Randomized Phase II Study to Assess the Role of Nivolumab as Single Agent to Eliminate Minimal Residual Disease and Maintain Remission in Acute Myelogenous Leukemia (AML) Patients After Chemotherapy (REMAIN TRIAL)
Phase: PHASE2
Conditions: Acute Myeloid Leukemia
Sponsor: National Cancer Institute (NCI)

i 
SUMMARY OF CHANGES – Protocol 
 
For Protocol Amendment # 19 
 
NCI Protocol #: 9706 
Local Protocol #: CIRB 15-0185 
 
Protocol Date:    04/25/2023 
 
The following changes were made to protocol Amendment #18: 
 
# 
Section # 
Description 
1. 
n/a 
Protocol version date updated to 4/25/2023 
2. 
Cover Page 
PI updated from Hongtao Liu to Wendy Stock.   
3.  
Cover Page 
Responsible study coordinator updated to remove J Peterson and add 
CTSO contact information.  
 
4. 
Cover Page 
Responsible Research Nurse contact information deleted. 

1 
NCI Protocol #: 
9706 
 
Local Protocol #: 
CIRB15-0185 
 
ClinicalTrials.gov Identifier: NCT02275533 
 
TITLE:  RANDOMIZED PHASE II STUDY TO ASSESS THE ROLE OF NIVOLUMAB 
AS SINGLE AGENT TO ELIMINATE MINIMAL RESIDUAL DISEASE AND 
MAINTAIN REMISSION IN ACUTE MYELOGENOUS LEUKEMIA (AML) PATIENTS 
AFTER CHEMOTHERAPY (REMAIN TRIAL) 
 
CORRESPONDING  
ORGANIZATION: 
EDDOP-IL057/ University of Chicago Comprehensive Cancer 
Center  
 
Principal Investigator: 
Wendy Stock, MD 
University of Chicago Comprehensive Cancer Center  
5841 S. Maryland Ave. MC 2115 
Chicago, IL 60637 
Telephone: 773-834-8982 
Fax: 773-702-0963 
E-mail address:  wstock@medicine.bsd.uchicago.edu 
 
 
PARTICIPATING ORGANIZATIONS: 
  
LAO-NCI/ National Cancer Institute LAO 
LAO-11030 / University Health Network Princess Margaret Cancer Center LAO  
 
LAO-CA043 / City of Hope Comprehensive Cancer Center LAO  
LAO-CT018 / Yale University Cancer Center LAO  
 
LAO-MA036 / Dana-Farber - Harvard Cancer Center LAO  
 
LAO-MD017 / JHU Sidney Kimmel Comprehensive Cancer Center LAO  
 
LAO-MN026 / Mayo Clinic Cancer Center LAO  
 
LAO-NC010 / Duke University - Duke Cancer Institute LAO  
 
LAO-NJ066 / Rutgers University - Cancer Institute of New Jersey LAO  
 
LAO-OH007 / Ohio State University Comprehensive Cancer Center LAO  
 
LAO-PA015 / University of Pittsburgh Cancer Institute LAO  

NCI Protocol #: 9706 
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LAO-TX035 / University of Texas MD Anderson Cancer Center LAO  
 
EDDOP / Early Drug Development Opportunity Program 
 
NON-MEMBER COLLABORATORS  
n/a 
 
 
 
 
NCI-Supplied Agent):  Nivolumab (BMS-936558, MDX-1106, and ONO-4538) (NSC 
#748726) 
Other Agent(s):  Not applicable 
IND #:   
IND Sponsor:  DCTD, NCI 
 
PROTOCOL TYPE / VERSION # / VERSION DATE: 
Initial Protocol 09/17/2014 
 
Consensus Review 12/04/2014 
 
 
Follow Up Review 01/07/2015 
 
Follow Up Review 02/16/2015 
       
Follow Up Review 03/06/2015             
Follow Up Review 04/21/2015              
 
 
 
Initial Review Approval 05/05/2015             
Amendment #1 08/13/2015 
Amendment #2 10/23/2015 
Amendment #3 02/26/2016; 03.11.2016 
Amendment #4 07/07/2016 
Amendment #5 01/17/2017 (disapproved) 
Statistician: 
 
 
Theodore Karrison, PhD  
The University of Chicago  
5841 S. Maryland Ave.,  
MC 2007,  
R-314 Chicago, Illinois 60637  
Phone: 773-702-9326  
Fax: 773-702-1979  
tkarrison@health.bsd.uchicago.edu 
Lead Site Contact Details:  
For all regulatory inquires contact the general 
e-mail at 
PCCC_Regulatory@bsd.uchicago.edu 
 
For registration and data related inquires 
contact the general e-mail at  
phaseIIcra@medicine.bsd.uchicago.edu 
 
 
 
Responsible Study Coordinator(s):  
CTSO Quality Unit  
The University of Chicago  
5841 S. Maryland Ave. MC1140  
Chicago, IL 60637  
Phone: 773-702-9885 
Fax: 773-702-4889 
phaseIIcra@medicine.bsd.uchicago.edu 
  
 

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Amendment #6 01/25/2017 
Amendment #7 08/30/2017 
Amendment #8 02/20/2018 
Amendment #9 08/13/2018 (disapproved) 
Amendment #10 08/16/2018 (disapproved) 
Amendment #11 08/21/2018 
Amendment #12 01/29/2019 (disapproved) 
Amendment #13 02/21/2019 (disapproved) 
Amendment #14 03/22/2019 
Amendment#15 07/16/2019 (disapproved) 
Amendment#16 08/06/2019 
Amendment#17 09/24/2019 (in response to CIRB approved with modifications comments to Amendment #16) 
Amendment #18 03/12/2021 (response to RRA to update CAEPR to v2.4) 
Amendment #19 04/25/2023 (PI Change) 
 
 
 

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SCHEMA 
 
 
 
AML patients in first 
remission after 
induction or 
consolidation 
chemotherapy (not 
candidate for SCT) 
Screening and randomization 
 
Observation Arm 
Treated with Nivolumab 
(3mg/kg IV q2weeks till 
disease progression or 
intolerable toxicities or 
closure of the study 
D i s e a s e  R e l a p s e 
Off study  
Nivolumab Arm 
(3mg/kg IV q2weeks x46 
doses)  
Off study  

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TABLE OF CONTENTS 
 
SCHEMA .........................................................................................................................................3 
1. 
OBJECTIVES ......................................................................................................................8 
1.1 
Primary Objectives...................................................................................................8 
1.2 
Secondary Objectives...............................................................................................8 
1.3 
Exploratory Objectives ............................................................................................8 
2. 
BACKGROUND .................................................................................................................8 
2.1 
AML - Standard treatment .......................................................................................8 
2.2 
Immunotherapy of cancer ........................................................................................8 
2.3 
The role of negative co-stimulatory regulators in AML ..........................................9 
2.4 
CTEP IND Agent: Nivolumab ...............................................................................10 
2.5 
Rationale to utilize the PD-1-blocking antibody, Nivolumab, to eliminate 
MRD and maintain remission in AML patients after 
induction/consolidation chemotherapy ..................................................................14 
2.6 
Correlative Studies Background ............................................................................14 
3. 
PATIENT SELECTION ....................................................................................................15 
3.1 
Eligibility Criteria ..................................................................................................16 
3.2 
Exclusion Criteria ..................................................................................................19 
3.3 
Inclusion of Women and Minorities ......................................................................20 
4. 
REGISTRATION PROCEDURES (Rostered Protocol Model)........................................21 
4.1 
Investigator and Research Associate Registration with CTEP ..............................21 
4.2 
Site Registration .....................................................................................................22 
4.3 
Patient Registration ................................................................................................23 
4.4 
General Guidelines.................................................................................................25 
5. 
TREATMENT PLAN ........................................................................................................25 
5.1 
Agent Administration.............................................................................................25 
5.2 
General Concomitant Medication and Supportive Care Guidelines ......................26 
5.3 
Duration of Therapy ...............................................................................................26 
5.4 
Duration of Follow Up ...........................................................................................27 
5.5 
Criteria for Removal from Study ...........................................................................27 
5.6 
Criteria to Resume Treatment ................................................................................28 
5.7 
Treatment of Nivolumab-Related Infusion Reactions ...........................................29 
5.8 
Treatment with Nivolumab after Relapse in the Control Arm ..............................30 
6. 
DOSING DELAYS/DOSE MODIFICATIONS ...............................................................31 
7. 
ADVERSE EVENTS:  LIST AND REPORTING REQUIREMENTS ............................36 
7.1 
Comprehensive Adverse Events and Potential Risks List (CAEPR) for 
BMS-936556 ( Nivolumab, MDX-1106, NSC 748726) ........................................36 
7.2 
Adverse Event Characteristics ...............................................................................41 

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7.3 
Expedited Adverse Event Reporting ......................................................................42 
7.4 
Routine Adverse Event Reporting .........................................................................43 
7.5 
Secondary Malignancy...........................................................................................44 
7.6 
Second Malignancy ................................................................................................44 
8. 
PHARMACEUTICAL INFORMATION ..........................................................................44 
8.1 
CTEP IND Agent: Nivolumab (NSC 748726) ......................................................44 
9. 
BIOMARKER, CORRELATIVE, AND SPECIAL STUDIES ........................................48 
9.1 
Integral Laboratory ................................................................................................48 
9.2 
Integrated Correlative Studies................................................................................48 
9.3 
Exploratory Correlative Studies .............................................................................50 
9.4 
Special Studies .......................................................................................................55 
10. 
STUDY CALENDAR .......................................................................................................56 
11. 
MEASUREMENT OF EFFECT........................................................................................57 
11.1 
Antitumor Effect – AML .......................................................................................57 
11.2 
Other Response Parameters: ..................................................................................59 
12. 
DATA REPORTING / REGULATORY REQUIREMENTS ...........................................59 
12.1 
Data Reporting .......................................................................................................60 
12.2 
CTEP Multicenter Guidelines ................................................................................62 
12.3 
Collaborative Agreements Language .....................................................................62 
13. 
STATISTICAL CONSIDERATIONS...............................................................................63 
13.1 
Study Design/Endpoints.........................................................................................63 
13.2 
Sample Size/Accrual Rate ......................................................................................67 
13.3 
Stratification Factors ..............................................................................................68 
13.4 
Analysis of Secondary Endpoints ..........................................................................68 
13.5 
Reporting and Exclusions ......................................................................................69 
14. 
Study Status Updates and Study Closure ...........................................................................70 
14.1 
Definitions of Study Status Changes .....................................................................70 
14.2 
Responsibility for Filing Protocol Status Update Forms .......................................72 
REFERENCES ..............................................................................................................................73 
APPENDIX A 
PERFORMANCE STATUS CRITERIA ..................................................78 
APPENDIX B 
Blood Sample Collection Form .................................................................79 
APPENDIX C 
Bone Marrow Aspiration Collection Form ................................................81 
APPENDIX D 
Archived Bone Marrow Biopsy Acquisition Form ....................................83 
APPENDIX E 
Archived Bone Marrow Acquisition Form ................................................84 

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APPENDIX F 
SHIPPING INSTRUCTIONS ....................................................................85 
 
APPENDIX G 
Ficoll Isolation of Lymphocytes from Bone Marrow Aspirates and  
Whole Blood Samples                                                                                                        81 
APPENDIX H 
Risks Associated with Pregnancy ..............................................................87 
APPENDIX I 
MANAGEMENT ALGORITHMS FOR ENDOCRINOPATHY, 
GASTROINTESTINAL, HEPATIC, NEUROLOGICAL, PULMONARY, RENAL, 
AND SKIN ADVERSE EVENTS .....................................................................................93 
 
 

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1. OBJECTIVES 
 
1.1 
Primary Objectives 
 
To evaluate and compare the progression free survival rate after randomization in the two 
treatment arms (Nivolumab vs. observation) 
 
1.2 
Secondary Objectives 
 
1.2.1 To determine and compare the overall survival rates in the two arms 
1.2.2 To determine and compare the incidence of non-relapse mortality in the two arms 
1.2.3 To evaluate the toxicities of Nivolumab as maintenance 
 
1.3 
Exploratory Objectives 
 
1.3.1 To analyze PD-LI expression on AML cells from peripheral blood and/or bone marrow 
samples at diagnosis if available and at the time of study enrollment 
1.3.2 To monitor AML MRD by WT1 PCR at enrollment and at subsequent defined time points 
in the Nivolumab-treated and control groups 
1.3.3 To perform an exploratory analysis on the frequencies, absolute numbers and subsets of T 
cells (including regulatory T cells) in the Nivolumab-treated and control groups with an 
emphasis on activation markers 
1.3.4 To perform deep sequencing of TCR- and TCR- chains on polyclonal T cells at 
baseline and at subsequent time points in the Nivolumab and control groups 
 
 
2. BACKGROUND 
 
2.1 
AML - Standard treatment   
Standard intensive chemotherapy regimens induce complete remission in the majority of adults 
AML patients under age 60, but maintenance of durable remissions remains a challenge. Post-
remission strategies to eradicate minimal residual disease (MRD) after induction chemotherapy 
include repeated cycles of cytarabine-based consolidation chemotherapy (Mayer, Davis et al. 
1994), autologous stem cell transplantation (ASCT) (Farag, Ruppert et al. 2005) and allogeneic 
stem cell transplantation (Allo-SCT) (Cassileth, Harrington et al. 1998). For AML patients who 
are not candidates for SCT, the current standard of care is observation after 
induction/consolidation chemotherapy. Unfortunately, more than 50% of patients eventually will 
experience a disease relapse (Mayer, Davis et al. 1994). The outcome of older AML patients (>= 
60 years) is more dismal with an estimated 2 year progression free survival (PFS) of 
approximately 20% without SCT (Mrozek, Marcucci et al.). Thus, new post-remission strategies 
are needed in order to improve the long-term outcomes of AML patients. 
 
2.2 
Immunotherapy of cancer  
Over the past two decades, evidence has mounted suggesting that the immune system can play an 
active role in the elimination of malignant cells. However, although spontaneous anti-tumor 
immune responses are generated in a subset of patients, immune-mediated eradication of 

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established tumors is rare. These observations suggest the existence of immune evasion 
mechanisms which are activated in the cancer-bearing host which are capable of potent 
suppression of the anti-tumor immune response (Gajewski, Meng et al. 2006).  A growing 
number of putative immune evasion mechanisms have been characterized, including extrinsic 
suppression by regulatory T cells (Tregs), T cell anergy, diminished T cell activation by 
engagement of negative co-stimulatory T cell molecules, such as PD-1 and CTLA-4 (reviewed in 
(Gajewski, Meng et al. 2006; Zou 2006; Colombo and Piconese 2007)). Recently, strategies 
aimed at reversing immune escape pathways have led to impressive clinical results, particularly 
those which involve blocking so-called “checkpoint” inhibitors of T cell function, such as 
CTLA-4, and the PD-1/PD-L1 pathway (Hamid, Robert et al. ; Weber, Kudchadkar et al. ; 
Wolchok, Kluger et al. ; Bashey, Medina et al. 2009; Hodi, O'Day et al. 2010). 
 
2.3 
The role of negative co-stimulatory regulators in AML 
Up-regulation of negative T cell co-stimulatory receptors, such as CTLA-4 and PD-1, on tumor-
specific T cells clearly inhibit their effector function in solid malignancies, such as melanoma.  
Administration of antibodies which block these receptors, or their ligands in the case of PD-1, 
leads to clinical benefit in a subset of patients with melanoma and other solid tumors (Hamid, 
Robert et al. ; Weber, Kudchadkar et al. ; Wolchok, Kluger et al. ; Bashey, Medina et al. 2009; 
Hodi, O'Day et al. 2010).  In pre-clinical AML models, blocking CTLA-4 in combination with a 
peptide-based vaccine has led to enhanced CTL responses and prolonged survival, providing 
rationale for targeting this receptor in AML patients.   
 
With regard to the PD-1/PD-L1 pathway, we have generated pre-clinical data to suggest that this 
pathway is involved in immune evasion in AML. Our group (Dr. Justin Kline working with Dr. 
Thomas Gajewski) has published that the PD-1/PD-L1 pathway was involved in immune evasion 
in a murine AML model.  When C1498 AML cells were inoculated into animals, they grew 
progressively and apparently evaded immune destruction (Zhang, Gajewski et al. 2009). Low 
levels of PD-L1 expression were found on C1498 cells grown in vitro.  Exposure of AML cells 
to IFN led to significant PD-L1 up-regulation
Figure 1A) and (Figure 2A in (Zhang, 
Gajewski et al. 2009)). In addition, PD-L1 expression was up-regulated on C1498 cells when 
analyzed directly ex vivo (Figure 1B and Figure 2B in (Zhang, Gajewski et al. 2009)). PD-1-/- 
mice challenged with C1498 cells generated augmented antitumor T-cell responses, showed 
decreased AML burden in the blood and other organs, and survived significantly longer than 
wild-type mice did (Figure 1C and Figure 4C in (Zhang, Gajewski et al. 2009)). Similar results 
were obtained with a PD-L1 blocking antibody (Zhang, Gajewski et al. 2009). To determine 
whether PD-L1 was expressed on human AML cells, bone marrow samples from AML patients 
were analyzed for expression patterns of PD-L1. Three staining patterns were observed: 1) no 
PD-L1 expression on either CD34+ or CD34- cells, 2) expression of PD-L1 on the CD34-, but 
not on CD34+ cells, and 3) PD-L1 expression on CD34+ AML cells, but not on CD34- cells 
(Figure 1D).  These preliminary data support a hypothesis that PD-L1 expression on a subset of 
AML cases might serve as a possible baseline biomarker for response to PD-1-blocking 
antibodies, as has been demonstrated in melanoma patients (Topalian, Hodi et al.).  The 
identification of PD-L1 expression on human AML cells, in addition to pre-clinical data 
demonstrating a negative impact of the PD-1/PD-L1 pathway on anti-leukemia immunity; 
provide strong rationale for testing PD-1 blockade in AML patients.  

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Figure: PD-L1 is expressed on murine and human AML cells and promotes immune 
evasion in a pre-clinical AML model. (A) PD-L1 expression on C1498 AML cells by flow 
cytometry either at baseline or after in vitro exposure to IFN-  for 48 hours. The shaded 
histogram represents staining with an isotope control antibody. The dashed line represents PD-
L1 expression at baseline, and the solid line represents PD-L1 expression after exposure to IFN-
. (B) PD-L1 expression on C1498 AML cells following IV inoculation into C57BL/6 mice. (C) 
C1498 AML cells were inoculated into syngeneic control or PD-1-/- mice and survival was 
assessed. (D) Three patterns of PD-L1 expression on bone marrow samples from AML patients. 
Bone marrow samples were stained with anti-CD34 and anti-PD-L1 antibodies and analyzed by 
flow cytometry. Representative plots are shown above. 
 
2.4 
CTEP IND Agent: Nivolumab 
Nivolumab (BMS-936558, MDX-1106, and ONO-4538) is a fully human monoclonal 
immunoglobulin G4 (IgG4) antibody (HuMAb) that is specific for human programmed death-1 
(PD-1, cluster of differentiation 279 [CD279]) cell surface membrane receptor (Investigator 
Brochure, 2013).  PD-1 is a negative regulatory molecule that is expressed transiently following 
T-cell activation and on chronically stimulated T cells characterized by an “exhausted” 
phenotype.  Nivolumab binds to cynomolgus monkey PD-1 but not mouse, rat, or rabbit 
molecules.  Clinical activity of nivolumab has been observed in patients with melanoma, non-
small cell lung cancer (NSCLC), and renal cell carcinoma (RCC).   The combination of 
nivolumab and ipilimumab (anti-cytotoxic T lymphocyte associated antigen-4 [anti-CTLA-4]) in 
a phase 1/2 trial showed markedly enhanced clinical activity with an acceptable safety profile in 
melanoma patients (Wolchok et al., 2013).   
 
The clinical use of monoclonal antibodies to T-cell inhibitory receptors has provided 
transformative information on the nature of the immune system and cancer.  An emerging picture 
suggests that endogenous immune responses can mediate effective tumor regression and/or 
improved survival even in patients with large volume tumors resistant to other forms of therapy.  
Some of the unique features of this type of therapy, based largely on experience in advanced 
melanoma, include:  improved overall survival (OS) with or without radiographic responses or 
improved progression-free survival (PFS); responses that may be delayed or occur after 
radiographic disease progression; combinations of immune modulators with enhanced or novel 

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activities (in the example of ipilimumab and nivolumab); and toxicity that is almost exclusively 
immune or inflammatory in nature.  It is not yet clear what factors determine responses and 
which components of the immune system are needed for this to occur.  It seems likely that both 
memory helper and effector cells would be needed to sustain long-term responses. Increasing 
emphasis has been placed on understanding the relationships of the tumor, celluar infiltrate, and 
immunologic milieu surrounding each tumor.   
 
PD-1, a 55-kDa type 1 transmembrane protein, is a member of the CD28 family of T-cell co-
stimulatory receptors that include Ig super family member CD28, CTLA-4, inducible co-
stimulator (ICOS), and B and T lymphocyte attenuator (BTLA) (Investigator Brochure, 2013).  
PD-1 is transiently but highly expressed on activated T cells functioning to limit immune 
effectors at the site of activation.  Chronic stimulation may prevent the re-methylation of the PD-
1 gene leading to continuous expression and characterizes a state of “exhausted” T cells that lose 
function and proliferative capacity while enhancing a suppressive tumor microenvironment.  PD-
1 may act together with other T-cell modulating molecules, including CTLA-4, TIM-3, 
lymphocyte-activation gene 3 (LAG-3) as well as indoleamine-pyrrole 2,3-dioxygenase 1 (IDO-
1), cytokines, and transforming growth factor beta (TGF-beta).   
 
Two ligands specific for PD-1 have been identified:  PD-ligand 1 (PD-L1, also known as B7-H1 
or CD274, expressed on tumor, antigen-presenting cells [APCs], and dendritic cells [DCs]) and 
PD-L2 (also known as B7-DC or CD273, expressed on endothelial cells).  The interaction of PD-
1 with PD-L1 and PD-L2 results in negative regulatory stimuli that down-modulate the activated 
T-cell immune response through SHP-1 phosphastase.   
 
PD-1 knockout mice develop strain-specific lupus-like glomerulonephritis (C57BL/6) and 
cardiomyopathy (BALB/c).  In transplantable tumor models that expressed PD-1 and LAG-3 on 
tumor-infiltrating CD4+ and CD8+ T cells dual anti-LAG-3/anti-PD-1 antibody treatment cured 
most mice of established tumors that were largely resistant to single antibody treatment (Woo et 
al., 2012).  Despite minimal immunopathologic sequelae in PD-1 and LAG-3 single knockout 
mice, dual knockout mice abrogated self-tolerance with resultant autoimmune infiltrates in 
multiple organs, leading to eventual lethality. 
 
PD-L1 expression is found on a number of tumors, and is associated with poor prognoses based 
on OS in many tumors, including melanoma (Taube et al., 2012), renal (Thompson et al., 2004; 
Thompson et al., 2005; Thompson et al., 2006), esophageal (Ohigashi, et al. 2005), gastric (Wu 
et al., 2006), ovarian (Dong et al., 2003), pancreatic (Nomi, et al., 2007), lung (Zitvogel, et al., 
2006), and other cancers (Investigator Brochure, 2014). 
 
The PD-1/PD-L1 axis plays a role in human infections, particularly in hepatitis C virus (HCV) 
and human immunodeficiency virus (HIV).  In these cases, high expression levels of PD-1 were 
found in viral-specific CD8+ T cells that also display a non-responsive or exhausted phenotype.  
Non-responsive PD-1-high T cells were observed in simian immunodeficiency virus (SIV) 
infection in rhesus macaques.  Treatment of SIV-infected macaques with an anti-PD-1 mAb (3 
mg/kg x4) resulted in decreased viral loads and increased survival along with expanded T cells 
with increased T-cell functionality.   

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2.4.1 Nonclinical Development of Nivolumab 
 
In intravenous (IV) repeat-dose toxicology studies in cynomolgus monkeys, nivolumab 
alone was well tolerated (Investigator Brochure, 2013).  Combination studies have 
highlighted the potential for toxicity when combined with ipilimumab, MDX-1408, and 
BMS-986016.  Nivolumab bound specifically to PD-1 (and not to related members of the 
CD28 family such as CD28, ICOS, CTLA-4, and BTLA) with a Kd = 3.06 nM.  A 
surrogate rat anti-mouse PD-1 antibody (4H2) was derived and expressed as chimeric 
IgG1 murine antibody.  Antitumor activity was seen for several tumor models, including 
colon carcinoma and fibrosarcoma. 
 
2.4.2 Clinical Development of Nivolumab 
 
Nivolumab is being evaluated as monotherapy and in combination with cytotoxic 
chemotherapy, other immunotherapy (such as ipilimumab), anti-angiogenesis therapy, 
and targeted therapies in completed and ongoing BMS-sponsored clinical trials in 
NSCLC, melanoma, RCC, hepatocellular carcinoma (HCC), gastrointestinal (GI) 
malignancies including microsatellite instability (MSI) in colorectal cancer, and triple-
negative breast cancer (TNBC) with an expanding group of indications (Investigator 
Brochure, 2013).  In addition, two investigator-sponsored trials (ISTs) of nivolumab in 
combination with a peptide vaccine in melanoma are being conducted in the adjuvant 
setting and advanced disease.  
 
Seven nivolumab studies were conducted in Japan, including six studies in advanced 
solid tumors and recurrent or unresectable stage III/IV melanoma sponsored by Ono 
Pharmaceuticals Co. Ltd., and one IST in recurrent or advanced platinum-refractory 
ovarian cancer.   
 
2.4.3 Pharmacokinetics 
 
Pharmacokinetics (PK) of nivolumab was linear in the range of 0.3 to 10 mg/kg, with 
dose-proportional increases in maximum serum concentration (Cmax) and area under the 
concentration-time curve from time zero to infinity (AUC0-∞), with low to moderate inter-
subject variability observed at each dose level (Investigator Brochure, 2013).  Clearance 
of nivolumab is independent of dose in the dose range (0.1 to 10 mg/kg) and tumor types 
studied.  Body weight normalized dosing showed approximately constant trough 
concentrations over a wide range of body weights.  The mean terminal elimination half-
life of BMS-936558 is 17 to 25 days consistent with the half-life of endogenous IgG4.   
 
2.4.4 Efficacy 
 
In a phase 1 (1, 3, and 10 mg/kg nivolumab doses) dose-escalation study the 3 mg/kg 
dose was chosen for expanded cohorts. Among 236 patients, objective responses (ORs) 
(complete or partial responses [CR or PR]) were seen in NSCLC, melanoma, and RCC.  
ORs were observed at all doses (Sznol et al., 2013).  Median OS was 16.8 months across 
doses and 20.3 months at the 3 mg/kg dose.  Median OS across all dose cohorts was 9.2 

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months and 9.6 months for squamous and non-squamous NSCLC, respectively (Brahmer 
et al., 2013).  In the RCC cohort, median duration of response was 12.9 months for both 
doses with 5 of the 10 responses lasting ≥1 year (Drake et al., 2013).   
 
In an advanced melanoma phase 1 study, nivolumab and ipilimumab were administered 
IV every 3 weeks for 4 doses followed by nivolumab alone every 3 weeks for 4 doses 
(concurrent regimen) (Wolchok et al., 2013).  The combined treatment was subsequently 
administered every 12 weeks for up to 8 doses.  In a sequenced regimen, patients 
previously treated with ipilimumab received nivolumab every 2 weeks for up to 48 doses.  
In the concurrent regimen (53 patients), 53% of patients had an OR at doses 1 mg/kg 
nivolumab and 3 mg/kg ipilimumab, with tumor reduction of 80% or more (modified 
World Health Organization [mWHO] criteria).  In the sequenced-regimen (33 patients), 
the objective response rate (ORR) was 20%. 
 
In a phase 1 study of nivolumab plus platinum-based doublet chemotherapy (PT-doublet) 
in chemotherapy-naïve NSCLC patients, 43 patients were treated with nivolumab + PT-
doublet (Rizvi et al., 2013).  No dose-limiting toxicities (DLTs) were reported and 
total/confirmed ORRs were 43/33%, 40/33%, and 31/31% in 
nivolumab/gemcitabine/cisplatin, nivolumab/pemetrexed/cisplatin, and 
nivolumab/carboplatin/paclitaxel arms, respectively. 
 
2.4.5 Toxicology 
 
A maximum tolerated dose (MTD) of nivolumab was not defined (Topalian et al., 2012).  
Serious adverse events (SAEs) occurred in 32 of 296 patients (11%) similar to the 
immune-related inflammatory events seen with ipilimumab:  pneumonitis, vitiligo, 
colitis, hepatitis, hypophysitis, and thyroiditis (with noted pulmonary toxicity resulting in 
3 deaths. Renal failure, symptomatic pancreatic and DM, neurologic events, and 
vasculitis have also been reported.).   In combination with ipilimumab in the concurrent-
regimen group (Wolchok et al., 2013), grade 3 or 4 treatment-related events were noted 
in 53% of patients.  Skin rash represents the majority of these events.  
 
The possible side effects of Nivolumab are summarized below (included in the 
Investigator Brochure).  
 
The following are the key safety findings for the subjects with advanced malignancies 
mainly solid tumors.  
 
• Drug-related AEs (any grade) were reported in 75.2%, and drug-related Grade 3-4 
AEs were reported in 17.0% of subjects. 
- The most frequently reported drug-related AE was fatigue (28.1%). Other drug-
related AEs Grade 3-4 reported in more than 2 subjects were pneumonitis (1.3%), 
lymphopenia (1.3%), diarrhea (1.0%), abdominal pain (1.0%), CD4 lymphocytes 
decreased (1.0%), and hypophosphatemia (1.0%). 
 
• The most frequently reported drug-related SAE was pneumonitis (7 subjects, 2.3%). 

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Drug-related Grade 3-4 pneumonitis was reported in 4 (1.3%). 
 
• The most frequently reported drug-related select AE categories (any grade) were skin 
(24.5%), GI (14.1%), and endocrine (9.5%). 
- AEs belonging to the pulmonary and renal select AE categories were unexpected, 
drug-related toxicities associated with the use of nivolumab. 
- AEs belonging to select AE categories were generally manageable and reversible 
with the use of immunosuppressants. 
 
Below is the safety experience from the single agent nivolumab Phase 1 trial in patients 
with relapsed or refractory hematologic malignancies (included in the Investigator 
Brochure). 
  
• The most frequently reported drug-related AEs (> 10% of subjects) were fatigue 
(12.6%) and rash (10.7%). The majority were Grade 1-2 in severity. 
• The most frequently reported drug-related SAE was pneumonitis (4.9%). 
• AEs leading to discontinuation were reported for 15.5% of subjects. The most 
frequently reported AE leading to discontinuation were pneumonitis and nodule (1.9%, 
each). 
• Thirteen subjects have died, with 1 death reported due to study drug toxicity (1 
subject in the 3 mg/kg treatment group died due to Grade 5 pneumonitis with onset 10 
days after the subject received the only dose of nivolumab). 
 
It is very important to monitor AEs for the patients on the Nivolumab treatment, since 
getting medical treatment right away may keep these problems from becoming more 
serious. The local investigators or treating physicians need to be aware the main AEs, and 
should not hesitate to contact the study chair for any question and management. The 
management of AEs will be further discussed in Section 5 (Treatment Plan, especially 
section 5.6: Criteria to resume treatment) and Section 6: Dose delay/Dose modification.   
  
2.4.6 Pharmacodynamics/Biomarkers 
 
Tumor-cell expression (melanoma) of PD-L1 was characterized in combination with 
ipilimumab with the use of IHC staining and pharmacodynamics changes in the 
peripheral-blood absolute lymphocyte count (Wolchok et al., 2013).  With PD-L1 
positivity defined as expression in at least 5% of tumor cells, biopsy specimens from 21 
of 56 patients (38%) were PD-L1–positive.  Among patients treated with the concurrent 
regimen of nivolumab and ipilimumab, ORs were observed in patients with either PD-
L1–positive tumor samples (6 of 13 patients) or PD-L1–negative tumor samples (9 of 
22).  In the sequenced regimen cohorts, a higher number of overall responses was seen 
among patients with PD-L1–positive tumor samples (4 of 8 patients) than among patients 
with PD-L1–negative tumor samples (1 of 13) suggesting the possibility that these tumors 
have higher response rates to the combination. The relationship between PDL-1 
expression and responses may not be present in patients treated with the combination.  
Tissue expression of PDL-2, interferon-γ (IFN- γ), IDO, and T cell CD8+ are of current 
interest.  Until more reliable data based on standardized procedures for tissue collection 

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and assays are available, PD-L1 status cannot be used to select patients for treatment at 
this time.   
 
The combination of ipilimumab with nivolumab has been reported to result in improved 
responses in advanced melanoma marked by time to response, number of responses, 
depth and duration of responses,  PFS, and OS compared to single agent ipilimumab 
(Wolchok et al., 2013).  
 
For RCC results have been reported (Hammers et al., 2014). 
 
The combination is being evaluated in other disease settings typically with 3mg/kg 
nivolumab and 1mg/kg ipilimumab q 3 weeks x 4 induction doses.  
 
See Section 8.1.2 for drug information.  
 
2.5 
Rationale to utilize the PD-1-blocking antibody, Nivolumab, to eliminate MRD and 
maintain remission in AML patients after induction/consolidation chemotherapy 
Based on our exciting pre-clinical results of PD-1/PD-L1 blockade (Zhang, Gajewski et al. 
2009), along with our data demonstrating that PD-L1 is expressed on AML cells in a subset of 
patients, a logical strategy to eliminate MRD in AML is to enhance the host anti-leukemia 
immune response by targeting PD-1 (Nivolumab). Thus, we propose a multi-center randomized 
phase II study to assess the efficacy of Nivolumab as maintenance therapy for AML patients who 
are not candidates for stem cell transplantation. Nivolumab will be administered IV at 3mg/kg 
every 2 weeks for forty-six doses.  The duration of Nivolumab treatment will be 2 years 
following randomization based on the experience in melanoma patients (Wolchok, Kluger et al.). 
Patients will be followed for disease relapse, adverse events, non-relapse mortality and survival. 
The primary objective will be progression free survival at 2 years. Secondary objectives will be 
OS, and evaluation of adverse events following Nivolumab administration. Exploratory 
objectives will include: determination of PD-LI expression on CD34+ and CD34- cells within the 
bone marrow as predictive biomarker; eradication of MRD assessed by quantitative RT-PCR of 
WT-1 during the Nivolumab treatment; immune responses including WT1 specific CTLs, Treg 
changes; and TCR repertoire sequencing to determine the TCR clonal evolution during 
Nivolumab treatment. 
 
2.6 
Correlative Studies Background 
 
2.6.1 WT1 as a marker of minimal residual disease in AML 
 
Wilm’s tumor 1 (WT1) is a zinc finger transcription factor that is normally expressed in tissues 
of mesodermal origin during embryogenesis. WT1 is over-expressed in various hematological 
malignancies, including AML (80-100% of cases).  WT1 mRNA levels in the peripheral blood 
and bone marrow correlate with disease status during and after treatment, and elevation of WT1 
after chemotherapy and stem cell transplant predicts clinical relapse. Thus, WT1 RT-PCR is now 
being used to monitor minimal residual disease (MRD) during treatment of AML (Reviewed in 
(Grimwade, Vyas et al. 2010; Dominietto 2011)).  
 

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Dr. Wendy Stock’s lab assessed MRD for a GVAX vaccine study in patients with AML using 
quantitative RT-PCR detection of WT1 expression. Although > 90% of patients who achieved 
clinical complete remission (CR) had significant declines in WT1 in both blood and bone 
marrow, 95% of patients had persistently detectable WT1 levels in bone marrow demonstrating 
clear MRD in AML patients in CR (Figure 3 in (Borrello, Levitsky et al. 2009)). Importantly, the 
achievement of a significant decrease or elimination of WT1 transcripts during the course of 
vaccination correlated with significant superior relapse-free survival (table 3 in (Borrello, 
Levitsky et al. 2009)). Furthermore, patients who relapsed during or following vaccination had a 
significant increase in WT1 expression, while patients in continuous CR maintained very low or 
undetectable WT1 levels (Figure 4 in (Borrello, Levitsky et al. 2009)).  
 
WT1 MRD monitoring appears to correlate well with remission status in conventionally-treated 
AML patients as well.  Among AML patients in clinical CR after induction and consolidation 
therapy, 80-90% and 30-40% of patients continue to have detectable WT1 transcript in the 
marrow or blood, respectively (Miyawaki, Hatsumi et al.). WT1 levels in the peripheral blood 
can predict relapse after CR, and its levels after consolidation therapy are closely correlated with 
disease free survival (DFS), OS, and early relapse. The DFS rate at 3 years was 20% and 50% 
for WT1 MRD positive and negative patients, respectively (Miyawaki, Hatsumi et al.). Thus, 
WT1 monitoring is a useful tool not only for predicting relapse in AML patients in clinical CR, 
but also for monitoring response to post-remission immunotherapeutic strategies, such as 
vaccination and possibly after post-remission treatment with PD-1 blocking antibodies.   
 
2.6.2 TCR sequencing during immunotherapy 
Recent advances in deep sequencing technology make it possible to characterize the T cell 
receptor (TCR) repertoire generated following immunotherapy, such as WT1 peptide vaccination 
(i.e. the “clonality” of the response). Preliminary studies have demonstrated biased usage of 
TCR-V gene families in WT1 peptide vaccinated patients (Ochsenreither, Fusi et al. ; 
Ochsenreither, Fusi et al. ; Tanaka-Harada, Kawakami et al.), which was confirmed by our own 
TCR sequencing data in a patient who received WT1 peptide vaccination post SCT (Figure 4 in 
preliminary result section).  Dr. Yusuke Nakamura at our institution is one of the pioneers in the 
assessment of genetic variations (VNTR and SNP markers) and whole-genome analysis in cancer 
cells. Using a novel sequencing approach, we have been working together to understand the TCR 
“landscape” during immune-based therapies for AML. TCR sequencing of peripheral blood 
and/or bone marrow samples during immunotherapy, such as PD-1 blockade, will allow us to 
assess for an emerging T cell repertoire in patients who respond versus those who do not. This 
type of analysis may also be useful to identify high-affinity TCRs for use in down-stream 
adaptive T cell therapy approaches for AML, as well as to enable the identification of new AML 
associated antigens. 
 
 
3. PATIENT SELECTION 
 
3.1 
Eligibility Criteria  
 
 

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3.1.1 AML patients in first CR (CR1) or first CRi after induction and/or consolidation 
chemotherapy; except young (<60 years) AML patients in European LeukemiaNet 
favorable group. (Since young AML patients in the European LeukemiaNet favorable 
group have excellent 2 year PFS at around 64%, further maintenance therapy might not 
provide additional benefit; thus the current trial will exclude young favorable group AML 
patients (Mrozek, Marcucci et al.). Patients could receive any cycle consolidation or no 
consolidation per the discretion by the treating physician.  
 
3.1.2 Within 60 days after bone marrow biopsy confirmed remission after the patients recover 
from their last course of chemotherapy, the goal to consent the eligible patient prior to the 
remission confirmation bone marrow biopsy at the end of the planned chemotherapy. 
Ideally, the research samples will be collected during the bone marrow biopsy, and the 
patient will be enrolled to the study within 2 weeks of the bone marrow biopsy. 
  
If there is delay to enroll the patient after the bone marrow biopsy and research sample 
collection, it is ok not to repeat bone marrow biopsy within 4 weeks, after the last bone 
marrow biopsy, if there is no sign of disease relapse.  
 
A repeat bone marrow biopsy should be done if the delay of enrollment is more than 4 
weeks after the last bone marrow biopsy.  
 
Patients with confirmed remission within 60 days after the last bone marrow biopsy, 
without research samples collection, should have a repeat bone marrow biopsy conducted 
within two weeks prior to enrolling on the study. 
 
3.1.3 Patient is not a candidate for stem cell transplant due to advanced age or co-morbidities; 
or the enrollee does not have donor available; or the enrollee declines stem cell transplant 
due to personal belief; or stem cell transplant is not standard of care based on the risk 
category of disease 
 
3.1.4 Age ≥18 years.  Because no dosing or adverse event data are currently available on the 
use of nivolumab in patients <18 years of age, children are excluded from this study, but 
will be eligible for future pediatric trials. 
 
3.1.5 ECOG/Karnofsky performance status of 0 or 1 (Karnofsky ≥70%, see Appendix A). 
 
3.1.6 Life expectancy of greater than 6 months 
 
3.1.7 Patients must have acceptable organ and marrow function as defined below: 
 
 
− 
leukocytes 
≥1,500/mcL 
− 
absolute neutrophil count 
≥1,000/mcL 
− 
platelets 
≥50,000/mcL or recovery to the baseline count 
− 
total bilirubin 
≤1.5× institutional upper limit of normal (ULN) 
(except patients with Gilbert Syndrome, who can have total bilirubin <3.0 mg/dL) 

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− 
AST(SGOT)/ALT(SGPT) 
≤2.5× ULN  
− 
Amylase and lipase  
≤1.5× ULN without any symptoms of pancreatitis 
− 
 
− 
Serum creatinine 
≤1.5× ULN  
OR 
− 
creatinine clearance (CrCl) 
≥50 mL/min (if using the Cockcroft-Gault formula 
below): 
Female CrCl = (140 - age in years) x weight in kg x 0.85 
72 x serum creatinine in mg/dL 
Male CrCl = (140 - age in years) x weight in kg x 1.00  
72 x serum creatinine in mg/dL  
 
 
 
3.1.8 The effects of nivolumab on the developing human fetus are unknown.  For this reason, 
women of child-bearing potential (WOCBP) and men must agree to use adequate 
contraception (hormonal or barrier method of birth control; abstinence) prior to study 
entry and for the duration of study participation.  WOCBP should use an adequate 
method to avoid pregnancy for 23 weeks) after the last dose of investigational drug 
Nivolumab.  Women of childbearing potential must have a negative serum or urine 
pregnancy test (minimum sensitivity 25 IU/L or equivalent units of HCG) within 24 
hours prior to the start of nivolumab.  Women must not be breastfeeding.  Men who are 
sexually active with WOCBP must use any contraceptive method with a failure rate of 
less than 1% per year.  Men receiving nivolumab and who are sexually active with 
WOCBP will be instructed to adhere to contraception for a period of 31 weeks after the 
last dose of investigational product.  Women who are not of childbearing potential (i.e., 
who are postmenopausal or surgically sterile as well as azoospermic men) do not require 
contraception.   
 
Women of childbearing potential (WOCBP) is defined as any female who has 
experienced menarche and who has not undergone surgical sterilization (hysterectomy or 
bilateral oophorectomy) or who is not postmenopausal.  Menopause is defined clinically 
as 12 months of amenorrhea in a woman over 45 in the absence of other biological or 
physiological causes.  In addition, women under the age of 55 must have a documented 
serum follicle stimulating hormone (FSH) level less than 40 mIU/mL.   
 
WOCBP receiving nivolumab will be instructed to adhere to contraception for a period of 
23 weeks after the last dose of investigational product.  Men receiving nivolumab and 
who are sexually active with WOCBP will be instructed to adhere to contraception for a 
period of 31 weeks after the last dose of investigational product.  These durations have 
been calculated using the upper limit of the half-life for nivolumab (25 days) and are 
based on the protocol requirement that WOCBP use contraception for 5 half-lives plus 30 
days and men who are sexually active with WOCBP use contraception for 5 half-lives 
plus 90 days. 
 
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participating in this study, she (or the participating partner) should inform the treating 
physician immediately. 
 
 
3.1.9 Ability to understand and the willingness to sign a written informed consent document. 
 
3.2 
Exclusion Criteria 
 
3.2.1 Patients who have had chemotherapy or radiotherapy within 4 weeks (6 weeks for 
nitrosoureas or mitomycin C) prior to entering the study or those who have not recovered 
from adverse events (AEs) due to agents administered more than 4 weeks earlier.   
 
3.2.2 Patients who are receiving any other investigational agents. 
 
3.2.3 Patients should be excluded if they have had prior treatment with an anti-PD-1, anti-PD-
L1, anti-PD-L2, anti-CTLA-4 antibody, or any other antibody or drug specifically 
targeting T-cell co-stimulation or immune checkpoint pathways 
 
3.2.4 Patients with known CNS involvement may be excluded because of poor prognosis and 
concerns regarding progressive neurologic dysfunction that would confound the 
evaluation of neurologic and other adverse events.  However, if CNS disease is cleared 
before the treatment with Nivolumab, patients could be allowed if no permanent CNS 
damage.  
 
3.2.5 History of severe hypersensitivity reaction to any monoclonal antibody. 
 
3.2.6 Uncontrolled intercurrent illness including, but not limited to, ongoing or active infection, 
symptomatic congestive heart failure, unstable angina pectoris, cardiac arrhythmia, or 
psychiatric illness/social situations that would limit compliance with study requirements. 
 
3.2.7 Pregnant women are excluded from this study because Nivolumab is an agent with the 
potential for teratogenic or abortifacient effects.  Because there is an unknown but 
potential risk for adverse events in nursing infants secondary to treatment of the mother 
with Nivolumab, breastfeeding should be discontinued if the mother is treated with 
Nivolumab.   
 
3.2.8 Patients with known history of testing positive for human immunodeficiency virus (HIV) 
or known acquired immunodeficiency syndrome (AIDS) might be enrolled if the viral 
load by PCR is undetectable with/without active treatment and absolute lymphocyte 
count >= 350/ul.  
 
Patients with a positive test for hepatitis B virus surface antigen (HBV sAg) or hepatitis 
C virus ribonucleic acid (HCV antibody) indicating acute or chronic infection might be 
enrolled if the viral load by PCR is undetectable with/without active treatment. 
 

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3.2.9 Patients with active autoimmune disease or history of autoimmune disease that might 
recur, which may affect vital organ function or require immune suppressive treatment 
including systemic corticosteroids, should be excluded.  These include but are not limited 
to patients with a history of immune related neurologic disease,  multiple sclerosis, 
autoimmune (demyelinating)  neuropathy, Guillain-Barre syndrome, myasthenia gravis; 
systemic autoimmune disease such as SLE, connective tissue diseases, scleroderma, 
inflammatory bowel disease (IBD), Crohn’s, ulcerative colitis, hepatitis; and patients 
with a history of toxic epidermal necrolysis (TEN), Stevens-Johnson syndrome, or 
phospholipid syndrome should be excluded because of the risk of recurrence or 
exacerbation of disease.  Patients with vitiligo, endocrine deficiencies including 
thyroiditis managed with replacement hormones including physiologic corticosteroids are 
eligible.  Patients with rheumatoid arthritis and other arthropathies, Sjögren’s syndrome 
and psoriasis controlled with topical medication and patients with positive serology, such 
as antinuclear antibodies (ANA), anti-thyroid antibodies should be evaluated for the 
presence of target organ involvement and potential need for systemic treatment but 
should otherwise be eligible. 
 
3.2.10 Patients are permitted to enroll if they have vitiligo, type I diabetes mellitus, residual 
hypothyroidism due to autoimmune condition only requiring hormone replacement, 
psoriasis not requiring systemic treatment, or conditions not expected to recur in the 
absence of an external trigger (precipitating event). 
 
3.2.11 Patients should be excluded if they have a condition requiring systemic treatment with 
either corticosteroids (>10 mg daily prednisone equivalents) or other immunosuppressive 
medications within 14 days of study drug administration.  Inhaled or topical steroids and 
adrenal replacement doses <10 mg daily prednisone equivalents are permitted in the 
absence of active autoimmune disease.  Patients are permitted to use topical, ocular, intra-
articular, intranasal, and inhalational corticosteroids (with minimal systemic absorption).  
Physiologic replacement doses of systemic corticosteroids are permitted, even if <10 
mg/day prednisone equivalents.  A brief course of corticosteroids for prophylaxis (e.g., 
contrast dye allergy) or for treatment of non-autoimmune conditions (e.g., delayed-type 
hypersensitivity reaction caused by contact allergen) is permitted. 
 
3.2.12 Patients who have had evidence of active or acute diverticulitis, intra-abdominal abscess, 
GI obstruction and abdominal carcinomatosis which are known risk factors for bowel 
perforation should be evaluated for the potential need for additional treatment before 
coming on study. 
 
 
3.3 
Inclusion of Women and Minorities 
 
NIH policy requires that women and members of minority groups and their subpopulations be 
included in all NIH-supported biomedical and behavioral research projects involving NIH-
defined clinical research unless a clear and compelling rationale and justification establishes to 
the satisfaction of the funding Institute & Center (IC) Director that inclusion is inappropriate 
with respect to the health of the subjects or the purpose of the research.  Exclusion under other 

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circumstances must be designated by the Director, NIH, upon the recommendation of an IC 
Director based on a compelling rationale and justification.  Cost is not an acceptable reason for 
exclusion except when the study would duplicate data from other sources.  Women of 
childbearing potential should not be routinely excluded from participation in clinical research.  
Please see http://grants.nih.gov/grants/funding/phs398/phs398.pdf. 
 
We will try to enroll equal number of patients from different genders (the final ratio of male / 
female should be very close since there is significant difference of AML in different gender). We 
will encourage the enrollment of patients from different race and ethnicity without any 
discrimination, but more Caucasians still might be enrolled to the study. Please see the detail in 
Planned Enrollment Report under Section 13.2. 
 
4. REGISTRATION PROCEDURES (ROSTERED PROTOCOL MODEL) 
 
4.1 
Investigator and Research Associate Registration with CTEP 
 
4.1.1 CTEP Registration Procedures 
 
Food and Drug Administration (FDA) regulations and National Cancer Institute (NCI) 
policy require all investigators participating in any NCI-sponsored clinical trial to register 
and to renew their registration annually. 
 
Registration requires the submission of: 
 
• a completed Statement of Investigator Form (FDA Form 1572) with an original 
signature 
• a current Curriculum Vitae (CV) 
• a completed and signed Supplemental Investigator Data Form (IDF) 
• a completed Financial Disclosure Form (FDF) with an original signature 
 
Fillable PDF forms and additional information can be found on the CTEP website at 
http://ctep.cancer.gov/investigatorResources/investigator_registration.htm 
 
4.1.2 CTEP Associate Registration Procedures / CTEP-IAM Account 
The Cancer Therapy Evaluation Program (CTEP) Identity and Access Management 
(IAM) application is a web-based application intended for use by both Investigators (i.e., 
all physicians involved in the conduct of NCI-sponsored clinical trials) and Associates 
(i.e., all staff involved in the conduct of NCI-sponsored clinical trials). 
 
Associates will use the CTEP-IAM application to register (both initial registration and 
annual re-registration) with CTEP and to obtain a user account. 
 
Investigators will use the CTEP-IAM application to obtain a user account only. (See 
CTEP Investigator Registration Procedures above for information on registering with 
CTEP as an Investigator, which must be completed before a CTEP-IAM account can be 
requested.) 

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An active CTEP-IAM user account is needed to access all CTEP and CTSU (Cancer 
Trials Support Unit) websites and applications, and is critical to the conduct of this study, 
including document access, patient enrollment, and clinical data submission. 
 
Additional information can be found on the CTEP website at 
http://ctep.cancer.gov/branches/pmb/associate_registration.htm. 
 
4.1.3 For Questions and Support 
 
For questions about Investigator Registration, please contact the CTEP Investigator 
Registration Help Desk: pmbregpend@ctep.nci.nih.gov. 
 
For questions about Associate Registration or CTEP-IAM Account Creation, please 
contact the CTEP Registration Help Desk: ctepreghelp@ctep.nci.nih.gov. 
 
4.2 
Site Registration 
 
This study is supported by the NCI Cancer Trials Support Unit (CTSU). 
 
Each investigator or group of investigators at a clinical site must obtain Institutional Review 
Board (IRB) approval for this protocol and submit all required regulatory documents (including 
any protocol specific documents) to the CTSU Regulatory Office before they can be approved to 
enroll patients. 
 
The CTSU Regulatory Office tracks receipt of these documents in the CTSU Regulatory Support 
System (RSS), reviews for compliance, and transmits site approval data to CTEP. 
 
Sites participating on the NCI CIRB initiative and accepting CIRB approval for the study are not 
required to submit separate IRB approval documentation to the CTSU Regulatory Office for 
initial, continuing, or amendment review.  However, sites must submit a Study Specific 
Worksheet for Local Context (SSW) to the CIRB (via IRBManager) to indicate their intention to 
open the study locally. The CIRB’s approval of the SSW is then communicated to the CTSU 
Regulatory Office for compliance in the RSS. The Signatory Institution must inform the CTSU 
which CIRB approved institutions aligned with the Signatory Institution are participating in the 
study so that the study approval can be applied to those institutions. Other site registration 
requirements (e.g., laboratory certifications, protocol-specific training certifications, or modality 
credentialing) must be submitted to the CTSU Regulatory Office or compliance communicated 
per protocol instructions. 
 
4.2.1 Downloading Regulatory Documents 
 
Site registration forms may be downloaded from the 9706 protocol page located on the 
CTSU Web site.  Permission to view and download this protocol is restricted and is based 
on person and site roster data housed in the CTSU RSS.  To participate, Investigators and 
Associates must be associated with the Corresponding or Participating protocol 

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organization in the RSS.  
 
• Go to https://www.ctsu.org and log in using your CTEP IAM username and 
password 
• Click on the Protocols tab in the upper left of your screen  
• Either enter the protocol # in the search field at the top of the protocol tree, or 
• Click on the By Lead Organization folder to expand, then select EDDOP-IL-057, 
and protocol #9706 
• Click on LPO Documents, select the Site Registration documents link, and 
download and complete the forms provided.  (Note: For sites under the CIRB 
initiative, IRB data will automatically load to RSS.) 
 
4.2.2 Requirements For 9706 Site Registration: 
 
• CTSU IRB Certification (for sites not participating via the NCI CIRB) 
• CTSU IRB/Regulatory Approval Transmittal Sheet (for sites not participating via 
the NCI CIRB)  
• Local informed consent document 
 
Submit required forms and documents to the CTSU Regulatory Office, where it will be 
entered and tracked in the CTSU RSS.   
CTSU Regulatory Office 
1818 Market Street, Suite 1100 
Philadelphia, PA 19103 
Phone:  1-866-651-2878 
Fax:  215-569-0206 
E-mail:  CTSURegulatory@ctsu.coccg.org (for regulatory document submission only) 
 
4.2.3 Checking Site Registration Status 
 
Sites can check the status of their registration packets by querying the Site Registration 
subtab of the members’ section of the CTSU Web site.  (Note: Sites will not receive 
formal notification of regulatory approval from the CTSU Regulatory Office.) 
• Go to https://www.ctsu.org and log in using your CTEP IAM username and 
password. 
• Click on the Regulatory tab at the top of your screen. 
• Click on the Site Registration subtab. 
• Enter your 5-character CTEP Institution Code and click on Go. 
Note: If possible, please allow three working days for site registration approval before 
attempting to enroll your first patient. 
 
4.3 
Patient Registration* 
*Please notify the University of Chicago general e-mail at 
phaseIIcra@medicine.bsd.uchicago.edu when you have a potential patient that you would 
like to screen for the study and to obtain a pre-registration ID for screening BMBX 

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samples. 
 
 
 
4.3.1 OPEN / IWRS 
 
Patient enrollment will be facilitated using the Oncology Patient Enrollment Network 
(OPEN).  OPEN is a web-based registration system available to users on a 24/7 basis.  It 
is integrated with the CTSU Enterprise System for regulatory and roster data interchange 
and with the Theradex Interactive Web Response System (IWRS) for retrieval of patient 
registration/randomization assignment.  Patient enrollment data entered by Registrars in 
OPEN / IWRS will automatically transfer to the NCI’s clinical data management system, 
Medidata Rave. 
 
 
For this protocol, sites must first contact the lead institution (University of Chicago) prior 
to registering the patient in OPEN/IWRS so that the patient can be randomized. They 
should send an e-mail to the UC Data Manager at phaseIIcra@medicine.bsd.uchicago.edu 
containing the following information: 
 
Clinical site name 
Patient’s first and last initials 
Date of birth 
Age group:  <60  or  >=60 years 
AML risk category:   Adverse, Intermediate-1, Intermediate-2, or Favorable 
 
The UC Data Manager will obtain the randomized treatment assignment from REDCap. The 
UC Data Manager will then login to OPEN/IWRS, enter the assigned treatment, and reserve a 
slot for the patient.  He/she will then notify the clinical site that a slot has been reserved, and 
the site will access OPEN/IWRS and complete registration of the patient. 
 
The OPEN system will provide the site with a printable confirmation of registration and 
the treatment assigned to the patient (Nivolumab or Observation).  Please print this 
confirmation for your records.  
 
4.3.2 OPEN/IWRS User Requirements 
 
OPEN/IWRS users must meet the following requirements: 
• Have a valid CTEP-IAM account (i.e., CTEP username and password). 
• To enroll patients or request slot reservations:  Be on an ETCTN Corresponding 
or Participating Organization roster with the role of Registrar. 
• To approve slot reservations or access cohort management:  Be identified to 
Theradex as the “Client Admin” for the study.  
• Have regulatory approval for the conduct of the study at their site. 
 
Prior to accessing OPEN/IWRS, site staff should verify the following: 

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• All eligibility criteria have been met within the protocol stated timeframes. Site 
staff should use the registration forms provided on the CTSU web site as a tool to 
verify eligibility. 
• If applicable, all patients have signed an appropriate consent form and HIPAA 
authorization form. 
 
4.3.3 OPEN/IWRS Questions? 
 
Further instructional information on OPEN is provided on the OPEN tab of the CTSU 
website at https://www.ctsu.org or at https://open.ctsu.org.  For any additional questions 
contact the CTSU Help Desk at 1-888-823-5923 or ctsucontact@westat.com.  
 
Theradex has developed a Slot Reservations and Cohort Management User Guide, which 
is available on the Theradex website:  http://theradex.com/CTMS/Downloads.aspx. This 
link to the Theradex website is also on the CTSU website OPEN tab. For questions about 
the use of IWRS for slot reservations, contact the Theradex Helpdesk: 609-619-7802 or 
Theradex main number 609-799-7580; CTMSSupport@theradex.com. 
 
4.4 
General Guidelines 
 
Following registration, patients should begin protocol treatment within 14 days. Issues that 
would cause treatment delays should be discussed with the lead site Principal Investigator.  If a 
patient does not receive protocol therapy following registration, the patient’s registration on the 
study may be canceled.  The UC Study Coordinator should be notified of cancellations as soon 
as possible at: phaseIIcra@medicine.bsd.uchicago.edu 
 
5. TREATMENT PLAN 
 
5.1 
Agent Administration 
 
Treatment will be administered on an outpatient basis.  Reported adverse events and potential 
risks are described in Section 7.  Appropriate dose modifications are described in Section 6.  No 
investigational or commercial agents or therapies other than those described below may be 
administered with the intent to treat the patient's malignancy. 
 
 
5.1.1 Nivolumab 
 
Nivolumab will be given every two weeks (±2 days) (two weeks is equivalent to one 
cycle) at a dose of 3 mg/kg.  Patients may be dosed no less than 12 days from the 
previous dose of drug.  
 
The dosing calculations should be based on the actual body weight.  If the patient’s 
weight on the day of dosing differs by >10% from the weight used to calculate the 
original dose, the dose must be recalculated.  All doses should be rounded to the nearest 
milligram if dose rounding is allowed as per institutional guidelines.  There will be no 

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dose modifications allowed.   
 
Nivolumab is to be administered as a 30-minute (+/- 10 minutes) IV infusion, using a 
volumetric pump with a  0.2-1.2 micron in-line filter at the protocol-specified dose.  The 
drug can be diluted with 0.9% normal saline for delivery but the total drug concentration 
of the solution cannot be below 1 mg/mL.  It is not to be administered as an IV push or 
bolus injection.  At the end of the infusion, flush the line with a sufficient quantity of 
normal saline.   
 
 
5.2 
General Concomitant Medication and Supportive Care Guidelines 
 
There is no clear evidence to demonstrate a potential for interaction of Nivolumab with other 
concomitantly administered drugs through the cytochrome P450 system. 
 
5.3 
Duration of Therapy 
 
In the absence of treatment delays due to adverse event(s), treatment may continue for 46 cycles 
(92 weeks) or until one of the following criteria applies: 
 
• 
Disease progression, 
 
• 
Intercurrent illness that prevents further administration of treatment, 
 
• 
Unacceptable adverse event(s) which include the following (see also section 6 and 
specific algorithms in Appendix H): 
 
o Any grade 4 events. 
 
o Grade 3 drug-related autoimmune or inflammatory events including uveitis, 
pneumonitis, diarrhea, colitis, neurologic adverse events, hypersensitivity 
reaction, or infusion reaction of any duration requires discontinuation except as 
noted below: 
: 
o Any other grade 3 non-skin, drug-related AE lasting >7 days including fatigue. 
 
o Any grade 3 or 4 drug-related laboratory abnormality or electrolyte abnormality, 
not associated with underlying organ pathology, that does not require treatment 
except for electrolyte replacements does not require treatment discontinuation.  
 
o Grade 3 or 4 amylase or lipase abnormalities that are not associated with diabetes 
mellitus (DM), associated liver or gall bladder inflammation clinical 
manifestations of pancreatitis and which decrease to < Grade 2 within 1 week of 
onset may resume study treatment when resolved.   
 
o Any grade 2 drug-related uveitis or eye pain or blurred vision that does not 

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respond to topical therapy and does not improve to Grade 1 severity within the re-
treatment period OR requires systemic treatment 
 
o Patients requiring > two dose delays for the same type of event should go off 
protocol therapy. 
 
o Any adverse event, laboratory abnormality, or intercurrent illness which, in the 
judgment of the investigator, presents a substantial clinical risk to the subject with 
continued study drug dosing  
 
o Any dosing interruption lasting >6 weeks, with the following exceptions:   
 
Patients being tapered after high dose corticosteroids over one month 
followed by a two-week observation period will be allowed an additional 
two weeks to restart treatment ( a maximum  eight week interruption).  
Dosing interruptions >6 weeks that occur for non-drug-related reasons 
may be allowed if approved by the Investigator.  Prior to re-initiating 
treatment in a subject with a dosing interruption lasting >6 weeks, the 
Principal Investigator must be consulted. 
 
o 
Tumor assessments should continue as per protocol even if dosing is interrupted.  
 
o 
Any patients who require additional immune suppressive treatment beyond 
steroids should go off study treatment 
 
 
• 
Patient decides to withdraw from the study, or 
 
• 
General or specific changes in the patient's condition render the patient unacceptable 
for further treatment in the judgment of the investigator. 
 
5.4 
Duration of Follow Up 
 
Patients will be followed for up to two years from the enrollment after removal from study or 
until death, whichever occurs first.  Patients removed from study for unacceptable adverse 
event(s) will be followed until resolution or stabilization of the adverse event. Patients who have 
come off treatment/observation after two years on study without relapse will continue to be 
followed every 6 months for the first year and yearly until disease relapse or death (whichever 
occurs first). 
 
5.5 
Criteria for Removal from Study 
 
Patients will be removed from study when any of the applicable criteria, including relapse of 
disease, adverse events, patient withdrawal or inability to follow study protocol as listed in 
Section 5.4.  The reason for study removal and the date the patient was removed must be 
documented in the Case Report Form. 

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5.6 
Criteria to Resume Treatment 
 
Restarting applies only to grade 2 events and some grade 3 events (skin rash and thyroiditis). The 
section should emphasize stopping treatment and starting steroids earlier to obtain resolution 
with the possibility for restarting rather than waiting for higher grade events. 
 
For non-autoimmune or non-inflammatory events patients may resume treatment with study drug 
when the drug-related AE(s) resolve to Grade ≤1 or baseline value, with the following 
exceptions: 
 
• Patients may resume treatment in the presence of Grade 2 fatigue 
• Evaluation to exclude any additional immune mediated events endocrine, GI, and liver / 
pancreas function as clinically indicated must be made prior to restarting.  
• Non-drug-related toxicity including hepatic, pulmonary toxicity, diarrhea, or colitis, must 
have resolved to baseline before treatment is resumed. 
 
If the criteria to resume treatment are met, the patient should restart treatment no sooner than the 
next scheduled time point per protocol.  However, if the treatment is delayed past the next 
scheduled time point per protocol the treatment should resume at the earliest convenient point 
that is within the six week delay period.  
 
If treatment is delayed for >6 weeks, (>8 weeks for patients on a steroid taper), the patient must 
be permanently discontinued from study therapy, except as specified in Section 5.4 (Duration of 
Therapy). 
 
For patients treated with corticosteroids for autoimmune side effects: 
 
Grade 2 events must resolve to < Grade 1 before considering retreatment.  
 
All patients treated with steroids for grade ≥2 events should have nivolumab held until resolution 
to < Grade 1 for at least 2 weeks following complete removal from steroid treatment except for 
maintenance replacement doses for adrenal insufficiency (preferably no greater than 10mg 
prednisone equivalent daily).   
 
All patients treated with steroids for grade ≥3 events should have nivolumab discontinued.  
Patients with grade 3 thyroiditis and skin rash may continue therapy as for grade 2 events with 
resolution and stable replacement treatment. 
 
Patients with hepatitis, pancreatitis, pneumonitis, and colitis are at risk for exacerbation with 
retreatment if there is residual inflammation and should resolve to Grade 0 or baseline before 
retreatment.  Baseline can mean the initial grade i.e. grade <1 where permitted on study. 
 
Patients with thyroiditis or hypopituitarism who are stable as above may be restarted with 
replacement hormones including thyroid hormone and physiologic doses only of corticosteroids.  
Please note that grading and for hypophysitis with symptoms of headache, visual or neurologic 

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changes or radiologic evidence of pituitary enlargement and other CNS events such as aseptic 
meningitis or encephalitis should be considered grade 3 events.  
 
New immune related events or exacerbation of existing events during steroid treatment or taper 
suggest the presence of ongoing immune activation and should require permanent 
discontinuation of nivolumab.  
 
A patient who is treated with steroids, evaluated, and found to not have an autoimmune or 
inflammatory event requiring steroid treatment, may be restarted if asymptomatic off steroids for 
2 weeks and other restarting criteria are met.  
 
Prior to starting corticosteroids or hormone replacement for any reason, appropriate endocrine 
testing including cortisol, ACTH, TSH and T4 must be drawn if clinically feasible  to document 
baseline function and distinguish the pituitary  from  peripheral organ dysfunction and later from 
steroid (or thyroid)  treatment associated  ACTH ( or TSH)  suppression.  Steroids should be 
started prior to obtaining results based on clinical indications.  
 
 
5.7 
Treatment of Nivolumab-Related Infusion Reactions 
 
Since nivolumab contains only human immunoglobulin protein sequences, it is unlikely to be 
immunogenic and induce infusion or hypersensitivity reactions.  However, if such a reaction 
were to occur, it might manifest with fever, chills, rigors, headache, rash, urticaria, angioedema, 
pruritis, arthralgias, hypo- or hypertension, bronchospasm, or other symptoms.  
 
All Grade 3 or 4 infusion reactions should be reported as an SAE if criteria are met.  Infusion 
reactions should be graded according to NCI CTCAE version 5.0 guidelines. 
 
Treatment recommendations are provided below and may be modified based on local treatment 
standards and guidelines as medically appropriate: 
 
For Grade 1 symptoms:  (Mild reaction; infusion interruption not indicated; intervention not 
indicated) 
 
Remain at bedside and monitor subject until recovery from symptoms.  Infusion rate may be 
slowed.  If the infusion is interrupted, then restart the infusion at 50% of the original infusion 
rate when symptoms resolve; if no further complications ensue after 30 minutes, the rate may be 
increased to 100% of the original infusion rate.  Monitor patient closely.   
 
The following prophylactic premedications are recommended for future infusions:  
diphenhydramine 50 mg (or equivalent) and/or paracetamol 325 to 1000 mg (acetaminophen) at 
least 30 minutes before additional nivolumab administrations, slowing infusion rate as above. 
 
For Grade 2 symptoms:  (Moderate reaction requires therapy or infusion interruption but 
responds promptly to symptomatic treatment [e.g., antihistamines, non-steroidal anti-
inflammatory drugs, narcotics, corticosteroids, bronchodilators, IV fluids]; close observation for 

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recurrence and treatment  medications may need to be continued for 24-48 hours).   
 
Stop the nivolumab infusion, begin an IV infusion of normal saline, and treat the subject with 
diphenhydramine 50 mg IV (or equivalent) and/or paracetamol 325 to 1000 mg (acetaminophen); 
remain at bedside and monitor patient until resolution of symptoms.  Corticosteroid or 
bronchodilator therapy may also be administered as appropriate.  If the infusion is interrupted, 
then restart the infusion at 50% of the original infusion rate when symptoms resolve; if no further 
complications ensue after 30 minutes, the rate may be increased to 100% of the original infusion 
rate.  Monitor patient closely.  If symptoms recur, then no further nivolumab will be 
administered at that visit.  Administer diphenhydramine 50 mg IV, and remain at bedside and 
monitor the patient until resolution of symptoms.  The amount of study drug infused must be 
recorded on the electronic case report form (eCRF).  The following prophylactic premedications 
are recommended for future infusions: diphenhydramine 50 mg (or equivalent) and 
(acetaminophen) (or paracetamol) 325 to 1000 mg should be administered at least 30 minutes 
before additional nivolumab administrations.  If necessary, corticosteroids (recommended dose: 
up to 25 mg of IV hydrocortisone or equivalent) may be used. 
 
For Grade 3 or Grade 4 symptoms: (Severe reaction), Grade 3 symptoms: prolonged [i.e., not 
rapidly responsive to symptomatic medication and/or brief interruption of infusion]; recurrence 
of symptoms following initial improvement; hospitalization indicated for other clinical sequelae 
[e.g., renal impairment, pulmonary infiltrates]).   
 
Grade 4 symptoms: (life threatening; pressor or ventilatory support indicated).  
Immediately discontinue infusion of nivolumab.  Begin an IV infusion of normal saline, and treat 
the subject as follows.  Recommend bronchodilators, epinephrine 0.2 to 1 mg of a 1:1,000 
solution for subcutaneous administration or 0.1 to 0.25 mg of a 1:10,000 solution injected slowly 
for IV administration, and/or diphenhydramine 50 mg IV with methylprednisolone 100 mg IV 
(or equivalent), as needed.  Patient should be monitored until the investigator is comfortable that 
the symptoms will not recur.  Nivolumab will be permanently discontinued.  Investigators should 
follow their institutional guidelines for the treatment of anaphylaxis.  Remain at bedside and 
monitor patient until recovery from symptoms.  In the case of late-occurring hypersensitivity 
symptoms (e.g., appearance of a localized or generalized pruritis within 1 week after treatment), 
symptomatic treatment may be given (e.g., oral antihistamine, or corticosteroids). 
 
Please note that late occurring events including isolated fever and fatigue may represent the 
presentation of systemic inflammation.  Please evaluate accordingly.  
 
5.8 
Treatment with Nivolumab after Relapse in the Control Arm 
If the patients in the observation arm choose to get Nivolumab at the time of relapse; they will be 
followed for toxicities, mortality not related to AML progression and survival, but analyzed 
separately. Correlative study samples will be collected following the schedules for the patients 
randomized on the Nivolumab arm. 
 
For the patients in the control arm who cross over to receive Nivolumab treatment, the primary 
endpoint will be overall response rate (ORR) (including CR2, CR2i, partial response, and stable 
disease) after eight q2weeks treatment. Patients who progress during the treatment will be taken 

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off the study. Patients can receive a maximum of 46 doses (92 weeks). 
 
The International Working Group (IWG) response criteria were developed to assess the activity 
of drugs in AML. According to the criteria, relapse following complete remission is defined as 
reappearance of leukemic blasts in the peripheral blood or the finding of more than 5% blasts in 
the bone marrow, not attributable to another cause, or extramedullary relapse. 
 
Patients may be permitted to be treated with Nivolumab at the time to disease relapse in the 
control arm as long as they meet the inclusion criteria, except that the blood count requirement 
will be lower as listed below,  and the following criteria: 
 
• Patients must be clinically stable with no significant change in performance status 
due to disease progression 
• No indication for immediate alternative treatment 
• Patient assessed by the treating physician is going to have clinical benefit and 
tolerates study drug.  The assessment of clinical benefit should take into account 
whether the subject is clinically stable or deteriorating and likely or unlikely to 
receive further benefit from continued treatment. In general, the relapsed AML 
will not require urgent chemotherapy.. Hydroxyurea could be used to control 
AML prior to initiating Nivolumab treatment. The dose and duration of 
hydroxyurea would be based on the current practice for controlling AML prior to 
treatment  
• The cell counts will meet the following criteria at the time of first dose of 
Nivolumab:  
− 
leukocytes 
≥1,000/mcL 
− 
absolute neutrophil count 
≥500/mcL 
− 
platelets 
≥20,000/mcL  
 
6. DOSING DELAYS/DOSE MODIFICATIONS 
Below are DOSE DELAY tables for Nivolumab for the following adverse events 
 
Please refer to the Nivolumab Investigator Brochure or Appendix H to the protocol for toxicity 
management algorithms which include specific treatment guidelines.  These algorithms should 
be followed unless there are specific clinical circumstances which the treating physician 
indicates variations or alternative treatment is needed.  
 
Generally we strongly encourage early evaluation, withholding drug, and appropriate treatment 
as indicated in the management tables and following event specific guidelines.   
 
ALL OTHER 
EVENTS* 
Management/Next Dose for Nivolumab 
≤ Grade 1 
No change in dose 
Grade 2 
Hold until ≤ Grade 1 OR baseline Resume at same dose level. 
Grade 3 
Hold* until ≤ Grade 1 continue at investigator discretion  
Grade 4 
Off protocol therapy 
* Not agent related,  or agent related non-immunologically mediated 

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ALL OTHER 
EVENTS* 
Management/Next Dose for Nivolumab 
Recommended management:  As clinically indicated  
ALL OTHER 
EVENTS** 
Management/Next Dose for Nivolumab 
≤ Grade 1 
No change in dose  
Grade 2 
Hold until ≤ Grade 1 OR baseline* When  resolved  <  or  following 
steroids resume at same dose level . 
Grade 3 
 Off protocol therapy (exceptions noted in 5.4)  
Grade 4 
Off protocol therapy 
** immunologically mediated 
Recommended management:  As clinically indicated  
 
Skin Rash and Oral 
Lesions 
 
Management/Next Dose for Nivolumab 
≤ Grade 1 
No change in dose  
Grade 2 
Hold until 1≤ Grade resolved.  Consider steroid treatment > 7 days 
Resume at same dose level.  
Grade 3 
Hold until ≤ Grade 1.  Resume at same level at investigator discretion  
Grade 4 
Off protocol therapy 
*Patients with purpuric or bullous lesions must be evaluated for vasculitis, Steven-Johnson syndrome, 
TEN, and autoimmune bullous disease including oral lesions of bullous pemphigus/pemphagoid.  
Pruritus may occur with or without skin rash and should be treated symptomatically if there is no 
associated liver or GI toxicity.  Note skin rash typically occurs early and may be followed by additional 
events particularly during steroids tapering.   
Recommended management:  AE management guidelines 
 
 
Liver Function AST  
Management/Next Dose for Nivolumab 
≤ Grade 1 
Hold until UNL or baseline.   Resume at same dose level. 
Grade 2 
Hold until UNL or baseline.  Resume at same dose level. Consider 
steroid treatment  > 7 days 
Grade 3 
Off protocol therapy  
Grade 4 
Off protocol therapy 
Continued treatment of active immune mediated heapatitis may exacerbates ongoing inflammation.  
Holding drug to evaluate LFT changes and early treatment are recommended.   
LFT changes may occur during steroid tapers from other events and may occur together with other GI 
events including cholecystitis/pancreatitis. 
Recommended management:  see Hepatitic AE  management algorithm 
 
 
Diarrhea/ Colitis  
Management/Next Dose for Nivolumab 
≤ Grade 1 
Hold until Grade 0 or baseline.  No change in dose 
Grade 2 
Hold until Grade 0 or baseline.  Consider steroid treatment  > 7 days 
Grade 3 
Off protocol therapy. 

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Diarrhea/ Colitis  
Management/Next Dose for Nivolumab 
Grade 4 
Off protocol therapy 
See GI AE Algorithm for management of symptomatic colitis.  
Patients with grade 2 symptoms but normal colonoscopy and biopsies may be retreated after resolution.  
Patients who require steroids should be taken off study treatment.  
 Please evaluate pituitary function prior to starting steroids if possible without compromising acute care.   
Evaluation for all patients for additional causes includes C. diff, acute and self-limited infectious and 
foodborne illness, ischemic bowel, diverticulitis, and IBD.  
Recommended management:  see GI AE management Algorithm 
 
 
Pancreatitis 
Amylapse/Lipase  
Management/Next Dose for Nivolumab 
≤ Grade 1 
Hold dose until grade 0 
Grade 2 
Hold until Grade 0.  Resume at same dose level if asymptomatic 
 
Grade 3 
Hold* until Grade 0.  Resume at same dose level if asymptomatic. 
*Patients who develop symptomatic pancreatitis or DM should be taken 
off treatment  
Grade 4 
Off protocol therapy; Asymptomatic grade 4 amylase/lipase may be re-
treated as in Grade 3. 
Patients may develop symptomatic and radiologic evidence of pancreatitis as well as DM and 
DKA.  Lipase elevation may occur during the period of steroid withdrawal and with other 
immune mediated events or associated with colitis, hepatitis, and patients who have 
asymptomatic lipase elevation typically have self-limited course and may be retreated.  
For treatment management of symptomatic pancreatitis please follow the Hepatic Adverse 
Event Management Algorithm  
 
 
Pneumonitis 
Management/Next Dose for Nivolumab 
≤ Grade 1 
Hold dose pending evaluation and resolution to ≤ Grade 0 or baseline 
including baseline pO2.  Resume no change in dose after pulmonary 
and/or ID consultation  
Grade 2 
Hold dose pending evaluation. Resume no change in dose after 
pulmonary and/or ID consultation if lymphocytic pneumonitis is 
excluded.  Off study if steroids are required. ^  
Grade 3 
Hold dose pending evaluation. Resume no change in dose after 
pulmonary and/or ID consultation only  if  lymphocytic pneumonitis is 
excluded. Off protocol therapy  
Grade 4 
Off protocol therapy 
Distinguishing inflammatory pneumonitis is often a diagnosis of exclusion for patients who do not 
respond to antibiotics and have no causal organism identified including influenza.  Most patients with 
respiratory failure or hypoxia will be treated with steroids.  Bronchoscopy may be required and analysis 
of lavage fluid for lymphocytic predominance may be helpful.  Patients with new lung nodules should 
be evaluated for sarcoid like granuloma.  Please consider recommending seasonal influenza killed 
vaccine for all patients. 
Recommended management:  See Pulmonary Adverse Event Management Algorithm  

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Other GI N-V  
Management/Next Dose for Nivolumab 
≤ Grade 1 
No change in dose.  
Grade 2 
Hold pending evaluation for gastritis duodenitis and other immune 
adverse events or other causes.  Resume at same dose level after 
resolution to ≤ Grade 1.   
Grade 3 
Hold pending evaluation   until ≤ Grade 1.  Resume at same dose level. 
If symptoms do not resolve within 7 days with symptomatic treatment  patients 
should go off protocol therapy 
Grade 4 
Off protocol therapy 
Patients with grade 2 or 3 N-V should be evaluated for upper GI inflammation and other immune related 
events.   
 
Fatigue  
Management/Next Dose for Nivolumab 
≤ Grade 1 
No change in dose. 
Grade 2 
No change in dose 
Grade 3 
Hold until ≤ Grade 2.  Resume at same dose level 
Grade 4 
Off protocol therapy 
Fatigue is the most common adverse event associated with immune checkpoint therapy.  Grade 
2 or greater fatigue should be evaluated for associated or underlying organ involvement 
including pituitary, thyroid, and hepatic, or muscle (CPK) inflammation  
 
Neurologic events  
 
Management/Next Dose for Nivolumab 
≤ Grade 1 
Hold dose pending evaluation and observation. ^ Resume with no 
change in dose. * 
Grade 2 
Hold dose pending evaluation and observation. ^ Hold until ≤ Grade 
1.*Off protocol therapy if treatment with steroids is required. ^ Resume 
at same dose level for peripheral isolated n. VII (Bell’s palsy)^  
Grade 3 
Off protocol therapy  
Grade 4 
Off protocol therapy  
*Patients with any CNS events including aseptic meningitis, encephalitis, symptomatic 
hypophysitis, or myopathy, peripheral demyelinating neuropathy, cranial neuropathy (other 
than peripheral n. VII), GB syndrome, myasthenia gravis should be off study.   
Recommended management:  See Neurologic Adverse Event Management Algorithm  
 
 
Endocrine 
Hypophysitis 
Adrenal 
Insufficiency  
 
Management/Next Dose for Nivolumab 
≤ Grade 1 
Asymptomatic TSH elevation *  Hold pending evaluation, endocrine 
consult  
Grade 2 
Hold until patients are on a stable replacement hormone regimen.  If 

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Endocrine 
Hypophysitis 
Adrenal 
Insufficiency  
 
Management/Next Dose for Nivolumab 
treated with steroids patients must be stable off steroids for two weeks. 
Resume at same dose level.  
Grade 3 
Off study treatment. 
Grade 4 
Off protocol therapy 
Note all patients with symptomatic pituitary enlargement, exclusive of hormone deficiency, but 
including severe headache or enlarged pituitary on MRI should be considered grade 3 events.  Isolated 
thyroid or testosterone deficiency may be treated as grade 2 if there are no other associated deficiencies 
and adrenal function is monitored.  
Please evaluate pituitary function before beginning steroid therapy or replacement therapy of any kind.  
*Note patients with thyroiditis may be retreated on replacement therapy.  Patients must be evaluated to 
rule out pituitary disease prior to initiating thyroid replacement. 
Recommended management: See Endocrine Management Algorithm  
 
 
 
 
Fever  
Management/Next Dose for Nivolumab 
≤ Grade 1 
 
Continue nivolumab at same dose. If no improvement after 5-7 days or 
worsening, then treat as Grade 2.  
 
Grade 2 
Hold until ≤ Grade 1.  Resume at same dose level. 
Grade 3 
Hold until ≤ Grade 1.  Resume at same dose level. 
Grade 4 
Off treatment 
Patients with fever should be evaluated as clinically appropriate.  Patients may experience 
isolated fever during infusion reactions or up to several days after infusion.  Evaluation over the 
course of 1-2 weeks should be done for other autoimmune events that may present as fever  
See section 5.    infusion reactions   
 
If treatment is delayed >6 weeks, >8 weeks for patients on high dose steroids with recommended 
4 weeks taper and 2 week observation, the patient must be permanently discontinued from study 
therapy, except as specified in Section 5.7 (Criteria to Resume Treatment.) 
  
Patients requiring a delay of >6 weeks, >8 weeks for patients on high dose steroids with required 
4 weeks minimal taper and 2 week observation, should go off protocol therapy.  
Patients requiring > two dose delays for the same event should go off protocol therapy. 
 
Prior to starting corticosteroids or hormone replacement for any reason, appropriate endocrine 
testing including cortisol, ACTH, TSH and T4 must be obtained to document baseline.  
 
Patients may be dose-delayed for evaluation and restarted depending on results.  
 

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Any patient started on corticosteroids initially who is determined to not require steroids 
treatment for an autoimmune adverse event may resume therapy after a 2 week observation 
period without further symptoms at the discretion of the PI or investigator. 
 
Please refer to the Drug modification table for cardiomyopathy myocarditis:  
 
 
 
Cardiac *   
Management/Next Dose for BMS-936558 (Nivolumab) + 
Ipilimumab Cardiac Toxicities  
≤ Grade 1  
Hold dose pending evaluation and observation.**  Evaluate for signs and 
symptoms of CHF, ischemia, arrhythmia or myositis.  Obtain history EKG, CK 
(for concomitant myositis), CK-MB.  Repeat troponin, CK and EKG 2-3 days.  If 
troponin and labs normalize may resume therapy.   If labs worsen or 
symptoms develop then treat as below.  Hold pending evaluation  
Grade >2 with 
suspected 
myocarditis 
Hold dose.**  Admit to hospital.  Cardiology consult.  Rule out MI and other 
causes of cardiac disease.  Cardiac Monitoring.  Cardiac Echo.  Consider 
cardiac MRI and cardiac biopsy.  Initiate high dose methylprednisolone.  If no 
improvement within 24 hours, add either infliximab, ATG or tacrolimus. 
Consult algorithm for more details.  Resume therapy if there is a return to 
baseline and myocarditis is excluded or considered unlikely. 
Grade >2 with 
confirmed 
myocarditis   
Off protocol therapy.  Admit to CCU (consider transfer to nearest Cardiac 
Transplant Unit).  Treat as above.  Consider high dose methylprednisolone   
Add ATG or tacrolimus if no improvement.   Off treatment. 
 
 
*Including CHF, LV systolic dysfunction, Myocarditis, CPK, and troponin  
**Patients with evidence of myositis without myocarditis may be treated according as 
“other event”   
Note:  The optimal treatment regimen for immune mediated myocarditis has not been 
established.  Since this toxicity has caused patient deaths, an aggressive approach is 
recommended.   
 
▪ 
Drug will be held for grade 2 cardiac dysfunction pending evaluation 
▪ 
Drug will be permanently discontinued for grade  3 or 4 cardiac dysfunction and grade 2 events that 
do not recover to baseline or that reoccur 
▪ 
Missed doses of study drug are not made up. If drug was held for an adverse event (or other non-
medical reason), site must indicate that no drug was given for that cycle. 
▪ 
Treatment with steroids as clinically indicated  
▪ 
Add the table as follows in the treatment  modification and AE management section 
 
 
7. ADVERSE EVENTS:  LIST AND REPORTING REQUIREMENTS  
 
Adverse event (AE) monitoring and reporting is a routine part of every clinical trial.  The 
following list of AEs (Section 7.1) and the characteristics of an observed AE (Section 7.2) will 
determine whether the event requires expedited reporting via the CTEP Adverse Event Reporting 
System (CTEP-AERS) in addition to routine reporting. 
 
7.1 
Comprehensive Adverse Events and Potential Risks List (CAEPR) for BMS-936556 
(Nivolumab, MDX-1106, NSC 748726) 
 
The Comprehensive Adverse Events and Potential Risks list (CAEPR) provides a single list of reported and/or 

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potential adverse events (AE) associated with an agent using a uniform presentation of events by body system. In 
addition to the comprehensive list, a subset, the Specific Protocol Exceptions to Expedited Reporting (SPEER), 
appears in a separate column and is identified with bold and italicized text. This subset of AEs (SPEER) is a list of 
events that are protocol specific exceptions to expedited reporting to NCI (except as noted below). Refer to the 
'CTEP, NCI Guidelines: Adverse Event Reporting Requirements' 
http://ctep.cancer.gov/protocolDevelopment/electronic_applications/docs/aeguidelines.pdf for further clarification. 
Frequency is provided based on 2069 patients. Below is the CAEPR for BMS-936558 (Nivolumab, MDX-1106). 
NOTE: Report AEs on the SPEER ONLY IF they exceed the grade noted in parentheses next to the AE in the 
SPEER.  If this CAEPR is part of a combination protocol using multiple investigational agents and has an AE 
listed on different SPEERs, use the lower of the grades to determine if expedited reporting is required. 
 
     Version 2.4, December 2, 20201 
 
 Adverse Events with Possible  
 Relationship to Nivolumab 
 (CTCAE 5.0 Term) 
[n= 2069] 
 
 
 Specific Protocol 
Exceptions to Expedited 
Reporting (SPEER)  
 
Likely (>20%) 
Less Likely (<=20%) 
Rare but Serious (<3%) 
 
 
BLOOD AND LYMPHATIC SYSTEM DISORDERS 
  
 
Anemia 
 
 Anemia (Gr 3) 
CARDIAC DISORDERS 
  
 
 
Cardiac disorders - Other 
(cardiomyopathy) 
  
 
 
Myocarditis 
  
 
 
Pericardial tamponade2 
  
 
 
Pericarditis 
  
ENDOCRINE DISORDERS 
  
 
Adrenal insufficiency3 
 
  
 
Hyperthyroidism3 
 
  
 
Hypophysitis3 
 
  
 
Hypothyroidism3 
 
  
EYE DISORDERS 
  
 
 
Blurred vision 
  
 
 
Dry eye 
  
 
 
Eye disorders - Other (diplopia)3   
 
 
Eye disorders - Other (Graves 
ophthalmopathy)3 
  
 
 
Eye disorders - Other (optic 
neuritis retrobulbar)3 
  
 
 
Eye disorders - Other (Vogt-
Koyanagi-Harada) 
  
 
Uveitis 
 
  
GASTROINTESTINAL DISORDERS 
  
 
Abdominal pain 
 
 Abdominal pain (Gr 2) 
 
Colitis3 
 
  
 
 
Colonic perforation3 
  
 
Diarrhea 
 
 Diarrhea (Gr 3) 
 
Dry mouth 
 
 Dry mouth (Gr 2) 
 
 
Enterocolitis 
  
 
 
Gastritis 
  
 
 
Mucositis oral 
  

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38 
 
 Adverse Events with Possible  
 Relationship to Nivolumab 
 (CTCAE 5.0 Term) 
[n= 2069] 
 
 
 Specific Protocol 
Exceptions to Expedited 
Reporting (SPEER)  
 
Likely (>20%) 
Less Likely (<=20%) 
Rare but Serious (<3%) 
 
 
 
Nausea 
 
 Nausea (Gr 2) 
 
Pancreatitis4 
 
  
GENERAL DISORDERS AND ADMINISTRATION SITE CONDITIONS 
  
Fatigue 
 
 
 Fatigue (Gr 3) 
 
Fever 
 
 Fever (Gr 2) 
 
Injection site reaction 
 
 Injection site reaction (Gr 2) 
HEPATOBILIARY DISORDERS 
  
 
 
Hepatobiliary disorders - Other 
(immune-mediated hepatitis) 
  
IMMUNE SYSTEM DISORDERS 
  
 
 
Allergic reaction3 
  
 
 
Autoimmune disorder3 
  
 
 
Cytokine release syndrome5 
  
 
 
Immune system disorders - 
Other (GVHD in the setting of 
allotransplant)3,6 
  
 
 
Immune system disorders - 
Other (sarcoidosis)3 
  
INJURY, POISONING AND PROCEDURAL COMPLICATIONS 
  
 
Infusion related reaction7 
 
  
INVESTIGATIONS 
  
 
Alanine aminotransferase 
increased3 
 
 Alanine aminotransferase 
increased3 (Gr 3) 
 
Aspartate aminotransferase 
increased3 
 
 Aspartate aminotransferase 
increased3 (Gr 3) 
 
Blood bilirubin increased3 
 
 Blood bilirubin increased3 
(Gr 2) 
 
CD4 lymphocytes decreased 
 
 CD4 lymphocyte decreased 
(Gr 4) 
 
Creatinine increased 
 
  
 
Lipase increased 
 
  
 
Lymphocyte count decreased 
 
 Lymphocyte count 
decreased (Gr 4) 
 
Neutrophil count decreased 
 
  
 
Platelet count decreased 
 
  
 
Serum amylase increased 
 
  
METABOLISM AND NUTRITION DISORDERS 
  
 
Anorexia 
 
  
 
 
Hyperglycemia 
 Hyperglycemia (Gr 2) 
 
 
Metabolism and nutrition 
disorders - Other (diabetes 
mellitus with ketoacidosis)3 
  
MUSCULOSKELETAL AND CONNECTIVE TISSUE DISORDERS 
  
 
Arthralgia 
 
  

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 Adverse Events with Possible  
 Relationship to Nivolumab 
 (CTCAE 5.0 Term) 
[n= 2069] 
 
 
 Specific Protocol 
Exceptions to Expedited 
Reporting (SPEER)  
 
Likely (>20%) 
Less Likely (<=20%) 
Rare but Serious (<3%) 
 
 
 
 
Musculoskeletal and connective 
tissue disorder - Other 
(polymyositis) 
  
 
 
Myositis 
  
 
 
Rhabdomyolysis 
  
NERVOUS SYSTEM DISORDERS 
  
 
 
Encephalopathy3 
  
 
 
Facial nerve disorder3 
  
 
 
Guillain-Barre syndrome3 
  
 
 
Myasthenia gravis3 
  
 
 
Nervous system disorders - 
Other (demyelination 
myasthenic syndrome) 
  
 
 
Nervous system disorders - 
Other (encephalitis)3 
  
 
 
Nervous system disorders - 
Other (meningoencephalitis) 
  
 
 
Nervous system disorders - 
Other (meningoradiculitis)3 
  
 
 
Nervous system disorders - 
Other (myasthenic syndrome) 
  
 
 
Peripheral motor neuropathy 
  
 
 
Peripheral sensory neuropathy   
 
 
Reversible posterior 
leukoencephalopathy 
syndrome3 
  
RENAL AND URINARY DISORDERS 
  
 
 
Acute kidney injury3 
  
 
 
Renal and urinary disorders - 
Other (immune-mediated 
nephritis) 
  
RESPIRATORY, THORACIC AND MEDIASTINAL DISORDERS 
  
 
Pleural effusion3 
 
  
 
Pneumonitis3 
 
  
 
 
Respiratory, thoracic and 
mediastinal disorders - Other 
(bronchiolitis obliterans with 
organizing pneumonia)3 
  
SKIN AND SUBCUTANEOUS TISSUE DISORDERS 
  
 
 
Erythema multiforme3 
  
 
Pruritus3 
 
 Pruritus3 (Gr 2) 
 
Rash maculo-papular3 
 
 Rash maculo-papular3 (Gr 2) 
 
 
Skin and subcutaneous tissue 
disorders - Other (bullous 
pemphigoid) 
  

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 Adverse Events with Possible  
 Relationship to Nivolumab 
 (CTCAE 5.0 Term) 
[n= 2069] 
 
 
 Specific Protocol 
Exceptions to Expedited 
Reporting (SPEER)  
 
Likely (>20%) 
Less Likely (<=20%) 
Rare but Serious (<3%) 
 
 
 
Skin and subcutaneous tissue 
disorders - Other (Sweet's 
Syndrome)3 
 
  
 
Skin hypopigmentation3 
 
  
 
 
Stevens-Johnson syndrome 
  
 
 
Toxic epidermal necrolysis 
  
 
1This table will be updated as the toxicity profile of the agent is revised.  Updates will be distributed to all 
Principal Investigators at the time of revision.  The current version can be obtained by contacting 
PIO@CTEP.NCI.NIH.GOV.  Your name, the name of the investigator, the protocol and the agent should 
be included in the e-mail. 
 
2Pericardial tamponade may be related to possible inflammatory reaction at tumor site. 
 
3Nivolumab being a member of class of agents involved in the inhibition of “immune checkpoints”, may 
result in severe and possibly fatal immune-mediated adverse events probably due to T-cell activation and 
proliferation.  This may result in autoimmune disorders that can include (but are not limited to) 
autoimmune hemolytic anemia, acquired anti-factor VIII immune response, autoimmune aseptic 
meningitis, autoimmune hepatitis, autoimmune nephritis, autoimmune neuropathy, autoimmune 
thyroiditis, bullous pemphigoid, exacerbation of Churg-Strauss Syndrome, drug rash with eosinophilia 
systemic symptoms [DRESS] syndrome, facial nerve disorder (facial nerve paralysis), limbic encephalitis, 
hepatic failure, pure red cell aplasia, pancreatitis, ulcerative and hemorrhagic colitis, endocrine disorders 
(e.g., autoimmune thyroiditis, hyperthyroidism, hypothyroidism, autoimmune hypophysitis/hypopituitarism, 
thyrotoxicosis, and adrenal insufficiency), sarcoid granuloma, myasthenia gravis, polymyositis, and 
Guillain-Barre syndrome. 
 
4Pancreatitis may result in increased serum amylase and/or more frequently lipase. 
 
5Cytokine release syndrome may manifest as hemophagocytic lymphohistiocytosis with accompanying 
fever and pancytopenia. 
 
6Complications including hyperacute graft-versus-host disease (GVHD), some fatal, have occurred in 
patients receiving allo stem cell transplant (SCT) after receiving Nivolumab.  These complications may 
occur despite intervening therapy between receiving Nivolumab and allo-SCT. 
 
7Infusion reactions, including high-grade hypersensitivity reactions which have been observed following 
administration of nivolumab, may manifest as fever, chills, shakes, itching, rash, hypertension or 
hypotension, or difficulty breathing during and immediately after administration of nivolumab. 
 
 
Adverse events reported on Nivolumab trials, but for which there is insufficient evidence to 
suggest that there was a reasonable possibility that Nivolumab caused the adverse event: 
 
BLOOD AND LYMPHATIC SYSTEM DISORDERS - Leukocytosis 
CARDIAC DISORDERS - Atrial fibrillation; Atrioventricular block complete; Heart failure; Ventricular 
arrhythmia 
EAR AND LABYRINTH DISORDERS - Vestibular disorder 
EYE DISORDERS - Eye disorders - Other (iridocyclitis); Optic nerve disorder; Periorbital edema 
GASTROINTESTINAL DISORDERS - Constipation; Duodenal ulcer; Flatulence; Gastrointestinal 

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disorders - Other (mouth sores); Vomiting 
GENERAL DISORDERS AND ADMINISTRATION SITE CONDITIONS - Chills; Edema limbs; Malaise; 
Pain 
HEPATOBILIARY DISORDERS - Bile duct stenosis 
IMMUNE SYSTEM DISORDERS - Anaphylaxis; Immune system disorders - Other (autoimmune 
thrombotic microangiopathy); Immune system disorders - Other (limbic encephalitis) 
INFECTIONS AND INFESTATIONS - Bronchial infection; Lung infection; Sepsis; Upper respiratory 
infection 
INVESTIGATIONS - Blood lactate dehydrogenase increased; GGT increased; Investigations - Other 
(protein total decreased); Lymphocyte count increased; Weight loss 
METABOLISM AND NUTRITION DISORDERS - Dehydration; Hyperuricemia; Hypoalbuminemia; 
Hypocalcemia; Hyponatremia; Hypophosphatemia 
MUSCULOSKELETAL AND CONNECTIVE TISSUE DISORDERS - Back pain; Musculoskeletal and 
connective tissue disorder - Other (musculoskeletal pain); Musculoskeletal and connective tissue disorder 
- Other (polymyalgia rheumatica); Myalgia; Pain in extremity 
NEOPLASMS BENIGN, MALIGNANT AND UNSPECIFIED (INCL CYSTS AND POLYPS) - Neoplasms 
benign, malignant and unspecified (incl cysts and polyps) - Other (Histiocytic necrotizing lymphadenitis) 
NERVOUS SYSTEM DISORDERS - Dizziness; Headache; Intracranial hemorrhage 
PSYCHIATRIC DISORDERS - Insomnia 
RENAL AND URINARY DISORDERS - Hematuria; Renal and urinary disorders - Other (tubulointerstitial 
nephritis) 
RESPIRATORY, THORACIC AND MEDIASTINAL DISORDERS - Bronchospasm; Cough; Dyspnea; 
Hypoxia 
SKIN AND SUBCUTANEOUS TISSUE DISORDERS - Alopecia; Dry skin; Hyperhidrosis; Pain of skin; 
Photosensitivity; Rash acneiform; Skin and subcutaneous tissue disorders - Other (rosacea) 
VASCULAR DISORDERS - Flushing; Hypertension; Hypotension; Vasculitis 
 
 
Note: Nivolumab in combination with other agents could cause an exacerbation of any adverse event 
currently known to be caused by the other agent, or the combination may result in events never 
previously associated with either agent. 
 
 
7.2 
Adverse Event Characteristics  
 
CTCAE term (AE description) and grade:  The descriptions and grading scales found in the 
revised NCI Common Terminology Criteria for Adverse Events (CTCAE) version 5.0 will be 
utilized for AE reporting.  All appropriate treatment areas should have access to a copy of the 
CTCAE version 5.0.   
•  CTCAE v5.0 documents, including a mapping document, are available on the CTEP website: 
https://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm . 
 
• For expedited reporting purposes only:   
- 
AEs for the agent that are bold and italicized in the CAEPR (i.e., those listed in the 
SPEER column, Section 7.1.1) should be reported through CTEP-AERS only if the 
grade is above the grade provided in the SPEER. 
- 
Other AEs for the protocol that do not require expedited reporting are outlined in 
section 7.3.4. 
 
• Attribution of the AE: 
- 
Definite – The AE is clearly related to the study treatment. 

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- 
Probable – The AE is likely related to the study treatment. 
- 
Possible – The AE may be related to the study treatment. 
- 
Unlikely – The AE is doubtfully related to the study treatment. 
- 
Unrelated – The AE is clearly NOT related to the study treatment. 
 
7.3 
Expedited Adverse Event Reporting 
 
7.3.1 Expedited AE reporting for this study must use CTEP-AERS (CTEP Adverse Event 
Reporting System), accessed via the CTEP Web site (https://eapps-
ctep.nci.nih.gov/ctepaers).  The reporting procedures to be followed are presented in the 
“NCI Guidelines for Investigators: Adverse Event Reporting Requirements for DCTD 
(CTEP and CIP) and DCP INDs and IDEs” which can be downloaded from the CTEP 
Web site 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/adverse_events.htm
).  These requirements are briefly outlined in the tables below (Section 7.3.3). 
 
In the rare occurrence when Internet connectivity is lost, a 24-hour notification is to be 
made to CTEP by telephone at 301-897-7497.  Once Internet connectivity is restored, the 
24-hour notification phoned in must be entered electronically into CTEP-AERS by the 
original submitter at the site. 
 
7.3.2 CTEP-AERS is programmed for automatic electronic distribution of reports to the 
following individuals:  Principal Investigator and Adverse Event Coordinator(s) (if 
applicable) of the Corresponding Organization, the local treating physician, and the 
Reporter and Submitter.  CTEP-AERS provides a copy feature for other e-mail recipients. 
Please also copy all adverse events to the following email address: 
phaseIIcra@medicine.bsd.uchicago.edu   
 
The Coordinating Center of the Corresponding Organization is responsible for submitting 
to the CTSU documentation of AEs that they deem reportable for posting on the CTSU 
protocol web page and inclusion on the CTSU bi-monthly broadcast. 
 
7.3.3 Expedited Reporting Guidelines 
 
Use the NCI protocol number and the protocol-specific patient ID assigned during trial 
registration on all reports. 
 
Note:  A death on study requires both routine and expedited reporting regardless of 
causality, unless as noted below.  Attribution to treatment or other cause must be 
provided. 
 
Death due to progressive disease should be reported as Grade 5 “Neoplasms benign, 
malignant and unspecified (including cysts and polyps) - Other (Progressive 
Disease)” under the system organ class (SOC) of the same name.  Evidence that the death 
was a manifestation of underlying disease (e.g., radiological changes suggesting tumor 
growth or progression: clinical deterioration associated with a disease process) should be 

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submitted.  
 
Late Phase 2 and Phase 3 Studies: Expedited Reporting Requirements for Adverse Events 
that Occur on Studies under an IND/IDE within 30 Days of the Last Administration of the 
Investigational Agent/Intervention1, 2 
FDA REPORTING REQUIREMENTS FOR SERIOUS ADVERSE EVENTS (21 CFR Part 312) 
NOTE:  Investigators MUST immediately report to the sponsor (NCI) ANY Serious Adverse Events, whether or not 
they are considered related to the investigational agent(s)/intervention (21 CFR 312.64) 
An adverse event is considered serious if it results in ANY of the following outcomes:   
1) 
Death 
2) 
A life-threatening adverse event  
3) 
An adverse event that results in inpatient hospitalization or prolongation of existing hospitalization for ≥ 24 
hours  
4) 
A persistent or significant incapacity or substantial disruption of the ability to conduct normal life functions  
5) 
A congenital anomaly/birth defect.  
6) 
Important Medical Events (IME) that may not result in death, be life threatening, or require hospitalization 
may be considered serious when, based upon medical judgment, they may jeopardize the patient or 
subject and may require medical or surgical intervention to prevent one of the outcomes listed in this 
definition. (FDA, 21 CFR 312.32; ICH E2A and ICH E6). 
 
ALL SERIOUS adverse events that meet the above criteria MUST be immediately reported to the NCI via electronic 
submission within the timeframes detailed in the table below. 
Hospitalization 
Grade 1 
Timeframes 
Grade 2 
Timeframes 
Grade 3 Timeframes 
Grade 4 & 5 
Timeframes 
Resulting in 
Hospitalization  
≥ 24 hrs 
10 Calendar Days 
24-Hour 5 
Calendar Days 
Not resulting in 
Hospitalization  
≥ 24 hrs 
Not required 
10 Calendar Days 
NOTE:  Protocol specific exceptions to expedited reporting of serious adverse events are found in the Specific 
Protocol Exceptions to Expedited Reporting (SPEER) portion of the CAEPR 
Expedited AE reporting timelines are defined as: 
o 
“24-Hour; 5 Calendar Days” - The AE must initially be submitted electronically within 24 hours of learning 
of the AE, followed by a complete expedited report within 5 calendar days of the initial 24-hour report. 
o 
“10 Calendar Days” - A complete expedited report on the AE must be submitted electronically within 10 
calendar days of learning of the AE. 
1Serious adverse events that occur more than 30 days after the last administration of investigational 
agent/intervention and have an attribution of possible, probable, or definite require reporting as follows:  
Expedited 24-hour notification followed by complete report within 5 calendar days for: 
• 
All Grade 4, and Grade 5 AEs 
Expedited 10 calendar day reports for: 
• 
Grade 2 adverse events resulting in hospitalization or prolongation of hospitalization  
• 
Grade 3 adverse events 
2For studies using PET or SPECT IND agents, the AE reporting period is limited to 10 radioactive half-lives, 
rounded UP to the nearest whole day, after the agent/intervention was last administered.  Footnote “1” above 
applies after this reporting period. 
Effective Date:  May 5, 2011 
 
7.4 
Routine Adverse Event Reporting 
 
All Adverse Events must be reported in routine study data submissions.  AEs reported 

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expeditiously through CTEP-AERS must also be reported in routine study data 
submissions. 
 
Adverse event data collection and reporting, which are required as part of every clinical trial, are 
done to ensure the safety of patients enrolled in the studies as well as those who will enroll in 
future studies using similar agents.  AEs are reported in a routine manner at scheduled times 
during the trial using Medidata Rave.   
7.5 
Secondary Malignancy 
 
A secondary malignancy is a cancer caused by treatment for a previous malignancy (e.g., 
treatment with investigational agent/intervention, radiation or chemotherapy). A secondary 
malignancy is not considered a metastasis of the initial neoplasm. 
 
CTEP requires all secondary malignancies that occur following treatment with an agent under an 
NCI IND/IDE be reported expeditiously via CTEP-AERS.  Three options are available to 
describe the event:  
 
• Leukemia secondary to oncology chemotherapy (e.g., acute myelocytic leukemia 
[AML])  
• Myelodysplastic syndrome (MDS)  
• Treatment-related secondary malignancy  
 
Any malignancy possibly related to cancer treatment (including AML/MDS) should also be 
reported via the routine reporting mechanisms outlined in each protocol.  
 
  
7.6 
Second Malignancy 
 
A second malignancy is one unrelated to the treatment of a prior malignancy (and is NOT a 
metastasis from the initial malignancy).  Second malignancies require ONLY routine AE 
reporting unless otherwise specified. 
 
8. PHARMACEUTICAL INFORMATION 
 
A list of the adverse events and potential risks associated with the investigational agent 
administered in this study can be found in Section 7.1. 
 
8.1 
CTEP IND Agent: Nivolumab (NSC 748726) 
 
Amino Acid Sequence:  4 polypeptide chains, which include 2 identical heavy chains 
with 440 amino acids and 2 identical light chains. 
 
Other Names:  BMS-936558, MDX1106 
 
Classification:  Anti-PD-1MAb 
 

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M.W.:  146,221 daltons 
 
Mode of Action:  Nivolumab targets the programmed death–1 (PD-1, cluster of 
differentiation 279 [CD279]) cell surface membrane receptor.  PD-1 is a negative 
regulatory receptor expressed by activated T and B lymphocytes.  Binding of PD-1 to its 
ligands, programmed death–ligand 1 (PD-L1) and 2 (PD-L2), results in the down-
regulation of lymphocyte activation.  Nivolumab inhibits the binding of PD-1 to PD-L1 
and PD-L2.  Inhibition of the interaction between PD-1 and its ligands promotes immune 
responses and antigen-specific T-cell responses to both foreign antigens as well as self-
antigens. 
 
Description:  Nivolumab Injection is a clear to opalescent, colorless to pale yellow 
liquid; light (few) particulates may be present.  The drug product is a sterile, 
nonpyrogenic, single-use, isotonic aqueous solution formulated in sodium citrate 
dihydrate, sodium chloride, mannitol, diethylenetriaminepentacetic acid (pentetic acid) 
polysorbate 80 (Tween® 80), and water for injection.  Dilute solutions of hydrochloric 
acid and/or sodium hydroxide may be used for pH adjustment (pH 5.5-6.5) 
 
How Supplied:  Nivolumab is supplied by Bristol-Myers Squibb and distributed by the 
Pharmaceutical Management Branch, CTEP/DCTD/NCI as 100 mg vials (10 mg/mL) 
with a 0.7 mL overfill.  It is supplied in 10 mL type I flint glass vials, with fluoropolymer 
film-laminated rubber stoppers and aluminum seals. 
 
Preparation:    Nivolumab injection can be infused undiluted (10 mg/mL) or diluted with 
0.9% Sodium Chloride Injection, USP or 5% Dextrose.  When the dose is based on 
patient weight (i.e., mg/kg), nivolumab injection can be infused undiluted or diluted to 
protein concentrations as low as 0.35 mg/mL. For patients weighing less than 40 
kilograms (kg), the total volume of infusion must not exceed 4 mL per kg of patient 
weight. During drug product preparation and handling, vigorous mixing or shaking is to 
be avoided.   
Nivolumab infusions are compatible with polyvinyl chloride (PVC) or polyolefin 
containers and infusion sets, and glass bottles. 
 
 
 
Storage:  Vials of Nivolumab injection must be stored at 2°-8°C (36°-46°F) and 
protected from light and freezing.  The unopened vials can be stored at room temperature 
(up to 25°C, 77°F) and room light for up to 48 hours. 
If a storage temperature excursion is identified, promptly return Nivolumab to 2°C-8°C 
and quarantine the supplies.  Provide a detailed report of the excursion (including 
documentation of temperature monitoring and duration of the excursion) to 
PMBAfterHours@mail.nih.gov for determination of suitability. 
 
 
 

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Stability:  Shelf-life surveillance of the intact vials is ongoing. 
 
The administration of undiluted and diluted solutions of Nivolumab must be completed 
within 24 hours of preparation.  If not used immediately, the infusion solution may be 
stored up to 24 hours in a refrigerator at 2°-8°C (36°-46°F) and a maximum of 8 hours of 
the total 24 hours can be at room temperature (20°-25°C, 68°-77°F) and room light.  The 
maximum 8-hour period under room temperature and room light conditions includes the 
product administration period. 
 
CAUTION:  The single-use dosage form contains no antibacterial preservative or 
bacteriostatic agent.  Therefore, it is advised that the product be discarded 8 hours after 
initial entry.  
 
Route of Administration:  Intravenous infusion.  Do not administer as an IV push or 
bolus injection.   
 
Method of Administration:  Administer through a 0.2 micron to 1.2 micron pore size, 
low-protein binding polyethersulfone membrane in-line filter. 
 
Potential Drug Interactions:  The indirect drug-drug interaction potential of nivolumab 
was assessed using systemic cytokine modulation data for cytokines known to modulate 
CYP enzymes.  There were no meaningful changes in cytokines known to have indirect 
effects on CYP enzymes across all dose levels of nivolumab. This lack of cytokine 
modulation suggests that nivolumab has no or low potential for modulating CYP 
enzymes, thereby indicating a low risk of therapeutic protein-drug interaction. 
 
 
Availability 
 
Nivolumab is an investigational agent supplied to investigators by the Division of Cancer 
Treatment and Diagnosis (DCTD), NCI. 
 
Nivolumab is provided to the NCI under a Collaborative Agreement between the 
Pharmaceutical Collaborator and the DCTD, NCI (see Section 12.3). 
 
8.1.1 Agent Ordering and Agent Accountability 
 

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8.1.1.1 
NCI-supplied agents may be requested by the Principal Investigator (or their 
authorized designee) at each participating institution.  Pharmaceutical Management 
Branch (PMB) policy requires that agent be shipped directly to the institution where 
the patient is to be treated.  PMB does not permit the transfer of agents between 
institutions (unless prior approval from PMB is obtained).  The CTEP-assigned 
protocol number must be used for ordering all CTEP-supplied investigational agents.  
The responsible investigator at each participating institution must be registered with 
CTEP, DCTD through an annual submission of FDA Form 1572 (Statement of 
Investigator), Biosketch, Agent Shipment Form, and Financial Disclosure Form 
(FDF).  If there are several participating investigators at one institution, CTEP-
supplied investigational agents for the study should be ordered under the name of one 
lead investigator at that institution. 
Submit agent requests through the PMB Online Agent Order Processing (OAOP) 
application.  Access to OAOP requires the establishment of a CTEP Identity and 
Access Management (IAM) account and the maintenance of an “active” account 
status, a “current” password, and active person registration status.  For questions 
about drug orders, transfers, returns, or accountability, call or email PMB any time.  
Refer to the PMB’s website for specific policies and guidelines related to agent 
management. 
 
8.1.1.2 
Agent Inventory Records – The investigator, or a responsible party designated by the 
investigator, must maintain a careful record of the receipt, dispensing and final 
disposition of all agents received from the PMC using the appropriate NCI 
Investigational Agent (Drug) Accountability Record (DARF) available on the CTEP 
forms page.  Store and maintain separate NCI Investigational Agent Accountability 
Records for each agent, strength, formulation and ordering investigator on this 
protocol.  
 
8.1.1.3 
Useful Links and Contacts 
• CTEP Forms, Templates, Documents:  http://ctep.cancer.gov/forms/ 
• NCI CTEP Investigator Registration:  RCRHelpDesk@nih.gov 
• PMB policies and guidelines: 
http://ctep.cancer.gov/branches/pmb/agent_management.htm  
• PMB Online Agent Order Processing (OAOP) application:  
https://ctepcore.nci.nih.gov/OAOP 
• CTEP Identity and Access Management (IAM) account: 
https://ctepcore.nci.nih.gov/iam/  
• CTEP IAM account help:  ctepreghelp@ctep.nci.nih.gov  
• IB Coordinator:  IBCoordinator@mail.nih.gov  
• PMB email:  PMBAfterHours@mail.nih.gov 
• PMB phone and hours of service: (240) 276-6575 Monday through Friday 
between 8:30 am and 4:30 pm (ET) 
 
 

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9. BIOMARKER, CORRELATIVE, AND SPECIAL STUDIES 
 
9.1 
Integral Laboratory  
 
N/A 
 
 
9.2 
Integrated Correlative Studies (Mandatory studies) 
 
9.2.1 Analysis of PD-LI expression on AML cells from peripheral blood and/or bone 
marrow samples at diagnosis if available and at the time of study enrollment. 
 
Background: As discussed in the background section 2.3, to determine whether PD-L1 was 
expressed on human AML cells, bone marrow samples from AML patients were analyzed for 
expression patterns of PD-L1. Three staining patterns were observed: 1) no PD-L1 expression on 
either CD34+ or CD34- cells, 2) expression of PD-L1 on the CD34-, but not on CD34+ cells, 
and 3) PD-L1 expression on CD34+ AML cells, but not on CD34- cells.  These preliminary data 
support a hypothesis that PD-L1 expression on a subset of AML cases might serve as a possible 
baseline biomarker for response to PD-1-blocking antibodies, as has been demonstrated in 
melanoma patients (Topalian, Hodi et al.). We intend to test the expression of PD-LI inside the 
bone marrow environments and in the blood to determine if PD-LI expression will predict the 
response to the Nivolumab treatment. 
Method: Bone marrow core biopsy samples at the time of diagnosis (when available), study 
enrollment, 3, 6, 12 months on the study and at the end of the study either due to relapse or at 24 
months on the study will be collected from patients on each study arm.  Immunohistochemistry 
(IHC) staining using anti-PD-L1 and CD34 antibody will be conducted by BMS at the central lab 
if the test is ready at the time of the initiation of the study. Otherwise, PD-LI expression will be 
done by Flow cytometry using bone marrow aspiration and peripheral blood at Dr. Kline’s 
laboratory at University of Chicago as demonstrated in the Background section 2.3. These results 
will be used to determine if expression level of PD-L1 on bone marrow cells will predict 
patient’s response to Nivolumab treatment using logistic regression modeling. 
Analysis: The expression of PD-LI will quantified using the staining intensity or the mean 
fluorescence intensity from 0 to 1+ to 4+. To determine association between PD-LI expression and 
clinical response, patients will be divided based on response to Nivolumab maintenance therapy: 
responders (continuing CR) versus non-responders (disease relapse on Nivolumab). The differences 
will be assessed using a conventional T-test, followed by a modified T-test employed using the 
eBayes function in limma (R version 9.2). 
 
9.2.1.1 
Collection of Specimen(s): 
5-10* unstained bone marrow core biopsy slides (x3) (at time of diagnosis, at the time of enrollment, 
and at the time of relapse) and 5ml bone marrow aspiration in green top tube (x3) (at the time of 
enrollment and at the time of disease relapse) will be collected for PD-LI expression detection.  
* More slides (up to 20 total) may be requested if samples are not adequate for testing purposes.   
 
For bone marrow core biopsy unstained slides: The bone marrow core biopsy will processed 

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locally by the department of pathology at the participating institution as part of the standard of care. 
5-10 unstained bone marrow core biopsy slides will be shipped to University of Chicago medical 
center and stored for future PD-LI IHC staining ..  
 
For bone marrow aspiration samples: 5-10ml bone marrow aspiration will be collected using Green 
top tube. The fresh samples will be shipped on ice pack same day or overnight to University of 
Chicago central lab (Dr. Stock’s lab) for sample processing.  For centers with ability to process the 
samples, mononuclear cells (MNC) could be isolated using Ficoll method and  then be frozen in 
90% fetal calf serum/10% DMSO and stored in liquid nitrogen before shipment in batches.  Fresh 
samples should be sent with ice pack, and frozen samples should be sent on dry ice. All 
specimens must be accompanied by a specimen transmission (Appendix C-F) form that includes:  
Patient name  
Hospital record number  
NCI protocol number  
Sample type (aspirate, core biopsy, peripheral blood etc)  
Date when biopsy was obtained 
 
9.2.1.2 
Handling of Specimens (See also Appendix F) 
The shipment of all human tissue samples must comply with appropriate regulations as specified 
by the carrier. Fresh samples should be sent with ice pack, and frozen samples should be sent on 
dry ice. At a minimum, all samples must be packaged within two containers with absorbent 
material between containers to control any spill or leakage. The outer container must be puncture 
resistant (e.g., cardboard mail tube, corrugated cardboard box). A biohazard sticker must be 
affixed to both the inner and outer containers.  
 
9.2.1.3 
Shipping of Specimen(s) 
All bone marrow core biopsy blocks and bone marrow aspiration must be accompanied by a 
pathology report and a sample transmission form (appendix C-F) and shipped the following 
address:     
 
 
  
SHIPPING METHODS 
 
1. 
The fresh samples will be shipped on ice pack same day or overnight to University of 
Chicago Dr. Wendy Stock laboratory for sample processing.  
 
2. 
For centers with ability to process the samples, MNC could be isolated using Ficoll 
method and then be frozen in 90% fetal bovine serum/10% DMSO and stored in liquid nitrogen 
or -80ºC freezer before shipment in batches to Dr. Wendy Stock laboratory 
a. 
Lab Address: 
W. Stock Lab 
900 East 57th Street 
KCBD Room 9160 LB49 
Chicago, IL 60637 
b. 
Contact Number: 
773-795-3727 
 
3. 
Archival tissue slides or blocks should be sent to the University of Chicago Phase II 

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50 
Multi-site Coordinator for storage. 
a. 
Address: 
5841 S. Maryland Ave., MC 2115 
Chicago, IL 60637 
b. 
Contact number: 
773-834-1746 
 
 
Please send an email along with the shipment tracking information to Bartholomew Eisfelder at 
beisfelder@medicine.bsd.uchicago.edu Noreen Fulton at nfulton1@bsd.uchicago.edu (fresh 
samples) and copy the Phase 2 general e-mail phaseIIcra@medicine.bsd.uchicago.edu 
for tissue and slides. Please do NOT ship samples on Fridays as there will not be anyone here to 
receive the package or process the sample. 
 
 
9.2.1.4 
Site Performing Correlative Study:  
For Flow cytometry, it will be done by the lab of Dr. Kline at the University of Chicago. 
For IHC, it will be done by the by the lab of Dr. Kline or the hematopathology department at the 
University of Chicago. 
 
9.3 
Exploratory Correlative Studies 
All the exploratory correlative studies are mandatory. 
 
9.3.1.1 
Minimal residual disease (MRD) monitor  
 
Background: WT1 as a marker of minimal residual disease in AML 
Wilm’s tumor 1 (WT1) is a zinc finger transcription factor that is normally expressed in tissues 
of mesodermal origin during embryogenesis. WT1 is over-expressed in various hematological 
malignancies, including AML (80-100% of cases).  WT1 mRNA levels in the peripheral blood 
and bone marrow correlate with disease status during and after treatment, and elevation of WT1 
after chemotherapy and stem cell transplant predicts clinical relapse. Thus, WT1 RT-PCR is now 
being used to monitor minimal residual disease (MRD) during treatment of AML (Reviewed in 
(Grimwade, Vyas et al. 2010; Dominietto 2011)).  
 
Dr. Wendy Stock’s lab assessed MRD for a GVAX vaccine study in patients with AML using 
quantitative RT-PCR detection of WT1 expression. Although > 90% of patients who achieved 
clinical complete remission (CR) had significant declines in WT1 in both blood and bone 
marrow, 95% of patients had persistently detectable WT1 levels in bone marrow demonstrating 
clear MRD in AML patients in CR (Figure 3 in (Borrello, Levitsky et al. 2009)). Importantly, the 
achievement of a significant decrease or elimination of WT1 transcripts during the course of 
vaccination correlated with significant superior relapse-free survival (table 3 in (Borrello, 
Levitsky et al. 2009)). Furthermore, patients who relapsed during or following vaccination had a 
significant increase in WT1 expression, while patients in continuous CR maintained very low or 
undetectable WT1 levels (Figure 4 in (Borrello, Levitsky et al. 2009)).  
 
WT1 MRD monitoring appears to correlate well with remission status in conventionally-treated 

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AML patients as well.  Among AML patients in clinical CR after induction and consolidation 
therapy, 80-90% and 30-40% of patients continue to have detectable WT1 transcript in the 
marrow or blood, respectively (Miyawaki, Hatsumi et al.). WT1 levels in the peripheral blood 
can predict relapse after CR, and its levels after consolidation therapy are closely correlated with 
disease free survival (DFS), OS, and early relapse. The DFS rate at 3 years was 20% and 50% 
for WT1 MRD positive and negative patients, respectively (Miyawaki, Hatsumi et al.). Thus, 
WT1 monitoring is a useful tool not only for predicting relapse in AML patients in clinical CR, 
but also for monitoring response to post-remission immunotherapeutic strategies, such as 
vaccination and possibly after post-remission treatment with PD-1 blocking antibodies.   
 
Method: Assessment of MRD: MRD, measured by quantitative RT-PCR (qPCR) for WT1 
transcripts will be monitored in peripheral blood and bone marrow samples during the course of 
the study. Briefly, total RNA will be extracted from blood and bone marrow MNC, and cDNA 
synthesized using standard techniques. Amplifications of patient samples, K562 cell line cDNA 
(WT1-expressing), and no-template controls will be performed in triplicate. WT1 expression 
levels will be detected using a transcript-specific primer and probe set. To compensate for 
differences in RNA integrity and cDNA synthesis efficiency, the absolute WT1 transcript copy 
number will be normalized to the endogenous control gene, ABL. The limit of normalized WT1 
transcript quantification is 10-3 WT1/ABL. Disappearance of detectable WT1 transcript will be 
considered a complete molecular response (clearance of MRD). A one log decline in WT1 
transcript will define a partial molecular response.  Molecular stability will be defined by no 
reduction or increase in WT1 transcript level by more than one log (10 fold).  Molecular 
progression will be defined by greater than one log increase in WT1 transcript. Molecular 
response and progression will be tabulated by treatment arm and analyzed using ordinal logistic 
regression. 
Analysis: To determine association between MRD and clinical response, patients will be divided 
based on response to Nivolumab maintenance therapy: responders (MRD-) versus non-responders 
(MRD+). The differences will be assessed using a conventional T-test, followed by a modified T-test 
employed using the eBayes function in limma (R version 9.2). 
 
9.3.1.2 
Collection of Specimen(s) 
10-20ml bone marrow aspiration and peripheral blood will be collected using Green top tube. 
The bone marrow samples will be collected 5 times at the time of scheduled bone marrow 
biopsies on the study (At enrollment, week 13, 25, 53 on the study, and at the time clinically 
indicated bone marrow to evaluate relapse/off study). (See the study calendar for the detail). 
 
The peripheral blood samples will be collected on the same day of the scheduled bone marrow 
biopsy, an additional peripheral blood will be collected after 2 months on the study (week 9).  
 
For bone marrow aspiration samples: 10-20ml bone marrow aspiration will be collected. The 
fresh samples will be shipped on ice pack same day or overnight to University of Chicago central 
lab (Dr. Stock’s lab) for sample processing.  For centers with ability to process the samples, 
MNC could be isolated using Ficoll method and  then be frozen in 90% fetal calf serum/10% 
DMSO and stored in liquid nitrogen or -80ºC freezer before shipment in batches. 
 
For peripheral blood samples: The samples will be shipped on ice pack same day or overnight to 

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University of Chicago central lab (Dr. Stock’s lab) for sample processing.  For centers with 
ability to process the samples, MNC will be isolated per the standard procedure. Briefly, blood 
will be diluted 1:1 with PBS. Diluted blood will be layered on top of 5mls ficoll. Specimens will be 
spun at 350g for 20 minutes at room temperature. The buffy coat layer containing MNC will be 
collected and washed two times with PBS. MNC will then be frozen in 90% fetal calf serum/10% 
DMSO and stored in liquid nitrogen or -80ºC freezer before shipment in batches.. 
 
9.3.1.3 
Handling of Specimens(s) 
Same as 9.2.1.2. 
  
9.3.1.4 
Shipping of Specimen(s) 
Same as 9.2.1.3 
 
9.3.1.5 
Site(s) Performing Correlative Study 
It will be done by the lab of Dr. Stock at the University of Chicago. Dr. Christopher 
Hourigan (Myeloid Malignancies, NHLBI) will be a collaborator for the MRD 
correlate section. 
 
9.3.2 T cell subset analysis including Tregs 
 
Background: A recent study demonstrated that human AML cells constitutively express the 
immuno-suppression enzyme indoleamine 2, 3-dioxygenase (Curti, Aluigi et al. 2007) and co-
culture with IDO positive AML leukemia cells with lymphocytes could transform CD3+CD25- T 
cells to CD3+CD25+ T cells in vitro, which function as Treg cells because they do not proliferate, 
do not produce interleukin IL–2, and inhibit naive T-cell proliferation. These data suggest that 
AML leukemic cell-induced regulatory T cells might play a role in the pathogenesis of refractory 
or relapsed AML (Curti, Pandolfi et al. 2007).  It has been demonstrated that the frequency of 
CD4+CD25+ Treg in peripheral blood in AML patients is significantly higher when compared 
with healthy individuals (Wang, Zheng et al. 2005). A recent study demonstrated that Treg cell 
numbers prior to chemotherapy correlated with the response to chemotherapy. Specifically, 
patients who achieved a CR after induction chemotherapy had a significantly lower Treg 
frequency at diagnosis compared with patients who did not respond. Even though it needs to be 
proven that the observed high suppressor activity levels of Treg in patients with CR are related to 
higher relapse rates, it suggests that immunotherapy approaches to deplete or down-regulation of 
functional activity of Tregs are attractive to consider for patients with AML(Szczepanski, 
Szajnik et al. 2009). 
 
There is clear data to demonstrated that immune negative regulators, such as CTLA4 (reviewed 
in (Bour-Jordan, Esensten et al.), and PD1-PDL1 axis (Amarnath, Mangus et al.); could regulate 
the function of T regulatory cells. We intent to monitor the T cell subset changes during 
immunotherapy with PD1 antibody in the AML patients. Since Nivolumab will target PD1 on 
the T cells, we will also monitor the expression of PD1 on the T cells prior to the treatment, 
during the treatment and at the time of disease relapse. 
 
 
 
Method: Assessment of effect of Nivolumab on T cell subsets and T cell activation status, 

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53 
and PD1 expression on the T cells:  T cell frequencies and will be enumerated following 
staining of PBMC samples with anti-CD4, anti-CD8 and anti-FoxP3 staining and flow 
cytometry.  Absolute numbers will be enumerated by multiplying the frequency of CD8+, CD4+ 
and CD4+FoxP3+ (Tregs) cells by the total WBC from each sample.  T cell activation status will 
be performed in parallel, and include the following markers: HLA-DR, CD69, ICOS, and CD25. 
PD1 expression on T cells especially CD8 positive T cells will be assessed by flow cytometry.  
Others may be included depending on preliminary results.   
 
Analysis: Differences in pre- and post-treatment levels, as well as comparisons between blood 
samples, will be compared using a paired t-test. Changes in levels of responders versus non-
responders will be analyzed using a non-parametric Wilcoxon rank-sum test. 
 
9.3.2.1 
Collection of Specimen(s) 
5-10ml peripheral blood will be collected using Green top tube. 
The peripheral blood samples will be collected 6 times on the study. (At enrollment, week 9, 13, 
25, 53 on the study, and at the time of clinically indicated bone marrow to evaluate relapse/off 
study). 
 
For PD1 expression on the T cells, the samples will only be collected 3 times (at enrollment, 
week 9 after treatment and at the time of relapse). 
 
The fresh samples will be shipped on ice pack same day or overnight to University of Chicago 
central lab (Dr. Stock’s lab) for sample processing.  For centers with ability to process the 
samples, MNC will be isolated per the standard procedure. Briefly, blood will be diluted 1:1 with 
PBS. Diluted blood will be layered on top of 5mls ficoll. Specimens will be spun at 350g for 20 
minutes at room temperature. The buffy coat layer containing MNC will be collected and washed 
two times with PBS. MNC will then be frozen in 90% fetal calf serum/10% DMSO and stored in 
liquid nitrogen or -80ºC freezer before shipment in batches. 
 
9.3.2.2 
Handling of Specimens(s) 
Same as 9.2.1.2 
 
9.3.2.3 
Shipping of Specimen(s) 
Same as 9.2.1.3 
 
9.3.2.4 
Site(s) Performing Correlative Study : 
The tests will be conducted at the Human Immune Monitoring facility directed by Dr. 
Thomas Gajewski at University of Chicago. 
 
9.3.3 Deep sequencing of TCR- and TCR- chains on polyclonal T cells  
 
Background: Recent advances in deep sequencing technology make it possible to characterize 
the T cell receptor (TCR) repertoire generated following immunotherapy, such as WT1 peptide 
vaccination (i.e. the “clonality” of the response). Preliminary studies have demonstrated biased 
usage of TCR-V gene families in WT1 peptide vaccinated patients (Ochsenreither, Fusi et al. ; 
Ochsenreither, Fusi et al. ; Tanaka-Harada, Kawakami et al.), which was confirmed by our own 
TCR sequencing data in a patient who received WT1 peptide vaccination post SCT (Figure 4 in 

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54 
preliminary result section).  Dr. Yusuke Nakamura at our institution is one of the pioneers in the 
assessment of genetic variations (VNTR and SNP markers) and whole-genome analysis in cancer 
cells. Using a novel sequencing approach, we have been working together to understand the TCR 
“landscape” during immune-based therapies for AML. TCR sequencing of peripheral blood 
and/or bone marrow samples during immunotherapy, such as PD-1 blockade, will allow us to 
assess for an emerging T cell repertoire in patients who respond versus those who do not. This 
type of analysis may also be useful to identify high-affinity TCRs for use in down-stream 
adaptive T cell therapy approaches for AML, as well as to enable the identification of new AML 
associated antigens. 
 
Method: TCR Sequencing: To determine whether Nivolumab treatment leads to changes in the 
“clonality” of the polyclonal T cell repertoire, cDNA libraries for TCRA and TCRB genes will be 
constructed by polymerase chain reaction (PCR) using a primer set designed for adaptor 
sequence and C segment region in the TCR genes. Following that, emulsion-PCR will be 
conducted on the Ion Torrent system (Life Technologies, Carlsbad, CA) that templates clonal 
copies of DNA libraries onto the Ion Sphere Particles. The complex of DNA and particle will be 
deposited onto semiconductor chips for sequencing by the Ion Torrent PGM sequencer (Life 
Technologies, Carlsbad, CA).  This analysis will be performed on selected patient peripheral 
blood and marrow samples.   
 
Analysis: Differences in pre- and post-treatment levels, as well as comparisons between blood 
samples will be compared using a paired t-test. Changes in levels of responders versus non-
responders will be analyzed using a non-parametric Wilcoxon rank-sum test. 
 
9.3.3.1 
Collection of Specimen(s) 
5-10ml bone marrow aspiration and peripheral blood will be collected using Green top tube. 
The bone marrow samples will be collected 5 times at the time of scheduled bone marrow 
biopsies on the study (At enrollment, week 13, 25, 54 on the study and at the time clinically 
indicated bone marrow to evaluate relapse/off study). (See the study calendar for the detail). 
 
The peripheral blood samples will be collected on the same day of the scheduled bone marrow 
biopsy, an additional peripheral blood will be collected after 2 months (week 9) on the study. 
 
For bone marrow aspiration samples: 5-10ml bone marrow aspiration will be collected. The fresh 
samples will be shipped on ice pack same day or overnight to University of Chicago central lab 
(Dr. Stock’s lab) for sample processing.  For centers with ability to process the samples, MNC 
could be isolated using Ficoll method and  then be frozen in 90% fetal calf serum/10% DMSO 
and stored in liquid nitrogen or -80ºC freezer before shipment in batches. 
For peripheral blood samples: The fresh samples will be shipped on ice pack same day or 
overnight to University of Chicago central lab (Dr. Stock’s lab) for sample processing.  For 
centers with ability to process the samples, MNC will be isolated per the standard procedure. 
Briefly, blood will be diluted 1:1 with PBS. Diluted blood will be layered on top of 5mls ficoll. 
Specimens will be spun at 350g for 20 minutes at room temperature. The buffy coat layer containing 
MNC will be collected and washed two times with PBS. MNC will then be frozen in 90% fetal calf 
serum/10% DMSO and stored in liquid nitrogen or -80ºC freezer before shipment in batches. 
 

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9.3.3.2 
Handling of Specimens(s) 
Same as 9.2.1.2 
9.3.3.3 
Shipping of Specimen(s) 
Same as 9.2.1.3 
9.3.3.4 
Site(s) Performing Correlative Study   
The study will be done at the laboratory of Dr. Yusuke Nakamura at University of 
Chicago. 
 
9.4 
Special Studies 
 
n/a 
 

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56 
10. STUDY CALENDAR 
Baseline evaluations are to be conducted within 2 week prior to start of protocol therapy.  In the 
event that the patient’s condition is deteriorating, laboratory evaluations should be repeated 
within 72 hours prior to initiation of the next cycle of therapy. 
 
Study Calendar for Nivolumab and Observation Arms 
 
Pre- 
Study 
(Day -14 to 0) 
Wk  
1& 
(±1 Wk) 
Wk 
9@ 
(±1 Wk) 
Wk 
13 
(±1 Wk) 
Wk 
25 
(±1 Wk) 
Wk 
53 
(±1 Wk) 
Off Study or 
at the time 
of clinically 
suspected 
relapse H β 
NivolumabA 
 
B 
B 
B 
B 
B 
 
Informed consent 
X 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
Concurrent meds 
X 
X 
X 
X 
X 
X 
X 
Physical exam 
X 
X 
X 
X 
X 
X 
X 
Vital signs 
X 
X 
X 
X 
X 
X 
X 
Height 
X 
 
 
 
 
 
 
Weight 
X 
X 
X 
X 
X 
X 
X 
Performance status 
X 
X 
X 
X 
X 
X 
X 
CBC w/diff, plts C* 
X 
X 
X 
X 
X 
X 
X 
Serum chemistry D* 
X 
X 
X 
X 
X 
X 
X 
EKG (as indicated)Ω 
X 
 
 
 
 
 
 
HIV and hepatitis test E 
X 
 
 
 
 
 
 
Adverse event evaluation 
X 
X 
X 
X 
X 
X 
X 
B-HCG 
XF 
 
 
 
 
 
XF 
Bone marrow biopsy F (mandatory) 
X# 
 
 
X 
X 
X 
X 
Correlative studies: (Mandatory) 
 
 
 
 
 
 
 
PD-L1 staining/mutation  load 
(Bone marrow core biopsy slides 
or Bone marrow aspiration) 
X 
 
 
 
 
 
X 
WT1 MRD detection/digital PCR 
panel  (5-10cc peripheral blood at 
each time point and  5-10cc bone 
marrow aspiration on the day of 
scheduled bone marrow biopsy)   
X 
 
X 
X 
X 
X 
X 
T cell subsets analysis /CyTOF 
analysis. (5-10cc peripheral blood 
at each time point and  5-10cc bone 
marrow aspiration on the day of 
scheduled bone marrow biopsy 
X 
 
X 
X 
X 
X 
X 
T cell PD-1 expression (5-10cc 
peripheral blood in green top tube) 
X 
 
X 
 
 
 
X 
TCR sequencing (5-10cc 
peripheral blood and 5-10cc bone 
marrow aspiration if available) 
X 
 
X 
X 
X 
X 
X 

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57 
A: 
Nivolumab only given to patients on treatment arm, same calendar of events should be followed for all patients on study aside from the 
biweekly Nivolumab treatments. If a patient on observation relapses, patient may elect to cross over to the Nivolumab arm and follow 
the same calendar as patients whom originally randomized to the Nivolumab arm, with exception to the Bone Marrow biopsy at 9 
weeks, which is ONLY required for patients on observation who have crossed over to Nivolumab. 
B:     Nivolumab:  3mg/kg IV every 2 weeks for 46 doses. 
C:     Patients on both arms will have CBCs done every 4 weeks for the first year and every 8 weeks in the second year.  An unexplained 
drop in the counts especially platelet should will mandatorily trigger an unscheduled bone marrow exam to assess for relapse. 
D: 
Albumin, alkaline phosphatase, total bilirubin, bicarbonate, BUN, calcium, chloride, creatinine, glucose, LDH, magnesium, 
phosphorus, potassium, total protein, SGOT [AST], SGPT [ALT], sodium , amylase, lipase, TSH (with reflexive Free T4 and Free T3) 
E:    If patient has history of HIV or Hepatitis B or C on or off treatment, or patient has positive antibody test, viral load by quantitative PCR 
should be ordered, and viral loading negativity is required to be eligible on the study. 
F:     Complete bone marrow exams (with flow, cytogenetics, and molecular MRD) should be done prior to Nivolumab, at 3, 6, and 12 
months, and then only as clinically indicated (e.g., appearance of peripheral blasts or dropping platelets), and at the time off  the study 
on both arms of the trial.   
G:    Serum pregnancy test (women of childbearing potential). A serum or urine pregnancy testing is required within 72 hrs of study 
enrollment or randomization. and one week after discontinuation from nivolumab  
H:     Off-study evaluation or at the time of clinically suspected relapse.   
* Laboratory testing prior to Nivolumab treatment (will be done before every 2 cycles of Nivolumab treatment; every 4 weeks): Within 72 
hrs prior to re-dosing to include CBC w/ differential, LFTs, BUN or serum urea level, creatinine, Ca, Mg, Na, K, Cl, Glucose, Phosphate. 
(amylase, lipase, TSH only if clinically indicated) 
# This bone marrow biopsy could be done anytime within 60 days after the last chemotherapy; as long as the correlative studies samples are 
obtained and there is no evidence of disease relapse from the time of bone marrow biopsy to the start of treatment on the trial. Screening 
bone marrow should be repeated within two weeks of starting on protocol treatment. 
&: Any labs within 2 weeks to the start of treatment could be used.  
@: for patients who crossed over to Nivolumab Arm, will have response re-assessment at Week 9, to determine if the patient will continue 
Nivolumab or not. 
Ω:EKG and ECHO cardiogram for any patients with a history of CHF or at risk because of underlying cardiovascular disease or exposure to 
cardiotoxic drugs as clinically indicated. For patients with evidence of CHF, MI, cardiomyopathy, or myositis cardiac evaluation including 
lab tests and cardiology consultations as clinically indicated including EKG, CPK, troponin, ECHO cardiogram”. 
β: Patients who have come off treatment/observation after two years on study without relapse will continue to be followed every 6 months 
for the first year and yearly for disease relapse and survival. If patients come off treatment prior to two years, patients should be followed 
according to the protocol: CBCs every month for the first year and every two months for the second year. Patient will then be followed every 
6 months for the first year and then yearly for disease relapse and survival. 
11. MEASUREMENT OF EFFECT 
 
11.1 
Antitumor Effect – AML 
 
Since all the patients initially enrolled on the study will be in morphologic complete remission 
(CR) or CRi, defined by the International Working Group Criteria (Dohner, Estey et al. ; Cheson, 
Bennett et al. 2003)  
 
Morphologic complete remission (CR): Bone marrow blasts < 5%; absence of blasts with Auer 
rods; absence of extramedullary disease; absolute neutrophil count > 1.0 x 109/L (1000/uL); 
platelet count >100 x 109/L (100 000/uL); independence of red cell transfusions. 
 
Morphologic complete remission with incomplete blood count recovery (CRi): Same as CR 
but ANC may be <1,000/µL and/or platelet count <100,000/µL in a cellular marrow (greater than 
≥ 20%). 
 
The patients on the study will be monitor for disease relapse and PFS and OS defined as below. 
Morphologic Relapse following complete response: is defined as bone marrow blasts ≥ 5%; or 
reappearance of blasts in the blood; or development of extramedullary disease. 
 
Consensus to determine disease relapse or progression: 
 
In general, the AML relapse on the study will be defined by morphologic relapse. 

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58 
 
In addition, cytogenetic relapse defined by the reappearance of a cytogenetic abnormality, in the 
absence of morphologic relapse, could be used to determine disease relapse on the study based 
on all the evaluable clinic data. The determination should be discussed with the study PI. 
 
Molecular relapse defined by reappearance of a molecular abnormality; and appearance of new 
dysplastic changes might be used to define disease relapse after the review of the clinical data 
and the discussion with the study PI. 
 
Progression-free survival (PFS): Defined as the time from randomization to disease relapse or 
death from any cause. Patients not known to have relapsed or died at last follow-up are censored 
on the date they were last examined.  
 
Overall Survival (OS) is defined for all patients of a trial; measured from the date of entry into a 
study to the date of death from any cause; patients not known to have died at last follow-up are 
censored on the date they were last known to be alive. 
 
Cumulative incidence of relapse (CIR): Defined for all patients achieving CR or CRi measured 
from the date of achievement of a remission until the date of relapse; patients not known to have 
relapsed are censored on the date they were last examined; patients who died without relapse are 
counted as a competing cause of failure. 
 
For the patient in the control arm that has disease relapse during the follow-up and cross over to 
be treated with Nivolumab. The response to Nivolumab treatment will be also evaluated by the 
International Working Group Criteria. 
 
Morphologic complete remission (CR): Bone marrow blasts < 5%; absence of blasts with Auer 
rods; absence of extramedullary disease; absolute neutrophil count > 1.0 x 109/L (1000/uL); 
platelet count >100 x 109/L (100 000/uL); independence of red cell transfusions. 
 
Morphologic complete remission with incomplete blood count recovery (CRi): Same as CR 
but ANC may be <1,000/µL and/or platelet count <100,000/µL in a cellular marrow (greater than 
≥ 20%). 
 
Partial Remission (PR): Requires all hematologic values for a CR (i.e. ANC ≥1,000/µL, platelet 
count ≥100,000/µL), but with a decrease of at least 50% in the percentage of marrow blasts to 5% 
to 25% in the bone marrow aspirate. 
 
Treatment Failure (TF): Any one of the following: 
 
 
Failure to achieve a CR, CRi or PR 
 
  
Failure due to resistant disease (patient survives ≥7 days post-chemotherapy with 
persistent disease in blood or bone marrow); Persistent disease is defined as an 
increase of at least 25% in the absolute number of leukemic cells in peripheral blood 
or bone marrow/aspirate, the development of extramedullary disease, or other 

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evidence of increased tumor burden. 
 
 
Failure due to aplasia [patient survives ≥7 days post-chemotherapy; death while 
cytopenic and aplastic (<20% marrow cellularity) bone marrow with no leukemic 
blasts] 
 
Failure due to indeterminate cause (patients who die <7 days post-chemotherapy; 
patients who die >7 days post-chemotherapy with no peripheral blood blasts, but no 
bone marrow examination; patients who do not complete the first course of Induction 
therapy) 
 
11.2 
Other Response Parameters: 
 
Cytogenetic CR (CRc): Reversion to a normal karyotype at the time of morphologic CR (or 
CRi) in cases with an abnormal karyotype at the time of diagnosis; based on the evaluation of 20 
metaphase cells from bone marrow. 
 
Molecular CR (CRm): No standard definition; but in general, it will be defined as 
disappearance of detectable molecule marker at the diagnosis, such as NPM1 mutation, FLT3-
ITD mutation and CEBP mutation et al at the time of morphologic CR (or CRi). 
 
Molecular and/or cytogenetic relapse is characterized by reappearance of a cytogenetic 
or molecular abnormality. 
 
Minimal residual disease (MRD) monitoring: In this protocol, MRD will be monitored by 
quantitative RT-PCR (qRT-PCR) of WT1 gene which will be normalized by control gene ABL. 
The limit of normalized WT1 transcript quantification is 10-3 WT1/ABL.  
 
MRD negative (MRD-): Disappearance of detectable WT1 transcript will be considered a 
complete molecular response (clearance of MRD).  
 
Partial MRD response: A one log decline in WT1 transcript will define a partial molecular 
response.   
 
Stable MRD: Molecular stability will be defined by no reduction or increase in WT1 transcript 
level by more than one log (10 fold).  
 
MRD progression: Molecular progression will be defined by greater than one log increase in 
WT1 transcript. 
 
12. DATA REPORTING / REGULATORY REQUIREMENTS 
 
Adverse event lists, guidelines, and instructions for AE reporting can be found in Section 7.0 
(Adverse Events: List and Reporting Requirements). 
 

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60 
12.1 
Study Oversight 
 
This protocol is monitored at several levels, as described in this section. The Protocol Principal 
Investigator is responsible for monitoring the conduct and progress of the clinical trial, including 
the ongoing review of accrual, patient-specific clinical and laboratory data, and routine and 
serious adverse events; reporting of expedited adverse events; and accumulation of reported 
adverse events from other trials testing the same drug(s). The Protocol Principal Investigator and 
statistician have access to the data at all times through the CTMS web-based reporting portal. 
 
During the phase 2 portion of the study, the Protocol Principal Investigator will have, at a 
minimum, quarterly conference calls with the Study Investigators and the CTEP Medical 
Officer(s) to review accrual, progress, and pharmacovigilance. Decisions to proceed to the 
second stage of a Phase 2 trial will require sign-off by the Protocol Principal Investigator and the 
Protocol Statistician through IWRS and Medidata Rave. 
 
All studies are also reviewed in accordance with the enrolling institution’s data safety monitoring 
plan. 
 
12.2 
Data Reporting 
 
Data collection for this study will be done exclusively through Medidata Rave.  Access to the 
trial in Rave is granted through the iMedidata application to all persons with the appropriate 
roles assigned in the Regulatory Support System (RSS).  To access Rave via iMedidata, the site 
user must have an active CTEP IAM account (https://eapps-ctep.nci.nih.gov/iam) and the 
appropriate Rave role (Rave CRA, Read-Only, or Site Investigator) on either the Corresponding 
Organization or Participating Organization roster at the enrolling site.   
 
Upon initial site registration approval for the study in RSS, all persons with Rave roles assigned 
on the appropriate roster will be sent a study invitation e-mail from iMedidata.  To accept the 
invitation, site users must log into the Select Login (https://login.imedidata.com/selectlogin) 
using their CTEP-IAM user name and password, and click on the “accept” link in the upper 
right-corner of the iMedidata page.  Please note, site users will not be able to access the study in 
Rave until all required Medidata and study specific trainings are completed.  Trainings will be in 
the form of electronic learnings (eLearnings), and can be accessed by clicking on the link in the 
upper right pane of the iMedidata screen.   
 
Users that have not previously activated their iMedidata/Rave account at the time of initial site 
registration approval for the study in RSS will also receive a separate invitation from iMedidata 
to activate their account. Account activation instructions are located on the CTSU website, Rave 
tab under the Rave resource materials (Medidata Account Activation and Study Invitation 
Acceptance).  Additional information on iMedidata/Rave is available on the CTSU members’ 
website under the Rave tab or by contacting the CTSU Help Desk at 1-888-823-5923 or by e-
mail at ctsucontact@westat.com. 
 
12.2.1 Method 
 

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This study will be monitored by the Clinical Trials Monitoring Service (CTMS).  Data 
will be submitted to CTMS at least once every two weeks via Medidata Rave (or other 
modality if approved by CTEP).  Information on CTMS reporting is available at 
http://www.theradex.com/CTMS.    On-site audits will be conducted three times annually 
(one annual site visit and two data audits).  For CTMS monitored studies, after users have 
activated their accounts, please contact the Theradex Help Desk at (609) 799-7580 or by 
email at ctms@theradex.com for additional support with Rave and completion of CRFs. 
 
 
12.2.2 Responsibility for Data Submission 
 
For ETCTN trials, it is the responsibility of the PI(s) at the site to ensure that all 
investigators at the ETCTN Sites understand the procedures for data submission for each 
ETCTN protocol and that protocol specified data are submitted accurately and in a timely 
manner to the CTMS via the electronic data capture system, Medidata Rave.   
 
Data are to be submitted via Medidata Rave to CTMS on a real-time basis, but no less 
than once every 2 weeks.  The timeliness of data submissions and timeliness in resolving 
data queries will be tracked by CTMS.  Metrics for timeliness will be followed and 
assessed on a quarterly basis.  For the purpose of Institutional Performance Monitoring, 
data will be considered delinquent if it is greater than 4 weeks past due. 
 
Data from Medidata Rave and CTEP-AERS is reviewed by the CTMS on an ongoing 
basis as data is received.  Queries will be issued by CTMS directly within Rave.  The 
queries will appear on the Task Summary Tab within Rave for the CRA at the ETCTN to 
resolve.  Monthly web-based reports are posted for review by the Drug Monitors in the 
IDB, CTEP.  Onsite audits will be conducted by the CTMS to ensure compliance with 
regulatory requirements, GCP, and NCI policies and procedures with the overarching 
goal of ensuring the integrity of data generated from NCI-sponsored clinical trials, as 
described in the ETCTN Program Guidelines, which may be found on the CTEP 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/adverse_events.htm
)  and CTSU websites. 
 
An End of Study CRF is to be completed by the PI.  CTMS will utilize a core set of 
eCRFs that are Cancer Data Standards Registry and Repository (caDSR) compliant 
(http://cbiit.nci.nih.gov/ncip/biomedical-informatics-resources/interoperability-and-
semantics/metadata-and-models).  Customized eCRFs will be included when appropriate 
to meet unique study requirements.  The PI is encouraged to review the eCRFs, working 
closely with CTMS to ensure prospectively that all required items are appropriately 
captured in the eCRFs prior to study activation.  CTMS will prepare the eCRFs with 
built-in edit checks to the extent possible to promote data integrity. 
 
CDUS data submissions for ETCTN trials activated after March 1, 2014, will be carried 
out by the CTMS contractor, Theradex.  CDUS submissions are performed by Theradex 
on a monthly basis.  The trial’s lead institution is responsible for timely submission to 
CTMS via Rave, as above. 

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Further information on data submission procedures can be found in the ETCTN Program 
Guidelines 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/adverse_events.htm
). 
 
   
12.3 
CTEP Multicenter Guidelines 
 
N/A. 
• . 
 
12.4 
Collaborative Agreements Language 
 
The agent(s) supplied by CTEP, DCTD, NCI used in this protocol is/are provided to the NCI 
under a Collaborative Agreement (CRADA, CTA, CSA) between the Pharmaceutical 
Company(ies) (hereinafter referred to as “Collaborator(s)”) and the NCI Division of Cancer 
Treatment and Diagnosis.  Therefore, the following obligations/guidelines, in addition to the 
provisions in the “Intellectual Property Option to Collaborator”  
(http://ctep.cancer.gov/industryCollaborations2/intellectual_property.htm) contained within the 
terms of award, apply to the use of the Agent(s) in this study: 
 
1. Agent(s) may not be used for any purpose outside the scope of this protocol, nor can 
Agent(s) be transferred or licensed to any party not participating in the clinical study.  
Collaborator(s) data for Agent(s) are confidential and proprietary to Collaborator(s) and shall 
be maintained as such by the investigators.  The protocol documents for studies utilizing 
Agents contain confidential information and should not be shared or distributed without the 
permission of the NCI.  If a copy of this protocol is requested by a patient or patient’s family 
member participating on the study, the individual should sign a confidentiality agreement.  A 
suitable model agreement can be downloaded from: http://ctep.cancer.gov. 
 
2. For a clinical protocol where there is an investigational Agent used in combination with 
(an)other Agent(s), each the subject of different Collaborative Agreements, the access to and 
use of data by each Collaborator shall be as follows (data pertaining to such combination use 
shall hereinafter be referred to as "Multi-Party Data”): 
 
a. NCI will provide all Collaborators with prior written notice regarding the existence and 
nature of any agreements governing their collaboration with NCI, the design of the 
proposed combination protocol, and the existence of any obligations that would tend to 
restrict NCI's participation in the proposed combination protocol. 
 
b. Each Collaborator shall agree to permit use of the Multi-Party Data from the clinical trial 
by any other Collaborator solely to the extent necessary to allow said other Collaborator 
to develop, obtain regulatory approval or commercialize its own Agent. 
 

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c. Any Collaborator having the right to use the Multi-Party Data from these trials must 
agree in writing prior to the commencement of the trials that it will use the Multi-Party 
Data solely for development, regulatory approval, and commercialization of its own 
Agent. 
 
3. Clinical Trial Data and Results and Raw Data developed under a Collaborative Agreement 
will be made available to Collaborator(s), the NCI, and the FDA, as appropriate and unless 
additional disclosure is required by law or court order as described in the IP Option to 
Collaborator (http://ctep.cancer.gov/industryCollaborations2/intellectual_property.htm).  
Additionally, all Clinical Data and Results and Raw Data will be collected, used and 
disclosed consistent with all applicable federal statutes and regulations for the protection of 
human subjects, including, if applicable, the Standards for Privacy of Individually 
Identifiable Health Information set forth in 45 C.F.R. Part 164. 
 
4. When a Collaborator wishes to initiate a data request, the request should first be sent to the 
NCI, who will then notify the appropriate investigators (Group Chair for Cooperative Group 
studies, or PI for other studies) of Collaborator's wish to contact them. 
 
5. Any data provided to Collaborator(s) for Phase 3 studies must be in accordance with the 
guidelines and policies of the responsible Data Monitoring Committee (DMC), if there is a 
DMC for this clinical trial. 
 
6. Any manuscripts reporting the results of this clinical trial must be provided to CTEP by the 
Group office for Cooperative Group studies or by the principal investigator for non-
Cooperative Group studies for immediate delivery to Collaborator(s) for advisory review and 
comment prior to submission for publication.  Collaborator(s) will have 30 days from the 
date of receipt for review.  Collaborator shall have the right to request that publication be 
delayed for up to an additional 30 days in order to ensure that Collaborator’s confidential and 
proprietary data, in addition to Collaborator(s)’s intellectual property rights, are protected.  
Copies of abstracts must be provided to CTEP for forwarding to Collaborator(s) for courtesy 
review as soon as possible and preferably at least three (3) days prior to submission, but in 
any case, prior to presentation at the meeting or publication in the proceedings.  Press 
releases and other media presentations must also be forwarded to CTEP prior to release.  
Copies of any manuscript, abstract and/or press release/ media presentation should be sent to: 
 
Email:  ncicteppubs@mail.nih.gov 
 
The Regulatory Affairs Branch will then distribute them to Collaborator(s).  No publication, 
manuscript or other form of public disclosure shall contain any of Collaborator’s confidential/ 
proprietary information. 
 
13. STATISTICAL CONSIDERATIONS 
 
13.1 
Study Design/Endpoints 
This is an open-label, randomized Phase II study to evaluate the efficacy of single agent of 
Nivolumab as maintenance treatment to prevent relapse in AML patients in first CR after induction 

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chemotherapy or consolidation chemotherapy.  Eighty patients with AML in first CR or CRi after 
induction and/or consolidation chemotherapy will be randomized into an observation arm 
(standard of care) or to Nivolumab treatment. Patients assigned to the Nivolumab arm will receive 
Nivolumab at 3mg/kg IV every 2 weeks for 2 years (46 doses). Patients on both arms will be 
monitored for relapse, adverse events and survival for a minimum of 2 years following 
randomization. During and after treatment, patients will be evaluated for toxicities, particularly 
immunologic adverse events including pneumonitis, autoimmune colitis, dermatitis etc. Peripheral 
blood and bone marrow samples will be collected before and during the treatment course for the 
correlative studies outlined above.  Complete blood counts and differential counts will be obtained 
every 4 weeks for the first year, and every 8 weeks for the second year on the study, for patients 
on both arms.  Bone marrow examination for disease assessment and correlative studies will be 
performed at the following time points: at the time of study enrollment; 3, 6 and 12 months 
following enrollment and/or at the time of study discontinuation due to toxicity or disease relapse. 
 
 

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Eligible patients will be randomized to observation arm vs Nivolumab arm with stratification by 
AML risk category (Adverse, Intermediate-1, Intermediate-2 and favorable) and age (<60 years, 
>=60 years). Patients randomized to the Nivolumab arm will receive IV Nivolumab at 3mg/kg 
every 2 weeks for forty-six doses (2 years). After that, patients will be followed up for adverse 
events, relapse and survival until at least 2 years after the last patient on the study is randomized. 
Patients who have come off treatment/observation after two years on study without relapse will 
continue to be followed every 6 months for the first year and yearly for disease relapse and 
survival. 
 
When the patients on the observation arm are confirmed to have disease relapse (defined as 
below), at the treating physicians’ discretion, the patients have the option to cross over to receive 
3mg/kg Nivolumab every 2 weeks in the same manner as the patients on the Nivolumab arm till 
disease progression or intolerable toxicities or closure of the trial. Upon disease relapse for 
patients on the observation arm, there will not be a randomization to be treated with Nivolumab 
or to be off the study. The decision to cross-over from the observation arm to be treated with 
Nivolumab at disease relapse will be made by the treating physician after discussion with the 
patient. 
The total duration of the treatment with Nivolumab for these patients will be no more than 2 
years after the accrual to the study is completed.  Before treatment with Nivolumab, these 
patients should still meet the inclusion criteria and exclusion criteria other than the disease status. 
Alternatively, these patients could be taken off study to pursue further salvage chemotherapy at 
any time at the treating physician’s discretion. However, all of these patients will be followed for 
survival. Many of the patients on the observation arm at the time of relapse might elect to receive 
salvage chemotherapy followed in some cases by stem cell transplantation. However, treatment 
with Nivolumab for early relapse in AML patients, especially elderly AML patients who are not 
candidates for intensive chemotherapy and SCT, might provide valuable clinical and scientific 
insight without affecting the primary objective of PFS of the study.  
 
If the patients in the observation arm choose to get Nivolumab at the time of relapse; they will be 
followed for toxicities, mortality not related to AML progression and survival, but analyzed 
separately. Correlative study samples will be collected following the schedules for the patients 
AML patients in first 
remission after 
induction or 
consolidation 
chemotherapy (not 
candidate for SCT) 
Screening and randomization 
 
Observation Arm 
Treated with 
Nivolumab 
(3mg/kg IV q2weeks till 
disease progression or 
intolerable toxicities or 
closure of the study 
D i s e a s e  R e l a p s e 
Off study  
Nivolumab Arm 
(3mg/kg IV q2weeks x 46  
doses)  
Off study  

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randomized on the Nivolumab arm. 
 
For the patients in the control arm who cross over to receive Nivolumab treatment, the primary 
endpoint will be overall response rate (ORR) (including CR2, CR2i, partial response, and stable 
disease) after eight q2weeks treatment. Patients who progress during the treatment will be taken 
off the study. 
 
Patients will have a baseline eligibility screen within 1-2 weeks prior to randomization, including 
medical history, physical examination, assessment of performance status, and assessment of 
hematology and chemistry parameters.  Baseline assessment of status of AML will depend on 
bone marrow aspirate and biopsy by morphology and flow cytometry.  Residual disease from 
cytogenetic and molecular analyses is allowed. Patient will have bone marrow biopsy prior to the 
randomization to confirm the status of AML, and obtain pre-treatment bone marrow samples 
along with peripheral blood samples for the correlative studies. The minimal residual disease 
(MRD) status will be determined by cytogenetics/FISH study; molecular studies if patients have 
identifiable disease markers, such as the NPM1 mutation. All the patients will also have 
quantitative RT-PCR to check for WT1 expression. The pre-treatment MRD status is not a 
requirement to be enrolled in the clinical trial. 
 
Bone marrow samples will be collected after four cycles of receiving every 2 weeks doses of 
Nivolumab to determine disease status, MRD status and to be used for correlative studies. 
Complete blood counts and differentials will be obtained monthly, and any appearance of 
peripheral blasts or unexpected drop in blood counts will trigger a bone marrow exam to evaluate 
for relapse.  Peripheral blood samples will be collected after four and eight every 2 weeks doses, 
at 1 year on the treatment and at the end of the treatment for correlative studies. All the patients 
will be followed up for clinical relapse or death. The relapse rate and progression free survival 
will be calculated for each arm; and in subset analysis for MRD+ or MRD- patients separately.  
In order to have comparison, patients on the observation arm will have all the scheduled sample 
collection. 
 
Analysis of primary endpoint:  
The primary endpoint is progression free survival (PFS). PFS is defined as the time from 
randomization to disease relapse or death from any cause. Patients alive and progression free will 
be censored at the date of the last negative bone marrow examination. Patients will be accrued 
over a two-year period and follow-up will continue for 2 years after the last patient is enrolled, 
providing a minimum of 2 years and maximum 4 years follow-up of each patient. Kaplan-Meier 
plots (Kaplan and Meier 1958) will be used to estimate PFS in each arm and a stratified log rank 
test (Peto, Pike et al. 1976) will be performed to compare the two groups. In addition, Cox 
proportional hazards models (Cox 1972) will be fit to provide estimates of the hazard ratio (HR) 
and associated 95% confidence interval (CIs), both unadjusted and adjusted for baseline 
covariates.  
 
This is a randomized Phase II study to compare relapse (progression) free survival between 
observation and Nivolumab maintenance treatment. According to the historical data, the 2 year 
PFS survival of AML after induction chemotherapy and 3-4 cycles high dose cytarabine 
consolidation is around 40%  in young patients (Mayer, Davis et al. 1994) and about 35% 

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including older patients extrapolated from the data of patients with primary AML enrolled onto 
CALGB first-line treatment trials who did not undergo allogeneic SCT in first complete 
remission per protocol (Figure 4A and 4C in (Mrozek, Marcucci et al.)). Since young AML 
patients in the European LeukemiaNet favorable group have excellent 2 year PFS at around 64%, 
further maintenance therapy might not provide additional benefit; thus the current trial will 
exclude young favorable group AML patients. The estimated 2 year PFS for all the AML 
patients excluding young favorable group is around 25% (Mrozek, Marcucci et al.). We expect 
that Nivolumab maintenance after induction and consolidation will improve the 2 year PFS rate 
from 25% to 45%.  Assuming exponential distributions, this corresponds to a HR of 0.58.   
 
Using a one-sided alpha of 0.10, to have 80% power to detect a HR of 0.58 would require 80 
patients (40 patients per arm) (Schoenfeld 1983).  This assumes 2 years of accrual and 2 years 
follow-up of the last patient. An interim futility analysis will be conducted after half the number 
of expected events (i.e., 31 of 62 expected) occur.  If, at this point, the observed HR does not favor 
the experimental arm, the study will be stopped for futility. This futility rule is associated with a 
minimal (<2%) reduction in power (Wieand, Schroeder et al. 1994).  
 
 
After reviewing the interim analysis results, the Data Monitoring Committee recommended that 
the trial be continued.  Accrual will proceed until the randomization of 80 eligible patients.  
 
Due to the heterogeneous population of AML patients, during the randomization, the patients will 
be stratified according to AML European LeukemiaNet risk categories (Adverse, intermediate-1, 
intermediate-2 or favorable) and age (<60 years vs. >= 60 years) (Mrozek, Marcucci et al.). 
Randomization sequences will be prepared by the statistician at University of Chicago and 
REDCap will be used for web-based treatment allocation.  
 
13.2 
Sample Size/Accrual Rate 
 
A total 82 (The first two patients enrolled on the trial were mistakenly not randomized and were 
administered nivolumab) will be enrolled into this randomized Phase II study.  The two patients 
mistakenly not randomized will not be included in primary efficacy outcome, however, they will 
be included overall safety analysis. 
 
We have agreement from all the cancer centers within our Phase II consortium to participate in 
this clinical trial. Our consortium consists of the following institutions: University of Chicago, 
University of Maryland, University of Michigan, and Northwestern University, NorthShore 
University Health System, Indiana University and all the community cancer centers in our Phase 
II consortium. We have strong record of collaboration with these institutions. We also have 
agreement from University of Wisconsin; City of Hope and Penn State Hershey Cancer Institute 
through University of California Davis Comprehensive Cancer Center P2C; Vanderbilt 
University through H Lee Moffitt Cancer Center P2C; Princess Margaret Cancer Center in 
Canada through University Health Network Princess Margaret Cancer Center P2C  to participate 
in the study. Thus, there will be at least 7-8 major institutions to assure timely accrual.  
 
Furthermore, all P2C will have the opportunity to participate in this trial via the ETCTN 

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mechanism.  
 
We anticipate enrollment of 3-4 patients/per month from all the participating institutions to 
accomplish the total accrual goal of 82 patients within 2 years. 
 
PLANNED ENROLLMENT REPORT 
Racial Categories 
Ethnic Categories 
Total 
Not Hispanic or Latino 
Hispanic or Latino 
 
Female 
Male 
Female 
Male 
 
American Indian/ Alaska 
Native 
1 
1 
 
 
2 
Asian 
2 
2 
 
 
4 
Native Hawaiian or 
Other Pacific Islander 
1 
1 
 
 
2 
Black or African 
American 
10 
10 
 
 
20 
White 
20 
20 
5 
5 
50 
More Than One Race 
1 
1 
 
 
2 
Total 
35 
35 
5 
5 
80 
PHS 398 / PHS 2590 (Rev. 08/12 Approved Through 8/31/2015)  
OMB No. 0925-0001/0002 
 
 
13.3 
Stratification Factors 
 
Due to the heterogeneous population of AML patients, during the randomization, the patients will 
be stratified according to AML European LeukemiaNet risk categories (Adverse, intermediate-1, 
intermediate-2 or favorable) and age (<60 years vs. >= 60 years) (Mrozek, Marcucci et al.). 
Randomization sequences will be prepared by the statistician at University of Chicago and 
REDCap will be used for web-based treatment allocation.  
 
13.4 
Analysis of Secondary Endpoints 
 
The secondary endpoints include: Overall Survival (OS), non-relapse mortality (NRM) and 
adverse effects of Nivolumab.  
 
OS is defined as the time from randomization to the date of death from any cause. NRM is 

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defined as the duration between the date of randomization and the date of patient death due to 
reasons other than relapse. 
 
For overall survival, Kaplan-Meier plots will be generated for each treatment arm and the curves 
will be compared using a log rank test. Kaplan-Meier estimates with 95% CI will be summarized 
every 6 month using Greenwood’s formula for the standard error of the Kaplan-Meier estimate.  
In addition to the Kaplan-Meier estimates, Cox proportional hazards models will be fit to 
estimate HR’s and to assess and adjust for the effects of covariates.  NRM will be analyzed using 
competing risks models (Gooley, Leisenring et al. 1999) with deaths due to relapse as a 
competing risk.  Cumulative incidence curves will be generated and treatment effects 
summarized using the sub-distribution hazard ratio (Fine and Gray 1999) with and without 
adjustment for covariates. 
 
Descriptive, subset analysis of PFS and OS will be performed for MRD+ and MRD- patients, 
respectively. 
 
Adverse Events Monitoring: 
Summary tables for adverse events (AEs) will include only AEs that started or worsened after 
randomization. The incidence of treatment-related adverse events (new or worsening from 
baseline after randomization) will be summarized by system organ class and/ or preferred term, 
severity (based on CTCAE grades), type of adverse event, and relation to study treatment by 
treatment group. Deaths reportable as SAEs and non-fatal serious adverse events will be listed by 
patient and tabulated by type of adverse event and treatment group.  All laboratory values will be 
converted into SI units and the severity grade calculated using Common Toxicity Criteria for 
Adverse Events (CTCAE).  All laboratory values will be listed by laboratory parameter and 
patient. Separate listings will display notable laboratory abnormalities (i.e. newly occurring 
CTCAE grade 3 or 4 laboratory toxicities).  
 
Toxicity rates will be compared between the two treatment arms via chi-square or Fisher’s exact 
test. Formal comparisons will be conducted at the time of the interim analysis (i.e., after 31 
events) and at completion of the trial.  However, we will also undertake more frequent safety 
monitoring to ensure that serious AEs (deaths or hospitalizations) or selected AEs are not 
occurring at an unacceptably high rate in the Nivolumab arm. The selected AEs will be those 
listed in Section 7.1 as having a possible relationship to Nivolumab based on previous trials, with 
the stipulation that they are of grade 4 or higher. First, all serious AEs are reviewed at our 
weekly Phase II monitoring conferences.  Second, safety reports containing comparative results 
will be generated quarterly and presented at the phase II conference.  If the rate of SAEs exceeds 
20% in the Nivolumab arm or the difference in rates between the Nivolumab and control arms is 
statistically significant at the p<0.01 level (to adjust for multiplicity), consideration will be given 
to stopping the trial.  Accrual will be suspended pending consultation between the principal and 
co-investigators to determine whether the trial should be terminated. 
 
13.5 
 Reporting and Exclusions 
 
13.5.1 Evaluation of Toxicity 
 

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All patients including participants randomized to the control arm will be evaluable for 
toxicity from the time of their first treatment with Nivolumab. 
 
13.5.2 Evaluation of Response 
 
All eligible participants who were randomized will be accounted for, during the 
assessment for response to treatment, since the primary analysis will be an intent-to-treat 
analysis.  Each patient will be assigned one of the following categories:  1) complete 
response, 2) partial response, 3) stable disease, 4) progressive disease, 5) early death from 
malignant disease, 6) early death from toxicity, 7) early death because of other cause, or 
9) unknown (not assessable, insufficient data).  [Note:  By arbitrary convention, category 
9 usually designates the “unknown” status of any type of data in a clinical database.] 
 
All of the patients who met the eligibility criteria should be included in the main analysis 
of the response rate.  Patients in response categories 4-9 should be considered to have a 
treatment failure (disease progression).  Thus, an incorrect treatment schedule or drug 
administration does not result in exclusion from the analysis of the response rate.  Precise 
definitions for categories 4-9 will be protocol specific.  
 
All conclusions should be based on all eligible patients.  Subanalyses may then be 
performed on the basis of a subset of patients, excluding those for whom major protocol 
deviations have been identified (e.g., early death due to other reasons, early 
discontinuation of treatment, major protocol violations, etc.).  However, these 
subanalyses may not serve as the basis for drawing conclusions concerning treatment 
efficacy, and the reasons for excluding patients from the analysis should be clearly 
reported.  The 95% confidence intervals should also be provided.  
 
 
14. STUDY STATUS UPDATES AND STUDY CLOSURE 
 
14.1 
Definitions of Study Status Changes 
 
14.1.1 Temporarily Closed to Accrual 
 
The study status is Temporarily Closed to Accrual when no patient slots are currently 
available, but there is the possibility that the trial will re-open for accrual (patient slots 
become available).  Sites are not permitted to accrue additional patients until CTEP is 
notified of Re-Activation. 
 
Study status will need to be changed to Temporarily Closed to Accrual when any of the 
following criteria are met: 
 
• Sites are notified by CTEP (via Request for Rapid Amendment [RRA]) of 
changes in the risk/benefit ratio that necessitate changes to the patient Informed 
Consent document.  Requested changes will be specified in the RRA and must be 
reviewed by the study’s IRB. 

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• CTEP and the lead investigator agree that unacceptable toxicities necessitate a 
discussion to change the dosing/regimen. 
• A protocol-defined benchmark has been achieved (such as an interim analysis 
before proceeding to the next stage).  
• Investigators encounter any of the stopping criteria described in Section 5.5. 
 
14.1.2 Closed to Accrual 
 
The study status is (permanently) Closed to Accrual when no more patient enrollment 
slots are available, and at least one patient is still actively receiving the study treatment.  
Sites are no longer permitted to enroll additional patients. 
 
Patient slots are no longer available when the following criteria are met: 
 
• The pre-specified number of evaluable patients has been successfully enrolled, 
treated, and evaluated. 
• The study treatment has failed to meet the pre-specified efficacy goal at the stage 
1 interim analysis. 
• CTEP and the investigators agree that unacceptable toxicities preclude further 
enrollment. 
• Investigators encounter any of the stopping criteria described in Section 5.5. 
 
14.1.3 Closed to Accrual and Treatment 
 
The study status is Closed to Accrual and Treatment when no more patient enrollment 
slots are available and no patients are currently receiving the study treatment.  Patients 
may still be enrolled on the protocol only for the purposes of follow-up. 
 
Patient accrual and treatment will be permanently halted when any of the following 
criteria are met: 
 
• Enrollment was previously closed (study status of “Closed to Accrual”), and no 
patients are receiving the study treatment. 
• CTEP and the investigators agree that unacceptable toxicities preclude further 
enrollment.  In this case, CTEP and the investigators must collaborate to alter the 
regimen or to halt the study treatment altogether as soon as it can be safely done 
for patients currently receiving treatment.  
 
CTEP and Theradex must be notified when patients are no longer receiving treatment 
[i.e., when the last patient(s) to be receiving treatment is/are no longer receiving the study 
regimen for any reason]. 
 
14.1.4 Closed to Follow-Up 
 
The study is considered Closed to Follow-Up when all protocol-defined follow-up 
procedures have been completed for all patients who have not been removed from the 

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study for other reasons.  That is, there are no outstanding follow-up procedures to be 
performed as mandated by the protocol. 
 
CTEP does not need to be notified of a status change to “Closed to Follow Up.” 
 
14.1.5 Complete 
 
Study is considered Complete if it has been at least thirty (30) days since the last patient 
follow-up evaluation.   
 
A citation to a final study report (manuscript, meeting abstract, etc.) is required with the 
submission of the Protocol Status Update Form to CTEP PIO.   
 
14.1.6 Future plan:  
 
If this Phase II study demonstrates that Nivolumab  can prolong 2 year PFS as 
maintenance therapy compared to observation alone, we will then plan to conduct a 
randomized Phase III trial to compare Nivolumab vs observation aiming for FDA 
approval of Nivolumab for the indication of maintenance treatment in AML patients. 
 
14.2 
Responsibility for Filing Protocol Status Update Forms 
 
CTEP must be notified of all study status changes in Section 14.1 (except for Closed to Follow-
Up) by the Corresponding Organization via Protocol Status Update Form, available from the 
CTEP website at http://ctep.cancer.gov/protocolDevelopment/default.htm#amendments.    
 
Theradex must be notified as soon as all patients are off treatment (i.e., when study status 
changes to Closed to Accrual and Treatment).  Theradex will produce a report within 90 days of 
this notification. 
 
 

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Dohner, H., E. H. Estey, et al. "Diagnosis and management of acute myeloid leukemia in adults: 
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Dominietto, A. (2011). "Minimal residual disease markers before and after allogeneic 
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Estey, Elihu. “Acute Myeloid Leukemia: 2016 Update on Risk-Stratification and 
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Farag, S. S., A. S. Ruppert, et al. (2005). "Outcome of induction and postremission therapy in 
younger adults with acute myeloid leukemia with normal karyotype: a cancer and leukemia 
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Fine, J. P. and R. J. Gray (1999). "A proportional hazards model for the subdistribution of a 
competing risk." Journal of the American Statistical Association 94(446): 496-509. 
 
Gajewski, T. F., Y. Meng, et al. (2006). "Immune resistance orchestrated by the tumor 
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competing risks: New representations of old estimators." Statistics in Medicine 18(6): 695-706. 
 
Grimwade, D., P. Vyas, et al. (2010). "Assessment of minimal residual disease in acute myeloid 
leukemia." Curr Opin Oncol 22(6): 656-63. 
 
Hamid, O., C. Robert, et al. "Safety and tumor responses with lambrolizumab (anti-PD-1) in 
melanoma." N Engl J Med 369(2): 134-44. 
 
Hammers, H.J., E.R. Plimack, J.R. Infante, et al.  (2014).  Phase I study of nivolumab in 
combination with ipilimumab in metastatic renal cell carcinoma (mRCC).  J Clin Oncol.  
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Hodi, F. S., S. J. O'Day, et al. (2010). "Improved survival with ipilimumab in patients with 
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Investigator Brochure.  (2014).  Nivolumab, BMS-936558, MDX1106.  Version 13, 21 July 
2014. Bristol-Myers Squibb.  Lawrenceville, NJ.   
 

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Ipilimumab Investigator Brochure.  (2014).  BMS-734016, MDX010.  Version 17, 10 March 
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Miyawaki, S., N. Hatsumi, et al. "Prognostic potential of detection of WT1 mRNA level in 
peripheral blood in adult acute myeloid leukemia." Leuk Lymphoma 51(10): 1855-61. 
 
Mrozek, K., G. Marcucci, et al. "Prognostic significance of the European LeukemiaNet 
standardized system for reporting cytogenetic and molecular alterations in adults with acute 
myeloid leukemia." J Clin Oncol 30(36): 4515-23. 
 
Nomi, T., M. Sho, T. Akahori, et al.  (2007).  Clinical significance and therapeutic potential of 
the programmed death-1 ligand/programmed death-1 pathway in human pancreatic cancer.  Clin 
Cancer Res.  13:2151-2157. 
 
Ochsenreither, S., A. Fusi, et al. ""Wilms Tumor Protein 1" (WT1) peptide vaccination-induced 
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ligand-1 and programmed death-1 ligand-2 expression in human esophageal cancer.  Clin Cancer 
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Rizvi, N.A., S.J. Antonia, L.Q.M. Chow, et al.  (2013).  A phase I study of nivolumab (anti-PD-
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chemotherapy-naive non-small cell lung cancer (NSCLC) patients (pts).  J Clin Oncol.  
31:A8072. 
 
Sznol, M., H.M. Kluger, F.S. Hodi, et al.  (2013).  Survival and long-term follow-up of safety 
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(anti-PD-1; BMS-936558; ONO-4538).  J Clin Oncol.  31:ACRA9006. 
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Szczepanski, M. J., M. Szajnik, et al. (2009). "Increased frequency and suppression by regulatory 
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Cancer Sci 101(3): 594-600. 
 
Taube, J.M., R.A. Anders, G.D. Young, et al.  (2012).  Colocalization of inflammatory response 
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Thompson, R.H., M.D. Gillett, J.C. Cheville, et al.  (2004).  Costimulatory B7-H1 in renal cell 
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Thompson, R.H., M.D. Gillett, J.C. Cheville, et al.  (2005).  Costimulatory molecule B7-H1 in 
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Thompson, R.H., S.M. Kuntz, B.C. Leibovich, et al.  (2006).  Tumor B7-H1 is associated with 
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Topalian, S.L., F.S. Hodi, J.R. Brahmer, et al.  (2012).  Safety, activity, and immune correlates 
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Wang, X., J. Zheng, et al. (2005). "Increased population of CD4(+)CD25(high), regulatory T 
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Weber, J. S., R. R. Kudchadkar, et al. "Safety, efficacy, and biomarkers of nivolumab with 
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Wolchok, J.D., H. Kluger, M.K. Callahan, et al.  (2013).  Nivolumab plus ipilimumab in 
advanced melanoma.  N Engl J Med.  369:122-133. 
 
Woo, S.R., M.E. Turnis, M.V. Goldberg, et al.  (2012).  Immune inhibitory molecules LAG-3 
and PD-1 synergistically regulate T-cell function to promote tumoral immune escape.  Cancer 
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108:19-24. 

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Zhang, L., T. F. Gajewski, et al. (2009). "PD-1/PD-L1 interactions inhibit antitumor immune 
responses in a murine acute myeloid leukemia model." Blood 114(8): 1545-52. 
 
Zitvogel, L., A. Tesniere, and G. Kroemer.  (2006).  Cancer despite immunosurveillance: 
immunoselection and immunosubversion.  Nat Rev Immunol.  6:715-727. 
 
Zou, W. (2006). "Regulatory T cells, tumour immunity and immunotherapy." Nat Rev Immunol 
6(4): 295-307. 
 
 
 

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APPENDIX A 
PERFORMANCE STATUS CRITERIA 
 
ECOG Performance Status Scale 
Karnofsky Performance Scale 
Grade 
Descriptions 
Percent 
Description 
0 
Normal activity.  Fully active, able 
to carry on all pre-disease 
performance without restriction. 
100 
Normal, no complaints, no evidence 
of disease. 
90 
Able to carry on normal activity; 
minor signs or symptoms of disease. 
1 
Symptoms, but ambulatory.  
Restricted in physically strenuous 
activity, but ambulatory and able 
to carry out work of a light or 
sedentary nature (e.g., light 
housework, office work). 
80 
Normal activity with effort; some 
signs or symptoms of disease. 
70 
Cares for self, unable to carry on 
normal activity or to do active work. 
2 
In bed <50% of the time.  
Ambulatory and capable of all 
self-care, but unable to carry out 
any work activities.  Up and about 
more than 50% of waking hours. 
60 
Requires occasional assistance, but 
is able to care for most of his/her 
needs. 
50 
Requires considerable assistance and 
frequent medical care. 
3 
In bed >50% of the time.  Capable 
of only limited self-care, confined 
to bed or chair more than 50% of 
waking hours. 
40 
Disabled, requires special care and 
assistance. 
30 
Severely disabled, hospitalization 
indicated.  Death not imminent. 
4 
100% bedridden.  Completely 
disabled.  Cannot carry on any 
self-care.  Totally confined to bed 
or chair. 
20 
Very sick, hospitalization indicated. 
Death not imminent. 
10 
Moribund, fatal processes 
progressing rapidly. 
5 
Dead. 
0 
Dead. 
 
 
 

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APPENDIX B 
BLOOD SAMPLE COLLECTION FORM 
The University of Chicago  
 
 
 
Protocol# CIRB15-0185 
Blood Sample Collection Form    
 
Clinician/Research Nurse: Please Fill Out 
Blood Samples 
 
Patient Name: ___________________  
Week#_______ 
 
Patient Protocol ID #:______________  
Date Blood Obtained: _________________  
 
Date of Birth: _____________________  
Attending Physician: ______________  
 
Institution: _________________  
 
Cross-over samples (COS)? (circle): yes / no 
 if YES Cross-over samples: label COS-CXDX on tubes 
 
Date consent was signed: ___________   Day Started on the clinical protocol: ___________ 
 
 Visit  
Collection Tubes to 
use:  
Date 
Drawn  
Fresh Samples 
(shipped on ICE 
pack) Date Sent 
Processed MNC 
(shipped on Dry 
Ice) 
Date Sent 
Please check 
all sent 
samples  
Pre-study 
Screening 
(Four green 
top tubes 
(5-10cc each)) 
1. WT1 MRD 
 
2. T cell subsets 
 
3. TCR sequencing  
 
4. T cell PD-1 
Expression 
 
 
 
□  WT1 MRD 
 
□ T cell subsets 
 
□ TCR 
sequencing 
 
□ T cell PD-1 
Expression 
Week 9  
(Four green 
top tubes 
(5-10cc each)) 
1. WT1 MRD 
 
2. T cell subsets 
 
3. TCR sequencing 
 
4. T cell PD-1 
Expression  
 
 
 
□  WT1 MRD 
 
□ T cell subsets 
 
□ TCR 
sequencing 
 
□ T cell PD-1 
Expression 
Week 13 
(Three green 
top tubes 
(5-10cc each)) 
1. WT1 MRD 
 
2. T cell subsets 
 
3. TCR sequencing 
 
 
 
□  WT1 MRD 
 
□ T cell subsets 
 
□ TCR 
sequencing 
 
Week 25 
(Three green 
top tubes 
1. WT1 MRD 
 
2. T cell subsets 
 
 
 
□  WT1 MRD 
 
□ T cell subsets 

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(5-10cc each)) 
 
3. TCR sequencing 
 
□ TCR 
sequencing 
Week 53 
(Three green 
top tubes 
(5-10cc each)) 
1. WT1 MRD 
 
2. T cell subsets 
 
3. TCR sequencing 
 
 
 
□  WT1 MRD 
 
□ T cell subsets 
 
□ TCR 
sequencing 
 
□ T cell PD-1 
Expression 
At the time of 
off study 
(Four green 
top tubes 
(5-10cc each)) 
1. WT1 MRD 
 
2. T cell subsets 
 
3. TCR sequencing  
 
4. T cell PD-1 
Expression  
 
 
 
□  WT1 MRD 
 
□ T cell subsets 
 
□ TCR 
sequencing 
 
□ T cell PD-1 
Expression 
 
 
 

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APPENDIX C 
BONE MARROW ASPIRATION COLLECTION FORM 
 
The University of Chicago  
 
 
Protocol# CIRB15-0185 
Bone Marrow Aspiration Collection Form 
 
 
Clinician/Research Nurse: Please Fill Out 
Bone Marrow Aspiration Sample 
 
Patient Name: ___________________  
Week#_______________ 
 
Patient Protocol ID #:______________  
Date Obtained: _________________  
 
Date of Birth: _____________________  
Attending Physician: ______________  
 
Institution: _________________  
 
Cross-over samples (COS)? (circle): yes / no 
 if YES Cross-over samples: label COS-CXDX on tubes 
 
Date consent was signed: ___________   Day Started on the clinical protocol: ___________ 
 
 Visit  
Collection Tubes to 
use:  
Date 
Drawn  
Fresh 
samples 
(shipped on 
Ice pack)  
Processed MNC 
(Shipped on Dry 
Ice)  
Please check all 
sent samples  
Pre-study 
Screening 
(Four green 
top tubes 
(5-10cc 
each)) 
1. PD-L1 flow 
cytometry 
 
2. WT1 MRD 
 
3. T cell subsets/CyTOF 
 
4. TCR sequencing  
 
 
 
 
Week 13 
(Three green 
top tubes 
(5-10cc 
each)) 
1. WT1 MRD 
 
2. TCR sequencing 
 
3. T cell subsets/CyTOF 
 
 
 
 
 
Week 25 
(Three green 
top tubes 
(5-10cc 
each)) 
1. WT1 MRD 
 
2. TCR sequencing 
 
3. T cell subsets/CyTOF 
 
 
 
 
 
Week 53 
(Three green 
top tubes 
(5-10cc 
each)) 
1. WT1 MRD 
 
2. TCR sequencing 
 
3. T cell subsets/CyTOF 
 
 
 
 
 

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At the time of 
off study 
(Three green 
top tubes 
(5-10cc 
each)) 
1. WT1 MRD 
 
2. TCR sequencing 
 
3. T cell subsets/CyTOF 
 
 
 
 
 
 
 
 

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APPENDIX D 
ARCHIVED BONE MARROW BIOPSY ACQUISITION FORM 
 
The University of Chicago   
 
 
 
Protocol# 
Archived Bone Marrow Core Biopsy Acquisition Form 
 
 
 
  
Clinician/Research Nurse: Please Fill Out 
Bone Marrow Core Biopsy Tissue (Blocks/Unstained slides) 
 
Patient Name: ___________________  
Week#_______________ 
 
Patient Protocol ID #:______________  
Date Obtained: _________________  
 
Date of Birth: _____________________  
Attending Physician: ______________  
 
Origin of Tissue: __________________  
Institution: _________________  
 
Cross-over samples (COS)? (circle): yes / no 
 if YES Cross-over samples: label COS-CXDX on tubes 
 
Date consent was signed: ___________   Day Started on the clinical protocol: ___________ 
 
Researcher: Please Fill Out  
 
Date Samples received: _________________ 
 Data entered into Database: Yes      No  
 
Name of Data Manager informed: _______________________ Date Informed:_____________  
 
Visit  
Collection  
Date of biopsy 
At the diagnosis 
of initial AML 
 
1. block ____ 
 
2. Unstained Slides______  
 
 
At the time of 
study screening  
 
1. block ____ 
 
2. Unstained Slides______ 
 
 
After AML 
relapse, at the 
time of cross 
over to get 
Nivolumab 
 
1. block ____ 
 
2. Unstained Slides______ 
 
 
 
 
 

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APPENDIX E 
ARCHIVED BONE MARROW ACQUISITION FORM 
The University of Chicago  
    
 
 
Protocol# 
Archived Bone Marrow Aspiration Acquisition Form 
 
 
 
  
Clinician/Research Nurse: Please Fill Out 
Bone Marrow Aspiration (frozen bone marrow cells) 
 
Patient Name: ___________________  
Week#_______________ 
 
Patient Protocol ID #:______________  
Date Obtained: _________________  
 
Date of Birth: _____________________  
Attending Physician: ______________  
 
Origin of Tissue: __________________  
Institution: _________________  
 
Cross-over samples (COS)? (circle): yes / no 
 if YES Cross-over samples: label COS-CXDX on tubes 
 
Date consent was signed: ___________   Day Started on the clinical protocol: ___________ 
 
 
Researcher: Please Fill Out  
 
Date Samples received: _________________ 
 Data entered into Database: Yes      No  
 
Name of Data Manager informed: _______________________ Date Informed:_____________  
 
Visit  
Collection  
Date of biopsy 
Date samples 
received   
Date enter to 
database 
At the diagnosis 
of initial AML 
 
Frozen cell vial ________ 
 
 
 
 
Note: 
 
 
 
 
 
 
 
 
 
 
 
 

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APPENDIX F 
SHIPPING INSTRUCTIONS  
 
SHIPPING INSTRUCTIONS  
All shipments must contain a completed Sample Identification form and Tracking form.  
 
****Each site will be required to pay for shipping using their per- patient amounts, and 
really should only batch ship once per patient.  
 
* Green top tubes should be collected and processed as per Section 9.  
 
Paraffin sections can be sent at room temperature.  
 
The blood samples should be labeled with the patient protocol ID#, Week identifiers:  
 
 
SHIPPING METHODS 
 
1. The fresh (unprocessed) samples will be shipped on ice packs, NOT dry ice, same day or 
overnight to University of Chicago Wendy Stock laboratory for sample processing.  
 
2. For centers with ability to process the samples, MNC could be isolated using Ficoll 
method and then be frozen in 90% fetal bovine serum/10% DMSO and stored in liquid 
nitrogen or -80ºC freezer before shipment in batches to Wendy Stock laboratory 
a. Lab Address: 
W. Stock Lab 
900 East 57th Street 
KCBD Room 9160 LB49 
Chicago, IL 60637 
b. Contact Number: 
773-795-3727 
 
3. Archival tissue slides or blocks should be sent to the University of Chicago Phase II 
Consortium for storage. Please include a copy of patient pathology report with 
submission of slides, or send in email 
a. Address: 
 
Phase II Consortium 
5841 S. Maryland Ave., MC 2115 
Chicago, IL 60637 
b. Contact number: 
773-834-1746 
Email: phaseIIcra@medicine.bsd.uchicago.edu 
 
**Prior to shipment (cells, and tissue) please EMAIL Bartholomew Eisfelder at 
beisfelder@medicine.bsd.uchicago.edu, Noreen Fulton at nfulton1@bsd.uchicago.edu (fresh 

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samples) and phaseIIcra@medicine.bsd.uchicago.edu (tissue and slides) to notify that samples 
are being sent. This is a requirement.   
 
*** Please ship preferentially on Monday, Tuesday or Wednesday, to allow for arrival prior to 
weekends.  
 
Instructions for Shipping Specimens  
The Federal Aviation Administration (FAA) is an arm of the Department of Transportation 
(DOT). The DOT requires that anyone involved in the transport of hazardous materials (such as 
infectious substances, diagnostic specimens, genetically modified organisms, biological 
products, and dry ice) needs to comply with the Federal transportation law (49 CFR 172.700). 
These regulations are applicable to anyone who handles, offers for transport, and transports 
dangerous goods or causes dangerous goods to be transported. Please comply with federal 
regulations 49 CFR and IATA 1.5.  
 
Helpful Hints for Shipping  
Dry Ice: Packing Instruction  
Processed MNC of blood and bone marrow cell pellet. 
 
1. Please fill cooler at least half full with dry ice and cover specimen bag with dry ice to prevent 
thawing.  
2. Enclose embedded tissue or serum/plasma vial in a sealed biohazard bag.  
3. Include an absorbent pad within primary sealed biohazard bag.  
4. Place the primary bag in a secondary bag.  
5. Box (cooler) must be able to release gas build-up.  
6. Indicate the dry ice weight in kg on both the waybill and on the box.  
7. Include the marking "Dry Ice, UN1845" on the box.  
8. Place a Class 9 label on the box.  
 
Training courses and additional information about shipping requirements can be also found at:  
http://www.saftpak.com/  
http://hazmat.dot.gov  
 
Whole Blood MNC and cells from bone marrow aspiration 
-Please write the study number, patient initials, cycle number, and date on tubes.  
-Store in liquid nitrogen.  
 
Please note, the only thing shipped on dry ice should by already isolated/ frozen 
mononuclear cells (MNC) or RNA lysates. Unprocessed whole blood and bone marrow 
aspirates should be shipped on ice packs, NOT dry ice. 
  
Bone Marrow Core Biopsy Tissue  
Preferably, whole Tissue blocks sent to University of Chicago and processed. Blocks will be 
returned. Alternatively, please process paraffin embedded tissue as follows:  
1. For IHC- Unstained formalin fixed paraffin embedded tissue sections should be cut at 4-5 
micron thickness. Cut 20 serial sections and store at room temperature.  

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APPENDIX G 
FICOLL ISOLATION OF LYMPHOCYTES FROM BONE 
MARROW ASPIRATES AND WHOLE BLOOD SAMPLES 
 
CONSUMABLES: 
• 25, 10, and 1 mL pipettes 
• Gauze  
• 1.5 mL tubes 
• 50 mL conical tubes 
• 15 mL conical tubes 
• 15 & 50 mL conical tube racks 
• Transfer pipette 
• RPMI media or PBS (stored at 4ºC) 
• Ficoll-Paque Plus (GE Healthcare) (stored at 4ºC) 
• Isopropanol 
• 70% ethanol 
• RNA STAT-60 or TRIzol 
• Freezing Media (90% FBS, 10% DMSO; stored at -20 ºC) 
• Liquid waste container 
 
EQUIPMENT: 
• Biological Safety Cabinet 
• Centrifuge 
• Pipette aid 
• Automated cell counter 
• Controlled rate freezing container (e.g., Mr. Frosty, CoolCell, etc.) 
 
PROCEDURE 
 
1. Once samples have been received, information on patient sheet including study name, 
MRN, patient ID and/or initials, date of sample, sample timepoint (pretreatment, etc.) and 
tissue type (BM/PB) should be cross-checked with the labels on the samples. 
 
2. Prepare centrifuge: 400 g, room temperature with brakes OFF. 
 
3. Biological safety cabinet should be UV-sterilized for at least 15 minutes and wiped down 
with 70% ethanol prior to beginning. 
 
4. Label all 50, 15, and 1.5 mL conical tubes with sample ID and tissue type. 
 
5. Aliquot 20 mL cold Ficoll-Paque Plus to a 50 mL tube for each sample.  
 
6. Using gauze, carefully open PB/BM samples. Immediately discard cap and gauze in 
biohazardous waste. 

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7. Pour up to 20 mL of PB/BM sample into empty 50 mL conical tubes. Record volume on 
sample sheet. 
• If sample volume is greater than 20 mL, split sample equally into two 50 mL 
conical tubes. 
 
8. Add cold RPMI or PBS to sample to bring volume up to 30 mL using a 25 mL pipette, 
and mix three times.  
 
9. Set Pipet-Aid to SLOW. Overlay sample onto Ficoll dropwise. Hold pipette tip at a 
slanted angle on the inside edge of the conical tube while dispensing at a steady and 
constant rate without mixing/penetrating Ficoll layer. 
 
10. Centrifuge for 30 minutes at 400 g, room temperature with brakes OFF. 
 
 
 
 
 
 
 
 
 
 
11. Following centrifugation, use a transfer pipette to transfer entire lymphocyte layer into an 
empty 15 mL tube.  If samples of the same tissue type were split in Step 8, they may be 
recombined to one 15 mL tube. 
 
12. Wash by adding RPMI or PBS to the 15 mL conical tube containing lymphocytes to 
bring volume up to 14 mL. 
 
13. Centrifuge for 15 minutes at 400 g, room temperature with brakes ON.  
 
 
14. Remove supernatant using a transfer pipette. Resuspend cells with RPMI/PBS and count 
cells using automated cell counter. 
 
15. Depending on the number of cells, samples may be frozen for viability and lysed for 
RNA directly. If cell yields are poor, RNA takes priority. Guidelines for this study 
are: 
 
 
 
 
Sample 
Minimum 
Cell Number 
Reagent Used 
Parameters 

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WT1 MRD* 
2 x 106 
RNA STAT-60 or 
TRIzol 
<5 x 106 cells – lyse with 0.5 mL 
5-10 x 106 cells – lyse with 1 mL 
T-Cell Subset 
2 x 106 
Freezing Media 
(90% FBS, 10% 
DMSO) 
1 mL Freezing Media per cryovial 
No more than 10 x 106 cells per vial 
TCR 
Sequencing* 
5 x 106 
RNA STAT-60 or 
TRIzol 
<5 x 106 cells – lyse with 0.5 mL 
5-10 x 106 cells – lyse with 1 mL 
 
*If cell yield is poor, cells for RNA (WT1 and TCR sequencing) can be combined into 
one lysate. 
 
 
16. For cells frozen for viability (T-Cell Subset): 
 
• Centrifuge cells at 400 g, room temperature for 5 minutes. 
 
• Remove supernatant and resuspend with Freezing Media. Aliquot 1 mL into each 
labeled cryovial. 
 
• Place cryovials in controlled rate freezing container, and store at -80ºC overnight. 
Transfer to liquid nitrogen storage the following morning. 
 
 
17. If cells are going to be made into RNA lysates: 
 
• Centrifuge cells at 400 g, room temperature for 5 minutes. 
 
• Label 1.5 mL tubes: 
 
                  Acc. # and Patient ID 
                  GITC (Tissue Type (BM or PB)) 
                  Date 
 
• Remove supernatant and lyse cells with RNA STAT-60 or TRIzol according to 
parameters listed above. 
• <5 x 106 cells should be lysed with 0.5 mL RNA STAT-60 or TRIzol 
• 5-10 x 106 cells should be lysed with 1 mL RNA STAT-60 or TRIzol 
 
• Store lysates at -80ºC. 
 
 
 
 
 
 

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APPENDIX H 
RISKS ASSOCIATED WITH PREGNANCY 
 
Risks Associated with Pregnancy:  
The toxicity of Nivolumab was tested in the pregnant cynomolgus monkeys (Investigator 
Brochure.  (2014). While Nivolumab was well tolerated by the pregnant monkeys, in the 
offspring, maternal nivolumab administration was associated with fetal/neonatal mortality 
characterized by: 1) dose-dependent increases in third trimester fetal losses and increased 
neonatal mortality at higher dose. The cause(s) of these fetal losses and infant prematurity could 
not be determined. The teratogenic effect of Nivolumab in humans cannot be ruled out. 
Therefore, a risk minimization plan to prevent pregnancy must be observed. 
 
Criteria for females of childbearing potential (FCBP)  
This protocol defines a female of childbearing potential as a sexually mature woman who: 1) has 
not undergone a hysterectomy or bilateral oophorectomy or 2) has not been naturally 
postmenopausal (amenorrhea following cancer therapy does not rule out childbearing potential) 
for at least 24 consecutive months (i.e., has had menses at any time in the preceding 24 
consecutive months).  
 
Counseling  
For a female of childbearing potential, Nivolumab is contraindicated unless all of the following 
are met (i.e., all females of childbearing potential must be counseled concerning the following 
risks and requirements prior to the start of Nivolumab):  
•  
She understands the potential teratogenic risk to the unborn child  
•  
She understands the need for effective contraception, without interruption, throughout 
the entire duration of study treatment, dose interruption and 23 weeks after the end of study 
treatment  
•  
She should be capable of complying with effective contraceptive measures  
•  
She is informed and understands the potential consequences of pregnancy and the 
need to notify her study doctor immediately if there is a risk of pregnancy  
•  
She understands the need to commence the study treatment as soon as study drug is 
dispensed following a negative pregnancy test  
•  
 

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•  
She acknowledges that she understands the hazards and necessary precautions 
associated with the use of Nivolumab  
 
The investigator must ensure that for females of childbearing potential:  
• 
 Complies with the conditions for pregnancy risk minimization, including confirmation 
that she has an adequate level of understanding  
• 
 Acknowledge the aforementioned requirements  
 
The effect of Nivolumab on sperm is not clear at this time. Male patients taking Nivolumab must 
meet the following conditions (i.e., all males must be counseled concerning the following risks 
and requirements prior to the start of Nivolumab):  
•  
Understand the potential teratogenic risk if engaged in sexual activity with a pregnant 
female or a female of childbearing potential  
•  
Understand the need for the use of a condom even if he has had a vasectomy, if 
engaged in sexual activity with a pregnant female or a female of childbearing potential.  
 
Contraception  
 
Females of childbearing potential (FCBP) enrolled in this protocol must agree to use two reliable 
forms of contraception simultaneously or to practice complete abstinence from heterosexual 
contact during the following time periods related to this study: 1) for at least 28 days before 
starting study drug; 2) while participating in the study; 3) during dose interruptions; and 4) for at 
least 23 weeks after study treatment discontinuation.  
 
The two methods of reliable contraception must include one highly effective method and one 
additional effective (barrier) method. FCBP must be referred to a qualified provider of 
contraceptive methods if needed. The following are examples of highly effective and additional 
effective methods of contraception:  
 
Highly effective methods:  
− Intrauterine device (IUD)  
− Hormonal (birth control pills, injections, implants)  
− Tubal ligation  
− Partner’s vasectomy  
 
Additional effective methods:  
− Male condom  
− Diaphragm  
− Cervical Cap 
 
 
 
 

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Pregnancy testing  
Medically supervised pregnancy tests with a minimum sensitivity of 25 mIU/mL must be 
performed for females of childbearing potential, including females of childbearing potential who 
commit to complete abstinence, as outlined below.  
 
Before starting study drug  
 
Female Patients:  
Women of childbearing potential must have a negative serum or urine pregnancy test (minimum 
sensitivity 25 IU/L or equivalent units of HCG) within 24 hours prior to the start of nivolumab. 
The patient may not receive study drug until the study doctor has verified that the results of these 
pregnancy tests are negative.  
 
Male Patients:  
Must practice complete abstinence or agree to use a condom during sexual contact with a 
pregnant female or a female of childbearing potential while participating in the study, during 
dose interruptions and for at least 31 weeks (or 7 months) following study drug discontinuation, 
even if he has undergone a successful vasectomy.  
 
 
 
 
 
 

93 
APPENDIX I 
MANAGEMENT ALGORITHMS FOR ENDOCRINOPATHY, 
GASTROINTESTINAL, HEPATIC, NEUROLOGICAL, 
PULMONARY, RENAL, AND SKIN ADVERSE EVENTS

94 
 

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97 

98 

99 

100 
 
 
 

101 
APPENDIX J 
AML RISK CATEGORY GUIDELINES 
 
Prognostic Group 
Subsets 
Favorable 
Inv (16) or t(16;16);t(8;21); NK 
NPM1+FLT3 ITD‐;NK and 
CEBPA+/+ 
Intermediate -1 
NK and NPM‐/FLT3 ITD‐ or 
CEBPA+/‐ 
Intermediate- 2 
Cytogenetic abnormalities not in 
“favorable” or “adverse” 
groups;FLT3 ITD positive 
Adverse 
−5, −7,5q‐, abn 3q,17p,11q (other 
than 9;11), t(6;9) complex cyto; 
Insufficient metaphases for analysis; 
mutations in TP53, ASXL1, RUNX1 
  
 
 
Estey, Elihu (2016)