Clinical trial NCT03260881
Conditions: Type2 Diabetes Coronary Artery Disease
Keywords: Epicardial Fat
Sponsor: University of Miami
Start date: 2018-09-01; Completion date: 2024-09-05
Enrollment: 38 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Epicardial adipose tissue (EAT) is the visceral fat of the heart. EAT could locally affect the coronary arteries through local secretion of pro-inflammatory cytokines. EAT plays a role in the development of the coronary artery disease (CAD). EAT is a highly enriched with genes involved in inflammation. Given its rapid metabolism and simple measurability, as first developed by Iacobellis, EAT serves as target for medications targeting the fat. Glucagon-like peptide-1 agonists (GLP-1A) are anti-diabetic medications with recently suggested cardio-protective properties. Liraglutide, a GLP-1A, has recently shown to reduce the cardiovascular risk. Iacobellis'group found that EAT thickness decreased by an unprecedented 36% after 12 weeks of treatment with liraglutide. Remarkably, Iacobellis'group found for the first time that human EAT express GLP-1 Receptor (GLP-1R). GLP-1A effects may be therefore visceral fat specific and target EAT. Based on these preliminary data, we hypothesize that treatment with liraglutide will significantly and rapidly reduce EAT inflammation. Decreased EAT inflammation can reduce the burden of the coronary plaques. We will test our hypothesis in a 12-week randomized, double-blind, placebo-controlled, interventional study in 40 patients with type 2 diabetes mellitus (T2DM), and CAD, with an acceptable glycemic control on their current diabetes regimen who require elective coronary artery bypass graft (CABG) regardless of their participation in the study. A minimum time frame of 3-week treatment will be considered to detect significant changes in the study endpoints. Inclusion criteria for body fat markers will rule out the confounding effect of different body fast distribution at baseline. Study subjects will be randomized in two groups of 20 patients to receive additional liraglutide or to remain on current treatment/ placebo prior to cardiac surgery. CAD subjects not allocated to liraglutide will be started on a supervised low-calorie diet (LCD) to achieve approximately 5% of weight loss after from a minimum of 3 weeks up to 12 weeks to avoid the confounding effect of weight loss on the study outcomes. Fat samples will be collected during cardiac surgery after up to 12 weeks of treatment either with liraglutide or placebo and processed for analysis of mRNA and protein expression of EAT and SAT inflammatory genes such as Tumor Necrosis Factor-alpha (TNF-α) and Interleukin 6 (IL-6), and GLP-1R.

Detailed description:

Arms: arm 1: • L-group will be started on liraglutide. Liraglutide will be started and administered for from a minimum of 4 weeks up to 12 weeks prior to CABG with a starting dose of 0.6 mg (after a least one week) and subsequent increments to 1.2 mg (after a least one week) and to 1.8 mg (after at least a week on 1.2 mg). The dose of 1.8 mg daily will be maintained until the end of the 12-week study. Other and current diabetes treatment will be continued arm 2: placebo will be administered in addition to current treatment prior to the CABG with a starting dose of 0.6 mg (after a least one week) and subsequent increments to 1.2 mg (after a least one week) and to 1.8 mg (after at least a week on 1.2 mg). D-group will be started on a supervised low calorie diet (LCD) to achieve approximately 5% of weight loss after from a minimum of 4 weeks up to 12 weeks.
Interventions: intervention 1: Liraglutide Pen Injector [Victoza] intervention 2: matching liraglutide-placebo pre-filled pens
Intervention descriptions: intervention 1: Study subjects will be randomized in two groups of 20 patients to receive additional liraglutide, (L-group) or to remain on current treatment or placebo (D-group). intervention 2: Study subjects will be randomized in two groups of 20 patients to receive additional liraglutide, (L-group) or to remain on current treatment or placebo (D-group).

Protocol documents:
Prot_SAP_000.pdf: 
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Protocol Title 
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Effects of Liraglutide on Epicardial Fat Pro-Inflammatory Genes 
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in Type 2 Diabetes and Coronary Artery Disease 
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INVESTIGATOR-SPONSORED STUDY PROPOSAL
 NCT03260881
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U1111-1190-9987
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Gianluca Iacobellis MD, PhD
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Professor of Medicine, 
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Director of University of Miami Hospital Diabetes Service,
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Division of  Endocrinology, Diabetes and Metabolism, Department of 
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Medicine, University of Miami, Miller School of Medicine,
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Miami, FL, USA
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Correspondence to:
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Gianluca Iacobellis MD PhD
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Professor of Medicine
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Division of Endocrinology, Diabetes and Metabolism
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University of Miami, Miller School of Medicine
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1400 NW 10th Ave, 
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Dominion Tower suite 805-807
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Miami, FL, 33136 
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USA 
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e-mail: giacobellis@med.miami.edu
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FAX number:  305 243 9487
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Abstract
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Epicardial adipose tissue (EAT) is the visceral fat of the heart. EAT could locally affect 
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the coronary arteries through local secretion of pro-inflammatory cytokines. EAT plays a 
38
role in the development of the coronary artery disease (CAD). EAT is a highly enriched 
39
with genes involved in inflammation.  Given its rapid metabolism and simple 
40
measurability, as first developed by Iacobellis, EAT serves as target for medications 
41
targeting the fat. Glucagon-like peptide-1 agonists (GLP-1A) are anti-diabetic medications 
42
with recently suggested cardio-protective properties.  Liraglutide, a GLP-1A, has recently 
43
shown to reduce the cardiovascular risk.  Iacobellis‘group found that EAT thickness 
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decreased by an unprecedented 36% after 12 weeks of treatment with liraglutide. 
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Remarkably, Iacobellis’group found for the first time that human EAT express GLP-1 
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Receptor (GLP-1R). GLP-1A effects may be therefore visceral fat specific and target EAT. 
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Based on these preliminary data, we hypothesize that treatment with liraglutide will 
48
significantly and rapidly reduce EAT inflammation. Decreased EAT inflammation can 
49
reduce the burden of the coronary plaques. We will test our hypothesis in a 12-week 
50
randomized, double-blind, placebo-controlled, interventional study in 40 patients with type 
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2 diabetes mellitus (T2DM), and CAD, with an acceptable glycemic control on their current 
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diabetes regimen who require elective CABG regardless of their participation in the study. 
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A minimum time frame of 3-week treatment will be considered to detect significant 
54
changes in the study endpoints. Inclusion criteria for body fat markers will rule out the 
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confounding effect of different body fast distribution at baseline. Study subjects will be 
56
randomized in two groups of 20 patients to receive additional liraglutide or to remain on 
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current treatment/ placebo prior to cardiac surgery. EAT samples will be collected during 
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cardiac surgery and processed for analysis of mRNA and protein expression of EAT 
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inflammatory genes such as Tumor Necrosis Factor-alpha (TNF-α) and Interleukin 6 (IL-
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6), and GLP-1R. 
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Table of Contents
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Background
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EAT Anatomy and Physiology: EAT is the visceral fat depot of the heart. 1-3 EAT and 
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intra-abdominal fat have the same embryogenesis and both evolve 
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from brown fat (BAT). EAT is supplied by branches of the 
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coronary arteries and no muscle fascia separates the fat depot and 
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the myocardium (as depicted in the microscopic figure of human 
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EAT).  Hence, as the two tissues share the same microcirculation, 
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a direct cross-talk between the EAT and the myocardium has been 
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highly suggested. Under physiological conditions EAT could serve as a buffer, absorbing 
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fatty acids and protecting the heart against high fatty acids levels, as a lipid storage and 
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local energy source channelling fatty acids to the myocardium and as BAT to defend the 
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myocardium against hypothermia. Under pathological conditions EAT releases factors that 
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promote harmful coronary artery and myocardial changes. EAT is an extremely active 
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organ that produces both pro-inflammatory, such as TNF-α, IL-6, and anti-inflammatory 
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adipokines. 1-3   Given its anatomical proximity to the heart, EAT may interact locally and 
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modulate the myocardium and coronary arteries through paracrine or vasocrine secretion 
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of bioactive molecules.  EAT transcriptome is also unique when compared to subcutaneous 
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fat. EAT is a highly inflammatory tissue enriched with genes involved in inflammation, 
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endothelial function, coagulation, immune signalling and apoptosis.4 
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EAT and Coronary Artery Disease: EAT plays a significant role in the development and 
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progression of CAD.5-6 EAT could alter the coronary arteries via multiple pathways 
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including macrophage activation, inflammatory response, oxidative stress and plaque 
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destabilization.  Epicardial adipocytes display an intrinsic pro inflammatory and 
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atherogenic profile. A dense inflammatory infiltrate, mainly represented by macrophages, 
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is commonly detected in EAT of subjects with CAD. Regardless of the pathway, 
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inflammatory cells secreted by the EAT surrounding the adventitia may stimulate the 
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proliferation of vasa vasorum and ultimately cause intramural changes. EAT has been 
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largely associated with the severity of CAD and its association with CAD was confirmed 
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in several large population studies. The relationship of EAT thickness and coronary artery 
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disease is driven by local mechanisms and partially independent of coronary calcification. 
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EAT as therapeutic target: EAT thickness can be visualized and measured with standard 
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echocardiography, as first developed and validated by Iacobellis. 7-8 (echo-free space 
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within the red dot from the parasternal view). In addition to 
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the easy accessibility and excellent reproducibility, as 
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reported by the majority of the studies using this technique, 
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echocardiographic EAT independently reflects the intra-
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abdominal visceral fat, measured with Magnetic resonance 
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imaging (MRI) and the intra-myocardial lipid content, 
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calculated 
with 
MR-spectroscopy.9 
Given 
its 
rapid 
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metabolism, organ fat specificity and simple objective measurability, EAT can serve as 
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target for medications targeting the adipose tissue.10-15
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Cardiovascular effects of Liraglutide:  liraglutide, an analogue of glucagon-like peptide-
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1 (GLP-1), is indicated for the treatment of type 2 diabetes mellitus. In addition to its well 
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established glucose-lowering effect, liraglutide causes a modest weight loss. Liraglutide 
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has also been shown to provide cardio-protective effects beyond the glycemic control, 
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although the mechanisms are still unclear. Very recently, the Liraglutide Effect and Action 
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in Diabetes: Evaluation of Cardiovascular Outcome Results (LEADER) trial showed that 
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patients treated with liraglutide had a lower risk of first occurrence of cardiovascular death, 
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nonfatal myocardial infarction, nonfatal stroke and lower risks of death from 
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cardiovascular causes in comparison to placebo.16
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Preliminary Data 
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Liraglutide 
targeting 
EAT: 
GLP-1 
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activation targeting EAT has been recently 
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evaluated.11-13 Our group recently evaluated 
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the effect of liraglutide on EAT in a 24-week 
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interventional 
case-controlled 
study 
in 
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overweight/obese subjects with T2DM on 
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metformin monotherapy.11 Individuals were 
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randomized in two groups to receive 
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additional liraglutide up to 1.8 mg sc once 
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daily or to remain on Metformin. Our results showed that ultrasound measured EAT 
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thickness decreased from 10.2±2 to 6.9±1.9 and 5.8±1.9 mm (p< 0.001) after 12 and 24 
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weeks, respectively, accounting for approximately 40% of reduction at 12 and 24 weeks 
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(graphic on the right), whereas there was no significant EAT reduction in the Metformin 
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group.  EAT shrunk independently of overall weight loss and improved glucose control. A 
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milder, as compared to our results, yet noticeable (-13%), reduction of EAT thickness was 
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recently observed after 12 weeks of treatment either with liraglutide or exenatide in a 
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smaller group of patients with type 2 diabetes. 12 However, EAT results in this study were 
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pooled together making difficult to discriminate differences between the two agents.
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EAT as pro-inflammatory organ: EAT of CAD subjects is 
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enriched with pro-inflammatory genes, as our recent microarray 
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analysis (microarray table on the left) showed. 4 Pro-
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inflammatory cytokines are secreted and transported from EAT 
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into the coronary lumen through vasocrine and paracrine 
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pathways. 1-3
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10.5 
mm
6.3 mm
5.8 mm
base
3 months
6 months
EAT changes with LIRAGLUTIDE
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We recently showed that diabetic EAT was mainly enriched in 
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inflammatory genes, such Interleukin-6 (IL-6) (graphic on the right) 
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and Tumor Necrosis Factor-alpha (TNF-α).17 Our study suggests a 
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unique and novel atherogenic pathway in diabetes mediated by EAT 
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inflammatory profile.
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EAT expresses GLP-1 receptor (GLP-1R)
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Whether EAT expressed GLP- 1 receptor (GLP-1R) was unknown. 
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We recently performed a RNA-sequencing (RNA-seq analysis) and 
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Quantitative real-time RT-PCR (qRT-PCR) to evaluate the presence 
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of GLP-1R in EAT obtained from 8 subjects with coronary artery 
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disease and type 2 diabetes mellitus undergoing elective cardiac 
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surgery. Immunofluorescence was also performed on EAT and SAT 
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samples using Mab 3f52 against GLP-1R. RNA-seq analysis showed 
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that EAT expresses GLP-1R gene. qRT-PCR analysis confirmed 
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GLP-1R expression by using two different sets of intron-spanning 
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primers (graphic on the right). Immunofluorescence showed that 
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GLP-1R is present and more abundant in EAT than SAT 
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(immunofluorescence images). 18 
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In the upper quadrants (A), the immunofluorescence images (starting from the left) show definite higher signal in GLP-1R antibody-
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treated EAT, irrespective of diabetes, when compared to GLP-1R antibody-treated SAT and to not treated with primary antibody EAT 
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(last image on the right). Lower quanrants (B) images show the amplification of the GLP-1R signal using biotinylated anti mouse IgG, 
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horseradish peroxidase-streptavidin and alexa fluor 647 tyramide. DAPI was used to label the nucleus and autofluorescence of the tissue 
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was collected at 488nm excitation.  
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So, for the first time, we found that human EAT express GLP-1R. GLP-1A effects may be 
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therefore visceral fat specific and target EAT. However, the mechanisms mediating the 
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actions of GLP-1 analogues on adipose tissue are still unclear. Liraglutide may improve 
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EAT insulin sensitivity and microcirculation, stimulate EAT thermogenesis and adipocyte 
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browning.19-22
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Short term effects of Liraglutide on inflammation
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The experimental study by Shiraki et al showed a rapid effects of liraglutide in reducing 
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TNF-α-induced oxidative stress and inflammation in endothelial cells (25). A rapid anti-
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inflammatory effects of liraglutide on endothelial cells was also reported by Krasner et al  
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Liraglutide reduced the inflammatory responses to TNFα and LPS stimulation, with a 
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significant reduction of protein expression of the adhesion molecules VCAM-1 and E-
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Selectin, and THP-1 monocyte adhesion in cultured human aortic endothelial cells (26). 
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Additionally, we provided evidence that epicardial fat expresses the GLP-1R protein and 
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mRNA, such as the heart. Hence, liraglutide is more likely to have a direct effect on 
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epicardial fat transcriptome than mediated by weight loss or glycemic control. 
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The LEAD trials showed a decrease in mean weight ranging from 1 to 3.2 kg (2.2 to 7.04 
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pounds) over the course of 26 or 52 weeks with liraglutide.
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Significance We believe that this study proposal may have a great relevance and impact 
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in the research and clinical management of CAD. From a pathophysiological perspective, 
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this study proposal can provide data that will contribute the understanding of the complex 
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mechanisms that lead to the development and progression of CAD. Understanding whether 
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EAT plays a metabolic role in CAD is key. This study can provide evidences of the recently 
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suggested cardioprotective mechanisms of Liraglutide. Treatment with Liraglutide and 
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other GLP1-A may reduce EAT inflammation and revert EAT to its physiological function 
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and therefore to a cardioprotective role. From a clinical perspective, this study proposal 
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can provide data that will open new avenues and have an important impact in the 
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management of CAD. Liraglutide may become an effective treatment of patients with CAD 
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and diabetic cardiovascular complications. EAT can be easily measured, as we first 
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developed, and then targeted by Liraglutide, other GLP-1A or other anti-inflammatory 
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drugs in patients with CAD.   
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Innovation This proposal contains a number of novel concepts with important clinical 
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applications. For the first time this project will evaluate the idea that GLP-1 activation may 
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reduce EAT inflammation in subjects with T2DM and CAD. This is will be the first time 
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that liraglutide effects will be evaluated on EAT samples obtained from subjects with 
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T2DM and CAD. These important points will be addressed in well-studied population of 
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subjects, as was never done before. Based on pre-existing data, the expected liraglutide 
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effects on the epicardial adipose tissue inflammatory genes, primary endpoint, may not 
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necessarily be time dependent. The shorter term of treatment that may occur in some 
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patients may actually rule out the potential confounding effect of the liraglutide-associated 
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weight loss on the study outcomes
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Study Rationale Liraglutide, may have properties that go beyond the glucose-lowering 
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effects, as the recent LEADER trial showed that Liraglutide reduces the risk of major 
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cardiovascular events. Hence, our timely proposal will help the understanding of the 
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mechanisms of the cardio-protective effects of Liraglutide. As our preliminary data are 
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showing that a) EAT is highly inflammatory fat depot affecting the coronary arteries, b) 
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Liraglutide causes a massive and rapid reduction of EAT thickness, c) EAT expresses GLP-
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1R our hypotheses can be likely proven in this study. Our proposal would provide stronger 
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evidences for using GLP-1As in the treatment of CAD. In addition, as EAT is an easily 
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measurable and modifiable risk factor, as we first developed, it can be targeted by 
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Liraglutide. The randomized, double-blind, placebo-controlled study design will allow for 
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an independent and unbiased effect of Liraglutide on study results. In addition, actions to 
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equalize weight loss and maintain similar glucose control between study arms will rule out 
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the confounding effects of these factors on the study outcomes. 
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Specific Objectives
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Primary Objective: 
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
to test the hypothesis that liraglutide can reduce the pro-inflammatory 
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transcriptome profile of EAT of subjects with T2DM and CAD. 
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Secondary Objectives: 
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
to assess the hypothesis that liraglutide will reduce the ultrasound measured EAT 
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thickness in subjects with T2DM and CAD. 
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
to evaluate the hypothesis that liraglutide can produce more anti-inflammatory 
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effects in the EAT as comparted to SAT of subjects with T2DM and CAD. 
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
to test the hypothesis that EAT mRNA and protein expression of GLP-1R will be 
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greater in subjects receiving liraglutide as compared to those continuing their 
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standard therapy
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Research Design and Methods
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Study Hypotheses
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Hypothesis for Primary Objective: We hypothesize that EAT inflammatory gene mRNA 
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and protein expression will be lower in subjects receiving additional therapy with 
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liraglutide as compared to those who will continue with their current treatment. 
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Hypothesis for Second Objectives: based on our biomolecular and clinical preliminary 
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data, we hypothesize that a) EAT thickness will respond and decrease more on liraglutide 
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than on standard therapy, b) inflammatory genes and protein expression will be different 
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in EAT vs SAT c) EAT GLP-1R expression will be greater in patients on liraglutide vs 
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those on standard therapy.
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Endpoints
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Primary Endpoint: 
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
Change from baseline in EAT inflammation (measured as mRNA and protein 
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expression of TNF-α and IL-6) within a minimum of 3 and up to 12 weeks of treatment
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Secondary Endpoints: Change from baseline within a minimum of 3 and up to 12 weeks of 
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treatment for the following parameters:
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Ultrasound measured EAT thickness 
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SAT inflammation (measured as mRNA and protein expression of TNF-α and IL-6) 
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EAT GLP-1R ( measured as mRNA and protein expression of GLP-1R)
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Study Type 
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This is an Investigator Sponsored Study (ISS).
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This will be a 12-week randomized, double-blind, placebo-controlled interventional study 
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in 40 patients with T2DM and CAD with an acceptable glycemic control on their current 
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diabetes regimen who require elective coronary artery bypass grafting (CABG) regardless 
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of their participation in the study. 
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A minimum time frame of 3-week treatment will be considered to detect significant 
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changes in the study endpoints. If not all subjects would be able to complete the 12 
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weeks, study outcomes will be evaluated at the 3 week time point. 
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Study Design Study subjects will be randomized in two groups of 20 patients to receive 
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additional liraglutide, (L-group) or to remain on current treatment or placebo (D-group). 
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
L-group will be started on liraglutide. Liraglutide will be started and administered 
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for 12 weeks prior to CABG with a starting dose of 0.6 mg (after a least one week) 
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and subsequent increments to 1.2 mg (after a least one week) and to 1.8 mg (after 
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at least a week on 1.2 mg). The dose of 1.8 mg daily will be maintained until the 
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end of the 12-week study. Other and current diabetes treatment will be continued. 
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In those study participants who may be impacted by CABG scheduling and 
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COVID-19, and cannot complete the 12-week treatment, we will consider a 
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minimum time frame of 3 weeks to detect significant changes in the study primary 
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endpoints. 
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
D-group: placebo will be administered in addition to current treatment prior to the 
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CABG with a starting dose of 0.6 mg (after a least one week) and subsequent 
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increments to 1.2 mg (after a least one week) and to 1.8 mg (after at least a week 
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on 1.2 mg). Patients will be started on a supervised diet to achieve approximately 
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5% of weight loss between 3 and 12 weeks. In those study participants who may be 
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impacted by CABG scheduling and COVID-19, and cannot complete the 12-week 
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treatment, we will consider a minimum time frame of 3 weeks to detect significant 
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changes in the study primary endpoints.
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Ultrasonographic measurement of EAT thickness will be performed at baseline and 
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after 12 weeks in all patients, regardless of the study arm.
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All groups will receive lifestyle and diabetes education, and supervised diet as 
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part of the standard care, to insure similar glycemic control between arms and 
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weight loss. Study patient will be also required to check and log their finger stick 
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blood glucose twice daily (fasting before breakfast and at bedtime) and report the 
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readings to the study coordinator on a bi-weekly base. 
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Rationale for study Design
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The randomized, double-blind, placebo-controlled study design will allow for an 
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independent and unbiased effect of liraglutide on study results. 
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Rationale for minimum study time line of 3 weeks. 
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The pre-operative clinic schedule may not always allow to wait the full 12 weeks between 
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the patient’s enrollment and the CABG, standard of care procedure. Dr Lamelas is the only 
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cardiac surgeon at UM performing elective CABGs. Once the patient is randomized and 
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randomly allocated to one of the study arms, the length of the treatment is set by the date 
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of standard of care CABGs. During the current COVID-19 pandemic, the number of 
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elective CABGs has been reduced and we cannot reschedule or delay the surgery 
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Study Population: Study group will be formed by 40 patients with T2DM and with 
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clinically and angiographically stable CAD who will undergo CABG surgery, as part of 
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their standard medical care. Cardiac surgery will be elective procedure in 
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hemodynamically stable patients taking their standard cardiac treatments and under the 
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care of the cardiologist.  
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Study team and site: Dr Iacobellis is Professor of Medicine, Division of 
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Endocrinology, Diabetes and Metabolism. Dr Iacobellis pioneered EAT echocardiographic 
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measurement and he is considered the leading expert in the epicardial fat. Dr Iacobellis will 
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lead and supervise liraglutide therapy, EAT measurement, analysis and interpretation of 
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the data. Dr Frasca, Research Assistant Professor, Dept. Microbiology and Immunology, 
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will lead and supervise adipose tissue analysis. Dr Claudia Martinez is associate professor 
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of Cardiology and contributes to patient recruitment. Dr Joseph Lamelas, professor of 
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cardiothoracic surgery will be the cardiac surgeons for this study. Low calorie diet (LCD) 
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and other nutritional aspects will be managed and supervised by the Division of 
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Endocrinology registered dietitian (RD), as part of the standard care.   University of Miami 
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has been the site of several National Health Institute (NIH) studies and many peer-reviewed 
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publications. The Division of Endocrinology Diabetes and Metabolism is fully equipped 
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and trained to visit, follow up and manage patients with diabetes. The Division of 
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Endocrinology provides multiple outpatient diabetes clinics at the Diabetes Research 
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Institute (DRI). Division of Endocrinology clinical laboratory is fully accredited and 
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equipped to perform state-of- the-art diagnostic tests assisting in the diagnosis and 
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management of diabetes.
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Inclusion Criteria 
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T2DM as defined by American Diabetes Association (ADA) criteria 
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Adult patients with T2DM who are indicated to receive liraglutide, not as first-line 
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therapy, in addition to diet and exercise to improve glycemic control 
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Hemoglobin A1c (HbA1c) ≤ 9% 
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Age ≥ 18 years old
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Body mass index (BMI) ≥ 27 Kg/m2 and/or waist circumference ≥ 102 cm (40 
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inches) in men and 88 cm (35 inches) in women, respectively. 
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Clinically and angiographically stable CAD who requires CABG as part of the 
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standard medical care, as CAD does not represent a contraindication for using 
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liraglutide. The stability of the CAD further warranties that study patients will not 
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be exposed to higher risk by using liraglutide.
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Exclusion Criteria 
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Patients with a personal or family history of medullary thyroid carcinoma or 
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patients with Multiple Endocrine Neoplasia syndrome type 2
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Patients with a prior serious hypersensitivity reaction to liraglutide
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Other contra-indications to liraglutide in accordance with risks and safety 
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information included in the latest updated prescribing information
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Type 1 diabetes, as defined by ADA criteria
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Current use of other GLP-1A, dipeptidyl peptidase 4 (DPP4) or Sodium Glucose 
367
transportes 2 (SGLT2) inhibitors, thiazolidinediones (TZDs), pramlintide and fixed 
368
prandial insulin.
369
-
Patients with unstable CAD, assessed by the Cardiology team and defined as new 
370
onset angina, rest angina, rapidly increasing or crescendo angina
371
-
History of diabetic ketoacidosis, pancreas or beta-cell transplantation, or diabetes 
372
secondary to pancreatitis or pancreatectomy; acute or chronic infective diseases, 
373
cancer or chemotherapy, history of pulmonary, renal or liver diseases, and drug 
374
abuse
375
-
Patients with chronic and acute inflammatory conditions such as sepsis, rheumatoid 
376
arthritis, ectopic dermatitis, asthma, ulcerative colitis. 
377
-
Current use of systemic corticosteroids in the 3 months prior this study.  
378
-
Pregnant women 
379
-
Women of childbearing potential who are not using adequate contraceptive 
380
methods (as required by local law or practice) 
381
382
Investinational drug
383
Liraglutide [rDNA origin] injection, solution for subcutaneous use. Liraglutide will be 
384
prescribed according to the same, current indications of the marketed Victoza.
385
386
Withdrawal Criteria
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1. The subject may withdraw at will at any time.
388
2. The subject may be withdrawn from the trial at the discretion of the investigator due to 
389
a safety concern or if judged non-compliant with trial procedures or included in 
390
contravention to the inclusion and/or exclusion criteria.
391
3. Subject diagnosed with acute pancreatitis by clinical and/or radiographic criteria.
392
4.  Pregnancy, positive pregnancy test prior to the enrollment or intention to become 
393
pregnant. 
394
5. 
395
6. Subjects who will not tolerate liraglutide dose increase to 1.2 or 1.8 mg will remain in 
396
the study at the highest tolerated dose
397
Subject Replacement
398
There will be no replacement of subjects in this trial.
399
400
Rationale for Study Population
401
We anticipate that approximately 400 patients will be screened in a 2-year period. We will 
402
recruit patients with T2DM and clinically and angiographically stable CAD who will 
403
undergo CABG surgery, as part of their standard medical care. Cardiac surgery will be 
404
elective procedure in hemodynamically stable patients taking their standard cardiac 
405
treatments and under the care of the cardiologist.  
406
407
Recruitment strategy: Participants will be recruited among the outpatient population who 
408
routinely refer to the Division of Cardiothoracic Surgery (Dr Lamelas and collaborators), 
409
Division of Cardiology (Dr Martinez) and Division of Endocrinology, Diabetes and 
410
Metabolism outpatient clinics (Dr Iacobellis), at University of Miami.
411
Visit Procedures
412
Study Timetable: This study will consist of two clinical visits prior the cardiac surgery 
413
and adipose tissue collection during the surgical intervention. Consecutive subjects with 
414
T2DM and CAD who are candidate for elective cardiac surgery regardless of their 
415
participation in the study will be selected, informed and consented.  Patients suitable to 
416
wait 12 weeks for cardiac surgery will be identified by the cardiology team and accordingly 
417
scheduled, as part of the standard care. Anthropometrics, blood draw for laboratory 
418
measures and echocardiographic measurements of EAT thickness will be performed at 
419
baseline and before cardiac surgery that will be scheduled 12 weeks after study enrollment 
420
and treatment. Adipose tissue collection will be collected during the cardiac surgery.
421
Study outcomes will be collected at 3 weeks in both groups in those patient who may not 
422
be able to complete the 12- week treatment due to CABG logistics and COVID-19 impact 
423
on scheduling 
424
425
Assessments for Efficacy
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Adipose tissue collection: EAT biopsy samples (average 0.5-1.0 g) will be taken, before 
427
heparin administration, near the proximal tract of the right coronary artery. Each tissue 
428
sample will be snap frozen in liquid nitrogen and stored at −80 °C until analysis in a freezer. 
429
The subcutaneous fat tissue (SAT) will be harvested at the site of the thoracic wound and 
430
preserved in the same manner.
431
Gene expression: in freshly isolated mature adipocytes (from EAT and SAT), extraction 
432
of total RNA, quantification of mRNA, c-DNA generation qRT-PCR will be performed for 
433
measuring expression of  the inflammatory genes (TNF-α, IL-6) and GLP-1R. 
434
Western Blot: Total proteins will be extracted from frozen EAT and SAT adipocytes and 
435
blotted for TNF-α, IL6, as previously described.
436
In addition to the mRNA expression of GLP1R, we will also measure protein expression 
437
of GLP-1R by western blotting. Total cell lysates from the EAT will be obtained as follows. 
438
Cells will be resuspended in M-PER (Mammalian Protein Extraction Reagent, Thermo 
439
Scientific), incubated on ice for 20 min, sonicated for a few seconds and then centrifuged 
440
(14,000 rpm, 15 min, 4°C) to obtain protein lysates which will  be stored at -80°C until 
441
use. Protein content will be determined by Bradford assay before western blotting runs. 
442
Antibodies will be anti-GLP1R Mab 3F52 (Novo Nordisk) and anti-GAPDH (GeneTex) 
443
as loading control.
444
Immunofluorescence 
445
To confirm GLP-1R protein expression, immunofluorescence will be performed on EAT 
446
and SAT samples using Mab 3f52 against GLP-1R, as previously described by Pyke et al 
447
23-24. This mouse monoclonal antibody has been extensively validated to show its 
448
specificity. Due to the relatively low abundance of GLP-1R the immunofluorescence will 
449
be performed using a tyramide signal amplification reagent (TSA kit#26 from Molecular 
450
probes) according to the manufacturer instructions. In summary, cryosection of the samples 
451
will be performed at -35 °C with 12 μm thickness using a Leica CM1850 UV cryostat 
452
(Wetzlar, Germany). The sections will be fixed for 20 min with 4% PFA, washed with PBS 
453
and dried at room temperature overnight. Sections will be washed 3 x with PBS and 
454
incubated with PBS with 3% BSA and 0.4% triton X for 1 hour. Sections will then 
455
incubated with Mab 3f52 (1:50 dilution) antibody overnight in PBS with 3% BSA and 
456
0.4% triton X. Sections will be then washed 4 times with PBS and incubated with 
457
biotinylated anti mouse IgG for 1 hour at room temperature, washed 4 times with PBS, 
458
then incubated at room temperature with horseradish peroxidase-streptavidin for another 
459
hour, washed 4 times with PBS and then incubated at room temperature with  alexa fluor 
460
647 tyramide  for 10 min. Sections will be further washed 3 times and incubated with DAPI  
461
(Thermo fisher, Waltham, MA) for 30 min. The samples will be washed 3 times with PBS 
462
and mounted on Mowiol 4-88 (Sigma-Aldrich, St Louis, MO). The tyramide signal can 
463
amplify from 10 to 200 times the immunofluorescence signal from low expressed proteins 
464
(Molecular probes, Eugene, OR). The fluorescence images will be acquired using a Zeiss 
465
LSM 710 confocal microscope (Oberkochen, Germany).
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Echocardiographic EAT Thickness: 
467
Echocardiogram for EAT thickness measurement will be performed at baseline and after 
468
12 weeks in all patients, regardless of the study arm. EAT thickness will be measured 
469
according to the method firstly described and validated by Iacobellis 7-8. Briefly, EAT will 
470
be identified as the echo-free space between the outer wall of the myocardium and the 
471
visceral layer of pericardium. EAT thickness will be measured perpendicularly on the free 
472
wall of the right ventricle at end-systole in three cardiac cycles. Parasternal long views 
473
allow the most accurate measurement of EAT on the right ventricle, with optimal cursor 
474
beam orientation. Maximum epicardial fat thickness will be measured at the point on the 
475
free wall of the right ventricle along the midline of the ultrasound beam, perpendicular to 
476
the aortic annulus, used as anatomical landmark for this view. The average value of three 
477
cardiac cycles will be considered.   
478
Reliability of echocardiographic measurement of EAT
479
Reliability of echocardiographic measurement of EAT will be assessed by the intra-class 
480
correlation coefficient. Inter- and intra-observer reproducibility will be evaluated by the 
481
intra- class correlation coefficient in all subjects. Echocardiograms will be read by Dr 
482
Iacobellis and an experienced cardiac imaging technician. Previously published studies 
483
have shown that intra- and inter-observer reproducibility of epicardial fat measurement was 
484
excellent.  Intra-class correlation coefficients varied from 0.90 to 0.98 and from 0.93 to 
485
0.98, respectively indicating good reproducibility and reliability. Both readers will be 
486
blinded to the subjects’ clinical data. 
487
Anthropometrics and Clinical measures: As standard of care, height (in cm) and weight 
488
(in kg) will be measured, and BMI will be calculated. Waist circumference (in cm or 
489
inches) will be measured as the minimum circumference between the lower rib margin and 
490
the iliac crest. Blood pressure will be also measured. 
491
Blood Measurements:Lipid panel, lipase, and amylase are not study end-points and will 
492
be collected if made available through a standard of care procedure at baseline and after 4 
493
or 12 weeks. 
494
Low calorie Diet (LCD) A low calorie diet program will be prescribed to study patients 
495
and controls. A RD who will reach out to patients to insure adherence to the dietary 
496
program will supervise the LCD. This will help patients to achieve approximately 5% of 
497
weight loss between 3 and 12 weeks. 
498
Assessments for Safety
499
Potential Risks to the Subjects
500
The risks of this study are minimal are mainly related to risks associated with the 
501
investigational drug liraglutide, echocardiogram, blood draw, and a potential breach of 
502
confidentiality.
503
Drug therapy: liraglutide, (Victoza®), is a GLP−1A indicated as an adjunct to diet and 
504
exercise to improve glycemic control in adults with T2DM. Liraglutide is taken once daily 
505
at any time of day, without regard to the timing of meals. Liraglutide is injected 
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subcutaneously in the abdomen, thigh, or upper arm. Liraglutide may cause mild and 
507
usually well-tolerable gastro-intestinal side effects (5%), such as nausea, constipation. 
508
Liraglutide is contraindicated in patients with a personal or family history of medullary 
509
thyroid cancer and in patients with Multiple Endocrine Neoplasia (MEN). Based on 
510
spontaneous postmarketing reports, acute pancreatitis has been observed in patients treated 
511
with liraglutide. Subjects will be instructed to contact the research staff if they develop new 
512
symptoms on the drugs. In case that an acute pancreatitis will be suspected, lab and imaging 
513
tests will be ordered. Marked or severe side effects will necessitate discontinuation of study 
514
drug.
515
Echocardiography: Diagnostic ultrasound imaging has been used in routine medical care 
516
in various forms for over 45 years. Over this period of time, no harmful effects have been 
517
identified from its use. Mild discomfort when the probe is pressed against the chest may 
518
be experienced, but otherwise there are no known risks.
519
Adipose tissue collection: Intra-operative collection of a very small sample of adipose 
520
tissue do not represent a clinical relevant risk to the study subjects 
521
Blood draw: Bruising at the intravenous puncture site may occur which will clear up in a 
522
few days. Rarely the skin or vein at the site can become infected. A total of approximately 
523
12 cc of blood will be drawn during the course of each of the patient visits which poses no 
524
significant risk to a subject who is not severely anemic. 
525
Confidentiality: Patient confidentiality is an important issue in any clinical research study. 
526
All stored blood will be deidentified using established protocols. Subject’s name will not 
527
be used in publications or data analyses nor will they be available for discussion by any 
528
investigators other than the treating physicians.
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PROTECTION AGAINST RISKS
531
The inclusion and exclusion criteria were designed to limit the possibility of adverse 
532
events. Any adverse event will be reported to the sponsor and Institutional Review Board 
533
(IRB). Subjects will be informed that they are free to withdraw at any point during the 
534
study. We will follow safeguards to minimize the risk to our subjects: a) we will carefully 
535
monitor response to medical treatment every 2 weeks by telephone contact, b) as standard 
536
clinical procedure women of reproductive age who are sexually active will undergo a urine 
537
pregnancy tests prior to participation in the study, c) female subjects whom are pregnant, 
538
breast-feeding, or not willing to use appropriate contraception at time of enrollment will 
539
not be included in the study. Participation in a research study introduces some inherent loss 
540
of privacy. However, full measure will be taken to protect the integrity of patient 
541
identifying information. All data used in the analysis and reporting of this evaluation will 
542
be without identifiable reference to the patient. Data used for reports or publications will 
543
never include identifiable information such as name, date of birth, social security number, 
544
address, or medical record number and in most cases will not include any identifiers other 
545
than sex, age, race, and diagnosis. 
546
Scheduling CABG timing
547
In patients with stable CAD found to have CAD anatomy that requires CABG, the timing 
548
and schedule for revascularization will be determined by Dr Lamelas. Patients suitable to 
549
wait between 3 and 12 week- period for revascularization will be identified and managed 
550
with optimal medical therapy, and scheduled for complete revascularization with CABG. 
551
Managing patients becoming unstable or developing acute events during the study
552
The inclusion and exclusion criteria will limit the possibility of recruiting patients with 
553
unstable CAD/angina, as defined before. Study patients showing unstable angina or 
554
developing acute cardiac events will be transferred promptly to the local emergency 
555
department for evaluation and treatment. If patient status becomes acute and emergent, the 
556
adverse event will be reported to the IRB.
557
Unblinding 
558
The investigator will follow the study’s randomization procedures. The blind should 
559
ordinarily be broken for serious and unexpected adverse experiences that are associated 
560
with the use of a drug to determine if there is a reasonable possibility that the experience 
561
may have been caused by the drug. Certain adverse events such as known consequences 
562
of the underlying disease under investigation or events common in the study population 
563
generally should not require unblinding. 
564
We feel that in the not likely case (given the inclusion criteria) that a study patient will 
565
become unstable or develop acute events during the study, this event would more likely 
566
fall in the known consequences of the underlying disease under investigation rather than 
567
caused by the study drug, and therefore should not require premature unblinding. 
568
Nevertheless, it will be upon the principal investigator’s best judgement and discretion to 
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break the blind. In this case, the principal investigator will promptly document and report 
570
to the IRB and Novo Nordisk any premature unblinding.
571
572
Subject Compliance
573
We estimate that the majority of the participants who will be considered eligible will 
574
complete the study. Attrition can be estimated as lower than 10%.  In the event that 
575
participants are lost to follow-up, the research coordinator will contact with the study 
576
participant by telephone. The research coordinator will be trained in effective telephone 
577
technique to maximize recruitment success. All groups will receive lifestyle and diabetes 
578
education, as part of the standard care, to insure similar glycemic control between arms. 
579
Study patients will be advised to monitor their capillary glucose twice daily, fasting in the 
580
morning and post-prandially.  Glucose profile will be tracked bi-weekly by the research 
581
coordinator by email or telephone. 
582
583
Statistical Considerations
584
POWER ANALYSIS
585
The statistical power (two-sided, α=0.05) of the study was calculated to detect statistically 
586
and clinically meaningful differences in the primary endpoint between the two study arms.  
587
Our preliminary data showed that EAT inflammatory genes, including IL-6 and TNF-α are 
588
upregulated in subjects with T2DM and CAD and higher in EAT as compared to SAT 4-17. 
589
From our recently published study, we calculated the average coefficient of variance (CV) 
590
for differential inflammatory genes to be 42%, but for only 5 T2DM patients. 17 As there 
591
are no data on the effects of liraglutide on EAT inflammation, the power analysis was based 
592
on our previous data and on the assumption that the difference in EAT TNF-α and IL-6 
593
expression between liraglutide treated and placebo treated patients would be clinically 
594
meaningful at 33% (approx. 1.3X fold). Based on these data, we propose to sample 40 
595
individuals. There will be 20 on liraglutide plus standard therapy versus 20 subjects on 
596
liraglutide-matching placebo plus standard therapy contributing EAT and SAT samples 
597
collected during cardiac surgery. Using the power set at 90% and α=0.05, and given the 
598
CV of 42%, this sample size will provide adequate power to detect the clinically significant 
599
expected difference (33%) in EAT TNF-α and IL-6 between the two study arms. 
600
Based on pre-existing data, this sample size will provide adequate power to detect 
601
statistically significant differences in EAT TNF-α and IL-6 between the two study arms in 
602
those patients who will receive treatment only for 3 weeks. The difference in EAT TNF-α 
603
and IL-6 between liraglutide treated and placebo treated patients is expected to be 
604
approximately 1X fold after 3 weeks of treatment.
605
606
The larger sample proposed in this current project would likely reduce the variability. To 
607
further assure consistency of results, mRNA and protein expression values will be log-
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transformed prior to statistical analyses unless they will be normally distributed. 
609
Housekeeping genes will be also used for both RT-PCR and western blot analyses. 
610
In those patients who will not complete the 12-week treatment, the analysis of EAT genes 
611
from the samples will be time adjusted
612
613
Statistical power was also calculated to detect statistically significant differences in the 
614
ultrasound measured EAT thickness, secondary endpoint,  between the two study arms Our 
615
group very recently showed that additional therapy with liraglutide for 12 weeks caused a 
616
reduction in ultrasound measured EAT from 9.6±2 to 6.2±1.5 mm (p< 0.001) in 54 patients 
617
with T2DM 11.  Given the reference value and the expected difference in EAT, the 
618
statistical power of the study (two-sided, α=0.05) was calculated. 
619
Outcome
Reference 
valueSD
Expected 
Difference
Detectable difference with 
80% power α=0.05
Detectable 
difference 
with 
>99% power α=0.05
EAT (mm)
9.62
-20-or -30%
-20%  
(absolute difference = 1.9)
-30% 
(absolute difference = 2.88)
620
621
Assuming a difference of approximately 2 mm (20%) in EAT to be clinically significant 
622
(based on our recent and previously published data 10-14), a convenience sample of 20 
623
individuals for each group will therefore provide at least the statistical power (80%) to 
624
detect an expected difference of at least -20% in the EAT before and after treatment with 
625
liragutide. Baseline standard Deviation (SDs) of 2 is expected to lower to approximately 
626
1.5 for the expected change. The expected change of SD will further assure significant 
627
statistical power of the comparative test.
628
629
In those study patients who will not be able to complete the 12-week treatment, we 
630
anticipate to see a 15% of EAT reduction after 3 weeks in patients who will be allocated to 
631
liraglutide. This anticipated 15% of EAT reduction will remain clinically significant. For 
632
example, a EAT thickness of 10 mm may likely decrease to 8.5 mm after 3 weeks of 
633
treatment with liraglutide, a clinically meaningful change for the patient cardio-metabolic 
634
profile.  
635
636
This sample size also considers the difficulties with recruitment in the real world clinics. 
637
Based on our previous studies in humans, some participants may be lost to follow-up. 
638
639
Student’s t-test or non-parametric (Mann–Whitney) tests, depending on whether the genes 
640
mRNA expression and protein present or not a normal distribution, will be used to compare 
641
the effects of treatment (liraglutide versus placebo) on the variables measured.  Student’s 
642
t-test will be also used to compare ultrasound measured EAT thickness before and after 
643
treatments. Continuous variables will be considered as age-adjusted, sex-adjusted and 
644
weight-adjusted means with their SDs or median, if values are skewed.  
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Univariate regression models will be performed to assess the correlation between clinical 
646
variables, including echocardiographic EAT thickness and blood tests, and EAT gene and 
647
protein expression.  Two-tailed p < 0.05 indicates statistical significance. 
648
649
Data Handling and Record Keeping
650
Each of the dedicated clinic exam rooms provides direct access to the clinical research 
651
study electronic data capture (EDC) eVelos system provided by the Clinical Research 
652
Informatics and Data Management Unit of the Center for Health Informatics and 
653
Bioinformatics at University of Miami (UM). The investigator and research coordinator 
654
will collect and manage the data. 
655
Data collection records with personal identifiers will be stored in locked file cabinets, at 
656
Dominion Tower suite 805, 1400 NW 10th Ave, Miami, FL, 33136.  Blood samples drawn 
657
and stored in conjunction with this study will not be labeled with information that could 
658
directly identify study subjects.   Presentation of the study results at scientific meetings or 
659
in publications will not identify subjects.  Access to research and confidential records will 
660
be limited to clinical investigators, research coordinators, and the IRB at UM.
661
Adipose tissue samples storing 
662
Each adipose tissue sample will be snap frozen in liquid nitrogen and stored at −80 °C until 
663
analysis in a freezer, in Dr Frasca’s lab, University of Miami, Dept of Microbiology, 1600 
664
NW 10th Ave Miami FL 33136-1015. Once the sample is taken, it will be unlinked from 
665
patient’s name. This will assure confidentiality and anonymity. Current or future research 
666
on adipose tissue samples will need to be approved by the local IRB.  
667
668
Ethical Considerations 
669
The study has been approved by the UM local institutional review board (IRB). The study 
670
will be conducted in accordance with the Declaration of Helsinki. The study will be 
671
conducted in accordance with the ICH GCP guidelines. The investigator will comply with 
672
all applicable regulatory and legal requirements, ICH GCP guidelines and the Declaration 
673
of Helsinki in obtaining and documenting the informed consent.
674
Informed Consent: Eligible and willing participant will be approached by the 
675
investigators or the research coordinator and consented. Written informed consent form 
676
will be obtained from the subjects before "baseline" activities begin. Each patient will sign 
677
and date the informed consent. As standard procedure, patients will be informed of the 
678
risks and complications related to cardiac surgery, regardless of their participation in the 
679
study. 
680
681
682
Study Schedule
683
Expected milestones: 
684

start of study: 09/01/2017
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
first patient first visit 09/01/2017
686

last patient last visit 10/31/2023
687

Planned completion of integrated final study report 12/2023
688
689
Study Drugs and Materials
690
Trial drugs
691
Trial drugs will be supplied as liraglutide and matching liraglutide-placebo pre-filled pens 
692
for subcutaneous injection, provided by Novo Nordisk. Pre-filled, multi-dose pen for 
693
subcutaneous injection delivers doses of 0.6 mg, 1.2 mg, or 1.8 mg (6 mg/mL, 3 mL). 
694
Labelling of trial drugs: All the pre-filled pens will be labeled “liraglutide/placebo” 
695
Trial drugs supply Trial drugs (liraglutide and matching placebo) will be supplied to each 
696
study subject by Novo Nordisk. 
697
Clinical Supply in Novo Nordisk Headquarter will revert with timeline for delivery of trial 
698
drugs when protocol is approved, and Novo Nordisk Headquarter will receive a signed 
699
contract and other additional information. Standard delivery is 4 months. 
700
Storage and Drug Accountability of Study Medication
701
The investigator will ensure the availability of proper storage conditions and record and 
702
evaluate the temperature. Prior to first use, trial drugs will be stored in a refrigerator 
703
between 36°F to 46°F (2°C to 8°C). After initial use of the trial drug pen, the pen can be 
704
stored for 30 days at controlled room temperature (59°F to 86°F; 15°C to 30°C) or in a 
705
refrigerator (36°F to 46°F; 2°C to 8°C). Trial drugs will be not dispensed to any person not 
706
enrolled in the study. Unused trial drugs will be stored separately from used trial 
707
medication.
708
Randomization and Blinding
709
This will be a double-blind, parallel group, placebo controlled study. The method of 
710
allocation generation will be a computerized random-number generator. The sequence will 
711
be generated by the process of restricted randomization. Computer-based randomization 
712
process will be managed by the UM Research pharmacy. 
713
714
Concomitant Illnesses and Medications
715
Background medications:
716
Metformin.   Metformin is considered background medication (non-investigational 
717
medicinal product) and will not be provided during the trial. Dose adjustments could occur 
718
during the trial at the investigator’s discretion.  As Iacobellis et al 12 recently showed no 
719
significant effects of metformin on EAT, we do not anticipate that metformin dose 
720
adjustment may influence the study outcomes.  
721
Long acting insulins. Insulin is considered background medication (non-investigational 
722
medicinal product) and will not be provided during the trial.  The total daily dose of insulin 
723
could be adjusted during the trial at the investigator’s discretion.  Iacobellis’ group recently 
724
performed a 24-week interventional study to compare EAT changes in insulin-naïve 
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inadequately controlled patients with type 2 diabetes following basal insulin initiation with 
726
detemir vs. glargine. Not significant changes in EAT between detemir and glargine were 
727
observed. 21 Based on these findings we do not anticipate that insulin treatment may 
728
influence the study outcomes.  
729
Sulfonylureas.  Sulfonylurea treatment is considered background medication (non-
730
investigational medicinal product) and will not be provided during the trial.  During the 
731
study, no up-titration of sulfonylurea dosage will be allowed.  Dose reduction of SU due to 
732
hypoglycemia may be allowed at the investigators discretion.  In the event of 
733
hypoglycemia, the dose of sulfonylurea can be reduced or the drug can be stopped at the 
734
investigator’s discretion.   
735
Adverse Events
736
In the case of an adverse event (AE), the investigator will comply with all local legal, 
737
regulatory, and IRB requirements. The investigator will report to IRB all adverse events 
738
including not serious and serious adverse events (SAE), suspected unexpected serious 
739
adverse reactions (SUSARs), serious adverse drug reactions (SADRs).  The investigator 
740
will report all SAEs, SUSARs, and SADRs at the same time such events are reported to 
741
regulatory authorities or within 15 days from the investigator becoming aware of such 
742
adverse events, whichever comes first.
743
The investigator will use the approved Updated Prescribing Information for Victoza®. The 
744
investigator will collect the following information at minimum for each of these events: 1. 
745
Study name2. Patient identification 3. Event (preferably a diagnosis) 4. Drug 5. Reporter 
746
identification. 6. Causality 7. Outcome 
747
Definitions An AE is any undesirable medical event occurring to a subject in a clinical 
748
trial, whether or not related to the trial product(s). This includes events reported from the 
749
first trial related activity after the subject has signed the informed consent and until post 
750
treatment follow-up period as defined in the protocol. The following should not be recorded 
751
as AEs, if recorded as medical history/concomitant illness at screening:• Pre-planned 
752
procedure, unless the condition for which the procedure was planned has    worsened from 
753
the first trial related activity after the subject has signed the informed consent • Pre-existing 
754
conditions found as a result of screening procedures
755
Clinical Laboratory Adverse Event: A clinical laboratory AE is any clinical laboratory 
756
abnormality regarded as clinically significant i.e. an abnormality that suggests a disease 
757
and/or organ toxicity and is of a severity, which requires active management, (i.e. change 
758
of dose, discontinuation of trial product, more frequent follow-up or diagnostic 
759
investigation).
760
Serious Adverse Event (SAE): A serious AE is an experience that at any dose results in 
761
any of the following:
762
• Death
763
• A life-threatening* experience
764
• In-patient hospitalization or prolongation of existing hospitalization
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 Suspicion of transmission of infectious agents must always be considered an SAE
766
• A persistent or significant disability/incapacity
767
• A congenital anomaly/birth defect
768
• Important medical events that may not result in death, be life-threatening*, or require 
769
hospitalisation may be considered an SAE when, based upon appropriate medical 
770
judgement, they may jeopardise the subject and may require medical or surgical 
771
intervention to prevent one of the outcomes listed in this definition
772
*The term life-threatening in the definition of SAE refers to an event in which the subject 
773
was at risk of death at the time of the event. It does not refer to an event which 
774
hypothetically might have caused death if it was more severe.
775
SADR: an adverse drug reaction (ADR) is an adverse event for which a causal relationship 
776
(Possible/Probable relation) between the study drug and the occurrence of the event is 
777
suspected. The ADR should be classified as serious if it meets one or more of the 
778
seriousness criteria. 
779
Medical Events of Special Interest (MESI): A MESI is (1) a medication error (e.g. wrong 
780
drug administration or wrong route of administration) or (2) a suspected transmission of an 
781
infectious agent via the product
782
Non-Serious Adverse Event: A non-serious AE is any AE which does not fulfil the 
783
definition of an SAE. Severity Assessment Definitions:
784
• Mild: Transient symptoms, no interference with the subject’s daily activities
785
• Moderate: Marked symptoms, moderate interference with the subject’s daily activities
786
• Severe: Considerable interference with the subject’s daily activities, unacceptable
787
Relationship to study medication Assessment Definitions:
788
• Probable: Good reasons and sufficient documentation to assume a causal relationship
789
• Possible: A causal relationship is conceivable and cannot be dismissed
790
• Unlikely: The event is most likely related to an etiology other than the trial product
791
 
792
Outcome Categories and Definitions: • Recovered: Fully recovered or by medical or 
793
surgical treatment the condition has returned to the level observed at the first trial related 
794
activity after the subject signed the informed consent• Recovering: The condition is 
795
improving and the subject is expected to recover from the event. This term should only be 
796
used when the subject has completed the trial • Recovered with sequelae: As a result of the 
797
AE, the subject suffered persistent and     significant disability/incapacity (e.g. became 
798
blind, deaf, paralysed). Any AE recovered with sequelae should be rated as an SAE• Not 
799
recovered• Fatal• Unknown
800
Collection, Recording and Reporting of Adverse Events: All events meeting the 
801
definition of an adverse event will be collected and reported from the first trial related 
802
activity after the subject has signed the informed consent and until the end of the post-
803
treatment follow-up period as stated in the protocol. 
804
Follow-up of Adverse Events: During and following a subject’s participation in a clinical 
805
trial, the investigator and institution will provide adequate medical care to the study subject 
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806
for any study-related adverse events, including clinically significant laboratory values 
807
related to the study. This medical care for study subjects will be provided regardless of 
808
their insurance status. All adverse events classified as serious or severe or 
809
possibly/probably related to the trial product will be followed until the subject has 
810
recovered and all queries have been resolved. For cases of chronic conditions follow-up 
811
until the outcome category is “recovered” is not required, as these cases can be closed with 
812
an outcome of “recovering” or “not recovered”. All other adverse events will be followed 
813
until the outcome of the event is “recovering” (for chronic conditions), or “recovered” or 
814
until the end of the post-treatment follow-up stated in the protocol, whichever comes first, 
815
and until all queries related to these AEs have been resolved.
816
Pregnancy: Pregnancy test must be negative prior to be enrolled in the study, as per 
817
standard care. However, study subjects will be instructed to notify the investigator 
818
immediately if they become pregnant. The investigator will report to IRB any pregnancy 
819
occurring during the trial period. Reporting of pregnancy by investigator will occur within 
820
the same timelines described above for reporting of Adverse Events.
821
Precautions/Over-dosage: Precautions and procedures will be observed in the event of 
822
overdose of liraglutide provided during the study. 
823
824
Liability and Subject Insurance
825
The investigator must state that during and following a subject’s participation in trial, the 
826
investigator and his/her institution will provide adequate medical care to the study subject 
827
for any study-related adverse events, including clinically significant laboratory values 
828
related to the study. This medical care for study subjects will be provided regardless of 
829
their insurance status.
830
The investigator will be responsible for the conduct of the study and agrees to defend, 
831
indemnify, and hold harmless Novo Nordisk, any of its parent companies, affiliates, or 
832
subsidiaries, and their respective officers, directors, employees, agents, representatives, 
833
distributors, salespersons, customers, licensees, and end-users from and against any claim, 
834
suit, demand, loss, damage, expense or liability imposed by any third party arising from or 
835
related to: (a) any breach of investigator's obligations; or (b) investigator’s negligent or 
836
grossly negligent use or willful misuse of the study drug, the results, or services derived 
837
there from.  This indemnification shall not apply in the event and to the extent that a court 
838
of competent jurisdiction or a duly appointed arbiter determines that such losses or liability 
839
arose as a result of Novo Nordisk’s gross negligence, intentional misconduct, or material 
840
breach of its responsibilities. 
841
Publication Plan
842
Data generated from the study will be published in highly ranked peer reviewed scientific 
843
journals, such as Diabetes, Diabetes Care, Obesity or Circulation and presented at national 
844
and international meetings, such as ADA 2018-19, ENDO Society 2018-19 and American 
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845
Heart Association  2018-19. Novo Nordisk will have any manuscripts for publication for 
846
review with a right to comment.
847
The investigator will register the study with a publicly assessable database such as 
848
clinicaltrials.gov.
849
850
REGULATORY CONSIDERATIONS
851
We plan to conduct the trial under an IND exemption. After reviewing 21 CFR 
852
312.2(b)(l) and FDA's '"Guidance for Clinical Investigators, Sponsors, and IRBs 
853
Investigational New Drug Applications (INDs) Determining whether human research 
854
studies can be conducted without an IND, September 2013",  we  believe an IND 
855
exemption is applicable to this study as it meets all the criteria, as reported in the table:
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857
21CFR 3 12.2 (B) EXEMPTIONS
858
(1) the clinical investigation of a drug product that is lawfully marketed in the 
859
united states is exempt from the requirements of this part if all the following 
860
apply:
861
(i) The investigation is not intended to 
be reported to FDA as a well-controlled 
study in support of a new indication for 
use nor intended to be used to support 
any other significant change in the 
labeling for the drug;
Meets criteria.
This clinical trial is not intended to be 
reported to the FDA in support of a 
new indication or change in Victoza 
labeling.
(ii) If the drug that is undergoing 
investigation is lawfully marketed as a 
prescription drug product, the 
investigation is not intended to support a 
significant change in the advertising for 
the_product.
Meets criteria.
This investigation is not intended to 
support a significant change in the 
Victoza advertising.
(iii) The investigation does not involve a 
route of administration or dosage level or 
use in a patient population or other factor 
that significantly increases the risks (or 
decreases the acceptability of the risks) 
associated with the product 
Meets criteria.
In this clinical trial liraglutide (and the 
matching placebo) will be administered 
according to the labeled route and dosage 
(Pre-filled, multi-dose pen for subcutaneous 
injection that delivers doses of 0.6 mg, 1.2 
mg, or 1.8 mg (6 mg/mL, 3 mL). In this 
clinical trial liraglutide (and the matching 
placebo) will be used according to the 
indications.  Liraglutide will be used in adults 
with type 2 diabetes (T2DM) in adjunct to diet 
and exercise advices and not as first-line 
therapy, as study patients will continue their 
standard 
diabetes 
therapy. 
Hence, 
liraglutide/placebo 
is 
prescribed 
under 
inclusion criteria that are the same as for 
marketed Victoza. Exclusion criteria for 
liraglutide are also the same as marketed 
Victoza.  This investigation will not increase 
the risks in the study population. 
(iv) The investigation is conducted in 
compliance with the requirements for 
institutional review set forth in part 56 and 
with the requirements for informed consent 
set forth in part 50;
Meets criteria.
This study will be reviewed and approved 
by the University of Miami IRB 
Institutional Review board prior to study 
start and will meet with the requirements for 
informed consent. 
(v) The investigation is conducted in 
compliance with the requirements of 21 
CFR 312.7.
Meets criteria.
Study Investigator, or any person acting on 
behalf of the investigator, will not represent 
in a promotional context that the study drug 
is safe or effective for the purposes for 
which it is under investigation or otherwise 
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promote the study drug.
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863
MAIN REFERENCES
864
865
1. Iacobellis G. Local and Systemic effects of the multifaceted Epicardial Adipose 
866
Tissue Depot. Nature Reviews Endocrinology 2015;11:363-371 PMID: 25850659
867
2. Iacobellis G, Bianco AC. Epicardial adipose tissue: emerging physiological, 
868
pathophysiological and clinical features. Trends Endocrinol Metab 2011; 22:450-
869
7. PMID: 21852149
870
3. Iacobellis G, Corradi D, Sharma AM. Epicardial adipose tissue: anatomic, 
871
biomolecular and clinical relationships with the heart. Nat Clin Pract Cardiovasc 
872
Med 2005; 2: 536-543 PMID: 16186852
873
4. McAninich E, Bianco AC and Iacobellis G. Epicardial Adipose Tissue Has a 
874
Unique Transcriptome that is Downregulated in Severe Coronary Artery Disease. 
875
Obesity, 2015; 23:1267-78. PMID: 25959145
876
5. Mahabadi AA, et al. Association of Epicardial Adipose Tissue With Progression of 
877
Coronary Artery Calcification Is More Pronounced in the Early Phase of 
878
Atherosclerosis: Results From the Heinz Nixdorf Recall Study. JACC Cardiovasc 
879
Imaging. 2014; 7, 909-916. 
880
6. Iacobellis G, Lonn E, Lamy A, Singh N, Sharma AM. Epicardial fat thickness and 
881
coronary artery disease correlate independently of obesity. Int J Cardiol. 
882
2011;146:452-4
883
7. Iacobellis G, Ribaudo MC, Assael F, Vecci E, Tiberti C, Zappaterreno A, Di Mario 
884
U, Leonetti F. Echocardiographic epicardial adipose tissue is related to 
885
anthropometric and clinical parameters of metabolic syndrome: a new indicator of 
886
cardiovascular risk. J Clin Endocrinol Metab 2003; 88:5163-5168 PMID: 
887
14602744
888
8. Iacobellis G, Willens HJ. Echocardiographic Epicardial Fat: A Review of Research 
889
and Clinical Applications. J Am Soc Echocardiogr 2009; 22:1311-1319. PMID: 
890
19944955
891
9. Malavazos AE, Di Leo G, Secchi F, Lupo EN, Dogliotti G, Coman C, Morricone 
892
L, Corsi MM, Sardanelli F, Iacobellis G. Relation of echocardiographic epicardial 
893
fat thickness and myocardial fat. Am J Cardiol. 2010;105:1831-5. PMID: 20538139
894
10. Iacobellis G. Epicardial fat: a new cardiovascular therapeutic target. Curr Opin 
895
Pharmacol. 2016;27:13-18 PMID: 26848943
896
11. Iacobellis G, Mohseni M, Bianco S, Banga PK. Liraglutide causes large and rapid 
897
Epicardial Fat reduction Obesity 2017; 25:311-316
898
12. Morano S, Romagnoli E, Filardi T, Nieddu L, Mandosi E, Fallarino M, Turinese I, 
899
Dagostino MP, Lenzi A, Carnevale V. Short-term effects of glucagon-like peptide 
900
1 (GLP-1) receptor agonists on fat distribution in patients with type 2 diabetes 
901
mellitus: an ultrasonography study. Acta Diabetol 2015; 52:727-32 
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902
13. Lima-Martínez MM, Paoli M, Rodney M, Balladares N, Contreras M, D'Marco L, 
903
Iacobellis G Effect of sitagliptin on epicardial fat thickness in subjects with type 2 
904
diabetes and obesity: a pilot study. Endocrine. 2016;51:448-55  PMID: 26233684
905
14. Elisha B, Azar M, Taleb N, Bernard S, Iacobellis G, Rabasa-Lhoret R. Body 
906
Composition and Epicardial Fat in Type 2 Diabetes Patients Following Insulin 
907
Detemir Versus Insulin Glargine Initiation. Horm Metab Res. 2015 Sep 4 
908
15. Sacks HS, Fain JN, Cheema P, Bahouth SW, Garrett E, Wolf RY. Inflammatory 
909
Genes in Epicardial Fat Contiguous with Coronary Atherosclerosis in the Metabolic 
910
Syndrome and Type 2 Diabetes: Changes associated with pioglitazone. Diabetes 
911
Care 2011, 34: 730-3 
912
16. Marso SP, Daniels GH, Brown-Frandsen K, Kristensen P, Mann JF, Nauck MA, 
913
Nissen SE, Pocock S, Poulter NR, Ravn LS, Steinberg WM, Stockner M, Zinman 
914
B, Bergenstal RM, Buse JB; LEADER Steering Committee; LEADER Trial 
915
Investigators Liraglutide and Cardiovascular Outcomes in Type 2 Diabetes. N Engl 
916
J Med. 2016;375:311-22
917
17. Camarena V, Wang G, Mohseni M, Salerno and Iacobellis G. Novel Atherogenic 
918
Pathways from the Differential Transcriptome Analysis of Diabetic Epicardial 
919
Adipose Tissue. Nutrition Metabolism and Cardiovascular Disease 2017 in press 
920
DOI: http://dx.doi.org/10.1016/j.numecd.2017.05.010
921
18. Iacobellis G, Camarena  V, Sant D, Wang G. Human Epicardial Fat Expresses Both 
922
Glucagon-Like Peptide 1 And 2 Receptors Genes Hormone and Metabolic 
923
Research accepted in press 2017
924
19. Faber R, Zander M, Pena A, Michelsen MM, Mygind ND, Prescott E. Effect of the 
925
glucagon-like peptide-1 analogue liraglutide on coronary microvascular function in 
926
patients with type 2 diabetes - a randomized, single-blinded, cross-over pilot study 
927
Cardiovasc Diabetol. 2015;14:41 PMID: 25896352
928
20. Noyan GLP-1R agonist liraglutide activates cytoprotective pathways and improves 
929
outcomes after experimental myocardial infarction in mice. Diabetes. 2009;58:975-
930
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931
21. Yang J, Ren J, Song J, Liu F, Wu C, Wang X, Gong L, Li W, Xiao F, Yan F, Hou 
932
X, Chen L. Glucagon-like peptide 1 regulates adipogenesis in 3T3-L1 
933
preadipocytes. Int J Mol Med 2013; 31:1429-35
934
22. Beiroa D, Imbernon M, Gallego R, Senra A, Herranz D, Villarroya F, Serrano M, 
935
Fernø J, Salvador J, Escalada J, Dieguez C, Lopez M, Frühbeck G, Nogueiras. 
936
GLP-1 agonism stimulates brown adipose tissue thermogenesis and browning 
937
through hypothalamic AMPK. Diabetes. 2014; 63:3346-58
938
23. Pyke C, Knudsen LB. The Glucagon-Like Peptide-1 Receptor—or Not?  
939
Endocrinology 2013;154:4-8.
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24. Pyke C, Heller RS, Kirk RK, Ørskov C, Reedtz-Runge S, Kaastrup P, Hvelplund 
941
A, Bardram L, Calatayud D, Knudsen LB. GLP-1 Receptor Localization in Monkey 
942
and Human Tissue: Novel Distribution Revealed With Extensively Validated 
943
Monoclonal Antibody. Endocrinology 2014; 155:1280-1290
944
25. Shiraki A, Oyama J, Komoda H, Asaka M, Komatsu A, Sakuma M, Kodama K, 
945
Sakamoto Y, Kotooka N, Hirase T, Node K. The glucagon-like peptide 1 analog 
946
liraglutide reduces TNF-?-induced oxidative stress and inflammation in endothelial 
947
cells. Atherosclerosis. 2012 Apr;221(2):375-82. 
948
26. Krasner NM, Ido Y, Ruderman NB, Cacicedo JM. Glucagon-like peptide-1 (GLP-
949
1) analog liraglutide inhibits endothelial cell inflammation through a calcium and 
950
AMPK dependent mechanism. PLoS One. 2014 May 16;9(5):e97554. 
951
952
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