Effect of Saxagliptin and Dapagliflozin on Endothelial Progenitor Cell in Patients With Type 2 Diabetes Mellitus

The Investigator hypothesize that Dapagliflozin will improve EPC number and function AND
Saxagliptin in addition to Dapagliflozin may have an additive effect to improve EPC number
and function even more than Dapa alone, compared to placebo.

In this proposal the investigator plan to conduct a placebo matched study with type 2
diabetes subjects on any doses of metformin or Insulin or a combination of both and has no
history of DPP4 ( Dipeptidyl Peptidase-4) DPP4 inhibitor, incretin mimetic or SGLT2 inhibitor
intake history. Participants will have known macrovascular complications (such as
Cardiovascular Disease (CVD), Cerebrovascular Accident (CVA), and Peripheral Vascular Disease
(PVD).

3 STUDY OBJECTIVES

PRIMARY OBJECTIVE:

CELLULAR BIOMARKER OF ENDOTHELIUM

The primary objective is to ascertain if 16 weeks of Dapa or Dapa+Saxa Combo therapy will
improve :

CD34+ cell number, CD34+ migratory function and CD34+ gene expression in type 2 diabetes with
CVD.

SECONDARY OBJECTIVE:

ARTERIAL STIFFNESS AND RENAL FUNCTION, NON-CELLULAR MARKERS OF ENDOTHELIUM To determine
whether use of Dapa or Dapa+Saxa Combo alters markers of endothelial function such as:
arterial stiffness measures (via tonometry), biochemical measures derived from plasma,
pertaining to endothelial function (hs-CRP, IL-6, TNF-alpha), renal function such as
proteinuria (microalbumin/creatinine ratio) and urine exosome study to determine podocyte
health. The secondary measures are indirect measures of endothelial inflammation in early
type 2 diabetes patients.

Effect on Arterial Stiffness:

I. Pulse Wave Analysis and Vascular Flow will be assessed using SphygmoCor CP system from
ATCOR as a measure of central arterial pressure and arterial stiffness.

II. Vessel health will be assessed by degree of arterial stiffness, using arterial tonometry.

III. The central and the aortic pressure is assessed by pulse wave analysis (PWA) and pulse
wave velocity (PWV).

Effect on Blood Biochemistry:

The Investigator believes cell based biomarkers are superior to traditional serum and plasma
biomarkers and the outcome report will be stronger if one can show positive correlation
between the two outcome measures. The Investigator therefore will be looking at:

I. Inflammation, apoptosis and anti-oxidant protein levels: Highly selective C-reactive
protein (hs-CRP), IL-6, TNF-alpha.

II. Plasma SDF1 alpha (ELISA) and GLP-1 and Ghrelin (ELISA) will be estimated to assess
endothelial health and factors that may influence CD34+ cell chemotaxis III. Podocyte health
via urine exosome analysis. IV. The glomerular filtration rate (GFR) will be estimated by
MDRD equation.

a. GFR = 141 X min (Scr/κ,1)α X max(Scr/κ,1)-1.209 X 0.993Age X 1.018 [if female] X 1.159 [if
African American]; where Scr is serum creatinine (mg/dL), κ is 0.7 for females and 0.9 for
males, α is -0.329 for females and -0.411 for males, min indicates the minimum of Scr/κ or 1,
and max indicates the maximum of Scr/κ or 1.32

TERTIARY OBJECTIVE:

METABOLISM MARKERS The tertiary objective is to determine whether use of Dapa or Dapa+Saxa
Combo alters body composition, fasting lipid profile, and levels of insulin, glucose, and
appetite controlling hormones.

Effect on Blood Biochemistry:

The Investigator believes cell based biomarkers are superior to traditional serum and plasma
biomarkers and the outcome report will be stronger if one can show positive co-relation
between the two outcome measures.

I. Fasting glucose, and insulin. a. Glycemic control will be evaluated by measuring fasting
blood glucose, insulin levels and HbA1c. Fasting blood glucose, insulin and lipid profile
will be used to assess insulin resistance.28,31 II. Lipid profile III. Appetite controlling
hormones via LabCorp: Leptin, Adiponectin IV. Appetite controlling hormones, via ELISA: GLP1,
Ghrelin

Effect of Dapa and Dapa+Saxa Combo on Body Habitus (Determination of body composition and
visceral fat) The Investigator plans to study cardio-metabolic effect of Dapa and Dapa+Saxa
Combo.

I. Using body composition scale:

1. Height and weight will be measured and the body mass index (BMI=kgm2) used as an
indicator of relative weight.

2. The body composition scale calculates body fat%, total body water%, fat free mass, etc.,
in addition to BMI.

The secondary outcome markers (arterial stiffness and renal outcome measures) and
tertiary outcome markers (serum biochemistry) are crucial in order to corroborate the
cellular findings with currently accepted clinical efficacy outcome measures such as
arterial stiffness and serum biochemistry. This design is similar to our recently
published manuscript on Saxagliptin and cellular outcome measures.

4 INVESTIGATIONAL PLAN

STUDY DESIGN AND DURATION

+/- 6 day window for visits

*Assessed at week 0, 8 and 16: Primary, Secondary & Tertiary Outcomes.

Week 20: A telephone call to subjects will be made 4 weeks after last dose of study
medication to determine if there have been any adverse events.

Inclusion Criteria:

1. Able to understand the study, and provide a signed & dated informed consent.

2. Diagnosis of Type 2 diabetes mellitus using criteria of the American Diabetes
Association.

3. 30-70 years old.

4. HbA1C 7 to 10%, both inclusive

5. BMI of 25 - 39.9 kg/m2 both inclusive.

6. Taking a stable dose (for 12 weeks) of Metformin (any dosage) and/or Insulin (any
dosage) for the treatment of T2DM

7. Patients with current Cardiovascular Disease (CVD) in tye 2 diabetes patients, defined
by ≥ 1 of the following:

1. MI >2 months prior

2. Multivessel CAD

3. Angina (intermittent or chronic)

4. Single vessel CAD with positive stress test or UA hospitalization in prior year

5. UA >2 months prior and evidence of CAD

6. Stroke >2 months prior

7. Occlusive PAD

8. Proteinuria of more than 30mg/dl

Exclusion Criteria:

1. Planned CV surgery or angioplasty in 1 month

2. Prior surgery with chronic malabsorption (eg, bariatric) in prior 1 year

3. Diagnosis of Type 1 diabetes mellitus

4. History of GAD antibody positive status

5. Uncontrolled Inflammatory Disease/Inflammatory drug use. **Evaluated by PI on
case-by-case basis**

6. Recent history of diabetic keto-acidosis in the past 3months, or recurrent history of
diabetic ketoacidosis (≥ 3 times)

7. Active bladder cancer

8. Active wounds (e.g. Diabetic ulcers) or recent surgery within 1 month

9. Untreated hyper/hypothyroidism

10. Women of child bearing potential who are not willing to use a contraceptive method to
avoid pregnancy for the 16 weeks of study duration

11. Women who are pregnant or breastfeeding

12. Implanted devices (eg. Pacemaker) that may interact with Tanita scale

13. Any other clinical condition that would jeopardize patients safety while participating
in this clinical trial Concomitant Medications

14. Taking any other oral anti-diabetic agent other than Metformin and/or Insulin for
their treatment of T2DM

15. Beginning statin medications in the past 1 month or change in statin dose in the past
1 month

16. Use of consistent long-term steroid medication (oral, inhaled, injected) within the
last 1 month

17. Treatment with a strong cytochrome P450 3A4 (CYP34A) or P-gp inducer (ie. Rifampin)

18. Subjects with a history of any serious hypersensitivity reaction to Dapagliflozin /
Saxagliptin or another SGLT-2 inhibitor/ DPP4 inhibitor Laboratory Findings

19. Uncontrolled hyperglycemia, defined as a fasting glucose >240 mg/dL (>13.3 mmol/L) at
screening.

20. Liver disease with ALT, AST or ALP x3 ULN

21. eGFR < 60 mL/min/1.73 m2 by MDRD equation in the past 3 months

22. Clinically significant RBC disorders such as hemoglobinopathies

23. Serum creatinine levels ≥1.8 mg/dL with estimated eGFP < 60 mL/min

24. Triglycerides > 450 mg/dL

25. Baseline Hematuria (judged by a urinalysis dipstick at screening) Social History

26. Active smokers

27. Chronic or persistent alcohol or drug abuse

28. Prisoners or subjects who are involuntarily incarcerated

29. Subjects who are compulsorily detained for treatment of either a psychiatric or
physical (eg. infectious disease) illness

30. Participation in another trial with an investigational drug within 30 days prior to
informed consent