offering a permanent therapeutic effect for all who inherit the target gene.
Possibility of eliminating some diseases from a particular family.
Also raises controversy:
Some people view this type of therapy as unnatural, and liken it to "playing God”.
Others have concerns about the technical aspects.
SOMATIC GENE THERAPY
Affects only the targeted cells in the patient, and is not passed to future generations.
Short-lived because the cells of most tissues ultimately die and are replaced by new cells.
Transporting the gene to the target cells or tissue is also problematic.
Appropriate and acceptable for many disorders, including cystic fibrosis, muscular dystrophy, cancer, and certain infectious diseases.
Types of somatic gene therapy
Ex vivo
cells are modified outside the body and then transplanted back in again
called ex vivo because the cells are treated outside the body
In vivo
genes are changed in cells when the cells are still in the body
called in vivo because the gene is transferred to cells inside the patient’s body
BASIC PROCESS OF GENE THERAPY
VIRAL VECTOR
NON VIRAL VECTOR
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GT utilizes the delivery of DNA into cells, which can be accomplished by a number of methods.
The two major classes of methods :
recombinant viruses – VIRAL VECTOR
naked DNA or DNA complexes – NONVIRAL VECTOR
Viruses have evolved a way of encapsulating and delivering their genes to human cells in a pathogenic manner. Scientists have tried to harness this ability by manipulating the viral genome to remove disease-causing genes and insert therapeutic ones .
VIRAL VECTOR
Virus bind to their hosts and introduce their genetic material into the host cell.
Plausible strategy for gene therapy, by removing the viral DNA and using the virus as a vehicle to deliver the therapeutic DNA.
The viruses used are altered to make them safe, although some risks still exist with gene therapy.
VIRUS
Many GT clinical trials rely on retroviruses or adenoviruses to deliver the desired gene.
Other viruses used as vectors include adeno-associated viruses, lentiviruses, pox viruses, alphaviruses, and herpes viruses.
Differ in how well they transfer genes to the cells they recognize and are able to infect, and whether they alter the cell’s DNA permanently or temporarily
TYPES OF VIRUS
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How It Works
A vector delivers the therapeutic gene into a patient’s target cell
The target cells become infected with the viral vector
The vector’s genetic material is inserted into the target cell
Functional proteins are created from the therapeutic gene causing the cell to return to a normal state
Picture
Viruses
Replicate by inserting their DNA into a host cell
Gene therapy can use this to insert genes that encode for a desired protein to create the desired trait
Four different types
Created double stranded DNA copies from RNA genome
The retrovirus goes through reverse transcription using reverse transcriptase and RNA
the double stranded viral genome integrates into the human genome using integrase
integrase inserts the gene anywhere because it has no specific site
May cause insertional mutagenesis
One gene disrupts another gene’s code (disrupted cell division causes cancer from uncontrolled cell division)
vectors used are derived from the human immunodeficiency virus (HIV) and are being evaluated for safety
Retroviruses
Adenoviruses
Are double stranded DNA genome that cause respiratory, intestinal, and eye infections in humans
The inserted DNA is not incorporate into genome
Not replicated though
Has to be reinserted when more cells divide
Ex. Common cold
Adeno-associated Viruses
Adeno-associated Virus- small, single stranded DNA that insert genetic material at a specific point on chromosome 19
From parvovirus family- causes no known disease and doesn't trigger patient immune response.
Low information capacity
gene is always "on" so the protein is always being expressed, possibly even in instances when it isn't needed.
hemophilia treatments, for example, a gene-carrying vector could be injected into a muscle, prompting the muscle cells to produce Factor IX and thus prevent bleeding.
Study by Wilson and Kathy High (University of Pennsylvania), patients have not needed Factor IX injections for more than a year
Herpes Simplex Viruses
Double stranded DNA viruses that infect neurons
Ex. Herpes simplex virus type 1
Non-viral Options
Direct introduction of therapeutic DNA
But only with certain tissue
Requires a lot of DNA
Creation of artificial lipid sphere with aqueous core, liposome
Carries therapeutic DNA through membrane
Chemically linking DNA to molecule that will bind to special cell receptors
DNA is engulfed by cell membrane
Less effective
Trying to introduce a 47th chromosome
Exist alongside the 46 others
Could carry a lot of information
But how to get the big molecule through membranes?
Problems with Gene Therapy
Short Lived
Hard to rapidly integrate therapeutic DNA into genome and rapidly dividing nature of cells prevent gene therapy from long time
Would have to have multiple rounds of therapy
Immune Response
new things introduced leads to immune response
increased response when a repeat offender enters
Viral Vectors
patient could have toxic, immune, inflammatory response
also may cause disease once inside
Multigene Disorders
Heart disease, high blood pressure, Alzheimer’s, arthritis and diabetes are hard to treat because you need to introduce more than one gene
May induce a tumor if integrated in a tumor suppressor gene because insertional mutagenesis
Are a tool commonly used by molecular biologists to deliver genetic material into cells.
Can be performed in vivo or in vitro.
Viruses have evolved specialized molecular mechanisms to efficiently transport their genomes inside the cells they infect.
Delivery of genes by a virus is termed transduction and the infected cells are described as transduced.
VIRAL VECTOR
Methods of non-viral gene delivery have also been explored using physical (carrier-free gene delivery) and chemical approaches (synthetic vector-based gene delivery).
NON VIRAL VECTOR
Physical approaches, including
Needle injection
Electroporation
Gene gun
Ultrasound
Hydrodynamic delivery
employ a physical force that permeates the cell membrane and facilitates intracellular gene transfer
PHYSICAL METHOD
The simplest method of non-viral transfection. Clinical trials carried out of intramuscular injection of a naked DNA plasmid have occurred with some success; however, the expression has been very low in comparison to other methods of transfection.
NAKED DNA
This success, however, does not compare to that of the other methods, leading to research into more efficient methods for delivery of the naked DNA such as electroporation and the use of a "gene gun", which shoots DNA coated gold particles into the cell using high pressure gas.
CHEMICAL METHODS THAT ENHANCE THE DELIVERY OF GENE THERAPY -lipoplexes & polyplexes-
DNA must be protected from damage & its entry into the cell must be facilitated
Plasmid DNA can be covered with lipids in an organized structure like a micelle or a liposomecomplexed with DNA it is called a lipoplex
3 types of lipids:
anionic (negatively charged)
neutral
cationic (positively charged)
LIPOPLEXES
Initially, anionic and neutral lipids :
-were used for the construction of lipoplexes for synthetic vectors.
-but,there is little toxicity associated with them,
-they are compatible with body fluids
-there was a possibility of adapting them to be tissue specific
-they are complicated turned to the cationic versions.
Cationic lipids, due to their positive charge,
-naturally complex with the negatively charged DNA.
-their charge they interact with the cell membrane
-endocytosis of the lipoplex occurs
-DNA is released into the cytoplasm.
-The cationic lipids also protect against degradation of the DNA by the cell.
In gene transfer into cancer cells, where the supplied genes have activated tumor suppressor control genes in the cell
decrease the activity of oncogenes.
useful in transfecting respiratory epithelial cells, so they may be used for treatment of genetic respiratory diseases such as cystic fibrosis.
Common uses of lipoplexes
Complexes of polymers with DNA are called polyplexes
consist of cationic polymers and their production is regulated by ionic interactions.
large difference compared to lipoplexes is that polyplexes cannot release their DNA load into the cytoplasm,
End= co-transfection with endosome-lytic agents such as inactivated adenovirus must occur (to lyse the endosome that is made during endocytosis, the process by which the polyplex enters the cell)
POLYPLEXES
Cure blindness of inherited condition
Leber’s conginetal amaurosis
- inherited disease caused by an abnormality in a gene called RPE65.
- The condition appears at birth or in the first few months of life and causes progressive worse and loss of vision.
GENE THERAPY CURES BLINDNESS
used harmless viruses
enable access to the cells beneath the retinas of patients
By using a very fine needle
-safe in an extremely fragile tissue and can improve vision in a condition previously considered wholly untreatable.
http://www.youtube.com/watch?v=d_YJZn-ft_Q
HOW IT WORKS??
it significantly improved the weakness of the symptoms such as tremors, motor skill problems, and rigidity
Main- overactive brain region: the subthalamic nucleus should be introduced with gene
that would produce GABA—an inhibitory chemical—then they could potentially quiet that brain region and alleviate tremors.
GENE THERAPY REDUCES PARKINSON’S DISEASE SYMPTOMS
Done with local anesthesia, used a harmless, inactive virus [AAV-2 GAD]
Deliver the GAD gene into patient’s subthalamic nucleus
The gene instructs cells to begin making GABA neurotransmitters to re-establish the normal chemical balance that becomes dysfunctional as the disease progresses
HOW IT WORKS??
Give a chance of a normal life to baby born with genetic disease.
Give hope of healthy life to cancer patient.
For certain disease that do not have any cure except gene therapy, it could save many lives
ADVANTAGES OF GENE THERAPY
The genetic testing, screening and research in finding the availability of certain gene is very controversy.
May increase rate of abortion if prenatal test regarding baby with genetic disease is done.
The cost is very high and the patient might need an insurance to cover the treatment.
Cosmetic industry may monopolized this gene therapy if it is used in enhancing beauty and in vanishing the aging effect, rather than used for treatment of a disease.
DISADVANTAGES OF GENE THERAPY
ETHICAL QUESTIONS SURROUNDING GENE THERAPY
How can “good” and “bad” uses of gene therapy be distinguished?
Who decides which traits are normal and which constitute a disability or disorder?
Will the therapy only benefit the wealthy due to its high cost?
Could the widespread use of gene therapy make the society less accepting of people who are different?
Should people be allowed to use gene therapy to enhance basic human traits such as height, intelligence, or athletic ability?
BIBLIOGRAPHY
Burdette, Walter J. The Basis for Gene Therapy. Springfield: Charles C Thomas, 2001.
Crayton, Stephanie. “First Clinical Trial Of Gene Therapy For Muscular Dystrophy Now Under Way.” Medical News Today. 1 April 2006. University of North Carolina at Chapel Hill. 11 November 2006 <www.medicalnewstoday.com>.
Gene Therapy. Human Genome Project Information. 18 November 2005. U.S. Department of Energy Office of Science, Office of Biological and Environmental Research, Human Genome Program. 12 September 2006 <http://www.ornl.gov/hgmis>.
McCormack, Matthew P. “Activation of the T-Cell Oncogene LMO2 after Gene Therapy for X-Linked Severe Combined Immunodeficiency.” The New England Journal of Medicine. http://content.nejm.org. 346: 1185-1193, Apr 18, 2002.
Peel, David. “Virus Vectors & Gene Therapy: Problems, Promises & Prospects.” Virus Vectors & Gene Therapy. 1998. Department of Microbiology & Immunology, University of Leicester. 11 November 2006 <http://www.tulane.edu/~dmsander/WWW/335/peel/peel2.html>.
Cavazzana-Calvo, M., et al., Gene therapy of human severe combined immunodeficiency (SCID)-X1 disease. Science, 2000. 288(5466): p. 669-72. Hacein-Bey-Abina, S., et al., Sustained correction of X-linked severe combined immunodeficiency by ex vivo gene therapy. New England Journal of Medicine, 2002. 346(16): p. 1185-1193.
Hacein-Bey-Abina, S., et al., Sustained correction of X-linked severe combined immunodeficiency by ex vivo gene therapy. New England Journal of Medicine, 2002. 346(16): p. 1185-1193
Fischer, A., S. Hacein-Bey, and M. Cavazzano-Calvo. 2002. Gene therapy of severe combined immunodeficiencies. Nature Reviews: Immunology 2:615-621.
What are the ethical issues surrounding gene therapy? - Genetics Home Reference. (2012, November 5). Genetics Home Reference - Your guide to understanding genetic conditions. Retrieved October 31, 2012, from http://ghr.nlm.nih.gov/handbook/therapy/ethics
Walters, L. (1999, February). Human Genome News Vol.10,No.1-2, February 1999. Oak Ridge National Laboratory. Retrieved November 3, 2012, from http://www.ornl.gov/sci/techresources/Human_Genome/publicat/hgn/v10n1/16walter.shtml
U.S. Department of Energy Genome Programs (2011, August 24). Gene Therapy. Oak Ridge National Laboratory. Retrieved November 3, 2012, from http://www.ornl.gov/sci/techresources/Human_Genome/medicine/genetherapy.shtml
Macer, D. R. (1992). Public Acceptance of Human Gene Therapy and Perceptions of Human Genetic Manipulation. Eubios Ethics Institute - HOME. Retrieved November 3, 2012, from http://www.eubios.info/Papers/HGT92.htm
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