A Pilot Study of the Combination of 5-azacitidine (5-AZA) and All-trans Retinoic Acid (ATRA) for Prostate Cancer (PCa) With PSA-only Recurrence After Definitive Local Treatment
Phase: PHASE2
Conditions: Prostatic Neoplasms, Prostate Neoplasms, Prostate Cancer
Sponsor: Icahn School of Medicine at Mount Sinai

A Pilot Study of the Combination of 5-azacitidine (5-AZA) and All-trans Retinoic Acid (ATRA) 
for Prostate Cancer (PCa) With PSA-only Recurrence After Definitive Local Treatment  
Dr. Vaibhav G Patel
NCT03572387
Document Date: Jan 17, 2023

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 .
A pilot study of the combination of 5-azacitidine (5-AZA) and all-trans retinoic acid 
(ATRA) for prostate cancer (PCa) with PSA-only recurrence after definitive local 
treatment
Principal Investigator:
Bobby Liaw, MD
Department of Medicine, Division of Hematology Oncology
Icahn School of Medicine at Mount Sinai
Co-Investigator(s):
Matthew Galsky, MD
Department of Medicine, Division of Hematology and Oncology
Icahn School of Medicine at Mount Sinai
Che-Kai Tsao, MD
Department of Medicine, Division of Hematology Oncology
Icahn School of Medicine at Mount Sinai
Biostatistician:
Rachel Jia
Department of Population Health and Policy
Icahn School of Medicine at Mount Sinai
Study Drug:
All-trans retinoic acid, 5-azacitidine
Funding Source: 
The V Foundation for Cancer Research – Translational Research Award
Initial version:
31MAR 2017
Current Version:
Version9.0
LIST OF ABBREVIATIONS
ADT
Androgen Deprivation Therapy
AE
Adverse Event
ASCO
American Society of Clinical Oncology
AST
Aspartate Aminotransferase
BM
Bone Marrow
Effective Date: 1/17/2023
End Date:1/16/2024

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CBC
Complete Blood Count
CMP
Comprehensive Metabolic Panel
CR
Complete Response
CT CA/P
Computed Tomography Scan of Chest Abdomen, and Pelvis
CTCAE
Common Terminology Criteria for Adverse Events
DLT
Dose Limiting Toxicity
DSMB
Data and Safety Monitoring Board
DTCs
Disseminated Tumor Cells
EC
Ethics Committee
ECG
Electrocardiogram
ECOG
Eastern Cooperative Oncology Group
FDA
Food and Drug Administration
GCP
Good Clinical Practice
HIPAA
Health Insurance Portability and Accountability Act
IB
Investigator Brochure
IND
Investigational New Drug
IRB
Institutional Review Board
LDH
Lactate Dehydrogenase
mCRPC
Metastatic Castration Resistant Prostate Cancer
MRI
Magnetic Resonance Imaging
NIST
National Institute of Standard and Technology
PFS
Progression Free Survival
PHI
Protected Health Information
PI
Principle Investigator
PR
Partial Response
PSA
Prostate Specific Antigen
PSADT
Prostate Specific Antigen Doubling Time
PCWG2
Prostate Cancer Working Group 2
RECIST
Response Evaluation Criteria in Solid Tumors
rPFS
Radiographic Progression Free Survival
RSI
Reported Safety Information
SAE
Serious Adverse Event
SD
Stable Disease
SRM
Standard Reference Material
TTP
Time to Tumor Progression
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End Date:1/16/2024

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STUDY SCHEMA
STUDY SUMMARY
Title
A pilot study of the combination of 5-azacitidine (5-AZA) and all-trans 
retinoic acid (ATRA) for prostate cancer with PSA-only recurrence
Short Title
Prostate Cancer Dormancy
Protocol Number
Phase
Pilot, phase 2
Methodology
Randomized, open label
Study Center(s)
Single-center
Objectives
Primary 
1) Evaluate disease progression-free rate at the end of Treatment 
Phase 1 between 5-AZA+ATRA and no-therapy.
2) Assess safety of the 5-AZA and ATRA combination therapy
Secondary:
1)
Evaluate time to disease progression from start of Treatment Phase 
1
2)
Correlate antitumor activity and disease progression with biomarkers 
of dormancy in BM aspirates and peripheral blood
Number of Subjects
20 evaluable patients; the target accrual number is set at 30 to account 
for screen failures.
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End Date:1/16/2024

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Diagnosis and Eligibility 
Criteria
Diagnosis: Histologically confirmed adenocarcinoma of the prostate
Main inclusion criteria: 1) Rising PSA 2) PSADT < 10 months prior to 
initiation of ADT 2)  3) Received definitive local treatment 4) Indication for 
ADT 5) No evidence of regional or active distant metastases, except for 
regional metastases where salvage radiation therapy is not an option
Main exclusion criteria: 1) Patients who have received ADT and/or other 
chemotherapy within 3 months prior to entering the study. 2) Patients who 
have had radiotherapy or surgery within 4 weeks prior to entering the 
study. Minimally-invasive procedures for the purpose of diagnosis or 
staging of the disease are permitted.
Study Product(s), Dose, 
Route, Regimen
5-azacitidine: 40 mg/m2 SQ on days 1-5
All-trans retinoic acid:  45 mg/m2 PO daily (divided into 2 doses) on days 
3-7
One cycle of treatment: 28 days
Duration of administration
Drugs will be administered on a 28-day cycle
Reference therapy
No reference therapy
Statistical Methodology
We will compare the disease progression-free rate at end of treatment 
phase 1 and time to disease progression from start of Treatment Phase 
1 between 5-AZA+ATRA and no-therapy. Descriptive summary will be 
provided for safety endpoints and dormancy marker panel. The effect of 
biomarker abundance on recurrence free survival will be determined 
using Kaplan-Meier curves stratified across biomarker abundance 
groups. 
Estimated enrollment 
period
12
Estimated study duration
24 months
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End Date:1/16/2024

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TABLE OF CONTENTS
STUDY SCHEMA .............................................................................................................4 
STUDY SUMMARY ..........................................................................................................4
1.
BACKGROUND AND RATIONALE...................................................................7
2.
STUDY OBJECTIVES ..........................................................................................8
3.
STUDY DESIGN ....................................................................................................8
4.
ELIGIBILITY CRITERIA .....................................................................................11
5.
DRUG INFORMATION .......................................................................................12
6.
ADVERSE EVENTS............................................................................................16
7.
FOLLOW UP, LABORATORY TESTING, AND DIAGNOSTIC IMAGING19
8.
STATISTICAL CONSIDERATIONS .................................................................21
9.
DATA HANDING AND RECORD KEEPING ..................................................22
10.
ETHICAL CONSIDERATIONS..........................................................................25
11.
FINANCIAL CONSIDERATIONS......................................................................26
12.
REFERENCES .....................................................................................................27
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1.
BACKGROUND AND RATIONALE
1.1
Disease Background
Prostate cancer (PCa) causes significant morbidity and mortality in the US, disproportionally affecting 
African American men. The initial treatment for PCa consists of surgical resection and radiation therapy 
(RT). While these approaches reduce the primary tumor burden, the majority of patients have clinical 
recurrence and eventually die of metastases several years later. One explanation is the presence of 
disseminated tumor cells (DTCs) that are resistant to chemotherapy and ADT, but maintain the capacity to 
reconstitute incurable metastatic disease later in life. In prior studies, DTCs have been shown to survive in 
BM of PCa patients in a dormant or quiescent (G0/G1 arrest) state and evade therapies.1 DTCs can be 
detected in the BM before overt disease is detected, suggesting that DTCs may help us understand how 
dormancy and reactivation to metastasis occurs. The mechanisms responsible for the onset of dormancy 
and the transition from DTC dormancy to reactivation have been mostly unknown.2 Thus, a window of 
opportunity to target quiescent residual DTCs for elimination or maintenance of dormancy is missed. Our 
team has pioneered the study of cancer dormancy to understand the biology of DTCs. Based on prior 
literature and our pre-clinical models, we have developed a treatment strategy with available FDA-
approved drugs (5-azacitidine and all-trans retinoic acid) to reprogram DTCs into a dormant state and 
ultimately, prolong time to clinical disease progression. 
1.2
Rationale
Our long-term goal is to target the biology of dormant PCa DTCs to prevent relapse of clinical disease. In 
pre-clinical models, specific micro-environmental cues have been shown to activate a transcription factor 
(TF) network, which drives epigenetic programs of quiescence and survival in dormant tumor cells.1, 3-5 
Specifically, ATRA could induce a high p38/ERK activity ratio and dormancy in previously proliferative 
DTCs through TGF2 signaling in the BM and through upregulation of BMP7.6 The p38 activation also 
induces TFs DEC2 and NR2F1 to induce a dormant state.4 Overall, the dormancy TF network serves as a 
signature to predict longer metastasis-free periods and to identify dormant DTCs recovered from BM of 
prostate cancer patients. Interestingly, many dormancy genes are epigenetically silenced and can be re-
expressed using a short-term pulse of low dose 5-AZA, a DNA methylating agent and DNMT1 inhibitor.7, 8 
When we introduced 5-AZA and ATRA in malignant prostate cancer cell lines for three days, many cells 
displayed upregulation of dormancy biomarkers and the response was maintained for at least 2 weeks.1
Our pre-clinical work led us to design a “reprogramming therapy” combining two available FDA-approved 
drugs (5-AZA, ATRA) to reprogram proliferative cells into stable induced dormant cancer cells (iDCCs).1 
Prior studies with the drugs of interest were performed in castration-sensitive PCa patients with prior 
exposure to ADT and with fast PSA doubling time (PSADT).9-11 We will test if 5-AZA+ATRA extends the 
progression-free period and delays biochemical and clinical recurrence via induction of PCa cell dormancy. 
This study will allow us to temporally understand the impact of the reprogramming therapy on the delay in 
chemical and clinical progression, and to correlate clinical activity with biomarkers of dormancy in DTCs.
Currently, patients who have received definitive local treatment with either RT or surgery undergo serial 
measurements of PSA to detect early disease recurrence. Such monitoring leads to the identification of 
men with PSA-only (biochemical) recurrence without evidence of clinical symptoms, or radiographic 
evidence of recurrent or disseminated disease. Androgen deprivation therapy (ADT) is the standard of care 
for the initial systemic therapy, however the optimal timing for initiation of ADT remains controversial due to 
paucity of clinical trial data in this population.12 Furthermore, ADT causes side effects such as hot flashes, 
loss of libido, decreased muscle mass, fatigue, and osteoporosis, which can significantly lower the quality 
of life in otherwise asymptomatic men.
Leuprolide is considered standard of care for prostate cancer with PSA-only recurrence.  One dose of 
Leuprolide has a transient effect of decreasing PSA to negligible levels resulting in a similar baseline for 
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both groups. Based on historical data, in measuring primary endpoint at 12 weeks, we do not expect 
Leuprolideto impact disease progression. 
To this end, we will conduct a randomized, pilot study in recurrent PCa patients with biochemical 
recurrence, as defined by rising PSA, of the combination of 5-azacitidine and all-trans retinoic acid to delay 
disease progression.
1.3
Correlative Studies
There is prior literature that shows that DTCs in head and neck squamous cell carcinoma4 and PCa 
models6 could persist in a dormant state in the BM. Mechanistically, we found that TGF2 mRNA was 
enriched in naïve BM and in PCa DTCs from patients with no evidence of disease for up to 18 years vs. 
DTCs from advanced metastatic patients.3, 4 Upregulation of TGF2 signaling leads to activation of p38 
and its associated quiescence program involving DEC2 and NR2F1.1 Furthermore, we showed that NR2F1 
induces RARb and patients with high levels of NR2F1 and RARb showed longer disease free periods than 
those with low levels of these transcription factors.1, 6 We also found that inducible knock down of NR2F1 
reactivated DTCs in different organs.1
DNA promoter methylation and/or changes in histone-H3 post-translational modifications (PTMs) 
contribute to NR2F1 silencing proliferative tumor cells and thus, drugs can be used to remodel the 
epigenome and restore dormancy.1 ATRA was sufficient to induce NR2F1 and RARb but only transiently, 
suggesting that other epigenetic changes might be needed to sustain dormancy.1 Whole genome changes 
in histone-H3 PTMs occur after low-dose 5-AZA and can be used to rewrite the epigenome for growth 
suppression.7, 8 Many aggressive cancer cells including prostate cells treated with 5-AZA+ATRA displayed 
upregulation of dormancy biomarkers including NR2F1 and RARb mRNAs.1 We hypothesize that the 
reprogramming of residual prostate cancer with 5-AZA and ATRA will be a new strategy to prevent or 
significantly delay time to clinical recurrence compared to current therapies.
2.
STUDY OBJECTIVES
To our knowledge, no study has evaluated the combination of 5-AZA and ATRA in recurrent PCa. Our 
hypothesis is that this combination can reprogram DTCs to activate the dormancy pathway and thus delay 
disease progression, and can be safely tolerated.
2.1
Primary Objectives
The co-primary objectives of the study are:
o
To compare disease progression-free rate at end of Treatment Phase 1 between 5-AZA + ATRA and 
no-therapy.
o
To assess safety of the 5-AZA and ATRA combination therapy
2.2
Secondary Objectives 
The secondary objectives are:
o
To compare time to disease progression from start of Treatment Phase 1 during the 24-month study 
period between 5-AZA+ATRA and no-therapy. 
o
To correlate antitumor activity and progression with biomarkers of dormancy in BM aspirates and 
peripheral blood.
3.
STUDY DESIGN
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3.1
General Design
This is a prospective, open-label, randomized, pilot study of reprogramming therapy in patients with 
recurrent PCa based on rising PSA only. Reprogramming therapy will include 5-azacitidine and all-trans 
retinoic acid. Expected duration of subject participation is 24 months.
All study will be assigned in a 1:1 randomization using computer random number generator to either the ‘5-
AZA+ATRA’ group or the ‘no therapy’ group. All enrollees will receive Leuprolide 7.5 mg x 1 on Day 1. On 
Day 29, they will initiate treatment phase 1. During this phase, patients in the ‘5-AZA + ATRA’ group will 
receive treatment on a 28-day cycle, in the absence of prohibitive toxicities, for 3 cycles. During treatment 
phase 2, the ‘no therapy’ group will receive 5-AZA + ATRA on a 28-day cycle, in the absence of prohibitive 
toxicities, for 3 cycles. The treatment effect will be determined via comparing the intra-individual disease 
progression outcome at the end of the treatment phase 1, with period effect being separated through 
randomization.17 After treatment phase 1 and phase 2, all patients will be followed for a total of 24 months 
from the start of the study or until the events leading to discontinuation as outlined in Section 7.1 are 
observed. 5-AZA will be given subcutaneously on days 1-5 at a dose of 40 mg/m2. ATRA, dosed at 45 
mg/m2 daily (divided into 2 doses), will be taken orally on days 3-7 of each cycle.
3.2
Primary study endpoints
The primary clinical endpoints of this study are the disease progression-free rate at the end of treatment 
phase 1 and safety of the combination of 5-AZA and ATRA. Disease progression is defined as a composite 
of PSA or radiographic progression. 
3.2.1
PSA Progression
PSA Progression is as defined by PCWG2 criteria.13 Baseline PSA will be defined as the serum PSA level 
measured prior to start of treatment.
In patients who have a decline in PSA value from baseline, progression is defined by:
o
An increase in PSA by 25% above the nadir, AND
o
An increase in PSA by a minimum of 2 ng/ml, or an increase in PSA to the pre-treatment PSA value, 
AND
o
Confirmation by a second PSA at least 3 weeks apart, AND
o
There is no objective evidence of disease response.
In patients whose PSA value has not declined from baseline, progression is defined by:
o
An increase in PSA by 25% above either the pre-treatment level, or the nadir PSA level (whichever is 
lowest), AND
o
An increase in PSA by a minimum of 2 ng/ml, AND
o
Confirmation by a second PSA at least 3 weeks apart, AND
o
There is no objective evidence of disease response.
3.2.2
Radiographic Progression
Radiographic progression will be evaluated by Response Evaluation Criteria in Solid Tumors (RECIST) 
version 1.1 criteria, with definitions of CR, PR, SD, and PD for target and non-target lesions as defined 
below.  Baseline imaging defined as the CT and bone scan obtained prior to starting treatment. All 
baseline evaluations should be performed as closely as possible to the beginning of treatment and never 
more than 6 weeks before the beginning of the treatment.
Lesions are categorized as either measurable or non-measurable based on the criteria below.  All 
measurements should be taken and recorded in metric notation using a ruler or calipers. Tumor lesions 
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that are situated in a previously irradiated area will not be considered measurable. The same method of 
assessment and the same technique should be used to characterize each identified and reported lesion at 
baseline and during follow-up.
Target Lesions
All measurable lesions up to a maximum of 5 lesions per organ and 10 lesions in total, representative of all 
involved organs, should be identified as target lesions and recorded and measured at baseline. Target 
lesions should be selected on the basis of their size (lesions with the longest diameter) and their suitability 
for accurate repeated measurements (either by imaging techniques or clinically). A sum of the longest 
diameters (LD) for all target lesions will be calculated and reported as the baseline sum LD. The baseline 
sum LD will be used as reference by which to characterize objective tumor response.
Non-Target Lesions
All other lesions (or sites of disease) should be identified as non-target lesions and should also be 
recorded at baseline.  Non-target lesions include measurable lesions that exceed the maximum numbers 
per organ or total of all involved organs as well as non-measurable lesions. Measurements of these lesions 
are not required, but the presence or absence of each should be noted throughout follow-up.
3.2.2.1
Response Criteria for Target Lesions
Complete Response (CR): Disappearance of all target lesions.  Any pathological lymph nodes (whether 
target or non-target) must have reduction in short axis to <10 mm.
Partial Response (PR): At least a 30% decrease in the sum of the longest diameter (LD) of target lesions, 
taking as reference the baseline sum LD.
Progressive Disease (PD): At least a 20% increase in the sum of the LD of target lesions, taking as 
reference the smallest sum LD recorded since the treatment started.  The sum must also demonstrate an 
absolute increase of at least 5 mm.  The appearance of one or more new lesions is also considered 
progression.
Stable Disease (SD): Neither sufficient shrinkage to qualify for PR nor sufficient increase to qualify for PD, 
taking as reference the smallest sum LD since the treatment started.
3.2.2.2
Response Criteria for Non-Target Lesions
Complete Response (CR): Disappearance of all non-target lesions and normalization of tumor marker 
level.  All lymph nodes must be non-pathological in size (<10 mm short axis).
Incomplete Response/Stable Disease (SD): Persistence of one or more non-target lesion(s) and/or 
maintenance of tumor marker level above the normal limits.
Progressive Disease (PD): Appearance of two or more new lesions and/or unequivocal progression of 
existing non-target lesions.
Although a clear progression of “non-target” lesions only is exceptional, in such circumstances, the opinion 
of the treating physician should prevail, and the progression status should be confirmed at a later time by 
the Principal Investigator.
3.2.2.3
Outcomes Based on Radionuclide Bone Scans
The subjectivity in interpreting serial changes in radionuclide bone scan is well recognized.  The primary 
outcome will be whether the scan is stable or improved vs. worse or with evidence of progression.  
Changes in intensity will not be used as an outcome measure.  
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Stable/Improved: A stable or improved classification requires that no new lesions appear or that new pain 
has not developed in an area that was previously visualized.
3.2.3
Safety
Safety and tolerability of the combination of 5-AZA and ATRA will be assessed by the recording of adverse 
events, monitoring of vital signs and physical examinations, safety laboratory evaluations, and 12-lead 
ECG. Adverse events will be graded according to the Common Terminology Criteria for Adverse Events 
(CTCAE) version 4.03.14 This will be assessed at baseline examination and every 4 weeks. 
3.3
Secondary Study Endpoints
Secondary end points include time to tumor progression from start of treatment phase 1 and correlation of 
antitumor activity and tumor progression with levels of dormancy biomarkers.
3.3.1
Time to disease progression
Time to disease progression is defined as a composite of either PSA or radiographic progression, 
whichever occurs first. Please refer to 3.2 for further details.
3.3.2
Dormancy biomarkers
We will measure the dormancy biomarkers TGF2, BMP7, BMP4, GAS6, retinoic acid, NR2F1, in BM 
and/or blood supernatants of our study patients. We will correlate the levels of these biomarkers with 
clinical response to treatment and disease progression. 
4.
ELIGIBILITY CRITERIA
4.1
Patient Eligibility
Eligibility waivers are not permitted. Subjects must meet all of the inclusion and must not meet any of the 
exclusion criteria to be registered to the study. Study treatment may not begin until a subject is registered.
4.1.1 Inclusion Criteria
o
Histologically confirmed adenocarcinoma of the prostate
o
Rising PSA
o
PSADT < 10 months prior to initiation of ADT
o
No evidence of regional or active distant metastases, except for regional metastasis where salvage 
radiation therapy is not an option
o
Indication for ADT after receiving definitive local therapy
o
Males ≥ 18 years.
o
ECOG performance status of < 2
o
Men must agree to use a condom and not father a child or donate sperm for the duration of the study 
and for 90 days after completion of therapy
o
Ability to understand and the willingness to sign a written informed consent
o
Ability to adhere to the study visit schedule and requirements of the protocol
4.1.2 Exclusion Criteria
o
Patients who have received ADT and/or other chemotherapy within 3 months prior to entering the 
study.
o
Patients who have had radiotherapy or surgery within 4 weeks prior to entering the study. Minimally-
invasive procedures for the purpose of diagnosis or staging of the disease are permitted.
o
Patients may not be receiving any other investigational agents.
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o
Patients with known brain metastases should be excluded from this clinical trial because of their poor 
prognosis and because they often develop progressive neurologic dysfunction that would confound the 
evaluation of neurologic and other adverse events.
o
History of allergic reactions attributed to compounds of similar chemical or biologic composition to 5-
AZA and ATRA.
o
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.
o
Significant active cardiac disease within the previous 6 months
o
Inadequate organ and marrow function as defined below:
- leukocytes
< 3,000/mcL
- absolute neutrophil count
< 1,500/mcL
- platelets
< 100,000/mcl
- total bilirubin
above normal institutional limits
- AST(SGOT)/ALT(SPGT)
> 2.5 X institutional upper limit of normal
- creatinine 
above normal institutional limits
4.2
Pretreatment Evaluation
All patients must sign a written informed consent form before study specific screening procedures are 
performed.  Informed consents may be obtained up to 30 days prior to day 1 of study.  Screening 
procedures to evaluate patient eligibility for the study will be conducted within 30 days prior to day 1.  The 
pretreatment evaluation includes a complete medical history and physical examination, laboratory studies 
including complete blood count (CBC) with differential, comprehensive metabolic panel (CMP), lactate 
dehydrogenase (LDH), PSA, testosterone panel, and baseline EKG. A bone scan, and CT chest 
abdomen/pelvis will also be performed up to 60 days prior to Day 1 of treatment. 18F-Fluciclovine 
(Axumin) PET/CT may be substituted for pre-treatment CT chest/abdomen/pelvis and bone scan and can 
be obtained up to 60 days prior to Day 1 of treatment. If the patient meets eligibility, they will return on Day 
1 for Leuprolide administration.
5.
DRUG INFORMATION
5.1
5-azacitidine
Please see product package insert for complete details. 
5.1.1
Availability
5-azacitidine is commercially available
5.1.2
Formulation
5-azacitidine is available as a reconstituted suspension in vials containing 100 mg of 5-azacitidine. 
Commercial supplies of 5-azacitidine will be used for the study
5.1.3
Storage, preparation, and administration
Azacitidine is associated with a moderate emetic potential; antiemetics are recommended to prevent 
nausea and vomiting.
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SubQ: The manufacturer recommends equally dividing volumes >4 mL into 2 syringes and injecting into 2 
separate sites; however, policies for maximum SubQ administration volume may vary by institution; 
interpatient variations may also apply. Rotate sites for each injection (thigh, abdomen, or upper arm). 
Administer subsequent injections at least 1 inch from previous injection sites; do not inject into tender, 
bruised, red, or hard areas. Allow refrigerated suspensions to come to room temperature (up to 30 
minutes) prior to administration. Resuspend by inverting the syringe 2 to 3 times and then rolling the 
syringe between the palms for 30 seconds.
5.1.4
Mechanism of Action
Promotes hypomethylation of DNA to restore normal gene differentiation and proliferation.
5.1.5
Drug dosing rationale
The proposed dosing of 5-AZA is based on the FDA-approved doses for MDS. 5-AZA has been tested in 
phase II trials of PCa patients. Sonpavde et al10 conducted a phase II trial in 26 patients with CRPC and 
PSADT < 3 months. A dose of 40 mg/m2 SQ on days 1-5 was administered every 28 days.  The primary 
endpoint was prolongation of PSADT by greater than 3 months and was noted in 19 patients (55%).  PSA 
declines were noted in 14 of 36 evaluable patients.  Median PSADT was 2.8 months after treatment, 
compared with 1.5 months prior to starting (p<0.01).  
5-azacitidine is generally well tolerated, as demonstrated by a phase II trial in chemo-naive castration-
resistant metastatic prostate cancer.10 The toxicities were generally mild and manageable. In those 
patients with toxicities, the most common were fatigue (41.2%), constipation (41.2%), nausea (35.3%), 
injection site reaction (32.4%), vomiting (26.5%), anorexia (20.6%), anemia (20.6%), and neutropenia 
(17.6%). The most common grade 3 toxicities were fatigue (11.8%) and neutropenia (5.9%). There were 
no grade 4 toxicities. Only 4 out of 36 patients discontinued therapy due to toxicities.
5.1.6
Adverse Events
Incidence rates of adverse events associated with 5-azacitidine are provided in the product package insert. 
The most common adverse events treatment are listed below.
Cardiovascular: Peripheral edema (7% to 19%), chest pain (16%)
Central nervous system: Fatigue (13% to 36%), rigors (26%), headache (22%), dizziness (19%), anxiety 
(5% to 13%), depression (12%), malaise (11%), pain (11%), insomnia (9% to 11%)
Dermatologic: Erythema (7% to 17%), pallor (16%), skin lesion (15%), skin rash (10% to 14%), pruritus 
(12%), diaphoresis (11%)
Endocrine & metabolic: Weight loss (≤16%), pitting edema (15%), hypokalemia (6% to 13%)
Gastrointestinal: Nausea (48% to 71%), vomiting (27% to 54%), constipation (34% to 50%), diarrhea 
(36%), anorexia (13% to 21%), abdominal pain (11% to 16%), abdominal tenderness (12%)
Hematologic & oncologic: Thrombocytopenia (66% to 70%; grades 3/4: 58%), anemia (51% to 70%; 
grades 3/4: 14%), neutropenia (32% to 66%; grades 3/4: 61%), leukopenia (18% to 48%; grades 3/4: 
15%), bruise (19% to 31%), petechia (11% to 24%), febrile neutropenia (14% to 16%; grades 3/4: 13%), 
bone marrow depression (nadir: days 10 to 17; recovery: days 28 to 31)
Local: Injection site reactions (14% to 29%): Erythema (35% to 43%; more common with IV 
administration), pain (19% to 23%; more common with IV administration), bruising (5% to 14%)
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Neuromuscular & skeletal: Weakness (29%), arthralgia (22%), limb pain (20%), back pain (19%), myalgia 
(16%)
Respiratory: Cough (11% to 30%), dyspnea (5% to 29%), pharyngitis (20%), epistaxis (16%), 
nasopharyngitis (15%), upper respiratory infection (9% to 13%), pneumonia (11%), rales (9% to 11%)
Miscellaneous: Fever (30% to 52%)
5.2
All-trans retinoic acid
Please see product package insert for complete details. 
5.2.1
Availability
ATRA is commercially available
5.2.2
Formulation
ATRA is available as an oral capsule of 10 mg.
5.2.3
Storage, preparation, and administration
Dosage calculations will be rounded to the nearest 10 mg increment and given in two divided doses per 
day.
ATRA will be taken orally with meals.
5.2.4
Mechanism of Action
Promotes hypomethylation of DNA to restore normal gene differentiation and proliferation.
5.2.5
Drug dosing rationale
The proposed dosing of ATRA is based on the FDA-approved doses for acute promyelocytic leukemia. 
ATRA has been studied in several phase II trials. Trump et al11 gave ATRA 50 mg/m3 daily  x 14 days 
every 3 weeks to 17 CRPC patients and saw no clinical responses. However, Kelly et al9 showed in a trial 
of ATRA 45 mg/m2 daily combined with cis-retinoic acid + IFN that 4 patients with hormone-sensitive PCa 
had stabilization in PSA kinetics. 
ATRA has been studied in multiple phase II trials of PCa patients.9, 11 In both studies, the toxicities were 
mild and manageable.  In the study by Trump et al., only one patient required a 50% dose reduction due to 
intolerable headaches occurring on day 1 of therapy. Most patients reported xeroderma, cheilosis, and 
mild conjunctivitis. Two men developed painful balanitis controlled with emollients. Six patients had 
headaches; two had mild and four had moderate headaches. No other important toxicities were noted. In 
the study by Kelly et al., only one patient required dose reduction, which was for fatigue. Xeroderma and 
chelitis were common adverse events, which were successfully treated with skin emollients. Other mild 
toxicities (grade 1 or 2) included elevated transaminases (79%), headaches (57%), constipation (50%), 
and nausea (29%). The most common hematologic toxicities were mild anemia and thrombocytopenia 
except for two patients, who had grade 3 anemia.
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5.2.6
Adverse Events
Incidence rates of adverse events associated with ATRA are provided in the product package insert. The 
most common adverse events are listed below.
Cardiovascular: Peripheral edema (52%), chest discomfort (32%), edema (29%), cardiac arrhythmia 
(23%), flushing (23%), hypotension (14%), hypertension (11%), localized phlebitis (11%)
Central nervous system: Headache (86%), malaise (66%), shivering (63%), pain (37%), dizziness (20%), 
anxiety (17%), paresthesia (17%), depression (14%), insomnia (14%), confusion (11%)
Dermatologic: Xeroderma (≤77%), skin rash (54%), diaphoresis (20%), pruritus (20%), alopecia (14%), 
skin changes (14%)
Endocrine & metabolic: Hypercholesterolemia (≤60%), hypertriglyceridemia (≤60%), weight gain (23%), 
weight loss (17%)
Gastrointestinal: Dry mucous membranes (≤77%), nausea (≤57%), vomiting (≤57%), gastrointestinal 
hemorrhage (34%), abdominal pain (31%), mucositis (26%), diarrhea (23%), anorexia (17%), constipation 
(17%), dyspepsia (14%), abdominal distention (11%)
Hematologic & oncologic: Hemorrhage (60%), leukocytosis (40%), disseminated intravascular coagulation 
(26%)
Hepatic: Increased liver enzymes (50% to 60%)
Infection: Infection (58%)
Neuromuscular & skeletal: Ostealgia (77%), APL differentiation syndrome (≤25%), myalgia (14%)
Ophthalmic: Eye disease (17%), visual disturbance (17%)
Otic: Otalgia (23%; ear fullness)
Renal: Renal insufficiency (11%)
Respiratory: Upper respiratory complaint (63%), dyspnea (60%), respiratory insufficiency (26%), pleural 
effusion (20%), pneumonia (14%), rales (14%), wheezing (expiratory: 14%)
Miscellaneous: Fever (83%)
5.2.7
Drug-drug interactions
ATRA is a major CYPC28 substrate:
CYP2C8 Inducers (Strong): May increase the metabolism of CYP2C8 Substrates (High risk with Inducers). 
Management: Consider an alternative for one of the interacting drugs. Some combinations may be 
specifically contraindicated. Consult appropriate manufacturer labeling. 
CYP2C8 Inhibitors (Moderate): May decrease the metabolism of CYP2C8 Substrates (High risk with 
Inhibitors). 
Management: Monitor during therapy.
CYP2C8 Inhibitors (Strong): May decrease the metabolism of CYP2C8 Substrates (High risk with 
Inhibitors). 
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Management: Consider an alternative for one of the interacting drugs. Some combinations may be 
specifically contraindicated. Consult appropriate manufacturer labeling.
Table 1. Therapeutic agents: Dosing and administration schedule
Agent
Dose
Route
Schedule
Cycle 
Length 
5-AZA
40 mg/m2
SQ
Days 1-5 
x 12 
weeks
ATRA
45 mg/m2 daily 
(divided into 2 
doses)
PO
Days 3-7 
x 12 
weeks
4 weeks 
(28 
days)
Note: Height and weight will be obtained at Day 1 of each treatment cycle to determine treatment dosing. 
The daily dosing for ATRA will be rounded to the nearest 10 mg increment and given in two divided doses 
per day. 
Table 2. Administration schedule per cycle
Cycle
Drug
D1
D2
D3
D4
D5
D6
D7
D8-28
5-AZA
X
X
X
X
X
ATRA
X
X
X
X
X
The treatment schedule maybe adjusted for up to 7 days if any scheduling conflicts related to patient 
availability and/or holidays.
6.
ADVERSE EVENTS
An adverse event or experience is defined as any symptom, sign, illness, or untoward experience 
(including a clinically significant laboratory finding classified as grade ≥ 3 by the National Cancer Institute’s 
Common Terminology Criteria for Adverse Events [CTCAE]14 that develops or worsens during the study, 
whether the event is considered related to study drug, and should be recorded only after the first dose of 
study drug is taken.  Serious adverse events are recorded from the time the informed consent form is 
signed.
6.1
Definitions
Serious adverse event (SAE): any untoward medical occurrence that at any dose: 

Results in death,

Is life threatening, (Note: the term “life-threatening” refers to an event/reaction in which the patient 
was at risk of death at the time of the event/reaction; it does not refer to an event/ reaction which 
hypothetically might have caused death if it were more severe),

Requires inpatient hospitalization or results in prolongation of existing hospitalization,

Results in persistent or significant disability/incapacity,

Is a congenital anomaly/birth defect, or

Is a medically important event or reaction. Medical and scientific judgment should be exercised in 
deciding whether other situations should be considered serious, such as important medical events 
that might not be immediately life-threatening or result in death or hospitalization, but might 
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jeopardize the patient or might require intervention to prevent one of the other outcomes listed in 
the definition above.
Related Adverse Event, i.e. Adverse Drug Reaction (ADR): There is a reasonable possibility per the 
IST/ISS sponsor that the product may have caused the event.
Unexpected Adverse Drug Reaction: An adverse reaction, the nature or severity of which is not consistent 
with the applicable product information (e.g., Investigator's Brochure for an unapproved investigational 
product or package insert/summary of product characteristics for an approved product). An expected ADR 
with a fatal outcome should be considered unexpected unless the local/regional product labeling 
specifically states that the ADR might be associated with a fatal outcome.
6.2
Management of Study Drug Events and Drug Dosing Modifications
Any patient who receives treatment on this protocol will be evaluated for toxicity. Each patient will be 
assessed for the development of toxicity. Toxicity will be assessed per the NCI Common Toxicity Criteria 
for Adverse Events (CTCAE), version 4.03.14 Dose adjustments should be made according to the system 
showing the greatest degree of toxicity. Concurrent participation in another clinical trial or treatment with 
any other anti-cancer therapy is not permitted. The Investigator may prescribe any other concomitant 
medications as deemed necessary.
The proposed dosing of 5-AZA is based on the FDA-approved doses for MDS. 5-azacitidine is generally 
well tolerated, as demonstrated by a phase II trial in chemo-naive castration-resistant metastatic prostate 
cancer.10 The toxicities were generally mild and manageable and only 4 out of 36 patients discontinued 
therapy due to toxicities. ATRA has been studied in multiple phase II trials of PCa patients.9, 11 In both 
studies, the toxicities were mild and manageable. Only one patient required dose reduction in each trial.
Sequential boundaries will be used to monitor the toxicity rate.14 In this trial, we conservatively plan that the 
accrual will be halted if there is sufficient evidence that the toxicity rate exceeds the acceptable rate of 10% 
by more than 15% (unacceptable toxicity rate of 25%). The discrete toxicity boundary values are reported 
in following Table 4. The cumulative number of patients experiencing the toxicities described in Table 6 
and 7 will be compared with Pocock’s stopping boundary values and if greater than the associated 
boundary value, then the trial will stop early. This toxicity monitoring plan controls the overall type I error 
rate of 10%, i.e., probability of stopping trial early under acceptable toxicity rate of 10%, and 80% power, 
i.e., probability of stopping trial early under unacceptable toxicity rate of 25%.
Table 4: Progressive toxicity boundary rules for cumulative number of patients experiencing toxicity.
Looks
5
10
15
20
Boundary
1
2
3
4
If among the 20 patients, 2 or more of the first 5, 3 or more of the first 10, 4 or more of the first 15, or 5 or 
more of the first 20 patients experience recurrent grade 3 or 4 hematological or non-hematological 
toxicities defined in Table 6 and 7 or any toxicity deemed unacceptable by the patient or investigator 
during the 12-week treatment window and up to 4 weeks after treatment window, we will stop the trial 
early.  
The statistical operating characteristics of the above toxicity monitoring rule are as follows:
Table 5: Probability of stopping early based on the true toxicity rate.
Toxicity 
probability
10%
13%
16%
19%
22%
25%
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Early 
stopping 
probability
14%
24%
36%
48%
60%
71%
6.2.1
Hematologic Toxicities
Table 6. Hematologic toxicity dose reductions for 5-AZA
Grade
ANCa
Platelets
Hemoglobin
Action
1
<LLN to 
1,500/L 
<LLN to 
75,000/L 
< LLN to 10 g/dL
None
2
1000-
1499/L
50,000 to 
74,999/L 
8 – 10 g/dL
None 
3
500-
999/L
25,000 to 
49,999/L 
< 8 g/dL
-1st Occurrence: Hold current dose 
until ANC ≥ 1,000/L, platelets ≥ 
50,000/L, and hemoglobin > 8 g/dL. 
Do not replace missed doses. Restart 
next treatment at 30 mg/m2 dose. 
-2nd Occurrence: Discontinue protocol 
therapy.
-If therapy held > 3 weeks: 
Discontinue protocol therapy. 
4
<500/L
<25,000/L
Life-threatening 
consequences; urgent 
intervention indicated
-1st Occurrence: Hold current dose 
until ANC ≥ 1,000/L, platelets ≥ 
50,000/L, and hemoglobin > 8 g/dL.  
Do not replace missed doses. Restart 
next treatment at 30 mg/m2 dose. 
-2nd Occurrence: Discontinue protocol 
therapy.
-If therapy held > 3 weeks: 
Discontinue protocol therapy. 
aNote: G-CSF (Filgrastim) 5 mcg/kg/day may be added for low ANC on day of treatment 
BEFORE a dose reduction is instituted at treating physician’s discretion. This will be continued 
until anticipated nadir has passed and ANC > 1000. Neulasta® is NOT allowed.
6.2.2
Non-hematologic toxicities
5-AZA will be continued at the original dose if the adverse event is grade 0-2. For grade 3-4 toxicities related 
to study drug, 5-AZA will be held until improvement of symptoms to grade 2 or below. After the first episode, 
the dose will be reduced by 25%. After the second episode, the protocol therapy will be discontinued. The 
investigator will determine whether the toxicity is related to study drug. The investigator may prescribe any 
concomitant medications as necessary, except for other anti-cancer therapy, for the management of adverse 
effects. 
ATRA will be continued at the original dose if the adverse event is grade 0-2. For grade 3-4 toxicities related 
to study drug, ATRA will be held until improvement of symptoms to grade 2 or below. After the first episode, 
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the dose will be reduced by 25%. After the second episode, the protocol therapy will be discontinued. The 
investigator will determine whether the toxicity is related to study drug. The investigator may prescribe any 
concomitant medications as necessary, except for other anti-cancer therapy, for the management of adverse 
effects. 
Table 7. Non-hematologic toxicity dose reductions for 5-AZA and ATRA
NCI CTC Grade
5-AZA
ATRA
0-2
No change from original 
starting dose
No change from original 
starting dose
3 -4
Hold until resolved to < 
Grade 2, then reduce to 
30 mg/m2
Hold until 
resolved to < 
Grade 2, then 
reduce to 34 
mg/m2
Second episode of 
grade 3 or 4 toxicity
Remove subject from 
trial
Remove subject from 
trial
The administration of 5-AZA will be reported in a log book by the clinical trial staff. All patients will receive a 
study medication diary to log medication administration and adverse events for both 5-AZA and ATRA (See 
Appendix A).
7.
FOLLOW UP, LABORATORY TESTING, AND DIAGNOSTIC IMAGING
Assessments performed exclusively to determine eligibility for this study will be done only after obtaining 
informed consent. All screening procedures must be performed within 30 days prior to registration unless 
otherwise stated in section 4.2.
Baseline:
Assessments performed for clinical indications (not exclusively to determine study eligibility) may be used 
for baseline values even if the studies were done before informed consent was obtained as outlined in 
section 4.2.
Treatment Phase(s):
Patients will be evaluated on day 1 of each cycle for assessment of adverse events and laboratory values. 
Peripheral blood will also be collected at day 1 of study prior to Leuprolide administration and day 1 of 
each cycle to perform dormancy signature assays. Patients will have an option to undergo BM biopsy to 
obtain aspirate at the end of the study period for dormancy signature assays. A restaging bone scan and 
CT A/P will be performed after every 3 cycles.
Follow Up Period:
Patients will be assessed with physical examination, AE monitoring, and laboratory values. CT 
abdomen/pelvis and bone scan assessments will be performed as deemed appropriate by the clinical 
investigator. Patients will continue with follow up for total of 24 months from day 1 of study, unless there is 
evidence of radiographic progression and/or initiation of other cancer-directed therapies. 
Table 8. Assessment schedule
 Procedures
Treatment Phase(s)
Follow Up
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Physical Examination 
Q4 weeks + 7 days 
Q12 weeks + 7 days 
Vital Signs/Weight 
Q4 weeks + 7 days 
Q12 weeks + 7 days 
CBC with differential 
Q4 weeks + 7 days++ 
Q12 weeks + 7 days 
CMP 
Q4 weeks + 7 days 
Q12 weeks + 7 days 
PSA 
Q4 weeks + 7 days 
Q12 weeks + 7 days 
Testosterone 
Q4 weeks + 7 days 
Q12 weeks + 7 days 
LDH 
Q4 weeks + 7 days 
Q12 weeks + 7 days 
AE Monitoring 
Q4 weeks + 7 days 
Q12 weeks + 7 days 
CT A/P 
Q12 weeks + 7 days 
as per clinical investigator 
Bone Scan 
Q12 weeks + 7 days 
as per clinical investigator 
Blood samples  for dormancy 
biomarkers*^
Q4 weeks + 7 days 
Q12 weeks + 7 days 
++In order to evaluate for cytopenias while on 5-AZA, patients must obtain CBC with differential 
on Day 5 and Day 12 (+/- 3 days) while on treatment arm for all three cycles. Patients have the 
option to obtain the CBC at their nearest local laboratory facility and then send results to clinical 
trial investigators.
* 10 CC of whole blood should be placed in one green top (heparinized) tube following the 
timeline in the protocol. Blood should be collected in addition to bone marrow (optional).  Label 
the tube with the patient’s ID#, the date of collection, diagnosis, relapse (Y/N), and the source of 
the material (i.e. blood). The tubes should be kept on ice and handed over immediately to 
Aguirre-Ghiso Lab for processing. 
^ Optional - 10 CC of bone marrow aspirate will be collected by Bobby Liaw/co-investigators at 
the end of 24-month study period. It should be placed in one green top (heparinized) tube.  Same 
procedures as above. 
7.1
Duration of Therapy
Treatment may continue for the 3-cycle treatment phase or until:

Inter-current illness that prevents further administration of treatment

Unacceptable adverse event(s) of the study regimen despite appropriate dose 
reduction and best supportive management

Patient decides to withdraw from the study

Noncompliance with study regimen or follow up appointments

General or specific changes in the patient’s condition render the patient 
unacceptable for further treatment in the judgment of the investigator.
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The principal investigator will be notified and a case report form will be completed including the reason for 
study removal and the date the patient was removed. The patient should be followed up per protocol.
8.
STATISTICAL CONSIDERATIONS
8.1
Study Design/Study Endpoints
The primary endpoints will be the disease progression-free rate at the end of the treatment phase 1 and 
safety of the combination of 5-AZA and ATRA. Disease progression is defined by composite of PSA or 
radiographic progression relative to period baseline (start of treatment phase 1), whichever occurs first. 
PSA levels will be measured on day 1 of each cycle. For assessment of PSA progression, see section 
3.2.1 above. CT A/P and bone scan will be performed every 3 cycles. For assessment of radiographic 
progression, see section 3.2.2 above. Safety will be assessed by the recording of adverse events, 
monitoring of vital signs and physical examinations, safety laboratory evaluations, and 12-lead ECG. 
Adverse events will be graded per the Common Terminology Criteria for Adverse Events (CTCAE) version 
4.0.14 Therapy will be continued at the original dose if the adverse event is grade 0-2. For grade 3-4 
toxicities, therapy will be held until improvement of symptoms to grade 2 or below. The dose will then be 
reduced by 25% of original for each episode. A maximum of 2 dosage reductions will be allowed. The third 
occurrence will result in removal of the patient from the study.
The secondary endpoints will be time to tumor progression from start of treatment phase 1 and the 
measurement of dormancy biomarker levels in relation to disease states: tumor progression and disease 
progression. To determine the relation between dormancy biomarkers and disease states, every 3 cycles, 
we will measure the dormancy biomarkers, TGF2, BMP7, BMP4, GAS6, retinoic acid, NR2F1, in BM 
aspirates and peripheral blood. 
8.2
Sample Size and Accrual
The primary endpoint is disease progression free rate at the end of treatment phase 1. Disease 
progression is defined as either PSA progression or radiographic progression relative to baseline. The 
baseline value is defined as the PSA level at the beginning of treatment phase 1 (Day 29 of study). The 
previous Phase II studies on 5-AZA and ATRA individually demonstrated their efficacy in delaying PSA rise 
among prostate cancer patients. Preclinical in-vivo study demonstrated that in adjuvant setting following 
primary tumor surgery, animals treated with the drug combination for 4 weeks have 73% reduction on the 
median cell count of disseminated tumor cells in lung comparing to control animals (Personal 
communications between Dr. Aguirre-Ghiso and Dr. Jia). 
Thus, we assume the difference in progression free rates at end of the treatment periods between drug 
combination and no-therapy ranges between 30% to 60% and listed various scenarios of progression-free 
rates in Table 1. We use McNemar test statistic to calculate the range of sample size required to 
differentiate the treatment from no-therapy under each scenario.15, 18 The maximum sample size is 
required when the PSA rise is independent in the two treatment periods for the same patient. The 
minimum sample size is required when the PSA progression status in the two treatment periods is highly 
concordant.  With the planned 20 patients in total, we would have >80% power to detect the difference of 
PSA progression rate of 20% vs 60%, with two-sided type I error of 10%. All calculation was done using 
PASS software.
Table 1: Sample size using McNemer test statistic, with 80% power and 10% two-sided type I error 
rate.
Sample size
P1=0.6
P2=0.7
P2=0.8
P1=0.2
14-19
8-10
10-13
P1=0.25
16-26
12-16
9-11
P1=0.3
19-35
14-20
10-13
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With accrual rate of 2 patients per month, we expect to finish enrollment in 10 months. Patients who did 
not finish 8 weeks of drug combination treatment will be replaced but will included for toxicity stopping rule. 
8.3
Randomization Schema
Since prior PSA doubling time is a known prognostic factor to disease progression, the randomization will 
be stratified on PSA doubling time: PSADT<=3 months vs PSADT>3 months. A stratified block 
randomization of block size 1 will be used19, 20 and separate randomization lists are generated in different 
strata (See appendix). The enrollment will continue until total number of 20 evaluable patients is reached. 
The target accrual number is set at 30 to account for screen failures.The maximum imbalance in each 
stratum is 1 and maximum total imbalance across strata is 2.
8.4
Data analyses
Analysis on efficacy and safety:
Patients will receive at least 8 weeks of treatment in order to be eligible for efficacy analysis. Patients who 
discontinued due to toxicity after 8 weeks of treatment will be included in efficacy analysis. Descriptive 
statistics such as mean/median/range and frequency will be provided for continuous and categorical 
variables separately, and the distributions of period baseline variables are contrasted between 5-AZA+ATRA 
and placebo using Wilcoxon statistics and Fisher’s exact test statistics for continuous and categorical 
variables separately. Waterfall plot will be provided for PSA change relative to period baseline for all patients 
for each treatment arm and period. Analysis will follow the two-step procedure prescribed in17 for randomized 
crossover study16 for binary endpoint. To test for carryover effect, Wilcoxon rank-sum test stratified by prior 
PSADT will be performed to assess whether the sum of outcome variables from both periods is different 
between the randomization arms. If the test yields insignificant results, a similar test will be performed to 
compare the within-patient outcome difference between period 1 and 2 between the randomization arms to 
assess the treatment effect. If the carryover effect is significant, the primary analysis will be based on the 
progression free rates at the end of first treatment period using Chi-square test.  In addition, general linear 
mixed model will be applied to assess the treatment effect, carryover effect, and period effect, adjusting for 
the stratification factor prior PSADT. The time to disease progression is defined as time to either PSA 
progression or radiographic progression whichever occurs first since discontinuation of ADT. Time to event 
endpoints will be summarized using Kaplan-Meier method. The time to event endpoints will be estimated 
using rank preserving failure time model (RPFTM) to obtain the counter-factual event times assuming cross-
over did not occur (R package “rpfstm”).21 20 
Descriptive summary will be provided for the safety endpoints. All patients who received at least one dose 
of study drug (treated set) will be included in the safety analysis.  
Correlative analysis and biomarkers:
The micro-environmental biomarkers and dormancy biomarkers collected in this study will be described with 
summary statistics (mean, median, standard deviation, range) at each time point, and compared between 
the 5-AZA+ATRA and placebo using the Wilcoxon rank sum test statistics. Distribution of biomarkers will be 
examined and linear mixed model with random effects will be applied to analyze the biomarker trajectories 
over time. The biomarkers will be correlated with disease progression outcomes. Biomarker analysis will be 
performed on all patients with at least one biomarker measure at one time point (biomarker set).
9.
DATA HANDING AND RECORD KEEPING
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9.1
Electronic Data Capture (EDC) System
All eCRFs will be entered directly into a web-based electronic research application portal known as eRAP. 
eRAP’s role-based access control security and audit capability ensures that the research data is protected 
from unauthorized access, modification, and exposure. Key features of this web-based database system 
include allowing access and data entry from multiple sites, with each site having a separate pool of data as 
necessary. All data stored in the eRAP system is backed up daily. Detailed audit services include any data 
field level changes, who made the changes, when the changes were made, the old value and the new 
value. Data from eRAP can easily be extracted to excel or flat text file formats for easy import into SAS or 
other statistical software packages.
9.2
Verification of EDC System
Once the database has been set up, the PI will test the system. The testing will include confirmation of the 
proper functioning of valid value checks, subject ID generation, derived variable computations, data 
extracts, and system reports. 
9.3
Entering data
Since electronic CRFs will be used, data entry will be conducted onsite by clinicians who have been 
granted appropriate access to do so.
9.4
Data Validation Process
As part of the data validation process, edit check programs will be embedded in the database thru valid-
value, valid-range and missing value alerts specified for each field as necessary. Valid-value edit checks 
include:
o
Requiring the user to enter a coded value taken from a list that is presented on the CRF. If a value not 
on the list is entered the user will be prompted to check the value and enter a value on the list.
o
Requiring the user to enter a value within a particular range of possible values for a continuous 
measure. If a value outside the range is entered the user will be prompted to check the value and re-
enter.

In addition to the univariate field alerts specified above, there will be multivariate 
alerts built into the database design:
o
Confirming that only valid options are selected in a ‘choose all that apply’ multiple choice field, where 
the range of options deemed valid depends on some other parameter.
o
Confirming that diastolic blood pressure reading is less than the associated systolic blood pressure 
reading.
o
Confirming that “other, specify” is completed when “other” is selected.
o
Confirming that eligibility criteria are met.
o
As well as cross-module alerts such as:

Comparing the dates and times of all assessment time points to confirm that they 
occur in an appropriate sequence.
 
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9.5
Data Cleaning and Discrepancy Management
Any discrepancies identified through this process will be highlighted and corrected in the system.   
9.6
Data Security
The EDC database is approved by Mount Sinai IT Security and HIPAA and operates using high-end 
servers located in the Secured Mount Sinai Data Center. All data stored in the eRAP system is backed up 
daily. Access to the database is controlled by policies requiring the PI to authorize user and user roles. 
Quick view audit service allows authorized user to see who accessed records for view or edit and detailed 
audit services include seeing any data field level changes, who made the changes, when the changes 
were made, as well as the value before and after the change.
9.7
Quality Control Procedures
Comprehensive edit checks will be used to clean data. Patient data will be entered continuously. All 
changes to the data and the database structure will be recorded in an automatic audit trail. Random 
checks will be done by the PI to ensure data accuracy and completeness. A final database will be declared 
when all data has been entered, the data entry verified, the data validated and the database defined as 
clean. After declaration of a final database the data will be exported from eRAP to excel for import to SAS 
and both the database and the SAS datasets will be locked and protected from changes. All statistical 
analyses for the final analysis will be performed on the locked SAS datasets. 
9.8
Medical and Adverse Event Coding
Coding of Adverse Events occurring during the study will be performed by the Clinical Research Manager 
according to the revised CTCAE v4.0 while coding of concomitant medications, prior anti-cancer therapy 
and further therapy will be performed according to the World Health Organization-Drug Dictionary 
Enhanced (WHO-DDE). These dictionaries contain the respective classifications of adverse events and 
drugs in proper classes. When dictionary entries and verbatim terms (either for adverse events for 
medications) do not directly match, the PI will review the entry as well as other related information such as 
comment fields to identify the most appropriate match. In the event there are compound verbatim events 
(either AEs or medications) listed in the CRF, such events must be split into more than one record for 
purposes of medical coding. The PI will be responsible for splitting the compound events into separate 
entries.
9.9
Confidentiality
Information about study subjects will be kept confidential and managed according to the requirements of 
the Health Insurance Portability and Accountability Act of 1996 (HIPAA).  Those regulations require a 
signed subject authorization informing the subject of the following: 
•
What protected health information (PHI) will be collected from subjects in this study
•
Who will have access to that information and why
•
Who will use or disclose that information
•
The rights of a research subject to revoke their authorization for use of their PHI. 
In the event that a subject revokes authorization to collect or use PHI, the investigator, by regulation, 
retains the ability to use all information collected prior to the revocation of subject authorization.  For 
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subjects that have revoked authorization to collect or use PHI, attempts should be made to obtain 
permission to collect at least vital status (i.e. that the subject is alive) at the end of their scheduled study 
period.
9.10
Records Retention
The Investigator must retain drug disposition records (if applicable), source documents, and case histories 
designed to record all observations and other data pertinent to the investigation (e.g. case report form) for 
the maximum period required by applicable regulations and guidelines, or Institution procedures.
If a change in the PI occurs, the records shall be transferred to a mutually agreed upon designee (e.g., 
another Investigator, IRB).
9.11
Subject Privacy
Patients will be informed of the extent to which their confidential health information generated from this 
study may be used for research purposes. Following this discussion, they will be asked to sign the HIPAA 
form and informed consent documents. The original signed document will become part of the patient’s 
medical records, and each patient will receive a copy of the signed document. The use and disclosure of 
protected health information will be limited to the individuals described in the informed consent document.
It is the responsibility of the research staff to ensure that protocol subjects received, understands, and 
signs the informed consent document before enrolling the patient onto this trial. Personnel must provide a 
HIPAA form and obtain acknowledgment before the subject participates in this study.
All subject data will be identified by a subject identification number and subject initials only, to protect the 
subject’s privacy. The data will be blinded accordingly in all data analysis. However, in compliance with 
federal guidelines, the investigator will permit a representative from Mount Sinai Health System audit 
committee to review that portion of the subject’s medical record that is directly related to the study. This will 
include all relevant study documentation including medical histories to verify eligibility, laboratory test 
results to verify transcription accuracy, X-ray reports, admission/discharge summaries for 
hospital/outpatient admissions while the subject is on-study and autopsy reports for deaths occurring 
during the study. As part of the required content of informed consent, the subject will be informed that his 
medical record may be reviewed. Should access to the medical record require a separate waiver or 
authorization, it is the PI’s responsibility to obtain such permission from the patient in writing before the 
subject is entered into the study.
10.
ETHICAL CONSIDERATIONS
This study is to be conducted according to US and international standards of Good Clinical Practice (FDA 
Title 21 part 312 and International Conference on Harmonization guidelines), applicable government 
regulations and Institutional research policies and procedures.
This protocol and any amendments will be submitted to a properly constituted independent Ethics 
Committee (EC) or Institutional Review Board (IRB), in agreement with local legal prescriptions, for formal 
approval of the study conduct.  The decision of the EC/IRB concerning the conduct of the study will be 
made in writing to the investigator and a copy of this decision will be provided to the sponsor before 
commencement of this study.  The investigator should provide a list of EC/IRB members and their affiliate 
to the sponsor.
All subjects for this study will be provided a consent form describing this study and providing sufficient 
information for subjects to make an informed decision about their participation in this study.  See 
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Attachment D for a copy of the Subject Informed Consent Form.  This consent form will be submitted with 
the protocol for review and approval by the EC/IRB for the study.  The formal consent of a subject, using 
the EC/IRB-approved consent form, must be obtained before that subject undergoes any study procedure.  
The consent form must be signed by the subject or legally acceptable surrogate, and the investigator-
designated research professional obtaining the consent. 
11.
FINANCIAL CONSIDERATIONS
See attached clinical trial and correlative study budget spreadsheets (appendix B). Funding was obtained 
from the Jimmy V Foundation – Translational Research Award Grant. Drug funding for 5-AZA and ATRA 
will be obtained from the Tisch Cancer Institute Grant.
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End Date:1/16/2024