Clinical trial NCT03765229
Conditions: Melanoma
Keywords: PD-1
Sponsor: UNC Lineberger Comprehensive Cancer Center
Start date: 2019-03-22; Completion date: 2023-07-31
Enrollment: 11 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
Cancers develop in two different ways. First, cancer cells can become invisible to the immune system by stop having proteins on their surface that are required for the immune system to recognize them. In this scenario, tumors do not attract any immune cells (e.g. white blood cells) whatsoever or they do not attract specialized white blood cells against cancer cells, called lymphocytes. White blood cells are the type of immune cells that attack foreign cells, such as cancer cells or normal cells infected with viruses or bacteria. Second, cancer cells can still grow side-to-side with white blood cells but are able to hide from them. As a result, the white blood cells cannot find and attack the cancer cells. Different types of cancers have different chance of having immune cells in the tumor. For example, the possibility that immune cells are within skin melanomas is almost 50% whereas the possibility in melanoma of the eye is only 10%.

As a result, the first goal of this study is to understand whether entinostat can make a melanoma tumor more visible to the immune system. To see whether entinostat makes tumor more visible to the immune system, participants will have a mandatory tumor biopsy 3 weeks after starting entinostat therapy. Tumor tissue collected before and after participating in this study will be compared to see if there are more immune cells in the tumor after receive entinostat. The second goal of the study is to see if giving a combination of entinostat and pembrolizumab can shrink melanoma tumors of patients who did not have immune cells in tumors prior to treatment. The study will determine how many subjects cancer has become better or not changed 6 months after subjects have started treatment on the study. We will also determine what type of side effects occur in subjects receiving entinostat and pembrolizumab to look at the safety of this combination.

The investigators will also look at any changes in the DNA of melanoma before the study begins. As a result of these changes in DNA, there are often see differences in the proteins that work to create other proteins. In addition, the study will look into how entinostat may make melanoma cells more visible to the immune system by comparing proteins in tumors before and after treatment. Finally, the study will see if this treatment changes the numbers and types of immune cells that are found in the blood by comparing blood at different time points while patients are on the study.

Detailed description:
This is an exploratory (n=10-12 evaluable patients), open-label, single-arm, phase II study in patients with 'non-inflamed' unresectable regional or distant metastatic melanomas irrespective of prior treatment with PD-1/PD-L1 pathway inhibitors. The primary endpoint is to assess the incidence of histopathologic conversion of non-inflamed melanomas from patients with metastatic melanoma to 'inflamed' melanoma following single-agent entinostat "priming" (entinostat monotherapy). More specifically, patients with metastatic melanomas that have no evidence of tumor-infiltrating or tumor-associated lymphocytes (TIL/TAL), based on standard hematoxylin and eosin (H\&E) staining of representative tumor sections (non-inflamed melanomas) will receive weekly entinostat for 3 weeks in cycle 1 (entinostat monotherapy; 5mg PO, qwk on D1, D8, D15 of cycle 1; cycle length = 21 days). Mandatory tumor tissue biopsies will be performed in the end of cycle 1, beginning of cycle 2 (day 21±2 days), immediately before treatment with concurrent entinostat (once weekly × 3) and pembrolizumab (200 mg q3wks) in cycles 2-9 (see section 5.2.1 drug dosing schema). Correlative studies will be performed at baseline and in the end of cycle 1, beginning of cycle 2 (day 21±2 days) to assess whether: (a) 3 weeks of entinostat monotherapy converts TIL/TAL-absent melanomas to TIL/TAL-present by histopathologic (H\&E stain) analysis, (b) 3 weeks of single-agent entinostat induces changes in gene expression profiling by assessing distinct signatures as defined by RNA-sequencing signatures (RNA-seq; 'immune-high', innate anti-PD-1 resistance, and epigenetic 1-3), (c) 3 weeks of single-agent entinostat induces changes in histone-accessible DNA by performing formaldehyde-assisted isolation of regulatory elements (FAIRE)4, (d) conversion of non-inflamed to inflamed melanomas by single-agent entinostat and/or antitumor response to the entinostat-pembrolizumab combination is associated with a baseline signature that consists of distinct somatic mutation gene profile, global histone modification profile in melanoma tissue or peripheral blood mononuclear cells (PBMC), and/or abundance of entinostat targets in melanoma cells (HDAC 1, 2, 3, and 11). Up to 12 evaluable patients will be treated with entinostat plus pembrolizumab for up to 27 weeks (approximately 6 months) based on clinical benefit. Patients who continue to have clinical benefit at 27 weeks may continue therapy with pembrolizumab or any other PD-1/PD-L1 inhibitor, as per standard of care.Secondary exploratory endpoints involve: (a) assessment of antitumor response of entinostat administered concurrently with pembrolizumab at week 10 (or earlier if patient progresses), based on response rate per Response Evaluation Criteria in Solid Tumors version 1.1 (RECIST v1.1). (b) assessment of the progression-free survival (PFS) rate at 27 weeks (approximately 6 months) of the entinostat-pembrolizumab combination. (c) determine the safety of the entinostat-pembrolizumab combination in patients with metastatic melanoma per NCI Common Terminology Criteria for Adverse Events (NCI-CTCAE). (d) assessment of other exploratory biomarkers in tumor tissue and peripheral blood.

Arms: arm 1: Subjects in this trial will be administered entinostat, 5mg PO, once weekly (D1, D8, D15 of a 21-day cycle) starting on day 1 of study treatment. Pembrolizumab, 200 mg will be administered intravenously (IV) every 3 weeks and initiated at cycle 2 after the mandatory research tumor biopsy in the end of cycle 1, beginning of cycle 2 (day 21±2 days),. Combination therapy with both agents will continue if subject is receiving clinical benefit from therapy for up to 27 weeks (8 cycles of combination therapy or approximately 6 months). Study therapy will be discontinued for intolerable toxicity, disease progression or for other reasons at the discretion of the investigator.
Interventions: intervention 1: Entinostat intervention 2: Pembrolizumab
Intervention descriptions: intervention 1: Oral drug 5 mg taken once weekly for up to nine 3-week (21 day) cycles. Take on an empty stomach i.e., at least 2 hours after a meal and at least 1 hour before the next meal. On days when entinostat is administered on the same day as pembrolizumab, entinostat should be taken before the pembrolizumab infusion. intervention 2: 200 mg IV starting on Day 22 (cycle 2, Day 1) given every 3 weeks for up to 8 cycles.

Protocol documents:
Prot_SAP_000.pdf: 
LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
PROTOCOL AMENDMENT 2 
LCCC 1729: Breaking Innate PD-1 Inhibitor (PD1i) Resistance Using 
Epigenetic Modifiers; Antitumor Efficacy and Correlative Analyses of 
Entinostat plus Pembrolizumab in Non-Inflamed Metastatic Melanoma (MM) 
AMENDMENT INCORPORATES: 
X     Editorial, administrative changes (IRB approval) 
X     Scientific changes (IRB approval)         
X       Therapy changes (IRB approval) 
X     Eligibility Changes (IRB approval) 
Rationale for amendment: 
The primary purpose of this amendment is to update the dose modifications for 
pembrolizumab. The definition of a serious adverse event  (SAE) was updated. The 
schedules for PBMC sample collection and scans were updated. Inclusion Criteria 
were updated to provide guidance on when adequate recovery is achieved after a 
pre-study treatment of radiotherapy or surgery. Prohibited medications sections 
were updated to clarify the allowance of fentanyl for the study biopsy procedure. 
Dose 
modification 
and 
toxicity 
management 
for 
pembrolizumab 
and 
pembrolizumab combinations has been updated. Clarified discrepancies in timing 
for peripheral blood draws throughout and pre-treatment imaging. Disease 
assessment at cycle was added. 
Summary of Changes 
Editorial Changes 
1. Sections 1.7.2.4, 3.3.2, 5.1, and 6.5.1: made edits to clarify the timing of
peripheral blood draws.
2. Section 7.1:
a.
Removed PBMC collection at cycles 7, 8, 9 to remain consistent
with the rest of the protocol.
b. Removed footnote #6 from the top row of column D1, Cycles 7, 8,
9 as it was accidentally not removed previously.
3. Section 7.5.1: Added PBMC collection to remain consistent with the
protocol
4. Section 7.8.1: Section updated to correct discrepancy in pre-tretment
imaging from 4 weeks to 21 days.


LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
 
5. Section 9.1.1: The definition of an SAE as previously described was 
updated to align with the FDA’s definition. 
6. Updated the week 9 designation to week 10 throughout the protocol. 
7. Appendix A and Appendix B: To indicate that the prohibition of fentanyl 
does not apply to the study biopsy procedure. 
 
Scientific Changes 
1. Sections 6.3.6 and 7.4.7: Updated to add D1 of cycle 8.  
2. Section 7.1: Time and Events Table: Added whole body CT assessment at 
week 8 in the table. Footnote#8 updated with this information. 
3. Section 7.4.7: Added in disease assessment at D1 of cycle 8. 
 
Therapy Related-Changes 
1. Section 5.3.2: Updated dose modifications for pembrolizumab 
a. Dose modification and toxicity management table has been replaced 
with an updated table. 
 
Eligibility Changes 
1. Criterion 4.1.2: Updated criterion to clarify that subjects must recover to 
Grade ≤ 1. 
 
The attached version dated December 15, 2021, incorporates the above revisions. 
 
 


LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
 
 
 
 
PROTOCOL AMENDMENT 1 
 
 
LCCC 1729: Breaking Innate PD-1 Inhibitor (PD1i) Resistance Using 
Epigenetic Modifiers; Antitumor Efficacy and Correlative Analyses of 
Entinostat plus Pembrolizumab in Non-Inflamed Metastatic Melanoma (MM) 
 
 
AMENDMENT INCORPORATES: 
X     Editorial, administrative changes (IRB approval) 
X     Scientific changes (IRB approval)         
        Therapy changes (IRB approval) 
X     Eligibility Changes (IRB approval)  
 
Rationale for amendment:  
This amendment clarifies the process for analyzing biopsy samples, as well as the 
timing of certain laboratory tests.  The amendment also removes unnecessary 
enrollment restrictions regarding subjects with hypertension, diabetes mellitus, 
and lung metastases. 
 
 
Summary of Changes 
 
Editorial/Administrative changes 
• Biopsy timing had been stated inconsistently in the protocol (Day 21 vs. 22, 
with a window of +/- 2 days).  The timing has been made consistent 
throughout the protocol as Day 22 +/- 2 days. 
• Coagulation studies described in section 4.1.9 were not captured in the Time 
and Events Table in section 7.1, and have now been added to the table. 
• The contact information for Syndax SAE reporting has been updated in 
Section 8.4.2, in accordance with a prior administrative letter. 
• Minor edits and clarifications have been made. 
 
Scientific Changes 
1. The process of analyzing biopsy tissue has been updated in sections 1.7.2, 
4.1.7, 6.3.7 and Appendix C. 
 
Therapy Changes 
2. Urinalysis will only be required at screening and at Cycle 1, and at other 
times as clinically indicated. 
 
Eligibility Changes 


LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
 
• Patients whose only metastasis is in the lung were previously excluded.  
This exclusion has been removed from section 4.1.6. 
• Patients with uncontrolled hypertension were previously excluded per 
section 4.2.6.  This exclusion has been removed. 
• Patients with diabetes mellitus (HbA1c >9.0 within 15 days from first dose 
of entinostat) were previously excluded per section 4.2.6.  This exclusion 
has been removed. 
 
The attached version dated July 29, 2019, incorporates the above revisions 
 
 




LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
 
 
LCCC1729: Breaking Innate PD-1 Inhibitor (PD1i) Resistance Using 
Epigenetic Modifiers; Antitumor Efficacy and Correlative Analyses of 
Entinostat plus Pembrolizumab in Non-Inflamed Metastatic Melanoma (MM) 
 
 
 
Principal Investigator 
Stergios Moschos, MD 
Department of Medicine, Division of Hematology/Oncology 
University of North Carolina at Chapel Hill 
3116 Physicians Office Building 
170 Manning Drive, CB# 7305 
Chapel Hill, NC 27599-7305 
PHONE: (919) 843-7713 
FAX: (919) 966-6735 
Email: moschos@med.unc.edu 
 
 
 
 
 
Signature Page 
 
The signature below constitutes the approval of this protocol and the attachments, and 
provides the necessary assurances that this trial will be conducted according to all 
stipulations of the protocol, including all statements regarding confidentiality, and 
according to local legal and regulatory requirements and applicable U.S. federal regulations 
and ICH guidelines. 
 
Principal Investigator (PI) Name: _______________________ 
 
PI Signature: ___________________ 
 
Date: ____________________ 
 
Version date:  December 15, 2021 
Amendment Number 2 / Version #: 1  
 
 


LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
 
LIST OF ABBREVIATIONS  
 
ADME 
Absorption, distribution, metabolism, and excretion 
ADP 
Adenosine diphosphate 
AE 
Adverse event 
AJCC 
American Joint Committee on Cancer 
ALC 
Absolute lymphocyte count 
ALP 
Alkaline phosphatase 
ALT 
Alanine aminotransferase 
ANC 
Absolute neutrophil count 
aPTT 
Activated partial thromboplastin time 
AST 
Aspartate aminotransferase 
ATP 
Adenosine triphosphate 
AUC 
Area under the curve 
β-HCG 
Beta –human chorionic gonadotropin 
BRAF 
Murine sarcoma viral oncogene homolog B 
Cmax 
Maximum concentration 
CBC 
Complete blood count 
CDKN2A 
Cyclin dependent kinase inhibitor 2A 
CL 
Chloride 
CLIA 
Clinical Laboratory Improvement Amendments 
CO2 
Bicarbonate 
COPD 
Chronic obstructive pulmonary disease 
CpG 
Cytosine phosphate-diester Guanine 
CPO 
Clinical Protocol Office 
CR 
Complete response 
CrCl 
Creatinine clearance 
CRF 
Case report form 
CT 
Computer tomography 
CTLA 
Cytotoxic T Lymphocyte-Associated 
CYP 
Cytochrome P450 
D1 
Day 1 
dL 
Deciliter 
DSMC 
Data Safety Monitoring Committee 
EC50 
Effective concentration 50% (concentration that gives half-maximum effect) 
ECOG 
Eastern Cooperative Oncology Group 
eCRF 
Electronic case report form 
EPO 
Erythropoietin 
FACS 
Fluorescence-activated cell sorting 
FAIRE 
Formaldehyde-assisted isolation of regulatory elements 
FDA 
Food and Drug Administration 
FFPE 
Formalin-fixed paraffin-embedded 
5-FU 
5-fluorouracil 
GCP 
Good clinical practice 
GFR 
Glomerular filtration rate 
GNAQ/GNAS 
Guanine nucleotide binding protein alpha G(q) / alpha subunit  
Gy 
Gray 
H3/4 
Histone 3 or Histone 4 
HAT 
Histone acetyl transferases 
HDAC /HDACi 
Histone deacetylase / Histone deacetylase inhibitor 
H&E 
Hematoxylin & Eosin 
Hgb 
Hemoglobin 


LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
 
HIPAA 
Health Insurance Portability and Accountability Act 
HIV 
Human immunodeficiency virus 
H3K 
Histone 3 kinase 
H3K18Ac 
Acetylated histone 3 lysine 18 
H4K12Ac 
Acetylated histone 4 lysine 12 
H4R3diMe 
Dimethylated histone 3 lysine 4 
H3K27triMe 
Trimethylated histone 3 lysine 27 
Hr 
Hour 
IB 
Investigator’s Brochure  
IC50 
Inhibitory concentration 50% (inhibits an effect by 50%) 
ICP 
Immune checkpoint proteins 
IDO 
Indoleamine 2,3, dioxygenase 
IDS 
Investigational Drug Service  
IF 
Immunofluorescence 
IFN / IFNγ 
Interferon / Interferon gamma 
Ig 
Immunoglobulin 
IHC 
Immunohistochemistry 
IL-2 
Interleukin-2 
IMGF 
Immune Genomics Facility 
INR 
International Normalized Ratio 
IPI-N 
Ipilumumab-naive 
IPI-T 
Ipilumumab-treated 
IPRES 
Innate PD-1 resistance 
IRB 
Institutional Review Board 
ITIM  
Immunoreceptor tyrosine-based inhibition motif   
ITISM 
Immunoreceptor tyrosine-based switch motif 
IU 
International unit 
IV 
Intravenous 
K 
Potassium 
Kg 
Kilogram 
LDH 
Lactate dehydrogenase 
(m) 
Mucosal 
(m/a) 
Mucosal/acral 
MAPK 
Mitogen-activated protein kinase 
MB 
Methyl binding 
MDSC 
Myeloid-derived suppressor cell 
Me1, Me2, or Me3 
Mono, Di- or tri-methylation 
MEK 
Mitogen-activated protein kinase  
Mg 
Milligram 
MHC 
Major histocompatibility class 
Min 
Minute 
miRNA 
Micro-Ribonucleic acid 
MITF 
Microphthalmia-associated transcription factor 
mL 
Milliliter 
M 
Metastatic melanoma 
MRI 
Magnetic Resonance Imaging 
M-W-F /T-Th 
Monday, Wednesday, Friday / Tuesday, Thursday 
Na 
Sodium 
NCI-CTCAE 
National Cancer Institute–Common Terminology Criteria for Adverse Events 
NK 
Natural killer 
NFκB 
Nuclear factor kappa Beta 
NOAEL 
No observed adverse event level 
NRAS 
Neuroblastoma sarcoma viral oncogene homolog 


LINEBERGER COMPREHENSIVE CANCER CENTER 
LCCC1729 
CLINICAL ONCOLOGY RESEARCH PROGRAM 
 
December 15, 2021 
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL 
Amendment 2 
 
NSCLC 
Non-small cell lung cancer 
NYHA 
New York Heart Association 
(o) 
ocular 
ORR 
Overall response rate 
OS 
Overall survival 
PBMC 
Peripheral blood mononuclear cells 
PD 
Progressive disease 
PD-1 or PD-L1 
Programmed death-1 or Programmed death ligand 1 
PD1i 
Programmed death-1 inhibitor 
PE 
Physical Exam 
PFS 
Progression-free survival 
Pgp 
P-glycoprotein 
PKC 
Protein kinase C 
PO 
Per os (by mouth or orally) 
PR 
Partial response 
PT 
Prothrombin time 
QW 
Quaque  (once a week or once weekly) 
Q3 weeks 
Every 3 weeks 
RECIST 
Response evaluation criteria in solid tumors 
RNA 
Ribonucleic acid 
RNA-Seq 
Ribonucleic acid sequencing 
RR 
Response rate 
SAE 
Serious adverse event 
SD 
Stable disease 
SmPc 
Summary of product characteristics 
SUSAR 
Serious unexpected adverse reaction 
T½ 
Half-life 
T4 
Thyroxine 4 
TCR 
T-cell receptor 
TCGA 
Cancer Genome Atlas Project 
T1DM 
Type 1 diabetes mellitus 
Th1 / Th2 
T-helper 1 / T-helper 2 
TIL/TAL 
Tumor-infiltrating lymphocyte / Tumor-associated lymphocyte 
Tmax 
Time to maximum concentration 
TNFα 
Tumor necrosis factor alpha 
TPL 
Translational Pathology Laboratory 
Treg 
T-regulatory  
TSH 
Thyroid stimulating hormone 
UGT 
Uridine diphosphate glucuronosyltransferase 
ULN 
Upper limit of normal 
µM 
Micromolar 
UNC-CH 
University of North Carolina at Chapel Hill 
UVR 
Ultraviolet radiation 
V-type 
Variable type 
Wk 
Week 
WOCBP 
Woman of childbearing potential 
 


LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
i 
TABLE OF CONTENTS 
LIST OF ABBREVIATIONS ............................................................................ vii 
1.0 
BACKGROUND AND RATIONALE .......................................................4 
1.1 
Study Synopsis ......................................................................................................4 
1.2 
Disease Background ..............................................................................................5 
1.3 
Current Standard of Care .......................................................................................6 
1.4 
Investigational Treatments ....................................................................................8 
1.5 
Pembrolizumab (MK-3475) ................................................................................15 
1.6 
Rationale for Clinical Study ................................................................................20 
1.7 
Correlative Studies ..............................................................................................21 
2.0 
STUDY OBJECTIVES ..............................................................................25 
2.1 
Primary Objective ...............................................................................................25 
2.2 
Secondary Objectives ..........................................................................................25 
2.3 
Exploratory Objectives ........................................................................................25 
3.0 
Criteria for Evaluation / Study Endpoints ..............................................26 
3.1 
Primary Endpoint ................................................................................................26 
3.2 
Secondary Endpoints ...........................................................................................26 
3.3 
Exploratory Endpoints .........................................................................................26 
4.0 
PATIENT ELIGIBILITY .........................................................................27 
4.1 
Inclusion Criteria .................................................................................................27 
4.2 
Exclusion Criteria ................................................................................................30 
5.0 
TREATMENT PLAN ................................................................................32 
5.1 
Schema ................................................................................................................32 


LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
ii 
5.2 
Treatment Dosage and Administration ...............................................................33 
5.3 
Toxicities and Dosing Delays/Dose Modifications .............................................35 
5.4 
Concomitant Medications/Treatments/Supportive Care Allowed ......................45 
5.5 
Prohibited Medications/Treatments ....................................................................45 
5.6 
Rescue Medications and Supportive Care ...........................................................46 
5.7 
Other Modalities or Procedures ...........................................................................53 
5.8 
Duration of Therapy ............................................................................................53 
5.9 
Duration of Follow Up ........................................................................................53 
5.10 
Study Withdrawal ................................................................................................54 
6.0 
DRUG INFORMATION ...........................................................................55 
6.1 
Entinostat - Investigational Agent .......................................................................55 
6.2 
Pembrolizumab (Keytruda™) - Commercial Drug .............................................57 
6.3 
Clinical Assessments ...........................................................................................60 
6.4 
Clinical Laboratory Assessments ........................................................................62 
6.5 
Correlative Studies ..............................................................................................64 
7.0 
EVALUATIONS AND ASSESSMENTS .................................................65 
7.1 
Time and Events Table ........................................................................................65 
7.2 
Pre-Screening Assessments .................................................................................67 
7.3 
Screening Assessments .......................................................................................67 
7.4 
Study Treatment Assessments .............................................................................67 
7.5 
Post Treatment Follow-Up Assessments .............................................................70 
7.6 
Handling of Bio specimens Collected for Correlative Research .........................71 
7.7 
Assessment of Safety ..........................................................................................72 
7.8 
Assessment of Efficacy .......................................................................................72 


LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
iii 
8.0 
ADVERSE EVENTS .................................................................................74 
8.1 
Definitions ...........................................................................................................74 
8.2 
Documentation of non-serious AEs or SARs ......................................................76 
8.3 
SAEs or Serious SARs ........................................................................................76 
8.4 
Adverse Event Reporting ....................................................................................76 
8.5 
Data and Safety Monitoring Plan ........................................................................79 
9.0 
STATISTICAL CONSIDERATIONS .....................................................80 
9.1 
Study Design/Study Endpoints ............................................................................80 
9.2 
Sample Size, Accrual and Duration of Accrual ..................................................81 
9.3 
Data Analysis Plans .............................................................................................81 
10.0 
STUDY MANAGEMENT .....................................................................84 
10.1 
Institutional Review Board (IRB) Approval and Consent ..................................84 
10.2 
Required Documentation.....................................................................................85 
10.3 
Registration Procedures.......................................................................................85 
10.4 
Data Management and Monitoring/Auditing ......................................................85 
10.5 
Adherence to the Protocol ...................................................................................85 
10.6 
Amendments to the Protocol ...............................................................................87 
10.7 
Record Retention .................................................................................................87 
10.8 
Obligations of Investigators ................................................................................87 
11.0 
REFERENCES .......................................................................................88 
12.0 
APPENDICES ........................................................................................97 
APPENDIX A  PROHIBITED MEDICATIONS FOR ENTINOSTAT .....................97 
APPENDIX B  PATIENT HANDOUT-PROHIBITED MEDICATIONS .................98 
APPENDIX C  ARCHIVED TUMOR TISSUE FOR ENROLLMENT ....................99 


LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
4 
1.0 
BACKGROUND AND RATIONALE 
1.1 
Study Synopsis 
This is an exploratory (n=10-12 evaluable patients), open-label, single-arm, phase 
II study in patients with ‘non-inflamed’ unresectable regional or distant metastatic 
melanomas irrespective of prior treatment with PD-1/PD-L1 pathway inhibitors. 
The primary endpoint is to assess the incidence of histopathologic conversion of 
non-inflamed melanomas from patients with metastatic melanoma to ‘inflamed’ 
melanoma following single-agent entinostat “priming” (entinostat monotherapy). 
More specifically, patients with metastatic melanomas that have no evidence of 
tumor-infiltrating or tumor-associated lymphocytes (TIL/TAL), based on standard 
hematoxylin and eosin (H&E) staining of representative tumor sections (non-
inflamed melanomas) will receive weekly entinostat for 3 weeks in cycle 1 
(entinostat monotherapy; 5mg PO, qwk on D1, D8, D15 of cycle 1; cycle length = 
21 days). Mandatory tumor tissue biopsies will be performed in the end of cycle 1, 
beginning of cycle 2 (day 22±2 days), immediately before treatment with 
concurrent entinostat (once weekly × 3) and pembrolizumab (200 mg q3wks) in 
cycles 2-9 (see section 5.2.1 drug dosing schema). Correlative studies will be 
performed at baseline and in the end of cycle 1, beginning of cycle 2 (day 22±2 
days) to assess whether: (a) 3 weeks of entinostat monotherapy converts TIL/TAL-
absent melanomas to TIL/TAL-present by histopathologic (H&E stain) analysis, 
(b) 3 weeks of single-agent entinostat induces changes in gene expression profiling 
by assessing distinct signatures as defined by RNA-sequencing signatures (RNA-
seq; ‘immune-high’, innate anti-PD-1 resistance [IPRES], and  epigenetic 1-3), (c) 
3 weeks of single-agent entinostat induces changes in histone-accessible DNA by 
performing formaldehyde-assisted isolation of regulatory elements (FAIRE)4, (d) 
conversion of non-inflamed to inflamed melanomas by single-agent entinostat 
and/or antitumor response to the entinostat-pembrolizumab combination is 
associated with a baseline signature that consists of distinct somatic mutation gene 
profile, global histone modification profile in melanoma tissue or peripheral blood 
mononuclear cells (PBMC), and/or abundance of entinostat targets in melanoma 
cells (HDAC 1, 2, 3, and 11). Up to 12 evaluable patients will be treated with 
entinostat plus pembrolizumab for up to 27 weeks (approximately 6 months) based 
on clinical benefit. Patients who continue to have clinical benefit at 27 weeks may 
continue therapy with pembrolizumab or any other PD-1/PD-L1 inhibitor, as per 
standard of care. 
 
Secondary exploratory endpoints involve: (a) assessment of antitumor response of 
entinostat administered concurrently with pembrolizumab at week 10(or earlier if 
patient progresses), based on response rate per Response Evaluation Criteria in 
Solid Tumors version 1.1 (RECIST v1.1). (b) assessment of the progression-free 
survival (PFS) rate at 27 weeks (approximately 6 months) of the entinostat-
pembrolizumab combination. (c) determine the safety of the entinostat-
pembrolizumab combination in patients with metastatic melanoma per NCI 
Common Terminology Criteria for Adverse Events (NCI-CTCAE). (d) assessment 


LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
5 
of other exploratory biomarkers in tumor tissue and peripheral blood (see 
‘Secondary Correlative Studies”). 
 
1.2 
Disease Background 
1.2.1  Melanoma Heterogeneity 
Melanoma is derived from malignant transformation of melanocytes, neural crest-
derived cells that produce melanin and at least for the skin protect basal cells 
of the skin from the effects of ultraviolet radiation (UVR).  Cutaneous melanoma 
represents the most common melanoma subtype (approximately 95%); other non-
cutaneous melanomas are mucosal (1.3% total; conjunctival, sinonasal cavity, 
anorectal, oral cavity)5, and ocular (3-5%; choroidal, iris, and ciliary body)6. 
Alternative to the clinical-anatomic heterogeneity, melanomas can be 
histogenetically classified as epithelium-associated (high UVR-induced  lentigo 
maligna, desmoplastic; low UVR-induced non-chronic sun damage melanomas, 
spitzoid melanomas; mucosal; and acral melanomas) and non-epithelium-
associated (uveal, visceral, blue-nevus-like, melanoma in congenital nevus). 
Irrespective of the method of classification, melanoma subtypes exhibit stark 
molecular differences (e.g. high incidence of BRAF codon V600, NRAS codon 12, 
13, 61 mutations in cutaneous; low incidence of BRAFV600, NRAS codon 12, 13, 
51 mutations in mucosal; high incidence of cKIT mutations in acral; high incidence 
of GNAQ/GNAS mutations in ocular melanomas)7-10 as well as differences in 
underlying host immune response9-11. For example, the Cancer Genome Atlas 
Project (TCGA) in Cutaneous Melanoma (n=329) has shown that approximately 
50% of melanomas bear the ‘inflamed’ phenotype that is defined by the presence 
of a pleomorphic immune cell population that includes lymphocytic and other 
stromal infiltrate and contains various co-stimulatory and co-inhibitory immune 
checkpoint proteins8. In contrast, the TCGA Project in Primary Ocular Melanoma 
(n=80) showed that the incidence of TIL was only in 13% of primary ocular 
melanoma specimens 11. Finally, a single institution analysis in primary oral 
mucosal melanomas showed that high density of TIL was present in only 22% of 
patients 12. Interestingly, the heterogeneity of host immune response across 
different melanoma subtypes is a poor prognostic factor. More specifically, 
melanomas with high density of TIL have better overall survival and vice versa. 
The stark differences in the somatic mutation burden, distinct somatic mutations, 
and host immune response among different melanoma subtypes 7,10,13 suggests that 
aberrations other than genetic changes may play a role. An increasing number of 
reports summarize that epigenetic dysregulation by means of microRNA 
expression, gene hypermethylation, and histone modification plays a role across 
various melanoma subtypes 14,15.  
 
 


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1.2.2  Differential Responses of Various Melanoma Subtypes to Available Systemic 
Treatments; Interplay with Underlying Biology and Host Immune Response 
Existing systemic treatments for metastatic melanoma exhibit different clinical 
benefit across different melanoma subtypes, and irrespective of treatment type (i.e. 
immunotherapies versus targeted therapies). For example, a large proportion of 
patients with metastatic cutaneous melanoma who receive any type of systemic 
treatment have both high response rates and durable clinical benefit [concurrent 
dabrafenib and trametinib, response rate (RR) 67%, 3-year OS rate 44%; 
single-agent ipilimumab, RR 7-19%, 3-year OS rate 22%; single-agent 
nivolumab, RR 43.7-44%, 3-year OS rate 35%; single-agent pembrolizumab 
RR 33%, 3-year OS rate 38-43%; concurrent nivolumab and ipilimumab RR 
52-57.7%, 2-year OS rate67%] 16-21. However, RR in mucosal/acral (m/a) and 
ocular (o) melanoma to immune checkpoint inhibitors is lower [single-agent 
ipilimumab (m/a)0-8.3%, single-agent pembrolizumab (m/a)25%, concurrent 
pembrolizumab 
plus 
ipilimumab 
(m/a)20%; 
single-agent 
nivolumab 
(m)23.2%, concurrent nivolumab plus ipilimumab (m) 37.2%; single-agent 
PD-1 inhibitors (a) 32%, single-agent PD-1 inhibitors (m)23%; single-agent 
D-1/PD-L1 inhibitors (o) 2.6%; concurrent ipilimumab plus nivolumab (o)0] 
22-26. Lower responses have also been seen in mucosal or acral melanomas that are 
BRAFV600-mutant to MAPK inhibitors (BRAF inhibitors (m)38.1%, 
(a)20%) 27. Interestingly, presence of TIL has been predictive of response to 
BRAF inhibitors (BRAFV600-mutant), ipilimumab, and PD-1 inhibitors in 
cutaneous metastatic melanoma28-32. Given that available systemic treatments have 
better clinical benefit in melanomas with pre-existing host immune response 
(inflamed) and the incidence of inflamed melanomas differs according to 
melanoma subtype (e.g. in the two melanoma TCGA projects inflamed cutaneous 
melanomas were approximately 51% whereas inflamed ocular melanomas were 
12% respectively), these data imply that such treatments may have clinical benefit 
in a subset of all melanomas. It is therefore accurate to postulate that by being able 
to alter the biology of melanomas towards a more immunogenic/’inflamed’ 
phenotype, it is possible to augment the response of immunotherapies to non-
inflamed melanomas which do not usually respond to systemic treatments. 
Such strategies are expected to expand the therapeutic benefit in the non-‘inflamed’ 
melanomas.  
 
1.3 
Current Standard of Care 
Multiple systemic treatments have now been approved in metastatic cutaneous 
melanoma, such as immunotherapies {high-dose bolus IL-233, CTLA-4 inhibitors 
(ipilimumab)34, PD-1 inhibitors (pembrolizumab, nivolumab)35-37], targeted 
therapies [BRAF inhibitors (dabrafenib, vemurafenib) and MEK inhibitors 
(trametinib, cobimetinib)38,39]}, and chemotherapies (dacarbazine)40. BRAF 
inhibitors and MEK inhibitors are used in combination (i.e. vemurafenib plus 
cobimetinib and dabrafenib with trametinib) because of their superiority over 
single-agent BRAF inhibitors in patients with BRAFV600-mutant metastatic 


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melanoma17,39. No predictive biomarkers exist for any immunotherapy type, and 
BRAFV600 mutation status or any other distinct somatic mutation does not 
influence response to such treatments36,37. However, underlying host immune 
response alone and/or in association with intratumoral expression of PD-L1 in the 
vicinity of TIL may predict better responses and clinical benefit to any 
immunotherapy, in particular to single-agent PD-1 inhibitors29,32. Concurrent 
blockade of CTLA-4 (ipilimumab) and PD-1 (nivolumab or pembrolizumab) has 
shown to have higher antitumor responses compared to single-target blockade and 
with toxicity that is life-threatening, but dependent on the ipilimumab dose 21,41. 
However, no overall survival (OS) benefit was seen in the randomized study 
between concurrent ipilimumab-nivolumab versus nivolumab alone, except for the 
unplanned subgroup analysis of patients with PD-L1 negative tumors42.  
There are no established guidelines regarding the sequence of systemic therapies 
(immunotherapies first followed by targeted therapies or vice versa), which is 
currently the primary aim of the intergroup EA6134 randomized ongoing study. 
Upfront single-agent CTLA-4 or PD-1 inhibitors followed by MAPK inhibitors 
conferred longer OS compared to the reverse sequence in patients with 
BRAFV600-mutant melanoma 43,44. Guidelines regarding upfront concurrent PD-1 
and CTLA-4 blockade, as opposed to single PD-1 blockade, are not defined at this 
time, although symptomatic disease or fast melanoma kinetics favor their combined 
use. It appears though that resistance to one immunotherapy type (ipilimumab, PD-
1 inhibitor, or high dose bolus IL-2) does not predict cross-resistance to any other 
immunotherapy type 45,46. Upfront treatment with PD-1 inhibitors (as opposed to 
ipilimumab) is associated with longer OS than upfront treatment with ipilimumab47.  
Non-cutaneous melanoma subtypes are conventionally treated similarly to 
cutaneous melanoma, although most registration studies did not include non-
cutaneous melanoma subtypes. As summarized in section 1.2.2, immunotherapies 
consistently underperform in the non-cutaneous melanoma subtypes (and acral 
melanoma), suggesting the need to improve the therapeutic benefit of 
immunotherapies. 
As of June 2018, efforts to improve the clinical benefit of immune checkpoint 
inhibitors in melanoma focus on the following strategies: 
a) combine PD-1/PD-L1 pathway inhibitors with MAPK inhibitors in 
BRAFV600-mutant (NCT02908672, atezolizumab plus vemurafenib plus 
cobimetinib48; NCT02902042, pembrolizumab, encorafenib, binimetinib), 
b) combine PD-1/PD-L1 pathway inhibitors with other non-CTLA-4 blocking 
antibody therapies49, 
c) 
combine 
PD-1/PD-L1 
pathway 
inhibitors 
with 
drugs 
targeting 
immunoregulatory components of the tumor microenvironment  (e.g. IDO 
inhibitors50), and 
d) adoptive T cell therapy in combination with lymphodepletion (fludarabine 
plus cyclophosphamide) and high dose bolus IL-251. 


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These approaches are expected to enhance the therapeutic benefit of immune 
checkpoint inhibitors by bypassing issues of local immune tolerance. However, 
with the exception of treatment strategies that directly target melanoma (e.g. MAPK 
inhibitors) immunotherapeutic-based strategies are not expected to have an effect, 
if melanomas are not immunogenic 52. Based on the estimate from TCGA in 
cutaneous melanomas, lack of immunogenicity is seen in up to 50% of cutaneous 
melanoma cases 13. Cancer immunogenicity and antigenicity is a complex 
phenomenon that dynamically changes as cancers develop, establish and progress. 
While during early stages of cancer development both immunoreactive and non-
immunoreactive tumor clones may be present, the process of cancer immunoediting 
may gradually eliminate the former whereas the latter may be left unchecked and 
grow 53.  Another important issue is that immunogenicity and antigenicity are not 
strictly dependent on the number of somatic mutations, although admittedly various 
studies have shown a moderate but significant benefit from immune checkpoint 
inhibitors in patients with high neoantigen burden and across various cancers 54. 
There is a growing number of reports that resistance to various FDA-approved 
treatments in metastatic melanoma –both targeted therapies and immunotherapies– 
is also transcriptionally/epigenetically mediated 55. Therefore, it is tempting to 
speculate that drugs which affect transcription may potentially render less 
immunoreactive tumors to more immunoreactive.  
  
1.4 
Investigational Treatments 
 
1.4.1  Epigenetic Dysregulation and not Somatic Mutations Per Se May Account 
for Poor Immunogenicity of Melanomas and Resistance to PD-1 Inhibitors 
Understanding resistance to PD-1 inhibitors, the systemic therapy with the largest 
clinical benefit-to-toxicity ratio in melanoma, is of great importance to expand the 
clinical benefit in metastatic melanoma and other cancers. With the possible 
exception of BRCA-2, no genes were more frequently mutated in responders versus 
non responders to PD-1 inhibitors 2,56. Transcriptional analysis of tumors collected 
from patients who responded versus those who resisted PD-1 inhibitors showed that 
gene signatures associated with mesenchymal transition (AXL, ROR2, WNT5A, 
LOXL2, 
TWIST2, 
TAGLN, 
FAP, 
loss 
of 
E-cadherin), 
immunosuppression/angiogenesis 
(IL10, 
VEGFA, 
VEGFC), 
monocyte/macrophage chemotactic genes (CCL2, CCL7, CCL8, CCL13), and low 
MHC class II expression was associated with poor response to PD1 inhibitors 2,57,58. 
This suggests that epigenetic, in addition to genetic changes, by melanoma cells 
may drive the PD-1 inhibitor resistance phenotype. 
 
1.4.2  Overview of Epigenetics with Focus on Histone Deacetylases 
Epigenetic regulation involves heritable modifications to DNA that alter gene 
expression and chromatin structure without changes to the underlying nucleotide 


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DNA sequence. This non-genetic type of regulation shifts chromatin between two 
interchangeable states: closed (hetero-chromatin) and open (eu-chromatin). 
Depending on the chromatic structure genes can be switched ‘on’ and ‘off’ by 
allowing/restricting access and/or function of components of the transcriptional 
machinery (RNA polymerase and DNA-binding transcription factors)59. Epigenetic 
dysregulation is now a well-established mechanism of melanoma development and 
progression. Epigenetic changes include: (1) DNA methylation in CpG 
dinucleotides enriched in regions known as ‘CpG islands’. This process is more 
permanent, because methylated gene promoters along with the recruitment of bulky 
MB methyl-CpG-binding proteins by means of direct steric hindrance block 
binding of transcription factors and RNA polymerase leading up to a closed 
heterochromatin state, and therefore transcriptional repression/silencing. For 
example, the promoter of cell cycle genes corresponding to the CDKN2A gene 
locus is hypermethylated in cutaneous and uveal melanomas 60. In addition, 
promoters of more than 50 other genes with tumor suppressive function are 
hypermethylated during melanoma development, progression, and metastases 
(reviewed in 61). To make matters more complex, hypomethylation, both global as 
well as gene-specific, can also occur; in the case of melanoma, the promoter of 
genes encoding several cancer testis and melanoma-associated antigens can be 
hypomethylated resulting in increased expression (reviewed in 61). (2) histone 
modifications which are more dynamic, diverse, and are mediated by multiple 
mechanisms, such as post-translational modifications of histone proteins 
(acetylation, methylation, phosphorylation, and ubiquitination), ATP-dependent 
chromatin remodeling complexes, histone variant exchanges, and the action of non-
coding RNA (e.g. miRNA). Histone methylation is more complex to predict the 
effect in gene transcription because it is dependent on the extent of methylation 
[e.g. mono- (me1), di- (me2), or tri-methylation (me3)]. It is important to mention 
that promoter methylation can affect histone activity and, vice versa, histone 
acetylation affects promoter methylation levels62.  
Reversible acetylation of lysine residues on the tails and other more internal 
residues of histone proteins, H3 and H4 (e.g. H3K9, H3K14, H4K5, H4K16), is the 
best studied type of histone modification and is catalyzed by the transfer of acetyl 
group from acetyl-CoA to the ε-amino group at the terminal of the lysine side chain 
by means of histone acetyl transferases (HAT). The addition of the acetyl group 
neutralizes the lysine’s positive charge, thereby weakening the interaction with the 
negatively charged DNA, and leads to a more open chromatin structure allowing 
easier access to transcription factors and increased transcription. Histone 
acetylation also influences transcription by acting as a binding target for histone 
reader groups and regulates (prevents) deposition of other histone marks, such as 
the transcriptionally permissive H3K4me3 or the repressive H3K27me3 
modification63. On the other hand, these acetyl groups are removed by histone 
deacetylases (HDACs). HDACs are divided into 4 classes on the basis of structural 
homology to yeast HDACs, mechanism of enzyme activity, and cellular 
localization. The ‘classic’, or type I, are ubiquitously expressed in all cell types and 
mostly localized to the nucleus (1, 2, 3, and 8). Type II are more restricted in distinct 


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cell types (smooth muscle, heart, pancreas, and brain) and shuttle between the 
nucleus and cytosol (4, 5, 6, 7, 9, and 10). Type III (NAD-dependent sirtuins 1-7) 
possess dual enzymatic activity that includes mono-ADP-ribosyltransferase, in 
addition to the HDAC activity. In addition, type III HDACs have various 
subcellular organizations other than the nucleus (cytoplasm, mitochondria, and 
nucleolus). Class IV HDACs have similar localization to class II (HDAC11)64.  
The clinical significance of HDACs in cancer is that HDAC1, 2, 3, and 6 are 
upregulated in various cancers. Compared to normal skin and nevi, primary 
cutaneous melanomas overexpress HDAC1, 2, and 365. However, in metastatic 
cutaneous melanoma, expression of class I HDACs can be upregulated, but without 
any prognostic significance 66. Finally, HDACs sand HATs have histone-
independent targets, such as HSP90, TP53, and the NFκB subunit p65 with 
significant cellular effects. For example, hyperacetylation of HSP90 via class I 
HDAC inhibition may lead to degradation of signaling molecules (c-RAF, Akt) or 
upstream receptor tyrosine kinases (ERBB1 and ERBB2) and may therefore 
account for resistance to MAPK inhibitors. Hyperacetylation of TP53 may 
similarly alter melanoma resistance to therapy whereas NF-κB activation may alter 
cytokine production, anti-apoptotic protein transcription, and immune response67.   
1.4.3  Histone Acetylation, HDAC Inhibitors and Direct Effects in Melanoma Cells 
As expected, HDAC inhibitors (HDACi) modulate expression of thousands of 
genes in melanoma.  More specifically, HDACi inhibit expression of anti-apoptotic 
proteins survivin, bcl-XL, blc-2, Mcl-1, XIAP, reduce expression of cFLIP, an 
inhibitor of TNF-related apoptosis, and induce expression of pro-apoptotic proteins 
BIM, BAX, BAK, and CDKN1A68. Equally important, class I HDACi may alter 
immunogenicity by upregulating expression of melanoma differentiation antigens, 
class I and class II MHC antigens, antigen processing and presenting machinery 
(TAP1, TAP2, LMP2, LMP7, tapasin, CIIA), costimulatory molecules (CD40, 
CD86), ligands of activating receptor on NK cells (MICA, MICB), and PD-L1/PD-
L2 genes in vitro 69-72. The ultimate effect of HDAC inhibition in melanoma can 
be suppression of cell proliferation, increased immunogenicity, and increased 
apoptosis.  
 
1.4.4  Histone Acetylation, HDAC Inhibitors and Effects on Host Immune 
Response 
It is important to emphasize that DNA methylation and histone modifications 
regulate development, differentiation, activation, and memory lineage function of 
host immune response 73. In particular, selective demethylation of several specific 
CpG dinucleotides in the promoter region of IFN-γ, IL-3 and IL-13 genes regulate 
T helper cell differentiation into Th1, Th2, Th17 and induction of regulatory T cells. 
Signaling through cytokines activates transcription factors (STAT4, STAT6, 
GATA-4, T-bet), which facilitates chromatin remodeling (long-range H4 
acetylation, H3K9 trimethylation) at the enhancer regions of T helper cell-specific 
genes and ultimately influences activation and T cell differentiation (reviewed in 
74).  


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Not surprisingly, HDACi have pleiotropic effects on immune cells. Therefore, the 
net effect remains ill-defined as HDACi can also impair immune surveillance and 
induce hematologic toxicity in cancer patients depending on the dose, type and 
selectivity of HDACi used. For example, panobinostat can have differential effects 
on T cell viability and function, depending on the dose used 75,76. Similar dose-
dependent results were seen with entinostat 77. Nevertheless, several positive effects 
of HDACi were seen, such as the activation and inhibition of apoptosis of CD4+ 
cells78, CD8+ cells79, macrophages80, T regulatory cells81, and myeloid-derived 
suppressor cells (MDSC) 82. 
 
1.4.5  Preclinical Evidence about the Role of Histone Deacetylation Inhibition in 
Melanoma 
Treatment with a pan-HDACi (LBH589) enhanced survival and proliferation of 
tumor antigen-specific T cells and suppressed the T regulatory (Treg) population 
when coadministered with gp100-specific T cells in a B16 melanoma model 79. The 
pan-HDACi lost its antitumor effect in immunodeficient as opposed to 
immunocompetent mice 70. Concurrent LBH589 (15mg/kg, M-W-F) and PD-1 
blockade (3mg/kg, Tu-Th) in C57BL/6 mice showed delayed growth and enhanced 
survival of B16-F10-bearing melanoma tumors69, similar to results seen in a lung 
adenocarcinoma model83. Entinostat has effects in MDSC 82,84, Treg 81, and 
dendritic cells85. When administered as a single-agent orally no antitumor 
responses were seen in patients with metastatic melanoma86.  
 
1.4.6  Entinostat 
 
1.4.6.1 Non-Clinical studies 
Entinostat (pyridine-3-yl)methyl((4-[(2-aminophenyl)carbamoyl]phenyl)methyl) 
carbamate), or SNDX-275, MS-275 is an orally available inhibitor of HDACs 1, 2, 
3, and 11 [IC50 (nM) 124, 139, 187, 427, respectively]. The selectivity of entinostat 
for these HDAC isoforms may account for its better safety and efficacy compared 
to other nonselective pan-HDAC inhibitors87,88. Exposure of various cancer cell 
lines to entinostat resulted in accumulation of hyperacetylated histones at 
concentration between 0.3-1 µM and gene expression changes in various cell 
processes, such as cell growth, apoptosis, differentiation, cell communication, 
regulation of transcription, cell signaling, and chromosome organization. Entinostat 
demonstrated broad, dose-dependent, tumor-inhibitory effects against cancer cell 
lines with IC50 values ranging between 0.04-4.71 µM. Entinostat showed tumor 
growth inhibition in human tumor xenografts from diverse cancer types, such as 
melanoma, non-small cell lung cancer, and breast cancer. The maximum tolerated 
dose in the animal experiments was 50mg/kg daily. 
As predicted by its epigenetic mechanism of action, entinostat may restore 
sensitivity in resistant cancer cell lines in vitro and in vivo when combined with 
various classes of anticancer agents, such as chemotherapies, anti-estrogens, EGFR 
inhibitors, DNA methyltransferase (DNMT) inhibitors and immunotherapies. 


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Examples of entinostat in combination with hormonal therapies (aromatase 
inhibitor, selective estrogen modulators) and DNA methylase transferase inhibitors 
(‘dual epigenetic blockade’) are provided in the Investigators’ Brochure and 
elsewhere89,90.  
Proof-of-principle experiments in syngeneic murine models that bear poor or 
modestly immunogenic tumors suggest that resistance to concurrent CTLA-4 and 
PD-1 blockade can be restored with entinostat. In these models primary targets of 
entinostat were actually circulating granulocytic MDSC, which are known to be 
immunosuppressive, whereas the function of CD8+ cells remained intact82. In other 
preclinical models entinostat reduced transcription of FoxP3 in circulating Treg, an 
important transcription factor for survival and function of this other 
immunosuppressive population81. In other clinical models entinostat can induce 
favorable changes in effector T cell response against cancer cells91. The entinostat-
induced reduction of circulating MDSC was confirmed in a clinical study of 
entinostat in combination with aromasin compared with aromasin alone 84. 
Reduction of circulating Tregs and monocytic MDSC was also seen in a clinical 
trial of entinostat administered in combination with high dose bolus IL-2 in patients 
with advanced renal cell carcinoma 92. Entinostat also showed single-agent activity 
against xenograft melanoma models93 and in combination with temozolomide65 and 
interleukin-294.  
Entinostat administered in rats at doses comparable to those currently used in 
clinical studies had no effects in various physiologic systems (cardiovascular, 
central nervous, respiratory, GI, and urogenital), with the exception of diuresis and 
increased excretion of Na+, K+, Cl-, and creatinine.  
Pharmacokinetic studies of entinostat administered across various animal models 
show considerable differences with respect to oral bioavailability, serum half-life 
(t ½), area-under-curve (AUC) exposure, peak serum concentrations (Cmax), time-
to-peak serum concentrations (tmax). Food decreased absorption of entinostat. The 
highest serum concentrations were seen in liver, kidney, spleen, and glandular 
tissue. Entinostat was moderately stable in preparation of human hepatocytes and 
liver microsomes. Reversible inhibition by entinostat was demonstrated for 
CYP2C8 and CYP3A4 and glycuronidation was the major method of metabolism. 
Entinostat is excreted equally by urine and biliary tract.  
Toxicologic studies of entinostat across various animals suggested effects on the 
skin, atrophy of lymphatic organs, myelosuppression, GI toxicity, loss of appetite, 
weight loss, changes in renal function, hypercholesterolemia, reduced fertility, and 
teratogenicity.  
 
1.4.6.2 Effects of Entinostat in Humans 
As of September 29 2016, Entinostat has been administered in >1,000 cancer 
patients across pharma-sponsored studies (Syndax, Syndax predecessors, NCI, 
Kyowa Hakko Kirin Co., Ltd -the company that bears development and 
commercialization rights of entinostat in Japan and Korea). This number does not 


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include patients who were treated as part of investigator-initiated studies. In 
particular, clinical studies investigated entinostat as monotherapy [6 studies had 
reportable data at the time; 4 studies in patients with solid tumors (n=128), and 2 
studies in patients with hematologic malignancies (n=89)] and in combination with 
other systemic treatments [22 studies had reportable data at the time; 17 studies in 
patients with solid tumors (n=535) and 5 in patients with hematologic malignancies 
(n=205)]. 
1.4.6.2.1 Pharmacokinetic Studies 
Pharmacokinetic data, which were compiled from 4 studies, showed linear 
dose-dependent pharmacokinetics based on single and repeated dose 
administrations. Entinostat absorption is facilitated by low pH in the stomach 
and its metabolism is mediated by UGT 1A4, but not by CYP enzymes. 
Entinostat mean terminal t1/2 was approximately 194 hours based on the 
radiolabeled absorption, distribution, metabolism, and excretion (ADME) study 
with the majority of the radioactive dose excreted in urine (71%) and feces 
(12%). Entinostat and/or its metabolites partition preferentially into 
erythrocytes, with greater affinity to binding sites on erythrocytes relative to 
those in plasma. In fact, the slow dissociation rate of entinostat from 
erythrocytes may account for the long t½ of entinostat.  
1.4.6.2.2 Safety in Human Clinical Studies 
An integrated analysis across all industry-sponsored and NCI-sponsored studies 
with available safety data showed that single-agent entinostat was well tolerated 
at the doses and schedules investigated. The most frequent adverse effects (AE), 
predominantly grade 1-2, included fatigue, nausea, vomiting, anorexia, 
diarrhea, hematologic abnormalities (anemia, thrombocytopenia, neutropenia, 
leukopenia), 
and 
metabolic 
abnormalities 
(hypoalbuminemia, 
hypophosphatemia, hyperglycemia, and hyponatremia). Certain types of AEs 
were more frequent in patients with solid tumors than in those with hematologic 
malignancies; for example, hypoalbuminemia (59%), fatigue (58%), nausea 
(57%), hypophosphatemia (45%), anemia (42%), and thrombocytopenia (41%) 
were more frequent in patients with solid tumors whereas thrombocytopenia 
(52%), anemia (36%), neutropenia (36%), fatigue (35%), and nausea (34%) 
were more frequent in patients with advanced hematologic malignancies. 
Similar patterns and incidence for serious adverse events (SAEs, grade 3 and 4) 
were seen for all AEs in patients with solid tumors versus hematologic 
malignancies. The most SAEs occurring in >5% of patients with solid tumors 
included fatigue (13%), dyspnea (9%), dehydration (9%), infection (9%), 
anemia (7%) and neutropenia (7%). The most common SAEs in patients with 
hematologic malignancies were neutropenic infection (15%) and febrile 
neutropenia (12%). Fatal (grade 5) events were febrile neutropenia, respiratory 
failure, neutropenic infection, and disease progression; almost all occurred in 
patients with hematologic malignancies. 
Entinostat has been combined with other systemic treatments [aromatase 
inhibitors (n=111), erlotinib (n=99), 5-azacytidine (n=319), other agents (13-


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cis-retinoic acid, interleukin-2, sorafenib, lapatinib, trastuzumab, nivolumab, 
ipilimumab; n=211)]. In general, AEs were seen regardless of indication, such 
as fatigue, nausea, anemia, thrombocytopenia, leukopenia, diarrhea, vomiting, 
and neutropenia. Again, patients with hematologic malignancies trended to 
have higher incidences of both AEs and SAEs (62% versus 45%). The incidence 
of SAEs was also high in patients receiving entinostat in combination with 
azacytidine (43%). The most frequent SAEs that occurred in entinostat-based 
studies in 535 patients with solid tumors were decreased absolute lymphocyte 
count (7%), fatigue (7%), dyspnea (6%), and anemia (5%). Fatal AEs were 
reported in 15% of the 740 patients treated with entinostat in combination with 
other agents and were at least twice more frequent in patients with hematologic 
malignancies compared to those with solid tumors. However, approximately 
90% of the fatal AEs were attributed to disease progression.  
1.4.6.2. Efficacy in Cancer Clinical Studies 
Table 1 shows several phase I/II studies of entinostat administered in various 
schedules alone or in combination with systemic therapies with different 
mechanisms of action (hormonal, targeted, dual epigenetic blockade, 
immunomodulators). Entinostat exhibited clinical benefit either as monotherapy or 
in combination with other therapies, including cancer types with resistance to 
standard of care therapies. Based on these results entinostat (5mg po qwk) is 
currently being investigated in combination with exemestane (25mg po qd) in 
patients with advanced hormone receptor-positive, HER2-negative breast cancer 
with prior disease progression on a non-steroidal aromatase inhibition as part of a 
randomized double-blind (entinostat vs. placebo) 600-patient phase III study 
(E2112, NCT02115282). In particular, entinostat has shown promising activity in 
combination with high dose bolus IL-2 in renal cell carcinoma as well as with 
pembrolizumab in patients with PD-1 inhibitor refractory non-uveal melanoma. 
Entinostat is currently being evaluated in combination with avelumab (epithelial 
ovarian cancer, NCT02915523), pembrolizumab (MDS, after DNMTi therapy 
failure, NCT02936752; NSCLC and mismatch repair-proficient colorectal cancer, 
NCT02437136; uveal melanoma, NCT02697630; relapsed/refractory lymphomas, 
NCT03179930), atezolizumab (triple negative breast cancer, NCT02708680), 
atezolizumab plus bevacizumab (renal cell carcinoma, NCT03024437), and with 
nivolumab-ipilimumab (HER2-negative breast cancer, NCT02453620).  
 
Drug 
Cancer Type (stage) 
Schedule 
Aim 
N 
Efficacy 
Ref 
En  
Melanoma, nonuveal 
(IV) 
3mg d1, d15 
7mg d1, d8, d15 
 
RR/TTP 
28 
0% RR  
TTP 50d 
(mdn) 
86 
En 
HD, 
relapsed/refractory 
10mg (or 15mg) qow 
15mg qwk (3/4 wks) 
RR 
49 
12% 
95 
Ex ± 
En 
Breast cancer, ER+, 
postmenopausal (IV) 
En 5mg qwk  
Ex 25mg qd,  
PFS 
130 
Ex+En 4.3m 
(mdn) 
Ex  2.3m 
(mdn) 
96 


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Drug 
Cancer Type (stage) 
Schedule 
Aim 
N 
Efficacy 
Ref 
Er + 
En 
NSCLC resistant to 
chemotx, no prior 
EGFRi (IIIB/IV) 
En 10mg  d1, d15 
Er 150mg qd 
4m PFS rate 
132 
Er+En 18% 
Er 20% 
97 
Lap + 
trast + 
En 
Breast cancer, fail 
trast, prior lap (IV) 
En 12mg qow 
Lap 1,000mg qd 
Trast 86mg/kg 
q3wk 
Clinical 
benefit at 6m 
(CR+PR+SD) 
22 
54% 
98 
Aza + 
En 
Metastatic refractory 
CRC (IV) 
En 7mg qwk OR d3, 
d10 
Aza 40mg/m2 d1-5 & 
d8-10 
RR 
47 
0% 
99 
Aza + 
En 
Breast cancer, HR+ or  
TNBC (IV) 
En 7mg qwk d3, d10 
Aza 40mg/m2 d1-5 & 
d8-10 
RR 
52 
HR      5% 
TNBC 0% 
100 
Aza ± 
En 
MDS, AML 
Aza 50 mg/m2/d qd 
d1-1 
En 4 mg/m2/d d3 
d10. 
Rate hem nl 
149 
Aza 32% 
Aza + En 27% 
101 
Aza ± 
En 
NSCLC recurrent 
advanced 
En 7mg d3 d10  
Aza 30 or 40mg/m2/d 
sc 
RR 
19 
21% 
102 
GM-
CSF + 
En 
MDS 
relapsed/refractory 
AML 
En 4mg/m2 d1, d8, 
d15, d22 
GM-CSF 
125µgr/m2/qd sc 
RR 
28 
39% 
103 
Pembro 
+ En 
Non-uveal melanoma, 
PD1-inhibtor 
refractory (IV) 
En 3 OR 5mg qwk 
Pembro 200mg q3wk 
RR 
13 
30% 
104 
hdIL-2 
+ En 
RCC (IV) 
En 5mg qwk 
IL-2 (600,000u/kg 
q8hrs) 
RR 
44 
34% 
92 
 
Table 1. Phase I/II studies of entinostat alone or in combination with various other anticancer treatments. Only 
efficacy data are shown. Abbreviations: En, entinostat; PK, pharmacokinetics; RR, response rate; TTP, time-to-
progression; mdn, median; HD, Hodgkin’s disease; Ex, exemestane; ER+, estrogen receptor positive; PFS, progression-
free survival; NSCLC, non-small cell lung cancer; chemotx, chemotherapy; EGFRi, inhibitor targeting epidermal growth 
factor receptor; Lap, lapatinib; trast, trastuzumab; CR, complete response; PR, partial response; SD, stable disease; Aza, 
azacytidine; CRC, colorectal cancer; HR, hormone receptor positive; TNBC, triple-negative breast cancer; 
myelodysplastic syndrome; AML, acute myelogenous leukemia; rate hem nl, rate of hematologic normalization; hdIL-2, 
high dose bolus IL-2; RCC, renal cell carcinoma. 
1.5 
Pembrolizumab (MK-3475) 
Pembrolizumab (MK-3475, KeytrudaTM) is a potent and highly selective 
intravenous humanized mAb of the immunoglobulin (Ig) G4/kappa isotype that 
directly blocks the interaction between PD-1 and its ligands, PD-L1 and PD-L2. 
This blockade enhances functional activity of the target lymphocytes to facilitate 
an antitumor immune response, leading to tumor regression and immune rejection 
of the tumor. Pembrolizumab was approved by the U.S FDA as a frontline therapy 
in metastatic melanoma, metastatic non-small cell lung (NSCLC) cancers, recurrent 
or metastatic squamous cell carcinoma of the head and neck, recurrent locally 


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advanced or metastatic gastric/gastroesophageal junction adenocarcinoma whose 
tumors express PD-L1,  adult or mismatch repair-deficient solid tumors that have 
progressed following prior treatment, locally advanced or metastatic urothelial 
carcinoma who have disease progression during or following platinum-containing 
chemotherapy, in combination with pemetrexed and carboplatin for treatment of 
patients with previously untreated metastatic non-squamous non-small cell lung 
cancer, adult and pediatric patients with refractory classical Hodgkin lymphoma or 
those who have relapsed after 3 or more prior lines of therapy as outlined in the 
package 
insert 
available 
at 
http://www.merck.com/product/usa/pi_circulars/k/keytruda/keytruda_pi.pdf. 
 
The importance of intact immune surveillance in controlling outgrowth of 
neoplastic transformation has been known for decades 105.  Accumulating evidence 
shows a correlation between TILs in cancer tissue and favorable prognosis in 
various malignancies 106-110.  In particular, the presence of CD8+ T-cells and the 
ratio of CD8+ effector T-cells / FoxP3+ Treg cells correlates with improved 
prognosis and long-term survival in several solid tumors. 
 
The PD-1/PD-L1 pathway is a major co-inhibitory immune checkpoint pathway 
that suppresses effector T cell immune response in various cancers.  PD-1 is one of 
the several co-inhibitory immune checkpoint proteins that is normally upregulated 
on the cell surface of activated T-cells under physiologic conditions to 
downregulate unwanted or excessive immune responses against exogenous stimuli 
(viruses, bacteria, cancer), including autoimmune reactions.  PD-1 is encoded by 
the gene (PDCD1) and is an Ig superfamily member related to CD28 and CTLA-4. 
The structure of its murine counterpart reveals that it is a type I transmembrane 
glycoprotein that contains an Ig variable-type (V-type) domain responsible for 
ligand binding and a cytoplasmic tail that is responsible for binding of signaling 
molecules.  The cytoplasmic tail of PD-1 contains two tyrosine-based signaling 
motifs, an immunoreceptor tyrosine-based inhibition motif (ITIM) and an 
immunoreceptor tyrosine-based switch motif (ITSM). Following T-cell 
stimulation, PD-1 recruits the tyrosine phosphatases SHP-1 and SHP-2 to the ITSM 
motif within its cytoplasmic tail, leading to the dephosphorylation of effector 
molecules, such as CD3ζ, PKCθ and ZAP70, all of which are involved in the T-cell 
signaling cascade 111-114.  The mechanism by which PD-1 down modulates T-cell 
responses is similar to, but distinct from that of CTLA-4, as both molecules regulate 
an overlapping set of signaling proteins [13; 14].  PD-1 is expressed on activated 
lymphocytes, including peripheral CD4+ and CD8+ T-cells, B-cells, Treg and 
natural killer cells 115,116. 
 
The ligands for PD-1 (PD-L1 and PD-L2) are constitutively expressed or can be 
induced in a variety of cell types, including non-hematopoietic tissues as well as in 
various tumors 117-120.  Both ligands are type I transmembrane receptors containing 
both IgV- and Ig (constant) C-like domains in the extracellular region and short 
cytoplasmic regions with no known signaling motifs.  Binding of either PD-1 ligand 


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to PD-1 inhibits T-cell activation triggered through the T-cell receptor.  PD-L1 is 
expressed at low levels across various non-hematopoietic tissues, most notably on 
vascular endothelium, whereas PD-L2 protein is only detectable on antigen-
presenting cells found in lymphoid tissue or chronic inflammatory environments.  
PD-L2 is thought to control immune T-cell activation in lymphoid organs, whereas 
PD-L1 dampens unwarranted T-cell function in peripheral tissues 121.  Although 
healthy organs express little, if any, PD-L1, several cancers, including melanoma 
can overexpress PD-L1 by means of amplification of the gene locus that contains 
the PD-L1 gene 122. It is important to emphasize that this mechanism of 
upregulation is independent from the reactive expression of PD-L1 in response to 
host immune response, which may have important therapeutic implications 123.  
1.5.1 Pre-clinical Studies of Pembrolizumab  
Pembrolizumab (MK-3475) strongly enhances T-lymphocyte immune responses 
in cultured blood cells from healthy human donors, cancer patients, and 
primates. In in vitro assays using T-cells isolated from human donor blood cells, 
pembrolizumab induces production of interleukin-2 (IL-2), tumor necrosis factor 
alpha (TNFα), interferon gamma (IFNγ), and activates T cells in subnanomolar 
EC50 concentrations (i.e. concentrations where 50% of the maximum effect is 
achieved; ~0.1 - 0.3 nM).  It is important to emphasize that pembrolizumab 
potentiates existing immune responses only in the presence of antigen and 
specifically activate T-cells 124. 
 
Using a murine antibody against PD-1 (mDX400), PD-1 blockade has been shown 
to significantly inhibit tumor growth in a variety of syngeneic murine tumor 
models. In these mouse experiments, mDX400 treatment is synergistic with 
chemotherapeutic agents, such as gemcitabine and 5-fluorouracil (5-FU), and 
combination therapy results in increased complete tumor regression rates in vivo 
124. 
 
The safety of pembrolizumab was characterized in the one-month, repeated-dose 
toxicity study in cynomolgus monkeys when administered as intravenous (IV) 
doses of 6, 40 or 200 mg/kg once a week (a total of five doses) and in the 6-month 
repeat-dose toxicity study in the same species when administered as IV doses of 
6, 40 or 200 mg/kg every other week (a total of 12 doses).  Pembrolizumab was 
well tolerated with a systemic exposure [area under the curve (AUC)] of up to 
~170,000 μg/day/mL over the course of the one-month study, and with an AUC of 
up to approximately 67,500 μg/day/mL over the course of the 6-month study. No 
findings of toxicological significance were observed in either study and the No 
Observed Adverse Event Level (NOAEL) was ≥200 mg/kg.   In addition, no 
findings of toxicological relevance were observed in the in vitro tissue cross-
reactivity study using human and cynomolgus monkey tissues 124. 
1.5.2 Summary from Relevant Clinical Studies of Pembrolizumab in Melanoma 
In the largest ever conducted disease-specific phase I study of an experimental drug 
(n=655, KEYNOTE-001) pembrolizumab was administered in various schedules 


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in patients with advanced metastatic melanoma 35,125. This phase I study was 
essentially comprised of 4 different cohorts. The first (n=135) was a study in which 
patients were treated with 3 different dose schedules; 2mg/kg q3wks, 10mg/kg 
q3wks, and 10mg/kg q2wks, irrespective of prior treatment with ipilimumab. The 
second (n=173) was a randomized study of patients who have previously received 
and have become resistant to ipilimumab (ipilimumab -treated; IPI-T), defined as 
administration of at least 2 ipilimumab doses and equivocal progressive disease 
within 6 months of first ipilimumab dose; patients were randomized to 2mg/kg 
q3wks versus 10mg/kg q3wks. The third (n=103) was a randomized study of 
patients who have never received ipilimumab treatment (IPI naïve; IPI-N).  The 
fourth (n=244) was a randomized study of patients who have been IPI-N and IPI-T 
to either 10mg/kg q3wks vs. 10mg/kg q2wks. At a cutoff date of April 18, 2015, 
the median duration of follow-up was 15 months (range 8-29) and 244 patients 
(37%) were still receiving pembrolizumab. IPI-T (n=342) as opposed to IPI-N 
(n=313) patients had received a smaller number of pembrolizumab infusions (8 
versus 11 doses). No treatment-related deaths were observed in this trial and no 
differences in grade 3-4 treatment-related events were observed between IPI-T 
versus IPI-N. The most frequent immune-mediated adverse events were 
hypothyroidism (any grade, 7.5%; grade 3-4, 0.2%), hyperthyroidism (2.3%, 
0.3%), pneumonitis (2.7%, 0.3%), colitis (1.7%, 1.1%), hepatitis (0.6%, 0.3%), 
nephritis (0.5%, 0.3%), and uveitis (0.9%, 0%). As of October 18, 2014, and with 
a median follow-up of 21 months, 581 patients had measurable disease per central 
RECIST v 1.1 at baseline. In this combined cohort of IPI-T and IPI-N overall 
response rates (ORR) were assessed at 33% whereas complete response (CR) was 
8%. Subgroup analyses revealed that M1b disease, tumor burden less than the 
median seen in this study, no prior treatment with ipilimumab, and normal serum 
LDH were associated with RR greater than 33% whereas prior treatment with 
BRAF inhibitors, tumor burden higher than the median seen in this study, M1c 
disease, and elevated serum LDH were associated with a significantly lower 
incidence of ORR. In this subgroup analysis, there were no significant differences 
in ORR between the 2mg/kg q3wk (n=143), 10mg/kg q3wk (n=272), and 10mg/kg 
q2wks (n=166).  The median PFS was 4.4 months (PFS rate at 12 months was 0.35), 
whereas the median OS was 22.8 months (OS rate at 12 and 24 months was 0.66 
and 0.49, respectively)126. 
 
When pembrolizumab was administered as first-line therapy in patients with 
advanced cutaneous melanoma, CR and ORR were 13.5% and 45.1%, respectively, 
whereas the corresponding median PFS was 13.8 months and OS was 31.1 months, 
respectively. In this study tumor blocks from 67% of the first 411 patients who were 
enrolled into this study were evaluable for immunohistochemical (IHC) detection 
of PD-L1 in the tumor. Allred proportion scores (APS) were randomly set for 
expression of PD-L1; APS=0, corresponds to 0% staining, APS=2 corresponds to 
1-10% staining, APS=3 corresponds to 10-33% staining, APS=4 corresponds to 
34%-66% staining, and APS=5 corresponds to 66-100% staining. Patients with 
APS=0 or 1 (n=28, n=24, respectively) had ORR <10% each, patients with APS=2 
(n=72) had ORR ~ 20%, patients with APS=3 (n=54) had ORR 40%, patients with 


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APS=4 (n=32) had ORR ~70%, and patients with APS=5 (n=32) had ORR ~ 50% 
35. Pembrolizumab monotherapy had lower clinical benefit when administered in 
patients with metastatic ocular or metastatic mucosal melanoma23,24. The lower 
responses in these two non-cutaneous melanoma subtypes was attributed to the 
lower frequency of ‘inflamed’ types. Pembrolizumab was safe and showing 
promising antitumor activity in 18 patients with asymptomatic, up to 20-mm active 
melanoma brain metastases (22% incidence of intracranial antitumor response)127.  
 
KEYNOTE-006 was a randomized study in which patients with advanced 
melanoma and no prior treatment with CTLA-4, PD-1, and PD-L1 inhibitors were 
randomized to pembrolizumab 10mg/kg administered every 2 wks (n=279) or every 
3 wks (n=277) versus standard of care ipilimumab (3mg/kg, n=278). In the 
intention-to-treat population the estimated 6-month PFS rate for patients receiving 
pembrolizumab every 2 or 3 weeks was 47.3% and 46.4%, respectively, whereas 
the corresponding 6-month PFS rate for ipilimumab-treated patients was 26.5%. 
Median estimates of PFS were 5.5 months, 4.1 months, and 2.8 months, 
respectively. At the time of data cut-off for the second interim analysis, which was 
driven by a minimum follow-up duration of 12 months for all patients, one-year 
estimates for OS were 74.1%, 68.4%, and 58.2%, respectively. At a longer follow-
up (median 22.9 months) median OS was not reached for any of the pembrolizumab 
arms but was 16 months for the ipilimumab arm128. The results from this study 
suggest that pembrolizumab administered at 10mg/kg irrespective of the schedules 
tested was superior to ipilimumab in the front-line setting (i.e. no prior treatment 
with co-inhibitory immune checkpoint inhibitors) 35. 
 
KEYNOTE-002 is a randomized study in which patients with advanced melanoma 
who have previously received and have become resistant to ipilimumab and MAPK 
pathway inhibitors (if BRAFV600-mutant) were randomized to receive 
pembrolizumab, 2mg/kg every 3wks (n=180), pembrolizumab, 10mg/kg every 3 
wks (n=181), and investigator’s choice chemotherapy (n=171; paclitaxel plus 
carboplatin, paclitaxel, carboplatin, dacarbazine, or oral temozolomide). At a 
median follow-up duration of 10 months pembrolizumab administered at either 
2mg/kg or 10mg/kg showed significant improvement in PFS compared to 
investigator’s choice chemotherapy (hazard ratio or 0.57 and 0.50, respectively). 
RR in patients who received pembrolizumab at 2mg/kg, 10mg/kg versus 
investigator’s choice chemotherapy were 21%, 25%, and 4%, respectively, whereas 
the median PFS was 5.4 months, 5.8 months, and 3.6 months, respectively. 
Incidence of grade 3-4 treatment-related AEs was higher in those given 
chemotherapy (26%) than in those given pembrolizumab at 2mg/kg (11%) and 
pembrolizumab at 10mg/kg (14%). The most common grade 3-4 treatment-related 
AEs in the pembrolizumab 2 mg/kg group were fatigue, generalized edema, and 
myalgia whereas in the pembrolizumab 10mg/kg group common grade 3-4 events 
were hypopituitarism, colitis, diarrhea, decreased appetite, hyponatremia, and 
pneumonitis. In the pre-specified subgroup analysis of PFS the only subgroup that 
did not appear to significantly benefit from pembrolizumab 2mg/kg was the 
BRAFV600-mutant group 37. 


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In summary, pembrolizumab administered at 10mg/kg provides better clinical 
benefit than the standard of care ipilimumab in the front-line setting (i.e., no prior 
co-inhibitory immune checkpoint inhibitors). Although the FDA-approved 2mg/kg 
every 3 weeks dose was never directly compared head-to-head against ipilimumab, 
data from KEYNOTE-001 and KEYNOTE-002 show that there is no significant 
difference between pembrolizumab administered at 2mg/kg and 10mg/kg every 3 
wks. Currently, pembrolizumab is administered at 200mg IV, flat/fixed dose every 
3 weeks. 
 
1.6 
Rationale for Clinical Study 
No immunotherapies in metastatic cutaneous melanoma have achieved an 
antitumor response rate more than 60%. Furthermore, combined ipilimumab and 
nivolumab has not resulted in longer OS compared with single-agent nivolumab in 
a randomized phase III study42. The ‘up to 60% ceiling effect’ of combined PD-
1/CTLA-4 blockade can be explained by the fact that pre-existing TIL and 
immunogenicity are required for most of these immunotherapies to have an effect, 
which was set to approximately 50% in the case of cutaneous melanomas13. The 
requirement for underlying immune response for clinical benefit from 
immunotherapies can be better recognized indirectly among other non-cutaneous 
melanoma subtypes, in which the incidence of underlying immune response is 
much lower compared to that seen in cutaneous melanoma. Unless we attempt to 
develop strategies to augment immunogenicity of non-‘inflamed’ melanomas, the 
clinical benefit from any immunotherapy regimen will predominantly remain 
limited to inflamed melanomas only. Although immunogenicity is to some extent 
defined by the number of high-avidity neoantigens, it is also regulated at the 
transcriptional level.  
Epigenetic modifiers that affect gene expression via changes in promoter 
methylation or chromatin (e.g. histone) structure have shown to augment 
immunogenicity of cancer cells129. HDACi are expected to augment 
immunogenicity of melanoma cells in a subgroup that is likely not to be defined by 
somatic mutations in most genes (overall burden and type) but rather by melanoma 
biology that is defined by distinct changes in DNA promoter methylation and 
regulatory elements that are regulated by histone marks (acetylation and/or 
methylation); these changes will ultimately be reflected by the RNA/miRNA 
expression profiling 58. We predict that a subset of non-inflamed melanomas treated 
with entinostat will ultimately become immunogenic and that this conversion can 
be predicted by a combined biomarker that involves: (a) a handful of somatic 
mutations in epigenetic modifiers and epigenetic mediators (e.g. OCT4, NANOG, 
LIN28, SOX2, KLF4), which have been previously identified in cutaneous 
melanomas130, (b) DNA promoter methylation profiling, and (c) changes in 
chromatin accessibility of distinct regulatory DNA elements. 
Clinical trials in metastatic melanoma using single-agent HDACi have not shown 
any major antitumor responses 86,131. In addition, two trials using HDACi in 


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combination with PD-1 inhibitors were recently reported104,132. While the 
entinostat-based treatment combination with pembrolizumab showed activity in a 
portion of patients with PD-1 inhibitor resistance, the panobinostat trial did not 
show any clinical benefit in patients with PD-1 inhibitor-naïve melanoma. We 
interpret these results as follows: 
a. Potency and selectivity of the HDACi matters. Entinostat (a class I HDACi) is 
more selective than panobinostat (pan-HDACi) yet more potent for class I HDACs 
133.  
b. The type of immune checkpoint inhibitor that the HDACi is concurrently 
administered with may matter. In the panobinostat trial, ipilimumab was used. 
c. Given that entinostat/pembrolizumab-treated patients with PD-1 inhibitor 
refractory melanoma did respond, identification of pretreatment and on-treatment 
changes in response to entinostat at the tumor tissue level should be pursued in 
parallel with investigations that explore entinostat effects on various peripheral 
blood immune cell (PBMC) subsets. Given the complex changes that HDACi 
induce, a comprehensive –omics strategy focusing on epigenetic changes (DNA 
methylation profiling, profiling of regulatory DNA elements that are free from 
nucleosomes, RNA-seq) at baseline and on-treatment may be important for 
identifying a biomarker. 
 
Dose Rationale: We will use the standard of care FDA-approved dose of 
pembrolizumab (200 mg, once every 3 weeks) along with the entinostat 
administered at 5 mg once weekly (qwk). This dose of entinostat was safe and well 
tolerated in combination with pembrolizumab in an ongoing phase 1b/2 study104. 
104.  
1.7 
Correlative Studies 
The study contains a single integral tumor tissue biomarker and several exploratory 
tumor tissue and peripheral correlative biomarkers.  
1.7.1 Integral Biomarker. Assessment of TIL/TAL in representative H&E-stained 
sections from archived tumor tissues to determine eligibility for the trial 
during screening.  
Since the primary endpoint of the study is to assess the incidence of conversion of 
non-inflamed vs. inflamed melanoma following 3 weeks of entinostat 
monotherapy, we will use a cost-efficient assay to determine eligibility, namely 
assessment of TIL/TALs in archived metastatic melanoma specimens. Since the 
presence of TIL/TALs can be heterogeneous within the same tissue as well as 
across different metastatic sites, the following are requirements for archived tumor 
tissue eligibility: 
• The amount of surface area corresponding to melanoma devoid of stroma 
should be at least 1cm2. This ensures sufficient tumor to be screened for 
presence of TIL/TALs, yet it is NOT too restrictive for patients who do not 
have “sufficiently large” archived tumor specimen for analysis. 


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• No more than 20% of necrosis is allowed; this ensures that high quality 
specimens are available for analysis. 
• At least 60% of viable melanoma cells are present; this ensures that the 
melanoma cell compartment is sufficiently larger than the stromal 
component. 
• Must have available archival tissue and subjects must have consented to 
allow collection of archived tumor blocks from previous surgeries 
confirming or treating unresectable stage III or distant metastatic disease. If 
more than one archived tumor block is available, only one block is required 
to be analyzed for the presence of TIL/TALS. Archived tumor tissues must 
fulfill the following criteria based on representative H&E-stained tissue 
sections: (1) the tumor surface area must be ≥1cm2 (which can be the sum 
tumor surface from more than metastatic blocks, if necessary), (2) ≤20% of 
necrosis, (3) the ratio of viable tumor cells to tumor-associated stroma 
should be ≥60/40, and (4) if TILs are present based on H&E stained 
sections, they must be ≤1% of the total number of cells in the specimen. 
• If more than one archived metastatic melanoma specimen is available 
during the 21-day screening period, only one block is required to be 
analyzed for the presence of TIL/TALs. 
1.7.2 Exploratory Biomarkers-Tumor Tissue 
 
1.7.2.1 RNA-seq (both baseline and in the end of cycle 1, beginning of cycle 2 
(day 22±2 days), immediately before concurrent entinostat-pembrolizumab 
treatment day 22 ± 2 days).  
RNA will be extracted from FFPE tissue sections from both archived in the end 
of cycle 1, beginning of cycle 2 (day 22±2 days), immediately before treatment 
for sequencing analysis. RNA-seq on paired baseline and on day 22±2 treatment 
tissues will assist in: (a) identifying transcription signatures that are associated 
with non-inflamed melanomas, such as IPRES and immune low signatures 
(baseline), type I interferon response, IPRES and epigenetic signatures (day 22 
versus baseline) 2,3,13. Secondly, (b) perform a somatic mutation calling 
(expressed mutations) analysis, in particular for genes encoding for epigenetic 
modifiers (e.g. SMARCA4, PBRM1, ARID1A, ARID2, ARID1B, DNMT3A, 
TET2, MLL1/2/3, NSD1/2, EZH2, SETD2, BRD4) and epigenetic regulators 
(e.g. OCT4, NANOG, LIN28, SOX2, KLF4). We predict that inactivating 
mutations in any of these genes may have impact in chromatin accessibility, as 
has been previously published in renal cell carcinoma from UNC-CH 
investigators 134, and therefore these tumors may differentially respond to 
HDAC inhibition. RNA-seq is also ideal for detection of aberrant splicing and 
expressed immunogenic neoepitopes 135 (Dr. Benjamin Vincent UNC-CH, 
unpublished data). This process can therefore help us study immunogenicity of 
tumors that may have a low somatic tumor burden. It is therefore quite possible 
that HDAC inhibition may change RNA expression of genes that may have low 
or absent expression at baseline due to widespread RNA processing defects, but 


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upon treatment with a HDACi, RNA expression of these genes may be 
significantly upregulated leading to increased expression of neoepitopes. 
Thirdly, (c) analyze the immune receptor repertoire (richness, observed V-J 
rearrangements; clonality, evenness of the repertoire based on frequencies of 
specific V-J rearrangements) within tumors. The T-cell receptor (TCR) 
repertoire in peripheral blood has been previously associated with response to 
immune checkpoint inhibitors 136-139. We postulate that TCR and B-cell receptor 
(BCR) repertoire analysis within tumor tissue will yield more specific 
TCR/BCR clones that are associated with immune response in the end of cycle 
1, beginning of cycle 2 (day 22±2 days), immediately before treatment samples 
from the prospective responders, compared to the prospective non-responders. 
 
 
1.7.2.2 Formaldehyde-assisted isolation of regulatory elements (FAIRE; 
baseline and on day 22).  
In tumors, organization of the chromatin landscape offers a comprehensive 
overview of the rewiring of transcriptional networks that accompany processes 
such as cellular differentiation, tumor growth, metastasis, and therapeutic 
resistance, which can be more reliable in predicting cell behavior than gene 
expression profiles alone140,141.  Further, it has been shown that chromatin 
accessibility changes can be detected in cancer cells following treatment with small 
molecule inhibitors142-144.  Therefore, tumor-specific chromatin signatures may 
offer remarkable diagnostic and biomarker potential.  We will use a protocol 
developed by the Pattenden, Dayton, and Davis labs at UNC-CH to extract 
DNA accessible chromatin from formalin-fixed, paraffin-embedded (FFPE) 
tissue. These DNA regions harbor regulatory elements (active transcriptional 
start sites, transcriptional enhancers, insulators, silencers, and locus control 
regions). Chromatin signatures will be determined by performing FAIRE, a 
biochemical technique that purifies accessible chromatin from formalin-fixed 
samples, followed by high-throughput sequencing. 
 
1.7.2.3 Tissue-imaging studies 
1.7.2.3.1 Expression of PD-L1 (melanoma cells and immune cells), MHC 
class I (melanoma cells) and CD8+ cells at baseline and on day 22, using 
multicolor immunofluorescence (IF). If our integrated biomarker of 
assessing TIL/TAL is correct, we do not anticipate detecting any CD8+ cells 
at baseline; in fact, a proportion of these non-inflamed melanomas may have 
no expression of MHC class I. However, in the end of cycle 1, beginning of 
cycle 2 (day 22±2 days), immediately before concurrent pembrolizumab-
entinostat treatment, we anticipate that HDAC inhibition in those 
melanomas that have TIL/TAL present, based on the H&E analysis (integral 
biomarker), to also contain CD8+ cells. In addition, we anticipate that 
melanoma cells in patients who respond to concurrent entinostat-
pembrolizumab will express both MHC class I+ and PD-L1, which have 
been previously shown to be upregulated in response to HDACi.  
 


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1.7.2.3.2 Expression of global histone modification markers: acetylated 
histone 3 lysine 18 (H3K18Ac), acetylated histone 4 lysine 12 (H4K12Ac), 
dimethylated histone 4 arginine 3 (H4R3diMe), dimethylated histone 3 
lysine 4 (H3 K4diMe), (H3K27triMe) in tumor specimens by single-color 
immunohistochemistry (IHC) will be important due to previous reports 
about association between global histone modification levels and 
prognosis145,146. Both archived/baseline and day 22 samples will be stained 
to assess both prediction of response to entinostat as well as entinostat-
associated changes in global histone modifications. We anticipate that at 
least the global histone acetylation markers H3K18Ac and H4K12Ac to be 
high at baseline in a subset of patients whose melanomas convert from ‘non-
inflamed’ to ‘inflamed’ and that 21±2 days of entinostat priming will 
significantly suppress H3K18Ac and H4K12Ac. Lack of suppression of the 
histone acetylation markers may suggest inadequate target inhibition by 
entinostat. 
 
1.7.2.3.3 Expression of entinostat targets HDAC1, HDAC2, HDAC3 in 
archived-baseline tumor tissues only by single-color IHC will be examined 
due to reports about heterogeneous expression of HDAC1, HDAC2, and 
HDAC3 in melanoma 66. We would like to investigate whether expression 
of any of these targets, HDAC1, HDAC2, HDAC3, is associated with 
treatment response to the entinostat-pembrolizumab combination. 
 
1.7.2.4 Exploratory Biomarkers-Peripheral blood mononuclear cells 
(PBMC; baseline, in the end of cycle 1, beginning of cycle 2 (day 22±2 days), 
immediately before concurrent entinostat-pembrolizumab; week 10; and 
end of treatment): 
a. It is quite possible that entinostat will have an indirect 
immunomodulatory effect on immune cell subsets, much like the 
suppression of peripheral blood MDSC in breast cancer patients84. We 
plan to perform multi-parameter flow cytometry on PBMCs to identify 
various immunoregulatory cell subsets (e.g. monocytic and granulocytic 
MDSCs, T reg cells). 
b. Assess Histone H3 and H4 acetylation in PBMCs by peripheral blood 
multi-parameter flow cytometry 147,148. In the ENCORE 301 trial 
patients bearing peripheral immune cell subsets (T cells, B cells, 
monocytes) that were found to be hyperacetylated and were treated with 
entinostat and exemestane lived longer compared to non-acetylators. It 
would be interesting to see if TIL/TAL-absent melanomas that convert 
from absent TIL/TALs before entinostat treatment to TIL/TAL-present 
tumors following 21 days of entinostat monotherapy exhibit (a) higher 
levels of H3 and H4 acetylation in peripheral blood compared to non-
converters at baseline b) undergo the greatest changes in acetylation 
between baseline and day 22, immediately prior to pembrolizumab 
treatment.  
 










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System 
Laboratory Value 
International Normalized Ratio (INR) or 
Prothrombin Time (PT) 
 
 
 
 
Activated Partial Thromboplastin Time 
(aPTT) 
≤1.5xULN, unless subject is receiving 
anticoagulant therapy as long as PT or PTT is 
within therapeutic range of intended use of 
anticoagulants 
 
≤1.5xULN, unless subject is receiving 
anticoagulant therapy, as long as PT or PTT is 
within therapeutic range of intended use of 
anticoagulants 
*Note: Hematology and other lab parameters that are ≤ grade 2 BUT still meet 
criteria for study entry are allowed. Furthermore, changes in laboratory parameters 
during the study should not be considered AEs unless they meet criteria for dose 
modification(s) of study medication outlined by the protocol in Section 5.3.1 and/or 
worsen from baseline during therapy.   
4.1.10 A female of childbearing potential must have a negative serum pregnancy test 
during screening and a negative urine pregnancy test within 3 days prior to 
receiving the first dose of study drug. If the screening serum test is done within 3 
days prior to receiving the first dose of study drug, a urine test is not required. A 
female of childbearing potential must agree to use effective contraception during 
the study and for 120 days after the last dose of study drug.  A female of non-
childbearing potential defined as (by other than medical reasons): 
 ≥45 years of age and has not had menses for >2 years, 
 
 Amenorrheic for <2 years without a hysterectomy and oophorectomy and a 
follicle-stimulating hormone value in the postmenopausal range upon pre-study 
(screening) evaluation, 
 Post hysterectomy, oophorectomy or tubal ligation. Documented hysterectomy 
or oophorectomy must be confirmed with medical records of the actual 
procedure or confirmed by an ultrasound. Tubal ligation must be confirmed 
with medical records of the actual procedure; otherwise the patient must be 
willing to use 2 adequate barrier methods throughout the study, starting with 
the screening visit through 120 days after the last dose of entinostat, 
 If male, agrees to use an adequate method of contraception starting with the 
first dose of study drug through 120 days after the last dose of entinostat, 
 See section 5.6.2 for information on acceptable methods of contraception. 
4.1.11 Experienced resolution of toxic effect(s) of the most recent prior anti-cancer 
therapy to Grade ≤1 (except alopecia or neuropathy) by CTCAE v5.0. Exceptions 


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are patients who may have developed autoimmune-type side effects that require 
permanent hormonal replacement from previous therapies.  
4.1.12 If the patient underwent major surgery or radiation therapy, these procedures must 
have occurred at least 15 days prior to the first dose of entinostat. In addition, 
patients must have recovered from the toxicity and/or complications from the 
intervention to Grade ≤1. 
4.1.13 Subjects must be willing and able to comply with study procedures based on the 
judgement of the investigator or protocol designee.   
4.2 
Exclusion Criteria 
All subjects meeting any of the exclusion criteria at baseline will be excluded from 
study. 
4.2.1 Is receiving systemic steroid therapy or any other form of immunosuppressive 
therapy for autoimmune side effects related to previous use of immunotherapies for 
melanoma. Exceptions include episodic (up to 7 days) use of systemic steroids for 
common conditions while on study treatment (e.g. COPD exacerbation, poison 
ivy), use of corticosteroids as replacement doses for adrenal or pituitary 
insufficiency. 
4.2.2 Has a known history of tuberculosis (Bacillus Tuberculosis) or human 
immunodeficiency virus (HIV 1/2 antibodies). 
4.2.3 Hypersensitivity to pembrolizumab or any of its excipients. Allergy to benzamide 
or inactive components of entinostat. 
4.2.4 Has known history of biopsy-proven (non-infectious) pneumonitis that required 
systemic steroids, or any evidence of current pneumonitis. 
4.2.5 Conditions that would preclude adequate absorption of oral medications 
(malabsorption, significant nausea and vomiting, resection of >100-cm of proximal 
small bowel, resection of >200-cm of distant small bowel).  
4.2.6 Has a history or current evidence of any condition, therapy, or laboratory 
abnormality that might confound the results of the trial, interfere with the subject’s 
participation for the full duration of the trial, or is not in the best interest of the 
subject to participate, in the opinion of the treating investigator, including but not 
limited to: 
i. 
Myocardial infarction or arterial thromboembolic events within 6 months 
prior to screening or severe or unstable angina, New York Heart Association 
(NYHA) Class III of IV disease, or a QTc interval > 470 msec, 
ii. 
Active infection requiring systemic antibiotic therapy by the first day of 
entinostat treatment, 
iii. 
Known psychiatric or substance abuse disorders that would interfere with 
cooperation with the requirements of the trial. 


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4.2.7 If female, is pregnant or breastfeeding, or, if male, expecting to conceive or father 
children within the projected duration of the trial, starting with the pre-screening or 
screening visit through 120 days after the last dose of trial treatment. 
4.2.8 Known active hepatitis B (e.g., hepatitis B surface antigen-reactive) or hepatitis C 
(e.g., hepatitis C virus ribonucleic acid [qualitative]). Patients with past hepatitis B 
virus (HBV) infection or resolved HBV infection (defined as the presence of 
hepatitis B core antibody [HBc Ab] and absence of HBs Ag) are eligible. HBV 
DNA test must be performed in these patients prior to study treatment if known 
history of viral hepatitis. Patients positive for hepatitis C virus (HCV) antibody are 
eligible, only if polymerase chain reaction is negative for HCV RNA. 
4.2.9 Has received a live vaccine within 30 days of planned start of study therapy. 
Note:  Seasonal influenza vaccines for injection are generally inactivated flu 
vaccines and are allowed; however intranasal influenza vaccines (e.g., Flu-Mist®) 
are live-attenuated vaccines and are not allowed. 
4.2.10 History of prior malignancy, with the exception of the following: 
• Non-melanoma skin cancers, non-invasive bladder cancer, and carcinoma in situ of 
the cervix, 
• Prior history of prostate cancer provided the patient is not undergoing active 
systemic treatment other than hormonal therapy and has documented PSA that is 
undetectable (<0.2ng/mL), 
• Papillary thyroid cancer, even if patients may have just completed thyroidectomy 
within the last 2 years, have not received adjuvant radioactive iodine therapy, and 
were only recently diagnosed with asymptomatic papillary thyroid cancer and their 
surgery is pending, 
• Chronic lymphocytic leukemia (CLL) provided patient has isolated lymphocytosis 
(Rai stage 0) and does not require systemic treatment [for “B” symptoms, Richter’s 
transformation, lymphocyte doubling time (<6 months), lymphadenopathy or 
hepatosplenomegaly], 
• Lymphoma, hairy-cell leukemia, or myelodysplasia, provided that patient is not on 
active systemic treatment and is in complete remission, as evidenced by PET/CT 
scans and bone marrow biopsies for at least 3 months, 
• History of any other malignancy provided patient has completed therapy and is free 
of disease for ≥ 2 years. If patient had other malignancy within the last 2 years from 
which he, or she, may have been completely cured by surgery alone, he may be 


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considered to be enrolled on condition that the risk of development of distant 
metastatic disease based on the most recent AJCC staging system is less than 30%. 
4.2.11 Has known active (i.e. previously untreated) parenchymal central nervous system 
(CNS) metastases that are symptomatic, and/or more than one lesion with the 
largest diameter being > 5-mm and/or require antiepileptic drugs or systemic 
corticosteroids for management of intracranial symptoms. Patients with 
carcinomatous meningitis are also excluded.  Exceptions are: 
Subjects with previously treated brain metastases provided they are stable (i.e. 
without evidence of progression by brain MRI or head CT with IV contrast) for at 
least 2 weeks prior to the first dose of entinostat. Any neurologic symptoms must 
have returned to baseline, and have no evidence of new or enlarging brain 
metastases, and are not using ongoing systemic corticosteroids for management of 
intracranial symptoms for at least 7 days prior to first dose of entinostat, 
Patients with active (i.e. not treated with stereotactic radiosurgery), single, 
asymptomatic, up to 5-mm in largest diameter brain metastases (measured either 
by brain MRI with IV contrast or head CT with IV contrast measuring within 2 
weeks prior to the first dose of entinostat). 
4.2.12 Currently participating and receiving study therapy or has participated in a study of 
an investigational agent and received study therapy or used an investigational 
device within 4 weeks of the first dose of entinostat.   
4.2.13 Subject is receiving prohibited medications or treatments as listed in section 5.5 of 
the protocol that cannot be discontinued/replaced by an alternative therapy.  
4.2.14  Prior treatment with HDACi for their melanoma.   
 
5.0 
TREATMENT PLAN 
5.1 
Schema 
 
Eligible subjects must have histologically confirmed AJCC stage III/IV melanoma 
of any subtype and sufficient, archived metastatic tumor tissue to confirm absence 


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UNIVERSITY OF NORTH CAROLINA 
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of TIL/TAL in representative H&E-stained tumor tissues. Subjects are eligible even 
if they have received any prior PD-1/PD-L1 pathway inhibitors or any other 
systemic treatments, including immunotherapies, on condition that they have not 
received HDACi for their melanoma. They must have recovered from any side 
effects associated with prior anticancer treatments. Patients will be ‘primed’ with 
entinostat, 5mg PO qwk, for 3 weeks. In the end of cycle 1, beginning of cycle 2 
(day 22±2 days), immediately before concurrent entinostat-pembrolizumab 
treatment, they will undergo a mandatory melanoma biopsy after which they will 
receive up to 4 infusions of standard of care pembrolizumab along with weekly 
administration of entinostat. Restaging radiographic studies will be performed on 
week 10and then every 6 weeks until week 27.  Peripheral blood will be collected 
at baseline; in the end of cycle 1, beginning of cycle 2 (day 22±2 days), immediately 
before concurrent entinostat-pembrolizumab treatment; on week 10 after 3 
infusions (cycles) of pembrolizumab; and at the end of treatment, either at disease 
progression or week 27, or if intolerable toxicity occurs that requires permanent 
discontinuation of pembrolizumab and/or entinostat. Subjects will be considered to 
have completed all trial requirements if: 
(a) Their disease has progressed by RECIST v1.1 criteria within less than 10 
weeks from the first dose of entinostat, 
(b) They develop toxicity related to pembrolizumab or entinostat, but still 
complete sample collection (peripheral blood and tumor tissue) and tumor 
assessments by at least week 3, 
(c) Complete the entire 27 weeks of combination therapy and study-related 
assessments without experiencing disease progression by RECIST v1.1 criteria 
or clinical grounds. Those subjects experiencing clinical benefit after 27 weeks 
may continue pembrolizumab therapy per standard of care. 
5.2 
Treatment Dosage and Administration 
Subjects in this trial will be administered entinostat, 5mg PO, once weekly (D1, 
D8, D15 of a 21-day cycle) starting on day 1 of study treatment. Pembrolizumab, 
200 mg will be administered intravenously (IV) every 3 weeks and initiated at cycle 
2 after the mandatory research tumor biopsy in the end of cycle 1, beginning of 
cycle 2 (day 22±2 days). Combination therapy with both agents will continue if 
subject is receiving clinical benefit from therapy for up to 27 weeks (8 cycles of 
combination therapy or approximately 6 months).  Study therapy will be 
discontinued for intolerable toxicity, disease progression or for other reasons at the 
discretion of the investigator.   
 
 


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Agent 
Required 
Premedications; 
Precautions 
Dose 
Route 
Schedule 
Cycle 
Length 
Entinostat 
Take on an empty 
stomach i.e., at least 2 
hours after a meal and at 
least 1 hour before the 
next meal. 
5 
mg3 
Oral (PO) 
  
Start on study D1 
of cycle 1 
and administer 
once weekly (i.e., 
D1, D8, D15 of 
each cycle) for 9 
cycles qwk) (±1 
day) 
 
 
Every 
21* 
days  
 
  
Pembrolizumab 
Risk for immune-
mediated1 and infusion-
related2 reactions  
200 
mg3 
 
IV; administer over 
30 min  
(range -5, +10 min) 
Start on study 
D22 (± 2 days) of 
the study (i.e., D1 
of cycle 2) 
and administer 
every 21 (± 2) 
days (q3wk) x 8 
cycles (i.e., cycles 
2-9) 
1. See Section 5.3.2 for management guidelines for immune-mediated AEs. 
2. See Section 5.6.1 for management guidelines for infusion-related AEs 
3. On days when entinostat is administered on the same day as pembrolizumab, entinostat should 
be taken before the pembrolizumab infusion. 
 
*Entinostat monotherapy for cycle 1 and combination therapy should be initiated on D1 of cycle 2 and 
continue until withdrawal for toxicity, withdrawal for patient preference, disease progression or the subject 
completes 27 weeks of protocol-directed therapy. Entinostat cannot be continued beyond the 27-week 
duration of study-directed therapy for the trial, irrespective of treatment response. 
 
*Pembrolizumab will be started on D22 of the study (i.e., D1 of cycle 2) and may continue for up to 24 weeks 
in combination with entinostat (i.e. cycles 2-9). Following completion of the trial therapy at 27 weeks and on 
condition that there is no disease progression, pembrolizumab can be continued per investigator discretion, 
as per standard of care guidelines. 
5.2.1 Drug Dosing Schema 
 


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Figure 1. Study medications should be administered during the weeks denoted above.  See 
the schedule of assessments in Section 7.1 for further details. On days when entinostat is 
administered on the same day with pembrolizumab, entinostat should be administered 
before the infusion of pembrolizumab. 
 
5.3 
Toxicities and Dosing Delays/Dose Modifications 
Any subject who receives at least one dose of entinostat on this protocol will be 
evaluable for toxicity.  Each patient will be assessed periodically for development 
of any toxicity according to the Time and Events table in section 7.1.  Toxicity will 
be assessed according to the NCI-CTCAE v5.0. No dosing adjustments will be 
made for pembrolizumab, but delays in dosing or discontinuation of 
pembrolizumab and entinostat may be required. Depending on the side effects 
related to pembrolizumab administration, there would be either treatment delays 
(i.e., delays up to 3 days of scheduled dose if AE(s) occur prior to cycle 3 or 4, up 
to 7 days of scheduled dose allowed if AE(s) occurs prior to cycles 5 and 6; delays 
up to 15 days of scheduled dose allowed for pembrolizumab infusions if AE(s) 
occurs for cycle 7, 8 or 9 of combination treatment).  If pembrolizumab delays go 
beyond what is allowed, subject will skip the particular dose and a note will be 
made regarding the reason. If pembrolizumab is permanently discontinued, then 
entinostat must also be discontinued. Entinostat dosing guidelines and delays are 
provided in the section below.  
5.3.1 Entinostat Dosing Modification Guidelines 
All dose modifications should be based on the AEs requiring the greatest 
modification and also meet criteria for dose adjustment. AEs should be properly 
documented in source documents. Investigators may take a more conservative 
approach than the guidelines outlined below based on clinical judgment that is in 
the best interest of the subject. 
Management of toxicities that are at least possibly related to entinostat, with 
toxicities graded by the Investigator according to the NCI, CTCAE, version 5.0 
should be managed as follows:  
Non-hematologic Toxicity 
Toxicity 
Dose Modifications for Entinostat 
Grade 4 
Administer symptomatic remedies/start prophylaxis. 
Hold dose until recovery to Grade 1 or baseline under the following directions: 
1. 
If recovered within 4 weeks of onset (i.e.: ≤3 missed doses), 
resume study drug as follows: 
• 
If receiving 5 mg, restart study drug at 3 mg, 
• 
If receiving 3 mg, restart study drug at 2 mg, 
• 
If receiving 2 mg, discontinue study treatment. 
2. 
If not recovered within 4 weeks, permanently discontinue study drug. 


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Non-hematologic Toxicity 
Toxicity 
Dose Modifications for Entinostat 
Grade 3 
Administer symptomatic remedies/ start prophylaxis. Hold dose until recovery to 
Grade 1 or baseline under the following directions:  
1) If recovered within 1 week, resume study drug at prior dose. If not 
recovered within 1 week, continue to hold dose.  
2) If recovered within 2-4 weeks, resume study drug as follows:  
•       If receiving 5 mg, restart study drug at 3 mg,  
•       If receiving 3 mg, restart study drug at 2 mg, 
•       If receiving 2 mg, permanently discontinue study drug. 
3) If not recovered within 4 weeks, permanently discontinue study drug.  
Recurrence of the 
same ≥ Grade 3 
toxicity, despite 
dose reduction 
If the same ≥ Grade 3 event recurs: 
1) Administer symptomatic remedies/ start prophylaxis. Hold1 dose until 
recovery to Grade 1 or baseline. 
2) If recovered within 2 weeks, resume study drug as follows: 
a. 
    If receiving 5 mg, restart study drug at 3 mg, 
b.     If receiving 3 mg, restart study drug at 2 mg, 
c. 
    If receiving 2 mg, permanently discontinue study drug. 
3) If the same ≥ Grade 3 event recurs (i.e., third occurrence) despite 
entinostat dose reduction to 2 mg, as described above, discontinue 
study drug. 
≤Grade 2 
Administer symptomatic remedies/start prophylaxis. 
Dosing of study drug may be interrupted at the Investigator’s discretion. 
•     If dose is held for 4 consecutive weeks, permanently discontinue 
study drug1. 
•     If toxicity resolves, resume entinostat at the original dose. 
If greater than 50% of doses are missed during any 6-week period, discontinue from study 
drug treatment. Entinostat must also be discontinued in case of permanent discontinuation 
of pembrolizumab. 
Hematologic Toxicity 
Toxicity 
Dose Modifications for Entinostat 
≥Grade 3 
neutropenia, 
≥Grade 3 
uncomplicated 
thrombocytopenia, 
or Grade 2 
complicated 
thrombocytopenia 
Administer symptomatic remedies/start prophylaxis. 
Hold dose1 until recovery to Grade 1 or study baseline under the following direction:  
1) If not recovered by next scheduled dose, skip the dose. If recovered by 
next scheduled dose, resume study drug at prior dose. 
2) If receiving 2 mg dose, and not recovered by either of the next two 
scheduled doses, permanently discontinue study treatment. Otherwise, 
skip each dose. If recovered for either of these doses, resume study 
drug as follows: 
a. 
   If receiving 5 mg, restart study drug at 3 mg, 
b.    If receiving 3 mg, restart study drug at 2 mg. 
3) If not recovered within 4 weeks, permanently discontinue study drug. 
Recurrence of the 
same hematologic 
toxicity 
If the same hematologic toxicity recurs:  
1)  Administer symptomatic remedies/start prophylaxis. Hold1 dose until recovery 
to Grade 1 or baseline.  
2) If recovered within 2 weeks, resume study drug as follows:  
• 
If receiving 5 mg, restart study drug at 3 mg,  
• 
If receiving 3 mg, restart study drug at 2 mg,  
• 
If receiving 2 mg, permanently discontinue study drug.  
3) If the same ≥ Grade 3 event recurs (i.e., third occurrence), despite entinostat 
dose reduction to 2 mg as described above, permanently discontinue study drug.  


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If greater than 50% of doses are missed during any 6-week period, discontinue from study 
drug treatment. 
5.3.2 Dose Modification and toxicity management for immune-related AEs 
associated with pembrolizumab and combination therapy  
AE (both non-serious and serious) associated with pembrolizumab exposure, 
including coadministration with additional compounds, may represent an 
immunologic etiology. These immune-related AEs (irAEs)  may occur shortly after 
the first dose or several months after the last dose of pembrolizumab/combination 
treatment and may affect more than one body system simultaneously. Therefore, 
early recognition and initiation of treatment is critical to reduce complications.  
Based on existing clinical study data, most irAEs were reversible and could be 
managed 
with 
interruptions 
of 
pembrolizumab/combination 
treatment, 
administration of corticosteroids and/or other supportive care. For suspected irAEs, 
ensure adequate evaluation to confirm etiology or exclude other causes. Additional 
procedures or tests such as bronchoscopy, endoscopy, skin biopsy may be included 
as part of the evaluation. Dose modification and toxicity management guidelines 
for irAEs associated with pembrolizumab/combination treatment are provided in 
the table below. Pembrolizumab must be withheld for drug-related toxicities and 
severe or life-threatening AEs as per the table provided below. See Section 5.6 for 
supportive care guidelines, including use of corticosteroids and additional guidance 
on management of immune-mediated events (i.e., pneumonitis, diarrhea/colitis, 
Type 1 diabetes mellitus, hypophysitis, hyper/hypothyroidism, hepatotoxicity, 
renal insufficiency/nephritis, Steven’s-Johnson Syndrome, and myocarditis). 
 
Dosing interruptions are permitted in the case of medical/surgical events or 
logistical reasons not related to study therapy (e.g., elective surgery, radiation 
therapy, unrelated medical events, patient vacation, and/or holidays). Subjects 
should be placed back on study therapy within 3 weeks (i.e., 21 days) of the 
scheduled interruption, unless otherwise discussed with the Sponsor. The reason 
for interruption should be documented in the patient's study record. If 
pembrolizumab is permanently discontinued, entinostat must be discontinued as 
well. 
 
Attribution of Toxicity: 
 
When study interventions are administered in combination, attribution of an 
adverse event to a single component is likely to be difficult.  Therefore, while the 
investigator may attribute a toxicity event to the combination,  to entinostat alone 
or to pembrolizumab alone], for adverse events listed in the table below, both 
interventions must be held according to the criteria in the table below: Dose 
Modification and Toxicity Management Guidelines for Immune-Related Adverse 
Events Associated with Pembrolizumab and IO Combinations. 
 
Holding Study Interventions: 
 


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When study interventions are administered in combination, if the AE is considered 
immune-related, both interventions should be held according to recommended dose 
modifications. 
 
Restarting Study Interventions: 
 
Participants may not have any dose modifications (no change in dose or schedule) 
of pembrolizumab in this study, as described in the table below. 
• If the toxicity does not resolve or the criteria for resuming treatment are not 
met, the participant must be discontinued from all study interventions. 
• If the toxicities do resolve and conditions are aligned with what is defined 
in the table below, the combination of entinostat and pembrolizumab may 
be restarted at the discretion of the investigator. In these cases where the 
toxicity is attributed to the combination or to entinostat alone , re-initiation 
of pembrolizumab as a monotherapy may be considered at the principal 
investigator’s discretion . 












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5.3.3 Combined Entinostat and Pembrolizumab Dosing Guidelines 
In the ongoing study of concurrent pembrolizumab – entinostat administration in 
patients with solid tumors, no overlapping toxicities have been identified between 
these two drugs except for the following: fatigue, arthralgia, AST/ALT elevation, 
rash, and diarrhea. Overall, the treatment regimen has been tolerated well 104.  
 
• If any AEs with grade ≥3 are equally attributed to both pembrolizumab and 
entinostat, reduce entinostat and pembrolizumab as per dosing guidelines in 
sections 5.3.1 and 5.3.2.  
 
• As with any new treatment, we will also monitor for any AEs that have not 
been observed with this combination (unexpected SARs will be reported 
per guidelines) under study. 
 
• If pembrolizumab is permanently discontinued, then entinostat must also be 
discontinued. 
 
5.4 
Concomitant Medications/Treatments/Supportive Care Allowed 
All treatments that the investigator considers necessary for a subject’s welfare may 
be administered at his/her discretion, in keeping with the community standards of 
medical care.  All concomitant medications will be recorded on the case report form 
(CRF) including all prescriptions, over-the-counter (OTC), non-exclusionary 
herbal supplements, IV medications and IV fluids. If changes occur during the trial 
period, documentation of drug dosage, frequency, route, and date may also be 
included on the CRF. 
 
All concomitant medications received within 28 days before the first dose of trial 
treatment and 30 days after the last dose of trial treatment should be recorded.   
5.5 
Prohibited Medications/Treatments 
Subjects with pre-existing hypersensitivity to pembrolizumab or other PD-1/PD-
L1 pathway inhibitors are disqualified from participation in the trial. 
 
Subjects are prohibited from receiving the following therapies during the Screening 
and Treatment Phase (including retreatment for post-complete response relapse) of 
this trial: 
• Antineoplastic systemic chemotherapy 
• Targeted therapy (e.g. MAPK inhibitors), or biological therapy  
• Immunotherapy not specified in this protocol 
• Radiation therapy to multiple lesions or major surgery  
Note: Radiation therapy or minor surgery to a single pre-existing but currently 
growing symptomatic extracranial lesion may be considered on an exceptional 
case-by-case basis after consultation with SYNDAX. The patient must have 


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measurable disease outside the radiated or operated field. Administration of 
palliative radiation therapy to a new lesion will be considered clinical 
progression. 
• Live vaccines within 30 days prior to the first dose of trial treatment and 
while participating in the trial.  Examples of live vaccines include, but are 
not limited to, the following: measles, mumps, rubella, varicella/zoster, 
yellow fever, rabies, BCG, and typhoid vaccine. 
• Systemic glucocorticoids with the exceptions specified in section 4.2.1, 
• Any other HDACi, including valproic acid, 
• DNA methyltransferase inhibitors, 
• Any additional anticancer agents, such as chemotherapy, immunotherapy, 
targeted therapy, biological response modifiers will not be allowed, even if 
utilized as treatment of non-cancer indications, 
• Any other investigational agents.  
• Traditional herbal medicines; these therapies are not fully studied and their 
use may result in unanticipated drug-drug interactions that may cause or 
confound the assessment of toxicities. 
• Sensitive substrates of CYP1A2, CYP2C8, CYP3A with a narrow 
therapeutic window (see APPENDIX A  PROHIBITED MEDICATIONS), 
• Drugs that are known to inhibit or induce P-gp (see APPENDIX A  
PROHIBITED MEDICATIONS). 
 
There are no prohibited therapies during the Post-Treatment and Follow-up Phases. 
5.6 
Rescue Medications and Supportive Care 
Subjects should receive appropriate supportive care measures, as deemed necessary 
by the treating investigator. Suggested supportive care measures for the 
management of AEs with potential immunologic etiology are outlined below. 
Where appropriate, these guidelines include the use of oral or IV treatment with 
corticosteroids as well as additional anti-inflammatory agents if symptoms do not 
improve with administration of corticosteroids. Note that several courses of steroid 
tapering may be necessary as symptoms may worsen when the steroid dose is 
decreased. For each disorder, attempts should be made to rule out other causes, such 
as metastatic disease, bacterial or viral infection, which might require additional 
supportive care. The treatment guidelines will be applied when the investigator 
determines the events to be related to pembrolizumab.  
NOTE: If after the evaluation the event is determined unrelated, the investigator 
does not need to follow the treatment guidance (as outlined below). Refer to Section 
5.3.2 for dose modification guidelines.  
It may be necessary to perform conditional procedures, such as bronchoscopy, 
endoscopy, or skin photography as part of evaluation of the event.  
 


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• Pneumonitis:  
Diagnosis of pneumonitis may require consultation with pulmonology and should 
not be solely relying on radiographic imaging. If possible, bronchoscopy with 
bronchoalveolar lavage and/or lung biopsy should be performed. 
o For Grade 2 events, treat with systemic corticosteroids (prednisone 1-
2mg/kg daily, or its equivalent). When symptoms improve to Grade 1 or 
less, steroid taper should be started and continued over no less than 4 weeks. 
o For Grade 3-4 events, immediately treat with intravenous steroids.  
Administer additional anti-inflammatory measures, as needed. 
o Add prophylactic antibiotics for opportunistic infections in the case of 
prolonged steroid administration. 
• Diarrhea/Colitis:  
Subjects should be carefully monitored for signs and symptoms of enterocolitis 
(such as diarrhea, abdominal pain, blood or mucus in stool, with or without fever) 
and of bowel perforation (such as peritoneal signs and ileus). Given that entinostat 
may also present with diarrhea, consultation with gastroenterology for possible 
colonoscopy should be considered. An attribution to specific drug (entinostat or 
pembrolizumab) 
should 
be 
sought 
versus 
a 
combination 
of 
interventions/maneuvers (e.g. entinostat hold and/or rechallenge). 
o All subjects who experience diarrhea/colitis should be advised to drink 
liberal quantities of clear fluids.  If sufficient oral fluid intake is not feasible, 
fluid and electrolytes should be substituted via IV infusion.  For Grade 2 or 
higher diarrhea, consider GI consultation and endoscopy to confirm or rule 
out colitis. 
o For Grade 2 diarrhea/colitis attributed to pembrolizumab, administer oral 
corticosteroids (prednisone 1-2mg/kg daily, or its equivalent).  
o For Grade 3 or 4 diarrhea/colitis attributed to pembrolizumab, treat with 
intravenous steroids followed by high dose oral steroids.   
o When symptoms attributed to pembrolizumab improve to Grade 1 or less, 
steroid taper should be started and continued over no less than 4 weeks. 
o For diarrhea associated with entinostat, please consult section 5.3.1. 
• Type 1 diabetes mellitus (T1DM, if new onset, including diabetic ketoacidosis) 
or ≥ Grade 3 hyperglycemia in patients with pre-existing diabetes, if associated 
with ketosis (ketonuria) or metabolic acidosis 
o For T1DM or Grade 3-4 hyperglycemia 


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 Insulin replacement therapy is recommended for T1DM and for 
Grade 3-4 hyperglycemia associated with metabolic acidosis or 
ketonuria.  
 Evaluate patients with serum glucose and a metabolic panel, urine 
ketones, glycosylated hemoglobin, and C-peptide. 
• Hypophysitis: 
o For Grade 2 events, treat with corticosteroids (prednisone 1-2mg/kg daily, 
or its equivalent). When symptoms improve to Grade 1 or less, steroid taper 
should be started and continued over no less than 4 weeks. Replacement of 
appropriate hormones may be required as the steroid dose is tapered. 
o For Grade 3-4 events, treat with an initial dose of IV corticosteroids 
followed by oral corticosteroids. When symptoms improve to Grade 1 or 
less, steroid taper should be started and continued over no less than 4 weeks. 
Replacement of appropriate hormones may be required as the steroid dose 
is being tapered. 
• Hyperthyroidism or Hypothyroidism:  
Thyroid disorders can occur at any time during treatment. Monitor patients for 
changes in thyroid function (at the start of treatment, periodically during treatment, 
and as indicated based on clinical evaluation) and for clinical signs and symptoms 
of thyroid disorders. 
o Grade 2 hyperthyroidism events (and Grade 2-4 hypothyroidism): 
 In hyperthyroidism, non-selective β-blockers (e.g. propranolol) are 
suggested as initial therapy if subject has a resting heart rate >90 
beats/min. If free T4 is 30% more than institutional ULN, patient is 
required to start on anti-thyroid medication (e.g. propylthiouracil, 
methimazole, tapazole). 
 In hypothyroidism, thyroid hormone replacement therapy, with 
levothyroxine or liothyronine, is indicated per standard of care. 
o Grade 3-4 hyperthyroidism  
 Treat with an initial dose of IV corticosteroids followed by oral 
corticosteroids. When symptoms improve to Grade 1 or less, steroid 
taper should be started and continued over no less than 4 weeks. 
Replacement of appropriate hormones may be required as the 
steroid dose is being tapered. 
 


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• Hepatic: 
Given that both entinostat and pembrolizumab can induce transaminitis, a liver 
biopsy can be considered to exclude autoimmune hepatitis, because management 
can be vastly different. 
o For Grade 1-2 events attributed to pembrolizumab, monitor liver function 
tests more frequently until returned to baseline values (consider weekly). 
 Treat with IV or oral corticosteroids 
o For Grade 3-4 events attributed to pembrolizumab, treat with IV 
corticosteroids for 24 to 48 hours.  
o When symptoms improve to Grade 1 or less, a steroid taper should be 
started and continued over no less than 4 weeks. 
o If transaminitis is attributed to entinostat, please consult section 5.3.1. 
• Renal Failure or Nephritis: 
Although autoimmune interstitial nephritis can occur with pembrolizumab but not 
entinostat, electrolyte abnormalities due to electrolyte wasting can occur with 
entinostat. Please consider urinalysis (with urine culture if WBCs in urine are 
present) and urine electrolytes (Na, Cl, Mg, Phos, Ca2+, CO2) if unexplained low 
serum electrolytes are observed. Consult nephrology if there is suspicion of 
autoimmune interstitial nephritis attributed to pembrolizumab. 
o For Grade 2 events attributed to pembrolizumab, treat with corticosteroids. 
o For Grade 3-4 events attributed to pembrolizumab, treat with systemic 
corticosteroids. 
o When symptoms improve to Grade 1 or less, steroid taper should be started 
and continued over no less than 4 weeks.  
• Steven-Johnson Syndrome (SJS) and Toxic Epidermal Necrolysis (TEN): 
o For signs or symptoms of SJS or TEN, withhold pembrolizumab and 
entinostat and refer the patient for specialized care for assessment and 
treatment. 
o If SJS or TEN is confirmed, permanently discontinue pembrolizumab and 
entinostat. 
• Cardiac Immune-mediated myocarditis: 
Although both entinostat and pembrolizumab can affect heart function, AEs can be 
different (e.g. entinostat can cause cardiac arrhythmias whereas pembrolizumab 
can cause myocarditis). Please consult cardiology if any suspicion for cardiac 


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abnormalities and hold both drugs until cardiology clearance is complete. For 
suspected immune-mediated myocarditis, ensure adequate evaluation to exclude 
other etiologies, and administer corticosteroids as appropriate. 
5.6.1 Management of Infusion Reactions for Pembrolizumab  
Signs and symptoms usually develop during or shortly after drug infusion and 
generally resolve completely within 24 hours of completion of infusion. The table 
below outlines guidelines for subjects who experience an infusion-related reaction 
to pembrolizumab administration. 
NCI CTCAE Grade 
Treatment 
Premedication at 
subsequent dosing 
Grade 1 
Mild reaction; infusion 
interruption not indicated; 
intervention not indicated 
Increase monitoring of vital signs as 
medically indicated until the subject is 
deemed medically stable in the opinion of 
the investigator. 
None 
Grade 2 
Requires infusion interruption but 
responds promptly to symptomatic 
treatment (e.g., antihistamines, 
NSAIDS, narcotics, IV fluids); 
prophylactic medications indicated 
for ≤ 24 hrs 
Stop Infusion and monitor symptoms. 
Additional appropriate medical therapy may 
include but is not limited to: 
IV fluids, antihistamines, NSAIDS, 
acetaminophen, narcotics. 
Increase monitoring of vital signs as 
medically indicated until the subject is 
deemed medically stable in the opinion of 
the investigator. 
If symptoms resolve within 1 hour of 
stopping drug infusion, the infusion may be 
restarted at 50% of the original infusion rate 
(e.g. from 100 mL/hr to 50 mL/hr).  
Otherwise dosing will be held until 
symptoms resolve and the participant 
should be premedicated for the next 
scheduled dose. 
Participants who develop Grade 2 
toxicity despite adequate premedication 
should be permanently discontinued 
from further study drug treatment. 
Subject may be pre-
medicated 1.5h (± 30 
minutes) prior to 
infusion of 
pembrolizumab with: 
 
Diphenhydramine 50 
mg po (or equivalent 
dose of antihistamine). 
 
Acetaminophen 500-
1000 mg po (or 
equivalent dose of 
antipyretic). 


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NCI CTCAE Grade 
Treatment 
Premedication at 
subsequent dosing 
Grades 3 or 4 
Grade 3: 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: Life-threatening; pressor 
or ventilatory support indicated 
Stop Infusion. 
Additional appropriate medical therapy may 
include but is not limited to: 
IV fluids, antihistamines, NSAIDS, 
acetaminophen, narcotics, oxygen, pressors, 
corticosteroids, epinephrine**. 
Increase monitoring of vital signs as 
medically indicated until the subject is 
deemed medically stable in the opinion of 
the investigator. 
Hospitalization may be indicated. 
**In cases of anaphylaxis, epinephrine 
should be used immediately. 
Subject is permanently discontinued 
from further trial treatment 
administration. 
No subsequent dosing 
Appropriate resuscitation equipment should be available at the bedside and a physician readily available 
during the period of drug administration. For further information, please refer to the Common Terminology 
Criteria for Adverse Events v5.0 (CTCAE) at http://ctep.cancer.gov. 
5.6.2 Contraception 
Acceptable Contraceptive Methods   
Failure rate of >1% per year when used consistently and correctly.  
● 
Male or female condom with or without spermicide 
● 
Cervical cap, diaphragm or sponge with spermicide 
Highly Effective Contraceptive Methods That Are User Dependent a  
Failure rate of <1% per year when used consistently and correctly. 
● 
Combined (estrogen- and progesterone-containing) hormonal contraception b 
○ 
Oral  
○ 
Intravaginal  
○ 
Transdermal  
○ 
Injectable 
● 
Progestogen-only hormonal contraception b 
○ 
Oral  
○ 
Injectable  
Highly Effective Methods That Have Low User Dependency  
Failure rate of <1% per year when used consistently and correctly. 
● 
Progesterone-only contraceptive implant b, c  
● 
Intrauterine hormone-releasing system (IUS) b  
● 
Intrauterine device (IUD) 
● 
Bilateral tubal occlusion 
● 
Vasectomized partner  
A vasectomized partner is a highly effective contraception method provided that the partner is the sole 
male sexual partner of the WOCBP and the absence of sperm has been confirmed. If not, an additional 
highly effective method of contraception should be used.  
● 
Sexual abstinence  
Sexual abstinence is considered a highly effective method only if defined as refraining from heterosexual 
intercourse during the entire period of risk associated with the study treatment. The reliability of sexual 
abstinence needs to be evaluated in relation to the duration of the study and the preferred and usual lifestyle 
of the participant.)  


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Notes:  
Use should be consistent with local regulations regarding the use of contraceptive methods for participants 
of clinical studies. 
a) Typical use failure rates are lower than perfect-use failure rates (i.e., when used consistently and 
correctly).  
b) If hormonal contraception efficacy is potentially decreased due to interaction with study treatment, 
condoms must be used in addition to the hormonal contraception during the treatment period and for at 
least 120 days, (corresponding to time needed to eliminate study treatment plus 30 days for study 
treatments with genotoxic potential) after the last dose of entinostat. 
c) If locally required, in accordance with Clinical Trial Facilitation Group (CTFG) guidelines, acceptable 
hormonal contraceptives are limited to those which inhibit ovulation.  
 
5.6.3 Use in Pregnancy 
If a subject inadvertently becomes pregnant while on treatment with 
pembrolizumab/entinostat, the subject will immediately be removed from the 
study.  The site will contact the subject at least monthly and document the subject’s 
status until the pregnancy has been completed or terminated.  The outcome of the 
pregnancy will be reported to SYNDAX without delay and within 24 hours to the 
Sponsor and within 2 working days to SYNDAX, if the outcome is a serious 
adverse experience (e.g., death, abortion, congenital anomaly, or other disabling or 
life-threatening complication to the mother or newborn).  
  
The study investigator will make every effort to obtain permission to follow the 
outcome of the pregnancy and report the condition of the fetus or newborn to the 
Sponsor. If a male subject impregnates his female partner, the study personnel at 
the site must be informed immediately and the pregnancy reported to SYNDAX, as 
described above and in Section 8.4. 
5.6.4 Overdose of Pembrolizumab 
For purposes of this study, an overdose of pembrolizumab will be defined as any 
dose of 1,000 mg or greater (≥5 times the indicated dose).  No specific information 
is available on the treatment of overdose of pembrolizumab. In the event of 
overdose, the participant should be observed closely for signs of toxicity.  
Appropriate supportive treatment should be provided, if clinically indicated. 
 
If an AE(s) is associated with (“results from”) the overdose of pembrolizumab, the 
AE(s) is reported as a SAE, even if no other seriousness criteria are met.  
 
5.6.5 Overdose of Entinostat 
There is no information on overdose of entinostat.  Any SAE that occurs at any 
dose level of entinostat should be reported to SYNDAX Pharmaceuticals. Refer to 
section 8.4 for additional details on AE reporting requirements. 
 


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5.7 
Other Modalities or Procedures 
5.7.1 On-Entinostat Treatment Biopsy (Day 22 (± 2 days)) 
A mandatory on-entinostat treatment tumor biopsy will be performed in the end of 
cycle 1, beginning of cycle 2 (day 22±2 days), immediately before concurrent 
entinostat-pembrolizumab treatment. Depending on the location of the tumor, the 
biopsy can be a core, excisional, or punch and performed as an outpatient procedure 
at the bedside (if palpable; e.g. subcutaneous, cutaneous, or palpable lymph node 
or CT-guided by interventional radiology). The tumor must be accessible for biopsy 
in a location considered safe for biopsy. Lung metastases should not be biopsied 
due to risk of pneumothorax. 
5.7.2 Peripheral Blood for Translational Endpoints 
Approximately 50 mL of peripheral blood will be collected for the purpose of 
analyzing the translational endpoint outlined in section 6.5.1.  
5.8 
Duration of Therapy 
In the absence of treatment delays due to AE, treatment may continue for up to 27 
weeks or until: 
• Disease progression, 
• Inter-current illness that prevents further administration of treatment, 
• Unacceptable AE(s), 
• Pregnancy, 
• Patient decides to withdraw from study treatment, OR 
• General or specific changes in the patient’s condition render the patient 
unacceptable for further treatment in the judgment of the investigator, 
• Subject completes maximum number of treatment cycles (i.e., 9 cycles / 27 
weeks of treatment) allowed per protocol, 
• Subject is lost to follow-up. 
 
Subjects receiving clinical benefit after the study assessment period (6 months) may 
continue pembrolizumab off-study at the investigator’s discretion, as per standard 
of care guidelines. 
 
5.9 
Duration of Follow Up 
Patients who have not progressed on pembrolizumab plus entinostat after 27 weeks 
of treatment will be followed every 3 months (± 2 weeks) for up to 2 years following 
treatment initiation (C1D1). Follow ups include phone calls or visits. The following 
information will be recorded: (a) details regarding systemic treatment for their 
melanoma that they are currently on (type, response if any, duration on treatment), 
(b) whether they have progressed (and when) from current PD-1 inhibitor 
treatment, (c) whether treatment-related AEs that patients may have developed 
during the 27 weeks of entinostat-pembrolizumab treatment have resolved, and 
when, (d) whether they are alive or not; if dead, precise date of death.  


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Patients who have progressed on pembrolizumab plus entinostat within the 27 
weeks of the study duration will be followed every 3 months (± 2 weeks) for up to 
2 years following treatment initiation. Follow up may include phone calls or visits. 
The following information will be recorded: (a) systemic treatment for their 
melanoma they are currently on (type, response if any, duration on systemic 
anticancer treatments, systemic anticancer treatment-related grade 2 or higher 
AEs), (b) whether treatment-related AEs that patients may have developed during 
the year of entinostat-pembrolizumab treatment have resolved, and when, (c) 
whether they are alive or not; if dead, precise date of death.  
 
Patients removed from study treatment for unacceptable AEs will be followed for 
resolution or stabilization of the adverse event(s). All patients (including those 
withdrawn for AEs) should be followed after removal from study treatment as 
stipulated in the protocol.   
5.10 
Study Withdrawal 
If a patient decides to withdraw from the study treatment (and not just from protocol 
therapy), an effort should be made to complete and report study assessments as 
thoroughly as possible.  At the time of withdrawal, the investigator should attempt 
to establish as completely as possible the reason for the study treatment withdrawal.  
1. 
The patient should be asked if they are willing to allow for the abstraction 
of relevant information from their medical record to meet the long-term 
follow-up (e.g., survival endpoints) objectives outlined in the protocol.   
2. 
A complete final evaluation at the time of the patient’s study treatment 
withdrawal should be obtained with an explanation of why the patient is 
withdrawing from the study.   
3. 
If the patient is noncompliant and does not return for an end of study follow-
up assessment, this should be documented in the eCRF as lost to follow-up.  
4. 
If the reason for removal of a patient from the study is an AE, the principal 
specific event will be recorded on the eCRF per reporting guidelines.  
 
Note: Inappropriately high number of patient withdrawals from protocol therapy or 
from the study (i.e. >4 patients) can render the study un-interpretable; therefore, 
unnecessary withdrawal of patients should be avoided. 
 
 
 


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6.0 
DRUG INFORMATION 
6.1 
Entinostat - Investigational Agent  
6.1.1 Description 
Entinostat is a synthetic orally available class I selective HDACi that belongs to the 
class of substituted pyridylcarbamates with a molecular weight of 376.4. Entinostat 
belongs to the pharmacologic class of antineoplastic agents. 
6.1.2 Supplier/How Supplied/Storage/Handling 
Investigational supply of entinostat will be provided at no cost to the patient by 
Syndax Pharmaceuticals. 
Entinostat is an oral drug supplied by SYNDAX as pink to light red (1 mg) or 
yellow (5 mg) polymorph B coated tablets. Each tablet contains mannitol, sodium 
starch glycolate, hydroxypropyl cellulose, potassium bicarbonate, and magnesium 
stearate as inert fillers. The film coating consists of hypromellose, talc, titanium 
dioxide, and ferric oxide pigments (red and yellow) as colorants. Entinostat is to be 
stored at controlled room temperature (15ºC to 25ºC) in a secure, locked storage 
area to which access is limited.  Entinostat is to be protected from light and not to 
be exposed to extremes of temperature (greater than 30ºC or less than 5ºC). The 
pharmacist should dispense the investigational material to the patient at appropriate 
intervals throughout the study in childproof containers. 
6.1.3 Dosage and Administration 
Entinostat is to be taken on an empty stomach, at least 2 hours after a meal and at 
least 1 hour before the next meal.  If entinostat is vomited, dosing should not be re-
administered but instead the dose should be skipped. 
 
For weekly (or less frequent) dosing: If an entinostat dose is missed, it may be taken 
up to 48 hours after the scheduled dosing time. If it is not taken within the 48-hour 
window, the dose should not be taken and should be counted as a missed dose. The 
patient should take the next scheduled dose per protocol.   
6.1.4 Entinostat Compliance 
Treatment compliance for entinostat dosing will be assessed at the end of each 
cycle, defined as 3 weeks (21 days). The first cycle on study will consist only of 
entinostat (mono-therapy). All 21-day cycles of treatment on study thereafter will 
include entinostat in combination with pembrolizumab. Patients will complete the 
IRB-approved dosing diary for documenting date and time of each weekly 
entinostat dose. Subjects will be instructed to bring all study IP (including partially 
used bottles) to each visit, including their subject dosing diary. The dates of any 
missed doses will also be recorded in the subject dosing diary. Site staff will 
perform accountability of the returned drug. Patient compliance will be assessed. 


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Site staff must ensure that the patient clearly understands the directions for self-
administration of study drug and complies with the schedule provided. 
6.1.5 Drug Accountability 
The investigator, or designee, is responsible for keeping accurate records of the 
clinical supplies received from the company sponsor or designee. The total amount 
of drug dispensed to subjects and amount remaining at trial conclusion the will be 
documented. The amount dispensed, amount returned, date dispensed, date 
returned, and lot number is to be recorded for each patient as applicable. The 
amount of drug dosed and start date for each dose will be recorded. An accurate 
and current accounting of the dispensing and return of entinostat for each subject 
will be maintained on an ongoing basis by a member of the study site staff.  The 
amount of study drug dispensed and returned for individual subjects will be 
recorded in the Investigational Drug Accountability Record.   
6.1.6 Return and Retention of Study Drug 
Upon completion or termination of the study, all unused and/or partially used 
investigational product will be destroyed at the site per institutional policy (e.g., 
UNC IDS drug destruction policy). It is the Investigator’s responsibility to arrange 
for disposal of all empty containers, provided that procedures for proper disposal 
have been established according to applicable federal, state, local and institutional 
guidelines and procedures, and provided that appropriate records of disposal are 
kept. 
6.1.7 AEs Associated with Investigational Drug 
As of July 1, 2016, entinostat has been given to >1,055 cancer patients in 31 clinical 
research studies in the mono-therapy setting and combination setting with another 
agent(s).   
 
The following side effects occurred in more than 20% of patients with cancer, 
receiving entinostat alone. These events may or may not be related to entinostat.   
• Fatigue (tiredness), 
• Nausea, 
• Thrombocytopenia, 
• Anemia,  
• Vomiting, 
• Neutropenia, 
• Diarrhea, 
• Leukopenia, 
• Feeling not hungry (decreased appetite), 
• Hypophosphatemia, 
• Hyponatremia, 
• Hypoalbuminemia,  
• Headache. 
 


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The following side effects were seen in ≥10% to 20% of patients, with cancer, 
treated with entinostat alone. These events may or may not be related to entinostat.   
• Shortness of breath or difficulty breathing, 
• Constipation, 
• Hyperglycemia, 
• Hypocalcemia, 
• Hypokalemia, 
• Swelling in the legs, feet and/or ankles, 
• Increase in serum alkaline phosphatase, 
• Leukopenia, 
• Abdominal pain, 
• Fever, 
• Cough, 
• Muscle pain, 
• Salty, metallic taste,  
• Indigestion, 
• Dehydration. 
 
The following SAE occurred in ≥ 5% of patients with cancer, receiving entinostat 
alone. These events may or may not be related to entinostat.  
• Fatigue, 
• Shortness of breath or difficulty breathing, 
• Decreased appetite, 
• Neutropenia, 
• Dehydration, 
• Anemia, 
• Fever, 
• Nausea, 
• Abdominal pain, 
• Diarrhea, 
• Infection, 
• Thrombocytopenia. 
 
Overall, the side effects of entinostat when given in combination with other anti-
cancer agents were similar to that seen when entinostat was given alone.  
 
6.2 
Pembrolizumab (Keytruda™) - Commercial Drug  
6.2.1 Description  
Pembrolizumab is a humanized monoclonal antibody that blocks the interaction 
between PD-1 and its ligands, PD-L1 and PD-L2. Pembrolizumab is an IgG4κ 
immunoglobulin with an approximate molecular weight of 140 kD. Full prescribing 
information for pembrolizumab is available at: 


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http://www.merck.com/product/usa/pi_circulars/k/keytruda/keytruda_pi.pdf 
6.2.2 Dosage and Administration: 
Pembrolizumab (200 mg) will be administered IV over 30 min (range of infusion 
time, 25-40 min) every 21 days (i.e., three weeks) in combination entinostat. 
Pembrolizumab should be given on D1 of each cycle, except for cycle 1, after 
entinostat dosing occurs assuming patient is not on a hold for entinostat.   
6.2.3 Storage and Stability: 
Solution for infusion vials should be stored at refrigerated conditions (2–8 °C) and 
protected from light. 
 
Clinical supplies must be stored in a secure, limited-access location under the 
storage conditions specified on the label.  Receipt and dispensing of trial medication 
must be recorded by an authorized person at the trial site. 
 
Handling and Disposal: Local requirements for disposal of hazardous drugs 
should be followed at each participating clinical site. 
 
Please see UNC policy on hazardous drugs:  
http://intranet.unchealthcare.org/intranet/hospitaldepartments/safetynet/policies/ha
zardousdrugs.pdf 
6.2.4 Adverse Events Associated with Commercial Drug 
The most common AEs (reported in ≥20% of patients in clinical trials of 
pembrolizumab) included fatigue, pruritus, diarrhea, decreased appetite, rash, 
pyrexia, cough, dyspnea, musculoskeletal pain, constipation, and nausea. The 
following warnings are associated with the use of pembrolizumab:  
 
 
Immune-Mediated Pneumonitis 
Pneumonitis occurred in ~3% of melanoma patients treated in clinical trials of 
pembrolizumab. The median time-to-development of pneumonitis was 5 months 
with a median duration of 4.9 months.  The one patient with Grade 3 pneumonitis 
required initial treatment with high-dose systemic corticosteroids (greater than or 
equal to 40 mg prednisone or equivalent per day) followed by a corticosteroid taper. 
Pneumonitis completely resolved in seven of the nine patients with Grade 2-3 
pneumonitis. 
 
Immune-Mediated Colitis 
Colitis occurred in 1% of melanoma patients treated in clinical trials of 
pembrolizumab. The median time-to-onset was 6.5 months with a median duration 
of 2.6 months. All three patients with Grade 2 or 3 colitis were treated with high-
dose corticosteroids (greater than or equal to 40 mg prednisone or equivalent per 
day). 
 
 


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Immune-Mediated Hepatitis  
Autoimmune hepatitis occurred in 0.5% of melanoma patients treated in clinical 
trials of pembrolizumab. The time-to-onset was 22 days for the case of Grade 4 
hepatitis, which lasted 1.1 months. The patient with Grade 4 hepatitis permanently 
discontinued pembrolizumab and was treated with high-dose (≥ 40 mg prednisone 
or equivalent per day) systemic corticosteroids followed by a corticosteroid taper. 
Both patients with hepatitis experienced complete resolution of the event.  
 
Immune-Mediated Hypophysitis  
Hypophysitis occurred in 0.5% of melanoma patients treated in clinical trials of 
pembrolizumab. The time-to-onset was 1.7 months for the patient with Grade 4 
hypophysitis and 1.3 months for the patient with Grade 2 hypophysitis. Both 
patients were treated with high-dose (greater than or equal to 40 mg prednisone or 
equivalent per day) corticosteroids followed by a corticosteroid taper and remained 
on a physiologic replacement dose.  
 
Renal Failure and Immune-Mediated Nephritis  
Nephritis occurred in 3 (0.7%) patients of melanoma patients treated in clinical 
trials of pembrolizumab, consisting of one case of Grade 2 autoimmune nephritis 
(0.2%) and two cases of interstitial nephritis with renal failure (0.5%), one Grade 3 
and one Grade 4. The time-to-onset of autoimmune nephritis was 11.6 months after 
the first dose of pembrolizumab (5 months after the last dose) and lasted 3.2 
months; this patient did not have a biopsy. Acute interstitial nephritis was 
confirmed by renal biopsy in two patients with Grades 3-4 renal failure. All three 
patients fully recovered their renal function with treatment with high-dose 
corticosteroids (≥40 mg prednisone or equivalent per day) followed by a 
corticosteroid taper. 
 
Immune-Mediated Hyperthyroidism  
Hyperthyroidism occurred in 5 (1.2%) of 411 melanoma patients treated in clinical 
trials of pembrolizumab. The median time-to-onset was 1.5 months and the median 
duration was 2.8 months (range 0.9 to 6.1 months). One of two patients with Grade 
2 and the one patient with Grade 3 hyperthyroidism required initial treatment with 
high-dose corticosteroids (greater than or equal to 40 mg prednisone or equivalent 
per day) followed by a corticosteroid taper. One patient required permanent 
discontinuation of pembrolizumab due to hyperthyroidism. All five patients with 
hyperthyroidism experienced complete resolution of the event. 
 
Immune-Mediated Hypothyroidism  
Hypothyroidism occurred in 34 (8.3%) of 411 melanoma patients treated in clinical 
trials of pembrolizumab. The median time-to-onset of hypothyroidism was 3.5 
months. All but two of the patients with hypothyroidism were treated with long-
term thyroid hormone replacement therapy. The other two patients only required 
short-term thyroid hormone replacement therapy. No patient received 
corticosteroids 
or 
discontinued 
pembrolizumab 
for 
management 
of 
hypothyroidism. Thyroid disorders can occur at any time during treatment.  


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Other Immune-Mediated Adverse Reactions 
Other clinically important immune-mediated adverse reactions can occur. The 
following clinically significant, immune-mediated adverse reactions occurred in 
less than 1% of patients treated with pembrolizumab, including exfoliative 
dermatitis, uveitis, arthritis, myositis, pancreatitis, hemolytic anemia, partial 
seizures arising in a patient with inflammatory foci in brain parenchyma, and 
adrenal insufficiency. 
 
Across clinical studies with pembrolizumab in approximately 2,000 patients, the 
following additional clinically significant, immune-mediated adverse reactions 
were reported in less than 1% of patients: myasthenic syndrome, optic neuritis, and 
rhabdomyolysis. 
 
Embryofetal Toxicity 
Pembrolizumab may cause fetal harm when administered to a pregnant woman. 
Animal models link the PD-1/PD-L1 signaling pathway with maintenance of 
pregnancy through induction of maternal immune tolerance to fetal tissue.  
 
Steven’s Johnson Syndrome (SJS) and Toxic Epidermal Necrolysis (TEN) 
The risk of SJS and TEN is reported at approximately 0.4–7 cases per million 
patient years in the general adult population. Independent risk factors include 
certain medications, such as anticonvulsants, sulfonamides, aminopenicillins, 
allopurinol, and NSAIDs. Non-medication triggers include infection, contrast 
media, and vaccinations. Malignancy is associated with an increased mortality rate 
in patients with SJS and TEN. 
 
Myocarditis 
A total of 6 cases of myocarditis have been reported in patients treated with 
pembrolizumab in clinical trials in an expanded access program. There was one 
fatal case reported in a clinical trial.  Immune-mediated myocarditis should be 
suspected if other causes of myocarditis, such as infection or prior radiation 
therapy, have been excluded. Risk factors include certain medications and 
treatment modalities, such as radiation, anthracycline, alkylating agents and, most 
recently, checkpoint inhibitors. 
 
6.3 
Clinical Assessments 
Clinical assessments will be performed as outlined in the Time and Events Table in 
Section 7.1. 
6.3.1 Concomitant Medications 
All concomitant medication and concurrent therapies will be documented at 
Baseline/Screening and throughout the study, as summarized in the Time and 
Events Table in Section 7.1. Dose, route, unit frequency of administration, and 
indication for administration and dates of medication will be captured. 


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6.3.2 Demographics  
Demographic information (date of birth, gender, race) will be recorded at 
Screening. 
6.3.3 Medical History  
Relevant medical history, including history of current disease, patient 
demographics, other pertinent history and information regarding underlying 
diseases will be recorded at screening and a focused medical history on 
symptoms/toxicity will be performed thereafter.  
6.3.4 Physical Examination 
For screening: A complete physical examination, including height (at screening 
only), weight, examination of skin, ECOG performance status and vital signs (i.e., 
weight, temperature, blood pressure, heart rate, respiratory rate, oxygen saturation) 
will be performed at screening and the first study visit #.  Qualified staff (MD, NP, 
RN, and PA) may complete the abbreviated physical exam (i.e., focused on 
symptoms/toxicity as noted below) at all other visits.   
 
After screening starting on D1/Cycle 1/Wk1:  physical exam should be focused on 
symptoms/toxicity including weight, examination of skin, ECOG performance 
status and vital signs thereafter.  New abnormal physical exam findings must be 
documented and will be followed by a physician or other qualified staff at the next 
scheduled visit. 
6.3.5 Adverse Events 
Events should be assessed per NCI-CTCAE criteria v5.0. Information regarding 
occurrence of AEs will be captured throughout the study. Duration (start and stop 
dates), severity/grade, outcome, treatment and relation to study drug will be 
recorded in the CRF.  
6.3.6 Disease Assessment 
This following test should be performed for assessment of melanoma disease. 
6.3.6.1 Computed tomography (CT) of the whole body i.e., neck (if applicable), 
chest, abdomen, pelvis with IV contrast (or abdominal/pelvis MRI if allergy to IV 
contrast)  
Baseline (screening) disease assessment should be obtained within 21 days of 
initiating study treatment. During the study, scans will be repeated as outlined in 
the Time and Events table in section 7.1 (i.e., at the end of week 10[D1 of cycle 4 
or Study day 64/Wk10], end of week 15 [D1 of cycle 6 i.e., Study day 106/Wk16], 
end of week 21 [D1 of cycle 8 or Study day 148/Wk22], and end of week 27).  
Scans obtained during study treatment may be performed within ±7 days of the 
scheduled study visit. 


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UNIVERSITY OF NORTH CAROLINA 
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6.3.6.2 Magnetic Resonance Imaging (MRI) of the brain  
MRI of the brain must be obtained at baseline (screening) and at end of treatment 
(i.e., at time of disease progression or at week 27); on any other visit only if 
clinically indicated, as per standard of care. If brain MRI is not feasible, patient 
can have a CT scan with IV contrast. 
6.3.7 Pre-Screening Pathology Assessment Period (pre-screening only) 
Given the limited 21-day screening period, and the requirement for absent TIL/TAL 
in melanoma tissues, archival metastatic tissue (locoregional, distant metastatic) 
can be requested and assessed before study enrollment (pre-screening).  There is 
not an official pre-screening period given that Dr. Googe, study’s pathologist, is 
UNC’s sole melanoma pathologist and routinely signs off his pathology reports, 
either from outside consultation slides or from in-house cases, along with a report 
about the presence of absence of tumor-infiltrating/-associated lymphocytes. If not 
available already, representative tumor sections from archived tumor blocks will be 
stained with H&E and will be assessed by Dr. Paul Googe, study pathologist, for 
assessment of quality using these criteria:  (1) the tumor surface area must be at 
least 1cm2 (which can be from a combination of metastatic blocks if necessary), (2) 
no more than 20% of necrosis, (3) the ratio of viable tumor cells to tumor-associated 
stroma should be at least 60/40, and (4) if TILs are present based on H&E stained 
sections, they must be ≤ 1% of the total number of cells in the specimen; the amount 
of tissue should be ≥1 cm2.  
If archival tissue is unavailable or insufficient, a fresh biopsy should be performed 
to confirm unresectable stage III or distant metastatic disease. If the patient 
qualifies, then they can sign the informed consent. 
6.3.8 Electrocardiogram (ECG) 
A 12-lead ECG will be performed at screening and on cycles 2, 4, 6, 8, as outlined 
in the Time and Events table. 
6.3.9 Patient Pill Diary (Entinostat) 
A patient pill diary will be checked at the beginning of each cycle and should be 
kept by subject for self-documentation of entinostat dosing (as outlined in the Time 
and Events table). A single 5 mg tablet is to be taken on day 1, day 8, and day 15 
of each cycle within the specified 21-day cycle windows. See Section 6.0 for 
information on entinostat drug supply, storage, handling and compliance. 
 
6.4 
Clinical Laboratory Assessments 
6.4.1 Hematology 
Blood will be obtained and sent to the clinical site hematology lab for assessment 
of a complete blood count with differential (i.e., hemoglobin, hematocrit, red blood 
cell count, white blood cell count, white blood cell differential, and platelet count), 
as outlined in the time and events table.  


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UNIVERSITY OF NORTH CAROLINA 
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Hematology labs should be performed pre-study and on D1 at every treatment cycle 
and at the end of treatment (i.e., D1C1, D1C2, D1C3, etc.), as outlined in the Time 
and Events Table. 
6.4.2 Blood Chemistry Profile 
Blood will be obtained and sent to the clinical site chemistry lab for determination 
of the following: sodium (Na) potassium (K), chloride (Cl), creatinine, blood urea 
nitrogen (BUN), aspartate aminotransferase (AST), alanine aminotransferase 
(ALT), total bilirubin, alkaline phosphatase, glucose, total protein, albumin, 
calcium, magnesium, phosphorus, lactate dehydrogenase (LDH), amylase, and 
lipase.  
 
All listed serum chemistries will be performed during screening, D1 of each 
treatment cycle, and at the end of treatment as outlined in the Time and Events 
table. 
6.4.3 Urinalysis  
A standard urinalysis test (Dipstick test) will be obtained at screening and at Cycle 
1, and at other times as clinically indicated, as outlined in the Time and Events table 
below. The analysis includes testing for the presence of proteins, glucose, ketones, 
hemoglobin, bilirubin, urobilinogen, nitrite, and leucocytes as well as testing of pH 
and specific gravity. 
6.4.4 Endocrine Test  
At the screening visit and then repeated on D1 of cycle 4/week 10 visit (section 
7.4.4) and D1 of cycle 6/week 16 visit (section 7.4.6): The endocrine test at baseline 
should include measurement of thyroid stimulating hormone (TSH), free thyroxine 
4 (fT4).  
 
During study treatment on D1 of cycles 7, 8 and 9 scheduled at weeks 19, 22, 25, 
respectively, and also at the end of study visit: the endocrine test will only need to 
include TSH and fT4. 
6.4.5 Pregnancy Test 
A pregnancy test (β-hCG) will be obtained from female subjects who are of 
childbearing age prior to their participation in the study.  A negative urine 
pregnancy test must be performed within 3 days prior to receiving the first dose of 
study drug i.e., within 72 hours of administration of the first dose of entinostat. 
 


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UNIVERSITY OF NORTH CAROLINA 
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6.5 
Correlative Studies  
6.5.1 Collection of Peripheral Blood Mononuclear Cells (PBMC) 
Peripheral blood (approximately 50 mL at each time point) will be collected for 
isolation of PBMCs using the Ficoll isolation technique for multiparameter flow 
cytometry (immunoregulatory subsets and assessment of H3 and H4 acetylation 
levels).  
 
Samples will be collected at  baseline; in the end of cycle 1, beginning of cycle 2 
(day 22±2 days), immediately before concurrent entinostat-pembrolizumab; week 
10; and end of treatment (i.e., at time of withdrawal for progression or other 
reasons) D105/wk16 visits, as outlined in the Time and Events table in Section 7.1. 
6.5.2 Mandatory On-Treatment Biopsy   
A mandatory (excisional, punch or CT-guided) biopsy will be obtained in the end 
of cycle 1, beginning of cycle 2 (day 22±2 days), immediately before concurrent 
entinostat-pembrolizumab treatment. 
 
 
6.5.3  Targeted panel somatic mutation sequencing 
If this has not been previously done as part of patient’s standard of care, archived 
tumor tissue will be submitted to Foundation Medicine (foundationmedicine.com) 
for FoundationOne testing. FoundationOne assesses somatic mutations across 
300 cancer-associated genes, computes tumor mutation burden, assesses expression 
of PD-L1, and calculates tumor infiltrating lymphocyte density, as per standard of 
care.  If patient’s tumor has been previously submitted for targeted panel 
sequencing using other commercially available tests these tests would be acceptable 
on condition that sequencing has been performed on  >100 genes and that  all 
exomes  of these genes have been sequenced (i.e.  not just hotspot mutations).   
Results from other non-FoundationOne targeted sequencing panels will be at the 
discretion of the principal Investigator.  Results are not required to be available at 
the time of initiation of study treatment.


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UNIVERSITY OF NORTH CAROLINA 
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7.0 
EVALUATIONS AND ASSESSMENTS 
7.1 
Time and Events Table 
(Note: Week 1 includes days 1-7; week 2 includes days 8-14; week 3 includes days 15-21, etc.)
Assessments1 
 
Pre-
screening 
Screening1 
D-21→-0 
Study Treatment3 
End 
of 
Tx4 
±7d 
Long-
term 
follow 
up4 
D1 
Wk1 
D22 
Wk4 
D43 
Wk7 
D64 
Wk10 
D85 
Wk13 
D106 
Wk16 
D127, D148, 
D169 
Wks19, 22, 25 
D1 
Cycle 1 
D1 
Cycle 2 
D1 
Cycle 3 
D1 
Cycle 4 
D1 
Cycle 5  
D1 
Cycle 6 
D1 
Cycles 7, 8, 9 
Informed 
Consent2 
X 
X 
 
 
 
 
 
 
 
 
 
History, PE 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Serum/Urine 
Pregnancy Test 
 
X 
X 
 
 
 
 
 
 
 
 
ECOG PS 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Concomitant 
Meds 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Hematology 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Serum Chemistry 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Endocrine tests 
 
X 
 
 
 
X 
 
X 
X5 
X 
 
Coagulation tests5 
 
X 
 
 
 
 
 
 
 
 
 
Urinalysis6 
 
X 
X 
 
 
 
 
 
 
 
 
12-lead ECG 
 
X 
 
X 
 
X 
 
X 
X 
X 
 
Toxicity 
Assessment 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
Brain MRI 
 
X 
 
 
 
 
 
 
 
X 
 
Whole body CT 
 
X 
 
 
 
X 
 
X 
X8 
X 
 
Patient pill diary 
(entinostat) 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
Entinostat7 
 
 
X 
X 
X 
X 
X 
X 
X8 
 
 
Pembrolizumab7 
 
 
 
X 
X 
X 
X 
X 
X8 
X9 
 
Blood for 
isolation of 
PBMCs 
 
 
X 
X 
 
X 
 
 
 
X 
 
Archival tissue 
assessment10 
X 
 
 
 
 
 
 
 
 
 
 
Fresh Tumor 
Biopsy 
 
 
 
X11 
 
 
 
 
 
 
 
FoundationOne 
 
X 
 
 
 
 
 
 
 
 
 


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Key to Footnotes  
 
1. All screening assessments should be obtained within 21 days before initiating study treatment, unless specified below: 
a. 
Screening labs obtained within 72 hours of D1 of study treatment do not need to be repeated on D1/cycle 1 visit. 
b. The serum pregnancy test should be obtained during screening and a urine pregnancy test should be obtained within 72 
hours before starting entinostat treatment on D1/cycle 1. 
2. Informed consent at pre-screening visit in order to access archival tissue or perform fresh tissue biopsy if archival tissue 
is unavailable or insufficient, to confirm unresectable stage III or distant metastatic disease. Main consent is to be signed 
at the screening visit. 
3. Study Treatment assessments: 
a. 
All study treatment visits should be performed within (± 1) day for the first two weeks of study therapy.  
b. For the cycle 1/day 2 visit (i.e., Study Day 22) visit the window is ± 2 days, 
c. 
Then ± 3 days for assessments scheduled on D1 of cycles 3 and 4 and then ± 7 days for assessments scheduled on D1 
of all subsequent cycles. 
4. a. Refer to section 7.5.1 for guidance on end of study treatment requirements for this study.  
b. Refer to section 7.5.2 for guidance on long-term follow up requirements for this study. 
5. Perform coagulation tests at screening, as described in section 4.1.9.  Perform at other visits if necessary per discretion 
of the investigator. 
6. Urinalysis at screening, at Cycle 1, and other times as clinically indicated. 
7. Entinostat and pembrolizumab should be dosed as outlined in Sections 5.2 and 5.2.1.  
8. Following radiographic disease assessment on week 16, patients will be seen by the investigator every 21 days for AE 
review and initiation of a new treatment cycle (i.e., every 3 weeks starting on D1 of weeks 19, 22, and 25 (see section 
7.4.7) and at the end of study treatment visit (see Section 7.5.1) for details of scheduled assessments. Radiographic 
disease assessment will occur at D1 of cycle 8 during this period. 
9. Following completion of the trial treatment (i.e., 9 cycles of protocol-directed therapy) at 28 weeks, pembrolizumab 
dosing may be continued per investigator discretion on condition that there is no disease progression and patient 
continues to receive clinical benefit. Treatment will be administered as per standard of care guidelines off study. 
10. If archival tissue is unavailable or insufficient, fresh biopsy should be performed in order to confirm unresectable stage 
III or distant metastatic disease. 
11. The biopsy should be performed at the end of cycle 1, beginning of cycle 2 (day 22±2 days), immediately before 
concurrent entinostat-pembrolizumab treatment.  
 


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7.2 Pre-Screening Assessments 
 
1. Pre-screening informed consent, 
2. Pathology assessment for eligibility, 
7.3 Screening Assessments 
 
3. Informed consent, 
4. Clinical evaluation:  complete history and complete physical exam, 
5. ECOG performance status, 
6. Concomitant medication review, 
7. Toxicity/safety assessment, 
8. Brain MRI (or CT scan with IV contrast, if applicable), 
9. Whole body CT (chest, abdomen and pelvis; neck if applicable). MRI of the abdomen/pelvis 
with IV contrast can be an acceptable equivalent, if clinically indicated 
10. 12-lead ECG, 
11. Targeted panel sequencing (FoundationOne), if it has not previously performed. Results 
are not required to be available at the time of study drug initiation. If patient’s tumor has 
been previously submitted for targeted panel sequencing using other commercially available 
tests these tests would be acceptable on condition that sequencing has been performed on  
>100 genes and that all exomes of these genes have been sequenced (i.e.  not just hotspot 
mutations).  Results from other non-FoundationOne targeted sequencing panels will be at 
the discretion of the principal Investigator.   
12. Laboratory studies: 
 
Pregnancy Test   
 
Hematology  
 
Serum Chemistries  
 
Urinalysis 
 
Endocrine test (screening visit) 
 
7.4 
Study Treatment Assessments 
7.4.1 Study Day 1 ± 1 day (D1 of Cycle 1/Week 1)  
1. Clinical evaluation: focused history and physical exam, 
2. ECOG performance status, 
3. Laboratory studies: (*do not repeat if pre-study assessments performed within 72 hours of 
this visit)  
 Hematology* 
 
Serum Chemistries* 
 
Urinalysis, including urine pregnancy test (as clinically indicated)* 
 
Collect peripheral blood for isolation of PBMCs (approximately 50 mL) 
4. Concomitant medication review, 
5. Toxicity/Safety Assessment,  
6. Patient pill diary review and patient training on diary instructions. 


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7. Entinostat self-administration with study staff present. **One 5 mg tablet is to be dosed 
weekly; dispense 2 additional 5-mg tablets for D8 and D15 home administration.  
 
7.4.2 Study Day 22 ± 2 days (D1 of Cycle 2/Week 4) 
1. Clinical evaluation: focused history and physical exam,  
2. ECOG performance status, 
3. 12-lead ECG, 
4. Laboratory studies:   
 
Hematology  
 
Serum Chemistries 
 
Collect peripheral blood for isolation of PBMCs (approximately 50 mL) 
5. Mandatory Fresh Tumor biopsy (excisional, punch, or CT-guided), 
6. Concomitant medication review,  
7. Toxicity Assessment,  
8. Patient pill diary check, 
9. Entinostat administration: 5mg PO to be administered after mandatory biopsy; also dispense 
two 5-mg pills for administration on D28 and D35, 
10. Pembrolizumab administration: 200 mg IV dose; to be administered after mandatory biopsy 
and after entinostat intake. 
 
7.4.3 Study Day 43 ± 3 days (D1 of Cycle 3/Week 7) 
1. Clinical evaluation: focused history and physical exam,  
2. ECOG performance status,  
3. Laboratory studies:   
 Hematology  
 Serum Chemistries 
4. Concomitant medication review, 
5. Toxicity Assessment,  
6. Check Patient pill diary, 
7. Entinostat administration in clinic and additional doses taken on cycle day 8 and cycle d15 
8. Pembrolizumab administration: 200 mg IV dose; to be administered after entinostat intake. 
 
7.4.4 Study Day 64 ± 7 days (D1 of Cycle 4/Week 10) 
1. Clinical evaluation:  focused history and physical exam,  
2. ECOG performance status,  
3. 12-lead ECG, 
4. Laboratory studies:   
 Hematology  
 Serum Chemistries  
  Endocrine panel: fT4/TSH 
 Collect peripheral blood for isolation of PBMCs and plasma (approximately 50 mL) 
5. Concomitant medication review, 
6. Toxicity/safety Assessment, 


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7. Disease assessment: whole body CT (chest, abdomen and pelvis; neck if applicable),  
8. Entinostat administration: in clinic and additional doses taken on cycle day 8 and cycle d15, 
9. Pembrolizumab administration: 200 mg IV dose; to be administered after entinostat intake. 
7.4.5 Study Day 85 ± 7 days (D1 of Cycle 5/Week 13) 
1. Clinical evaluation:  focused history and physical exam,  
2. ECOG performance status,  
3. Laboratory studies:   
• Hematology  
• Serum Chemistries 
4. Concomitant medication review, 
5. Toxicity Assessment, 
6. Patient pill diary check, 
7. Entinostat administration: in clinic and additional doses taken on cycle day 8 and cycle d15, 
8. Pembrolizumab administration: 200 mg IV dose; to be administered after entinostat intake. 
 
7.4.6 Study Day 106 ± 7 days (D1 of Cycle 6/Week 16) 
1. Clinical evaluation:  focused history and physical exam,  
2. ECOG performance status, 
3. 12-lead ECG 
4. Laboratory studies:   
o Hematology  
o Serum Chemistries 
o Endocrine panel:  fT4/TSH 
5. Concomitant medication review, 
6. Toxicity Assessment/safety,  
7. Disease assessment (chest, abdomen and pelvis; neck if applicable), 
8. Patient pill diary check, 
9. Entinostat administration: in clinic and additional doses on cycle day 8 and cycle d15, 
10. Pembrolizumab administration: 200 mg IV dose; to be administered after entinostat intake. 
 
7.4.7 Study Days 127, 148, 169 (D1 of Cycles 7, 8 and 9/Wks 19, 22, 25)  (within ± 7 days for 
each visit)* 
*Visits on pembrolizumab dosing infusion day 1 of wks 19, 22 and 25 
1. Clinical evaluation: focused history and physical exam,  
2. ECOG performance status, 
3. 12-lead ECG on cycle 8 
4. Laboratory studies:   
 Hematology 
 Serum Chemistries  
 Endocrine panel: fT4/TSH only 
5. Concomitant medication review, 
6. Toxicity/safety assessment, 


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7. Disease assessment: D1 of Cycle 8 (chest, abdomen and pelvis; neck if applicable), 
8. Patient pill diary check, 
9. Entinostat administration: in clinic and additional doses taken on cycle day 8 and cycle 
d15before pembrolizumab on weeks 19, 22, and 25 
10. Pembrolizumab administration: 200 mg IV dose; to be administered after entinostat intake 
on weeks 19, 22, and 25 
 
7.5 
Post Treatment Follow-Up Assessments 
7.5.1 End of Study Treatment Visit (Week 28, or earlier if disease progression, intolerable 
toxicity, patient withdraw) within + 7 days of stopping study treatment 
1. Clinical evaluation: focused history and physical exam,  
2. ECOG performance status,  
3. 12-lead ECG 
4. Laboratory studies:   
 Hematology  
 Serum Chemistries  
 Endocrine panel: include fT4/TSH only 
5. Disease assessment (chest, abdomen and pelvis; neck if applicable), 
6. Collect peripheral blood for isolation of PBMCs and plasma (approximately 50 mL) 
7. Brain MRI (if applicable), 
8. Concomitant medication review, 
9. Toxicity Assessment.  
7.5.2 Long Term Follow-up Assessments 
Patients who have not progressed on pembrolizumab plus entinostat after 27 weeks of treatment 
will be followed every 3 months (+/- 15 days) for up to 2 years post study treatment with phone 
calls or standard of care visits. The following information will be recorded:  
(a) details regarding systemic treatment for their melanoma that they are currently on (type, 
response if any, duration on treatment, treatment-related side effects),  
(b) whether they have progressed (and when) from current PD-1 inhibitor treatment,  
(c) whether treatment-related AEs that patients may have developed during the year of 
entinostat-pembrolizumab treatment have resolved, and when,  
(d) whether they are alive or not; if dead, precise date of death.  
 
Patients who progressed on pembrolizumab plus entinostat before completing 27 weeks of study 
therapy will be followed every 3 months (+/- 15 days) for up to 2 years since study initiation 
with phone calls or standard of care visits. The following information will be recorded:  
(a) systemic treatment for their melanoma they are currently on (type, response if any, duration 
on treatment, treatment-related side effects),  
(b) whether treatment-related AEs that patients may have developed during the year of 
entinostat-pembrolizumab treatment have resolved, and when,  
(c) whether they are alive or not; if dead, precise date of death. 
 


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Patients removed from study treatment for unacceptable AEs will be followed for resolution or 
stabilization of the adverse event(s). Following resolution of adverse events they will be 
followed every3 months (+/- 15 days) for up to 2 years since study initiation with phone calls 
or standard of care visits.  All patients (including those withdrawn for AEs) should be followed 
after removal from study treatment as stipulated in the protocol.   
 
7.6 
Handling of Bio specimens Collected for Correlative Research 
Bio specimens collected for this study will be stored in the Lineberger Comprehensive Cancer 
Center (LCCC) Tissue Procurement Facility (TPF), or, if needed, in a secure off-site storage 
facility. All bio specimens will be obtained in accordance with procedures outlined in the LCCC 
1729 Study Laboratory Manual and stored in containers with controlled access. Each sample 
will be assigned a unique code number and no identifiable personal health information (PHI) 
will be on the specimen label.  Information about the patient’s disease will be linked to the 
specimens stored in the repository database. TPF-associated research staff, LCCC 
Bioinformatics staff who support the TPF database and the LCCC Data Warehouse, and 
researchers with IRB-approval for access to PHI for each subject in this study will be able to 
link specimens to relevant medical information. Some results from laboratory analyses that 
occurred during the patient’s participation in the clinical study may also be included. This 
information may be important for understanding how the patient’s cancer developed and 
responded to treatment. 
Storage Time: 
1. 
The biospecimen will be used first and foremost for research purposes outlined within the 
confines of this protocol. Samples will be discarded/destroyed after relevant data are 
collected for this study, unless consent was obtained from the patient to use tissue for other 
research purposes (e.g., TPF consent form was signed by the patient).  In this circumstance, 
there is no time limit on how long biospecimens may be stored. 
2. 
The Principal Investigator must agree to abide by policies and procedures of the TPF 
facility and sign a letter of research agreement for ethical and appropriate conduct of their 
research that utilizes specimens obtained from the TPF facility (e.g., use of leftover 
specimens will require a protocol outlining the research plan for biospecimen use).  
 
Compliance Statement 
Biospecimen collection for this study will be conducted in full accordance to all applicable 
University of North Carolina at Chapel Hill (UNC-CH) Research Policies and Procedures and 
all applicable Federal and state laws and regulations including 45 CFR 46, and the Health 
Insurance Portability and Accountability Act (HIPAA) Privacy Rule. Any episode of 
noncompliance will be documented. 
 
The investigators will perform the study in accordance with this protocol, will obtain consent 
and assent (unless a waiver is granted), and will report unexpected problems in accordance with 
the UNC-CH IRB Policies and Procedures and all federal requirements. Collection, recording, 
and reporting of data will be accurate and will ensure the privacy, health, and welfare of research 
subjects during and after the study.  
 


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7.7 
Assessment of Safety  
Any patient who receives at least one dose of entinostat will be evaluable for toxicity.  Each 
patient will be assessed periodically for the development of any toxicity according to the Time 
and Events table.  Toxicity will be assessed according to the NCI CTCAEv5.0.   
7.8 
Assessment of Efficacy 
7.8.1 Assessment of Disease-Tumor Measurement Based on RECIST v1.1 
See the international criteria proposed by the Response Evaluation Criteria in Solid Tumors 
(RECIST) Committee, version 1.1 for additional details and clarifications149,150.  
 
Measurable disease will be defined as the presence of at least one measurable lesion that can be 
accurately measured in at least one dimension with the longest diameter a minimum size of: 
• >10 mm by CT scan (CT scan slice thickness no greater than 5 mm) 
• 10 mm caliper measurement by clinical exam (lesions which cannot be accurately measured 
with calipers should be recorded as non-measurable). 
• 20 mm by chest x-ray. 
 
For malignant-appearing lymph nodes on radiographic imaging that can be considered 
pathologically enlarged and measurable a lymph node must be ≥15 mm in short axis when 
assessed by CT scan with IV contrast performed with slice thickness ≤ 5 mm).  At baseline and 
in follow-up, only the short axis will be measured and followed.   
 
All other lesions, including small lesions (longest diameter <10 mm or pathological lymph 
nodes with ≥10 to <15 mm short axis) as well as truly non-measurable lesions, will be 
considered non-measurable.  Lesions considered truly non-measurable include: leptomeningeal 
disease; ascites; pleural/pericardial effusion; inflammatory breast disease; lymphangitic 
involvement of skin or lung, abdominal masses/abdominal organomegaly identified by physical 
exam that is not measurable by reproducible imaging techniques. 
 
All measurements should be recorded in metric notation, using calipers, if clinically assessed. 
All baseline evaluations should be performed as close as possible to the treatment start and 
never more than 21-days before the beginning of the treatment.  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.  Imaging-based evaluation should always be done rather than 
clinical examination, unless the lesion(s) being followed cannot be imaged but are assessable 
by clinical exam.  Clinical lesions will only be considered measurable when they are superficial 
and ≥10 mm diameter, as assessed using calipers (e.g. skin nodules). For the case of skin lesions, 
documentation by color photography, including a ruler to estimate the size of the lesions is 
recommended. 
7.8.2 Baseline Documentation of Target and Non-Target Lesions 
All measurable lesions up to a maximum of 5 lesions total (and a maximum of two lesions per 
organ) representative of all involved organs should be identified as target lesions and will be 
recorded and measured at baseline.   


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Target lesions should be selected on the basis of their size (lesions with the longer diameter), 
growth rate at the time of study enrollment (stable or regressing tumor lesions should be 
excluded, irrespective of other parameters), be representative of all involved organs, and, in 
addition, should be those that lend themselves to reproducible repeated measurements.  
 
A sum of the diameters (longest for non-nodal lesions, short axis for nodal lesions) for all target 
lesions will be calculated and reported as the baseline sum diameters. If lymph nodes are to be 
included in the sum, only the short axis is added into the sum.  The baseline sum diameters will 
be used as reference to further characterize the objective tumor response of the measurable 
dimension of the disease. 
 
All other lesions (or sites of disease), including pathological lymph nodes, should be identified 
as non-target lesions and should also be recorded at baseline. Measurements are not required 
and these lesions should be followed as “present” or “absent”, or in rare cases “unequivocal 
progression”.   
7.8.3 Evaluation of Target Lesions using RECIST 1.1 Criteria 
NOTE: In addition to the information below, also see section 4.3.2 in the international criteria 
proposed by the Response Evaluation Criteria in Solid Tumors (RECIST) Committee, version 
1.1 150 for special notes on the assessment of target lesions. 
 
Complete response (CR) − Disappearance of all target lesions.  Any pathological lymph node 
(whether target or non-target) must have decreased in short axis to <10 mm. 
 
Partial response (PR) −At least a 30% decrease in the sum of the LD of the target lesions taking 
as reference the baseline sum LD. 
 
Progressive Disease (PD) − At least a 20% increase in the sum of the LD of the target lesions 
taking as reference the smallest sum LD recorded since the treatment started including baseline 
if that is the smallest on study.  In addition to the relative increase of 20%, the sum must also 
demonstrate an absolute increase of at least 5 mm.  The appearance of one or more new lesions 
also constitutes PD. 
 
Stable disease (SD) − Neither sufficient shrinkage to qualify for PR nor sufficient increase to 
qualify for PD taking as references the smallest sum LD since the treatment started. 
7.8.4 Evaluation of Non-Target Lesions using RECIST 1.1 Criteria 
Complete response (CR) − Disappearance of all non-target lesions and normalization of tumor 
marker levels.  All LN must be non-pathological in size (<10 mm short axis).   
 
Non-complete response (non-CR)/non-progression (non-PD) − Persistence of ≥1 non-target 
lesion(s) or/and maintenance of tumor marker level above the normal limits. 
 
Progressive disease (PD) − Appearance of ≥1new lesions and/or unequivocal progression of 
existing non-target lesions. 


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7.8.5 Evaluation of Best Overall Response using RECIST 1.1 Criteria 
Please see review the RECIST v1.1 criteria for complete details150. The best overall response is 
the best response recorded from the start of the study treatment until the end of treatment, 
provided the confirmation criteria are met.  To be assigned a status of PR or CR, changes in 
tumor measurements must be confirmed by repeat studies that should be performed > 4 weeks 
after the criteria for response are first met. If a CR/PR cannot be confirmed, the original 
‘response’ should be considered stable disease.  The best overall response will be defined 
according to the following table: 
 
Overall 
Response First 
Time Point 
Overall Response 
Subsequent Time 
Point 
BEST Overall Response 
CR 
CR 
CR 
CR 
PR 
SD, PD, or PR1 
CR 
SD 
SD provided minimum criteria for SD duration met, otherwise, PD 
CR 
PD 
SD provided minimum criteria for SD duration met, otherwise, PD 
CR 
NE2 
SD provided minimum criteria for SD duration met, otherwise, 
NE2 
PR 
CR 
PR 
PR 
PR 
PR 
PR 
SD 
SD 
PR 
PD 
SD provided minimum criteria for SD duration met, otherwise, PD 
PR 
NE2 
SD provided minimum criteria for SD duration met, otherwise, 
NE2 
NE 
NE2 
NE2 
1 If a CR is truly met at first time point, then any disease seen at a subsequent time point, even disease meeting PR criteria relative 
to baseline, makes the disease PD at that point (since disease must have reappeared after CR).  Best response would depend on 
whether minimum duration for SD was met.  However, sometimes ‘CR’ may be claimed when subsequent scans suggest small 
lesions were likely still present and in fact the patient had PR, not CR at the first time point.  Under these circumstances, the 
original CR should be changed to PR and the best response is PR. 
2  NE=inevaluable 
 
8.0 
ADVERSE EVENTS 
8.1 
Definitions  
8.1.1 Adverse Event (AE) 
An AE is any untoward medical occurrence (e.g., an abnormal laboratory finding, symptom, or 
disease temporally associated with the use of a drug) in a patient or clinical investigation subject 
administered a pharmaceutical product and which does not necessarily have a causal 
relationship with this treatment.  An AE can therefore be any unfavorable and unintended sign 
(including an abnormal laboratory finding), symptom, or disease temporally associated with the 
use of a medicinal product, whether or not related to the medicinal product.   
 
Laboratory abnormalities that do not require dose modification for entinostat, hospitalization 
for elective surgery or routine clinical procedures that are not the result of an AE (e.g., surgical 
insertion of central line) need not be considered AEs and should not be recorded as an AE.  


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Disease progression should not be recorded as an AE, unless it is attributable by the investigator 
to the study therapy. 
8.1.2 Suspected Adverse Reaction (SAR) 
A suspected adverse reaction (SAR) is any AE for which there is a reasonable possibility that 
the drug is the cause.  Reasonable possibility means that there is evidence to suggest a causal 
relationship between the drug and the AE.  A suspected adverse reaction implies a lesser degree 
of certainty about causality than adverse reaction, which means any AE caused by a drug.   
 
Causality assessment to a study drug is a medical judgment made in consideration of the 
following factors: temporal relationship of the AE to study drug exposure, known mechanism 
of action or side effect profile of study treatment, other recent or concomitant drug exposures, 
normal clinical course of the disease under investigation, and any other underlying or concurrent 
medical conditions.  Other factors to consider in considering drug as the cause of the AE: 
1. Single occurrence of an uncommon event known to be strongly associated with drug 
exposure (e.g., angioedema, hepatic injury, Stevens-Johnson Syndrome). 
2. One or more occurrences of an event not commonly associated with drug exposure, but 
otherwise uncommon in the population (e.g., tendon rupture); often more than once 
occurrence from one or multiple studies would be needed before the sponsor could 
determine that there is reasonable possibility that the drug caused the event.   
3. An aggregate analysis of specific events observed in a clinical trial that indicates the events 
occur more frequently in the drug treatment group than in a concurrent or historical control 
group. 
8.1.3 Unexpected AE or SAR 
An AE or SAR is considered unexpected if its specificity or severity is not consistent with the 
applicable product information (e.g., Investigator’s Brochure [IB] for an unapproved 
investigational product or package insert/summary of product characteristics for an approved 
product).  Unexpected also refers to AEs or SARs that are mentioned in the IB as occurring with 
a class of drugs or as anticipated from the pharmacological properties of the drug, but are not 
specifically mentioned as occurring with the particular drug under investigation.   
8.1.4 Serious AE or SAR 
A SAE is one that at any dose (including overdose): 
• Results in death 
• Is life-threatening1 
• Requires inpatient hospitalization or prolongation of existing hospitalization 
• Results in persistent or significant disability or incapacity2 
• Is a congenital anomaly or birth defect 
• Is an important medical event3 
• Pregnancy 
 
1 “Life-threatening” means that the subject was at immediate risk of death at the time of the SAE; it does not refer 
to a SAE that hypothetically might have caused death if it were more severe. 
2 “Persistent or significant disability or incapacity” means that there is a substantial disruption of a person’s ability 
to carry out normal life functions. 


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3 Medical and scientific judgment should be exercised in deciding whether expedited reporting is appropriate in 
situations where none of the outcomes listed above occurred.  Important medical events that may not be 
immediately life-threatening or result in death or hospitalization but may jeopardize the patient or may require 
intervention to prevent one of the other outcomes listed in the definition above should also usually be considered 
serious.  Examples of such events include allergic bronchospasm requiring intensive treatment in an emergency 
room or at home, blood dyscrasias or convulsions that do not result in inpatient hospitalization, or the development 
of drug dependency or drug abuse.  A new diagnosis of cancer during the course of a treatment should be considered 
as medically important.  
 
The investigator will report serious, unexpected reactions to the FDA, Syndax, and the IRB in 
accordance with applicable laws and regulations.   
 
8.2 
Documentation of non-serious AEs or SARs 
For non-serious AEs or SARs, documentation must begin from day 1 of study treatment and 
continue until patient completes the 30-day follow-up period assessments. 
 
Collected information should be recorded in the eCRF and source documentation appropriately. 
Please include a description of the event, its severity or toxicity grade, onset and resolved dates 
(if applicable), and the relationship to the study drug. Documentation should occur at least 
monthly.  
8.3 
SAEs or Serious SARs  
8.3.1 Timing 
After informed consent but prior to initiation of study medications, only SAEs caused by a 
protocol-mandated intervention will be collected (e.g. SAEs related to invasive procedures such 
as biopsies, medication washout).  
 
For any other experience or condition that meets the definition of an SAE or a serious SAR, 
recording of the event must begin from day 1 of study treatment and continue through the  
30-day follow-up period after treatment is discontinued.  
8.3.2 Documentation and Notification 
SAEs or Serious SARs must be recorded in the SAE console within OnCore® for that patient 
within 24 hours of learning of its occurrence and will reported to SYNDAX.  
 
8.4 
Adverse Event Reporting  
8.4.1 IRB Reporting Requirements: 
UNC: 
• The UNC-IRB will be notified of all SAEs that qualify as an Unanticipated Problem as 
per the UNC IRB Policies using the IRB’s web-based reporting system (see section 
9.5.3) within 7 days of the Investigator becoming aware of the problem.   
 


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Pregnancy 
Pregnancies and suspected pregnancies (including a positive pregnancy test regardless of age 
or disease state) of a female subject occurring while the subject is on study, or within 30 days 
of the subject’s last dose of study should be recorded as SAEs.  The patient is to be discontinued 
immediately from the study.  
 
The female subject should be referred to an obstetrician-gynecologist, preferably one 
experienced in reproductive toxicity for further evaluation and counseling. 
 
The Investigator will follow the female subject until completion of the pregnancy, and must 
document the outcome of the pregnancy (either normal or abnormal outcome) and report the 
condition of the fetus or newborn to the UNC Study Coordinator. If the outcome of the 
pregnancy was abnormal (e.g., spontaneous or therapeutic abortion), the Investigator should 
report the abnormal outcome as an AE. If the abnormal outcome meets any of the serious 
criteria, it must be reported as an SAE.   
8.4.2 Syndax Reporting Requirements: 
The investigator must inform Syndax in writing using a SAE form or MEDWATCH 3500A 
form of any SAE within 24 hours of being aware of the event.  The date of awareness should be 
noted on the report.  The written report must be completed and supplied to Syndax within 24 
hours/1 business day at the latest on the following working day.  The initial report must be as 
complete as possible, including details of the current illness and (serious) AE, and an assessment 
of the causal relationship between the event and the investigational product(s).  SAE reports 
must include the Syndax Study ID Number, the Site Number, the Subject ID Number, and the 
name of the PI.  Information not available at the time of the initial report (e.g., an end date for 
the AE or laboratory values received after the report) must be documented on a follow-up report.  
A final report to document resolution of the SAE is required.  SAEs should be sent via email to:  
SyndaxSAEReporting@syndax.com 
8.4.3 FDA Expedited Reporting requirements for studies conducted under an IND:  
A sponsor must report any suspected adverse reaction that is both serious and unexpected to the 
FDA. The sponsor must report an AE as a suspected adverse reaction only if there is evidence 
to suggest a causal relationship between the drug and the adverse event, such as: 
1. 
A single occurrence of an event that is uncommon and known to be strongly associated 
with drug exposure (e.g., angioedema, hepatic injury, Stevens-Johnson Syndrome); 
2. 
One or more occurrences of an event that is not commonly associated with drug 
exposure, but is otherwise uncommon in the population exposed to the drug (e.g. tendon 
rupture); 
3. 
An aggregate analysis of specific events observed in a clinical trial (such as known 
consequences of underlying disease or condition under investigation or other events that 
commonly occur in the study population independent of drug therapy) that indicates 
those events occur more frequently in the drug treatment group than in a concurrent or 
historical control group. 
The sponsor must submit each IND safety report on FDA Form 3500A. Each notification to 
FDA must bear prominent identification of its contents, i.e., ‘IND Safety Report,’ and must be 
transmitted to the review division that has the responsibility for review of the IND. In each IND 


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safety report, the sponsor must identify all IND safety reports previously submitted to FDA 
concerning a similar suspected adverse reaction and must analyze the significance of the 
suspected adverse reaction in light of previous, similar reports or any other relevant information.  
 
Timing 
FDA must be notified of potential serious risks within 15 calendar days after the sponsor 
determines the event requires reporting. FDA must be notified of unexpected fatal or life-
threatening suspected adverse reactions as soon as possible but in no case later than 7 calendar 
days after the sponsor’s initial receipt of the information. If the results of a sponsor’s 
investigation show that an AE not initially determined to be reportable is reportable, the sponsor 
must report such suspected adverse reaction in an IND safety report as soon as possible, but in 
no case later than 15 calendar days after the determination is made. 
 
Follow-up 
The sponsor must promptly investigate all safety information it receives. Relevant follow-up 
information to an IND safety report must be submitted as soon as the information is available 
and must be identified as such, i.e., ‘Follow-up IND Safety Report.’ Additionally, upon request 
from FDA, the sponsor must submit to FDA any additional data or information that the agency 
deems necessary, as soon as possible, but in no case later than 15 calendar days after receiving 
the request.  
 
Notification of Investigators 
The sponsor must notify all participating investigators (i.e., all investigators to whom the 
sponsor is providing drug under its INDs or under any investigator’s IND) in an IND safety 
report of potential serious risks, from clinical trials or any other source, as soon as possible, but 
in no case later than 15 calendar days after the sponsor determines that the information qualifies 
for reporting.  
 
Process 
If the sponsor deems that an event is both a serious adverse reaction (SAR) AND unexpected, it 
must also (in addition to OnCore®) be recorded on the MedWatch Form 3500A as per 21 CFR 
312.32. Unexpected AEs or adverse reaction refers to an event or reaction that is not listed in the 
investigator’s brochure or is not listed at the specificity or severity that has been observed; or if 
an investigator’s brochure is not required or available, is not consistent with the risk information 
described in the general investigation plan or elsewhere in the current IND application.   
 
The MedWatch form should be submitted on MedWatch by the study coordinator.  
 
The MedWatch 3500a form can be accessed at: 
http://www.fda.gov/Safety/MedWatch/HowToReport/DownloadForms/default.htm.  
 
Note: (Please be sure and access form 3500a, and not form 3500).  
 
The MedWatch form should also be sent to the UNC-CH Regulatory Associate and the IND 
Specialist within 48 hours of the sponsor being aware of the event.  The Regulatory Associate 


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with the aid of the IND Specialist will submit the IND Safety Report via IND serial submission 
to the FDA review division. 
 
All IND safety reports must be submitted on Form 3500A and be accompanied by Form 1571. 
The FDA must be notified or any unexpected or life-threatening suspected adverse reactions as 
soon as possible, but no later than 7 calendar days of learning of the event. 
 
Additional Reporting Requirements 
The following additional items must be reported via IND safety report: 
1. 
Findings from other studies. The sponsor must report any findings from epidemiological 
studies, pooled analysis of multiple studies, or clinical studies, whether or not conducted 
under an IND, and whether or not conducted by the sponsor, that suggest a significant 
risk to humans exposed to the drug.  
2. 
Findings from animal or in vitro testing. The sponsor must report any findings from 
animal or in vitro testing, whether or not conducted by the sponsor, that suggest a 
significant risk in humans exposed to the drug, such as reports of mutagenicity, 
teratogenicity, or carcinogenicity, or reports of significant organ toxicity at or near the 
expected human exposure. 
3. 
Increased rate of occurrence of serious suspected adverse reactions. 
 
Additional Guidance 
Please refer to 21CFR312.32 and “Guidance for Industry and Investigators: Safety Reporting 
Requirements for INDs and BA/BE Studies” for additional information and reporting 
requirements. All IND Safety Reports will be submitted in accordance with regulations and 
guidance provided.  
 
8.5 
Data and Safety Monitoring Plan 
The Principal Investigator will provide continuous monitoring of patient safety in this trial with 
periodic reporting to the Data and Safety Monitoring Committee (DSMC).  
 
Meetings/teleconferences will be held at a frequency dependent on study accrual and in 
consultation with the study Biostatistician.  These meetings will include the investigators as 
well as protocol nurses, clinical research associates, regulatory associates, data managers, 
biostatisticians, and any other relevant personnel the principal investigators may deem 
appropriate.  At these meetings, the research team will discuss all issues relevant to study 
progress, including enrollment, safety, regulatory, data collection, etc. 
 
The team will produce summaries or minutes of these meetings. These summaries will be 
available for inspection when requested by any of the regulatory bodies charged with the safety 
of human subjects and the integrity of data including, but not limited to, the oversight (Office 
of Human Research Ethics (OHRE) Biomedical IRB, the Oncology Protocol Review 
Committee (PRC) or the North Carolina TraCS Institute Data and Safety Monitoring Board 
(DSMB).   
 


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The UNC-CH LCCC Data and Safety Monitoring Committee (DSMC) will review the study on 
a regular (quarterly to annually) basis, with the frequency of review based on risk and 
complexity as determined by the UNC-CH Protocol Review Committee.  The UNC PI will be 
responsible for submitting the following information for review: 1) safety and accrual data 
including the number of patients treated; 2) significant developments reported in the literature 
that may affect the safety of participants or the ethics of the study; 3) preliminary response data; 
and 4) summaries of team meetings that have occurred since the last report.  Findings of the 
DSMC review will be disseminated by memo to the UNC PI, PRC, and the UNC IRB and 
DSMB.   
 
9.0 
STATISTICAL CONSIDERATIONS 
This is a phase II study using one FDA-approved drug, pembrolizumab, and an investigational 
agent, entinostat.  
9.1 
Study Design/Study Endpoints 
All patients will receive pembrolizumab plus entinostat for up to 27 weeks. Drugs will be 
sequentially introduced; such that correlative analysis can be performed to investigate the 
indirect immunomodulatory effect of entinostat alone and in combination with pembrolizumab 
by performing serial peripheral blood collections (4 time points) and tumor collection (2 time 
points). Assessment of response to the treatment combination will be assessed at week 10 and 
then every 6 weeks of pembrolizumab treatment for up to 27-weeks.  
 
9.1.1  Definitions 
Evaluable patients are those who complete all activities by at least day 22, namely: (a) they have 
received at least one and up to 4 entinostat weekly doses on day 1 and day 8, 15 and 22 (b) they 
have received the first pembrolizumab infusion on day 22 (±2 days), and (c) they have 
completed day 22 (±2 days) mandatory tumor biopsy. The rationale for this is to answer the 
primary translational endpoint regarding entinostat-alone immunomodulatory effects in 
peripheral blood and tumor tissue with sufficient power.  
- SAE are events that meet FDA’s definition as defined in section 8.1.4 of the protocol 
irrespective of attribution to study drug(s). Subjects who receive at least one dose of entinostat 
are evaluable for safety. 
-ORR by RECIST 1.1 criteria is the sum of complete response and partial response. 
-PFS is measured from the date of enrollment on study to the date of documented progression. 
The rate of patients who are progression-free will be specifically calculated at 27-weeks 
(approximately 6 months) from study entry. 
-OS is measured from the date of enrollment on study to the recorded date of death. The rate of 
patients who are alive at 27-week (approximately 6 months) will be specifically calculated for 
the study. 
 
9.1.2  Correlative Studies 










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10.2 
Required Documentation 
Before the study can be initiated at any site, the following documentation must be provided to 
the Clinical Protocol Office (CPO) at the UNC-CH. 
• A copy of the official IRB approval letter for the protocol and informed consent 
• IRB membership list 
• CVs and medical licensure for the principal investigator and any sub-investigators who will 
be involved in the study. 
• Form FDA 1572 appropriately filled out and signed with appropriate documentation  
• Financial Disclosures  
• CAP and CLIA Laboratory certification numbers and institution lab normal values 
• Executed clinical research contract 
 
10.3 
Registration Procedures 
All patients must be registered with the CPO at the UNC-CH before enrollment to study.  Prior 
to registration, eligibility criteria must be confirmed with the UNC-CH Study Coordinator.  To 
register a patient, call the CPO at 919-966-4432 Monday-Friday 9:00 am – 5:00 pm EST or 
email the UNC Project Manager. 
 
10.4 
Data Management and Monitoring/Auditing 
The CPO of the UNC LCCC will serve as the coordinating center for this trial. Data will be 
collected through a web-based clinical research platform, OnCore®. All data will be collected 
and entered into OnCore® by research coordinators from UNC LCCC.  
 
The sponsor will provide direct access to source data/documents for trial-related monitoring, 
audits, IRB/IEC review, and regulatory inspection. As an investigator-initiated study, this trial 
will also be audited by the LCCC compliance committee every six or twelve months. 
 
The sponsor will provide direct access to source data/documents for trial-related monitoring, 
audits, IRB/IEC review, and regulatory inspection.  As an investigator initiated study, this trial 
will also be audited by the Lineberger Cancer Center compliance committee every six or twelve 
months, depending on the participation of affiliate sites. 
10.5 
Adherence to the Protocol 
Except for an emergency situation in which proper care for the protection, safety, and well-
being of the study patient requires alternative treatment, the study shall be conducted exactly as 
described in the approved protocol.   
10.5.1 Emergency Modifications 
UNC investigators may implement a deviation from, or a change of, the protocol to eliminate 
an immediate hazard(s) to trial subjects without prior UNC or their respective institution’s 
IRB/IEC approval/favorable opinion.   
 


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For any such emergency modification implemented, a UNC IRB modification form must be 
completed by UNC Research Personnel within five (5) business days of making the change.   
10.5.2 Single Patient/Subject Exceptions 
Eligibility single subject exceptions are not permitted for Lineberger Comprehensive Cancer 
Center Investigator-Initiated Trials under any circumstances. Other types of single-subject 
exceptions may be allowed if proper regulatory review has been completed in accordance with 
Lineberger Comprehensive Cancer Center’s Single Subject Exceptions Policy. 
 
10.5.3 Other Protocol Deviations/Violations 
According to UNC’s IRB, a protocol deviation is any unplanned variance from an IRB approved 
protocol that:  
• Is generally noted or recognized after it occurs, 
• Has no substantive effect on the risks to research participants, 
• Has no substantive effect on the scientific integrity of the research plan or the value of the 
data collected, 
• Did not result from willful or knowing misconduct on the part of the investigator(s).  
 
An unplanned protocol variance is considered a violation if the variance meets any of the 
following criteria:  
• Has harmed or increased the risk of harm to one or more research participants. 
• Has damaged the scientific integrity of the data collected for the study. 
• Results from willful or knowing misconduct on the part of the investigator(s). 
• Demonstrates serious or continuing noncompliance with federal regulations, State laws, or 
University policies. 
 
If a deviation or violation occurs, please follow the guidelines below: 
 
Protocol Deviations: UNC will record the deviation in OnCore®, and report to any sponsor or 
data and safety monitoring committee in accordance with their policies.  Deviations should be 
summarized and reported to the IRB at the time of continuing review. 
 
Protocol Violations: Violations should be reported by UNC personnel within one (1) week of 
the investigator becoming aware of the event using the same IRB online mechanism used to 
report Unanticipated Problems.   
 
Protocol Deviations: In the event a deviation from protocol procedures is identified, record the 
deviation in OnCore® and assess for risk of patient safety. 
 
Unanticipated Problems: 
Any events that meet the criteria for “Unanticipated Problems” as defined by UNC’s IRB must 
be reported by the study personnel using the IRB’s web-based reporting system.   
 


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10.6 
Amendments to the Protocol 
Should amendments to the protocol be required, the amendments will be originated and 
documented by the Principal Investigator at UNC.  It should also be noted that when an 
amendment to the protocol substantially alters the study design or the potential risk to the 
patient, a revised consent form might be required.   
 
The written amendment, and if required the amended consent form, must be sent to UNC’s IRB 
for approval prior to implementation.   
 
10.7 
Record Retention 
Study documentation includes all eCRFs, data correction forms or queries, source documents, 
Sponsor correspondence to Investigators, monitoring logs/letters, and regulatory documents 
(e.g., protocol and amendments, IRB correspondence and approval, signed patient consent 
forms). 
 
Source documents include all recordings of observations or notations of clinical activities and 
all reports and records necessary for the evaluation and reconstruction of the clinical research 
study. 
 
Government agency regulations and directives require that all study documentation pertaining 
to the conduct of a clinical trial must be retained by the study investigator.  In the case of a study 
with a drug seeking regulatory approval and marketing, these documents shall be retained for 
at least two years after the last approval of marketing application in an International Conference 
on Harmonization (ICH) region.  In all other cases, study documents should be kept on file until 
three years after the completion and final study report of this investigational study. 
 
10.8 
Obligations of Investigators 
The Principal Investigator is responsible for the conduct of the clinical trial at the site in 
accordance with Title 21 of the Code of Federal Regulations and/or the Declaration of Helsinki.  
The Principal Investigator is responsible for personally overseeing the treatment of all study 
patients.  The Principal Investigator must assure that all study site personnel, including sub-
investigators and other study staff members, adhere to the study protocol and all FDA/GCP/NCI 
regulations and guidelines regarding clinical trials both during and after study completion. 
 
The Principal Investigator will be responsible for assuring that all the required data will be 
collected and entered into eCRFs. Periodically, monitoring visits will be conducted and the 
Principal Investigator will provide access to his/her original records to permit verification of 
proper entry of data. At the completion of the study, all eCRFs will be reviewed by the Principal 
Investigator and will require his/her final signature to verify the accuracy of the data. 
 
 


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LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
97 
 
12.0 APPENDICES 
APPENDIX A  
PROHIBITED MEDICATIONS FOR ENTINOSTAT 
 
Examples of sensitive in vivo CYP substrates and CYP substrates with narrow therapeutic 
range are summarized below.  
  
Examples of substrates that may be affected by entinostat 
CYP Enzymes  
Substrates with narrow therapeutic range1 
CYP1A2  
Theophylline, tizanidine  
CYP2C8  
Paclitaxel 
CYP3A2  
Alfentanil, astemizole3, cisapride3, cyclosporine, dihydroergotamine, 
ergotamine, fentanyl, pimozide, quinidine, sirolimus, tacrolimus, 
terfenadine3  
1. CYP substrates with narrow therapeutic range refers to drugs whose exposure-response relationship 
indicates that small increases in their exposure levels by the concomitant use of CYP inhibitors may lead 
to serious safety concerns (e.g., torsade de pointes). 
2. Because a number of CYP3A substrates (e.g., darunavir, maraviroc) are also substrates of P-gp, the 
observed increase in exposure could be due to inhibition of both CYP3A and P-gp. Note: The prohibited 
use of fentanyl does not apply to the study biopsy procedure. 
3. Withdrawn from the United States market because of safety reasons. 
 
 
P-gp Inhibitors and Inducers 
 
Inhibitors 
Inducers 
Amiodarone, azithromycin, captopril, carvedilol, 
clarithromycin, conivaptan, diltiazem, dronedarone, 
felodipine, lopinavir, quercetin, ranolazine, ticagrelor,  
ritonavir, cyclosporine, verapamil, erythromycin, 
ketoconazole,  itraconazole, quinidine  
Avasimibe, carbamazepine, phenytoin, 
rifampin, St John’s Wort, tipranavir, 
ritonavir  
 
 
 


LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
98 
APPENDIX B  
PATIENT HANDOUT-PROHIBITED MEDICATIONS 
 
Prohibited Medications or those to be used with caution 
One of the medications you are receiving during this clinical trial is entinostat.   
Entinostat interacts with some drugs that are processed by your liver. Because of 
this, it is very important to tell your study doctors about all of your medicine before 
you start this study. It is also very important to tell them if you stop taking any 
regular medicine, or if you start taking a new medicine while you take part in this 
study. When you talk about your medicine with your study doctor, include medicine 
you buy without a prescription at the drug store (over-the counter remedy), or 
anything that you buy from the health food store or grocery store (herbal 
supplement). Many health care prescribers can write prescriptions. You must also 
tell your other prescribers (doctors, physicians’ assistants or nurse practitioners) 
that you are taking part in a clinical trial. Bring this paper with you.  
• Entinostat is processed by a certain enzyme in the liver called CYPs including 
CYP3A4.  Entinostat should not be given with drugs that are sensitive substrates 
of CYP1A2, CYP2C8, and CYP3A that have a narrow therapeutic window  
• You and healthcare providers who prescribe drugs for you must be careful about 
adding or removing any drugs in the above categories. 
• Before you start the study, your study doctor will work with your regular 
prescriber to switch the following medications if you are taking them: 
Theophylline, 
tizanidine, 
paclitaxel, 
alfentanil, 
astimizole, 
cisapride, 
cyclosporine, dihydroergotamine, ergotamine, fentanyl, pimozide, quinidine, 
sirolimus, tacrolimus, and terfenadine. (Note: The prohibited use of fentanyl 
does not apply to the study biopsy procedure) 
• You should also avoid drugs that are Pgp inhibitors or inducers: Amiodarone, 
azithromycin, captopril, carvedilol, clarithromycin, conivaptan, diltiazem, 
dronedarone, felodipine, lopinavir,quercetin, ranolazine,ticagrelor, ritonavir, 
cyclosporine, verapamil erythromycin, ketoconazole,  itraconazole, quinidine 
Avasimibe, 
carbamazepine, 
phenytoin, 
rifampin, 
St 
John’s 
Wort, 
tipranavir/ritonavir  
• Your regular prescribers should look at Appendix 12.1 in the study protocol and 
refer to this web site: http://medicine.iupui.edu/clinpharm/ddis/table.asp. Your 
study doctor may also have a list of medications for you to show your regular 
prescribers instead of, or in addition to, this website.   
• Other medicines can be a problem with your study drugs. 
• You should check with your doctor or pharmacist whenever you need to use an 
over-the-counter medicine or herbal supplement. 
 
Your regular prescriber should check a medical reference or call your study doctor before 
prescribing any new medicine for you. Your study doctor’s name is _______________ and 
he or she can be contacted at _______________________. 
 


LCCC 1729 
 
CONFIDENTIAL 
PI: Stergios Moschos, M.D. 
 
UNIVERSITY OF NORTH CAROLINA 
Amendment 2   
 
December 15, 2021 
 
99 
APPENDIX C  
ARCHIVED TUMOR TISSUE FOR ENROLLMENT 
 
Below is a sample letter (e-mail of this form is also acceptable) from the pathologist 
stating the adequacy of archived of tumor tissue for patient eligibility 
 
Dear Dr. Moschos, 
 
I have reviewed the hematoxylin and eosin-stained (H&E) slides corresponding to 
[SCREENING 
SUBJECT 
ID 
NUMBER 
and 
SUBJECT 
INITIALS]’s 
[PATIENT’S NUMBER OF AVAILABLE METASTATIC MELANOMA 
TISSUE BLOCKS) archived tumor block from the [PATIENT’S DIFFERENT 
PATHOLOGY ACCESION NUMBERS CORRESPONDIND TO PREVIOUS 
STANDARD OF CARE SURGICAL OR BIOPSY PROCEDURES]. I have 
confirmed that: 
 
The total tumor surface area from all the melanoma tissue blocks I have reviewed 
is at least 1 cm2.  will release 11-15 5-micron tissue section on charged slides to 
your institution for research analysis. 
 
The tumor contains no more than 20% necrosis  
 
The ratio of melanoma to stroma cells is at least 60:40. 
 
I attest that none of the H&E slides I have reviewed contains evidence of ≥ 1% 
tumor-infiltrating or tumor-associated mononuclear cells. 
 
I attest that the total volume of corresponding tumor tissue that is contained within 
these block(s) is sufficient to yield fifteen (15) 10-micron tumor sections for your 
molecular correlative studies (i.e. FoundationOne® testing, RNA-seq, FAIRE 
analysis) 
 
I attest that the total volume of corresponding tumor tissue that is contained within 
these block(s) is sufficient to yield addition ten (10) 5-micron tumor sections for 
your 
tumor 
imaging 
correlative 
studies 
(immunohistochemistry 
and/or 
immunofluorescence) 
 
 
Sincerely, 
 
Dr. Paul B. Googe 
[PRINTED NAME AND MAILING ADDRESS]