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Sitagliptin Improves Plasma Apolipoprotein Profile in Type 2 Diabetes: A Randomized Clinical Trial of Sitagliptin Effect on Lipid and Glucose Metabo- lism (SLIM) Study

Kyoko Tanimura-Inagaki, Mototsugu Nagao, Taro Harada, Hitoshi Sugihara, Shigeki Moritani, Jun Sasaki, Suminori Kono, Shinichi Oikawa, SLIM Study Investigators,

PII: S0168-8227(20)30369-7 DOI: https://doi.org/10.1016/j.diabres.2020.108119 Reference: DIAB 108119

To appear in: Diabetes Research and Clinical Practice

Received Date: 3 August 2019 Revised Date: 2 February 2020 Accepted Date: 10 March 2020

Please cite this article as: K. Tanimura-Inagaki, M. Nagao, T. Harada, H. Sugihara, S. Moritani, J. Sasaki, S. Kono, S. Oikawa, SLIM Study Investigators, Sitagliptin Improves Plasma Apolipoprotein Profile in Type 2 Diabetes: A Randomized Clinical Trial of Sitagliptin Effect on Lipid and Glucose Metabolism (SLIM) Study, Diabetes Research and Clinical Practice (2020), doi: https://doi.org/10.1016/j.diabres.2020.108119

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Sitagliptin Improves Plasma Apolipoprotein Profile in Type 2 Diabetes: A Randomized Clinical Trial of Sitagliptin Effect on Lipid and Glucose Metabolism (SLIM) Study

Kyoko Tanimura-Inagaki1, Mototsugu Nagao1, Taro Harada1, Hitoshi Sugihara1, Shigeki

Moritani2, Jun Sasaki3, Suminori Kono4, Shinichi Oikawa1,5*, ; SLIM Study

Investigators.

1 Department of Endocrinology, Diabetes and Metabolism, Graduate School of Medicine,

Nippon Medical School, Tokyo, Japan

2 Moritani Clinic, Tokyo, Japan

3 International University of Health and Welfare, Fukuoka, Japan

4 MedStat Corporation, Fukuoka, Japan

5 Fukujuji Hospital, Tokyo, Japan

*Correspondence Author:

Shinichi Oikawa

Fukujuji Hospital, 3-1-24 Matsuyama, Kiyose, 204-8522, Tokyo, Japan

Email: shinichi@nms.ac.jp

Short running title: Sitagliptin decrease apo CII, apo CIII, apo E, and apo B-48

Word count for the abstract: 202 words

Word count for the main body of the text: 3403 words

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Number of references: 48

Tables: 4

Figures: 1

Grant Support

The Kidney Foundation, Japan

Conflict of interest

All authors have nothing to disclose as it relates to the content of the current work.

Author contributions

KI: Data analysis, interpretation, and writing and editing of the manuscript; MN, TH, HS,

SM: Patient recruitment and follow-up, data acquisition, and critical revision of the

manuscript; JS, SO: Study design, patient recruitment and follow-up, data acquisition,

analysis and interpretation of data, and critical revision of the manuscript; SK: Study

design, analysis and interpretation of data, statistical analysis, critical revision of the

manuscript and study supervision.

3

Abstract

Aim: This study aims to evaluate the effect of dipeptidyl peptidase-4 inhibitors on lipid

metabolism in patients with type 2 diabetes mellitus (T2D).

Methods: This is a multicenter, open-labeled, randomized controlled study. T2D patients

with HbA1c 6.9–8.9% (52–74 mmol/mol) who were under treatment with sulfonylurea

were randomly allocated to either the sitagliptin group or the non-sitagliptin group.

Glucose and lipid metabolism parameters including apolipoproteins (apo), sterols, and

urinary albumin were assessed at baseline, 3, and 6 months of the treatment.

Results: A total of 164 patients completed the 6-month observation (n=81 for sitagliptin

and n=83 for non-sitagliptin). HbA1c decreased in the sitagliptin group but not in the

non-sitagliptin group. Serum TG and total, LDL and HDL cholesterol levels did not

change in either group. Apo B-48, apo CII, and apo CIII levels decreased in the sitagliptin

group, but not in the non-sitagliptin group. The change in urinary albumin was

significantly different between the groups with a preferable change in the sitagliptin group.

There were no changes in serum sterols levels in the two groups.

Conclusions: The treatment of sitagliptin for 6 months improves the metabolism of

glucose and chylomicron and reduces plasma levels of atherogenic lipoproteins in

patients with T2D.

Keywords

DPP-4 inhibitor, diabetes mellitus, apolipoproteins, sterol, urinary albumin

1. Introduction

Diabetes is a common risk factor for atherosclerotic diseases, and the risk of

coronary events is 2-4 times higher among diabetic patients compared to non-diabetic

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subjects [1-3]. Hyperglycemia and postprandial glycemic stress are specific triggers of

atherosclerosis in humans [4-7] and in animal models [8]. Dyslipidemia is an important

factor for atherosclerosis. Although statins decrease low-density lipoprotein (LDL)

cholesterol levels and prevent atherosclerotic cardiovascular diseases, complete

suppression of cardiovascular events is not attainable by lowering LDL cholesterol alone

[9-11]. Hypertriglyceridemia, therefore, is considered to be a part of the residual risk of

cardiovascular events [12-15].

The triglyceride (TG)-rich lipoproteins (TRLs) such as chylomicron (CM), very

low-density lipoprotein (VLDL), and their remnants are increased with

hypertriglyceridemia. CM particles are synthesized in the intestine and contribute to the

exogenous lipid pathway. Apolipoprotein B-48 (apo B-48) is a structural apolipoprotein

of CM and CM remnant, one apo B-48 protein being on one CM or CM remnant. The

serum level of apo B-48 is a marker of the amounts of CM remnant particles. CM remnant

is known to be atherogenic. Apo B-48 is found to be accumulated in human

atherosclerotic plaque [16]. Fasting apo B-48 concentrations were shown to correlate with

carotid intima-media thickness [17] and coronary artery disease [18].

Type 2 diabetes mellitus (T2D) drives the increase of atherogenic lipoproteins in

fasting and postprandial state. Dyslipidemia occurring in T2D is characterized by the

increase of TRLs and LDL [19] and the emergence of heterogeneous LDL particles as

the qualitative change [20]. Even though serum TG levels are not increased in T2D,

remnant particles are usually increased in relation to hyperglycemia [21, 22]. Remnant is

a therapeutic target for anti-atherosclerosis in T2D [23, 24].

Glucagon-like peptide-1 (GLP-1) is an incretin hormone, which enhances glucose-

dependent insulin secretion, suppresses glucagon secretion, delays gastric emptying and

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promotes satiety [25]. GLP-1 is rapidly degraded by dipeptidyl peptidase-4 (DPP-4).

DPP-4 inhibitors act in delaying the degradation of GLP-1, enhance the physiological

effects of GLP-1, and improves glycemic control. Recently, GLP-1 agonists and DPP-4

inhibitors have been reported to improve not only hyperglycemia but also dyslipidemia.

GLP-1 agonists decreased both fasting and postprandial TG [26] and DPP-4 inhibitors

decreased postprandial TG [27, 28]. DPP-4 inhibitors also decrease postprandial levels

of apo B and apo B-48 levels in patients with T2D [27, 28] and in healthy subjects [29,

30]. Thus, DPP-4 inhibitors decrease postprandial lipid levels. However, the effects of

the DPP-4 inhibitor on fasting lipids remain poorly defined. Although there are few

reports showing the effects of DPP-4 inhibitors on fasting TG, apo B, and apo B-48, such

reports include a limited number of subjects [31, 32]. We designed this current large-scale

study for determining the effect of DPP-4 inhibitor sitagliptin on fasting lipids and

apolipoproteins of uncontrolled T2D with sulfonylurea treatment.

2. Subjects and Methods

2.1. Study Design

The aim of this study is to investigate the effect of sitagliptin on lipid and glucose

metabolism. The trial is registered at UMIN (University Hospital Medical Information

Network) as SLIM Study (ID number UMIN000006511,

http://www.umin.ac.jp/english/). This study is a multicenter, prospective, open-labeled,

randomized controlled study. The study was carried out at 28 hospitals and clinics across

Japan. The patient recruitment and follow-up were carried out from April 2010 to

September 2014.

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Eligible patients were outpatients with T2D aged 20 years or older with HbA1c

between 6.9% (52 mmol/mol) and 8.9% (74 mmol/mol) who were under treatment with

sulfonylurea (SU) monotherapy for at least one month. The range of TG of the eligible

patients was 120-399 mg/dL. All subjects had been under treatment of either

pharmacological or non-pharmacological for T2D for more than 6 months. Patients were

ineligible if they took ongoing treatment with DPP-4 inhibitors, GLP-1 receptor agonists,

insulin, or lipid-lowering drugs (statins, fibrates, nicotinic acids, probucols, anion

exchange resin, eicosapentaenoic acid, docosahexaenoic acid, or ezetimibe). They were

also ineligible if they had chronic renal dysfunction (serum creatinine > 1.5 mg/dL),

severe liver dysfunction, infection, trauma, possible pregnancy, and a history of severe

ketosis or diabetic coma in the past 6 months. The eligible patients were reported to the

registration office and were randomly allocated to either the sitagliptin group or the non-

sitagliptin group. Patients of the sitagliptin group were administered with 50 mg of

sitagliptin once a day in addition to previously prescribed SU, and patients of the non-

sitagliptin group were treated with SU multiplication. Subjects with 7.0 and higher in

HbA1c were additionally medicated with SU or sitagliptin (100 mg once a day) in the

non-sitagliptin group or sitagliptin, respectively, to reduce HbA1c level below 7.0%. No

drugs for dyslipidemia were prescribed during the study period. This study conformed to

the principles outlined in the Declaration of Helsinki and was approved by the ethics

committees of Nippon Medical School Hospital. All patients provided written informed

consent.

2.2. Measurements

We took blood and urine samples under the fasting conditions at 0, 3, and 6 months

of treatment and blood samples were collected to the central laboratory. The blood

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pressure (BP) and body weight were measured at the same time of blood sampling. The

blood was centrifuged to separate serum for measurement of fasting levels of plasma

glucose (FPG), HbA1c, glycated albumin (GA), 1,5-anhydroglucitol (1,5-AG), fasting

serum insulin, serum total cholesterol (TC), TG, HDL cholesterol, serum small dense

LDL (sd-LDL), serum remnant like particle cholesterol (RemL-C), serum lipoprotein

lipase (LPL), serum apolipoproteins (apo AI, AII, B, CII, CIII, E, and B48), serum

creatinine, urinary albumin to creatinine ratio (ACR), and serum sterols (sitosterol,

campesterol, cholestanol, and lathosterol). Samples were analyzed as follows at SRL Inc.,

Tokyo. Apo AI, AII, B, CII, CIII, and E were determined by turbidimetric immunoassay

by using kits for Apo A-I, Apo A-II, Apo B, Apo C-II, Apo C-III, and Apo E Auto·N

"DAIICHI", (Sekisui Medical), respectively. The Autokit Micro Albumin (Fujifilm

Wako Pure Chemical) was used to measure ACR. TC and TG were determined by

cholesterol dehydrogenase UV method and enzymatic method, using Cholestest CHO

and Pure Auto S TG-N (Sekisui Medical), respectively. Pre-heparin LPL mass was

determined by enzyme-linked immunosorbent assay using LPL Elisa Daiichi kit (Sekisui

Medical), HDL-C and sd-LDL were measured by a direct assay using Cholestest N-HDL

(Sekisui Medical) and sd-LDL-EX Siken (Denka Seiken), respectively. LDL cholesterol

and non-HDL-C were calculated by Friedewald formula and the formula: TC  HDL-C,

respectively. Serum creatine, 1,5-AG, RemL-C, and GA were determined by visible

absorption spectrometry enzymatic method, using Detaminer L CRE, Detaminer L CRE

1,5-AG, Metabolead RemL-C (Kyowa medics), and Lucica GA-L (Asahi Kasei Pharma).

Plasma glucose was determined by the hexokinase UV method using Cicaliquid GLU L

(Kanto Kagaku). HbA1c was determined by latex agglutination turbidimetry using

RAPIDIA Auto HbA1c (Fujirebio Inc.) Sterols were assayed with gas chromatography

8

using GC-2010 (Shimazu Co.). Plasma apo B-48 levels were measured by

chemiluminescent enzymatic immunoassay using apo B-48 CLEIA (Fujirebio, Inc) as

previously reported method [33-35].

2.3 Sample size and statistical Analysis

While the primary endpoints were the changes in serum lipids and apolipoproteins,

we focused on the change in serum apo B in determining the sample size. We expected

serum apo B decreases of 6.0mg/dL in the sitagliptin group versus 2.9mg/dL in the SU

group with SD of 6mg/dL in each group based on the past literature [28, 36]. A total of

200 patients were a target of enrollment with allowance for dropouts based on the

calculated required sample size as 80 for each group with a two-sided significance level

of 5% and a power of 80%.

Data management and statistical analysis were performed by a company supporting

clinical research (La Nouvelle Place, Tokyo) using software IMP SPSS version 22. The

between-group comparison was done by the chi-square test for categorical variables and

by unpaired t-test for continuous variables. The paired t-test was used to determine the

within-group difference between baseline and after treatment. P values < 0.05 were

considered statistically significant. The results of statistical analyses were reviewed by a

statistical expert (S.K).

3. Results

3.1. Subject characteristics

The 189 patients were enrolled and randomized to the sitagliptin or non-sitagliptin

group. A total of 25 subjects were excluded because of protocol violation (n = 23), no

visit at baseline (n = 1) and withdrawal of informed consent (n = 1). Finally, 164 patients

9

(n = 81 in the sitagliptin group and n = 83 in the non-sitagliptin group) were analyzed as

shown in Figure 1. The baseline characteristics of the two groups are shown in Table 1

and 2. There were no significant differences at baseline in the clinical characteristics

between the sitagliptin and the non-sitagliptin group other than urinary albumin (p = 0.03),

apo CIII (p = 0.04), and apo E (p = 0.03). Although the difference was not statistically

significant, systolic BP (SBP) showed a higher trend in the sitagliptin group than in the

non-sitagliptin group.

3.2. Effect of sitagliptin treatment on clinical and biochemical parameters

At 3 months of treatment, HbA1c, GA, and 1,5-AG markedly improved in the

sitagliptin group, but not in the non-sitagliptin group (Table 3). Although there were no

changes in lipid and LPL levels, apo AI and apo CIII decreased substantially only in the

sitagliptin group, and the difference in the change of apo CIII was significant (p = 0.003)

between sitagliptin and non-sitagliptin group. SBP and diastolic BP (DBP) decreased

more markedly in the sitagliptin group than in the non-sitagliptin group; showing

significant differences in the BP changes between the two groups. The body weight

increased in the non-sitagliptin group, but not in the sitagliptin group; resulting in a

significant difference between the two groups (p = 0.001).

At 6 months of treatment, the changes in HbA1c, GA, and 1,5-AG from the baseline

were of almost the same magnitudes as observed at 3 months (Table 4). Apo CII and apo

CIII decreased in the sitagliptin group, but not in the non-sitagliptin group. The

differences in the changes of apo CII (p = 0.001) and apo CIII (p < 0.001) were

significantly different between the two groups. Apo B-48 significantly decreased at 6

months of treatment in the sitagliptin group, but not in the non-sitagliptin group; resulting

in a significant difference between the two groups (p = 0.02). A between-group difference

10

remained in the change of body weight, but not in the changes of SBP and DBP. Serum

sterol levels did not change in the two groups at either 3 or 6 months of the treatment.

4. Discussion

This first randomized controlled study examining the effects of sitagliptin on

plasma apolipoproteins in uncontrolled T2D demonstrated that sitagliptin treatment

significantly decreased fasting apo B-48, apo CII, and apo CIII, while sitagliptin did not

affect fasting TG and apo B. The decreases of apo B-48, apo CII, and apo CIII indicate

the improvement of TRLs metabolism.

This study showed a sitagliptin-induced reduction of apo B-48 in a fasting state

without decreasing TG and apo B by sitagliptin treatment. The results of a significant

decrease of apo B-48 contrasting to apo B change might suggest that the intestinal factor rather

than hepatic factor is more influenced by the DPP-4 inhibitor. The previous study showed that

sitagliptin reduced postprandial apo B-48, but not fasting hepatic apo B100 containing

lipoprotein particle production in healthy humans [29].　This result suggested that DDP-

4 inhibitors seem to affect the intestinal production of lipoproteins rather than hepatic

lipoprotein production [37]. GLP-1 is a gut-derived hormone, which secreted from the

intestine at a low basal level in fasting and increased two to threefold at a fed state. GLP-1

increases glucose-dependent insulin secretion, decreases postprandial glucagon levels,

slow gastric emptying, and cause satiety and calorific intake reduction. DPP-4 degrade

GLP-1 rapidly, therefore DPP-4 inhibitor enhances GLP-1 function and improves

glycemic control by contributing several organs. It is demonstrated the postprandial rise

of TG levels in the placebo was completely abolished by GLP-1 in a healthy human study

[38]. GLP-1 agonist exendin-4 decreased TG and cholesterol in VLDL and CM remnant

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sized lipoproteins and the postprandial apo B-48 levels were reduced after fat load. Some

studies showed DPP-4 inhibitor did not affect on TG and apo B levels. In a randomized

double-blind study of T2DM subjects, the change of lipid subfractions was compared

between vildagliptin and placebo. Vildagliptin had no effect on fasting total TG, total

cholesterol or serum apo B, but decreased the area under the curve of total plasma TG

following the test meal. Vildagliptin decreased apo B-48, but not apo B-100 in either the

CM or the VLDL-IDL subfractions [28]. TRL kinetics in respose to sitagliptin in healths

subjects was tested following a high-fat, liquid formul under pancreatic clamp. Sitagliptin

did not affect the plasma concentration or production of hepatic lipoproteins although it

decreased lipoprotein particle of intestinal origin [29].

There are possible mechanisms to be considered for the sitagliptin-induced

reduction of apolipoproteins. Apo B-48 is a constituent of CM and both Apo CII and apo

CIII are constituents of CM, VLDL, LDL, and HDL. Therefore, the reduction of apo B-

48, apo CII, and apo CIII in this study might reflect TRL reduction. The improvement of

glycemic control and the tendency of weight loss by sitagliptin also may affect the

reduction of TRL because the restoration of insulin sensitivity reduces fasting TG [37]

and the combination of weight loss and reduction of insulin resistance cause induction of

LPL activity [39, 40] as well as reduction of TG. Our findings of the reduction of apo

B-48, apo CII, and apo CIII by sitagliptin are supported by the previous observation in a

one-arm trial [32].

Apo CII and apo CIII are not only the structural protein but also have functions

contributing to TG metabolism. In addition to its function as an essential cofactor of LPL

to facilitate the hydrolysis of TG, apo CII inhibits the binding of VLDL to the LDL-

receptor related protein, which causes retention of TRL. It is reported that human apo CII

12

overexpression transgenic mice showed hypertriglyceridemia consistent with a reduction

of LPL mediated hydrolysis of VLDL-TG [41] and, in addition, that the completely apo

CII deficient subjects have chylomicronemia [42]. Thus, apo CII has both positive and

negative effects on TRL metabolism. The reduced level of apo CII observed in this study

would cause a reduction of TRL due to reduced apo B-48 levels. Apo CIII concentrations

were associated with the production of TRLs by facilitating TG assembly to VLDL,

retarding clearance of TRLs by inhibiting hepatic lipase and LPL, and decreasing hepatic

uptake of TRLs [43]. The reduction of apo CIII might improve TRL metabolism by

reducing TG assembly and improving TRL clearance. In addition, insulin resistance is

associated with apo CIII production [44]. The tendency of weight loss in the sitagliptin

group will improve insulin resistance and, in turn, will cause a reduction of apo CIII.

The previous study suggested that increasing of LPL mass and activity by

sitagliptin explained the degradation of TRLs[32]. But our data did not indicate the

significant changes in TG and LPL. It is well known that diabetes mellitus, in which

hyperglycemia and impaired insulin effect by both resistance and/or deficiency, induces

the increase of cholesterol absorption and the impaired lipoprotein degradation. In our

study, cholesterol absorption and synthesis markers had no change, which might be due

to the poor improvement of glycemic control because our subjects were very poor

glycemic control.

It is possible that lipid profile improvement is due to the class effect of DPP-4

inhibitors. A past report showed alogliptin [30], sitagliptin [32], and gemigliptin [45]

reduced apo B-48. However, the answer should be obtained after the comparison study

among various kinds of DPP-4 inhibitors.

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To our knowledge, this is the first report that addressed whether a DPP-4 inhibitor

affects serum sterols in a randomized controlled study. It had been expected that sterols,

markers of cholesterol kinetics, could be altered after the treatment with sitagliptin, but

neither cholesterol absorption (campesterol, sitosterol, and cholestanol) nor synthesis

(lathosterol) markers change in either sitagliptin or non-sitagliptin group. Overall,

sitagliptin, therefore, does not seem to affect cholesterol kinetics rather improve CM

metabolism in T2D.

ACR decreased in the sitagliptin group; thereby suggesting that sitagliptin

treatment is more preferable to improve albuminuria than adding sulfonylurea in patients

with uncontrolled T2D who have already been treated with sulfonylureas. The preferable

effect of DPP-4 inhibitor on urinary albumin excretion has been demonstrated in an open-

labeled prospective randomized study [46] and a multicenter randomized double-blind

placebo-controlled trial[47]. We consider the improvement of albuminuria could be

explained by the reductions of blood glucose levels and SBP by sitagliptin treatment.

This is the first multicenter randomized controlled study that showed sitagliptin

decreased fasting apo B-48 levels in uncontrolled diabetes subjects with SU. There are

some studies that showed DPP-4 inhibitors decreased apolipoproteins. However, there

are some strong points of our study. First, our study design is a randomized SU-controlled

study. Our enrolled subjects were diabetes patients who were already treated with SU.

The previous studies are placebo-controlled [28], placebo-controlled cross over [27, 48],

sitagliptin-one arm study [18]. There is no report that investigated the DPP-4 inhibitor

effect on apolipoprotein in a randomized SU-controlled study. The present study showed

that the addition of DPP-4 inhibitor to SU improved glycemic control and apo B-48 levels,

compared to the SU-controlled group. This finding suggests that the DPP-4 inhibitor is

14

useful in clinical practice to improve glycemic and CM metabolism in diabetes subjects

with SU. Second, our subjects were poorly controlled diabetes subjects. In contrast to it,

some previous studies enrolled healthy [29, 30] or mild diabetes subjects [27, 30, 31].

The diabetes-controlled state is very important to assess drug efficacy on lipid metabolism

because of insulin resistance and insulin secretion effect on lipid metabolism. Our study

showed the efficacy of the DPP-4 inhibitor even under poor-controlled diabetes with SU.

Third, our sample size would be more appropriate because the number of subjects was

set by statisticians depending on the magnitude of the hypothesized effect, the level of

significance desired, and the power desired. The sample number of previous studies that

investigate the DPP-4 inhibitor effect on apolipoproteins was smaller than in our study.

Fourth, our study was the first long term study that investigated the effect of DPP-4

inhibitor on apolipoproteins. The term of this study is 6 months, in contrast, the term of

most of the previous studies was from one day to 12 weeks. Fifth, our study is the first

report of a multicenter study, in contrast, previous studies are single-center studies. Sixth,

this is the first study that investigated the effect of DPP-4 inhibitor on plant sterols. From

these points, our study could give useful and novel information. Our result has shown that

the addition of DPP-4 inhibitor to SU preferably affects apo B-48 metabolism compare

to the addition of SU to the uncontrolled diabetes patients with already receiving SU, and

this result is meaningful in the daily medical care of diabetes for reducing cardiovascular

risk status.

There are several limitations in the present study. Firstly, there left a possibility that

the higher concentrations of plasma apo CIII and apo E and urinary albumin at baseline

in the sitagliptin group compared to the non-sitagliptin group affected in the improvement

of fasting apo B-48, apo CII, apo CIII, urinary albumin, while the patients were randomly

15

allocated into two groups. Secondly, glycemic control was improved only in the

sitagliptin group. Although the exact mechanism to cause this difference is to be

elucidated, we speculate that it can be affected by the improvement of lipoprotein

metabolism or by increasing the dosage of SU for uncontrolled diabetes patients who

suffered already SU in this study subjects. The result indicates that the treatment of

sitagliptin improves glycemic control and the lipoprotein metabolism for uncontrolled

diabetes patients who suffered already SU. Our results provide evidence to support the

notion that sitagliptin would be a suitable treatment option for uncontrolled diabetic

patients. Thirdly, the objectives of this study are only Japanese patients. Therefore, it

would be needed to confirm if the current finding is applicable to patients of other races.

5. Conclusions

Sitagliptin treatment with sulfonylureas improves CM metabolism and reduces

atherogenic lipoproteins accompanied by controlling glycemia in patients with

uncontrolled T2D. The effects of sitagliptin for dyslipidemia would be due to a

rearrangement of remnant metabolism and might affect vascular changes protectively.

Together with the preferable effect on albuminuria, the treatment with DPP-4 inhibitors

could be an ideal approach to prevent both micro- and macrovascular complications in

T2D.

Figure legend;

Fig. 1. Flowchart of subject enrollment

Acknowledgements

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This study was financially supported by The Kidney Foundation, Japan. We thank

all of the members of SLIM study group; Dr. Ishikawa T (Koishikawa Nippori Clinic,

Tokyo), Dr. Itagaki K (Itagaki Clinic, Tokyo), Dr. Okuda T (Okuda Clinic, Tokyo), Dr.

Takeda M (Takeda Clini, Tokyoy), Dr. Tanaka J (Kumanomae Clinic, Tokyo), Dr.

Memezawa H (Memezawa Clinic, Tokyo), Dr. Moriya H (Saito Clinic, Tokyo), Dr.

Yoshiyuki T (Yoshiyuki Clinic, Tokyo), Dr. Kimura K (Hachijyo Hospital, Tokyo), Dr.

Nakajima Y (Nakajima Clinic, Tokyo), Dr. Inazawa K (Kashiwa City Hospital, Chiba),

Dr. Shuto H (Minamikoshigaya-Kennshinkai Clinic, Saitama), Dr. Sasaki T (Sasaki

Clinic, Tokyo), Dr. Inomata Y (Kitakoiwa Ichoka Clinic, Tokyo), Dr. Ooki I (Okada

Clinic, Tokyo), Dr. Yamamura S (Yamamura Clinic, Tokyo), Ageta M (Ageta Clinic,

Miyazaki), Dr. Ikeda Y (Gosho Hospital, Fukuoka), Dr. Uchida Y (Saga Memorial

Hospital, Saga), Dr. Otonari T (Otonari Clinic, Fukuoka), Dr. Kuribayashi T (Koga

General Hospital, Miyazaki), Dr. Kobori S (Takuma Hospital, Kumamoto), Dr. Saikawa

T (Yufuin Kosei Hospital, Oita), Dr. Sawayama Y (Fukuoka Red Cross Hospital,

Fukuoka), Dr. Biro S (Tsukasa Health Care Hospital, Kagoshima), Dr. Yamamoto K

(Takagi Hospital, Fukuoka), and Dr. Fukushima N (Takagi Hospital, Fukuoka).

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Enrollment and randomization n=189

Sitagliptin treatment n=97

Non-sitagliptin treatment n=92

Excluded, n=16 • protocol violation (n=16)

Exluded, n=9 • no visit at baseline (n=1) • informed consent withdrawal (n=1) • protocol violation (n=7)

Analyzed n=81

Analyzed n=83

Fig. 1. Flowchart of subject enrollment

21

Table 1. Baseline characteristics of randomized patients

Sitagliptin (n=81) Non-sitagliptin (n=83)

Age (years) 64 (10) 64 (12)

Weight (kg) 68.3 (14.8) 68.6 (14.3)

Men, n (%) 45 (54) 45 (56)

Systolic BP (mmHg) 137 (11) 133 (12)

Diastolic BP (mmHg) 77 (9) 77 (8)

Serum creatinine (mg/dL) 0.71 (0.2) 0.74 (0.2)

Urinary albumin (mg/g Cr) 133 (306)* 55 (103)

hs-CRP (ng/mL) 2394 (5544) 2180 (5172)

Glucose metabolism parameters

HbA1c (%)

(mmol/mol)

7.5 (0.8)

58 (15)

7.5 (0.7)

58 (16)

Glycated albumin (%) 20.2 (4.0) 20.5 (3.6)

1,5-AG (μg/mL) 6.3 (4.4) 6.4 (4.9)

FPG (mg/dL) 160 (50) 169 (66)

Fasting insulin (μU/mL) 18.3 (37.1) 14.7 (16.0)

Type of sulfonylureas

Glimepiride (%) 88.7 93.3

Glibenclamide (%) 4.4 1.1

Gliclazide (%) 7.2 5.6

Data are expressed as mean (SD) or n (%), unless otherwise indicated. Abbreviations: BP, blood

pressure; HbA1c, glycosylated hemoglobin; 1,5-AG, 1,5-anhydroglucitol; FPG, fasting plasma

glucose; hs-CRP, high sensitive C-reactive protein. *p <0.050 for the between-treatment

difference as determined with use of paired t-tests or χ2 tests.

22

Table 2. Serum lipids and lipoproteins at baseline in the sitagliptin and non-sitagliptin group

Sitagliptin Non-sitagliptin

Total cholesterol (mg/dL) 208 (38) 210 (36)

Triglyceride (mg/dL) 185 (120) 163 (116)

HDL-cholesterol (mg/dL) 52 (14) 53 (13)

LDL-cholesterol (mg/dL) 125 (36) 126 (35)

Non-HDL-cholesterol (mg/dL) 157 (36) 157 (36)

RLP-C (mg/dL) 7.4 (5.1) 6.5 (5.2)

Sd LDL (mg/dL) 47 (19) 45 (19)

LPL (ng/mL) 46 (17) 44 (15)

Apolipoproteins

Apo AI (mg/dL) 146 (27) 150 (28)

Apo AII (mg/dL) 30.2 (4.9) 31.3 (5.9)

Apo B (mg/dL) 109 (24) 108 (25)

Apo CII (mg/dL) 5.1 (2.5) 4.6 (2.0)

Apo CIII (mg/dL) 11.8 (4.7)* 10.4 (3.6)

Apo E (mg/dL) 4.9 (1.5)* 4.5 (1.1)

Apo B-48 (μg/mL) 6.6 (5.2) 5.7 (4.0)

Plant sterols

Campesterol (μg/mL) 5.7 (3.2) 5.9 (2.9)

Sitosterol (μg/mL) 2.9 (2.0) 3.0 (1.9)

Cholestanol (μg/mL) 2.9 (1.0) 3.0 (1.0)

Lathosterol (μg/mL) 3.3 (1.6) 3.7 (2.8)

Data are expressed as mean (SD). Abbreviations: RLP-C, remnant-like particle cholesterol; Sd

LDL, small dense LDL; LPL, lipoprotein lipase; apo, apolipoprotein. *p <0.05 for the between-

treatment difference. Based on unpaired t-test.

23

Table 3. Changes in glucose and lipid parameters at 3 months of treatment from baseline in the

sitagliptin and the non-sitagliptin groups

Variable Sitagliptin Non-sitagliptin Between-

group

Mean (SD) p-value Mean (SD) p-value p-value

Weight (kg) 0.3 (1.7) 0.08 0.4 (1.2) 0.003 0.001

Systolic BP (mmHg) –5.4 (9.6) <0.0001 –1.1 (11.1) 0.36 0.009

Diastolic BP (mmHg) –2.8 (7.4) 0.001 –0.3 (8.3) 0.75 0.04

Urinary albumin (mg/g Cr) –24 (145) 0.15 19 (73) 0.03 0.02

FPG (mg/dL) –8.0 (58.1) 0.22 –0.5 (64.4) 0.48 0.76

HbA1c (%) –0.4 (0.7) <0.00001 0.1 (0.9) 0.32 <0.00001

GA (%) –1.9 (3.0) <0.00001 0.2 (3.4) 0.55 0.00005

1,5-AG (μg/mL) 2.3 (3.8) <0.00001 0.0 (3.6) 0.95 0.0001

Fasting insulin (μU/mL) 0.4 (16.8) 0.83 0.7 (14.8) 0.65 0.89

Total cholesterol (mg/dL) –1.6 (27.3) 0.78 0.0 (25.0) 1.00 0.70

Triglyceride (mg/dL) –4.2 (136) 0.78 2.6 (78) 0.77 0.69

HDL-cholesterol (mg/dL) –1.6 (7.4) 0.05 –0.2 (6.0) 0.78 0.17

LDL-cholesterol (mg/dL) 5.7 (28.3) 0.08 1.2 (23.0) 0.64 0.27

Non-HDL-cholesterol (mg/dL) 0.0 (24.6) 0.99 0.2 (23.1) 0.94 0.97

RLP-C (mg/dL) 0.3 (9.2) 0.77 0.2 (3.7) 0.55 0.96

Sd LDL (mg/dL) –0.7 (14.9) 0.69 –0.8 (12.6) 0.58 0.96

LPL (ng/mL) –0.5 (10.2) 0.65 0.7 (8.6) 0.45 0.40

Apo AI (mg/dL) –5.1 (15.9) 0.005 –1.3 (14.7) 0.43 0.11

Apo AII (mg/dL) –0.5 (3.9) 0.22 –0.7 (3.7) 0.09 0.80

Apo B (mg/dL) –1.5 (14.9) 0.37 –0.9 (14.5) 0.56 0.81

Apo CII (mg/dL) –0.3 (1.6) 0.07 0.1 (1.4) 0.40 0.05

Apo CIII (mg/dL) –1.3 (3.2) 0.0005 0.2 (3.1) 0.58 0.003

Apo E (mg/dL) –0.1 (1.0) 0.35 0.1 (0.9) 0.53 0.26

Apo B-48 (μg/mL) –0.8 (6.7) 0.26 0.2 (4.2) 0.69 0.24

Campesterol (μg/mL) 0.1 (1.9) 0.54 0.1 (1.6) 0.53 0.94

Sitosterol (μg/mL) 0.1 (1.2) 0.31 0.0 (1.0) 0.70 0.60

Cholestanol (μg/mL) 0.0 (0.8) 0.68 0.0 (0.8) 0.71 0.57

Lathosterol (μg/mL) 0.0 (1.6) 0.97 –0.2 (1.3) 0.16 0.37

24

Data are expressed as mean (SD). Abbreviations: BP, blood pressure; FPG, fasting plasma

glucose; HbA1c, glycosylated hemoglobin; GA glycated albumin; 1,5-AG, 1,5-anhydroglucitol;

RLP-C, remnant-like particle cholesterol; Sd LDL, small dense LDL; LPL, lipoprotein lipase;

apo, apolipoprotein.Based on paired t-test and unpaired t-test.

25

Table 4. Change in glucose and lipid parameters at 6 months treatment from baseline in the sitagliptin and the non-sitagliptin groups.

Variable Sitagliptin Non-sitagliptin Between-group

Mean (SD) p-value Mean (SD) p-value p-value

Weight (kg) –0.5 (2.3) 0.07 0.3 (1.7) 0.08 0.01

Systolic BP (mmHg) –3.6 (10.5) 0.002 –1.5 (10.8) 0.20 0.21

Diastolic BP (mmHg) –1.4 (7.4) 0.08 –0.6 (9.0) 0.54 0.53

Urinary albumin (mg/g Cr) –26 (142) 0.10 28 (170) 0.14 0.03

FPG (mg/dL) –8.0 (71.6) 0.32 5.3 (69.6) 0.49 0.23

HbA1c (%) –0.5 (0.8) 0.000 0.1 (1.0) 0.29 0.001

GA (%)

(mmol/mol)

–2.0 (3.1)

–22 (34)

0.000 –0.1 (3.6)

–1 (39)

0.73 0.000

1,5-AG (μg/mL) 2.9 (5.3) 0.000 0.2 (3.8) 0.58 0.0003

Fasting insulin (μU/mL) –3.1 (31.9) 0.38 1.8 (13.7) 0.24 0.20

Total cholesterol (mg/dL) 0.8 (28.3) 0.81 –0.3 (28.8) 0.91 0.81

Triglyceride (mg/dL) –18.2 (93.0) 0.08 4.4 (78.3) 0.61 0.09

HDL-cholesterol (mg/dL) 0.9 (9.1) 0.36 0.0 (7.4) 0.95 0.49

LDL-cholesterol (mg/dL) –0.2 (30.1) 0.96 –1.3 (26.7) 0.66 0.80

Non-HDL-cholesterol (mg/dL) –0.2 (26.4) 0.95 –0.4 (26.7) 0.89 0.96

RLP-C (mg/dL) –0.7 (4.9) 0.22 0.5 (3.6) 0.25 0.09

Sd LDL (mg/dL) –0.2 (14.1) 0.89 –1.0 (14.0) 0.51 0.71

LPL (ng/mL) 0.2 (10.4) 0.89 0.3 (9.5) 0.74 0.91

Apo AI (mg/dL) 0.3 (19.5) 0.89 –1.3 (15.6) 0.46 0.57

Apo AII (mg/dL) 0.2 (3.8) 0.66 –0.1 (3.2) 0.85 0.65

Apo B (mg/dL) –2.3 (16.5) 0.21 –2.0 (17.2) 0.31 0.88

Apo CII (mg/dL) –0.4 (1.3) 0.003 0.2 (1.2) 0.07 0.001

Apo CIII (mg/dL) –1.1 (2.7) 0.0003 0.4 (2.5) 0.14 0.0002

Apo E (mg/dL) –0.1 (0.8) 0.12 0.1(0.9) 0.33 0.08

Apo B-48 (μg/mL) –1.2 (4.5) 0.02 0.5 (3.9) 0.29 0.02

Campesterol (μg/mL) 0.1 (1.9) 0.54 0.1 (1.9) 0.65 0.90

Sitosterol (μg/mL) 0.2 (1.3) 0.17 0.1 (1.1) 0.36 0.63

Cholestanol (μg/mL) 0.1 (0.9) 0.19 0.0 (0.9) 0.63 0.56

Lathosterol (μg/mL) –0.1 (1.5) 0.74 –0.5 (1.6) 0.01 0.09

26

Abbreviations: BP, blood pressure; FPG, fasting plasma glucose; HbA1c, glycosylated hemoglobin, GA, glycated albumin; 1,5-AG, 1,5-anhydroglucitol; RLP-C, remnant-like particle cholesterol; Sd LDL, small dense LDL; LPL, lipoprotein lipase; apo, apolipoprotein. Based on paired t-test and unpaired t-test.