A Pilot Randomized Study to Assess the Effect and Safety Profile of Thymoglobulin® in Primary Cardiac Transplant Recipients: A 12-month, Single Center, Randomized, Open-label Study of Efficacy Comparing Immediate Treatment With and Without Thymoglobulin® 1.5 mg/kg/d for 5 Consecutive Days in Heart Transplant Recipients
Phase: PHASE2
Conditions: Heart Transplantation
Sponsor: Cedars-Sinai Medical Center

 
1 
 
CLINICAL STUDY PROTOCOL 
 
 
GZ-2016-11621 
 
 
 
A Pilot Randomized Study to Assess the Effect and Safety Profile  
of Thymoglobulin® in Primary Cardiac Transplant Recipients: A 12-month, 
multi-center, randomized, open-label study of efficacy comparing 
immediate treatment with and without Thymoglobulin
® 1.5 mg/kg/d for 5 
consecutive days in heart transplant recipients. 
 
 
 
 
 
 
 
Amendment 9.0 
 
Version Date:    17 August 2023 
 
IND#18177 
 
 
Principal Investigator: Jon Kobashigawa, MD 
Cedars-Sinai Medical Center  
 
 
Kaiser Principal Investigator: Jonathan Neyer, MD 
Kaiser Permanente Los Angeles Medical Center 
 
 
 
 
 
 
 
 
 
 

2 
 
 
 
 
 
 
Confidentiality Statement 
 
The information contained in this document is not to be disclosed in any way without the prior permission 
of the Principal Investigator. 

3 
 
 Protocol Summary 
 
Title 
A Pilot Randomized Study to Assess the Effect and Safety Profile of 
Thymoglobulin® in Primary Cardiac Transplant Recipients: A 12-month, multi 
center, randomized open-label study of efficacy comparing immediate 
treatment with and without Thymoglobulin® 1.5mg/kg/d for 5 consecutive 
days in heart transplant recipients 
Primary Objectives 
To describe between treatment groups the incidence of:  
• the composite primary endpoint of the development of de novo donor 
specific antibodies and ischemia on endomyocardial biopsy at 12 months 
post-transplantation. 
Secondary Objectives To describe between treatment groups: 
• Number of patients who develop cardiac allograft vasculopathy (CAV) 
(defined as a change ≥0.5mm in maximal intimal thickness (MIT) of the 
coronary arteries by intravascular ultrasound at 12 months as compared 
to baseline) 
• Correlations between significant changes in immune cell profiles and 
circulating antibodies., and their relation to any significant differences in 
clinical outcomes 
• Number of patients who experience acute cellular, antibody-mediated, 
hemodynamic compromise, and any-treated rejection within first 12 
months after transplantation 
• Number of patients with biopsy proven cellular rejection ≥2R, biopsy 
proven antibody mediated rejection ≥AMR1 and any-treated rejection at 
12 months post-transplantation 
• Number of acute cellular, antibody-mediated, hemodynamic compromise 
and any-treated rejection episodes per patient within the first 12 months 
post-transplantation 
• Freedom from acute cellular, antibody-mediated, hemodynamic 
compromise, and any-treated rejection by the ISHLT biopsy grading scale 
in the first 12 months post-transplantation 
• Time to first acute cellular, antibody-mediated, hemodynamic 
compromise, and any-treated rejection within the first 12 months 
• the incidence of Primary Graft Dysfunction (PGD) in the first 24 hours 
post-transplant 
• Patient and graft survival at 12 months post-transplantation 
• The types and number of patients with both fatal and non-fatal infectious 
complications (especially CMV infection) within the first 12 months post-
transplantation 
• Freedom from the development of circulating antibodies within the first 
12 months post transplantation, where circulating antibodies include 
donor specific antibodies (DSA), non-specific antibodies, and non-human 
leukocyte antigen antibodies 
• Change in coronary maximal intimal thickness, intimal area, intimal 
volume, vessel area, intimal index and percent atheroma volume (PAV) at 
matched sites by intravascular ultrasound at 12 months  
• Maintenance doses of mycophenolate mofetil, tacrolimus, sirolimus, and 
cumulative dose of corticosteroids at 12 months post-transplantation 
• Number of hospital days per patient, both during the transplant period 
and during the post-transplant period at 3 months, 6 months, and 1 year 

4 
 
• Number of patients requiring hospitalization by 3 months, by 6 months, 
or by 1 year post-transplantation 
Study Design 
Randomized and controlled, descriptive study 
Sample Size 
60 primary cardiac transplant recipients 
Number of Centers 
Two centers 
Selection Criteria 
• Primary cardiac transplantation 
• 18 to 74 years old 
• Men and non-pregnant women 
Study Medication 
Thymoglobulin® (rabbit anti-thymocyte globulin 25mg/5mL vials, IV) 
Comparator 
No induction therapy 
Test Dose/Duration 
• 1.5 mg/kg/24 hours X 5 doses I.V. 
• First dose administered within 24 hours after transplant if subject is 
randomized to receive Thymoglobulin. 
• Total duration 5 doses over 5 days 
Route of 
Administration 
Intravenous— 0.5mg/ml IV solution in D5W or NS. The First two doses infused 
over 8 hours.  All subsequent doses infused over 4-8 hours 
Primary Parameters 
of Efficacy 
• Incidence of the primary endpoint of the development of de novo donor 
specific antibodies and ischemia on endomyocardial biopsy at 12 months 
post-transplant 
Secondary 
Parameters of 
Efficacy 
To describe between groups: 
• Changes in percentages of various subsets of immune cells at pre 
transplant, 3 months, and 6 months.  Changes in percentages of 
various circulating antibodies at pre-transplant, 1 month, 3 months, 
6 months, and 12 months after transplant. 
• Freedom from cardiac allograft vasculopathy (CAV) (defined as a 
change ≥0.5mm in maximal intimal thickness (MIT) of the coronary 
arteries by intravascular ultrasound at 12 months as compared to 
baseline).Change in coronary maximal intimal thickness intimal area, 
intimal volume, vessel area, intimal index, and percent atheroma 
volume at matched sites by intravascular ultrasound at 12 months 
Freedom from acute cellular, antibody-mediated, hemodynamic 
compromise, and any-treated rejection within first 12 months after 
transplantation. 
• Freedom from acute cellular, antibody-mediated, hemodynamic 
compromise, and any-treated rejection episodes per patient within 
the first 12 months post-transplantation 
• Freedom from acute cellular, antibody-mediated, hemodynamic 
compromise, and any-treated rejection by the ISHLT biopsy grading 
scale in the first 12 months post-transplantation 
• Freedom from biopsy proven cellular rejection ≥2R, biopsy proven 
antibody mediated rejection ≥AMR1 and any-treated rejection at 12 
months post-transplantation 
• Time to first acute cellular, antibody-mediated, hemodynamic 
compromise, and any-treated rejection within the first 12 months 
post-transplantation 
• Freedom from primary graft dysfunction (PGD) in the first 24 hours 
post-transplantation 
• Patient and graft survival at 12 months post-transplantation 

5 
 
• Freedom from fatal and non-fatal infectious complications 
(especially CMV infection) within the first 12 months post-
transplantation   
• Freedom from the development of circulating antibodies within the 
first 12 months post-transplantation 
• Average maintenance doses of Mycophenolate Mofetil, tacrolimus, 
and cumulative dose of corticosteroids at 12 months post-
transplantation 
• Average number of hospital days per patient, both during the 
transplant period and during the post-transplant period at 3 months, 
6 months, and 1 year, 
• Freedom from hospitalizations by 3 months, 6 months, or 12 months 
post-transplantation. 
• Freedom from the development of circulating antibodies 
Main Parameters of 
Safety 
• Number of patients with clinical CMV infection 
• Incidence of adverse events, including infectious complications, and 
malignancies 
• Changes in laboratory values, especially absolute total white blood cell, 
platelet, and hemoglobin counts 
Randomization 
Patients qualifying for the study will be randomized in a 1:1 ratio to either 
Thymoglobulin® or No Induction or Control during the immediate post 
transplantation period after confirming the eligibility criteria for 
randomization.   
Duration 
The study was initiated in September 2018 and enrollment will continue until 
October 31st, 2024 The study will be completed 24 months after the last 
patient has entered. Study completion is expected by October 31, 2025. 
 
 
 

6 
 
GLOSSARY OF ABBREVIATIONS 
 
ABO 
The three blood types—A type, B type, and O type 
ACE 
Angiotensin Converting Enzyme 
AE 
Adverse Event 
AMR 
Antibody-Mediated Rejection 
ANOVA 
Analysis Of Variance 
ATG 
Anti-Thymocyte Globulin (of rabbit unless otherwise indicated) 
beta-HCG 
beta-Human Chorionic Gonadotropin 
bid 
taken twice a day 
BUN 
Blood Urea Nitrogen 
CAV 
Cardiac Allograft Vasculopathy 
CBC 
Complete Blood Count 
CI 
Cardiac Index 
CMV 
Cytomegalovirus 
CO2 
Carbon Dioxide 
CRF 
Case Report Form 
EBV 
Epstein Barr Virus 
ECG 
Electrocardiogram 
ELISA 
Enzyme Linked Immunosorbent Assay 
ESF 
Eligibility Screening Form 
FDA 
Federal Drug Administration 
GGT 
Gamma Glutamate Transferase 
HDL 
High-Density Lipoprotein 
HIPAA 
Health Insurance Portability and Accountability Act 
HIV-1 
Human Immunodeficiency Virus type 1 
HLA 
Human Leucocyte Antigen 
HMG CoA 
Reductase 
Inhibitor 
3-Hydroxy-3-Methyl-Glutaryl Coenzyme A Reductase Inhibitor 
(statins)  
ICU 
Intensive Care Unit 
IFN 
Interferon 
IgG 
Immunoglobulin G 
IL 
InterLeukin (a cytokine) 
IRI 
Ischemia-Reperfusion Injury 
ISHLT 
International Society for Heart and Lung Transplantation 
IU 
International Units 
IV 
IntraVenous 
IVUS 
Intravascular Ultrasound 
LDH 
Lactate DeHydrogenase 
LDL 
Low-Density Lipoprotein 
LVAD 
Left Ventricular Assist Device 
MHC 
Major HistoCompatibility 
MMF 
Mycophenolate MoFetil 
NIAID 
National Institutes of Allergy and Infectious Disease 
IC 
Infectious Complications 
PO/po 
Per Orum, taken orally or by mouth 
PBMC 
Peripheral Blood Mononuclear Cells 
PGD 
Primary Graft Dysfunction 
PRA 
Panel of Reactive Antibodies 

7 
 
PRT 
Panel of Reactive T-cells 
PRBC 
Packed Red Blood Cells 
PTLD 
Post-Transplant Lymphoproliferative Disorder 
qd 
Taken/administered once a day 
RBC 
Red Blood Cell Count 
RHC 
Right Heart Catheterization 
SAE 
Severe Adverse Events 
SCID 
Severe Combined ImmunoDeficiency 
SGOT 
Serum Glutamate Oxalacetate Transaminase 
SGPT 
Serum Glutamate Pyruvate Transaminase 
sirolimus 
= rapamycin 
SOC 
Standard Of Care 
tid 
taken three times a day 
TNF 
Tumor Necrosis Factor (a cytokine) 
VAD 
Ventricular Assist Device 
WBC 
White Blood Cell Count 
 

8 
 
TABLE OF CONTENTS 
1.
1. BACKGROUND
BACKGROUND 
................................
................................................................
................................................................
...........................................
........... 
13
13 
1.1 DISEASE BACKGROUND .............................................................................................................. 
13 
1.1.6 Donor Specific Antibodies 
......................................................................................................... 13 
1.1.2 Cardiac Allograft Vasculopathy ................................................................................................. 13 
1.1.3 Infection in Transplant Recipients 
............................................................................................. 16 
1.1.4 Post-transplant Lymphoproliferative Disease in Transplant Recipients 
.................................... 17 
1.1.5 Graft Tolerance 
.......................................................................................................................... 17 
1.1.6 Graft Rejection .......................................................................................................................... 18 
1.1.7 Ischemia-reperfusion injury, primary graft dysfunction, and significance to CAV and ATG ..... 20 
1.2 DRUG BACKGROUND 
.................................................................................................................. 
23 
1.2.1 Description and Mechanism of Action 
...................................................................................... 23 
1.2.2 Pharmacokinetics/pharmacodynamics 
..................................................................................... 24 
1.2.3 Clinical Trials 
.............................................................................................................................. 25 
1.2.3.1 
Human Clinical Trials of ATG in Renal Transplantation 
.................................................... 25 
1.2.3.2 
Human Clinical Studies of ATG in Cardiac Transplantation 
.............................................. 29 
1.3 RATIONALE FOR PERFORMING THE STUDY 
.................................................................................. 
35 
1.3.1 The Efficacy of ATG Induction in Cardiac Transplantation is Unknown 
..................................... 35 
1.3.2 Thymoglobulin has numerous mechanisms of immunosuppression 
........................................ 40 
1.4 SIGNIFICANCE OF STUDY............................................................................................................. 
41 
2.
2. STUDY OBJECTIVES
STUDY OBJECTIVES 
................................
................................................................
................................................................
....................................
.... 
42
42 
2.1 PRIMARY OBJECTIVES 
................................................................................................................. 
42 
2.2 SECONDARY OBJECTIVES 
............................................................................................................ 
42 
3.
3. STUDY DURATION AND STUDY EARLY TERMINATION
STUDY DURATION AND STUDY EARLY TERMINATION 
................................
...............................................
............... 
42
42 
3.1 PARTICIPANT COMPLETION 
........................................................................................................ 
43 
3.2 PREMATURE TERMINATION OF A PARTICIPANT FROM THE STUDY 
.............................................. 43 
3.3 PREMATURE DISCONTINUATION OF STUDY DRUG 
...................................................................... 
43 
3.4 STUDY STOPPING RULES 
............................................................................................................. 
43 
3.4.1 Post-Transplant Lymphoproliferative Disorder (PTLD) 
.............................................................. 44 
3.4.2 Infectious Complications 
........................................................................................................... 44 
3.4.3 Death ......................................................................................................................................... 44 
3.4.4 Rejection 
.................................................................................................................................... 44 
4.
4. CENTER OF STUDY
CENTER OF STUDY 
................................
................................................................
................................................................
.....................................
..... 
44
44 
5.
5. SELECTION CRITERIA
SELECTION CRITERIA ................................
................................................................
................................................................
................................. 
44
44 

9 
 
5.1 TOTAL NUMBER OF PATIENTS 
..................................................................................................... 
44 
5.2 RECIPIENT INCLUSION CRITERIA FOR STUDY ENTRY 
..................................................................... 
44 
5.3 RECIPIENT EXCLUSION CRITERIA FOR STUDY ENTRY 
.................................................................... 
45 
5.4  RECIPIENT INCLUSION CRITERIA FOR RANDOMIZATION ............................................................. 
45 
5.5  
RECIPIENT EXCLUSION CRITERIA FOR RANDOMIZATION ......................................................... 
45 
5.6 DONOR INCLUSION and EXCLUSION CRITERIA 
................................................................................ 
46 
6 DISEASE EVALUATION (EFFICACY AND SAFETY CRITERIA)
DISEASE EVALUATION (EFFICACY AND SAFETY CRITERIA) 
................................
.........................................
......... 
46
46 
6.1 PRIMARY EFFICACY PARAMETERS 
............................................................................................... 
46 
6.2 SECONDARY EFFICACY PARAMETERS 
.......................................................................................... 
46 
6.3 DIAGNOSIS AND TREATMENT OF CAV 
......................................................................................... 
47 
6.4 DIAGNOSIS AND TREATMENT OF REJECTION 
............................................................................... 
49 
6.4.1 Diagnosis of Rejection 
............................................................................................................... 49 
6.4.2 Definition of Hemodynamic Compromise 
................................................................................. 49 
6.4.3 Treatment of Cellular Rejection ................................................................................................ 50 
6.4.4 Treatment of Antibody Mediated Rejection ............................................................................. 50 
6.5 DIAGNOSIS OF ISCHEMIA-REPERFUSION INJURY 
......................................................................... 
51 
6.6 PHARMACOKINETIC/PHARMACODYNAMIC PARAMETERS .......................................................... 
51 
7.
7. STUDY DESIGN
STUDY DESIGN 
................................
................................................................
................................................................
...........................................
........... 
51
51 
7.1 DESIGN 
....................................................................................................................................... 
51 
7.2 STUDY PROCEDURES .................................................................................................................. 
53 
7.2.1 Screening/Baseline 
.................................................................................................................... 53 
7.2.3 Clinical Evaluations 
.................................................................................................................... 53 
7.3 RANDOMIZATION PROCEDURE AND ASSIGNMENT TO TREATMENT GROUP 
................................ 
54 
8.
8. STUDY MEDICATION
STUDY MEDICATION 
................................
................................................................
................................................................
..................................
.. 
54
54 
8.1 DRUG NAME, FORMULATION, & STORAGE ................................................................................. 
54 
8.2 PACKAGING AND LABELING 
........................................................................................................ 
54 
8.3 STUDY TREATMENT 
.................................................................................................................... 
55 
8.3.1 Rationale for Dose Selection 
..................................................................................................... 55 
8.3.2 Dosage Regimen and Dose Adjustment 
.................................................................................... 57 
8.3.3 Dose Modification 
..................................................................................................................... 58 
8.3.4 Route of Administration 
............................................................................................................ 59 

10 
 
8.3.5 Treatment Time Course 
............................................................................................................. 59 
8.4 DISPENSING AND ACCOUNTABILITY OF STUDY MEDICATION SUPPLIES ....................................... 
59 
8.5 CONCOMITANT TREATMENT ...................................................................................................... 
60 
8.5.1 Concomitant Immunosuppressive Therapy .............................................................................. 60 
8.5.2 Other Concomitant Therapy 
...................................................................................................... 60 
8.5.3 Mycophenolate Mofetil Dose Adjustment for Adverse Events 
................................................. 61 
8.5.4 Prophylactic Therapy 
................................................................................................................. 61 
8.5.5 Drug Interactions 
....................................................................................................................... 61 
9.
9. PREMATURE WITHDRAWAL
PREMATURE WITHDRAWAL 
................................
................................................................
......................................................
...................... 
62
62 
9.1 FOLLOW-UP 
................................................................................................................................ 
62 
10.
10. WARNINGS AND PRECAUTIONS
WARNINGS AND PRECAUTIONS 
................................
................................................................
................................................
................ 
62
62 
10.1 OVERDOSE OR EXAGGERATED RESPONSE 
............................................................................... 
64 
11.
11. SAFETY PARAMETERS
SAFETY PARAMETERS 
................................
................................................................
................................................................
................................ 
65
65 
11.1 MEDICAL HISTORY AND CLINICAL EVALUATIONS 
.................................................................... 
65 
11.2 LABORATORY PARAMETERS 
................................................................................................... 
65 
11.2.1 
Procedures in the Event of Significant Abnormal Laboratory Values 
................................... 66 
11.3 INFECTIOUS COMPLICATIONS 
................................................................................................. 
66 
12
12 ADVERSE EVENTS
ADVERSE EVENTS ................................
................................................................
................................................................
......................................
...... 
66
66 
12.1 Baseline Medical Condition 
.................................................................................................... 
66 
12.2 DEFINITION OF ADVERSE EVENTS 
........................................................................................... 
67 
12.2.1 
Adverse Event (AE) ............................................................................................................... 67 
12.2.2 
Suspected Adverse Reaction (SAR) 
....................................................................................... 67 
12.2.3 
Unexpected Adverse Event 
................................................................................................... 67 
12.2.4 
Serious Adverse Events 
......................................................................................................... 67 
12.3 GRADING AND ATTRIBUTION OF ADVERSE EVENTS ................................................................ 
67 
12.3.1 
Grading Criteria .................................................................................................................... 67 
12.3.2 
Attribution Definitions 
.......................................................................................................... 68 
12.4 COLLECTION AND RECORDING OF ADVERSE EVENTS 
............................................................... 
69 
12.4.1 
Collection Period................................................................................................................... 69 
12.4.2 
Collecting Adverse Events 
..................................................................................................... 69 
12.4.3 
Recording Adverse Events 
.................................................................................................... 69 
12.4.4 
Reporting of Serious Adverse Events 
.................................................................................... 69 
12.5 FOLLOW-UP OF ADVERSE EVENTS .......................................................................................... 
69 
12.5.1 
Planned Data and Safety Monitoring Reviews ..................................................................... 70 

11 
 
12.5.2 
Ad Hoc Data and Safety Reviews 
.......................................................................................... 70 
13.
13. MECHANISTIC ASSAYS
MECHANISTIC ASSAYS 
................................
................................................................
...............................................................
............................... 
70
70 
13.1 Blood for circulating antibodies 
.............................................................................................. 
70 
13.2 PRT ASSAY 
............................................................................................................................. 
70 
13.3 LYMPHOCYTE ANALYSIS 
......................................................................................................... 
71 
13.4 CYTOKINES 
............................................................................................................................. 
71 
14.
14. STATISTICAL CONSIDERATIONS
STATISTICAL CONSIDERATIONS 
................................
................................................................
.................................................
................. 
72
72 
14.1 SAMPLE SIZE CALCULATION 
.................................................................................................... 
72 
14.2 ANALYSIS PLAN 
...................................................................................................................... 
72 
14.2.1 
Primary Efficacy Parameters 
................................................................................................. 72 
14.2.2 
Secondary Efficacy Parameters ............................................................................................ 72 
14.2.3 
Definition of Parameters ...................................................................................................... 73 
14.2.4 
Statistical and Analytical Methods ....................................................................................... 74 
14.2.5 
Intent to Treat Population Analysis 
...................................................................................... 74 
14.3 SAFETY DATA ANALYSIS ......................................................................................................... 
74 
14.4 FINAL STUDY ANALYSIS 
.......................................................................................................... 
75 
15.  
15.   SOURCE DATA
SOURCE DATA................................
................................................................
................................................................
............................................
............ 
75
75 
15.1  SOURCE DATA 
........................................................................................................................ 
75 
15.2 ACCESS TO SOURCE DATA ...................................................................................................... 
75 
16.
16. QUALITY CONTROL AND QUALITY ASSURANCE
QUALITY CONTROL AND QUALITY ASSURANCE 
................................
.........................................................
......................... 
75
75 
16.1 DATA HANDLING 
.................................................................................................................... 
75 
17.
17. ETHICAL CONSIDERATIONS AND COMPLIANCE WITH GOOD CLINICAL PRACTICE
ETHICAL CONSIDERATIONS AND COMPLIANCE WITH GOOD CLINICAL PRACTICE 
....
.... 
76
76 
17.1 STATEMENT OF COMPLIANCE................................................................................................. 
76 
17.2 INFORMED CONSENT 
............................................................................................................. 
76 
17.3 PRIVACY AND CONFIDENTIALITY 
............................................................................................ 
76 
18.
18. REFERENCES
REFERENCES 
................................
................................................................
................................................................
..............................................
.............. 
77
77 
APPENDIX A
APPENDIX A 
................................
................................................................
................................................................
.......................................................
....................... 
88
88 

12 
 
APPENDIX B
APPENDIX B 
................................
................................................................
................................................................
.......................................................
....................... 
90
90 
SCHEDULE OF ASSESSMENTS
SCHEDULE OF ASSESSMENTS 
................................
................................................................
............................................................
............................ 
92
92 

13 
 
BACKGROUND 
 
1.1 
DISEASE BACKGROUND 
 
Cardiac transplantation is currently the procedure of choice for selected patients with end-stage 
heart disease that is not amenable to further medical intervention or conventional cardiac 
procedures.  Over the past 10 years, experience, research, and new drugs have together resulted 
in an increase in the median survival of cardiac transplant recipients to 11 years.
1  However, 
survival for transplant recipients is still limited by cardiac allograft vasculopathy (CAV), infectious 
complications, malignancy, and rejection.  Reports of spontaneous allograft tolerance emphasize 
the importance of understanding how the immune system interacts with the allograft and how 
the immune system might be manipulated to achieve allograft tolerance. Described below is the 
effect of each of these factors on recipient survival and what is known about the effect of 
antithymocyte globulin (ATG/Thymoglobulin
®) on each of these factors. 
 
1.1.6 Donor Specific Antibodies 
 
The development of donor-specific anti-HLA antibodies is an established risk factor for poor 
outcomes after heart transplantation. De novo DSA after heart transplant is associated with an 
increased risk of acute cellular rejection, antibody mediated rejection and CAV.
2 In a 
retrospective, single-center study of 243 patients who underwent heart transplantation 
between 1995 and 2004, 57 patients developed de novo DSA. Univariate analysis found that de 
novo and persistent DSA (found in two consecutive samples) were associated with increased 
mortality (HR 3.198, p = 0.0018; HR 4.351, p = 0.002). Similarly, on multivariate analysis, de 
novo, persistent DSA was strongly predictive of mortality (HR 4.331, p <0.0004). DSA were 
associated with biopsy proven rejection in the first year after heart transplant.
3 A large 
retrospective study of 950 heart transplant patients examining the effects of de novo DSA found 
that survival was 52% for patients developing de novo DSA in the first versus 70% in patients 
who developed no DSA at 15 years post-transplantation.
4 Another retrospective study 
examining a series of 71 heart transplant recipients, 7 of whom developed de novo DSA 
demonstrated an association with the development of CAV at 3 years post-transplant. Of the 
patients who developed CAV, 55% had Class II anti-HLA antibodies. The presence of these 
antibodies also correlated with death due to allograft failure.
5 A recent study examining biopsy-
proven AMR and outcomes found that 71% of patients with AMR had DSA and the presence of 
DSA with AMR increased the odds of graft dysfunction (OR 5.37, 95% CI 1.34 to 21.47, p = 
0.018).6 Another recent study of 122 consecutive heart transplant patients assessed the impact 
of developing de novo DSA on a composite primary endpoint of death or graft dysfunction. 
During a median follow-up of 3.3 years, 28% of patients developed de novo DSA. Development 
of antibodies to HLA-DQ was associated with an increased risk of the primary endpoint (HR 6.15, 
95% CI 2.57-14.75, p=0.001).7 
 
1.1.2 Cardiac Allograft Vasculopathy 
 
Cardiac allograft vasculopathy (CAV) accounts for a significant number of deaths in cardiac 
recipients after the first-year post-transplant.  In a recent registry report from the ISHLT, CAV 
was noted in 7.8% of patients at 1 year post-transplant, 30% of patients at 5 years, and 50% at 10 
years.
1  CAV is the third most common cause of death after 3 years post-transplant.  CAV carries 

14 
 
a poor
 prognosis with
 a
 mortality
 rate
 of
 50%
 within
 2
 years
 of
 diagnosis.
8 
 
CAV
 may
 occur
 as
 early
 
as one year after transplantation.  Disease appearing early following transplantation is more 
aggressive and associated with a worse prognosis, with two thirds of patients suffering coronary 
events within 5 years following the detection of CAV.
9-12  In one study, those with angiographic 
disease had a relative risk of any cardiac event of 3.4 and a relative risk of death of 4.6 compared 
with those without disease.
9  Due to the denervated transplanted heart, cardiac ischemia or 
infarction does not typically present with chest pain.  Patients with myocardial infarction 
frequently lack typical ECG changes due to either baseline abnormalities or to heterogeneous 
disease resulting from diffuse vasculopathy.  In general, the absence of symptoms and ECG 
findings often lead to lower utilization of revascularization therapies and consequently, worse 
outcomes such as heart failure, arrhythmia or sudden death.
9 
 
With the exception of re-transplantation, available treatments for CAV only slow its 
progression.  Because of the diffuse and concentric nature of CAV and its involvement in small 
vessels, percutaneous intervention and coronary artery bypass are not effective treatments.  
Medications which have been demonstrated to be efficacious in clinical trials include 
atorvastatin,
13 pravastatin,
14,15 simvastatin,
16 calcium channel blockers,
17-19 ACE inhibitors,
19 
aspirin,
20 vitamin C and vitamin E,
21 ganciclovir (in CMV mismatch),
22 and possibly sirolimus
23 and 
everolimus.24 
 
Modifiable
 risk
 factors
 include 
smoking
 cessation,
 control
 of
 diabetes,
 and
 freedom
 
from rejection. 
 
Re-transplantation for CAV is reasonable as the 1-year actuarial survival approaches the 1-year 
survival following primary transplants.  Patients having a second heart transplant do not have an 
increased risk for development of CAV in the second donor heart.
25,26  However, the scarcity of 
donor hearts both creates an ethical dilemma and limits re-transplantation to only a small 
minority of patients. 
 
CAV occurs both in recipients who were transplanted for non-ischemic heart disease as well as 
those who were transplanted for ischemic heart disease.  In contrast to the more focal lesions 
of conventional atherosclerosis, CAV exhibits a more diffuse nature in the form of concentric 
narrowing with frequent involvement of large and medium sized vessels as well as the 
microvasculature.
27  The disease involves only the allograft and spares the native arteries.  
Together these observations suggest an immunologic basis for CAV.  This is also consistent with 
early experimental evidence where hearts transplanted into a genetically different recipient were 
affected by CAV and those transplanted into a genetically identical recipient were spared. 
 
However, there are also non-immunological clinical risk factors for CAV, which have been 
consistently identified in a number of retrospective studies.  These include preservation injury, 
the cause of donor death, donor graft ischemic time, older donor age, early rejection, the host 
allo-immune
 response,
 and
 cytomegalovirus infection
9.
 
 Although
 not
 causative,
 the
 conventional 
risk factors for atherosclerosis, such as hypertension, hyperlipidemia, glucose intolerance, and 
oxidative stress, also accelerate
 the
 disease
 process.
  
Although
 the
 non-immunological
 risk
 factors
 
may
 seem
 diverse
 and
 un-related,
 one
 result
 in
 common
 is
 causing
 an
 increase
 in
 the
 susceptibility
 
of the endothelium to damage.  Therefore, currently it is believed that CAV results from an initial 
injury to the donor endothelium from ischemic events that occur pre- or peri- operatively.
28,29  
This
 injury
 is
 followed
 by
 intimal
 hyperplasia 
and
 the
 proliferation
 of
 vascular
 smooth
 muscle
 cells,
 
which is believed to reflect sustained activation of the recipient’s immune system against and 
ongoing immune response against donor MHC antigens that is perpetuated by subsequent 

15 
 
episodes of allograft rejection.
30  This is evidenced by a study in which the T-cell alloresponses to 
synthetic peptides corresponding to the hypervariable region of the mismatched HLA-DR antigen 
of the donor were prospectively followed in 34 heart transplant recipients from 1-3 years 
posttransplant.  Patients who developed CAV and who had more frequent rejection episodes (i.e. 
unable to develop tolerance) exhibited (a) a progressive
 diminution
 in the number
 of
 T-cell clones
 
recognizing
 the
 immunodominant
 donor
 alloantigen
 determinants
 and
 a
 gradual
 shift
 in
 the
 T-cell 
repertoire, leading to the new recognition of multiple non-dominant epitopes, which in turn 
perpetuated the rejection process; (b) stronger T-cell alloreactivity during the first 6 months 
posttransplant; and (c) persistent T-cell alloreactivity after the first 6 months.
31 
 
Experimental and clinical findings demonstrate that both humoral and cellular immune 
responses contribute to the pathogenesis of graft vasculopathy (see Section 1.1.5 below on 
humoral and cellular rejection).
32  Evidence for humoral involvement in clinical heart 
transplantation comes from the observation that circulating anti-HLA antibodies and anti-
endothelial cell surface antibodies have been associated with CAV.  When compared with control 
patients, those patients who develop AMR and have a higher frequency of AMR progress to CAV 
earlier and at an increased frequency. The vast majority of patients who demonstrate histological 
AMR suffer from CAV by 5 years after transplantation regardless of whether they have 
concomitant hemodynamic
 compromise.
33 
 
Animal
 studies
 support
 these
 observations.
 
 Russell
 et 
al. reported that hearts transplanted into B cell-deficient mice that are incapable of producing 
immunoglobulins develop only minor CAV.
34  Another animal study showed that hearts 
transplanted into immunodeficient SCID mice survived indefinitely with no significant 
vasculopathy.  However, vascular lesions rich in macrophages and NK cells developed in the SCID 
mice upon treatment with an antiserum reactive with donor antigens.
32  In rodent models of 
allograft vasculopathy arterial lesions are observed to first develop as endotheliitis and 
subsequently progress to smooth muscle cell-rich fibrosis.   
 
Cellular rejection also plays a significant role in the development of CAV.  In animal studies 
where knockout mice deficient in both cellular and humoral immunity or only humoral immunity 
were transplanted, CAV developed in knockout mice that were deficient only in humoral 
immunity, suggesting that T-cellular rejection does play a role in the development of CAV.
35  
Russell, et al. also showed that the histological features of the CAV produced in SCID mice 
transfused with antiserum are distinct from those lesions in mice which have preserved cellular 
and humoral activities.  T- cells, including CD4+, CD8+, and Mac-1+ cells, are seen in the arterial 
lesions of the mice with preserved cellular and humoral immunity.
36  These cells secrete a myriad 
of inflammatory markers, including cell adhesion molecules, cytokines, chemokines, growth 
factors, and macrophage activators.
37  In another study depletion of CD4+ T-cells (but not CD8+ 
T-cells) prevents arterial lesion formation in these rodent models.
38  In addition, interferon-
gamma and transcription factor stat 4 knockout animals (inflammatory cytokine products 
produced by TH1 cells) demonstrated decreased vasculopathy. 
 
The role of antibodies and AMR in the development of CAV is equivocal. Evidence from animal 
models of CAV have demonstrated that induction therapy that depleted B cells is associated with 
less
 severe
 CAV
 development.
39 
Whether
 induction
 therapy
 can
 prevent
 the
 development
 of
 AMR
 
is human subjects remains to be proven. A recent study examined 40 explanted hearts and 402 
endomyocardial biopsy (EMB) samples from these hearts before graft loss.40 Their work 
demonstrated that  47.5%  of  explanted,  
failing  allografts  had  undiagnosed  antibody  mediated 
rejection (AMR) years earlier. Immune modulation targeting the humoral response could, 

16 
 
therefore, play a role in preventing or slowing the progression of CAV. These finding were in 
agreement with an earlier 5-year  study  examining  the  
freedom  
from  CAV  
in  heart  transplant 
recipients with asymptomatic and untreated AMR.
41 CAV more frequently developed in patients 
with asymptomatic and untreated AMR than in controls. Another  study  examining  71  hear 
transplant recipients demonstrated that antibodies against class II HLA were strongly associated 
with the development of CAV by 3-years post transplantation.
5 
 
Whether
 antibody
 induction
 therapy
 affects
 the
 development
 of
 CAV
 remains
 controversial.
 
 Thus
 
far, controlled studies of monoclonal antibodies have not demonstrated a beneficial outcome in 
either CAV or rejection.  However, the role of ATG, a polyclonal antibody preparation, remains 
controversial. In contrast to the monoclonal antibodies, ATG, being a polyclonal agent, is able to 
target all of the potential mechanisms that contribute to the development of CAV—T-cell 
activation, B-cell activation,
 antibody formation,
 induction of tolerogenic cells,
 and
 modulation
 of 
lymphocyte-endothelium interactions.   These mechanisms will be discussed in more detail in the 
sections to follow. Several small retrospective studies have shown that the incidence of CAV in 
cardiac transplant recipients who received ATG induction therapy is significantly lower.
42-44  
Therefore,
 ATG
 induction
 may
 prevent
 the
 development
 of
 CAV
 and
 its
 mechanism
 may
 be
 related
 
to its polyclonal nature.   
 
1.1.3 Infection in Transplant Recipients 
 
Immunosuppression using triple drug therapy is currently the mainstay of prevention against 
cardiac allograft rejection.  However, while too little immunosuppression increases the risk for 
rejection, too much immunosuppression increases the risk for infection.  Currently there are no 
diagnostic tests that can guide the amount of immunosuppression needed to achieve a 
reasonable balance between rejection and infection. The amount of immunosuppression needed 
to prevent both infection and rejection is highly individual. Therefore despite the fact that newer 
immunosuppressive regimens have resulted in a trend towards a lower rejection rate in the 
United States, about 50% of cardiac transplant recipients are treated for infection within the first 
year after transplant.
45 
 
Cardiac
 allograft
 recipients
 are
 prone
 to
 acquiring
 persistent
 bacterial,
 fungal,
 and
 viral
 infections.
 
Infections within the first month post-transplant are usually nosocomial or donor-derived.  
Between one and six months post-transplant, levels of immunosuppression usually higher and 
thus the most common infections are either opportunistic or due to the activation of latent 
infection within the recipient.  Infections acquired greater than six months post-transplant are 
usually community-acquired.  The incidence and severity of post-transplant infections has been 
significantly
 altered
 in
 the
 past
 decade
 by
 a
 better
 understanding
 of
 the
 risk
 factors
 for
 infection
 
from research, implementation of anti-microbial prophylaxes and vaccinations both 
preemptively and universally, and improved methods for the detection of infection.   
 
Studies have demonstrated that changes to immunosuppression regimens either by the 
introduction of a new drug or the addition/subtraction of a drug can result in new patterns of 
infection. 
 For example, the use 
of
 azathioprine
 is
 associated
 with
 a
 lower
 rate
 of
 bacterial,
 fungal,
 
and aspergillus infections than use of proliferation signal inhibitors (PSI) such as everolimus or 
sirolimus, but the incidence of viral infection is significantly reduced in patients receiving PSI.
24,46  
In
 a
 study
 that
 compared
 everolimus 
to
 traditional
 immunosuppressant
 treatment,
 viral
 infections
 
occurred in 31.3% of subjects receiving azathioprine versus 14.8% and 17.1% for everolimus 1.5 

17 
 
mg/day and 3.0 mg/day groups, respectively (P<0.001).  Use of everolimus reduced the risk of 
CMV infection by almost one-third as compared with azathioprine-based therapy.
24 
 
Whether induction therapy with ATG results in an increased risk of infection when used 
concomitantly with standardized triple immune suppression therapy in cardiac transplant 
recipients
 remains
 controversial
 as
 a
 randomized
 controlled
 trial
 has
 not
 yet
 been
 performed
 that
 
addresses
 this
 question.
 
 Unfortunately,
 the
 initial
 studies
 of
 ATG
 induction
 therapy
 used
 to
 obtain
 
FDA approval in renal transplant recipients cannot be applied to cardiac transplant recipients.  
Even though the immunosuppressive drugs studied in combination with ATG were the same as 
those used routinely after cardiac transplantation, ATG was administered early after 
transplantation to the delay the initiation of the nephrotoxic immunosuppressants, whereas in 
cardiac transplantation there is usually no delay in the administration of immunosuppressive 
drugs.  However, there is evidence to suggest that the concurrent administration of ATG with 
the initiation of routine triple immunosuppressive drug therapy may be safe. Concomitant 
administration of ATG with corticosteroids, azathioprine, and cyclosporine in pediatric heart 
transplant recipients was not found to result in an increased rate of infection.
47  Furthermore, 
study of ATG induction given at the same time with corticosteroids, mycophenolate mofetil, 
and tacrolimus in adolescents undergoing renal transplantation also did not result in an 
increased rate of infection.
48 
 
1.1.4 Post-transplant Lymphoproliferative Disease in Transplant Recipients 
 
Post-transplant lymphoproliferative diseases (PTLD) are another adverse event unique to 
transplant and chronic immunosuppression.  PTLD is the most common malignancy found in the 
first-year post-transplant and this is thought to be due to higher immunosuppression during 
this period.  PTLD is a type of lymphoma associated with Epstein-Barr virus (EBV), heightened 
immunosuppression, and chronic antigenic stimulation.  As PTLD consists of a heterogeneous 
group of disorders, the treatment regimen cannot be standardized.  Currently, modalities used in 
the treatment of PTLD consist of reduction of immunosuppression, radiation, surgical excision, 
monoclonal antibodies, interferon-alfa, and chemotherapy. 
 
Whether
 ATG
 is
 associated
 with
 increased
 risk
 for
 PTLD
 remains
 controversial.
 
 Again,
 a
 controlled 
trial has not yet been performed, and so far the largest retrospective study conducted on this 
topic, which was based on the Scientific Registry of Transplant Recipients (SRTR), did not show 
any difference in the relative risk of developing PTLD between any specific lymphocyte-depleting 
antibodies.
49  
 
1.1.5 Graft Tolerance 
 
Transplant tolerance is defined as full acceptance of the allograft in the absence of chronic 
immunosuppression while maintaining appropriate responsiveness to pathogens.
50  
 A growing 
body  of  
literature  supports  the  existence  of  distinct  subsets  of  cells  within  each  arm  of  the 
immune system that play a role in both inducing and maintaining transplant tolerance in vivo.  
These regulatory cells fall into three main categories—T-regulatory cells (T-regs), T suppressor 
cells,
 and
 dendritic
 cells.
51 
 
T-regs are
 generally
 CD4+CD25+
 and
 can
 inhibit
 the
 activity
 of
 dendritic
 
cells, natural killer cells, and activated T and B cells.
51  T suppressors inhibit the immune system 
by rendering antigen presenting cells tolerogenic, producing immunosuppressive cytokines such 
as TGFß and IL-10, and inhibiting the activation of CD4+ and CD8+ effector memory cells, like T-

18 
 
reg.
50  Dendritic cells (DC) are another arm of the immune system which can mediate both 
rejection and tolerance in transplantation. The milieu in which the DC interact with T-cells can 
determine whether the immune system decides to reject or tolerate the organ.  Mature DC 
promote adaptive immunity and the activation of allogeneic T-cells, whereas immature DC are 
thought to promote tolerance.
50  
 
Historically it was thought that the underlying molecular mechanism of ATG was due to an 
overwhelming depletion of T-cells that then resulted in an inability of the host’s immune system 
to react with the donor graft.  However, in recent years human and animal studies have 
demonstrated the potential for ATG to generate tolerance.   
 
ATG
 has
 been
 shown
 to
 selectively
 delete
 activated
 T-cells
 and
 activated
 B-cells,
 to
 inhibit
 cytokine
 
release by dendritic cells and activated T-cells, and to modulate adhesion and cell-trafficking 
molecules, thereby suppressing the activation of key early host immune responses to the graft.
50  
In addition to these mechanisms there is evidence that ATG can also promote the phenotype and 
expansion of regulatory T-cells and immature DC.  Two studies have demonstrated that in vitro 
treatment of lymphocytes with ATG caused the selective expansion of T-regs by promoting the 
conversion of CD4+CD25- cells to CD4+CD25+ cells followed by rapid proliferation.
52  These ATG-
generated T-reg are capable of suppressing the immune responses of activated lymphocytes in 
vitro.
52,53 
 
Furthermore, 
adoptive
 transfer
 of
 ATG-treated
 lymphocytes
 into
 animal
 models
 of
 graft
 
versus host disease caused increased survival and this effect appeared to be from the inhibition 
of
 allogeneic
 CD8+
 T-cell expansion in
 vivo.
52 
ATG also promotes the expansion of the natural killer 
regulatory T-cells (NK-T). Mice pre-treated with total lymphoid irradiation (TLI) and ATG have 
significantly
 improved
 survival
 after
 bone
 marrow
 transplant.
 The
 effect
 is
 attributed
 to
 NK-T-cells 
because the effect of TLI/ATG pre-treatment is absent in mice that are NK-T deficient.
50 
 
Lastly, in support of ATG’s tolerogenic properties is the recent case report of a renal transplant 
recipient who received TLI/ATG pre-conditioning before bone marrow and renal transplantation 
at Stanford.  This patient has demonstrated any evidence of graft rejection or graft-versus-host 
disease and has been immunosuppressant-free for 28 months.
54 
 
1.1.6 Graft Rejection 
 
Despite the significant advances made in the past decade, graft rejection remains a significant 
barrier to the survival and long term outcomes of these patients.
55  Currently, some centers 
report at least one episode of high grade acute cellular rejection during the first year following 
transplantation in up to 45% of cardiac allograft recipients.
56  Approximately 20% of all 
recipients
 experience
 AMR
 at
 some
 time
 post-transplantation
 and
 this
 percentage
 has
 not
 been
 
positively influenced by the therapies that have been effective in reducing the incidence of 
cellular rejection.
57 
 
The consequences of acute and chronic rejection are donor graft dysfunction, graft failure, and 
the
 development
 of
 CAV,
 respectively,
 that
 may
 result
 in
 death
 or
 re-transplantation. 
 
The
 acute
 
hemodynamic compromise may not resolve even with prompt and successful medical 
intervention.  With multiple episodes of rejection or chronic rejection, the donor organ becomes 
progressively fibrotic and restrictive in physiology with deterioration in systolic function.  A 
rejection episode within the first month of cardiac transplant is an independent risk factor of 
death.
58  Patients  experiencing  one  or  more  clinical  
rejection  episode that requires steroid or 

19 
 
antibody therapy during the first month after transplantation are also under a significantly higher 
risk of developing recurrent rejection within two months as well as death from recurrent 
rejection.
58 
 Multiple studies have shown
 that recurrent cellular and
 AMR
 episodes
 are associated 
with
 an
 increased
 risk
 of
 CAV,
 which
 is
 currently
 the
 major
 cause
 of
 late
 death
 in
 cardiac
 recipients.
  
In fact those patients who have accumulated more than 0.75 rejections per year have the highest 
risk for the development of CAV
29 (see Section 1.1.2 for discussion of CAV).  The secondary 
negative consequences of rejection are related to treatment.  Rejection episodes are usually 
initially treated with high dose pulse steroids and increased immunosuppression, which 
predisposes the recipients to increased morbidity due to the side effects of these agents, such as 
infectious complications, malignancies, renal failure, bone loss, and glucose intolerance. 
 
Rejection can be classified as either hyperacute, cellular, and humoral.  Hyperacute rejection of a 
transplanted
 organ
 is
 due
 to
 the
 presence
 of
 pre-formed
 antibodies and
 has
 been
 greatly
 reduced
 
by preoperative screening for alloreactive antibodies and prospective, donor-specific 
crossmatching in sensitized recipients.  Nowadays,  hyperacute  rejection  
is  an  extremely  rare 
event.59 
 However,
 acute
 cellular
 rejection
 remains a
 significant
 problem.
 
 
Acute
 cellular rejection 
is the most common form of rejection, occurring days to months after transplantation and is 
responsible for the loss of 10-20% of allografts.
55  The activated T-lymphocyte plays a major role 
in the immune cascade that leads to (1) the recruitment and proliferation of various cell lines, 
including macrophages, B cells, helper T-cells, cytotoxic T-cells, natural killer cells, etc., (2) the 
secretion of various cytokines (TNF-alpha, IFN-gamma, IL-2, IL-4, IL-5, and IL-6)  and (3) the 
subsequent damage to donor endothelium and myocytes, which pathologically manifests as 
cellular rejection.
59 
 
AMR
 in
 cardiac
 transplantation
 is
 currently
 thought
 to
 represent
 the
 consequences
 of
 antibody-
induced and complement-mediated activation of endothelial cells, secretion of cytokines, 
increased endothelial cell adherence of leucocytes and subsequent ischemic damage to the 
graft.
59  Histologically, AMR in a biopsy specimen is defined by the presence of deposits in a 
vascular pattern that are comprised of immunoglobulin, complement, and fibrinogen deposits as 
visualized by immunofluorescence; endothelial cell swelling, distending capillaries, hemorrhage, 
and 
interstitial swelling as visualized by 
light microscopy; and positive 
immunoperoxidase staining 
of paraffin-embedded tissue.
33,60  The vascular deposits are hypothesized to represent immune 
complexes from a number of mechanisms, including the formation of host cytotoxic antibodies 
against donor MHC Class II antigens (HLA-DR) found on donor endothelium; and anti-endothelial 
cell antibody generation.  Clinically, patients with AMR have an increased risk of fatal irreversible 
rejection with acute hemodynamic compromise, the development of accelerated CAV, and 
death.
33,60 
 
The mainstay of therapy is the prevention of rejection by the initiation and maintenance of 
chronic immunosuppression using triple
 drug therapy
 soon after transplantation.
  Most centers 
utilize steroids in combination with two other immunosuppressive agents, usually a calcineurin 
inhibitor (cyclosporine or tacrolimus) with a purine synthesis inhibitor (azathioprine or 
mycophenolate mofetil).  Steroids reduce the transcription of many genes involved in 
inflammation and immunity by suppressing the activation of the transcriptional regulator nuclear 
factor kappa (NF-kB)  and  enhancing  the  expression  of  
inhibitory  factor  
intracellular  kappa  B 
(IkB).
61  Cyclosporine (also known as Sandimmune and Neoral) and tacrolimus both inhibit the 
function of calcineurin, thus blocking the upregulation of IL-2 transcription and subsequently 
preventing IL-2 production and activation of the T-cell.  Historically the advent of cyclosporine in 

20 
 
the 1980s accounts for the re-emergence of heart transplantation, which was prohibited and 
abandoned in the previous decade due to high rejection and mortality rates.  Besides the lower 
incidence of side effects such as gingival hypertrophy, hypertension, and hirsutism, tacrolimus is 
now favored over cyclosporine because of two trials in which tacrolimus treated heart transplant 
patients had significantly lower rejection rates as compared to cyclosporine treated groups.
59  
Azathioprine
 (also
 known
 as
 Imuran)
 and
 mycophenolate
 mofetil 
(also
 known
 as
 MMF) 
inhibit the 
proliferation of
 lymphocytes
 by
 interfering
 with
 purine
 synthesis.
 
 The
 target
 of
 MMF,
 the
 enzyme
 
IMPDH (inosine monophosphate dehydrogenase), is a unique step in the de novo pathway of 
purine synthesis in lymphocytes, which makes MMF more lymphocyte specific than 
azathioprine.
59 
 
MMF
 affects
 not
 only
 T
 lymphocyte
 activity,
 but
 is
 also
 associated
 with
 a
 reduction
 
of B lymphocytes and a downregulation of activation markers on B cells.
62  A number of 
comparison trials suggested that MMF is more efficacious than azathioprine in reducing severe 
rejection and 1 year mortality in heart transplant recipients, resulting in the increased utilization 
of MMF over azathioprine.
59  Besides their lipid lowering effects, statins also have anti-
inflammatory and immunosuppressive properties and have been shown to be associated with 
rejection prevention in heart transplant recipients.
14-16  Pravastatin, simvastatin, and atorvastatin 
are now routinely used in cardiac recipients.  Doses are titrated to patient tolerance rather than 
to an absolute lipid level. 
   
As already mentioned above, currently the main mode of therapy for acute rejection is high 
dose pulse steroids.  Acute rejection that is refractory to steroids may be treated with a repeat 
course of high dose pulse steroids, monoclonal antibodies, methotrexate, rabbit-antithymocyte 
globulin,
 total
 lymphoid
 irradiation,
 photopheresis,
 or
 plasmapheresis,
 depending
 on
 the
 protocol
 
of
 the
 institution.
 
 Immunosuppression
 protocols
 vary
 among
 institutions
 because
 there
 are
 very
 
few prospective, randomized trials in heart transplant recipients.  Likewise, perioperative 
induction
 therapy
 with
 monoclonal
 or
 polyclonal
 antibodies is also subject to the protocol
 of
 each 
individual transplant center and is recently estimated to be used in approximately 47% of cardiac 
allograft recipients world-wide.
55  Daclizumab  and  basiliximab  are  both  monoclonal  antibodies 
directed against the IL-2 receptor (IL-2R) and due to increased mortality seen in a randomized, 
double blinded, No Induction or Control controlled trial (unpublished data), daclizumab has fallen 
out of use. 
 
Multiple studies of antibody induction therapy, both prospective and retrospective have shown 
either superior or at least equivocal results of the polyclonal antibodies over monoclonal 
antibodies with better short and long term safety profiles.
63-66  However, whether induction 
therapy using polyclonal antibodies can prevent rejection in cardiac transplant recipients has 
yet to be definitively demonstrated.  Similar to CAV, the effect of ATG induction on rejection has 
been studied in several retrospective, uncontrolled studies.  The majority of these studies have 
demonstrated that ATG induction therapy prevents rejection in cardiac transplant 
recipients.
21,42,67 
 
1.1.7 Ischemia-reperfusion injury, primary graft dysfunction, and significance to CAV and 
ATG 
 
Two basic mechanisms play an important role in IRI: (a) systemic imbalance of oxidative 
stress/antioxidant status and (b) restoration of metabolic processes which trigger the 
immune/inflammatory responses.  

21 
 
It seems that reactive oxygen species (ROS) initiate and induce the adaptive alloimmune 
response (acute rejection) predominantly through activation of antigen-presenting cells.  
Furthermore, the ROS-induced injury contributes to the development of atherosclerosis of 
donor heart vessels (chronic rejection) through endothelial injury-induced proliferation of 
smooth muscle cells.
68  Loss of oxygen supply during the ischemic period and subsequent 
reperfusion of the graft trigger the loss of osmotic equilibrium and increased permeability of 
cellular membranes, which leads to cell necrosis and decreased overall organ function.
68-73  
Furthermore, the formation of reactive oxygen species that cause direct oxidative damage to 
nucleic acids, proteins, and lipids plays an important role in aggravating cell and tissue 
damage.
71,74,75  Tissue hypoxia is only one of the factors contributing to cellular damage related 
to ischemia-reperfusion during organ transplantation.  Reperfusion also triggers the expression 
of inflammatory cytokines and adhesion molecules that increases the rate of apoptosis in the 
reperfused tissue.
69,76 
The role of white blood cells, which are closely related to the development of inflammatory 
damage in ischemia-reperfusion, has been demonstrated in various studies.
77-79  Preservation 
and revascularization which occur early in the transplantation process initiate a cascade of 
molecular and cellular events which trigger the release of proinflammatory mediators and 
attract various cell types which infiltrate the tissues.
70,78  Leukocytes have been considered to be 
responsible for many pathophysiologic changes during IRI.
80-86  They may exacerbate tissue 
hypoxia by plugging capillaries77,86,87 and mediate direct cytotoxicity by producing oxygen 
radicals
88 and proteolytic enzymes.
84 The alteration of vascular resistance during ischemia-
reperfusion is another important role of these mediators causing IRI.
79 In fact, activation of 
components of the inflammatory response exacerbates the damage already caused by the 
oxidative radicals.
85  Therefore, already existing ischemia-induced damage is further 
exacerbated by cytotoxic cells and effects on adhesion molecules.
76,89 
 
Given the pathological processes detailed above, ischemia-reperfusion injury has the clear effect 
of damage to cardiac tissue, with clinical consequences.  In general, the risk of poor immediate 
post-transplant outcome directly correlates with the cold ischemia time.  Prolonged ischemic 
time directly correlates with poorer 30-day survival
90 and is acknowledged as a risk factor for 
Primary Graft Dysfunction (PGD),
91 which itself has a high mortality rate. 
 
The definition of PGD has been controversial in heart transplantation, but a standardized 
definition has emerged following an international consensus conference in April 2013.92  PGD is 
defined by either left or right ventricular failure that occurs within 24 h of surgery.  Severity is 
graded by the affected ventricle and the nature of circulatory support required.  Survival rates 
are poor in patients with high grade PGD, with retrospective studies demonstrating a 30 day 
survival of as low as 14%.
93  The ISHLT registry indicates PGD occurs in 2-26% of heart 
transplants, although exact rates are unclear due to the aforementioned previous lack of 
standardized criteria.
94 
 

22 
 
 
Refers to text above: defenition and severity scale for primary graft dysfunction (PGD) from the 2013 
ISHLT consensus conference
 
 
PGD is associated with poorer short (30 day) and long (1 and 5 year) term survival and with an 
increased need for re-transplantation.
95  Prolonged ischemic time is a risk factor for PGD and the 
subsequent development of CAV96-98, and correlates with increased risk and frequency of 
rejection.
99  These complications have economic consequences; studies actively demonstrate 
that a decrease in ischemic time would reduce costs through reduction of length of stay in ICU 
(patients with PGD have longer length of stay in ICU).
100  Additionally, re-transplantation is 
frequently the only option for patients with severe PGD, or later on, CAV; this incurs further 
costs. 
 
The mechanism of thymoglobulin in abrogating ischemia-reperfusion injury has been thought to 
result primarily from a direct effect on blocking the cell-to-cell interactions
69 and reducing the 
degree of leukocyte rolling and adhering along capillary endothelial surfaces.
101  This effect is 
due to down modulation of adhesion molecules and specific receptors which are responsible for 
these interactions (LFA-1, VLA-4, CCR5, and CCR7).
102  Thymoglobulin can also indirectly reduce 
inflammatory mediators and inhibit leukocyte-chemotaxis or chemokine receptor 
expression.
69,101,103  For that reason, inhibition of leukocyte homing and trafficking to the graft by 
binding to chemokine receptors is another way by which thymoglobulin affects IRI.
104  
Additionally, TG reduces the number of peripheral lymphocytes from the circulating pool by 
inducing T-cell depletion through complement-related lysis or activation associated apoptosis.
105  
Moreover, it causes anergy and functional impairment of non-depleted lymphocytes and 
prevents migration of memory T-cells.
102,105  Lopez et al. showed that the therapeutic effect of 
TG is not only due to T-cell depletion, but also due to generation of regulatory T-cell.
52  As a 
polyclonal agent, directed against molecules participating in IRI, it can minimize the IRI related 
problems in the grafted organ and subsequently preventing graft failure.
106,107 
 

23 
 
Some experimental studies have been published to show benefits of ATG in reducing IRI. Preville 
et al. performed an experimental study in a non-human primate model to investigate the extent 
of T-cell depletion in lymphoid tissue after ATG usage.  The purpose of this study was to 
establish a better concept of the mechanisms of action of ATG and to determine the appropriate 
dosage of ATG in different applications.  Using skin grafts and heart transplantation models, ATG 
treatment induced a dose-dependent lymphocytopenia and T-cell depletion in spleen and lymph 
nodes due to T-cell apoptosis.
105 
 
Beiras-Fernandez et al. performed another study on two different groups of primates 
(Cynomolgus monkeys); one group was treated with ATG and the other one without ATG. The 
study was designed to evaluate the effect of ATG on the prevention of apoptosis in reperfused 
limb after ischemia and to monitor its ability to increase lymphocyte apoptosis. There was a 
significant decrease of apoptotic cells in skeletal muscle, connective tissue, and endothelial cells 
in the ATG treated animals after 60 minutes of warm ischemia. Additionally, white blood cell 
(WBC) infiltration in muscles was reduced while the apoptosis of WBCs was increased. 
Furthermore, mononuclear cells in peripheral blood, expression of adhesion molecules, and 
tissue damage were significantly decreased in the ATG treated animals. The authors concluded 
that ATG not only increased the rate of apoptosis in WBCs, but also protected the reperfused 
tissue against IRI.
69 
 
Together the data indicate that IR-initiated inflammation contributes to poorer short-term 
outcomes, risk/frequency of rejection, and development of late cardiac allograft vasculopathy, 
and that ATG may be able to improve this through reduction of IR injury.  
 
1.2 
DRUG BACKGROUND 
 
1.2.1 Description and Mechanism of Action 
 
Polyclonal antibodies were introduced in the late 1960s for the induction phase of 
immunosuppression in heart transplantation.
50  Historically, many different preparations were 
produced  at,  and  used  exclusively  by,  
individual  transplant  centers, including anti-thymocyte, 
anti-lymphoblast, and anti-lymphocyte antibodies raised in rabbits, horses, and goats.  Over the 
past 40 years, rabbit antithymocyte globulin (ATG) has emerged as the most clinically 
efficacious of the polyclonal antibodies108.  ATG comprises a heterogeneous group of 
gammaglobulins (anti-CD2, CD3, CD4, CD8, CD18, HLA class I antigens and HLA-DR) that are 
produced by injecting human lymphocytes (or lymphocyte membranes) into a rabbit.
109  The 
rabbit then develops an immune response, which leads to the formation of gammaglobulins that 
are purified for pharmaceutical use in humans.  The mechanism of action by which polyclonal 
anti-lymphocyte preparations suppress immune responses is not fully understood.  ATG is a 
pan-T-cell agent that when administered reduces lymphocyte counts to about 10% of normal 
values in humans.  The lymphocytopenia is attributed to several mechanisms, including 
complement dependent cytolysis, cell-mediated antibody-mediated cytolysis, as well as 
opsonization and subsequent phagocytosis by macrophages of the reticuloendothelial system. 
 
“Induction
 immunosuppressive
 therapy”
 refers
 to
 the
 use
 of
 antithymocyte
 antibodies
 at
 the
 time
 
of transplantation.  It is hypothesized that ATG induction therapy may enhance graft tolerance 
by blocking T-cell activation and other immune cell functions against the allograft at the time 

24 
 
of transplantation.  It has been shown that the probability of developing a high-grade rejection is 
markedly decreased  six  months  after  heart  transplantation,  resulting  in  the  reduction  of 
immunosuppression over time.
110  In addition, some clinical data seem to suggest that aggressive 
early rejection prophylaxis may alter the predisposition to reject and tolerate 
long after the period 
of early prophylaxis.
110,111 
 
It
 is
 believed
 that
 this
 graft
 tolerance
 develops
 as
 a
 result
 of
 the
 gradual
 
emergence
 of
 suppressor
 mechanisms
 through
 the
 selective
 clonal
 deletion
 of
 alloreactive
 T-cells.  
In vitro experiments have shown that ATG has (1) the highest ability to induce the apoptosis and 
activation-induced cell death of CD4+ T-cells when compared with anti-IL2R antibodies; and (2) a 
dose dependent ability to inhibit the proliferation of resting T-cells that had been activated by 
anti-CD3 mouse antibody.
112  It is thought that that this may be the mechanism by which ATG 
enhances graft tolerance. 
 
Thymoglobulin
® (Genzyme) [rabbit anti-thymocyte globulin (ATG)] is a purified pasteurized, 
gamma immune globulin, obtained by immunization of rabbits with human thymocytes.  This 
immunosuppressive product contains  polyclonal  cytotoxic antibodies directed against antigens 
expressed on human T-lymphocytes.  Thymoglobulin
® (Genzyme) includes antibodies against T-
cell markers such as CD2, CD3, CD4, CD8, CD11a, CD18, CD25, CD44, CD45, HLA-DR, HLA Class I 
heavy  chains,  and  β2-microglobulin  (see  Appendix  F).   In vitro, Thymoglobulin® (Genzyme) 
(concentrations >0.1 mg/mL) mediates T-cell suppression effects via inhibition of proliferative 
responses to several mitogens.
113-115  In patients, T-cell depletion is usually observed within a day 
from initiating Thymoglobulin
® (Genzyme) therapy.
116-118 
 
Genzyme currently holds the license for Thymoglobulin® after acquiring SangStat in 2003.  
SangStat acquired  an  exclusive  license  in  1993  from  Pasteur  Merieux  Connaught  (PMC),  a 
subsidiary of Rhone Poulenc S.A., to market this drug in the United States and Canada.  
Thymoglobulin or Thymoglobulin(e)-Merieux  is  manufactured  and  marketed  outside  North 
America by 
IMTIX, the Pasteur Merieux Connaught transplantation division, and was used 
in North 
American studies prior to marketing clearance from the FDA, which it received on December 30, 
1998.  Thymoglobulin and Thymoglobulin(e)-Merieux  are  the  trade  names still currently used 
outside the United States.  Thus, to clarify, Thymoglobulin® (Genzyme), Thymoglobulin
® 
(Sangstat), Thymoglobulin, and Thymoglobulin(e)-Merieux are all trade names used for ATG 
produced using exactly the same protocol. 
 
1.2.2 Pharmacokinetics/pharmacodynamics 
 
After an intravenous dose of 1.25 to 1.5 mg/kg/day (over 4 hours for 7–11 days) 4–8 hours post-
infusion, Thymoglobulin
® levels were on average 21.5 µg/mL (10–40 µg/mL) with a half-life of 2–
3 days after the first dose, and 87 µg/mL (23–170 µg/mL) after the last dose.  A group of 79 renal 
transplant patients were treated with Thymoglobulin
®, 1.5 mg/kg/day for 6-14 days as part of a 
double-blinded trial comparing the efficacy of Thymoglobulin® and Atgam (horse anti-thymocyte 
globulin) for acute rejection
109. Serial serum samples from the patients were tested to determine 
the level of Thymoglobulin
® (i.e. rabbit IgG levels = total Thymoglobulin
®) and anti-Thymoglobulin 
using
 ELISAs.
 Antibodies
 binding
 to
 human
 lymphocytes
 (active
 Thymoglobulin),
 were
 determined
 
by flow cytometry; no correlation was seen between treatment efficacy and either active or 
total Thymoglobulin concentrations; the overall treatment success rate was 86%. 
Pharmacokinetics of total and active Thymoglobulin® were distinctly different; active 
Thymoglobulin
® disappeared much more rapidly: only 12% of patients had detectable active 
Thymoglobulin
® by day 90 compared to 81% of patients with detectable total Thymoglobulin® 

25 
 
percent active Thymoglobulin® decreased from a peak of 0.56-0.7% during treatment, to 0.07-
0.35% by day 21, and less than 0.14% by day 30. Thymoglobulin® and active Thymoglobulin
® 
concentrations were modeled by multiple regression. Using dose number and sensitization as 
independent variables, 47-76% of the variability seen in interpatient Thymoglobulin® levels could 
be explained, while for active Thymoglobulin® levels, the measured variables accounted for 13-
48% of the observed interpatient variation.  
 
The authors concluded that: (1) for a group of patients receiving primary Thymoglobulin® 
treatment (averaging nine full and one partial dose per patient), neither Thymoglobulin® nor 
active Thymoglobulin® levels are predictive of treatment outcome; (2) active Thymoglobulin
® 
disappears more rapidly from the circulation than total Thymoglobulin®; and (3) patients that 
develop anti-rabbit IgG antibodies clear Thymoglobulin
® and active Thymoglobulin
® more rapidly 
than unsensitized patients.
119  However, another study found that after multiple administrations, 
Thymoglobulin
® clearance decreases, resulting in an extended half-life.  Thymoglobulin
® can be 
detected in the serum of recipients up to 50 days after the last dose.
109  There are a few case 
reports of Thymoglobulin
® being detected in the serum for up to a year after the last dose. 
 
Thymoglobulin
® administration is associated with significant T-cell subset depletion, general 
leucopenia, anemia, and thrombocytopenia that recovers upon cessation of therapy (see graphs 
below).  However, persistent, mild leucopenia lasting up to a year after Thymoglobulin
® has been 
observed in some case reports, though the leucopenia could have also been attributed to other 
immunosuppressive agents, such as cyclosporine, mycophenolate mofetil, tacrolimus, etc. 
 
1.2.3 Clinical Trials 
 
Most of the randomized human clinical trials using ATG are in the area of renal transplantation.  
ATG has been demonstrated to be efficacious and safe for use in acute rejection in renal (adult 
and  pediatric)  transplantation,
109,120 intra-operative  induction  therapy  in  renal  
transplantation,
106,121 induction therapy in pediatric cardiac transplantation,
47 as well as 
outpatient infusion for acute renal rejection122.   
 
1.2.3.1 Human Clinical Trials of ATG in Renal Transplantation  
 
(a)
 
 The
 marketing
 clearance
 application
 for
 Thymoglobulin
® 
was
 based
 on
 a
 single
 double-blinded, 
randomized, multi-center Phase III trial, which as conducted at 28 leading U.S. renal transplant 
centers.  In this trial, Thymoglobulin® was compared to Atgam (horse-derived polyclonal 
antibody) in the treatment of renal transplant patients (n=163) with biopsy-proven Banff grade 
2
 (moderate),
 grade
 3
 (severe),
 or
 steroid-resistant
 grade
 1
 (mild)
 acute
 graft
 rejection. 
 
Patients 
were randomized to receive 7 to 14 days of Thymoglobulin® (1.5 mg/kg/day) or Atgam (15 
mg/kg/day).  For the entire study, the two treatment groups were comparable with respect to 
donor and recipient characteristics.  There were no significant differences between the two 
treatments with respect to (i) day 30 serum creatinine levels relative to baseline; (ii) 
improvement rate in post-treatment histology; (iii) one year post rejection Kaplan-Meier 
patient survival (Thymoglobulin® 93%, n=82 and Atgam 96%, n=80); (iv) day 30 and (v) one-year 
post rejection graft survival (Thymoglobulin® 83%, n=82 and Atgam 75%, n=80). 
 

26 
 
 
 
Refer to section (a) above: Overall 1-year post-rejection therapy actuarial graft survival (Kaplan-Meier 
method), by acute rejection severity for renal transplant recipients treated with either Thymoglobulin or 
Atgam. 
 
(b)  In another study also comparing the efficacy and safety of Thymoglobulin® to Atgam for 
induction in adult renal transplant recipients, 72 patients were randomized 2:1 in a double-
blinded
 fashion
 to
 receive
 Thymoglobulin® 
(n=72)
 at
 1.5
 mg/kg/day
 intravenously
 or
 Atgam
 (n=24)
 
at 15 mg/kg/day intravenously, intraoperatively, then daily for at least 6 days.  By 1 year after 
transplantation, 4% of Thymoglobulin
® treated  patients  experienced  acute  
rejection  compared 
with 25% of Atgam treated patient (p=0.14).  Both the rate and severity of acute rejection was 
significantly lower in with Thymoglobulin
® than Atgam and no recurrent rejection occurred with 
Thymoglobulin
® compared with 33% with Atgam (but p=NS).  Patient survival was not different, 
but the composite end point of freedom from death, graft loss, or rejection, the “event-free 
survival,” was superior with Thymoglobulin
® (94%) compared with Atgam (63%, p=0.0005).  
Fewer adverse events occurred with Thymoglobulin
® (p=0.013).  Leucopenia was more common 
with Thymoglobulin® than Atgam (56% vs 4%, p<0.0001) during induction.  The incidence of 
cytomegalovirus disease was less with Thymoglobulin
® than Atgam at 6 months (10% vs 33%, 
p=0.025).123 
 

27 
 
 
Refer to section (b) above: Graph A: Kaplan-Meier allograft survival. Allograft survival was superior with 
Thymoglobulin® compared with Atgam, P=0.021, log-rank test when graft losses from all causes were 
considered. Numbers in parentheses are the number of patients remaining at risk. Graph B: Kaplan-Meier 
event-free survival. Event-free survival, defined as freedom from rejection, death, and allograft loss, was 
superior with Thymoglobulin
® compared with Atgam. P=0.0005, log-rank test. Numbers in parentheses are 
the number of patients remaining at risk. 
 
(c)  An open, randomized, multicenter trial investigated induction therapy in non-
hyperimmunized patients receiving their first cadaveric renal allograft with either 
Thymoglobulin® (1.0-1.5 mg/kg/day [dose adjusted daily to keep CD2 or CD3 counts below 
20/mm3] plus delayed cyclosporine (n=50) or basiliximab [an inhibiting IL-2 receptor 
monoclonal antibody] (20mg/day on days 0-4) plus early cyclosporine (n=50).  All the patients 
were also treated with steroids and mycophenolate mofetil.  Patient and graft survival rates at 
12
 months
 were
 98
 and
 94%
 in
 the
 basiliximab
 group,
 respectively,
 compared
 with
 100
 and
 96%
 
in the Thymoglobulin
® group.  The incidences of biopsy-confirmed acute rejection (8% in both) 
and treatment failure (8% in Thymoglobulin
® and  14%  in  basiliximab)  were  not  significantly 
different.  The incidence of CMV infection and need for dialysis was also not statistically 
significantly different between the two groups.
124 
 

28 
 
Refer to text in section (c) above: 
 
 
(d)  Thymoglobulin® induction has been shown to be safe and effective in combination with 
triple immunosuppressive therapy for preventing early rejection in pediatric renal transplant 
recipients.  Seventeen pediatric renal transplant recipients (mean age 10.1+/-5.2 years) received 
either primary or second transplants between 1 August 1999 and 31 July 2001. One patient had 
primary
 allograft
 non-function
 secondary
 to
 vascular
 thrombosis.
 Two
 patients
 (12%)
 had
 delayed
 
allograft function. Immunosuppression consisted of Thymoglobulin® induction (1.5 mg/kg/dose 
for a mean number of doses 6+/-1.7) with tacrolimus (62%) or cyclosporine A (38%), 
mycophenolate mofetil, and prednisone. One year post transplant, patient and graft survival 
was
 100%
 and
 93%,
 respectively.
 No
 acute
 rejection
 episodes
 occurred
 during
 the
 first
 6
 months
 
after transplantation in any of the recipients. Additionally, no rejection episode occurred among 
the 14 patients followed for 1 year after transplant. The incidences of asymptomatic 
cytomegalovirus (CMV) and Epstein-Barr virus (EBV) seroconversion at 1 year in seronegative 
recipients with a seropositive donor were 100% of 4 patients and 0% of 4 patients, respectively. 
No symptomatic CMV or EBV infections and no post-transplant lymphoproliferative disease 
occurred in any patient over the course of 1 year of follow-up.125 
 

29 
 
Acute rejection episodes 
 
 6 months (16 patients) 
0 
 12 months (14 patients) 
0 
Estimated creatinine clearance 
 
Baseline (ml/min per 1.73 m
2) 
77.3±23.7 
 3 months 
77.9±24.2 
 6 months 
88.2±28.2 
 12 months 
74.2±14.8 
Refer to text in section (d) above:  Short-term outcome for 16 pediatric kidney transplant recipients who 
received Thymoglobulin
® induction therapy 
 
1.2.3.2 Human Clinical Studies of ATG in Cardiac Transplantation 
 
To date
 all
 studies of
 ATG
 in
 cardiac
 transplantation
 have
 been
 either
 uncontrolled,
 retrospective, 
or comparing another agent whose efficacy is unknown. 
 
(a)  A total of 484 primary cardiac transplanted patients received induction therapy with two 
different rabbit-ATG (Thymoglobuline-Merieux: n=342, ATG-Fresenius: n=142). All patients 
received immunosuppressive maintenance therapy with cyclosporine, azathioprine, and 
prednisolone. Cardiac rejection was assessed by serial endomyocardial biopsies. Surveillance of 
graft arteriosclerosis was performed by angiograms 1, 3, and 5 years after transplantation. Five-
year survival was significantly better in the Thymoglobuline group (76 vs. 60%). 
Thymoglobuline-Merieux
 patients
 had
 a
 lower
 rate
 of
 death
 from
 rejection
 (2.3
 vs.
 10%;
 P<0.01)
 
and graft arteriosclerosis (0.88 vs. 5.6%; P<0.01). After 5 years, freedom from rejection was 72% 
in the Thymoglobuline-Merieux  group  compared  
to  42%  
in  
the  ATG-Fresenius group (P<0.01). 
Graft arteriosclerosis appeared in 14% of Thymoglobuline patients and in 28% of ATG-Fresenius 
patients (P<0.01). Viral infections occurred more often in Thymoglobuline patients (53 vs. 39%, 
P<0.05) although there was no difference in appearance of cytomegalovirus disease (17 vs. 13%). 
Freedom from posttransplant malignant disease was comparable between the two groups
67. 
 

30 
 
 
 
 
 
Refer to text of section (a) above:  Actuarial survival and causes of death up to 10 years after cardiac 
transplantation comparing thymoglobuline versus ATG-fresenius 
 
(b)  A retrospective analysis of 163 patients who were administered a 3-day course of 
intravenous Thymoglobuline-Merieux immediately following heart transplantation from 1988 
to 1998 (called Group 1) compared with 48 patients transplanted from 1983 to 1987 and during 
an isolated period in 1994 where no induction therapy was used and intravenous and oral 
cyclosporine was used immediately following heart transplantation (called Group 2). Routine 
endomyocardial biopsies were performed in all patients. One, 5- and 10-year actuarial survival 
rate averaged 85%+/-3, 77%+/-4 and 67%+/-5 in Group 1 compared with 88%+/-5, 81%+/-6 and 
76%+/-6 in Group 2 (p = 0.5). At 1 year, the freedom rate from an episode of acute rejection 
averaged 43%+/-4 in Group 1 and 30%+/-7 in Group 2 (p = 0.03) and the freedom rate from an 
episode of infection averaged 44%+/-4 in Group 1 and 31%+/-7 in Group 2 (p = 0.2). At 1, 5 and 
10 years, the freedom rate from graft coronary artery disease averaged 93%+/-2, 68%+/-5 and 
50%+/-7 in Group 1 compared with 93%+/-4, 58%+/-8 and 30%+/-8 in Group 2 (p = 0.1) and the 
freedom rate from cancer averaged 98%+/-1, 91%+/-3 and 67%+/-8 in Group 1 compared with 

31 
 
100%,
 95%+/-3
 and 77%+/-8
 in Group 2
 (p
 =
 0.2).  There
 was no side
 effect related
 to the
 systemic 
injection of Thymoglobuline-Merieux.  Thus, after induction with Thymoglobuline-Merieux, the 
risk
 of
 infection
 and
 malignancy
 was
 not
 increased 
and
 there
 was
 a
 non-significant
 trend
 towards
 
a lower incidence of coronary atherosclerosis 5 and 10 years after transplantation.
42 
 
 
Refers to text in Section (b): Actuarial freedom rate from rejection.  Group 1=patients treated with 
thymoglobuline (Thymo). Group 2=patients not treated with thymoglobuline (Control) 
 
 
Refers to text in Section (b): Actuarial freedom rate from CAV. 
Group
 1=patients
 treated
 with
 thymoglobuline
 (Thymo).
 Group
 2=patients
 not
 treated
 with
 thymoglobuline
 
(Control). 
 

32 
 
 
Refers to text in Section (b): Actuarial freedom rate from infection.  Group 1=patients treated with 
thymoglobuline (Thymo). Group 2=patients not treated with thymoglobuline (Control). 
 
 
 
Refers to text in Section (b): Actuarial freedom rate from cancer. 
Group
 1=patients
 treated
 with
 thymoglobuline
 (Thymo).
 Group
 2=patients
 not
 treated
 with
 thymoglobuline
 
(Control). 
 
(c)  Another retrospective study examined the use of Thymoglobuline-Merieux in pediatric 
cardiac transplantation over a 13-year period in a single-center with respect to the short-term 
hematological effects as well as longer-term outcomes. The dose of Thymoglobuline given 
depended on baseline platelet count and was 2, 1.5, or 1 mg/kg per day over 5 days for the 
following platelet count groups: greater than 150,000/mm (normal group), 100 to 150,000/mm 
(mild thrombocytopenia group), and 50 to 100,000/mm (moderate thrombocytopenia group). 
Thirty
 children
 of
 median
 age
 14.2
 years
 were
 given
 a
 median
 cumulative
 dose
 of
 Thymoglobuline
 
of
 8
 mg/kg
 per
 patient;
 the
 moderate
 thrombocytopenia
 subgroup
 was
 given
 significantly
 less
 (6.4
 
mg/kg) (P=0.032).
 Immediate
 tolerability
 of
 Thymoglobuline was good,
 with no cases of
 first-dose 

33 
 
syndrome, anaphylaxis, or serum sickness. The platelet count decreased at the start of therapy, 
but recovered after discontinuation, and did not give rise to clinical concern. Patients were 
followed up for a median of 6.3 years (7 days-15.5 years); actuarial survival was 90%, 86%, and 
74.5%, respectively, at 1, 5, and 10 years. In the first year, 50% of patients suffered an episode of 
rejection. The overall incidence of infection in the month following transplantation was 40%. One 
lymphoma occurred at 5 months. The authors concluded that the use of Thymoglobuline-
Merieux in pediatric heart-transplant patients as part of an immunosuppressive protocol, with 
dose adjustment according to platelet levels, is effective in terms of a decreased rejection rate 
and
 improved
 patient
 survival,
 and
 safe
 in
 terms
 of
 the
 incidence
 of
 infections
 and
 malignancy.
21 
 
 
Refer to text in section (c):  Patients’ actuarial survival rates at 1, 5, and 10 years (n=30). 
 
 
Refer to text in section (c):  Freedom from cytomegalovirus with time (n=30). 
 

34 
 
(d)  In still yet another single-center, retrospective study, 31 consecutive heart transplant 
recipients (mean age, 7.8 years; median age, 9 years; range, 4 months-17 years), who all 
survived surgery had induction with Thymoglobuline-Merieux at age-dependent doses (1-1.5 
mg/kg/day between 0 and 1 year; 1.5-2 mg/kg/day from 1 year to 8 years; and 2.5 mg/kg/day >8 
years). Duration of treatment was 1 to 7 days. In patients <1 year old, the total number of 
lymphocytes was maintained
 at
 >500/mm
3,
 in
 patients between 1
 and
 8
 years
 old
 >300/mm3, and 
in patients >8 years old >150/mm
3. Thirty of 31 patients were alive at the end of follow-up. 
During the first 3 months, 3 Grade 3A and 10 Grade 1A (Working Formulation grading system) 
rejection episodes occurred. All reversed after steroid treatment. Eleven viral infections, 2 
bacterial infections, and 1 fungal infection occurred. Not all patients with infection were 
symptomatic but all responded successfully to treatment. Given that there was no comparison 
group, it is difficult to infer a possible increase in the rate of infection, but in the discussion the 
authors remarked that their rate of infection did not “substantially differ from risks reported in 
the literature for patients who did not receive induction therapy.”  One episode of post-
transplantation lymphoproliferative disease regressed after decreasing immunosuppression 
therapy and after acyclovir therapy.  The authors concluded that immunosuppression therapy 
with Thymoglobuline is safe even in early infancy.126 
 
(e)  In another retrospective, single-center study 30 patients (ages 8 months to 24 years) with 
end-stage
 heart
 failure
 underwent
 cardiac
 transplantation: 
12
 (40%)
 for
 postoperative
 end-stage 
heart failure, 9 (30%) as primary treatment for congenital heart disease, 5 (17%) for dilated 
cardiomyopathy, and 4 (13%) for restrictive/hypertrophic cardiomyopathy. Nineteen patients 
(63%) had undergone prior operations; 4 patients received transplants for failed Fontan 
procedures. Induction therapy with rabbit antithymocyte globulin (DCI Laboratories) was used 
routinely, and long-term immunosuppression was by cyclosporine and azathioprine alone. 
Rejection surveillance/diagnosis was based on echocardiographic criteria. Post transplantation 
follow-up ranges from 3 to 78 months. Operative mortality was 3.3% (1/30). No patients were 
diagnosed with either infection, accelerated allograft atherosclerosis, or post transplantation 
lymphoproliferative disease.
47 
 
(f) A retrospective, multicenter study in the United Kingdom was performed, encompassing over 
2,000 patients between 1995 and 2008, 1000 of whom had been inducted with ATG.  The study 
found no significant difference in survival at 10 years between the two groups: 56.2% in the ATG 
group versus 55.9% in the no-ATG group (p = 0.95). The investigators did note lower rates of 
rejection over the first year (incidence rate ratio, 0.76; 95% confidence interval [CI], 0.68–0.85, p 
< 0.01), but this potential benefit was accompanied by increased rates of infection.
127 

35 
 
 
Refer to text in section (e): Survival after transplantation.  There has been one operative death (3.3%) and 
no deaths in post-transplantation follow-up.  
 
 
1.3 
RATIONALE FOR PERFORMING THE STUDY 
 
1.3.1 The Efficacy of ATG Induction in Cardiac Transplantation is Unknown 
 
There is limited clinical research examining the efficacy of ATG at reducing the incidence of de 
novo donor specific antibody production after heart transplantation. A study in kidney 
transplantation
 demonstrated
 a
 lower
 incidence
 of
 de
 novo 
DSA
 in
 moderately
 sensitized
 patients. 
A series of 114 consecutive, moderately sensitized kidney transplant patients received either ATG 
(n = 85) or basiliximab (n = 29) induction. At 1 year post-transplant, patients who received ATG 
induction had a decreased risk of developing de novo DSA (HR 0.33, 95% CI 0.09-1.24) and the 
group as a whole had a lower sum total de novo DSA level as measured by MFI (455 versus 3,652 
p
 =
 0.02).
128 
In
 a
 single-center, retrospective 
study of 217 consecutive sensitized (PRA ≥ 10%) heart 
transplant recipients, patients were divided into those receiving ATG (n = 162) and those who did 
not (n = 55). Patients treated with ATG had greater freedom from de novo DSA development 
compared to no ATG at 1 year after transplant (86.4% versus 74.5%, p = 0.038).
129 
 
Ischemia-reperfusion injury (IRI) can damage the allograft vascular endothelium leading to 
vasculopathy. Animal models have demonstrated that ATG reduces IRI. A study of cynomolgus 
monkeys found that circulating leukocytes were lower in monkeys given ATG 30 minutes prior to 
reperfusion of ischemic limbs.
130 In a similar study, expression of adhesion molecules (ICAM-1, 
VCAM, PECAM, CD11b, CD62E) and inflammatory cytokines (IL-1, IL-6, TNF-α) were decreased in 
the ischemic limbs of cynomolgus monkeys treated with ATG 30 minutes prior to reperfusion.
131 
A human model utilizing human umbilical vein endothelial cells (HUVECs) corroborated these 
findings. HUVECs were bathed in a solution of ATG before and after stimulation with TNF-α. Cells 
that  were  not  bathed  
in  ATG  served  as  controls.  Adhesion  molecule  expression  (ICAM-1 and 
CD62E) was reduced in the ATG group.132 A study of human serum demonstrated that ATG 
modulates leukocyte responses by affecting cellular adhesion molecules and chemokine 
receptors.
102 Studies in heart transplant recipients are limited to a single retrospective study. A 
series of 330 consecutive heart transplant patients was divided into those who received ATG 

36 
 
induction and those who did not. Patients who received ATG induction (n = 129) had significantly 
greater freedom from any ischemia-reperfusion injury on biopsy in the first month after 
transplant compared with patients receiving no ATG (n = 201) (74.8% vs 63.5%, p = 0.019).
133 
 
Clinical evidence for the use of peri-operative induction therapy to prevent CAV and rejection in 
adult cardiac transplantation is limited. Whether to use cytolytic therapy to either prevent 
rejection or treat acute rejection has been based on personal preferences because, to date, there 
have not been any randomized clinical trials conducted to evaluate the efficacy of ATG induction 
therapy in cardiac transplantation. 
 
A
 small
 number
 of
 trials 
have examined the effects of 
induction therapy 
in cardiac transplantation.  
Two of these were randomized control trials looking at IL-2R antagonists versus no induction and 
showed a reduction in the risk of rejection but no survival benefit.
134,135   Two further trials 
compared ATG induction against IL-2R antagonist induction.  
 
However, the
 majority
 of
 the
 retrospective
 studies
 in
 heart
 transplant
 recipients
 do
 suggest
 that
 
ATG induction may be beneficial.  A large retrospective study examined 9,324 heart transplant 
recipients in the ISHLT database who received induction therapy with either ATG or basiliximab.  
The study found that patients treated with basiliximab had higher mortality rates, higher rates of 
infection, and suffered more episodes of graft failure than patients treated with ATG.
136 
 
 
 
Refers to text in above paragraph: Comparison of all-cause mortality probability between the basiliximab 
and ATG groups 
 

37 
 
A recent retrospective study compared 196 heart transplant recipients at a single center who 
received ATG induction therapy for renal insufficiency with patients who received no induction 
therapy. ATG appeared to dampen the formation for de novo DSAs in the first 12 months post-
transplant.
137 
 
In  another retrospective study 662 adult cardiac recipients who had received triple-drug 
immunosuppressive maintenance together with ATG induction therapy were evaluated for the 
presence and severity of CAV.  ATG induction therapy was shown to have a protective effect 
against the development of CAV.
43,138 
 
Multivariate Analysis of Risk Factors Affecting Development of CAV After Cardiac 
Transplantation 
 
Table refers to text in the paragraph above
43 
 
In another retrospective analysis, 163 adult patients transplanted between 1988 and 1998 who 
were
 administered
 a
 3-day 
course
 of
 ATG
 induction
 were
 compared
 with
 a
 cohort
 of
 patients
 who 
underwent the same immunosuppressive therapy without ATG induction between 1983 and 
1987.  ATG induction was associated with a significantly lower rate of acute rejection and a 
trend towards a decrease in the incidence of CAV.  Moreover, the risk of infection and cancer 
was not increased.
42  See next four figures: 
 
 
Refers to text in above paragraph: Actuarial freedom rate from rejection. 
Group 1=patients treated with thymoglobuline (Thymo). Group 2=patients not treated with 
thymoglobuline (Control) 

38 
 
 
Refers to text in above paragraph: Actuarial freedom rate from CAV.  Group 1=patients treated with 
thymoglobuline (Thymo). Group 2=patients not treated with thymoglobuline (Control). 
 
 
Refers to text in above paragraph: Actuarial freedom rate from infection.  Group 1=patients treated with 
thymoglobuline (Thymo). Group 2=patients not treated with thymoglobuline (Control). 
 

39 
 
 
Refers to text in above paragraph: Actuarial freedom rate from cancer.  Group 1=patients treated with 
thymoglobuline (Thymo). Group 2=patients not treated with thymoglobuline (Control). 
 
Another retrospective analysis of 40 adult cardiac recipients treated with 5 days of ATG induction 
therapy did not show a difference in morbidity, but also confirmed the relative safety of ATG in 
that there was no increase in the incidences of CAV, malignancy, or infection.
139 
 
Results of another retrospective study in which 117 adult patients undergoing cardiac 
transplantation were administered a 3-day course of ATG induction also showed a significantly 
lower early rejection rate.  Although patient survival was similar to results reported by others, 
ATG induction was again not found to be associated with an increased incidence of malignancy 
or infection.
140 
 
In a case series report 30 pediatric patients (age >6 months) who underwent cardiac 
transplantation
 between
 1988
 and
 1994
 at
 a
 center
 where
 ATG
 induction
 was
 used
 routinely
 were
 
followed.  Over the eight years, there were no re-transplantations, no diagnoses of CAV, and no 
occurrence of post-transplantation lymphoproliferative disease.
47 
 
 

40 
 
 
Refer to text in paragraph above: Survival after transplantation.  There has been one operative death 
(3.3%) and no deaths in post-transplantation follow-up. 
 
The utility and applicability of the retrospective studies discussed above is limited by either the 
lack of controls or the use of recipients from nearly a decade earlier as controls for comparison.  
It is well documented that the clinical practice of heart transplantation has changed significantly 
from decade to decade with a constant improvement in survival and decrease in rejection rate.
55  
Given this limitation one cannot attribute these outcomes purely to ATG induction therapy. 
 
Whether ATG induction in adult heart recipients should be used cannot be based on studies of 
non-heart organ transplants or even the literature on pediatric heart transplantation because 
the immunogenicity of the cardiac allograft differs from that of other organ allografts and the 
pediatric immune system is markedly different from that of the adult.
141  Lastly, it is well known 
that in vitro studies do not necessarily always translate into the predicted clinical outcome.  
Therefore,
 whether
 or
 not 
perioperative
 ATG
 induction
 therapy
 is
 clinically
 beneficial
 with
 respect
 
to decreasing rejection and preventing CAV remains theoretical. 
 
1.3.2 Thymoglobulin has numerous mechanisms of immunosuppression 
 
Some retrospective studies have suggested that aggressive early rejection prophylaxis using ATG 
may alter the predisposition to reject and tolerate the graft.
21  These beneficial effects were seen 
long after the period of early prophylaxis. These studies also correlated with other retrospective 
studies of ATG treated recipients which demonstrated long term improvement in survival and 
other
 end
 points
 such
 as
 rejection
 and
 CAV.
43,138 
 
As
 mentioned
 above
 in
 the
 “Disease
 Background”
 
(Section 1.1.1), CAV is thought to be associated with AMR and cellular rejection and related to 
both the time to onset of rejection as well as frequency of rejections.  Thus, it is reasonable to 
hypothesize that ATG may inhibit CAV by preventing rejection.  As discussed in Section 1.2.1, ATG 
may enhance graft tolerance (and thereby prevent acute rejection) by inducing the apoptosis and 
activation-induced cell death of CD4+ T-cells, inhibiting IL-2 receptor expression, and inhibiting 
the
 proliferation
 of
 activated
 T-cells.
112,114 
 
Another
 possible
 mechanism
 is through
 preventing
 the 
formation of anti-donor antibodies as ATGs have been found to inhibit B cell proliferation and 
differentiation into immunoglobulin-secreting cells.
113  Another possible mechanism is through 
inhibiting  
the  
formation  of  
immune  complex  deposits  by  directly  or  
indirectly  
inactivating  
the 

41 
 
various components of these deposits (i.e. complement, immunoglobulins, and fibrinogen [see 
Section 1.1.2]) and thus preventing the vascular injury of AMR.  Although ATG is believed to be a 
heterogeneous group of predominantly anti-T-cell gammaglobulins (anti-CD2, CD3, CD4, CD8, 
CD18), antibodies directed against HLA class I antigens, HLA-DR, and beta2 microglobulin are 
inexplicably present. Thus, strangely the activity of ATG appears to expand beyond the realm of 
merely
 modulating
 T-cell
 activity.
 From
 this
 one
 could
 possibly
 hypothesize
 that
 the
 direct
 binding
 
of ATG to the pre-/peri-operatively injured endothelium may shield the exposed donor antigens 
from the recipient’s immune system, thus pre-emptively arresting the initial events involved in 
development of CAV. 
 
Human and animal studies have
 demonstrated the ability
 of ATG to selectively delete activated T-
cells and activated B-cells, to inhibit cytokine release by dendritic cells and activated T-cells, and 
to modulate adhesion and cell-trafficking molecules, thereby suppressing the activation of key 
early host immune responses to the graft.
50  In addition to these mechanisms there is evidence 
that ATG can also promote the phenotype and expansion of regulatory T-cells and immature DC.  
Two recent studies have demonstrated that in vitro treatment of lymphocytes with ATG caused 
the selective expansion of T-regs by promoting the conversion of CD4+CD25- cells to CD4+CD25+ 
cells followed by rapid proliferation.
52  These ATG-generated T-reg are capable of suppressing the 
immune responses of activated lymphocytes in vitro.
52,53  Futhermore adoptive transfer of ATG-
treated lymphocytes into animal models of graft versus host disease caused increased survival 
and this effect appeared to be from the inhibition of allogeneic CD8+ T-cell expansion in vivo.
52  
ATG also promotes the expansion of the natural killer regulatory T-cells (NK-T). Mice pre-treated 
with total lymphoid irradiation (TLI) and ATG have significantly improved survival after bone 
marrow transplant.  The effect is attributed to NK-T-cells because the effect of TLI/ATG pre-
treatment is absent in mice that are NK-T deficient.
50 
 
1.4 
SIGNIFICANCE OF STUDY 
  
 
Despite major advances in medical therapy, CAV remains the major factor limiting long term 
survival in heart transplant recipients.  The limited options for these patients justify the need to 
seek potential alternative treatment options.   
 
The majority of the retrospective studies in both renal and heart transplant recipients suggest 
that Thymoglobulin
® induction has a protective effect against the development of graft 
vasculopathy.
42-44  Transplant vasculopathy is also seen in other transplanted solid organs such as 
the liver, kidney, and lung, known as vanishing bile duct syndrome, renal chronic rejection, and 
obliterative bronchiolitis, respectively.  Thus, the potential benefit of Thymoglobulin® induction 
therapy may not be limited to CAV.  Unfortunately, the utility and applicability of these 
retrospective studies are limited by either the complete lack of controls or the use of recipients 
from nearly a decade earlier as controls for comparison.  It is well documented that the clinical 
practice of solid organ transplantation has changed significantly from decade to decade with a 
constant improvement in survival and decrease in rejection rate.  With these severe limitations 
one cannot attribute these study outcomes to Thymoglobulin induction therapy.  Therefore, a 
prospective, randomized, controlled trial is needed to definitively establish Thymoglobulin’s 
potential efficacy in preventing CAV. 
  
 

42 
 
2. 
STUDY OBJECTIVES 
 
2.1 
PRIMARY OBJECTIVES 
 
(A) To describe between treatment groups the incidence of the composite primary endpoint of 
the development of de novo donor specific antibodies and ischemia on endomyocardial 
biopsy at 12 months post-transplantation 
 
2.2 
SECONDARY OBJECTIVES 
 
To compare between treatment groups:   
 
(A) Number of patients who develop cardiac allograft vasculopathy (CAV) (defined as a 
change ≥0.5mm in maximal intimal thickness (MIT) of the coronary arteries by 
intravascular ultrasound at 12 months as compared to baseline) 
(B) correlations between significant changes in immune cell profiles and circulating 
antibodies, and their relation to any significant differences in clinical outcomes 
(C) Number of patients who experience acute cellular, antibody-mediated, hemodynamic 
compromise, and any-treated rejection within first 12 months after transplantation 
(D) Number of patients with biopsy proven cellular rejection ≥2R, biopsy proven antibody 
mediated rejection ≥AMR1 and any-treated rejection at 12 months post-transplantation 
(E) Number of acute cellular, antibody-mediated, hemodynamic compromise and any-
treated rejection episodes per patient within the first 12 months post-transplantation 
(F) Freedom from acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection by the ISHLT biopsy grading scale in the first 12 months post-
transplantation 
(G) Time to first acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection within the first 12 months 
(H) the incidence of Primary Graft Dysfunction (PGD) in the first 24 hours post-transplant 
(I) Patient and graft survival at 12 months post-transplantation 
(J) the types and number of patients with both fatal and non-fatal infectious complications 
(especially CMV infection) within the first 12 months post-transplantation 
(K) freedom from the development of circulating antibodies within the first 12 months post 
transplantation, where circulating antibodies include donor specific antibodies (DSA), 
non-specific antibodies, and non-human leukocyte antigen antibodies 
(L) Change in coronary maximal intimal thickness, intimal area, intimal volume, vessel area, 
intimal index and percent atheroma volume (PAV) at matched sites by intravascular 
ultrasound at 12 months post-transplantation 
(M) 
Maintenance doses of mycophenolate mofetil, tacrolimus, sirolimus, and cumulative 
dose of corticosteroids at 12 months post-transplantation 
(N) Number of hospital days per patient, both during the transplant period and during the 
post-transplant period at 3 months, 6 months, and 1 year 
(O) Number of patients requiring hospitalization by 3 months, by 6 months, or by 1 year 
post-transplantation 
 
  
3. 
STUDY DURATION AND STUDY EARLY TERMINATION 
 

43 
 
3.1 
PARTICIPANT COMPLETION 
 
The study was initiated in September 2018 and enrollment will continue until October 31, 2024.  
The study will be completed 24 months after the last patient has entered.  Study completion is 
expected by October 31, 2025 
 
3.2 
PREMATURE TERMINATION OF A PARTICIPANT FROM THE STUDY 
 
Participants may be prematurely terminated from the study for the following reasons: 
 
1. The participant elects to withdraw consent from all future study activities, including 
follow-up. 
2. The participant is “lost to follow-up” (i.e., no further follow-up is possible because 
attempts to reestablish contact with the participant have failed).  
3. The participant dies. 
4. The Investigator and/or the Medical Monitor no longer believes participation is in the 
best interest of the participant. 
 
3.3 
PREMATURE DISCONTINUATION OF STUDY DRUG 
 
Study therapy will be prematurely discontinued for any participant if the participant has 
anaphylaxis, severe cytokine release syndrome or other hypersensitivity reactions.  The 
management of severe infusion reactions should be dictated by the signs and symptoms of the 
reaction. Medical personnel with experience in treating anaphylaxis should be available. ATG 
administration in patients with a history of ATG anaphylaxis is not recommended. 
 
Study therapy may also be prematurely discontinued for any participant if the investigator 
believes that the study treatment is no longer in the best interest of the participant, if the 
subject is judged non-compliant, or due to other safety concerns.  Participants who become 
pregnant during the study treatment period will discontinue investigational agents. 
 
 
3.4 
STUDY STOPPING RULES 
 
The principal investigator and Data and Safety Monitors will review safety data on an ongoing 
basis.  If a safety concern arises, enrollment and randomization of participants in the trial will be 
suspended pending Data and Safety review.  Subjects already randomized will continue to be 
treated per protocol. 
 
The criteria described below provide additional guidance for suspending trial 
enrollment/randomization based on the occurrence of selected adverse events. Because of the 
multiplicity of stopping guidelines each has been formulated to occur when the cumulative 
number of adverse event of concerns in either treatment group meets or exceeds a pre-defined 
threshold.  Selected adverse events of concern and their thresholds in this trial are: 
 
• 
Post-Transplant Lymphoproliferative Disorder (PTLD) 
• 
Infectious complications 
• 
Death 

44 
 
• 
Treated acute rejection with hemodynamic compromise  
 
If a criterion is satisfied, the trial will be placed on hold pending Data and Safety review. 
 
3.4.1 Post-Transplant Lymphoproliferative Disorder (PTLD) 
 
The study will be placed on hold pending Data and Safety review if there is an occurrence of any 
case of PTLD reported in any randomized subjects at any time during the study. 
 
3.4.2 Infectious Complications 
 
Infectious complications are defined as infections that require hospitalization or treatment with 
intravenous antibiotics. The study will be placed on hold pending Data and Safety review if the 
observed subject-based event rate in either group exceeds 30%. This rule will be met if the 
lower one-tailed 95% confidence limit on the estimated rate of infectious complications in either 
treatment group is greater than 0.30. 
 
3.4.3 Death 
 
Death from any cause is of concern early in the trial, when the threshold incidence rate of 
concern is 15%. That is, the rule will be met if the lower one-tailed 95% confidence limit on the 
estimated rate of death in either treatment group is greater than 0.15. 
 
3.4.4 Rejection 
 
Throughout the trial there would be safety concerns if the rate of subjects with any treated 
acute cellular rejection associated with hemodynamic compromise within 4 months of 
treatment exceeds 10%. This rule will be considered to have been met and the study will be 
placed on hold pending Data and Safety review if the lower one-tailed 95% confidence limit on 
the estimated acute rejection associated with hemodynamic compromise rate (within 4 weeks 
of treatment) is greater than 0.10. 
 
4. 
CENTER OF STUDY 
  
 
This study began as a single center study conducted at Cedars-Sinai Medical Center. As a multi-
center study, participating centers include Cedars-Sinai Medical Center and Kaiser Permanente 
Medical Center.    
 
5. 
SELECTION CRITERIA 
 
5.1 
TOTAL NUMBER OF PATIENTS 
 
A total of 60 patients will be randomized in the study. 
 
5.2 
RECIPIENT INCLUSION CRITERIA FOR STUDY ENTRY 
 
The following criteria will apply to the patients enrolled in this study: 
 

45 
 
(1) Men and non-pregnant women must be 18 to 74 years old 
(2) Women of childbearing potential must have a negative serum pregnancy test prior to 
transplantation.   
(3) Men with a female partner of childbearing age and women of childbearing potential 
must use two reliable forms of contraception simultaneously.  Effective contraception 
must be used before beginning study drug therapy, and for 4 months following 
discontinuation of study drug therapy. 
(4) Subjects must be willing and capable of understanding the purpose and risks of the 
study, and must sign a statement of informed consent 
 
 
5.3 
RECIPIENT EXCLUSION CRITERIA FOR STUDY ENTRY 
 
Patients meeting any one of the following criteria will be excluded from entering the study: 
 
(1) Allergy to Thymoglobulin. Thymoglobulin is contraindicated in patients with history of 
allergy or anaphylaxis to rabbit proteins or to any product excipients, or who have active 
acute or chronic infections which contraindicate any additional immunosuppression 
(2) Previous organ transplants 
(3) Patients receiving multiple organs 
(4) Patients with a BMI higher than 35  
(5) Patients with PRA ≥ 25%  
(6) Subjects with a Creatinine ≥ 2.0 mg/dl at time of transplant 
(7) History of a psychological illness or condition which would interfere with the patient’s 
ability to understand the requirements of the study 
(8) HIV-1, chronic Hepatitis B, or chronic Hepatitis C infection, or a history of Chagas 
disease.  
(9) Documented
 or
 strong
 suspicion
 for
 pre-operative
 active
 infection
 that
 has
 not
 yet
 been
 
adequately treated with the recommended course of antimicrobial therapy  
(10) Presence of any chronic myelosuppressive disease or agent that has resulted in either 
chronic leucopenia or chronic thrombocytopenia 
(11) Active peptic ulcer disease and Active GI bleeding  
(12) Patients who have received or require concomitant treatment with other 
investigational drugs within the past 30 days of transplant day (except for those listed 
in section 8.6 “Concomitant treatment”) 
(13) Patients with a history of AL amyloidosis (TTR amyloids) are permitted) 
(14) Current or planned participation in another investigational trial. 
 
5.4   RECIPIENT INCLUSION CRITERIA FOR RANDOMIZATION 
The following criteria will apply to the patients randomized in this study: 
 
1) Women of childbearing potential must have a negative serum pregnancy test prior to 
transplantation.   
 
5.5  RECIPIENT EXCLUSION CRITERIA FOR RANDOMIZATION 
Patients meeting any one of the following criteria will not be randomized: 
1. Patients requiring VAD upon completion of transplantation surgery.  

46 
 
2. White blood cell count ≤ 3000 /mm3, or platelets ≤ 75,000/mm
3, or hemoglobin ≤ 7g/dL 
after completion of heart transplant 
 
5.6 DONOR INCLUSION and EXCLUSION CRITERIA 
Screening of donor hearts will follow Cedars Sinai standard of care protocol and given the 
shortage of suitable donor hearts for cardiac transplantation, COVID-19 positive hearts, HCV 
positive and DCD donor heart might be considered and accepted. 
 
Decisions about whether to accept organs from donors who are HCV NAT positive must be made 
by transplant professionals and potential recipients through a shared decision-making process, 
on a case-by-case basis, when clinically appropriate. 
 
6 
DISEASE EVALUATION (EFFICACY AND SAFETY CRITERIA) 
 
6.1 
PRIMARY EFFICACY PARAMETERS 
 
The following primary efficacy parameters will be measured in all patients: 
 
(A) The incidence between groups of the development of de novo donor specific antibodies 
and ischemia on endomyocardial biopsy at 12 months post-transplant 
 
6.2 
SECONDARY EFFICACY PARAMETERS 
 
To compare between groups: 
 
(A) Secondary Parameters of Efficacy: Changes in percentages of various subsets of 
immune cells at pre transplant, 3 months and 6 months.  Changes in percentages of 
various circulating antibodies  at pre-transplant, 1 month, 3 months, 6 months, and 12 
months after transplant. 
(B) Freedom from cardiac allograft vasculopathy (CAV) (defined as a change ≥0.5mm in 
maximal intimal thickness (MIT) of the coronary arteries by intravascular ultrasound at 
12 months as compared to baseline) 
(C) Change in coronary maximal intimal thickness intimal area, intimal volume, vessel area, 
intimal index, and percent atheroma volume at matched sites by intravascular 
ultrasound at 12 months 
(D) Freedom from acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection within first 12 months after transplantation 
(E) Freedom from acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection episodes per patient within the first 12 months post-transplantation 
(F) Freedom from acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection by the ISHLT biopsy grading scale in the first 12 months post-
transplantation 
(G) Freedom from biopsy proven cellular rejection ≥2R, biopsy proven antibody mediated 
rejection ≥AMR1 and any-treated rejection at 12 months post-transplantation 
(H) Time to first acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection within the first 12 months post-transplantation 

47 
 
(I) Incidence of primary graft dysfunction (PGD) in the first 24 hours post-transplantation 
(J) Patient and graft survival at 12 months post-transplantation 
(K) The number of patients with both fatal and non-fatal infectious complications 
(especially CMV infection) within the first 12 months post-transplantation   
(L) Freedom from the development of circulating antibodies within the first 12 months 
post-transplantation 
(M) 
Average maintenance doses of Mycophenolate Mofetil, tacrolimus, and cumulative dose 
of corticosteroids at 12 months post-transplantation 
(N) Average number of hospital days per patient, both during the transplant period and 
during the post-transplant period at 3 months, 6 months, and 1 year, 
(O) Number of patients requiring hospitalization by 3 months, 6 months, or 12 months post-
transplantation. 
(P) Freedom from the development of circulating antibodies 
 
 
6.3  
DIAGNOSIS AND TREATMENT OF CAV 
  
 
Although most institutions use the cardiac angiogram to diagnose CAV, the limitations and failure 
of
 angiogram
 are
 well
 known.
142 
 
Intravascular
 ultrasound
 (IVUS)
 has
 been
 increasingly
 recognized
 
as a sensitive tool to assess the anatomy of the epicardial coronary arteries.  The procedure is 
performed at the time of the routinely scheduled angiogram and has been demonstrated to be 
safe and have reproducible findings with a correlation coefficient of 0.96 for inter-observer 
agreement.
143  
An
 early
 concern
 about
 the
 safety
 of
 IVUS
 was
 the
 possibility
 that
 catheter
 irritation 
might accelerate the atherosclerotic process.  These concerns have since been addressed.  One 
study evaluating 86 patients undergoing IVUS demonstrated that IVUS was not associated with 
the acceleration of arteriopathy in subsequent procedures.
144  Another study in the Journal of 
the American College of Cardiology confirmed these findings.
145  The reported complications of 
IVUS occur in <1.1% of procedures and include focal coronary spasm, vessel dissection, guidewire 
entrapment, and acute occlusion which may result in myocardial infarction.
146 
 
A study within 6 to 8 weeks after transplant allows the measurement of the recipient’s baseline 
atherosclerosis, which has been demonstrated to be not significantly changed at 2, 4, 6, or 8 
weeks post-transplant.
147  Although several measurements are available to analyze IVUS images, 
maximal intimal thickness (MIT), defined as the greatest distance from the intimal leading edge 
to the external elastic membrane, has been shown to be the most clinically useful measurement 
because of its high reproducibility and its use in predicting outcome.
148  A  maximal  intimal 
thickness of ≥ 0.5 mm is the current widely accepted definition of atherosclerosis.
142  Follow-up 
imaging at 1 year provides important information regarding the development of CAV as the 
change in the first year IVUS results as compared to baseline can predict the course of the 
disease.
143 
 
In
 two
 separate
 studies
 have
 demonstrated
 that
 a
 mean
 intimal
 thickening
 of
 >0.3
 mm
 
as measured by IVUS was associated with significantly worse 4 year overall survival (more death, 
myocardial infarction, and re-transplantation).
10,11  See the following two figures: 
 

48 
 
 
 
Plot showing freedom from development of subsequent angiographic coronary artery disease (angio CAD) 
in the sub-group of patients with normal angiograms at the time of intracoronary ultrasound according to 
intimal thickness of > 0.3 vs less or equal to 0.3 mm. In this analysis, the curve for the patient with intimal 
thickness of less or equal to 0.3 mm has been time-shifted by multiplying each data point by the ratio of a 
mean duration after transplantation in the two groups, i.e., 3.9/2.8 years. (open circle) indicates normal 
angiogram and intimal thickness less or equal to 0.3 mm; (closed circle), normal angiogram and intimal 
thickness > 0.3 mm
10.  
 
 
Relative Risks of Overall Mortality Associated With ICUS, Angiographic Findings, and Duration After 
Transplant in the Total Population10 
 
Two other IVUS studies reported that a >0.5 mm increase in intimal thickening in the first year of 
transplant (termed “rapidly progressive intimal thickening”) had significantly worse 5-year 
morbidity and mortality outcomes.
12,27 
 
A subset of transplant recipients will have a maximal intimal thickness measured at matched sites 
at
 4 
±
 2
 weeks
 and
 again
 at
 12
 months
 ±
 2
 weeks
 post-transplant. 
 Those whose change 
in maximal 
intimal thickness is ≥ 0.3 mm will be considered positive for CAV.  Those who have a change ≥ 0.5 
mm will be classified with “rapidly progressive CAV.”  As mentioned in Section 1.1.2 there is 
currently no treatment for CAV, only the slowing of its progression by the prompt treatment of 
rejection,
 modification
 of
 risk
 factors
 such
 as
 diabetes
 and
 smoking
 cessation,
 and
 the
 appropriate
 
institution and maintenance of the agents that have been demonstrated to slow its progression 
(i.e. mycophenolate mofetil, vitamins C & E, calcium channel blockers, HMG CoA reductase 
inhibitors, everolimus/sirolimus, ganciclovir when appropriate, and aspirin).   
 
 

49 
 
6.4 
DIAGNOSIS AND TREATMENT OF REJECTION 
 
6.4.1 Diagnosis of Rejection 
 
Acute cellular rejection may be diagnosed by an endomyocardial biopsy that has an ISHLT grade 
2R or worse, the presence of hemodynamic compromise requiring treatment (with or without 
biopsy), or endomyocardial biopsy with positive immunohistochemistry stains for antibody-
mediated rejection and significant hemodynamic compromise.  Patients must be treated for 
rejection according to one of the following regimens in Table  1  
(below  and  
in  Appendix  A-2) 
depending on the ISHLT biopsy grade and the presence of hemodynamic compromise. 
 
1) The date of onset of an episode of rejection will be defined according to the date of 
positive biopsy or treatment start date, whichever comes first 
2) All suspected rejections must be biopsied and documented in the case report form 
3) Endomyocardial biopsy to monitor allograft rejection will be performed according to the 
treating institution’s standard of care, which typically occurs at the following time 
points: 
 
Weeks 2 and 4 (Day 14 and 28) 
Months 2, 3, 6 & 12 (Days 56, 90, 180 & 360) 
 
For patients who require high surveillance monitoring, additional biopsies may be 
performed at the following timepoints: 
 
Weeks 1, 3 and 6 (Days 7, 21 and 42) 
Months 4, 5, 8, & 10 (Days 120, 150, 240 & 300) 
 
 
4) The ISHLT Standardized Grading System will be used to assess cellular and antibody-
mediated allograft rejection (see Appendix A-1) 
 
6.4.2 Definition of Hemodynamic Compromise 
 
Hemodynamic compromise is defined as either: 
 
1) Ejection fraction of ≤ 30% or a 0.20 absolute decrease from baseline, and the need for 
inotropic agents 
 
OR 
 
2) Fractional shortening ≤ 20% or a 25% decrease from baseline, and the need for inotropic 
agents 
PLUS 
 
3) Need for inotropic agents due to a Cardiac Index (CI) < 2.0 L/min/m2 or a 25% decrease 
from baseline 
 

50 
 
6.4.3 Treatment of Cellular Rejection 
 
All suspected rejections must be biopsied and documented in the case report form.  Rejection 
episodes will be treated according to the regimens specified in Table 1 (and Appendix A-2). 
 
A follow-up biopsy must be done within 2 weeks after the end of treatment for rejection.  
Resolution of rejection will be defined when no further treatment is required.  If the same or 
additional treatment is required for the current rejection (according to the regimen specified in 
Table 1), the rejection is still considered an ongoing episode.  A follow-up biopsy must be done 
within
 2
 weeks
 after
 the
 end
 of
 this
 treatment.
 
 A
 second
 episode
 of
 rejection
 may
 not
 be
 counted
 
until the first episode has resolved.  
 
Rejection episodes which fail to respond to the specified treatment course may be treated with 
the same course or treated with the next regimen as specified in the Table 1.  For example, a 
Grade 2R rejection episode treated with I.V. corticosteroids which fails to resolve may be treated 
with another course of corticosteroids as specified in Table 1 under 2R with no hemodynamic 
compromise or as specified in Table 1 under 2R with hemodynamic compromise. 
Immunosuppressants used to treat acute allograft rejection will be recorded on the case report 
form. 
 
 Table 1 
Hemodynamic compromise and/or clinical symptoms 
ISHLT biopsy grade 
Absent 
Present 
Grade 1R 
 
Mild 
No treatment 
1-2mg/kg oral corticosteroids for 3-5 days 
followed by 5-10 day taper or immediate 
return to Baseline oral corticosteroids 
and/or 
High dose i.v. corticosteroids with or without 
ATG therapy for 5 days 
Grade 2R 
 
Moderate 
1-4mg/kg oral corticosteroids 
for 3 to 5 days followed by 5 to 
10 day taper or immediate 
return to baseline oral 
corticosteroids 
or 
500mg to 1 g i.v. corticosteroids 
for 3 days followed by 
immediate return to baseline 
oral corticosteroids 
500mg - 1 gm i.v. corticosteroids for 3 days ± 
5 days of ATG therapy followed by taper or 
immediate return to Baseline oral 
corticosteroids  
and/or 
High dose i.v. corticosteroids with or without 
ATG therapy for 5 days 
Grade 3R 
Severe 
High dose i.v. corticosteroids +5 
days of ATG therapy 
High dose i.v. corticosteroids with or without 
ATG therapy for 5 days 
*Hemodynamic compromise is defined in section 6.4.2 
 
6.4.4 Treatment of Antibody Mediated Rejection 
 
Antibody mediated rejection (AMR) as defined by the 2013 ISHLT pathologic grading system (see 
appendix A-1) will be treated according to the standard of care. 
 

51 
 
6.5 DIAGNOSIS OF ISCHEMIA-REPERFUSION INJURY 
 
Ischemia-reperfusion injury (IRI) is hypoxic and anoxic tissue injury from inflammation during 
reperfusion after ischemia. The diagnosis of IRI is made on the basis of findings on the 
endomyocardial biopsy. The endomyocardial biopsy forms the standard of care to monitor the 
allograft for rejection and will be performed according to the treating institution’s standard of 
care, as outlined in Section 6.4.1.  
 
IRI is characterized on biopsy by myocyte necrosis and/or regions of myocyte dropout. Mild 
neutrophil infiltrates or no inflammation is seen, which contrasts with the mononuclear and 
eosinophilic infiltrates seen in acute cellular rejection. Signs of IRI are routinely screened for as 
part of the standard of care and reported as either: 
 
• Ischemic changes: up to 6 weeks post-transplant 
• Ischemic changes: late; related to allograft coronary disease 
 
 
6.6 
PHARMACOKINETIC/PHARMACODYNAMIC PARAMETERS 
 
No pharmacokinetic/pharmacodynamic evaluations will be performed.   
 
7. 
STUDY DESIGN 
 
7.1 
DESIGN 
 
This is a randomized, controlled, multi-center study to evaluate the efficacy of Thymoglobulin 
induction therapy in combination with Mycophenolate Mofetil, tacrolimus, and steroids in the 
prevention of CAV.  Approximately half of the patients will be randomized to receive a total of 5 
doses of Thymoglobulin during the study.   
 
For the subjects randomized to receive the study drug, the first two doses of Thymoglobulin will 
be administered at 1.5 mg/kg via intravenous infusion over 8 hours immediately upon arrival to 
the ICU post-operation [day 1 (POD#0) and day 2 (POD #1}.  Subsequent doses of 1.5 mg/kg will 
be administered on days 3, 4, and 5 via IV infusion over 4-8 hours. 
 
The subjects will be consented onto the study while on the UNOS waitlist.  The screening 
assessments may be drawn after consent is obtained and any time prior to transplant.    Patients 
who qualify for the study will be administered the following treatment regimens: 

 
52 
 
Table of Study Medication Administration 
Medication 
POD#0/ Day 1 
POD#1/ Day 
2  
POD#2/ 
Day3 
POD#3/ 
Day 4 
POD#4/ 
Day 5 
POD  
#30 
POD 
#90 
POD 
#180 
POD 
#365 
Thymoglobulin/ 
No Induction or 
Control* 
1.5 mg/ kg over 8 
hrs within 24 hours 
post Tx 
1.5 mg/ kg 
over 8 hrs 
1.5 mg/ 
kg over 
4-8 hrs 
1.5 mg/ kg 
over 4-8 
hrs 
1.5 mg/ 
kg over 
4-8 hrs 
Stopped after POD#4 
Mycophenolate 
Mofetil/MMF 
1.5 g bid 
Continue & adjust dose as indicated by side effects 
Tacrolimus 
 
1 to 4 mg bid 
Titrate to 
trough 
Titrate to 
trough 
Titrate to 
trough 
Trough 
10-15 
ng/mL 
Trough 8-
12 ng/mL 
Trough 5-
10 ng/mL 
Trough lowered to 5-
10 ng/mL if no 
rejection 
Corticosteroids 
125 mg IV 
Solumedrol q12 hrs 
x 3 doses 
Prednisone 
1 mg/kg/ 
day po 
divided into 
bid doses  
Taper by 
10 mg qd 
Taper to 
10 mg po 
bid 
 
Taper by 2 
mg q 1-2 
wks to 5 
mg po bid 
Taper by 2 
mg q 2 wks 
until 5 mg 
po qd 
Discontinue 
steroids 
 
HMG CoA Reductase 
Inhibitor** 
First dose initiated 
within 2 weeks of 
surgery 
Dose 
titrated up 
as tolerated 
Continued as tolerated 
Vitamin C** 
500 mg qd 
Continue as tolerated 
Vitamin E** 
400 IU qd 
Continue as tolerated 
Aspirin** 
81 mg qd 
Continue as tolerated 
* First dose within 24 hours post transplant 
** Dosages will be prescribed according to institution’s standards 

53 
 
Patients will be seen at Screening, Post op days Day 1-5, Day 7, 14, 21, 28, 42, 56, 90, 120, 
150, 180, 240, 300 and 365 (refer to Schedule of Activities for visit windows).  Laboratory 
parameters, diagnostic tests and clinical evaluations will be obtained as needed. 
 
All
 patients,
 including
 premature
 withdrawals,
 will
 be
 followed
 for
 a
 total
 of
 12
 months
 post
 study
 
start date for death, re-transplantation, acute cellular, antibody-mediated,  hemodynamic 
compromise, and any-treated rejection episodes regardless of whether they are receiving study 
medication. 
 
7.2 
STUDY PROCEDURES 
 
7.2.1 Screening/Baseline 
 
Prospective patients will be identified by the principal investigator and written informed consent 
must
 be
 obtained
 before
 the
 performance
 of
 any
 screening
 procedures.
  
Patients
 will
 be
 screened
 
within
 48
 hours
 of
 entry
 into
 the
 study.
 
 The
 following
 standard
 of
 care 
data will
 be
 extracted from 
the medical record: 
 
(A) Medical history 
(B) Body Weight 
(C) Laboratory tests: 
(1) CBC with differential, platelet count 
(2) Chemistry panel 
(3) Pregnancy test (serum) must be done prior to transplantation.  Urine test is allowed 
in addition to serum test in patients where serum results are delayed. 
(4) Panel of reactive antibodies (PRA) 
(5) HLA (DR) mismatch 
(6) CMV and EBV serologic status of the recipient and donor 
Blood and platelet loss will be monitored during the post-transplantation 
period and the following will be recorded: 
(a)  Number of PRBC and platelet transfusions  
(b) Amount of blood lost during first 48 hours post-transplant 
 
7.2.3  Clinical Evaluations 
 
The following standard of care data will be extracted from the medical record.  Please see the 
Schedule of Assessments for a complete outline of study visits and visit specific procedures. 
 
(A) Clinical Assessment—clinical evaluation of patients’ signs or symptoms of rejection, 
adverse events, infections, and malignancies (throughout entire inpatient stay post-
transplantation and at every clinic visit) 
(B) Endomyocardial biopsy  
Any patient who prematurely discontinues from the study will continue to have 
endomyocardial biopsies as is currently the standard for routine post-transplant care 
(C) Vital signs, including blood pressure, pulse, temperature (every visit) and body 
weight (all visits) 
(D) Laboratory evaluations - Record results closest to study visit date as outlined in the 
Schedule of Assessments: 

54 
 
a. CBC with differential, platelets, electrolytes, creatinine, BUN, lipid panel, and 
TACROLIMUS if completed per standard of care or as needed.  
b.  Immune profile and circulating antibodies  
(E) Concomitant medication:  All concomitant medications, particularly 
immunosuppressive therapy and prophylactic therapy should be reviewed with the 
patient at every visit and recorded on the CRF (all visits) 
(F) Adverse events (all visits).  See section 11.4.2 for definition of an adverse event 
(G) Intravascular ultrasound 
(H) Right Heart Catheterization 
(I) Echocardiogram 
 
7.3 RANDOMIZATION PROCEDURE AND ASSIGNMENT TO TREATMENT GROUP 
 
 
Patients consented to the study and meeting study entry and randomization inclusion and 
exclusion criteria will be randomized in a 1:1 ratio to either Thymoglobulin® or No Induction or 
Control during the immediate post transplantation period.   
 
Randomization will be implemented through RedCap by Cedars-Sinai. Patients who are 
randomized will receive a unique drug randomization number that corresponds to his/her unique 
study identification number. 
 
8. 
STUDY MEDICATION 
 
8.1 
DRUG NAME, FORMULATION, & STORAGE 
 
HOW SUPPLIED 
 
Thymoglobulin is available as sterile, lyophilized powder to be reconstituted with Sterile Water 
for Injection, USP (SWFI).  Each package contains a 10 mL vial freeze-dried Thymoglobulin (25 mg) 
NDC# 58468-0080-1. 
 
Reconstituted Thymoglobulin is physically and chemically stable for up to 24 hours at room 
temperature; however, room temperature storage is not recommended.  As Thymoglobulin 
contains no preservatives, reconstituted product should be used immediately. 
 
8.2 
PACKAGING AND LABELING 
 
The Thymoglobulin
® will be supplied by Genzyme and packaged in 25 mg vials.  Thymoglobulin® 
will be shipped to the Cedars-Sinai Investigational Pharmacy from Sanofi. The study drug will be 
labeled per CFR Title 21 312.6: “Caution: New Drug—Limited by Federal (or United States) law to 
investigational use,” and will contain the protocol name/# on the label. 
 
As per the manufacturer’s guidelines the study drug will be stored between 2 to 8 degrees C (36 
to 46 degrees F) away from light.  Study drug will not be frozen or used after the expiration date 
indicated
 on
 the
 label.
 
 Reconstituted
 vials
 of
 Thymoglobulin
® 
will
 be
 used
 within
 4
 hours.
 
 Infusion
 
solutions
 of
 Thymoglobulin
® 
must
 be
 used
 immediately.
 
 Any
 unused
 drug
 remaining
 after
 infusion
 
must be discarded. 

55 
 
 
 
8.3 
STUDY TREATMENT 
 
8.3.1 Rationale for Dose Selection 
 
Neither a dose response nor a time course study has been performed on cardiac transplant 
patients with Thymoglobulin®.  Thus, we have selected a dosing regimen of 1.5 mg/kg/day 
intravenously for 5 days based on some scientific data as well as current popular methodology.  
Thymoglobulin
® has been administered to over 40,000 renal and cardiac transplant recipients in 
Europe since 1985 and the most popular dose currently used for induction purposes is 1.5 
mg/kg/day for 4 days149.   As mentioned above in Section 1.2.2 no correlation has been seen 
between treatment outcomes and either active or total Thymoglobulin® concentrations in renal 
transplant patients who were dosed at 1.5 mg/kg/day for 6-14 days119.  This implies that there is 
probably no benefit to more than 6 days of therapy.   A number of studies measuring various T-
cell responses to patients receiving Thymoglobulin
®, have consistently demonstrated significantly 
lower absolute T-cell counts as well as significantly lower CD3, CD2, CD4, CD8, and CD19 by day 5 
of administration120,150-153, suggesting that the optimal dosing time course is probably 5 or 6 days.   
 
 
 
The effect of thymoglobulin (Thy, ),  anti-thymocyte globulin (ATG, ) or Minnesota anti-lymphoblast 
globulin (ALG, ) on CD3 T-cells over 10 days  of  flow  cytometric  analysis.  Data  are  expressed  as  the 
percentage of cells remaining relative to the preinduction level, represented on day 1 at 100%. Note that 
Thy
 was
 administered
 for
 only
 the
 first
 5
 days 
while
 ATG
 and
 ALG
 were
 administered
 over
 the
 full
 10
 days154 
 

56 
 
 
 
The effect of thymoglobulin (Thy, ), anti-thymocyte globulin (ATG, ) or Minnesota anti-lymphoblast 
globulin (ALG, ) on CD4 T-cells over 10 days  of  flow  cytometric  analysis.  Data  are  expressed  as  the 
percentage of cells remaining relative to the preinduction level, represented on day 1 at 100%. Note that 
Thy was administered for only the first 5 days while ATG and ALG were administered over the full 10 days. 
 
 
 
The effect of thymoglobulin (Thy, ),  anti-thymocyte globulin (ATG, ) or Minnesota anti-lymphoblast 
globulin (ALG, ) on CD8 T-cells over 10 days  of  flow  cytometric  analysis.  Data  are  expressed  as  the 
percentage of cells remaining relative to the preinduction level, represented on day 1 at 100%. Note that 
Thy
 was
 administered
 for
 only
 the
 first
 5
 days 
while
 ATG
 and 
ALG
 were
 administered
 over
 the
 full
 10
 days154 
 

57 
 
 
Refer to above
 paragraph:  Depletion of CD2, CD3, CD4, CD8,
 and CD19 positive cells was rapid, with counts 
falling to less than 30 cells/pL, and was sustained for several weeks after Thymoglobulin
®155. 
 
The Thymoglobulin
® package insert recommends a dose of 1.5 mg/kg/day for 7-14 days in the 
treatment of acute renal graft rejection.  Despite this recommendation, there are numerous 
studies in acute renal rejection in which Thymoglobulin
® was administered either at less than 1.5 
mg/kg/day and/or less than 7 days of therapy without a difference in clinical 
outcome.
106,121,123,125,154,156  However, one must keep in mind that the main purpose of 
Thymoglobulin
® induction therapy in renal transplantation is to be able delay the initiation of the 
nephrotoxic calcineurin inhibitors while the renal graft is recovering and at the same time obtain 
effective “nephro-safe” immunosuppression.  In most of the renal transplantation protocols 
Thymoglobulin is administered for an unspecified, open ended number of days until the renal 
graft has recovered from either surgery or rejection, which is usually more than 7 days.  Thus, the 
recommendation of 7-14 days seems justified for use in renal transplantation.  This study will 
incorporate Thymoglobulin
® in addition to the non-delayed introduction of triple 
immunosuppressive therapy immediately post-transplant with the objectives of reducing the 
incidence of acute rejection and CAV without increasing the risk of infection.  Thus, for the 
purposes
 of
 this
 study,
 the
 recommended
 7-14
 days
 of
 Thymoglobulin® therapy
 probably
 does
 not
 
apply. 
 
8.3.2 Dosage Regimen and Dose Adjustment 
 

58 
 
A total of 5 doses of Thymoglobulin® will be administered per recipient at 1.5 mg/kg/dose.  The 
appropriate dose should first be reconstituted. Transfer the contents of the calculated number of 
Thymoglobulin
® 
vials 
into the bag of 
infusion solution (saline or dextrose). Recommended volume: 
per one vial of Thymoglobulin use 50 mL of infusion solution (total volume usually between 50 to 
500 mL). Mix the solution by inverting  
the bag gently only once  or twice.  The first two doses 
should be administered over an 8 hour continuous intravenous  central infusion.  The 
reconstituted/diluted medication may be stored for up to 4 hours at 2 to 8 degrees C before 
administration.  Each patient should be premedicated with 650 mg of acetaminophen and 25 or 
50 mg of diphenhydramine (depends on the patient’s weight) and Methylprednisolone 125 mg IV 
approximately 60 minutes before each infusion of Thymoglobulin
®. The dose of diphenhydramine 
should be 25 mg for any patient less than 60 kg. If the patient is 60 kg or greater, a dose of 50mg 
is preferred. 
  
 
The first dose of Thymoglobulin
® will be administered at a dosage of 1.5 mg/kg IV upon arriving 
to the Intensive Care Unit after transplantation (called POD Day 0/Day #1) along with the first 
dose of IV methylprednisolone.  The first dose must be administered within 24 hours after 
transplantation in order for the patient to continue in the study.  The subsequent 4 doses also at 
1.5 mg/kg will be administered on days 2, 3, 4, and 5 every 24 hours from the start of the first 
dose.   
 
There has been one prospective, randomized, clinical trial of intraoperative versus postoperative 
initiation of Thymoglobulin
® therapy in renal transplant recipients where intraoperative 
administration was associated with significantly less initial delayed graft function
106. Although 
there may be a hypothetical advantage to administering Thymoglobulin® intraoperatively, there 
is concern that if side effects (such as anaphylaxis, see Section 10) were to occur intraoperatively 
or during transport, the limitations of the intra-operative setting would preclude the prompt and 
highly
 skilled
 intervention
 that 
is
 needed
 to
 treat
 the
 adverse
 event(s).
 Therefore, Thymoglobulin
® 
induction will be initiated immediately upon arrival to the Intensive Care Unit in this study.           
    
8.3.3 Dose Modification 
 
Any toxicity associated with the Thymoglobulin® infusion that is also considered by the physician 
to be a major threat to the patient mandates discontinuation of the antibody infusion and/or 
discontinuation of the course of therapy.   
 
The two main concerns are the development of leucopenia (WBC <2,000 cells/mm3) and 
thrombocytopenia (platelet count <50,000 cells/mm
3).  Doses will be modified as outlined in the 
tables below (obtained from Thymoglobulin
® package insert): 
 
Absolute WBC count 
Thymoglobulin® dose mg/kg/day 
>3,000 cells/mm3 
1.5 mg/kg/day 
2,000-3,000 cells/mm3 
0.75 mg/kg/day 
<2,000 cells/mm3 
Hold until WBC count >2,000  
 
Platelet count 
Thymoglobulin® dose mg/kg/day 
>75,000 cells/mm3     
1.5 mg/kg/day 

59 
 
50-75,000 cells/mm3 
0.75 mg/kg/day 
<50,000 cells/mm3 
 Hold until platelet count >50,000 
 
Holding Thymoglobulin treatment will be considered only if persistent and severe 
thrombocytopenia (< 50,000 cells/mm3) occurs or leukopenia (< 2,000 cells/mm3) develops. 
 
White blood cell and platelet counts will be monitored during and after Thymoglobulin therapy.  
Once the blood counts have recovered the recipient will resume therapy. 
 
Chills and fever may occur but generally can be controlled by the administration of 
diphenhydramine hydrochloride (25 to 50 mg IV every 6 hours as needed) and acetaminophen 
(325 to 650 mg PO every 4 hours as needed).  For severe fever and chills, meperidine (25 mg IV 
every 4 to 6 hours as needed) may also be required.  When any of the above non-life-threatening 
toxicities occur, the infusion should be interrupted until symptoms subside and then restarted at 
a slower rate. 
 
Respiratory  distress,  pain  
the  
flank  or  back,  or  hypotension  may  be  signs  of  anaphylaxis.  
  The 
infusion should be discontinued and not resumed, and the course of treatment should be 
terminated.  The above reactions must be entered on the case report form, and the patient will 
continue to be followed for safety for the duration of the study. 
 
Other minor toxicities can usually be managed with symptomatic treatment and slowing of the 
infusion. 
 
8.3.4 Route of Administration 
 
Thymoglobulin
® 
should
 be
 used
 under
 strict
 medical
 supervision
 in
 a
 hospital
 setting,
 and
 patients
 
should be carefully monitored during the infusion. The first dose should be infused within 24 
hours post-transplant into a high-flow vein. Follow the manufacturer’s instructions for the 
infusion administration set. Infuse through a 0.22 micrometer filter into a high-flow vein. 
 
Although
 ATG
 was
 administered
 intramuscularly
 in
 earlier
 studies,
 currently
 the
 intravenous
 route
 
is universally used, as according to the guidelines provided by Genzyme (see package insert).  
Intravenous Thymoglobulin
® has been demonstrated to be safe for use in the ambulatory 
setting.
122 
 
The drug will be administered via constant intravenous infusion within 24 hours (first dose) or 4 
hours (subsequent doses).  A central line will be used for administration of study drug.
122  
 
8.3.5 Treatment Time Course 
 
All patients randomized to study drug will receive 5 doses of Thymoglobulin
® unless an adverse 
event warrants dose reduction or temporary discontinuation.  The necessity of any dose 
adjustment should be discussed with the medical data monitor.  The first dose will be 
administered upon arrival to the Intensive Care Unit. 
 
8.4 
DISPENSING AND ACCOUNTABILITY OF STUDY MEDICATION SUPPLIES 
 

60 
 
Although Thymoglobulin® will be administered as standard of care, a drug administration log will 
be kept by the Study Coordinator and will identify the patient and the amount of medication 
administered to the patient.   
 
8.5 
CONCOMITANT TREATMENT 
 
Concomitant medications, including nitric oxide for pulmonary hypertension, will be permitted as 
necessary.   
 
8.5.1 Concomitant Immunosuppressive Therapy 
 
Standard supportive immunosuppressive therapy will be prescribed to all patients randomized in 
this study.  The standard concomitant therapy is described below (see Table of Study Medication 
Administration in Section 7.1): 
 
1)  Mycophenolate mofetil:  3.0 grams divided bid begun post-transplant, either IV or po as 
tolerated by patient.  Initial dose must be given within 24 hours post-transplant.  Dosing will be 
titrated based on recipient’s body size and any adverse side effects  
2)  TACROLIMUS:  Doses of 1-4 mg bid either IV or po will be administered on post-operative day 
#0 to achieve a target trough level of 10-15 ng/mL by post-operative day #5.  Target trough 
levels are 10-15 ng/mL for post-operative days #1-90 and 5-10 ng/mL thereafter. In patients 
with creatinine >0.2 mg/dL, delayed initiation of tacrolimus to post-operative day 3 will be done 
to allow recovery of renal function.
157 
3)  Corticosteroids:  125 mg IV methylprednisolone immediately post-operatively x 3 doses 
q12hrs, then switching to oral prednisone at 1.0 mg/kg/day po divided into bid doses that are 
rounded off to the next higher 5 mg increment.  For example, a 76 kg person should be dosed at 
38 mg po bid, which rounded off to the next 5 mg increment would be 40 mg po bid.  
(Equivalent dosing via an alternative route may be used if po is not tolerated or 
contraindicated).  Prednisone will be tapered by 10 mg qd until the dose of 10 mg po bid is 
reached.   
 
Patient will then be tapered by the following schedule (as per our institutional protocol) as 
tolerated by lack of evidence for rejection:   
 
Continue 10 mg po bid until post-operative day #30 
Taper by 2 mg every 1-2 weeks to achieve 5 mg po bid by day #90 
Taper by 2 mg every 2 weeks to achieve 5 mg po qd by day #180 
Steroids then discontinued  
 
8.5.2 Other Concomitant Therapy 
 
1)  HMG CoA Reductase Inhibition:  All patients will receive lipid lowering/anti-inflammatory 
therapy, either pravastatin or simvastatin, as per our institutional protocol.  Patients will receive 
one dose daily initiated at the time of oral administration or within 1-2 weeks of surgery.  
Dosages will be titrated up to a maximum dose 40 mg as tolerated by the patient.  If total 
cholesterol still exceeds 200 mg/dL at 40 mg of pravastatin, a stronger agent such as 
atorvastatin may be substituted and/or ezetimibe added. 

61 
 
2)  Calcium channel blockers or ACE inhibitors:  All patients who become hypertensive post-
transplant will receive a calcium channel blocker and/or an ACE inhibitor (amlodipine or 
nifedipine extended release), the dosage of which will be titrated according to the patient’s 
blood pressure. 
3)  Antioxidants:  All patients will be maintained on 400 IU of vitamin E and 500 mg vitamin C 
daily as according to our institutional protocol.  
4)  Aspirin:  All patients will receive at least 81 mg of aspirin daily as according to our 
institutional protocol.   
 
8.5.3 Mycophenolate Mofetil Dose Adjustment for Adverse Events 
 
Mycophenolate mofetil is known to cause leucopenia.  In the event of leucopenia (defined as a 
WBC count <2,000 cells/mm3) or another known adverse side effect of mycophenolate mofetil, 
mycophenolate mofetil doses may be reduced or interrupted.  Patients will not be withdrawn 
from  
the  study  because  of  extended  mycophenolate  mofetil dose reductions or interruptions.  
However, the investigator will attempt at his/her discretion to return the patient to the full 
maintenance dose upon resolution of the adverse event.  After a dose interruption, every effort 
will be made within 14 days, unless medically warranted, to re-instate mycophenolate mofetil in 
increments until the recommended mycophenolate mofetil dose is achieved.  If this is not 
possible,
 then
 Mycophenolate
 mofetil 
will
 be
 increased
 in
 stepwise
 increments
 of
 250
 mg until an 
optimal tolerable dose is found.   
 
8.5.4 Prophylactic Therapy 
 
All patients will also be prescribed prophylactic treatment as follows: 
 
1)  Antimicrobial prophylaxis 
a)  Bacterial and Pneumocystis carinii pneumonia prophylaxis:  Bactrim DS 2-4 tablets per 
week for 1 year.  For patients allergic to sulfa, dapsone at 50-100 mg orally, 
administered daily, or aerosolized pentamidine, 300 mg per month will be substituted 
for Bactrim DS. 
b)  Fungal prophylaxes:  Nystatin 5000 U qid or Mycelex troches for a minimum of 6 months 
2)  CMV prophylaxis: (CMV disease defined in Appendix B) 
All recipients who receive an organ from a CMV positive donor will be treated with 2 weeks of IV 
ganciclovir:  5-10 mg/kg IV ganciclovir for 14 days followed by oral ganciclovir (Cytovene) for a 
total of 6 months of therapy. 
Recipients that are CMV positive, who receive a CMV negative donor heart will receive 
ganciclovir/valganciclovir for 3 months.  
 
All concomitant medications will be reported in the case report form including start and stop 
dates.   
 
8.5.5 Drug Interactions 
  
 
The following medications have been administered in clinical trials with Thymoglobulin
® without 
incremental increase in adverse reactions:  daclizumab, cyclosporine, mycophenolate mofetil, 
ganciclovir, azathioprine, corticosteroids, sirolimus, and tacrolimus.    
 

62 
 
9. 
PREMATURE WITHDRAWAL 
  
 
Patients who prematurely discontinue treatment with the study therapy regimen will remain in 
the study and follow the visit schedule. Thymoglobulin® may be prematurely 
discontinued/terminated
 for
 any
 patient
 for
 life
 threatening
 reactions.
 The
 study
 therapy
 regimen
 
may also be prematurely discontinued for any participant if the investigator believes that the 
treatment is no longer in the best interest of the subject, if the subject is judged non-compliant, 
or due to safety concerns.  
 
 
9.1 
FOLLOW-UP 
 
All patients (including those who prematurely discontinued) will be followed for a total of 12 
months for the following data: death and re-transplantation, development of malignancies, and 
acute rejection episodes. 
 
If
 the
 reason
 for
 removal
 of
 a
 patient
 from
 the
 study
 is
 an
 adverse
 event
 or
 an
 abnormal
 laboratory
 
test result, the principal specific event or test will also be recorded on the case report form. 
 
10. 
WARNINGS AND PRECAUTIONS 
 
Thymoglobulin
® adverse events are generally manageable or reversible (see table below). 
Thymoglobulin
® should only be used by physicians experienced in immunosuppressive therapy.  
Medical surveillance is required during Thymoglobulin® infusion. Serious immune-mediated 
reactions have been reported with the use of Thymoglobulin
® and consist of anaphylaxis and 
cytokine release syndrome (CRS). Fatal anaphylaxis has been reported.  If anaphylaxis occurs, 
the infusion should be terminated immediately.  Medical personnel should be available to treat 
patients who experience anaphylaxis.  Emergency treatment such as 0.3 mL to 0.5 mL aqueous 
epinephrine (1:1000 dilution) subcutaneously and other resuscitative measures including 
oxygen, intravenous fluids, antihistamines, corticosteroids, pressor amines, and airway 
management, as clinically indicated, should be provided. Thymoglobulin® or other rabbit 
immunoglobulins should not be administered again for such patients. Severe, acute infusion-
associated reactions are consistent with CRS which is attributed to the release of cytokines by 
activated monocytes and lymphocytes. Severe acute CRS can cause serious cardiorespiratory 
events and/or death. Thrombocytopenia or neutropenia may result from cross-reactive 
antibodies and is reversible following dose adjustments (see Section 8.3.2.1).  During 
Thymoglobulin
® therapy, monitoring the lymphocyte count (i.e., total lymphocyte and/or T-cell 
subset) may help assess the degree of T-cell depletion.  For safety, WBC and platelet counts 
should also be monitored. 
 
Thymoglobulin
® infusion may produce fever and chills. To minimize these, the first two doses 
should be infused over a minimum of 8 hours and the rest of the doses (3-5) should be infused 4-
8 hours into a high flow vein. Also, premedication with corticosteroids (methyl prednisone), 
acetaminophen, and/or an antihistamine and/or slowing the infusion rate may reduce reaction 
incidence and intensity.  Adverse reactions more frequently reported following Thymoglobulin® 
infusion than following Atgam
® in a phase III randomized controlled trial: 
 
 

63 
 
Side effects 
Thymoglobulin® (n=82) # pts (%) 
Chills 
47 (57.3) 
Leukopenia  
47 (57.3) 
Headache 
33 (40.2) 
Abdominal pain 
31 (37.8) 
Hypertension 
30 (36.6) 
Nausea 
30 (36.6) 
Dyspnea 
23 (28.0) 
Hyperkalemia 
22 (26.8) 
Myalgia 
16 (19.5) 
Insomnia 
16 (19.5) 
Hypotension 
13 (15.9) 
Rash 
11 (13.4) 
Sweating 
11 (13.4) 
Malaise 
11 (13.4) 
Acne 
10 (12.2) 
Overdose 
5 (6.1) 
 
 
Infections, reactivation of infection, and sepsis have been reported after Thymoglobulin® 
administration in combination with other immunosuppressive agents. Appropriate antiviral, 
antibacterial, antiprotozoal, and/or antifungal prophylaxis is recommended. Infections over a 3-
month
 follow-up
 in
 the
 Phase
 III
 trial109 
are
 summarized
 in
 the
 table
 below,
 but
 the
 lack
 of
 a
 proper
 
control group makes it difficult to credit Thymoglobulin
® as the sole cause of these infections. 
 
Infections (Thymoglobulin® n=82) 
 
BODY SYSTEM 
Preferred Term 
% of 
patients 
# of 
patients 
Total 
Reports 
BODY AS A WHOLE 
36.6 
30 
36 
  Infection 
    Other 
    CMV 
  Sepsis 
   
30.5 
17.1 
13.4 
12.2 
 
25 
14 
11 
10 
 
26 
15 
11 
0 
DIGESTIVE 
6.1 
5 
5 
  Gastrointestinal 
  Moniliasis 
  Oral monoliasis 
  Gastritis 
 
4.9 
3.7 
1.2 
 
4 
3 
1 
 
4 
0 
1 
RESPIRATORY 
0.0 
0 
0 
  Pneumonia 
0.0 
0 
0 
SKIN 
4.9 
4 
4 

64 
 
  Herpes simplex 
4.9 
4 
4 
UROGENITAL 
18.3 
15 
15 
  Urinary tract infection 
18.3 
15 
15 
  Vaginitis 
0.0 
0 
0 
NOT SPECIFIED   
0.0 
0 
0 
  
 
The safety of immunization with attenuated live vaccines following Thymoglobulin
® therapy has 
not been studied; therefore, immunization with attenuated vaccines is not recommended for 
patients who have recently received Thymoglobulin
®. 
 
Use of immunosuppressive agents, including Thymoglobulin®, may increase the incidence of 
malignancy, including lymphoma or lymphoproliferative disorders (which may be virally 
mediated). These events have been associated with fatal outcome. The carcinogenic and 
mutagenic potential of Thymoglobulin® and its potential to impair fertility have not been studied. 
 
Reactions at the infusion site can occur and may include pain, swelling, and erythema. 
 
Thymoglobulin
® can stimulate the production of antibodies which crossreact with rabbit immune 
globulins.  Thymoglobulin
® has not been shown to interfere with any routine clinical laboratory 
tests which do not use immunoglobulins.  Thymoglobulin
® may interfere with rabbit antibody-
based immunoassays and with cross-match or panel-reactive antibody cytotoxicity assays.   
   
The safety of Thymoglobulin
® in pregnant women and nursing mothers has not yet been tested.  
It is not known whether this drug is excreted in human milk. Because many drugs are excreted in 
human  milk,  caution  should  be  exercised  when  Thymoglobulin® is administered to a nursing 
woman.  Women of childbearing potential must have a negative serum pregnancy test prior to 
transplantation.  Urine test is allowed in addition to serum test in patients where serum results 
are delayed.  Effective contraception must be used by women of childbearing potential before 
beginning Thymoglobulin® therapy and during therapy even where there has been a history 
infertility (unless due to hysterectomy) and up to 4 months following discontinuation of study 
medication.  Two reliable forms of contraception must be used simultaneously unless abstinence 
is the chosen method.   
  
 
If pregnancy occurs during the treatment or within the 12 months following discontinuation of 
study medication, the physician and patient should discuss the desirability of continuing the 
pregnancy.  All patients who become pregnant while receiving study medication or for 4 months 
following discontinuation of the study medication must be followed until termination or 
completion of the pregnancy.  The investigator must complete an SAE report in addition to 
completing the Pregnancy Outcome Page in the case report form. 
 
10.1 OVERDOSE OR EXAGGERATED RESPONSE 
 
Thymoglobulin
® overdosage may result in leukopenia (including lymphopenia and neutropenia) 
or thrombocytopenia, which can be managed with dose reduction.  The five doses of 
Thymoglobulin
® 
will
 be
 administered
 over
 five
 days
 in
 all
 patients
 while
 they
 are
 still
 in
 the
 hospital
 
as the average post-cardiac transplant patient at our facility stays a minimum of 6 days.  Thus, 

65 
 
accidental overdosing could occur in the setting of a nursing error, where the wrong dose is 
calculated and administered.  There is no antidote for Thymoglobulin
® overdose.  Treatment 
would consist of prompt cessation of the drug, close clinical observation, and monitoring of the 
blood
 count
 for
 profound
 thrombocytopenia,
 neutropenia,
 anemia,
 or
 lymphocytopenia
 and
 their
 
expected normalization upon discontinuation of therapy.  Platelet and blood transfusions may be 
necessary should the hemoglobin fall below 8 g/dL or the platelet count fall below 50,000 
cells/mm
3 and the patient is actively bleeding.  However, the benefit of a transfusion will need to 
be weighed against the risk of immune activation, and the decision to transfuse should be made 
by the transplant attending on a case-by-case basis.  Once the blood counts have recovered, the 
recipient will resume therapy based on the dosing table provided in Section 8.3.2.1. 
 
 
 
11. 
SAFETY PARAMETERS 
 
During the study, safety assessments will include: 
 
1) 
Clinical assessments, including weight and vital signs 
2) 
Incidence of adverse events, malignancies, infectious complications, and 
premature withdrawal due to adverse events 
3) 
Collection of selected concomitant medications and immunosuppressive 
therapies 
4) 
Laboratory assessments 
5) 
Graft loss and death 
6) 
Vital signs (temperature, respiration, blood pressure, and heart rate) will be 
noted preinfusion, immediately post-infusion, and 15 minutes post-infusion of 
Thymoglobulin
® and reported on the appropriate case report form. 
 
11.1 MEDICAL HISTORY AND CLINICAL EVALUATIONS 
 
A complete medical history will be performed during screening.  This assessment will include age, 
sex, race, weight, the etiology of the cardiac disease, information on ABO matching of recipient 
and donor, results of panel reactive antibody (PRA)—both the highest and most recent 
assessment, HLA (DR) mismatch, CMV and EBV serologic status of the recipient and donor, 
Hepatitis B and C, cold ischemic time, and age of donor organ.  Patients will be evaluated during 
the study for signs and symptoms including adverse events, infectious complications, 
malignancies, and acute rejection.  A clinical assessment will be performed as noted in the 
Schedule of Assessments. 
 
11.2 LABORATORY PARAMETERS 
 
The following routine laboratory tests will be performed at baseline and/or as noted in the 
Schedule of Assessments: 
 
Hematology:      
 
hemoglobin, hematocrit, RBC count, WBC with 
differential, platelet count 
Serum Chemistries:     
BUN, creatinine, SGOT, SGPT, GGT, calcium, 

66 
 
phosphorus, bilirubin (total and direct), total protein, 
albumin, glucose, alkaline phosphatase, LDH, uric acid, 
CO2, potassium, magnesium, sodium, and chloride 
Fasting lipid profile:   
Total cholesterol, LDL, HDL, HDL/LDL ratio, 
triglycerides 
Pregnancy test (serum):  
Serum pregnancy test must be performed 
prior to transplantation.  Results must be obtained 
and documented prior to  
transplantation.  Urine test is allowed in addition to  
serum test in patients where serum results are 
delayed.  Follow-up tests should be performed in 
the event of secondary amenorrhea. 
 
All laboratory analyses will be performed at the study institution’s designated laboratory.   
 
11.2.1 Procedures in the Event of Significant Abnormal Laboratory Values 
 
Abnormal laboratory values considered clinically significant by the Investigator other than those 
associated with the patient’s disease state, must be repeated as soon as possible and followed 
until they return to normal or an explanation is found.  If a clear explanation is established, it 
should be recorded on the case report form. 
 
11.3  INFECTIOUS COMPLICATIONS 
 
See Appendix B for classification criteria for infectious complications.  Infectious complications 
during
 the
 study
 will
 be
 documented
 on
 a
 specific
 IC 
case
 report
 form
 and
 not
 on
 an
 Adverse
 Event
 
case report form.  Information captured on the  case report form will include: date of onset, 
pathogen identified, and resolution, and action taken. 
 
12 
ADVERSE EVENTS 
  
  
 
12.1 Baseline Medical Condition 
 
It is not necessary to complete an Adverse Event (AE) page in the CRF for adverse medical 
conditions present during the screening period which do not worsen in either severity or 
frequency during the study.  These conditions should be adequately documented in the patient’s 
medical records, the CRF (i.e. previous medical conditions section) and any other medical 
documents. 
 
Adverse medical conditions present during the screening period which become worse following 
exposure to study drugs should be reported as adverse events.  They can also become Serious 
Adverse Events. 
 
Any
 intervention
 performed
 during
 the
 study
 that
 corresponds
 to
 a
 condition
 during
 the
 screening
 
period, should be recorded on the “Additional Observation” or “Concomitant Medication” pages 
of the CRF as appropriate. 
 

67 
 
12.2 DEFINITION OF ADVERSE EVENTS 
  
  
 
12.2.1 Adverse Event (AE) 
 
An adverse event (AE) is defined as any untoward or unfavorable medical occurrence associated 
with the use of a drug in humans, whether or not considered drug related (21 CFR 312.32(a)). An 
adverse event may include any unfavorable or unintended sign (including an abnormal 
laboratory finding), symptom, or disease temporally associated with the use of an 
investigational product (ICH E6, 1.2). 
 
12.2.2 Suspected Adverse Reaction (SAR) 
 
A suspected adverse reaction (SAR) is any adverse event for which there is a reasonable 
possibility that the investigational drug caused the adverse event. For the purposes of safety 
reporting, ‘reasonable possibility’ means there is evidence to suggest a causal relationship 
between the drug and the adverse event. A suspected adverse reaction implies a lesser degree 
of certainty about causality than adverse reaction, which means any adverse event caused by a 
drug (21 CFR 312.32(a)). 
 
12.2.3 Unexpected Adverse Event 
 
An adverse event or suspected adverse reaction is considered “unexpected” if it is not listed in 
the package insert or is not listed at the specificity, severity, or rate of occurrence that has been 
observed. 
 
12.2.4  Serious Adverse Events 
 
An adverse event or suspected adverse reaction is considered “serious” if, in the view of the 
investigator, it results in any of the following outcomes (21 CFR 312.32(a)): 
1. Death 
2. A life-threatening event: An AE or SAR is considered “life-threatening” if, in the view of 
the investigator, its occurrence places the subject at immediate risk of death. It does not 
include an AE or SAR that, had it occurred in a more severe form, might have caused 
death. 
3. Inpatient hospitalization or prolongation of existing hospitalization 
4. Persistent or significant incapacity or substantial disruption of the ability to conduct 
normal life functions 
5. Congenital anomaly or birth defect 
6. Important medical events that may not result in death, be life threatening, or require 
hospitalization may be considered serious when, based upon appropriate medical 
judgement, they may jeopardize the subject and may require medical or surgical 
intervention to prevent one of the outcomes listed above.  
12.3 GRADING AND ATTRIBUTION OF ADVERSE EVENTS 
 
12.3.1 Grading Criteria 
 

68 
 
The study site will grade the severity of adverse events experienced by the study subjects 
according to the criteria set forth in the National Cancer Institute’s Common Terminology 
Criteria for Adverse Events Version (CTCAE) (version 5.0 published Nov 27, 2017). This document 
(referred to herein as the NCI-CTCAE manual) provides a common language to describe levels of 
severity, to analyze and interpret data, and to articulate the clinical significant of all adverse 
events.  
 
Adverse events will be graded on a scale from 1 to 5 according to the following standards in the 
NCI-CTCAE manual: 
 
Grade 1 = Mild; asymptomatic or mild symptoms; clinical or diagnostic observations 
only; intervention not indicated. 
Grade 2 = Moderate; minimal, local or noninvasive intervention indicated; limiting age-
appropriate instrumental ADL*. 
Grade 3 = Severe or medically significant but not immediately life-threatening; 
hospitalization or prolongation of hospitalization indicated; disabling; 
limiting self care ADL**. 
Grade 4 = Life-threatening consequences; urgent intervention indicated. 
Grade 5 = Death related to AE. 
 
Activities of Daily Living (ADL) 
 
*Instrumental ADL refer to preparing meals, shopping for groceries or clothes, using the 
telephone, managing money, etc. 
 
**Self care ADL refer to bathing, dressing and undressing, feeding self, using the toilet, 
taking medications, and not bedridden. 
 
Adverse events will be collected from the time of the first protocol mandated procedure until 
the study completion, or until 30 days after the subject prematurely withdraws from the study. 
 
12.3.2 Attribution Definitions 
 
The relationship, or attribution, of an adverse event to the study therapy regimen or study 
procedure(s) will be determined by the site investigator and recorded on the appropriate AE 
eCRF. The relationship of an adverse event to study therapy regimen or procedures will be 
determined using the descriptors and definitions provided in the table below. 
 
NCI-CTCAE attribution of adverse events 
Code 
Descriptor 
Relationship 
Unrelated Category 
1 
Unrelated 
The adverse event is clearly not related. 
Related Categories 
2 
Possible 
The adverse event has a reasonable possibility to be 
related; there is evidence to suggest a causal 
relationship. 
3 
Definite 
The adverse event is clearly related 

69 
 
 
 
12.4 COLLECTION AND RECORDING OF ADVERSE EVENTS 
 
12.4.1 Collection Period 
 
Adverse events will be collected from the time of first protocol mandated procedure, until 
he/she completes study participation or until 30 days after he/she prematurely withdraws 
(without withdrawing consent) or is withdrawn from the study. 
 
12.4.2 Collecting Adverse Events 
 
Adverse events (including SAEs) may be discovered through any of these methods: 
• Observing the subject 
• Questioning the subject in an objective manner 
• Receiving an unsolicited complaint from the subject 
• Clinical or laboratory evaluation, or medical records demonstrating pregnancy or 
overdose with or without an adverse drug reaction 
• In addition, an abnormal value or result from a clinical or laboratory evaluation can also 
indicate an adverse event, as defined in section 11.3, Grading and Attribution of Adverse 
Events. 
12.4.3 Recording Adverse Events 
 
Throughout the study, the investigator will record adverse events and serious adverse events as 
described previously on the appropriate eCRF regardless of the relationship to study therapy 
regimen or study procedure. 
 
All adverse events must be recorded by the site on the appropriate AE/SAE eCRF as outlined in 
the terms of the contract. 
 
Once recorded, an AE/SAE will be followed until it resolves with or without sequelae, or until the 
end of study participation, or until 30 days after the subject prematurely withdraws or is 
withdrawn from the study, whichever occurs first. In summary, AEs/SAEs will be followed until 
they resolve or for 30 days after a subject is withdrawn or withdraws from the study. 
 
12.4.4 Reporting of Serious Adverse Events 
 
All investigators must report adverse events, including expedited reports, in a timely fashion to 
their respective IRBs in accordance with applicable regulations and guidelines. All adverse 
events will be reported to Sanofi as outlined in the terms of the contract. 
 
 
12.5 FOLLOW-UP OF ADVERSE EVENTS 
 
All adverse events must be documented and followed up until the event is either resolved or 
adequately explained, even after the patient has completed his/her trial treatment. 

70 
 
 
12.5.1 Planned Data and Safety Monitoring Reviews 
 
The Data and Safety Monitoring Board will review safety data at least yearly during planned Data 
and Safety Monitoring Meetings. Data for the planned safety reviews will include, at a minimum, 
a listing of all reported AEs and SAEs.   
 
The Data and Safety Monitors will be informed of an Expedited Safety Report in a timely manner. 
 
12.5.2 Ad Hoc Data and Safety Reviews 
 
In
 addition
 to
 the
 pre-scheduled
 data
 reviews
 and
 planned
 safety
 monitoring,
 the
 Data
 and
 Safety 
Monitors may be called upon for ad hoc reviews per the request of the principal investigator. 
 
 
13. 
MECHANISTIC ASSAYS 
 
The objective of the mechanistic studies is to determine how ATG affects the immune system, 
both cellular and humoral, and, in particular, how ATG alters the immune response to the 
allograft during and after induction therapy.  Mechanistic studies in the pre-transplant period 
will establish a baseline against which future laboratory results can be compared.  The post-
transplant mechanistic studies will elucidate the pathophysiologic and genetic mechanisms 
associated with the cellular and immune response against the allograft, the response to 
induction therapy, and graft rejection. 
 
The immune cells and circulating antibodies referred to in secondary efficacy parameter (A) will 
be defined and measured as follows: 
 
13.1 Blood for circulating antibodies 
 
Blood for circulating antibodies will be tested for by standardized alloantibody testing by Luminex 
and control sera.
158  We will use these standardized methods to define kinetics, specificities and 
relative binding strength (mean fluorescence intensity, MFI) of DSA in each patient. Serum 
samples will be obtained at pre-transplant, 1, 3, 6, and 12 months post-transplantation. The HLA 
specificity, class I vs. II, and relative binding strength of the anti-HLA IgG will be determined using 
Luminex-platform single HLA class I and class II antigen microbead assays in combination with 
phycoerythrin (PE) secondary antibody conjugates for the detection of human IgG.  To minimize 
any prozone effect (interfering factors which mask the antibody binding strength (MFI) serum 
samples
 will
 be
 diluted
 1:8
 according
 to
 established
 protocol
 to
 assess
 if
 there
 is
 an
 MFI
 increment 
after dilution. The Luminex single antigen beads will be analyzed using the LABScan™ 100 flow 
analyzer and an analysis program will assist in the assignment of the reaction strength (MFI).  The 
presence and relative strength of non-HLA specific antibodies will be determined by ELISA and 
Luminex-platform assays.  The final diagnosis of circulating antibodies will be made by an 
immunologist. 
 
13.2 PRT ASSAY 
 

71 
 
The panel of reactive T-cells (PRT) assay consists of PBMC from transplant candidates pre-
transplant being tested in IFN-gamma ELISPOT assays against a panel of HLA disparate B cell 
stimulator line.  Results of the PRT have previously correlated with worse 12-month outcomes in 
kidney transplant recipients, and our goal here is to observe whether this is the case in heart 
transplant recipients, as well as whether ATG affects this correlation. 
 
Regarding definition of a positive PRT, the optimal approach for quantifying and analyzing the 
results of the PRT assay, particularly for heart transplant recipients, remains unclear. We plan to 
use this opportunity to identify the optimal analytic approach.  Our published data indicate that 
any stimulator that induces >25 IFNγ ELISPOTs per 300,000 responder cells represents a positive 
response to that stimulator, and that the percentage of positive responses (out of the total 
number of stimulators tested) correlates with risk of developing AR or lower GFR after kidney 
transplantation.
159,160  An alternative analytic strategy involves summing the responses to each 
stimulator and correlating the total (sum) with outcome.
158  The optimal number of stimulators 
to be included in the assay has not been formally determined, although our published data 
strongly indicate that 6 randomly selected distinct stimulator lines is sufficient.161  We will define 
each result as either a) percentage of positive responses based on >25 or b) sum of the responses 
to each stimulator and calculate the results using all 10 stimulators.  We will initially handle each 
variable as continuous (e.g. no. of positive ELISPOTs, no. of lines to which the responses are >25) 
and in a secondary analysis use ROC curves to define a potential threshold effect (e.g. responses 
to > 60% of stimulators or total responses > 400 ELISPOTs correlate with outcome). Depending on 
the results we will test alternative definitions of a positive result against a single stimulator (e.g. 
>50 or >100). We will also randomly and repeatedly analyze sets of responses from 4-10 
stimulators from within the data to identify the lowest number of stimulators required to provide 
reliable correlations.
161 
 
13.3 LYMPHOCYTE ANALYSIS 
 
Each blood sample from each subject at each time point (Pre- Transplant, Month 3 and Month 6 
timepoints) will be analyzed by multiparameter flow cytometry to determine what proportion of 
the patient’s immune cells are naïve, T effector, T memory, or B-cells.  The immune cell 
populations will be identified by the following profiles: 
 
(1) Allogeneic T-cells will be grouped as either CD4+ or CD8+ and will be further sub-
classified as naïve, memory, or effector based on presence or absence of CD45RA, 
CD49a and CD62L.162-164 
(2) T regulatory cells will be defined as CD3+, CD127(lo), CD4+, and CD25+  and Foxp3.
165,166  
We will also 
include measurements of surface expressed and 
intracellular CTLA4, surface 
expressed (GITR) and intracellular pSTAT5 (among others) as these markers have been 
used by others as indirect correlates of T-reg function.
167 
(3) Accordingly CD8 suppressor cells will be defined as CD8+ CD3+ CD127(lo) and CD28-.
167 
(4) Dendritic cells or non-T-cell subsets will be defined as CD14-, CD56-, CD3-, CD19-.
168,169 
(5) B-cells will be defined as CD24+.  Pre-pro-B cells will be defined as CD24- CD43+.  Pro-B 
cells will be defined as CD24(int) CD43+.  pre-B cells will be defined as CD24(hi) CD43-. 
 
13.4 CYTOKINES 
 

72 
 
The production of pro-inflammatory markers (such as IL-1-beta, IL-2, IL-6, IL-17, TNF-alpha, IFN-
gamma), and anti-inflammatory cytokines (such as IL-10, TGF-beta) will be measured pre-
transplantation, then at 3 and 6 months. Post-transplant.  These markers are important as they 
mediate the immune response in graft rejection and CAV.  The cytokine milieu will give insight 
into the changes to the immune system from ATG induction therapy, its effectiveness, and the 
subsequent recovery of the immune system as the effects of ATG induction therapy dissipate.  
 
14. 
STATISTICAL CONSIDERATIONS 
 
14.1 SAMPLE SIZE CALCULATION 
 
The study is intended to serve as a preliminary study. Comparative analyses will be primarily 
descriptive due to a small sample size, with an aim of 60 patients, randomized 1:1 to the 
treatment arm or control arm. The study is adequately powered to assess treatment efficacy for 
the composite primary endpoint only. The secondary parameters are for descriptive purposes 
only. 
 
 
Sample size calculation for primary composite endpoint: 
 
Assuming that the incidences of the composite primary endpoint are 40% in the control arm and 
10% in the treatment arm, then under Fisher’s exact conditional test for two proportions, a 
sample size of 60 detects a reduction in the composite primary endpoint in the treatment arm 
with a power of 70%. 
 
14.2 ANALYSIS PLAN 
 
14.2.1 Primary Efficacy Parameters 
 
(A) Incidence of the primary composite endpoint of the development of de novo donor 
specific antibodies and ischemia on endomyocardial biopsy at 12 months post-
transplant  
 
  
14.2.2 Secondary Efficacy Parameters 
 
To compare between groups: 
 
(A) Changes in percentages of various subsets of immune cells at pre transplant, 3 months 
and 6 months.  Changes in percentages of various circulating antibodies at pre-
transplant, 1 month, 3 months, 6 months, and 12 months after transplant. 
(B) Freedom from cardiac allograft vasculopathy (CAV) (defined as a change ≥0.5mm in 
maximal intimal thickness (MIT) of the coronary arteries by intravascular ultrasound at 
12 months as compared to baseline) 
(C) Change in coronary maximal intimal thickness intimal area, intimal volume, vessel area, 
intimal index, and percent atheroma volume at matched sites by intravascular 
ultrasound at 12 months 
(D) Freedom from acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection within first 12 months after transplantation 

73 
 
(E) Freedom from acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection episodes per patient within the first 12 months post-transplantation 
(F) Freedom from acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection by the ISHLT biopsy grading scale in the first 12 months post-
transplantation 
(G) Freedom from biopsy proven cellular rejection ≥2R, biopsy proven antibody mediated 
rejection ≥AMR1 and any-treated rejection at 12 months post-transplantation 
(H) Time to first acute cellular, antibody-mediated, hemodynamic compromise, and any-
treated rejection within the first 12 months 
(I) Incidence of primary graft dysfunction (PGD) in the first 24 hours post-transplantation 
(J) Patient and graft survival at 12 months post-transplantation 
(K) The number of patients with both fatal and non-fatal infectious complications 
(especially CMV infection) within the first 12 months post-transplantation   
(L) Freedom from the development of circulating antibodies within the first 12 months 
post-transplantation 
(M) 
Average maintenance doses of Mycophenolate Mofetil, tacrolimus, and cumulative dose 
of corticosteroids at 12 months post-transplantation 
(N) Average number of hospital days per patient, both during the transplant period and 
during the post-transplant period at 3 months, 6 months, and 1 year, 
(O) Number of patients requiring hospitalization by 3 months, 6 months, or 12 months post-
transplantation. 
(P) Freedom from the development of circulating antibodies 
 
14.2.3 Definition of Parameters 
 
For both primary and secondary parameters, the 3-month assessment point will be defined as 
study day 90 ± 2 weeks, 6-month assessment point will be defined as study day 180 ± 2 weeks, 9-
month 
assessment
 point
 will
 be
 defined
 as
 study
 day
 270
 ± 
2
 weeks 
and
 the
 12-month
 assessment
 
point will be defined as study day 360 ± 4 weeks. 
 
For both the primary and secondary CAV parameters, the changes in maximal intimal thickness, 
averaged  maximal intimal thicknesses at 10 matched sites per patient, intimal area, intimal 
volume, vessel area, intimal index, and percent atheroma volume at the 12-month assessment 
point will be used for analysis. 
 
Patients with infectious complications will include: 
 
(1) 
Patients with viral infections, proven by positive serology and symptoms 
consistent with the suspected viral infection 
(2) 
Patients with documented bacterial, fungal, or parasitic infections 
 
 
The definition of rejection will be as follows: 
 
Patients with acute rejection will include: 
 
(1) 
Patients with acute cellular rejection that is biopsy proven with ISHLT grade 2R 
or worse histology 

74 
 
(2) 
Patients with acute antibody-mediated rejection that is biopsy proven with 
immunohistochemistry stains positive and concomitant significant 
hemodynamic compromise 
(3) 
Patients with hemodynamic compromise who are treated for acute cellular 
and/or antibody-mediated rejection, regardless of whether or not a biopsy is 
done and regardless of histologic findings if a biopsy is done 
(4) 
Patients who die within 12 months of transplantation before experiencing a 
primary end-point rejection 
(5) 
Patients who are lost to follow-up within 12 months after transplantation and 
have not experienced a primary endpoint rejection 
 
A rejection episode begins with a positive biopsy or with the start date of treatment for rejection, 
whichever comes first.  Resolution of rejection will be defined when no further treatment is 
required.  If continued treatment is required for the current rejection (according to the regimen 
specified in Table 1), the rejection is still considered an ongoing episode.  A second episode of 
rejection
 may
 not
 be
 counted
 until
 the
 first
 episode
 has
 resolved.
 
 Follow-up
 biopsies
 should
 occur
 
within 2 weeks (with an allowable window of 10 days to 21 days) after the end of treatment for 
the rejection. 
 
Time to first acute cellular, antibody-mediated, hemodynamic compromise, and any-treated 
rejection will be calculated from the date of transplantation to the date the patient first 
experiences acute rejection, date of death, date of re-transplantation, date of graft loss, or date 
of
 premature
 drop-out
 (for
 patients
 who
 are
 lost
 to
 follow-up), whichever
 comes
 first 
(see
 section
 
2.2.1 for definition of rejection).  If a patient never experiences any of these events while being 
followed, he/she will be censored at the last follow-up. 
 
Re-transplantation will be calculated form the date of original transplantation to the date of re-
transplantation or date of death, whichever comes first.  If a patient does not have a re-
transplantation or does not die, he/she will be censored at the last date of follow-up. 
 
Death will be calculated form the date of start of therapy to the date of death.  If a patient does 
not die while being followed, he/she will be censored at the last date he/she is known to be alive. 
 
14.2.4 Statistical and Analytical Methods 
 
A descriptive analysis of the endpoints will be applied owing to the small sample size. This is a 
multi-center pilot study and not designed, nor intended to provide data on the statistical 
significance of endpoints. 
 
14.2.5 Intent to Treat Population Analysis 
 
All patients randomized in the study will be involved in the statistical analyses.  The primary 
analysis population will be the intent-to-treat population. 
 
14.3 SAFETY DATA ANALYSIS 
 

75 
 
The safety population includes all patients who received at least one dose of drug (either 
Thymoglobulin
® or No Induction or Control) and for whom at least one post-baseline safety 
assessment was made. 
 
The safety parameters include clinical adverse events, malignancies, infectious complications, 
laboratory tests, and vital signs.  Incidence of clinical adverse events will be calculated overall, by 
body system, and by adverse event.  Incidence will also be presented by relationship to trial 
medication and severity.   
 
 
14.4 FINAL STUDY ANALYSIS 
 
A final study analysis will be prepared demonstrating descriptive statistics of the primary and 
secondary endpoints. 
 
15.   SOURCE DATA 
 
15.1  SOURCE DATA 
 
All study data should be verifiable by source documentation. Most often this will be present in 
the patient’s medical record. Except in rare instances, study documents (CRFs and data 
collection tools) are not source documents. “Shadow charts” – duplications of material from the 
medical record are not source documents. 
 
15.2 ACCESS TO SOURCE DATA 
 
The responsibilities of the investigator require that health authorities have access to (and may 
when required by applicable law copy) source documents. Unless required by the laws that 
permit copying of records, only the coded identity associated with documents or with other 
participant data may be copied (and all personally identifying information must be obscured). 
Authorized representatives as noted above are bound to maintain the strict confidentiality of 
medical and research information that is linked to identified individuals. 
 
16. 
QUALITY CONTROL AND QUALITY ASSURANCE 
 
The investigator is required to keep accurate records to ensure that the conduct of the study is 
fully documented. An internal auditor will regularly review the conduct of the trial, verify 
adherence to the protocol, and confirm the completeness, consistency, and accuracy of all 
documented data. 
 
16.1 DATA HANDLING 
 
The investigator is required to ensure that all CRFs are completed for every patient entered in 
the trial. All elements of data entry (i.e., time, date, verbatim text, and the name of the person 
performing the data entry) will be recorded with an audit trail to allow all data changes in the 
database to be monitored and maintained in accordance with federal regulations.  
 

76 
 
17. 
ETHICAL CONSIDERATIONS AND COMPLIANCE WITH GOOD CLINICAL PRACTICE 
 
17.1 STATEMENT OF COMPLIANCE 
 
The clinical study will be conducted using good clinical practice (GCP), as delineated in Guidance 
for Industry: E6 Good Clinical Practice Consolidated Guidance, and according to the criteria 
specified in the study protocol. Before study initiation, the protocol and the informed consent 
documents will be reviewed and approved by an appropriate IRB. Any amendments to the 
protocol or the consent materials must also be approved before they are implemented. 
 
 
17.2 INFORMED CONSENT 
 
The consent process provides information about the study to a prospective participant and 
allows adequate time for review and discussion prior to their decision. The principal investigator 
or designee will review the consent and answer questions. The prospective participant will be 
told that being in the trial is voluntary and that he or she may withdraw from the study at any 
time, for any reason. All participants (or their legally acceptable representative) must read, sign, 
and date a consent form before undergoing any study procedures. Consent materials must be 
presented in participants’ primary language or a short form may be used. A copy of the signed 
consent form must be given to the participant. 
 
The consent process is ongoing. The consent form must be revised when important new safety 
information is available, the protocol is amended, and/or new information becomes available 
that may affect participation in the study. 
 
17.3 PRIVACY AND CONFIDENTIALITY 
 
A participant’s privacy and confidentiality will be respected throughout the study. Each  
participant will be assigned a unique identification number and these numbers rather than 
names will be used to collect, store, and report participant information.  
 
  
 
 
 
 

77 
 
18. 
REFERENCES 
 
1. 
Lund LH, Edwards LB, Kucheryavaya AY, et al. The Registry of the International Society 
for Heart and Lung Transplantation: Thirty-second Official Adult Heart Transplantation 
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polyclonal anti-thymocyte globulins on the expression of adhesion and inflammation 
molecules after ischemia–reperfusion injury. Transplant immunology. 2009;20(4):224-
228. 
132. Walther S, Beiras-Fernandez A, Csapo C, et al. Influence of polyclonal antithymocyte 
globulins on the expression of adhesion molecules of isolated human umbilical vein 
endothelial cells. Paper presented at: Transplantation proceedings2010. 
133. Zarrini P, Aintablian T, Kittleson M, et al. Anti-thymocyte globulin protects against 
ischemia reperfusion injury in the immediate post heart transplant period. J Am Coll 
Cardiol. 2016;13(67):1452. 
134. Beniaminovitz A, Itescu S, Lietz K, et al. Prevention of rejection in cardiac transplantation 
by blockade of the interleukin-2 receptor with a monoclonal antibody. New England 
Journal of Medicine. 2000;342(9):613-619. 
135. Hershberger RE, Starling RC, Eisen HJ, et al. Daclizumab to prevent rejection after 
cardiac transplantation. New England Journal of Medicine. 2005;352(26):2705-2713. 

85 
 
136. Ansari D, Lund LH, Stehlik J, et al. Induction with anti-thymocyte globulin in heart 
transplantation is associated with better long-term survival compared with basiliximab. 
The Journal of Heart and Lung Transplantation. 2015;34(10):1283-1291. 
137. Rafiei M, Kittleson M, Patel J, et al. Anti-thymocyte gamma-globulin may prevent 
antibody production after heart transplantation. Transplantation Proceedings. 
2014;46(10):3570-3574. 
138. Zuckermann A, Ploner M, Czerny M, et al. Low incidence of graft arteriosclerosis after 
cardiac transplantation - risk factor analysis for patients with induction therapy. J Heart 
Lung Transplant. 2001;20(2):196. 
139. Lavee J, Bar-Zohar D, Har-Zahav Y, et al. Immunosuppression induction with rabbit 
antithymocyte globulin in heart transplantation patients: 6-year experience at the Sheba 
Medical Center. Transplantation Proceedings. 2000;32(4):737-739. 
140. Beaufigeau M, Dumesnil D, Carrier M, White M, Pelletier G. Seven-year experience with 
rabbit antithymocyte globulin after cardiac transplantation at the montreal heart 
institute. Transplantation Proceedings. 1997;29:10S-12S. 
141. Pietra BA, Boucek MM. Immunosuppression for pediatric cardiac transplantation in the 
modern era. Progress in Pediatric Cardiology. 2000;11(2):115-129. 
142. Tuzcu EM, Kapadia SR, Tutar E, et al. High Prevalence of Coronary Atherosclerosis in 
Asymptomatic Teenagers and Young Adults : Evidence From Intravascular Ultrasound. 
Circulation. 2001;103(22):2705-2710. 
143. Bocksch W, Wellnhofer E, Schartl M, et al. Reproducibility of serial intravascular 
ultrasound measurements in patients with angiographically silent coronary artery 
disease after heart transplantation. Coronary Artery Disease. 2000;11(7):555-562. 
144. Son R, Tobis JM, Yeatman LA, Johnson JA, Wener LS, Kobashigawa JA. Does use of 
intravascular ultrasound accelerate arteriopathy in heart transplant recipients? 
American Heart Journal. 1999;138(2):358-363. 
145. Ramasubbu K, Schoenhagen P, Balghith MA, et al. Repeated intravascular ultrasound 
imaging in cardiac transplant recipients does not accelerate transplant coronary artery 
disease. Journal of the American College of Cardiology. 2003;41(10):1739-1743. 
146. Batkoff BW, Linker DT. Safety of intracoronary ultrasound: data from a Multicenter 
European Registry. Cathet Cardiovasc Diagn. 1996;38(3):238-241. 
147. Tuzcu EM, Kapadia SR, Tutar E, et al. High prevalence of coronary atherosclerosis in 
asymptomatic teenagers and young adults: evidence from intravascular ultrasound. 
Circulation. 2001;103(22):2705-2710. 
148. Bocksch W, Wellnhofer E, Schartl M, et al. Reproducibility of serial intravascular 
ultrasound measurements in patients with angiographically silent coronary artery 
disease after heart transplantation. Coron Artery Dis. 2000;11(7):555-562. 
149.  Wahlers T. Cytolytic Induction Therapy in Heart and Lung Transplantation: The 
Protagonist Opinion. Transplantation Proceedings. 1998;30(4):1100-1103. 
150. Bell L, Girardin C, Sharma A, Goodyer P, Mazer B. Lymphocyte subsets during and after 
rabbit anti-thymocyte globulin induction in pediatric renal transplantation: sustained T 
cell depletion. Transplant Proc. 1997;29(7A):6S-9S. 
151. Schnetzler B, Leger P, Volp A, Dorent R, Pavie A, Gandjbakhch I. A prospective 
randomized controlled study on the efficacy and tolerance of two antilymphocytic 
globulins in the prevention of rejection in first-heart transplant recipients. Transpl Int. 
2002;15(6):317-325. 

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152. Lebeck LK, Chang L, Lopez-McCormack C, Chinnock R, Boucek M. Polyclonal 
antithymocyte serum: immune prophylaxis and rejection therapy in pediatric heart 
transplantation patients. J Heart Lung Transplant. 1993;12(6 Pt 2):S286-292. 
153. Lavee J, Bar-Zohar D, Har-Zahav Y, et al. Immunosuppression induction with rabbit 
antithymocyte globulin in heart transplantation patients: 6-year experience at the Sheba 
Medical Center. Transplant Proc. 2000;32(4):737-739. 
154. Brophy PD, Thomas SE, McBryde KD, Bunchman TE. Comparison of polyclonal induction 
agents in pediatric renal transplantation. Pediatric Transplantation. 2001;5(3):174-178. 
155. Bell L, Girardin C, Sharma A, Goodyer P, Mazer B. Lymphocyte subsets during and after 
rabbit anti-thymocyte globulin induction in pediatric renal transplantation: Sustained T 
Cell Depletion. Transplantation Proceedings. 1997;29:6S-9S. 
156. Akalin E, Ames S, Sehgal V, et al. Intravenous immunoglobulin and thymoglobulin 
facilitate kidney transplantation in complement-dependent cytotoxicity B-cell and flow 
cytometry T- or B-cell crossmatch-positive patients. Transplantation. 2003;76(10):1444-
1447. 
157. Cantarovich M, Giannetti N, Barkun J, Cecere R. Antithymocyte globulin induction allows 
a prolonged delay in the initiation of cyclosporine in heart transplant patients with 
postoperative renal dysfunction. Transplantation. 2004;78(5):779-781. 
158. Reed EF, Rao P, Zhang Z, et al. Comprehensive Assessment and Standardization of Solid 
Phase Multiplex-Bead Arrays for the Detection of Antibodies to HLA. American Journal 
of Transplantation. 2013;13(7):1859-1870. 
159. Poggio ED, Augustine JJ, Clemente M, et al. Pretransplant Cellular Alloimmunity as 
Assessed by a Panel of Reactive T Cells Assay Correlates With Acute Renal Graft 
Rejection. Transplantation. 2007;83(7):847-852. 
160. Poggio ED. Panel of Reactive T Cells as a Measurement of Primed Cellular Alloimmunity 
in Kidney Transplant Candidates. Journal of the American Society of Nephrology. 
2006;17(2):564-572. 
161. Sawinski D, Uribarri J, Peace D, et al. 25-OH-Vitamin D Deficiency and Cellular 
Alloimmunity as Measured by Panel of Reactive T Cell Testing in Dialysis Patients. 
American Journal of Transplantation. 2010;10(10):2287-2295. 
162. Picker LJ, Singh M, Zdraveski Z, et al. Direct demonstration of cytokine synthesis 
heterogeneity among human memory/effector T cells by flow cytometry. Blood. 
1995;86(4):1408-1419. 
163. Tomiyama H, Matsuda T, Takiguchi M. Differentiation of Human CD8+ T Cells from a 
Memory to Memory/Effector Phenotype. The Journal of Immunology. 
2002;168(11):5538-5550. 
164. Bank I, Book M, Ware R. Functional Role of VLA-1 (CD49A) in Adhesion, Cation-
Dependent Spreading, and Activation of Cultured Human T Lymphocytes. Cellular 
Immunology. 1994;156(2):424-437. 
165. Akimova T, Kamath BM, Goebel JW, et al. Differing Effects of Rapamycin or Calcineurin 
Inhibitor on T-Regulatory Cells in Pediatric Liver and Kidney Transplant Recipients. 
American Journal of Transplantation. 2012;12(12):3449-3461. 
166. van der Touw W, Cravedi P, Kwan Wh, Paz-Artal E, Merad M, Heeger PS. Cutting Edge: 
Receptors for C3a and C5a Modulate Stability of Alloantigen-Reactive Induced 
Regulatory T Cells. The Journal of Immunology. 2013;190(12):5921-5925. 
167. Manavalan JS. Alloantigen specific CD8+CD28- FOXP3+ T suppressor cells induce ILT3+ 
ILT4+ tolerogenic endothelial cells, inhibiting alloreactivity. International Immunology. 
2004;16(8):1055-1068. 

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168. Savary CA, Grazziutti ML, Melichar B, et al. Multidimensional flow-cytometric analysis of 
dendritic cells in peripheral blood of normal donors and cancer patients. Cancer 
Immunology, Immunotherapy. 1998;45(5):234-240. 
169. Sorg RV, Kögler G, Wernet P. Cord blood dendritic cells predominantly display a 
PRECURSOR-DC2-like phenotype but induce TH1 t-cell responses. Experimental 
Hematology. 2000;28(7):71-72. 
 

88 
 
APPENDIX A 
 
A-1 Standardized ISHLT Cardiac Biopsy Grading 
 
GRADE ACUTE CELLULAR REJECTION (2004) 
0 
No rejection 
1R 
Interstitial and/or perivascular infiltrate with up to 1 
focus of myocyte damage 
2R 
Two or more foci of infiltrate with associated 
myocyte damage 
3R 
Diffuse infiltrate with multifocal myocyte damage ± 
edema, ± hemorrhage ± vasculitis 
 
 
GRADE 
ANTIBODY MEDIATED REJECTION (2013) 
pAMR 0: Negative 
for pathological 
AMR 
Both histological and immunopathologic 
studies are negative 
pAMR 1 (H+): 
Histopathologic 
AMR alone 
Histological findings present and 
immunopathologic findings negative 
pAMR1 (I+): 
Immunopathologic 
AMR alone 
Histological findings negative and 
immunopathologic findings positive 
pAMR 2: 
Pathological AMR 
Both histological and immunopathologic 
findings are present 
pAMR 3: Severe 
pathological AMR 
Severe AMR with histopathologic findings 
of interstitial hemorrhage, capillary 
fragmentation, mixed inflammatory 
infiltrates, endothelial cell pyknosis 
and/or karyorrhexis, and marked edema 
 
 
 
A-2 Acute Rejection—Diagnosis and Treatment 
 
ISHLT 
BIOPSY 
GRADE 
NO HEMODYNAMIC 
COMPROMISE PRESENT 
HEMODYNAMIC  
COMPROMISE PRESENT 
1R 
No treatment 
High dose IV 500 mg corticosteroids for 
3-10 days 
OR 
5 days of ATG 1.5 mg/kg 
2R 
500 mg IV corticosteroids for 3 days 
followed by immediate return to 
baseline 
High dose IV 500 mg corticosteroids for 
3-10 days ± 5-10 day taper to baseline 
AND 
5 days of ATG 1.5 mg/kg 

89 
 
ISHLT 
BIOPSY 
GRADE 
NO HEMODYNAMIC 
COMPROMISE PRESENT 
HEMODYNAMIC  
COMPROMISE PRESENT 
3R 
High dose IV 500 mg corticosteroids 
with 5-10 day taper to baseline and 5 
days of ATG 1.5 mg/kg  
High dose IV 500 mg corticosteroids with 
5-10 day taper to baseline and 5 days of 
ATG 1.5 mg/kg  

 
90 
 
APPENDIX B 
 
CLASSIFICATION OF INFECTIOUS COMPLICATIONS  
 
A.  CMV 
 
CMV disease is defined as CMV infection supported by virologic evidence of active CMV replication by 
either viral culture (including shell-vial technique), or CMV antigenemia assay, or CMV PCR assay along 
with one of the following: 
 
CMV syndrome: defined
 as 
episodic
 fever
 and
 no response
 to antibiotics
 if used (i.e.,
 two spikes of
 fever
 
greater than 38 C during the 48 hours when antibiotics are being used); plus one of the following: 
• Malaise 
• Fall in neutrophil count over three consecutive daily measurements 
 
OR 
 
CMV hepatitis: Defined as evidence of CMV in the liver confirmed by liver biopsy. The biopsy must be 
characterized by one of more of the following:  
• Presence of cells with positive immunostaining or immunofluorescence or in situ hybridization 
for CMV or CMV inclusions 
• Histological evidence of CMV hepatitis 
 
In addition, at least one liver function test value (AST, alkaline phosphatase, or bilirubin) must be 
significantly outside the normal range. 
 
OR 
 
CMV gastroenteritis, esophagitis, or colitis confirmed by biopsy. The biopsy must be characterized by 
the presence of cells with positive immunostaining or immunofluorescence or in situ hybridization for 
CMV or CMV inclusions. In addition, one of the following must be present: 
• Signs and symptoms of upper gastrointestinal tract infection including: nausea, vomiting, 
anorexia, dysphagia, abdominal pain, or 
• Cramping, signs, and symptoms of colitis including persistent diarrhea, cramping or abdominal 
pain. 
 
OR 
 
CMV disease with involvement of the lung confirmed by Broncho-alveolar lavage (BAL) characterized 
by the following: 
• Presence of cells with positive immunostaining or immunofluorescence or in situ hybridization 
for CMV or CMV inclusions or positive viral culture. 
 
In addition, at least two of the following must be present: 
a) Dyspnea 
b) Interstitial infiltrates on chest x-ray 

 
91 
 
c) Requirement for supplemental oxygen and/or ventilator assistance or decreased pO2 (<80 
mmHg) with an increased A-a gradient (i.e., >20). 
OR 
 
CMV retinitis in one or both eyes based on dilated fundus exam by an ophthalmologist experienced in 
the diagnosis of CMV retinitis. 
 
 
B.  Aspergillus: 
       1.  Sputum 
 2.  Pulmonary or sinus invasive disease 
 3.  Disseminated or metastatic disease 
 
C.  Candida: 
 1.  Urinary tract infection 
 2.  Mucocutaneous (including oral, esophageal, rectal, and vaginal) 
 3.  Invasive tissue disease 
 4.  Fungemia/disseminated disease 
 
D.  Pneumocystis: 
 1.  Pulmonary disease 
 
E. Cryptococcus: 
 1.  Crytococcosis 
 
F.  Listeria: 
 1.  Listeriosis 
 
G.  Herpes zoster: 
 1.  Localized (1-2 dermatomes) 
 2.  Disseminated cutaneous disease 
 3.  Visceral disease 
 
H.  Herpes simplex: 
 1.  Herpes simplex 
 

 
92 
 
SCHEDULE OF ASSESSMENTS 
 
Screening
/Baseline¹ 
Day 
of3 Htx 
Visits Timeline 
 
STUDY 
WEEK/MONTH 
0 
W1 
W2 W3 W4 W6 W8 M3 M4 M5 M6 
M8 M10 M12 
STUDY DAY 
0 
1 
2 
3 
4 
5 7 14 21 28 42 56 907 1207 1507 1807 2407 3007 3657 
VISIT NUMBER 
0 
1 
2 
3 
4 
5 6 7 
8 
9 10 11 
12 
13 
14 
15 
16 
17 
18 
VISIT WINDOWS 
Prior to Tx 
Tx 
- 
- 
- 
- 
+/- 3 days 
+/- 4 days 
+/- 14 days 
+/- 30 days 
ASSESSMENT VISITS 
Medical History 
X 
 
Informed Consent 
X 
 
Randomization2 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Clinical Assessment 
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Immunosuppressant 
Therapy 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Study Drug 
Administration 
 
X6 
X6 
X6 
X6 
X6 
 
Pre-Treatment 
Medications 
 
X 
X 
X 
X 
X 
 
Antimicrobial 
Prophylaxis  
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Fungal prophylaxis  
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
 
CMV Prophylaxis  
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
 
Corticosteroids 
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
 
HMG CoA Reductase 
Inhibitor 
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Vitamin C, E 
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Aspirin 
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Concomitant 
Medications  
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
             SAFETY 
Vital Signs/Body 
weight 
 
X5 
X5 
X5 
X5 
X5 X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Adverse Events 
 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 

 
93 
 
 
Screening
/Baseline¹ 
Day 
of3 Htx 
Visits Timeline 
 
STUDY 
WEEK/MONTH 
0 
W1 
W2 W3 W4 W6 W8 M3 M4 M5 M6 
M8 M10 M12 
STUDY DAY 
0 
1 
2 
3 
4 
5 7 14 21 28 42 56 907 1207 1507 1807 2407 3007 3657 
VISIT NUMBER 
0 
1 
2 
3 
4 
5 6 7 
8 
9 10 11 
12 
13 
14 
15 
16 
17 
18 
VISIT WINDOWS 
Prior to Tx 
Tx 
- 
- 
- 
- 
+/- 3 days 
+/- 4 days 
+/- 14 days 
+/- 30 days 
Infectious 
Complications 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
EFFICACY  
Data Collection 
Rejection 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Echocardiogram
4 
 
 
 
 
 
 X10 X X10 X X10 
X 
X 
X 
X 
X 
X 
X 
X 
Endomyocardial 
Biopsy4,10 
 
 
 
 
 
 X10 X 
X10 X 
X10 
X 
X 
X10 X10 
X 
X10 
X10 
X 
Right Heart 
Catheterization4,11 
 
 
 
 
 
 X10 X 
X10 X 
X11 
 
X 
X10 X10 
X 
X10 
X10 
X 
Intravascular 
Ultrasound4,11 
 
 
 
 
 
 
 
 
 
 
X11 
 
 
 
 
 
 
 
X7 
Data Collection 
Patient/Organ Survival 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
LABORATORY TESTS 
Hematology (CBC with 
Diff) 4, 9  
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Serum Chemistry
4, 9  
X 
X 
X 
X 
X 
X X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Lipid Profile
9 
X 
 
 
 
 
 
X X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Blood Loss 
 
X 
X 
X 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
Serum Pregnancy Test 
(female) 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Trough levels 
(Tacrolimus) 
4 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X7 
X7 
X7 
X7 
X7 
X7 
X7 
HLA Donor Mismatch 
X 
 
 
 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X 
Cytomegalovirus Status 
(recipient and donor) 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
94 
 
 
Screening
/Baseline¹ 
Day 
of3 Htx 
Visits Timeline 
 
STUDY 
WEEK/MONTH 
0 
W1 
W2 W3 W4 W6 W8 M3 M4 M5 M6 
M8 M10 M12 
STUDY DAY 
0 
1 
2 
3 
4 
5 7 14 21 28 42 56 907 1207 1507 1807 2407 3007 3657 
VISIT NUMBER 
0 
1 
2 
3 
4 
5 6 7 
8 
9 10 11 
12 
13 
14 
15 
16 
17 
18 
VISIT WINDOWS 
Prior to Tx 
Tx 
- 
- 
- 
- 
+/- 3 days 
+/- 4 days 
+/- 14 days 
+/- 30 days 
Epstein-Barr Virus Status 
(recipient and donor)  
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Circulating Antibodies 
(PRA)  
X 
 
 
 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
X7 
Mechanistic 
Assessments8 
X 
 
 
 
 
 
 
 
 
 
 
 
X8 
 
 
X8 
 
 
 
¹ Screening and consenting – eligible subjects will be enrolled in the study while on the UNOS waitlist.  Screening labs may be drawn after consent obtained and 
at any time prior to heart transplantation.   
2 Patients consented to the study and meeting study entry and randomization inclusion  and  exclusion criteria will be randomized in a 1:1 ratio to either 
Thymoglobulin® or No Induction or Control during the immediate post transplantation period.   
3 Day of Heart Transplant 
4 For subjects repatriated to Kaiser 60 days after heart transplant, to be performed per KP standard of care protocol. 
5 The first 2 doses (post op Day 0 and 1) will be infused over 8 hours.  Vital signs every 15 minutes x 1 hour, then every 30 minutes x 2 hours, then every hour for 
the remaining infusion period.  Once infusions are completed, return to routine vital signs monitoring frequency. subsequent infusions (Days 3,4 &5) will be over 
a minimum period of 4-8 hours. 
6 Starting with 3
rd dose vital signs every hour during infusion.  
7 Study visits and procedures will be conducted at Kaiser locations per standard of care for patients that return their care back to Kaiser Permanente  
8 Kaiser patients will have blood drawn at their Kaiser location and the blood sample(s) will be shipped to Cedars-Sinai for the mechanistic assessments 
9 These laboratory tests will be collected as standard of care as required and as needed. If these labs are available, the data will be collected. However, if the labs 
are not collected as standard of care, this would not be considered a protocol deviation.  
10 May occur as SOC for patients who require high surveillance monitoring 
11 Early angiogram with IVUS will occur as standard of care between 5-8 weeks after transplant. The procedure will also include right heart catheterization, per 
local institutional SOC guidelines..