Clinical trial NCT02608905
Conditions: Type 2 Diabetes Mellitus
Sponsor: Baylor College of Medicine
Start date: 2015-11-01; Completion date: 2019-03-01
Enrollment: 17 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Sodium-glucose cotransporter 2 (SGLT-2) inhibitors reduce hyperglycemia and improve peripheral insulin sensitivity by ameliorating glucotoxicity. Insulin resistance in type 2 diabetes (T2DM) is associated with endothelial dysfunction and vascular inflammation. Thus strategies to improve insulin sensitivity and lower glucotoxicity may improve endothelial inflammation and vascular inflammation. However, the effects of these agents on vascular inflammation and endothelial function is not known in patients with type 2 diabetes although anti-inflammatory properties have been demonstrated in various animal models. In the present study the investigators will assess if dapagliflozin treatment for 12 weeks decreases monocyte inflammation and improves endothelial function in patients with type 2 diabetes on metformin monotherapy.

Detailed description:
The insulin-resistant state of type 2 diabetes mellitus (T2DM) is largely mediated by inflammatory pathways affecting skeletal muscle which is the primary site of whole body insulin resistance. Nuclear factor kappa B (NFkappaB) regulates pro-inflammatory cytokines which ultimately impair skeletal muscle insulin signaling and fatty acid oxidation; its activity reflects overall inflammatory tone in skeletal muscle. Recent human studies confirm that NFkappaB is elevated in the skeletal muscle of T2DM human subjects. Furthermore, the same inflammatory processes and signaling impairments contribute to worsening endothelial dysfunction, which is an independent predictor for future cardiovascular events in T2DM. In addition, these SGLT-2 Inhibitors reduce body weight, visceral adiposity, systolic and diastolic blood pressure, microalbuminuria, and oxidative stress. However, there are no studies examining the effects of SGLT-2 inhibitor therapy on NFkappaB and other inflammatory mediators in humans with T2DM. Moreover, no studies have examined the effect of SGLT-2 inhibitor therapy on endothelial function in this population. In the present study the investigators will assess whether dapagliflozin treatment for 12 weeks reduces monocyte inflammation and improves endothelial dysfunction in patients with type 2 diabetes on metformin monotherapy.

Arms: arm 1: Dapagliflozin 5 mg daily by mouth for 2 weeks followed by 10 mg by mouth daily for 10 weeks arm 2: Placebo tablets by mouth daily for 12 weeks
Interventions: intervention 1: Dapagliflozin intervention 2: Placebo
Intervention descriptions: intervention 1: Patients with Type 2 diabetes will be randomized to receive dapagliflozin 5 mg daily for 2 weeks followed by10 mg daily for 10 weeks by mouth or matching placebo for 12 weeks. All subjects will receive measurements of fasting plasma glucose, Free Fatty Acids, inflammatory markers and adipocytokines, monocyte inflammation, as well as ultrasound assessment of flow-mediated dilatation (FMD) of the brachial artery at baseline and after 12 weeks of drug treatment with either dapagliflozin or placebo. intervention 2: Patients with Type 2 diabetes will be randomized to receive dapagliflozin 5 mg daily for 2 weeks followed by10 mg daily for 10 weeks by mouth or matching placebo for 12 weeks. All subjects will receive measurements of fasting plasma glucose, Free Fatty Acids, inflammatory markers and adipocytokines, monocyte inflammation, as well as ultrasound assessment of flow-mediated dilatation (FMD) of the brachial artery at baseline and after 12 weeks of drug treatment with either dapagliflozin or placebo.

Protocol documents:
Prot_SAP_000.pdf: 
1(35) 
 
 
 
 
Drug Substance 
Dapaglifozin 
 
Study Number 
H-35985 
 
Version Number 
1.3 
 
Date 
12 April, 2017 
 
 
 
 
 
 
THE EFFECT OF DAPAGLIFLOZIN ON INFLAMMATION AND 
ENDOTHELIAL DYSFUCTION 
Principal Investigator: Mandeep Bajaj, MD  
Sponsor: Baylor College of Medicine, Houston, TX  
 
 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
2(35) 
TABLE OF CONTENTS 
PAGE
TITLE PAGE ........................................................................................................... 1 
TABLE OF CONTENTS ......................................................................................... 2 
LIST OF ABBREVIATIONS AND DEFINITION OF TERMS ............................ 6 
1. 
INTRODUCTION ................................................................................................... 7 
1.1 
Background .............................................................................................................. 7 
1.2 
Research hypothesis ................................................................................................. 7 
1.3 
Rationale for conducting this study ......................................................................... 7 
1.4 
Benefit/risk and ethical assessment.......................................................................... 8 
2. 
STUDY OBJECTIVES .......................................................................................... 10 
2.1 
Primary objective ................................................................................................... 10 
2.2 
Secondary objectives ............................................................................................. 10 
2.3 
Safety objective ...................................................................................................... 10 
2.4 
Exploratory objectives ........................................................................................... 10 
3. 
STUDY PLAN AND PROCEDURES .................................................................. 11 
3.1 
Overall study design and flow chart ...................................................................... 11 
3.2 
Rationale for study design, doses and control groups ............................................ 11 
4. 
SUBJECT SELECTION CRITERIA .................................................................... 11 
4.1 
Inclusion criteria .................................................................................................... 12 
4.2 
Exclusion criteria ................................................................................................... 12 
5. 
STUDY CONDUCT .............................................................................................. 12 
5.1 
Restrictions during the study ................................................................................. 13 
5.2 
Subject enrollment <<and randomization>> <<and initiation of 
investigational product>> ...................................................................................... 14 
5.2.1 
Procedures for randomization ................................................................................ 14 
5.3 
Procedures for handling subjects incorrectly enrolled<< or randomized>> 
<<or initiated on investigational product>> ......................................................... 14 
5.4 
Blinding and procedures for unblinding the study ................................................. 14 
5.4.1 
Methods for ensuring blinding ............................................................................... 14 
5.4.2 
Methods for unblinding the study .......................................................................... 14 
5.5 
Treatments .............................................................................................................. 15 
5.5.1 
Identity of investigational product(s) ..................................................................... 15 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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5.5.2 
Doses and treatment regimens ............................................................................... 15 
5.5.3 
Additional study drug ............................................................................................ 15 
5.5.4 
Labeling ................................................................................................................. 15 
5.5.5 
Storage ................................................................................................................... 15 
5.6 
Concomitant and post-study treatment(s) .............................................................. 16 
5.7 
Treatment compliance ............................................................................................ 16 
5.7.1 
Accountability ........................................................................................................ 16 
5.8 
Discontinuation of investigational product ............................................................ 16 
5.8.1 
Procedures for discontinuation of a subject from investigational product ............ 17 
5.9 
Withdrawal from study .......................................................................................... 17 
6. 
COLLECTION OF STUDY VARIABLES ........................................................... 18 
6.1 
Recording of data ................................................................................................... 18 
6.2 
Data collection at enrolment and follow-up ........................................................... 18 
6.2.1 
Enrollment procedures ........................................................................................... 18 
6.2.2 
Follow-up procedures ............................................................................................ 18 
6.3 
Efficacy .................................................................................................................. 18 
6.3.1 
<<Efficacy variable>> ........................................................................................... 19 
6.4 
Safety ..................................................................................................................... 19 
6.4.1 
Definition of adverse events .................................................................................. 19 
6.4.2 
Definitions of serious adverse event ...................................................................... 20 
6.4.3 
Recording of adverse events .................................................................................. 20 
6.4.4 
Reporting of serious adverse events....................................................................... 22 
6.4.5 
Laboratory safety assessment ................................................................................ 23 
6.4.6 
Physical examination ............................................................................................. 23 
6.4.7 
Vital signs .............................................................................................................. 23 
6.4.7.1 
Pulse and blood pressure ........................................................................................ 23 
6.4.8 
Other safety assessments ........................................................................................ 23 
6.5 
Patient reported outcomes (PRO) .......................................................................... 24 
6.5.1.1 
<<Name of PRO method or questionnaire>> ........................................................ 24 
6.6 
Pharmacokinetics ................................................................................................... 24 
6.6.1 
Collection of samples ............................................................................................. 24 
6.6.2 
Determination of drug concentration ..................................................................... 24 
6.7 
Pharmacodynamics ................................................................................................ 24 
6.7.1 
Collection of pharmacodynamic markers .............................................................. 24 
6.8 
Pharmacogenetics .................................................................................................. 24 
6.8.1 
Collection of pharmacogenetic samples ................................................................ 24 
6.9 
Health economics ................................................................................................... 24 
7. 
BIOLOGICAL SAMPLING PROCEDURES ....................................................... 24 
7.1 
Volume of blood .................................................................................................... 24 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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7.2 
Handling, storage and destruction of biological samples ...................................... 25 
7.2.1 
Pharmacokinetic and/or pharmacodynamic samples ............................................. 25 
7.2.2 
Pharmacogenetic samples ...................................................................................... 25 
7.3 
Labeling and shipment of biohazard samples ........................................................ 25 
7.4 
Chain of custody of biological samples ................................................................. 25 
7.5 
Withdrawal of informed consent for donated biological samples ......................... 25 
8. 
ETHICAL AND REGULATORY REQUIREMENTS ......................................... 25 
8.1 
Ethical conduct of the study ................................................................................... 25 
8.2 
Ethics and regulatory review.................................................................................. 26 
8.3 
Informed consent ................................................................................................... 26 
8.4 
Changes to the protocol and informed consent form ............................................. 27 
8.5 
Audits and inspections ........................................................................................... 27 
9. 
STUDY MANAGEMENT .................................................................................... 27 
9.1 
Training of study site personnel ............................................................................. 28 
9.2 
Monitoring of the study ......................................................................................... 28 
9.2.1 
Source data ............................................................................................................. 28 
9.3 
Study timetable and end of study ........................................................................... 28 
10. 
DATA MANAGEMENT ....................................................................................... 28 
11. 
EVALUATION AND CALCULATION OF VARIABLES ................................. 28 
11.1 
Calculation or derivation of efficacy variable(s) ................................................... 29 
11.2 
Calculation or derivation of safety variable(s) ....................................................... 29 
11.2.1 
Other significant adverse events (OAE) ................................................................ 29 
11.3 
Calculation or derivation of patient reported outcome variables ........................... 29 
11.4 
Calculation or derivation of pharmacokinetic variables ........................................ 29 
11.5 
Calculation or derivation of pharmacodynamic variable(s) ................................... 29 
11.5.1 
Calculation or derivation of the relationship between pharmacokinetic and 
pharmacodynamic variables ................................................................................... 29 
11.5.2 
Population analysis of pharmacokinetic/pharmacodynamic variables .................. 29 
11.6 
Calculation or derivation of pharmacogenetic variables........................................ 29 
11.7 
Calculation or derivation of health economic variables......................................... 29 
12. 
STATISTICAL METHODS AND SAMPLE SIZE DETERMINATION ............ 30 
12.1 
Description of analysis sets .................................................................................... 30 
12.1.1 
Efficacy analysis set ............................................................................................... 30 
12.1.2 
Safety analysis set .................................................................................................. 30 
12.2 
Methods of statistical analyses ............................................................................... 30 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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12.2.1 
Interim analyses ..................................................................................................... 31 
12.3 
Determination of sample size ................................................................................. 31 
12.4 
Data monitoring committee ................................................................................... 31 
13. 
IMPORTANT MEDICAL PROCEDURES TO BE FOLLOWED BY THE 
INVESTIGATOR .................................................................................................. 31 
13.1 
Overdose ................................................................................................................ 32 
13.2 
Pregnancy ............................................................................................................... 32 
13.2.1 
Maternal exposure .................................................................................................. 32 
13.2.2 
Paternal exposure ................................................................................................... 32 
14. 
LIST OF REFERENCES ....................................................................................... 33 
 
LIST OF TABLES 
Table 1: Visit Schedule 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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LIST OF ABBREVIATIONS AND DEFINITION OF TERMS 
The following abbreviations and special terms are used in this study Clinical Study Protocol. 
Abbreviation or 
special term 
Explanation 
 
 
T2DM 
 NFkB 
Type 2 Diabetes Mellitus 
Nuclear factor kappaB 
 
 
 
 
 
 
 
 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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1. 
INTRODUCTION 
The insulin-resistant state of type 2 diabetes mellitus (T2DM) is largely mediated by 
inflammatory pathways affecting skeletal muscle (1, 2), which is the primary site of whole 
body insulin resistance (3).  Nuclear factor kappa B (NFkB) regulates pro-inflammatory 
cytokines which ultimately impair skeletal muscle insulin signaling and fatty acid oxidation; 
its activity reflects overall inflammatory tone in skeletal muscle (2, 4).   Recent human studies 
confirm that NFkB is elevated in the skeletal muscle of T2DM human subjects (5-7).  
Furthermore, the same inflammatory processes and signaling impairments contribute to 
worsening endothelial dysfunction (8), which is an independent predictor for future 
cardiovascular events in T2DM (9). 
In the present study we will assess whether dapagliflozin reduces monocyte inflammation and 
improves endothelial dysfunction in patients with type 2 diabetes.  
 
1.1 
Background 
Multiple rodent models of T2DM show that 1-2 months of SGLT-2 inhibitor (including 
dapagliflozin) therapy results in greater insulin sensitivity and decreased levels of 
proinflammatory cytokines and free fatty acids (10-15).  Recent studies in T2DM human 
subjects show that 2-4 weeks of SGLT-2 inhibitor therapy improves insulin sensitivity despite 
an increase in endogenous glucose production (16, 17).  However, there are no studies 
examining the effects of SGLT-2 inhibitor therapy on NFkappaB and other inflammatory 
mediators in humans with T2DM.  Moreover, no studies have examined the effect of SGLT-2 
inhibitor therapy on endothelial function in this population.   We propose a study to elucidate 
the anti-inflammatory mechanisms related to SGLT-2 inhibitor therapy in T2DM. 
1.2 
Research hypothesis 
In T2DM subjects on metformin monotherapy, adding dapagliflozin for 12 weeks will reduce 
monocyte  inflammation and improve endothelial function, when compared to adding placebo 
for 12 weeks.  
1.3 
Rationale for conducting this study 
SGLT-2 Inhibitors reduce hyperglycemia and improve peripheral insulin sensitivity by 
ameliorating  glucotoxicity. Insulin resistance in type 2 diabetes is associated with endothelial 
dysfunction and vascular inflammation. Thus strategies to improve insulin sensitivity and 
lower glucotoxicity may improve endothelial inflammation and vascular inflammation. In 
addition, these SGLT-2 Inhibitors reduce body weight, visceral adiposity, systolic and 
diastolic blood pressure, microalbuminuria, and oxidative stress.  However, the effects of 
these agents on vascular inflammation and endothelial function is not known in  patients with 
type 2 diabetes although anti-inflammatory properties have been demonstrated in various 
animal models. In the present study we will assess if dapagliflozin decreases monocyte 
inflammation and improves endothelial function in patients with type 2 diabetes on metformin 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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monotherapy.  If our hypothesis is confirmed, further studies will be needed to assess the 
clinical significance of these findings on cardiovascular outcomes in patients with type 2 
diabetes and whether SGLT-2 inhibitors can reduce cardiovascular morbidity and mortality.  
1.4 
Benefit/risk and ethical assessment 
Dapagliflozin is associated with: 
1. Hypotension: Dapagliflozin causes intravascular volume contraction. Symptomatic 
hypotension can occur after initiating Dapagliflozin, particularly in patients with impaired 
renal function (eGFR <60 mL/min/1.73 m2), elderly patients, or patients on loop diuretics.  
2. Impairment in Renal Function: Dapagliflozin increases serum creatinine and decreases 
eGFR. Elderly patients and patients with impaired renal function may be more susceptible to 
these changes. Adverse reactions related to renal function can occur after initiating 
Dapagliflozin. Before initiating Dapagliflozin, renal function should be monitored and 
periodically thereafter. Dapagliflozin should be discontinued when eGFR is persistently <60 
mL/min/1.73 m2.  
3. Hypoglycemia with Concomitant Use with Insulin and Insulin Secretagogues: Insulin and 
insulin secretagogues are known to cause hypoglycemia. Dapagliflozin can increase the risk of 
hypoglycemia when combined with these agents. The dose of insulin or insulin secretagogue 
needs to be decreased to reduce the risk of hypoglycemia when used in combination with 
Dapagliflozin.  
4. Dapagliflozin increases the risk of genital mycotic infections especially in those patients 
with a previous history of genital mycotic infections. 
5. Increases in Low-Density Lipoprotein Cholesterol (LDL-C): Increases in LDL-C occur 
with Dapagliflozin.  
6. Bladder cancer: Across 22 clinical studies, newly diagnosed cases of bladder cancer were 
reported in 0.17% of dapagliflozin-treated patients and 0.03% of placebo/comparator-treated 
patients. After excluding patients in whom exposure to study drug was <1 year at the time of 
diagnosis of bladder cancer, there were 4 cases with dapagliflozin and no cases with 
placebo/comparator. Bladder cancer risk factors and hematuria (a potential indicator of pre-
existing tumors) were balanced between treatment arms at baseline. There were too few cases 
to determine whether the emergence of these events is related to dapagliflozin 
There are insufficient data to determine whether dapagliflozin has an effect on pre-existing 
bladder tumors. Dapagliflozin should not be used in patients with active bladder cancer. 
Furthermore, it needs to be used with caution in patients with a prior history of bladder cancer 
 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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In a pool of 12 placebo-controlled studies, the most common adverse reactions (≥5%) 
associated with Dapagliflozin 5 mg, 10 mg, and placebo respectively were female genital 
mycotic infections (8.4% vs 6.9% vs 1.5%), nasopharyngitis (6.6% vs 6.3% vs 6.2%), and 
urinary tract infections (5.7% vs 4.3% vs 3.7%).  
7. Diabetic Ketoacidosis: The regulatory authorities, such as the FDA and EMA, have issued 
communications regarding serious and sometimes life-threatening cases of diabetic 
ketoacidosis (DKA) in patients treated with the sodium-glucose cotransporter-2 inhibitors 
canagliflozin, dapagliflozin and empagliflozin.  
DKA is typically seen in patients with type 1 diabetes but may rarely happen in patients with 
type 2 diabetes and occurs as a consequence of absolute or relative insulin deficiency. The 
FDA communication notes that cases of DKA in patients on SGLT2 inhibitors often were 
atypical in that the patients’ blood sugar levels were only slightly elevated. Potential triggers 
for development of DKA included recognized risk factors such as major illness and reduced 
insulin doses, as well as reduced food and fluid intake. In the type 2 diabetes clinical 
development program phase II / III for dapagliflozin, there was one patient with a reported 
adverse event of diabetes ketoacidosis (who was on insulin treatment) out of a total of 5936 
dapagliflozin treated patients (6247 patient years). No cases of diabetes ketoacidosis were 
observed in the comparator groups with a total of 3403 patients (3638 patient years).  
i) The study team will advise patients to alert them and seek medical attention immediately if 
they experience symptoms consistent with DKA such as:  nausea, vomiting, abdominal pain, 
confusion, change in breathing pattern and unusual fatigue or sleepiness. 
ii) The study team will evaluate any patient experiencing the aforementioned signs or 
symptoms of DKA, even if glucose levels are not high. The study team will discontinue study 
drug if acidosis is confirmed and take standard clinical measures to treat DKA when it 
considers this to be a likely diagnosis.   
Use in Specific Populations 
-Pregnant Women: There are no adequate and well-controlled studies of Dapagliflozin in 
pregnant women. Consider appropriate alternative therapies, especially during the second and 
third trimesters.  
•Nursing Mothers: It is not known whether Dapagliflozin is excreted in human milk. Because 
of the potential for serious adverse reactions in nursing infants from Dapagliflzin, it is advised 
to discontinue nursing or discontinue Dapagliflozin. 
•Geriatric Use: A higher proportion of patients ≥65 years treated with Dapagliflozin had 
adverse reactions related to volume depletion and renal impairment or failure compared to 
patients treated with placebo. No Dapagliflozin dose change is recommended based on age. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
10(35) 
Yet, Phase III clinical trials showed that dapagliflozin reduced fasting and postprandial plasma 
glucose and significantly reduced HbA1c and was associated with a significant reduction in 
weight in patients with type 2 diabetes. Thus the benefits outweigh the risks of therapy.  
2. 
STUDY OBJECTIVES    
2.1 
Primary objective 
The primary objective is to examine the effects of 12 weeks of dapagliflozin therapy on 
monocyte inflammation i.e. NFkB (p65) protein in T2DM subjects. 
2.2 
Secondary objectives 
The secondary objectives are to examine the effects of 12 weeks of dapagliflozin therapy on 
plasma adipocytokines and endothelial function (as measured by flow mediated dilatation) in 
T2DM subjects. 
2.3 
Safety objective 
To study the safety of dapagliflozin in type 2 diabetes by monitoring for clinical events. 
2.4 
Exploratory objectives 
None 
3. 
STUDY PLAN AND PROCEDURES 
3.1 
Overall study design and flow chart (Table 1) 
This is a randomized double blind placebo controlled clinical trial. Prior to beginning the 
study, interested potential subjects will be prescreened via a telephone interview. If the subject 
meets criteria via telephone interview, he or she will be invited to participate in an hour-long 
outpatient screening visit (Visit 1). At Visit 1, subjects will be consented for participation in 
the study. We will then perform a complete history and physical exam and EKG. We will also 
draw the following baseline fasting labs: complete metabolic panel (glucose, BUN/Creatinine, 
liver function tests, electrolytes), lipid panel, hemoglobin A1C, complete blood count, 
urinanalysis, and urine pregnancy test (if applicable). Results will be reviewed. If the subject 
meets the inclusion and exclusion criteria, then he or she will be invited to participate in the 
trial. This is a single-center, randomized, placebo-controlled double-blinded prospective trial 
with a 1:1 allocation ratio. Patients with type 2 diabetes on metformin monotherapy will be 
enrolled as per the inclusion criteria. At the start of the study (Visit 2), all subjects will receive 
the following fasting baseline measurements: (i) plasma free fatty acids (FFA), plasma 
adiponectin, hsCRP, IL-1b, IL-6, TNF-alpha, 15-epi-lipoxin A4, glucose, and insulin; (ii) 
monocyte isolation from peripheral (venous) whole blood and quantification of monocyte 
NFkappaB (P65),TLR2, TLR4, SOCS-3, MYD88, and IkB-alpha, and IkB-beta protein as 
well as expression levels of CD36, MCP-1, MMP-9,VCAM1, ICAM1 and JNK; and (iii) 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
11(35) 
ultrasound assessment of flow-mediated dilatation (FMD) of the brachial artery per previously 
validated methods (18, 19). Each subject will be randomized to one of two arms: (1) addition 
of placebo pill daily for 12 weeks, or (2) addition of dapagliflozin 5 mg daily for 2 weeks 
followed by 10 mg po daily for 10 weeks (Visit 3). During the entire 12-week treatment 
period, subjects will return to clinic every 2-4 weeks (Visit 4-7) following an overnight fast 
for measurement of glucose, kidney function and electrolytes (BMP), body weight, and blood 
pressure. A urine pregnancy test (if applicable) will also be performed at each visit. At the end 
of the treatment period of 12 weeks (Visit 7), all subjects will again receive the following 
fasting measurements: (i) BMP, plasma free fatty acids (FFA), plasma lipids (including 
triglycerides), plasma adiponectin, hsCRP, IL-1b, IL-6, TNF-alpha, 15-epi-lipoxin A4, 
glucose, insulin, and HbA1c; (ii) monocyte isolation from peripheral (venous) whole blood 
and quantification of monocyte NFkappaB (P65), TLR2, TLR4, SOCS-3, MYD88, and IkB-
alpha, and IkB-beta protein as well as expression levels of CD36, MCP-1, MMP-9,VCAM1, 
ICAM1 and JNK; and (iii) ultrasound assessment of flow-mediated dilatation (FMD) of the 
brachial artery.   
 
3.2 
Rationale for study design, doses and control groups 
As the study hypothesis is to test whether dapagliflozin decreases vascular inflammation and 
improves endothelial function at a dose used in the treatment of type 2 diabetes. Patients with 
type 2 diabetes on metaformin therapy will receive either dapagliflozin or placebo orally for 
12 weeks. Dapagliflozin 5 and 10 mg daily is FDA approved for the treatment of type 2 
diabetes in patients on metformin therapy. 
4. 
SUBJECT SELECTION CRITERIA 
4.1 
Inclusion criteria 
For inclusion in the study subjects should fulfill the following criteria: 
1. 
Provision of informed consent prior to any study specific procedures 
2.            Men and women, ages 21 to 70 years.   
i) 
Women of childbearing potential (WOCBP) must be using an acceptable method of 
contraception to avoid pregnancy throughout the study in such a manner that the risk of 
pregnancy is minimized.  
ii) 
Women must have a negative serum or urine pregnancy test (minimum sensitivity 25 
IU/L or equivalent units of HCG) within 72 hours prior to the start of investigational product.  
iii) 
 Women must not be breastfeeding. 
 
3. 
Patients with Type 2 Diabetes Mellitus with the following parameters at study 
entry: hemoglobin A1C ranging from 6.5% to 9.0% and a fasting blood glucose ≤ 
200 mg/dL. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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4. 
Patients must be on a stable dose of Metformin therapy for 3 months; the dose of 
metformin will not change for the duration of the study. 
5. 
Patients are allowed, but not required, to be on statins, ACE-inhibitors, and 
angiotensin-receptor blockers at doses that have been stable for at least the last 3 
months prior to enrollment in the study. Doses will not be changed for the duration 
of the study. 
6. 
Patients must have a BMI between 27-40 kg/m2  
7. 
Patients must have a stable body weight (±4-5 pounds) for three months prior to 
enrollment in the study. 
8. 
Patients must have a Creatinine Clearance > 60 mL/min (calculated by Cockcroft-
Gault formula). 
9. 
Patients must have the following laboratory values: Hematocrit ≥ 34 vol%; S. 
creatinine < 1.5 mg/dl in men and 1.4 mg/dl in women and Cr Clearance > 60 
ml/min; and AST (SGOT) < 2.5 times ULN, ALT (SGPT) < 2.5 times ULN, 
alkaline phosphatase< 2.5 times ULN. 
4.2 
Exclusion criteria 
Subjects should not enter the study if any of the following exclusion criteria are fulfilled: 
1. History of Type 1 diabetes mellitus  
2.  Women who are pregnant or breastfeeding  
3.  Patients receiving lipid-lowering medications other than statins within the last 3 months  
4. Patient receiving SGLT-2 inhibitors, DPP-IV inhibitors, GLP-1 receptor agonists, 
thiazolidinediones, insulin, sulfonylureas, alpha-glucosidase inhibitors, corticosteroids, 
immunosuppressive therapy, thiazide or loop diuretics, or hormone replacement therapy 
within the last 3 months  
5. Patient must stop treatment with NSAIDs and antioxidant vitamin supplements at least one 
week prior to the start of the study  
6. Patients with diabetic gastroparesis  
7. Patients with current tobacco use  
8. Patients with active malignancy  
9. Patients with history of urinary bladder cancer  


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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10. Patients with a history of clinically significant heart disease (NYHA III or IV; more than 
non-specific ST-T wave changes on the EKG), peripheral vascular disease (history of 
claudication), or pulmonary disease (dyspnea on exertion of one flight or less; abnormal 
breath sounds on auscultation) will not be studied  
11.Subjects with a history of any serious hypersensitivity reaction to dapagliflozin 
12. Prisoners, or subjects who are involuntarily incarcerated  
13. Subjects who are compulsorily detained for treatment of either a psychiatric or physical 
(eg, infectious disease) illness. 
14. Patients with significant cardiac, hepatic or renal disease (Creatinine Clearance < 60 
mL/min calculated by Cockcroft-Gault formula) will be excluded. 
5. 
STUDY CONDUCT 
5.1 
Restrictions during the study 
1. Patients will not be allowed to take nonsteroidal anti-inflammatory drugs (NSAID) and 
antioxidant vitamin supplements during the study. 
5.2 
Subject enrollment and randomization and initiation of 
investigational product 
5.2.1 
Procedures for randomization 
Patients will be randomized to one of the two treatment groups: 1. Dapagliflozin or 2. Placebo. 
Randomization numbers (patient numbers) will be assigned strictly sequentially as patients 
become eligible for randomization. Randomization will be computer-generated by the research 
pharmacy, and the investigators will be blinded to the treatment assignments. The research 
pharmacy will provide the randomization number and the appropriate bottle numbers. 
5.3 
Procedures for handling subjects incorrectly enrolled<< or 
randomized>> <<or initiated on investigational product>> 
Enrolled patients who ultimately do not meet the inclusion/ exclusion criteria will not undergo 
further evaluation and be excluded from the study. A log of these subjects will be kept. 
 
5.4 
Blinding and procedures for unblinding the study 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
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5.4.1 
Methods for ensuring blinding 
The investigators will be blinded to the treatment assignments. The research pharmacy will 
provide the randomization number and the appropriate bottle numbers. 
5.4.2 
Methods for unblinding the study 
Blinding is critical to the integrity of this clinical study. However, in the event of a medical 
emergency or pregnancy in a subject, in which knowledge of the investigational product is 
critical to the subject's management, the blind for that subject may be broken by the treating 
physician.  
Before breaking the blind of an individual subject's treatment, the investigator should have 
determined that the information is necessary, ie, that it will alter the subject's immediate 
management. In many cases, particularly when the emergency is clearly not investigational 
product-related, the problem may be properly managed by assuming that the subject is 
receiving active product without the need for unblinding. 
 
 
 
5.5 
Treatments 
5.5.1 
Identity of investigational product(s) 
Investigational 
Product 
Manufacturer
IMP/
NIMP 
Type
Route of 
administratio
n 
Formulation
Comment
Dapagliflozin 5 
mg  
AstraZeneca 
IMP 
Active 
Drug  
Oral  
Film Coated 
Tablet   
Green, plain, 
diamond shaped 
Matching 
placebo for 
Dapagliflozin 
2.5mg and 5 mg 
AstraZeneca 
IMP 
Placebo  
Oral 
Film Coated 
Tablet   
Green, plain, 
diamond shaped 
Dapagliflozin 10 
mg  
AstraZeneca 
IMP 
Active 
Drug  
Oral  
Film Coated 
Tablet   
Green, plain, 
diamond shaped 
Matching 
placebo for 
Dapagliflozin 10 
mg  
AstraZeneca 
IMP 
Placebo  
Oral 
Film Coated 
Tablet   
Green, plain, 
diamond shaped 
 
AstraZeneca will provide the Sponsor with Dapagliflozin and matching placebo tablets. 
Dapagliflozin and matching placebo tablets will be supplied in bottles, each containing 35 
tablets. The tablets contain lactose, which may cause discomfort in lactose-intolerant 
individuals. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
15(35) 
5.5.2 
Doses and treatment regimens 
                Dapagliflozin 5 mg  once daily orally for 2 weeks followed by Dapagliflozin 10 mg 
once daily orally for 10 weeks/ Placebo PO once daily. 
5.5.3 
Additional study drug 
All patients will continue on metformin treatment at their current dose (dose at the time of 
screening). The dose of metformin will not be changed during the study. 
5.5.4 
Labeling 
Packaging and labeling will be in accordance with applicable local regulatory requirements to 
protect the blinded nature of the trial. Labeling will be done by the Sponsor. Labels will be 
prepared in accordance with Good Manufacturing Practice (GMP) and local regulatory 
guidelines. The labeling will not indicate whether the medication is dapagliflozin or placebo. 
Dapagliflozin and matching placebo tablets will be supplied in bottles, each containing 35 
tablets by AstraZeneca.  
Metformin will be provided as a prescription medication from the local pharmacy and billed 
to the patient or his insurance company. 
The study drugs must be swallowed whole with a liquid and not chewed, divided, dissolved, 
or crushed. 
5.5.5 
Storage 
All study drugs should be kept in a secure place and under appropriate storage conditions. The 
Investigational Product label on the bottle specifies the appropriate storage. 
5.6 
Concomitant and post-study treatment(s) 
1. 5. 
Patient must stop treatment with NSAIDs and antioxidant vitamin supplements at least 
one week prior to the start of the study. 
2. No change in the dose of metformin during the 12 week study period. 
5.7 
Treatment compliance 
Treatment compliance will be monitored by drug accountability as well as the patient’s medical 
record. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
16(35) 
5.7.1 
Accountability 
Subjects will be asked to bring any unused study drug to each study visit. 
5.8 
Discontinuation of investigational product 
Subjects MUST discontinue investigational product (and noninvestigational product at the 
discretion of the investigator) for any of the following reasons: 
•Withdrawal of informed consent (subject’s decision to withdraw for any reason). 
• 
Any clinical adverse event (AE), laboratory abnormality, or intercurrent illness which, 
in the opinion of the investigator, indicates that continued participation in the study is not in 
the best interest of the subject. 
• 
Pregnancy: Instruct WOCBP to contact the investigator or study staff immediately 
if they suspect they might be pregnant (eg, missed or late menstrual period) at any time during 
study participation. Institutional policy and local regulations should determine the frequency 
of on study pregnancy tests for WOCBP enrolled in the study. The investigator must 
immediately notify AZ if a study subject becomes pregnant.  
• 
Termination of the study by AZ 
• 
Loss of ability to freely provide consent through imprisonment or involuntarily 
incarceration for treatment of either a psychiatric or physical (eg, infectious disease) illness 
• 
Inability to comply with the protocol. 
All subjects who discontinue should comply with protocol specified follow-up procedures as 
outlined in section 6. The only exception to this requirement is when a subject withdraws 
consent for all study procedures or loses the ability to consent freely (ie, is imprisoned or 
involuntarily incarcerated for the treatment of either a psychiatric or physical illness). If a 
subject was withdrawn before completing the study, the reason for withdrawal must be 
entered on the appropriate case report form (CRF) page. 
 
5.8.1 
Procedures for discontinuation of a subject from investigational product  
Treatment with metformin should continue during the treatment period per current 
guidelines. At the end of the study, patients will continue with metformin therapy and 
follow up with their primary care physician.  
5.9 
Withdrawal from study 
Subjects are eligible to withdraw from this study at any time, for any reason, without 
impacting standard clinical care. Data collected up to the date of withdrawal from the 
study will still be used in the data analysis. 
The investigator and/or AstraZeneca may decide to withdraw a subject(s) from 
investigational product and/or other protocol-required therapies, protocol procedures, 
or the study as a whole at any time prior to study completion.  


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
17(35) 
Reasons for removal for treatment or the study can include any of the following:  
• Subject request  
• Safety concern (eg, due to an adverse event, ineligibility determined, protocol 
deviation, non-compliance, requirement for alternative therapy, protocol-specified 
criteria (list criteria), pregnancy)  
• Decision by sponsor (other than subject request or safety concern)  
• Death  
• Lost to follow-up  
 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
18(35) 
6. 
COLLECTION OF STUDY VARIABLES 
6.1 
Recording of data 
All data will be recorded in Case Report Forms (CRF) and stored in a secure area accessible 
only to the study investigators. 
6.2 
Data collection at enrolment and follow-up 
6.2.1 
Enrollment procedures 
The following procedures are to be completed after informed consent has been 
obtained:  
 Informed Consent Form signed  
 Medical history  
 Vital signs (eg, blood pressure, heart rate, respiration rate)  
 Review for Adverse Events and Serious Adverse Events (only AEs 
possibly related to study procedures and SAEs are collected)  
 Documentation of concomitant medications  
 Physical exam  
 Body height  
 Body weight 
 EKG 
 blood will be drawn for plasma free fatty acids (FFA), plasma lipids 
(including triglycerides), plasma adiponectin, hsCRP, IL-1b, IL-6, TNF-
alpha, 15-epi-lipoxin A4, glucose, insulin, and HbA1c; monocyte isolation 
from peripheral (venous) whole blood and quantification of monocyte 
NFkappaB (P65),TLR2, TLR4, SOCS-3, MYD88, and IkB-alpha, and IkB-
beta protein as well as expression levels of CD36, MCP-1, MMP-
9,VCAM1, ICAM1 and JNK 
 ultrasound assessment of flow-mediated dilatation (FMD) of the brachial 
artery  
 Randomization of treatment assignment 
 Investigational product  
 Local laboratory Assessments:  
Complete metabolic panel and CBC, HbA1C, Lipids 
Urine pregnancy test (females of childbearing potential only) 
6.2.2 
Follow-up procedures 
 Vital signs (e.g., blood pressure, heart rate, respiration rate)  
 Review for Adverse Events and Serious Adverse Events (only AEs 
possibly related to study procedures and SAEs are collected)  
 Blood will be drawn for fasting plasma glucose and kidney function tests 
(BMP) at each follow up visit (2-4 weeks interval). Urine pregnancy test 
(women of childbearing potential) will be performed at each follow up 
visit. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
19(35) 
 At the end of the study visit, blood will be drawn for plasma free fatty acids 
(FFA), plasma lipids (including triglycerides), plasma adiponectin, hsCRP, 
IL-1b, IL-6, TNF-alpha, 15-epi-lipoxin A4, glucose, insulin, and HbA1c; 
monocyte isolation from peripheral (venous) whole blood and 
quantification of monocyte NFkappaB (P65),TLR2, TLR4, SOCS-3, 
MYD88, and IkB-alpha, and IkB-beta protein as well as expression levels 
of CD36, MCP-1, MMP-9,VCAM1, ICAM1 and JNK. 
 Documentation of concomitant medications and investigational product 
compliance at each follow up visit. 
 ultrasound assessment of flow-mediated dilatation (FMD) of the brachial 
artery at the end of the study visit. 
 
6.3 
Efficacy 
Blood samples will be withdrawn from a peripheral vein by a qualified personnel. 
Ultrasound assessment of flow-mediated dilatation (FMD) of the brachial artery will be 
performed by the PI or a trained co-investigator. 
 
6.3.1 
Efficacy variable 
- Ultrasound assessment of flow-mediated dilatation (FMD) of the brachial artery 
-Plasma free fatty acids (FFA), plasma lipids (including triglycerides), plasma adiponectin, 
hsCRP, IL-1b, IL-6, TNF-alpha, 15-epi-lipoxin A4, glucose, insulin, and HbA1c; monocyte 
isolation from peripheral (venous) whole blood and quantification of monocyte NFkappaB 
(P65),TLR2, TLR4, SOCS-3, MYD88, and IkB-alpha, and IkB-beta protein as well as 
expression levels of CD36, MCP-1, MMP-9,VCAM1, ICAM1 and JNK. 
6.4 
Safety 
The Principal Investigator is responsible for ensuring that all staff involved in the study is 
familiar with the content of this section. 
6.4.1 
Definition of adverse events  
An adverse event is the development of an undesirable medical condition or the deterioration 
of a pre-existing medical condition following or during exposure to a pharmaceutical product, 
whether or not considered causally related to the product.  An undesirable medical condition 
can be symptoms (eg, nausea, chest pain), signs (eg, tachycardia, enlarged liver) or the 
abnormal results of an investigation (eg, laboratory findings, electrocardiogram).  In clinical 
studies, an AE can include an undesirable medical condition occurring at any time, including 
run-in or washout periods, even if no study treatment has been administered. 
The term AE is used to include both serious and non-serious AEs. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
20(35) 
6.4.2 
Definitions of serious adverse event 
A serious adverse event is an AE occurring during any study phase (ie, run-in, treatment, 
washout, follow-up), that fulfils one or more of the following criteria: 
 
Results in death 
 
Is immediately life-threatening 
 
Requires in-patient hospitalization or prolongation of existing hospitalization  
 
Results in persistent or significant disability/incapacity or substantial disruption of 
the ability to conduct normal life functions 
 
Is a congenital abnormality or birth defect 
 
Is an important medical event that may jeopardize the subject or may require 
medical intervention to prevent one of the outcomes listed above. 
The causality of SAEs (their relationship to all study treatment/procedures) will be assessed 
by the investigator(s) and communicated to AstraZeneca. 
6.4.3 
Recording of adverse events 
Time period for collection of adverse events 
Adverse events will be evaluated on a continuous basis while the patient is on study and until 
30 days after the last dose of study drug. 
Follow-up of unresolved adverse events 
Patients should be followed until all treatment-related adverse events have recovered to 
baseline or are deemed irreversible by the principal investigator. 
 
The following variables will be collected for each AE: 
 
AE (verbatim) 
 
The date when the AE started and stopped 
 
Whether the AE is serious or not 
 
Investigator causality rating against the Investigational Product (yes or no) 
 
Action taken with regard to investigational product 
 
AE caused subject’s withdrawal from study (yes or no) 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
21(35) 
 
Outcome. 
In addition, the following variables will be collected for SAEs: 
 
Date AE met criteria for serious AE 
 
Date Investigator became aware of serious AE 
 
AE is serious due to 
 
Date of hospitalization 
 
Date of discharge 
 
Probable cause of death 
 
Date of death 
 
Autopsy performed 
 
Causality assessment in relation to Study procedure(s) 
 
Causality assessment in relation to Other medication 
 
Causality assessment in relation to Additional Study Drug 
 
Description of AE. 
It is important to distinguish between serious and severe AEs.  Severity is a measure of 
intensity whereas seriousness is defined by the criteria in Section 6.4.2.  An AE of severe 
intensity need not necessarily be considered serious.  For example, nausea that persists for 
several hours may be considered severe nausea, but not a SAE. On the other hand, a stroke 
that results in only a limited degree of disability may be considered a mild stroke but would be  
Adverse Events based on signs and symptoms 
When collecting AEs, the recording of diagnoses is preferred (when possible) to 
recording a list of signs and symptoms.  However, if a diagnosis is known and 
there are other signs or symptoms that are not generally part of the diagnosis, the 
diagnosis and each sign or symptom will be recorded separately. 
Adverse Events based on examinations and tests 
Deterioration as compared to baseline in protocol-mandated laboratory values, or 
vital signs should therefore only be reported as AEs if they fulfill any of the SAE 
criteria or are the reason for discontinuation of treatment with the investigational 
product. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
22(35) 
If deterioration in a laboratory value/vital sign is associated with clinical signs and 
symptoms, the sign or symptom will be reported as an AE and the associated 
laboratory result/vital sign will be considered as additional information.  
Wherever possible the reporting investigator uses the clinical, rather than the 
laboratory term (eg, anemia versus low hemoglobin value). In the absence of 
clinical signs or symptoms, clinically relevant deteriorations in non-mandated 
parameters should be reported as AE(s s). 
Deterioration of a laboratory value, which is unequivocally due to disease 
progression, should not be reported as an AE/SAE.. 
Any new or aggravated clinically relevant abnormal medical finding at a physical examination 
as compared with the baseline assessment will be reported as an AE. 
Disease progression 
Disease progression can be considered as a worsening of a subject’s condition attributable to 
the disease for which the investigational product is being studied.  It may be an increase in the 
severity of the disease under study and/or increases in the symptoms of the disease.  The 
development of chest pain should be considered as disease progression and not an AE.  
Events, which are unequivocally due to disease progression, should not be reported as an AE 
during the study. 
6.4.4 
Reporting of serious adverse events  
Investigators and other site personnel must inform the FDA, via a MedWatch/AdEERs form, 
of any serious or unexpected adverse events that occur in accordance with the reporting 
obligations of 21 CFR 312.32, and will concurrently forward all such reports to AZ.  A copy 
of the MedWatch/AdEERs report must be faxed to AstraZeneca at the time the event is 
reported to the FDA.  It is the responsibility of the investigator to compile all necessary 
information and ensure that the FDA receives a report according to the FDA reporting 
requirement timelines and to ensure that these reports are also submitted to AstraZeneca at the 
same time. 
When reporting to AstraZeneca, a cover page should accompany the MedWatch/AdEERs 
form indicating the following: 
 
Investigator Sponsored Study (ISS) 
 
The investigator IND number assigned by the FDA 
 
The investigator’s name and address 
 
The trial name/title and AstraZeneca ISS reference number 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
23(35) 
Investigative site must also indicate, either in the SAE report or the cover page, the causality 
of events in relation to all study medications and if the SAE is related to disease progression, 
as determined by the principal investigator. 
Send SAE report and accompanying cover page by way of fax to AstraZeneca’s designated 
fax line: 1-866-984-7229 
Serious adverse events that do not require expedited reporting to the FDA need to be reported 
to AstraZeneca preferably using the MedDRA coding language for serious adverse events. 
AstraZeneca wants to receive SAE that not require expedited reporting quarterly as line 
listings. 
In the case of blinded trials, AstraZeneca will request that the Sponsor either provide a copy of 
the randomization code/ code break information or unblind those SAEs which require 
expedited reporting. 
All SAEs have to be reported to AstraZeneca, whether or not considered causally related to 
the investigational product.  All SAEs will be documented.  The investigator is responsible for 
informing the IRB and/or the Regulatory Authority of the SAE as per local requirements 
6.4.5 
Laboratory safety assessment 
Study drug toxicities will be assessed continuously by the following clinical and laboratory 
assessments. 
For blood volume see Section 7.1. 
6.4.6 
Physical examination 
 signs of volume depletion, rash. 
6.4.7 
Vital signs 
respiratory rate >18/ min 
6.4.7.1 
Pulse and blood pressure 
We will report HR >100 bpm, SBP <100 mmHg, DBP <60 mmHg. 
6.4.8 
Other safety assessments 
We will check kidney function (BUN/Creatinine) and electrolytes (sodium/potassium) and 
fasting plasma glucose (hypoglycemia/hyperglycemia) at screening and at 2-4 weeks interval 
during the treatment period. 
6.5 
Patient reported outcomes (PRO) 
Not applicable 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
24(35) 
6.5.1.1 
<<Name of PRO method or questionnaire>> 
Not applicable 
6.6 
Pharmacokinetics 
None. 
6.6.1 
Collection of samples 
None. 
6.6.2 
Determination of drug concentration 
None. 
6.7 
Pharmacodynamics 
None. 
6.7.1 
Collection of pharmacodynamic markers  
None. 
6.8 
Pharmacogenetics 
None. 
6.8.1 
Collection of pharmacogenetic samples 
None. 
6.9 
Health economics 
None.  
7. 
BIOLOGICAL SAMPLING PROCEDURES 
7.1 
Volume of blood 
A total of 115 ml of blood will be drawn during the entire study 
15 ml of blood will be drawn during the screening visit (Visit 1). 
30 ml of blood will be drawn at Visit 2. 
10 ml of blood will be drawn and Visit 4 and 5 and 6 
40 ml of blood will be drawn at Visit 7. 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
25(35) 
7.2 
Handling, storage and destruction of biological samples 
7.2.1 
Pharmacokinetic and/or pharmacodynamic samples 
None. 
7.2.2 
Pharmacogenetic samples 
None. 
7.3 
Labeling and shipment of biohazard samples 
Local laboratories will be used for analysis of all specimens collected at each 
participating affiliate site. Each sample will be labeled and coded to protect subject 
confidentiality. 
. 
7.4 
Chain of custody of biological samples 
Not applicable 
7.5 
Withdrawal of informed consent for donated biological samples 
Withdrawal of consent for a study means that the subject does not wish to receive further 
protocol-required therapies or procedures, and the subject does not wish to or is unable 
to continue further study participation. If the subject withdraws consent for donated 
samples, and it has not been processed, the request will be honored. If the subject 
withdraws consent for donated biological samples and it has already been processed, the 
data collected up to the date of withdrawal will be included in the analysis of the study. 
The investigator is to discuss with the subject appropriate procedures for withdrawal 
from the study.  
 
8. 
ETHICAL AND REGULATORY REQUIREMENTS 
8.1 
Ethical conduct of the study 
This study will be conducted in accordance with Good Clinical Practice (GCP), as defined by 
the International Conference on Harmonisation (ICH) and in accordance with the ethical 
principles underlying European Union Directive 2001/20/EC and the United States Code of 
Federal Regulations, Title 21, Part 50 (21CFR50). 
The study will be conducted in compliance with the protocol. The protocol, any amendments, 
and the subject informed consent will receive Institutional Review Board/Independent Ethics 
Committee (IRB/IEC) approval/favorable opinion before initiation of the study. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
26(35) 
All potential serious breaches must be reported to AstraZeneca immediately. A serious breach 
is a breach of the conditions and principles of GCP in connection with the study or the protocol, 
which is likely to affect, to a significant degree, the safety or physical or mental integrity of the 
subjects of the study or the scientific value of the study. 
Study personnel involved in conducting this study will be qualified by education, training, and 
experience to perform their respective tasks. This study will not use the services of study 
personnel where sanctions have been invoked or where there has been scientific misconduct or 
fraud (eg, loss of medical licensure; debarment). 
8.2 
Ethics and regulatory review 
Before study initiation, the investigator must have written and dated approval/favorable 
opinion from the IRB/IEC for the protocol, consent form, subject recruitment 
materials/process (eg, advertisements), and any other written information to be provided to 
subjects. The investigator will also obtain approval of the FDA prior to the start of the study. 
The trial will be listed on clinicaltrials.gov. The investigator should also provide the IRB/IEC 
with a copy of the Investigator Brochure or product labeling, information to be provided to 
subjects, and any updates. The investigator should provide the IRB/IEC with reports, updates, 
and other information (eg, expedited safety reports, amendments, and administrative letters) 
according to regulatory requirements or institution procedures. 
8.3 
Informed consent 
Investigators must ensure that subjects (or, in those situations where consent cannot be given 
by subjects, the legally acceptable representative) are clearly and fully informed about the 
purpose, potential risks, and other critical issues regarding clinical studies in which they 
volunteer to participate. Freely given written informed consent must be obtained from every 
subject (or, in those situations where consent cannot be given by subjects, the legally acceptable 
representative) before clinical study participation, including informed consent for any screening 
procedures conducted to establish subject eligibility for the study. 
Investigators must: 
1) Provide a copy of the consent form and written information about the study in the language 
in which the subject is most proficient prior to clinical study participation. The language 
must be non-technical and easily understood.  
2) Allow time necessary for subject or subject's legally acceptable representative to inquire 
about the details of the study. 
3) Obtain an informed consent signed and personally dated by the subject or the subject's 
legally acceptable representative and by the person who conducted the informed consent 
discussion.  


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
27(35) 
4) Obtain the IRB/IEC’s written approval/favorable opinion of the written informed consent 
form and any other information to be provided to the subjects, prior to the beginning of the 
study, and after any revisions are completed for new information. 
5) If informed consent is initially given by a subject’s legally acceptable representative or legal 
guardian, and the subject subsequently becomes capable of making and communicating their 
informed consent during the study, then consent must additionally be obtained from the 
subject. 
6) Revise the informed consent whenever important new information becomes available that 
is relevant to the subject's consent. The investigator, or a person designated by the 
investigator, should fully inform the subject or the subject's legally acceptable representative 
or legal guardian, of all pertinent aspects of the study and of any new information relevant 
to the subject's willingness to continue participation in the study. This communication 
should be documented.  
The consent form must also include a statement that AZ and regulatory authorities have direct 
access to subject records. 
The rights, safety, and well-being of the study subjects are the most important considerations 
and should prevail over interests of science and society. 
8.4 
Changes to the protocol and informed consent form 
All protocol amendments and revisions to the informed consent must be submitted to the AZ 
protocol manager and to the IRB/IEC. No protocol amendments will be implemented until 
written approval has been given by the IRB/IEC, except when necessary to eliminate an 
immediate hazard to study subjects. 
8.5 
Audits and inspections 
The Investigator will assure that local study staff cooperate with monitoring and audits. The 
Investigator agrees to allow auditing of all essential clinical study documents by AstraZeneca 
or its authorized representatives and inspection by the FDA or other appropriate regulatory 
authorities. Auditing visits by AstraZeneca will be scheduled with the appropriate staff at 
mutually agreeable times as applicable. 
 
 
 
 
9.0 STUDY MANAGEMENT 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
28(35) 
9.1 Training of study site personnel 
The Investigator will assure research activities, including those study-related duties delegated 
and will be performed by appropriately qualified individuals. The Investigator will assure that 
study staff will demonstrate due diligence in recruiting and screening study patients. 
9.2 Monitoring of the study 
9.21 Source data 
The PI and his team will monitor the study on a daily basis. All treatment emergent AEs will 
be recorded on source documents (i.e. original documents, data, and records).  AEs include 
those reported spontaneously by the subject and those noted incidentally or as observed by the 
investigator or study personnel.  All clinically significant abnormalities noted upon physical 
examination, or other diagnostic test results should be reported as an AE, except for baseline 
measurements that may be considered part of the medical history.  In addition, all clinically 
significant AEs that continue at Study Termination will be followed up by the investigator and 
evaluated with additional tests if necessary, until the underlying cause is diagnosed or 
resolution occurs.  All AEs will be evaluated for intensity and causal relationship with use of 
the study medication and/or study procedures by the investigator and reported to the Baylor 
IRB. All SAEs will be reported to the Baylor IRB and the sponsor within 24 hours. In 
addition, a safety report will be submitted to Baylor IRB annually. Any new information 
regarding the study medications will be submitted to Baylor IRB. 
9.3 Study timetable and end of study 
 Subject participation is anticipated to continue for 13 weeks days (+/- 3 days) 
 End of Treatment: will occur at week 13 (+/- 3 days). 
 End of Study Primary Completion: defined as all the randomized subjects in of the 
study have either completed all the scheduled visits or have early terminated from 
the study 
 End of Trial: defined as all enrolled subjects of the study have either completed all 
the scheduled visits or have early terminated from the study. 
10. DATA MANAGEMENT 
The investigator is responsible for complying with the requirements for all assessments 
and data collection (including subject’s not receiving protocol-required therapies) as 
stipulated in the protocol for each subject in the study. For subjects who withdraw 
prior to completion of all protocol-required visits and are unable or unwilling to 
continue the Schedule of Assessments the investigator or study staff can search 
publically available records [where permitted]) to ascertain survival status. This 
ensures that the data set(s) produced as an outcome of the study is/are as 
comprehensive as possible.  


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
29(35) 
11.EVALUATION AND CALCULATION OF VARIABLES 
<<>> 
11.1 
Calculation or derivation of efficacy variable(s) 
<<>> 
11.2 
Calculation or derivation of safety variable(s) 
<<>> 
11.2.1 
Other significant adverse events (OAE) 
<<>> 
11.3 
Calculation or derivation of patient reported outcome variables 
<<>> 
11.4 
Calculation or derivation of pharmacokinetic variables 
None 
11.5 
Calculation or derivation of pharmacodynamic variable(s) 
None 
11.5.1 
Calculation or derivation of the relationship between pharmacokinetic and 
pharmacodynamic variables  
None 
11.5.2 
Population analysis of pharmacokinetic/pharmacodynamic variables  
None 
11.6 
Calculation or derivation of pharmacogenetic variables 
None 
11.7 
Calculation or derivation of health economic variables 
None 
12.STATISTICAL METHODS AND SAMPLE SIZE DETERMINATION 
12.1 
Description of analysis sets 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
30(35) 
12.1.1 
Efficacy analysis set 
Between groups comparisons will be made using two-way analysis of variance (ANOVA.) to 
analyze changes in NFkappaB following dapagliflozin treatment in comparison to placebo 
(primary efficacy analysis). Secondary efficacy analysis will include the effect of 
dapagliflozin (versus placebo) on other markers monocyte inflammation. Pre and post 
treatment within a group will be analyzed using paired t-tests. Mann Whitney will be used for 
data which is not normally distributed. All statistical analysis will be performed using SAS 
(Cary, NC). 
12.1.2 
Safety analysis set 
The safety analysis set will include all consented patients regardless of whether or not primary 
outcome measures were observed. This data set will include all available data for patients who 
completed the study as well as those who were lost to follow-up, withdrew from the study, or 
otherwise did not complete the study. 
Adverse Events 
Adverse events will be coded using the current version of the Medical Dictionary for 
Regulatory Activities (MedDRA). Subject incidences of treatment-emergent adverse events, 
serious adverse events, treatment-related adverse events and adverse events leading to 
discontinuation of IP will be tabulated by system organ class and preferred term for each 
treatment group.  
Safety Laboratory Parameters  
Laboratory parameters will be summarized for each treatment group using descriptive 
statistics at each scheduled visit.  
Vital Signs  
Vital signs will be summarized for each treatment group using descriptive statistics at each 
scheduled visit.  
Concomitant medications of interest will be summarized for each treatment group.  
12.2 
Methods of statistical analyses 
Between groups comparisons will be made using two-way analysis of variance (ANOVA.) to 
analyze changes in NFkappaB following Dapagliflozin treatment in comparison to placebo 
(primary efficacy analysis). Secondary efficacy analysis will include the effect of 
Dapagliflozin (versus placebo) on other markers monocyte inflammation. Pre and post 
treatment within a group will be analyzed using paired t-tests. Mann Whitney will be used for 
data which is not normally distributed. All statistical analysis will be performed using SAS 
(Cary, NC). 
12.2.1 
Interim analyses 
None planned 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
31(35) 
12.3 
Determination of sample size 
This trial will be conducted as pilot study, as no prior studies have examined the effects of 
SGLT-2 inhibitor therapy on NFkB and other inflammatory mediators in humans with T2DM. 
Moreover, no prior studies have examined the effect of SGLT-2 inhibitor therapy on 
endothelial function in this population. We anticipate a 20% difference in NFkappaB protein 
between placebo and active treatment groups and 18 subjects in each study arm will yield 
significant differences between therapy and placebo in regard to our primary endpoint. Sample 
size is calculated using SigmaPlot software (San Jose, CA). For the two major outcome 
variables i.e. monocyte NFkappaB protein, the sample size of ~ 18 patients per treatment 
group (total of 36 patients) will have >90% power to detect a difference in the means of 20%, 
assuming a standard deviation of 15%, and with alpha= 0.025. Given that we expect about 
10% of the patients enrolled may not complete the 3 month study, we would like to enroll 40 
patients so that 36 patients may complete the study.  
12.4 
Data monitoring committee  
The study will be monitored by the PI on a daily basis. A Data Safety Monitoring Plan and 
annual reports will be submitted to the Baylor College of Medicine IRB. A Data Monitoring 
Committee will be constituted to monitor the study if directed by the BCM IRB.. 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
32(35) 
13.IMPORTANT MEDICAL PROCEDURES TO BE FOLLOWED BY 
THE INVESTIGATOR 
13.1 
Overdose  
If an overdose on an AstraZeneca study drug occurs in the course of the study, then 
investigators or other site personnel inform appropriate AstraZeneca representatives within 
one day, ie, immediately but no later than the end of the next business day of when he or she 
becomes aware of it. 
The designated AstraZeneca representative works with the investigator to ensure that all 
relevant information is provided to the AstraZeneca Patient Safety data entry site. 
13.2 
Pregnancy 
All outcomes of pregnancy should be reported to AstraZeneca. Women of child-
bearing potential must not be pregnant or nursing to participate in this study.  
However, in the event a woman became pregnant we must ensure patient safety. Each 
pregnancy in a patient on study treatment must be reported to AstraZeneca within 24 
hours of learning of its occurrence. The pregnancy should be followed up to determine 
outcome, including spontaneous or voluntary termination, details of the birth, and the 
presence or absence of any birth defects, congenital abnormalities, or maternal and/or 
newborn complications. 
 
13.2.1 
Maternal exposure 
If a pregnancy occurs in a female subject, while the subject is taking protocol-required 
therapies, it will be reported to AstraZeneca as specified above. In addition to reporting any 
pregnancies occurring during the study, investigators should monitor for pregnancies that 
occur after the last dose of protocol-required therapies and for an additional 15 weeks after the 
end of treatment with IP and report the pregnancy to AstraZeneca as specified above. 
13.2.2 
Paternal exposure 
If a pregnancy occurs in a female partner of a male subject, while the subject is taking 
protocol-required therapies, it will be reported to AstraZeneca as specified above. In addition 
to reporting any pregnancies occurring during the study, investigators should monitor for 
pregnancies that occur after the last dose of protocol-required therapies and for an additional 
15 weeks after the end of treatment with IP, report the pregnancy to AstraZeneca as specified 
above. 
 
 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
33(35) 
 
14.LIST OF REFERENCES 
 1. Feve B, Bastard JP. The role of interleukins in insulin resistance and type 2 diabetes 
mellitus. Nat Rev Endocrinol. 2009;5:305-11.  
2. Barma P, Bhattacharya S, Bhattacharya A, et al. Lipid induced overexpression of NF-
kappaB in skeletal muscle cells is linked to insulin resistance. Biochim Biophys Acta. 
2009;1792:190-200.  
3. DeFronzo RA, Jacot E, Jequier E, Maeder E, Wahren J, Felber JP. The effect of insulin on 
the disposal of intravenous glucose. results from indirect calorimetry and hepatic and femoral 
venous catheterization. Diabetes. 1981;30:1000-7.  
4. Boden G. Fatty acid-induced inflammation and insulin resistance in skeletal muscle and 
liver. Curr Diab Rep. 2006;6:177-81. 
 5. Green CJ, Pedersen M, Pedersen BK, Scheele C. Elevated NF-kappaB activation is 
conserved in human myocytes cultured from obese type 2 diabetic patients and attenuated by 
AMP-activated protein kinase. Diabetes. 2011;60:2810-9.  
6. Andreasen AS, Kelly M, Berg RM, Moller K, Pedersen BK. Type 2 diabetes is associated 
with altered NF-kappaB DNA binding activity, JNK phosphorylation, and AMPK 
phosphorylation in skeletal muscle after LPS. PLoS One. 2011;6:e23999.  
7. Reyna SM, Ghosh S, Tantiwong P, et al. Elevated toll-like receptor 4 expression and 
signaling in muscle from insulin-resistant subjects. Diabetes. 2008;57:2595-602.  
8. Eringa EC, Serne EH, Meijer RI, et al. Endothelial dysfunction in (pre)diabetes: 
Characteristics, causative mechanisms and pathogenic role in type 2 diabetes. Rev Endocr 
Metab Disord. 2013;14:39-48.  
9. Beckman JA, Creager MA, Libby P. Diabetes and atherosclerosis: Epidemiology, 
pathophysiology, and management. JAMA. 2002;287:2570-81.  
10. Macdonald FR, Peel JE, Jones HB, et al. The novel sodium glucose transporter 2 inhibitor 
dapagliflozin sustains pancreatic function and preserves islet morphology in obese, diabetic 
rats. Diabetes Obes Metab. 2010;12:1004-12. 
 11. Tahara A, Kurosaki E, Yokono M, et al. Effects of SGLT2 selective inhibitor ipragliflozin 
on hyperglycemia, hyperlipidemia, hepatic steatosis, oxidative stress, inflammation, and 
obesity in type 2 diabetic mice. Eur J Pharmacol. 2013;715:246-55.  


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
34(35) 
12. Chen L, Klein T, Leung PS. Effects of combining linagliptin treatment with BI-38335, a 
novel SGLT2 inhibitor, on pancreatic islet function and inflammation in db/db mice. Curr Mol 
Med. 2012;12:995-1004. 
 13. Thomas L, Grempler R, Eckhardt M, et al. Long-term treatment with empagliflozin, a 
novel, potent and selective SGLT-2 inhibitor, improves glycaemic control and features of 
metabolic syndrome in diabetic rats. Diabetes Obes Metab. 2012;14:94-6.  
14. Katsuno K, Fujimori Y, Ishikawa-Takemura Y, Isaji M. Long-term treatment with 
sergliflozin etabonate improves disturbed glucose metabolism in KK-A(y) mice. Eur J 
Pharmacol. 2009;618:98-104.  
15. Yamamoto K, Uchida S, Kitano K, et al. TS-071 is a novel, potent and selective renal 
sodium-glucose cotransporter 2 (SGLT2) inhibitor with anti-hyperglycaemic activity. Br J 
Pharmacol. 2011;164:181-91.  
16. Merovci A, Solis-Herrera C, Daniele G, et al. Dapagliflozin improves muscle insulin 
sensitivity but enhances endogenous glucose production. J Clin Invest. 2014;124:509-14.  
17. Ferrannini E, Muscelli E, Frascerra S, et al. Metabolic response to sodium-glucose 
cotransporter 2 inhibition in type 2 diabetic patients. J Clin Invest. 2014;124:499-508.  
18. Corretti MC, Anderson TJ, Benjamin EJ, et al. Guidelines for the ultrasound assessment of 
endothelial-dependent flow-mediated vasodilation of the brachial artery: A report of the 
international brachial artery reactivity task force. J Am Coll Cardiol. 2002;39:257-65.  
19. Uehata A, Lieberman EH, Gerhard MD, et al. Noninvasive assessment of endothelium-
dependent flow-mediated dilation of the brachial artery. Vasc Med. 1997;2:87-92. 
 
 
 
 
 
 
 
 
 


Clinical Study Protocol 
Drug Substance Dapaglifozin 
Study Number H-35985 
Edition Number 1.3 
Date 12 April, 2017 
35(35) 
 
 
 
 
 
 
 
Table 1: Time and Event Schedule for Visit Procedures 
 
 
 
 
 
Procedure                        
 
Visit 1 
(Screeni
ng) 
 
Visit 2 
 
 
Visit 3 
(randomiza
tion) 
 
Visit 4 
 
 
Visit 
5 
 
Visit 
6 
 
 
Visit 
7 
Study week                                 
-2 
-1 
0 
2 
4 
8 
12 
Obtain Informed Consent             
X 
 
 
 
 
 
 
Confirm Eligibility 
X 
 
 
 
 
 
 
 Medical History 
X 
 
 
 
 
 
 
Review Concomitant  
Medications 
X 
 
X 
X 
X 
X 
X 
Physical Examination 
X 
 
 
X 
X 
X 
X 
Vital Signs 
X 
X 
X 
X 
X 
X 
X 
CBC and 12-Lead Electrocardiogram 
X 
 
 
 
 
 
 
Pregnancy Test (a) 
X 
 
 
X 
X 
X 
X 
Urinalysis 
X 
 
 
 
 
 
 
Lipids, LFTs,  
 
X 
 
 
 
 
 
 
BMP 
X 
 
 
X 
X 
X 
X 
HbA1c 
X 
 
 
 
 
 
X 
Insulin/Cytokines/FFA/ 
 
X 
 
 
 
 
X 
Monocyte isolation studies/ 
Flow mediated dilatation by 
Ultrasound 
 
X 
 
 
 
 
X 
Randomize 
 
 
X 
 
 
 
 
Study Drug 
METFO
RMIN 
 
Start dapa 5 
mg or 
placebo 
Dapa10 
mg or 
placebo 
X 
X 
STOP 
Medication compliance check  
 
 
 
X 
X 
X 
X 
Assess for adverse events  
and Home Blood Glucose Monitoring 
Log 
 
 
X 
      X 
X 
X 
X