Conditions: Melanoma
Keywords: Resectable Stage IIIB, IIIC, and IVM1a
Allocation: NA | Intervention model: SINGLE_GROUP
Enrollment: 7
Healthy volunteers: no

Brief summary:
This is a single-arm, open label, Phase II study evaluating the safety and efficacy of neoadjuvant Nivolumab and HF10 in resectable stage IIIB, IIIC, and IVM1a melanoma.

Detailed description:

Arms:
arm 1: None

Interventions: intervention 1: Nivolumab intervention 2: HF10
Intervention descriptions: intervention 1: Nivolumab at a dose of 240 mg given as an IV infusion starting on day 0. It will be given every 14 days for a total of 7 infusions; Then participant will undergo surgery. Nivolumab will then be administered at a flat dose of 480 mg IV every 28 days for up to one year. intervention 2: 1 x 107th TCID50/mL, intratumoral injection to a single or multiple eligible tumors for a total of 5 mL; on days 0, 7, 14, 21, 28, 42, 56, 70, 84 for a total of 9 injections. All eligible tumors except one will be treated with HF10 up to the maximum volume allowed. The untreated tumor will be used as an untreated control lesion.

Protocol document:
Prot_SAP_003.pdf: 
 
Protocol name: Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable 
Stage IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
 
  
 
Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic 
Viral Therapy in Resectable Stage IIIB, IIIC, IVM1a Melanoma (Neo-NivoHF10) 
Trial ID: HCI102346/ IRB#102346  
Drug Manufacturer protocol # CA209-9F3 
NCT03259425 
 
Principal Investigator 
John Hyngstrom, MD 
Huntsman Cancer Institute/University of Utah 
2000 Circle of Hope Drive 
Salt Lake City, UT 84112 
801-585-0255 
john.hyngstrom@hci.utah.edu 
 
Sub-investigator(s) 
Brittany Thomas, PA-C 
Huntsman Cancer Institute/University of Utah 
brittany.thomas@hci.utah.edu 
 
Elizabeth Flores, PA 
Huntsman Cancer Institute/University of Utah 
elizabeth.flores@hci.utah.edu 
 
Carolyn Luckett, APRN 
Huntsman Cancer Institute/University of Utah 
carolyn.luckett@hci.utah.edu 
 
Siwen Hu-Lieskovan, MD 
Huntsman Cancer Institute/University of Utah 
Siwen.Hu-Lieskovan@hci.utah.edu  
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 2 of 61 
 
Kenneth Grossmann, MD 
Huntsman Cancer Institute/University of Utah 
Kenneth.grossmann@hci.utah.edu 
 
Statistician 
 
 
 
Drug Manufacturer  
Kenneth Boucher, PhD 
Huntsman Cancer Institute/University of Utah 
Ken.Boucher@hci.utah.edu 
 
Takara Bio Inc. 
Nojihigashi 7-4-38, Kusatsu 
Shiga 525-0058, Japan 
+81-77-565-6976  
 
Bristol-Myers Squibb 
PO Box 5100 
Wallingford, CT 06492-7660 
203-677-6000 
 
Investigational agent 
HF10  
Nivolumab (OPDIVO) 
 
 
IND Number 
 
17693  
 
 
 
 
 
 
 
 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 3 of 61 
 
Historical Protocol Versions 
Version 1: 25SEP2017 
Version 2: 08AUG2018 
Version 3: 06JUN2019 
Version 4: 22JUL2019 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 4 of 61 
 
TABLE OF CONTENTS 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Page 
PROTOCOL SIGNATURE PAGE……………………………….....................................9 
STUDY SUMMARY  …………………………………………………………………...10 
 
1 
OBJECTIVES ........................................................................................................... 16 
 
Primary Objectives and Endpoint ...................................................................... 16 
 
Secondary Objectives and Endpoint .................................................................. 16 
 
Exploratory Objective and Endpoint .................................................................. 16 
2 
BACKGROUND ...................................................................................................... 18 
 
Introduction ........................................................................................................ 18 
 
The Investigational Product ............................................................................... 18 
 
Nonclinical and Preclinical Studies.................................................................... 20 
 
Clinical Studies with HF10 ................................................................................ 24 
3 
Potential risks and benefits ....................................................................................... 27 
 
HF10 ................................................................................................................... 27 
 
Nivolumab .......................................................................................................... 28 
4 
STUDY DESIGN...................................................................................................... 29 
 
Description ......................................................................................................... 29 
 
Number of Patients ............................................................................................. 29 
 
Number of Study Centers ................................................................................... 29 
 
Study Duration ................................................................................................... 29 
5 
ELIGIBILITY CRITERIA........................................................................................ 30 
 
Inclusion Criteria ................................................................................................ 30 
 
Exclusion Criteria ............................................................................................... 31 
6 
TREATMENT PLAN ............................................................................................... 33 
 
Administration Schedule .................................................................................... 33 
 
Nivolumab .......................................................................................................... 34 
 
HF10 ................................................................................................................... 35 
 
Prohibited Concomitant Medications ................................................................. 35 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 5 of 61 
 
 
Duration of Therapy ........................................................................................... 36 
7 
TOXICITIES AND DOSEAGE MODIFICATION ................................................. 36 
 
Dose Modifications ............................................................................................ 36 
 
Subject Compliance............................................................................................ 39 
 
Patient Precautions for Minimizing Virus Exposure to Others .......................... 39 
 
Healthcare Provider Precautions ........................................................................ 39 
 
Supportive Care .................................................................................................. 40 
8 
STUDY CALENDAR .............................................................................................. 41 
9 
CRITERIA FOR EVALUATION AND ENDPOINT .............................................. 43 
 
Pathological response ......................................................................................... 43 
 
Safety .................................................................................................................. 43 
 
Recurrence Free Survival ................................................................................... 43 
 
Neo-Adjuvant Therapy Tumor Response .......................................................... 44 
 
irRC and RECIST 1.1 Concordance .................................................................. 45 
 
Overall Survival ................................................................................................. 47 
 
Correlative Biomarker Analysis ......................................................................... 47 
 
Complete Surgical Resection ............................................................................. 47 
 
Pathology ............................................................................................................ 47 
 
Stopping Rules ................................................................................................ 47 
10 STATISTICAL CONSIDERATIONS...................................................................... 47 
11 REGISTRATION GUIDELINES ............................................................................. 48 
12 DATA SUBMISSION SCHEDULE ........................................................................ 48 
13 SPECIAL INSTRUCTIONS – CORRELATIVE STUDIES ................................... 49 
 
Tissue Samples ............................................................................................... 49 
 
Blood Samples ................................................................................................ 49 
14 ETHICAL AND REGULATORY CONSIDERATIONS ........................................ 50 
 
Informed consent ............................................................................................ 50 
 
Institutional Review ........................................................................................ 50 
 
Data and Safety Monitoring Plan ................................................................... 50 
 
Adverse Events / Serious Adverse Events ...................................................... 50 
 
SAE Reporting Requirements ........................................................................ 52 
 
Reporting of Pregnancy .................................................................................. 54 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 6 of 61 
 
 
Protocol Amendments .................................................................................... 54 
 
Protocol Deviations ........................................................................................ 55 
 
FDA Annual Reporting .................................................................................. 55 
 
Clinical Trials Data Bank ............................................................................... 55 
 
Record Keeping .............................................................................................. 55 
15 BIBLIOGRAPHY ..................................................................................................... 56 
16 Appendix ................................................................................................................... 60 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 7 of 61 
 
LIST OF ABBREVIATIONS 
Abbreviation or 
Term1 
Definition/Explanation 
ACV 
Acyclovir  
AE 
Adverse event 
ALT 
Alanine aminotransferase 
ANCOVA 
Analysis of covariance 
ANOVA 
Analysis of variance 
APTT 
Activated partial thromboplastin time 
AST 
Aspartate aminotransferase 
AV 
Atrioventricular 
β-HCG 
Beta-human chorionic gonadotropin 
BID 
Twice daily 
BLQ 
Below limit of quantification 
BMI 
Body mass index 
BP 
Blood pressure 
BUN 
Blood urea nitrogen 
Ca++ 
Calcium 
CBC 
Complete blood count 
CFR 
Code of Federal Regulations 
CHF 
Congestive heart failure 
CI 
Confidence interval 
Cl- 
Chloride 
CLcr 
Creatinine clearance 
Cmax 
Maximum observed concentration 
Cmin 
Trough observed concentration 
CNS 
Central nervous system 
CR 
Complete response 
CRF 
Case report form 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 8 of 61 
 
Abbreviation or 
Term1 
Definition/Explanation 
CT 
Computed tomography 
CTCAE 
Common Toxicity Criteria for Adverse Events 
CV 
Coefficient of variation 
CYP 
Cytochrome P450 
D/C 
Discontinue 
ECOG 
Eastern Cooperative Oncology Group 
eCRF 
Electronic case report form 
DLT 
Dose Limiting Toxicity 
ECG 
Electrocardiogram 
Eg 
Exempli gratia (for example) 
FACS 
Fluorescence Activated Cell Sorting  
FDA 
Food and Drug Administration 
FDG-PET 
Fluorodeoxyglucose (FDG)-positron emission tomography (PET) 
GCP 
Good Clinical Practice 
GFR 
Glomerular filtration rate 
GGT 
Gamma glutamyl transferase 
GLP 
Good laboratory practice  
HBsAg 
Hepatitis B surface antigen 
HBV 
Hepatitis B virus 
HCO3- 
Bicarbonate 
HCV 
Hepatitis C virus 
HIV 
Human immunodeficiency virus 
HR 
Heart rate 
hr 
Hour or hours 
IC50 
Half maximal inhibitory concentration 
i.e. 
Id est (that is) 
IEC 
Independent ethics committee 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 9 of 61 
 
Abbreviation or 
Term1 
Definition/Explanation 
IFN 
Interferon 
INR 
International normalized ratio 
IRB 
Institutional review board 
irRC 
Immune-related response criteria  
IU 
International unit 
IV 
Intravenous, intravenously 
LDH 
Lactate dehydrogenase 
LLQ 
Lower limit of quantitation 
MedRA 
Medical Dictionary for Drug Regulatory Activities 
MRI 
Magnetic resonance imaging 
MRSD 
Maximum recommended starting dose 
MTD 
Maximum tolerated dose  
NOAEL 
No-observed-adverse-effect level 
NOEL 
No-observed-effect-level 
NSCLC 
Non-small cell lung cancer  
PD 
Pharmacodynamic(s) 
PFS 
Progression Free Survival 
PK 
Pharmacokinetic(s) 
PO 
Per os (administered by mouth) 
PR 
Partial response 
PT 
Prothrombin time 
PTT 
Partial thromboplastin time 
QC 
Quality control 
QD 
Once daily 
QTc 
QT interval corrected  
QTcF 
QT interval corrected using Fridericia equation 
RBC 
Red blood cell 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 10 of 61 
 
Abbreviation or 
Term1 
Definition/Explanation 
RFS 
Recurrence-free Survival 
SAE 
Serious adverse event 
SD 
Standard deviation or stable disease 
T1/2 
Terminal elimination half-life 
T3 
Triiodothyronine 
T4 
Thyroxine 
Tmax 
Time of maximum observed concentration 
TID 
Three times daily 
TSH 
Thyroid-stimulating hormone 
ULN 
Upper limit of normal 
ULQ 
Upper limit of quantitation 
UV 
Ultraviolet 
WBC 
White blood cell 
WOCBP 
Women of childbearing potential 
WONCBP 
Women of nonchildbearing potential 
All of these abbreviations may or may not be used in protocol.  
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 11 of 61 
 
PROTOCOL SIGNATURE 
 
 
I confirm that I have read this protocol, and I will conduct the study as outlined herein 
and according to the ethical principles stated in the latest version of the Declaration of 
Helsinki, the applicable ICH guidelines for good clinical practice, and the applicable laws 
and regulations of the federal government. I will promptly submit the protocol to the IRB 
for review and approval. Once the protocol has been approved by the IRB, I understand 
that any modifications made during the course of the study must first be approved by the 
IRB prior to implementation except when such modification is made to remove an 
immediate hazard to the subject. 
 
I will provide copies of the protocol and all pertinent information to all individuals 
responsible to me who assist in the conduct of this study. I will discuss this material with 
them to ensure that they are fully informed regarding the study treatment, the conduct of 
the study, and the obligations of confidentiality. 
 
This document is signed electronically through submission and approval by the Principal 
Investigator at Huntsman Cancer Institute in the University of Utah IRB Electronic 
Research Integrity and Compliance Administration (ERICA) system. For this reason, the 
Principal Investigator at Huntsman Cancer Institute will not have a hand-written 
signature on this signature page.  
 
Instructions to multi-site Principal Investigators at locations other than Huntsman 
Cancer Institute: SIGN and DATE this signature page and PRINT your name. Return 
the original, completed and signed, to the HCI Research Compliance Office. Retain a 
copy in the regulatory binder.  
 
 
____________________________________ 
_______________ 
Signature of Principal Investigator  
 
Date 
 
 
 
____________________________________ 
Principal Investigator Name (Print) 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 12 of 61 
 
 
 
 
____________________________________ 
Name of Institution 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 13 of 61 
 
STUDY SUMMARY 
 
Title 
Phase II Neoadjuvant trial of Nivolumab in Combination 
with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIC, IVM1a Melanoma (Neo-NivoHF10) 
Short Title 
Nivolumab and HF10 for the Neoadjuvant treatment of 
Advance Melanoma 
Protocol Number 
IRB #102346 
IND 
IND #17693 
Phase 
Phase II  
Design 
Multi-center, open-label phase II safety, tolerability and 
efficacy study.  
Study Duration 
1 year and 3 months for enrollment, 3 years for study 
completion. 
Study Center(s) 
Multi-center. This study will be conducted at the 
Huntsman Cancer Institute, and additional studies to be 
identified.  
Objectives 
Primary objective:  
Pathologic response including complete response after 12 
weeks of neoadjuvant treatment with Nivolumab and 
HF10. 
Secondary objectives:  
Recurrence-free survival, safety, biomarker analysis, 
overall survival, complete surgical resection. 
Number of Subjects 
This study is planned to enroll 20 patients with a 
possibility to increase enrollment based on observed 
pathologic response.  
Eligibility Criteria 
Inclusion  
1. Patients must be >18 years or older.  
2. Patients must have stage IIIB, IIIC, or IVM1a 
(equivalent staging at enrollment via AJCC 7th edition) 
metastatic melanoma which is eligible for complete 
surgical resection.  


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 14 of 61 
 
3. Prior systemic, regional and radiation anticancer 
therapies must have been completed at least three months 
prior to enrollment.  
4. Patients must be a candidate for intralesional therapy.  
5. Patients must have an Eastern Cooperative Oncology 
Group (ECOG) performance status of 0 or 1. 
6. Serum LDH level < 1.5 upper limit of normal (ULN) 
within 28 days prior to enrollment (for stage IIIB and 
IIIC). 
7. Patients have adequate organ function within 28 days 
prior to enrollment.  
Exclusion 
1. Patients with active visceral, central nervous system, or 
any bone metastases melanoma (Stage IVM1b or IVM1c).  
2. Patients whose primary diagnosis was ocular 
melanoma.  
3. Patients receiving anti-herpes medication (i.e., 
acyclovir, famciclovir, or valacyclovir) within 1 week 
prior to initiating HF10 treatment. Patients may not 
require intermittent or chronic systemic (intravenous or 
oral) treatment with an antiherpetic drug other than 
intermittent topical use.  
4. Patients who have an active herpetic skin lesion(s) or 
prior complications of HSV-1 infection.  
5. Uncontrolled intercurrent illness including, but not 
limited to, ongoing or active infection, symptomatic 
congestive heart failure, unstable angina pectoris, cardiac 
arrhythmia, or psychiatric illness/social situations that 
would limit compliance with study requirements, as 
determined by the investigator. 
6. Medical history of autoimmune disease (e.g. Crohn’s 
disease, ulcerative colitis) or other disease requiring 
systemic glucocorticoid or immunosuppressive therapy.  
7. Patients with clinically evident Human 
Immunodeficiency Virus (HIV), Hepatitis B Virus 
(HBV), Hepatitis C Virus (HCV), or Epstein-Barr Virus 
(EBV) infection are excluded. 
Study Product, Dose, 
Route, Regimen 
Neoadjuvant Nivolumab dose will be 240mg IV Q 2wks, 
and HF10 will be 1x107 TCID50/mL intratumoral injection 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 15 of 61 
 
for a maximum of 5mL per injection on days 0, 7, 14, 21, 
28, 42, 56, 70, 84 for a total of 9 injections.  
 
Adjuvant Nivolumab dose will be 480 mg IV Q4wks.  
Duration of administration A total of 7 IV infusions of Nivolumab and 9 intratumoral 
injections of HF10 will be administered neoadjuvantly. 
After surgery, patients will receive adjuvant Nivolumab 
every 4 weeks for one year.  
Statistical Methodology 
The primary endpoint of pathologic response including 
pCR is expected to be seen in 30% of patients, hence 6 of 
the planned 20 patients are expected to have a pathologic 
response including pCR.  
 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 16 of 61 
 
1 
OBJECTIVES 
 
Primary Objectives and Endpoint 
 Assess pathologic response rate (including complete response) after completion of 
12 weeks of neoadjuvant treatment with nivolumab and HF10. 
Endpoint: tumor viablitity will be assessed at surgery to assess pathologic 
response. 
 
Secondary Objectives and Endpoint 
 Assess recurrence-free survival 
Endpoint: recurrence after surgery will be assessed by radiologic scans 
scheduled per section 8 and confirmed by biopsy. 
 Assess overall survival 
Endpoint: patients will be followed for survival for one year after 
completion of adjuvant nivolumab. 
 Assess the response of tumors prior to surgery based on RECIST 1.1 
Endpoint: patients will be assessed radiographically with CT or MRI scan 
or assessed clinically. 
 Assess complete surgical resection 
Endpoint: patients will be assessed at surgery to determine if complete 
surgical resection was achievable after neo-adjuvant treatment with 
nivolumab and HF10. 
 Assess the safety and tolerability of neoadjuvant nivolumab and HF10 with 
adjuvant nivolumab in patients with metastatic melanoma. 
Endpoint: patients will be monitored for adverse events (AEs) and serious 
adverse events (SAEs) related to either or both nivolumab and HF10. 
 
Exploratory Objective and Endpoint  
 Assess tissue and blood biomarkers 
Endpoint:  tissue and blood collection will occur throughout study 
treatment to assess changes in tumor markers, environment, immune cell 
population, cytokines and cell free (cf) DNA. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 17 of 61 
 
 Assess concordance between irRC and RECIST 1.1 response assessments 
Endpoint: radiographic response assessment will be performed using 
RECIST 1.1, in addition to irRC, to explore concordance between the two 
response criteria. 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 18 of 61 
 
2 
BACKGROUND 
 
Introduction 
Patients with resectable stage IIIB, IIIC, or IVM1a melanoma still represent a patient 
population with significant unmet medical need as up to 60% have disease recurrence at 
18 months following surgical resection.1-3 Current adjuvant therapies in these patients 
benefit less than 30% of patients.4-9 Given the advances in the metastatic melanoma 
treatment landscape and the potential removal of antigen for immune system priming in 
the adjuvant setting, a key question remains: Does it make sense to treat these patients in 
the adjuvant setting or is neoadjuvant therapy more appropriate? Utilizing 
immunotherapies such as nivolumab combined with an intralesional oncolytic virus such 
as HF10 in the neoadjuvant treatment environment is likely to yield more clinical benefit 
because:  
1. Nivolumab and HF10 are well tolerated and have been shown to have efficacy in 
unresectable metastatic melanoma patients.10-13 
2. Tumor Antigen is present as an immune system target. The increased tumor 
burden in patients undergoing neoadjuvant treatment compared to patients who 
have had their tumor resected and are receiving adjuvant treatment may provide a 
more robust immune activation. 
3. There is potential synergy for added efficacy with an oncolytic virus in 
combination with a checkpoint inhibitor. Indeed, the precedent for the 
combination of Nivolumab and HF10 (an oncolytic virus) in melanoma has been 
established via other immune-oncology oncolytic virus studies (e.g. ipilimumab + 
HF10, ipilimumab + talimogene laherparepvec and pembrolizumab+talimogene 
laherparepvec) displaying initial robust efficacy and tolerable safety signals.13,14 
4. Nivolumab neoadjuvant clinical trials are currently on-going in non-small cell 
lung cancer and genitourinary cancers, while no such trials have started in 
resectable metastatic melanoma.  
 
The Investigational Product  
 HF10 
HF10 is an attenuated spontaneous mutant oncolytic HSV-1 strain that has not 
been modified by genetic recombination and contains no external genes. The 
genome structure of HF10 has numerous deletions and insertions, resulting in the 
lack of the functional expression of UL43, UL49.5, UL55 and UL56, when 
compared to HSV-1 strain 17, a reference strain of HSV-1.15  Additionally, HF10 
has amino acid changes in the sequence of the protein encoded by UL1, which is 
involved in the regulation of syncytium formation and exhibits a relatively high 
divergence. The attenuation of neurovirulence in HF10 is likely attributable to the 
lack of the UL56 gene. Although the exact underlying mechanism of attenuation 
of neurovirulence is not clear, it has been reported that the lack of the UL56 gene 
decreases HSV-1 pathogenicity without affecting the ability of the virus to 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 19 of 61 
 
replicate in most types of cultured cells.16 UL56 belongs to the tail-anchored type 
II membrane protein that can associate with the kinesin motor protein KIF1A, 
which is involved with axonal transport.17,18 Some defective mutations have also 
been detected in the other three genes. A 3,832 bp deletion at the UL and UL/IRL 
junction and a 2295 bp deletion and extensive rearrangement at the left end of the 
genome are also observed. Figure 1 shows the major differences in structure 
between HF10 and wild type HSV-1.  
 
Figure 1. 
 
 Nivolumab 
Immune checkpoint blockade is a rapidly advancing therapeutic approach in the 
field of immuno-oncology, and treatment with investigational agents targeting this 
mechanism has induced regressions in several types of cancer. Cytotoxic T-
lymphocyte–associated antigen 4 (CTLA-4) and programmed death 1 (PD-1) 
receptor are two important cellular targets that play complementary roles in 
regulating adaptive immunity. Whereas PD-1 contributes to T-cell exhaustion in 
peripheral tissues, CTLA-4 inhibits at earlier points in T-cell activation. 
Nivolumab is a fully human monoclonal immunoglobulin (Ig) G4 antibody that 
binds to the PD-1 cell surface membrane receptor, a negative regulatory molecule 
expressed by activated T and B lymphocytes. Inhibition of the interaction between 
PD-1 and its ligands promote immune responses and antigen-specific T cell 
responses to both foreign and self-antigens. PD-1 receptor blockade by nivolumab 
is a new approach for immunotherapy of tumors. Results from several clinical 
trials indicate that nivolumab is active in multiple tumor types. OPDIVO 
(nivolumab) is approved in the US for treatment of previously treated, 
unresectable or metastatic melanoma and previously treated, metastatic squamous 
non-small cell lung cancer, advanced renal cell carcinoma, classical Hodgkin 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 20 of 61 
 
lymphoma, and recurrent or metastatic squamous cell carcinoma of the head and 
neck. 
 
Nonclinical and Preclinical Studies 
 Nonclinical Pharmacology Studies  
In Vitro Studies  
In vitro studies using tumor and normal cell lines of human, mouse, and rat origin 
have shown that HF10 is capable of infection, replication, and cytotoxicity in a 
wide range of cell types. In vivo antitumor effects of HF10 were investigated 
using disseminated peritoneal tumor models and solid tumor models in 
immunocompetent mice. The mice treated with HF10 survived longer than 
control mice. Moreover, HF10-treated surviving mice that received a subsequent 
inoculation of the same tumor cell line survived longer than control mice, 
suggesting that HF10 contributed to acquisition of immunity against the tumors. 
Mechanisms of Action Studies  
As an attenuated, but replication-competent strain of HSV-1, HF10 has been 
shown to be capable of infecting a variety of mouse and human tumor cell lines. 
The virus replicates well in infected cells resulting in cell lysis. It was previously 
observed that the replication of HF10, unlike wildtype HSV-1 such as strain KOS 
and SP23, is highly restricted in murine macrophage RAW264 cells. Importantly, 
addition of anti-IFN α/β antibody results in complete lytic destruction of 
RAW264 cells.19 These results indicate that differences in the IFN-pathway 
activation between normal cells and cancer cells are involved in the selectivity of 
HF10. It is well known that most cancer cells have a defective IFN pathway and 
thereby exhibit no or little response to IFN. The replication of HF10 was not 
inhibited at a concentration of 1-100 units/mL of IFN-β in a variety of tumor cells 
tested. In contrast, normal skin fibroblast cells were drastically protected by 
relatively low concentrations of IFN-β. These results indicate that HF10 infection 
may induce IFN in vivo and protect normal tissues from virus lysis, whereas 
tumor cells are selectively killed due to the lack of IFN responsiveness. 
 In Vivo Studies – In Support of HF10 Single Agent Antitumor Activity 
In Vivo Efficacy of Repeat Intratumoral Administration of HF10 under 
Multiple Dose Levels against Mouse Melanoma in Immunocompetent DBA/2 
Mice 
The efficacy of repeated injections of HF10 at multiple dose levels against 
melanoma was evaluated. DBA/2 mice were subcutaneously inoculated with 
Clone M-3 melanoma cells (1 x 106 cells) and injected with 1 x 107, 1 x 106, or 1 
x 105 TCID50 /animal of HF10 on Days 5, 7, 9, 12, 14 and 16 after tumor 
inoculation. Results indicate that repeat intratumoral administration of multiple 
dose levels of HF10 in immunocompetent DBA/2 mice with the Clone M-3 
melanoma cells resulted in significantly smaller tumor volumes compared with 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 21 of 61 
 
control mice. Every HF10 treatment group showed statistically significant 
survival compared to that of vehicle treatment group. It is notable that the tumor 
mass completely disappeared in 7 of 10 mice in the HF10 group (1 x 107 TCID50), 
1 of 10 mice in the HF10 group (1 x 106 TCID50) and 5 of 10 mice in the HF10 
group (1 x 105 TCID50).  
In Vivo Antitumor Activity of HF10 against Mouse Cancer in 
Immunocompetent C3H and BALB/c Mice  
Nagoya University in Japan previously investigated the in vivo antitumor effects 
of single and repeated intratumoral, intraperitoneal, and intravesical HF10 
injections in immunocompetent mice with disseminated peritoneal tumors or solid 
tumors including: NfSa sarcoma 20,21, Colon-26 colorectal 22, MM102-TC breast 
23 and MBT-2 bladder 24. The results showed that mice treated with HF10 
survived longer and had smaller tumor volumes than control mice. Moreover, 
when surviving HF10-treated mice were inoculated again with the same tumor 
cell lines, they survived longer than control mice, suggesting that infection with 
HF10 may have resulted in acquisition of antitumor immunity in the HF10-treated 
mice. 
 In Vivo Studies – in Support of Combination Treatment with HF10 + 
Nivolumab Antitumor Activity 
The safety and efficacy of HF10 in combination with anti-PD-1 therapy has been 
evaluated in a preclinical Mouse M3 melanoma model. The M3 mouse melanoma 
cell line has a high sensitivity to HF10 and a moderate sensitivity to mouse anti-
PD-1 therapy. Tumor cells were inoculated to both flanks of mice to examine a 
systemic anti-tumor effect of the combination treatment. Seven days after tumor 
inoculation, HF10 (1 x 105 TCID50) was then intratumorally injected into a tumor 
on the left side. On day 8 and 11, 250 μg anti-mouse PD-1 antibody was 
administered intraperitoneally. Efficacy and survival was compared in 4 groups: 
controls, HF10 only, anti-mouse PD-1 only, and combination HF10 and anti-
mouse PD-1. All treatments had a significant reduction in tumor volume 
compared to controls. The greatest reduction in tumor volume was seen with the 
combination of HF10 and anti-mouse PD-1. Additionally, the combination 
therapy of HF10 and anti-mouse PD-1 antibody exerted an enhancement of the 
anti-tumor effect not only in the HF10-treated side but also in the non-treated 
side. Survival was also significantly improved in mice treated with combination 
compared to both control and monotherapy treated animals.  
In a study that investigated the effect of CD4+ T or CD8+ T cells the anti-tumor 
effect of the combination therapy was completely abrogated in the CD8+ depleted 
mice, while a trend towards an additive anti-tumor effect was observed in CD4+T 
cell depleted mice. These results were supported by the survival results. Taken 
together, it suggests that CD8+T cells are necessary for the anti-tumor effect in 
the combination therapy. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 22 of 61 
 
 Nonclinical Toxicology Studies of HF10 
In Vivo Toxicity Studies  
A comprehensive battery of toxicity and biodistribution studies (intratumoral, s.c., 
i.p., and i.v.) were performed in both normal and tumor-bearing mice aimed at 
characterizing the HF10 tissue distribution and defining a NOAEL in mice. 
A single intratumoral dose study demonstrated that HF10 was overall well 
tolerated (1.51 x 108, 6.76 x 104, and 5.89 x 103 TCID50/mouse), and promoted a 
dose-related extension of tumor cell death in mice with M3 melanoma tumors. 
Although some adverse pathology was noted in the high-dose (1.51 x 108 
TCID50/mouse) treated mice in ovaries, adrenal gland, and bone marrow, these 
findings did not extend to the lower dose levels of the study, and thus intratumoral 
administration of 5.89 x 103 TCID50/mouse and 6.76 x 104 TCID50/mouse were 
well tolerated. A NOAEL was established for HF10 at a dose of 6.76 x 104 
TCID50/mouse for intratumoral administration, the same route of the 
administration used in the clinical studies. This dose level equates to a level of 
1.89 x 108 TCID50/70 kg patient based on body weight. Therefore, the HF10 dose 
at 1 x 107 TCID50/mL up to 5 mL planned for the administration in the Phase II 
trial, is below the NOAEL established in mice for intratumoral administration. 
A single s.c. dose study in normal mice demonstrated that HF10 of up to 1.51 x 
108 TCID50 was well tolerated. Minor degenerative and/or inflammatory changes 
occurred at the injection site on Days 3 and 15, however these changes were 
recovered thereafter. On Day 3, most of the treated mice showed extramedullary 
hematopoiesis in the spleen and increased cellularity of the paracortex of axillary 
lymph nodes. These findings in the spleen and lymph node are considered to be 
secondary to the findings at the injection site. There was no treatment-related 
histopathology in any tissue on Day 30, thus demonstrating complete recovery. 
In the i.p. toxicity studies, adrenal necrosis was seen in mice given highest dose of 
HF10 (1.51 x 108 TCID50/mouse). Body weight losses at all doses tested (5.89 x 
103, 4.79 x105 and 1.51 x 108 TCID50/mouse) were recoverable by 14 days post 
dose. Changes in hematology and blood chemistry composition reflected the 
expected response to viral infection and were considered to be non-adverse. The 
NOAEL was considered to be 4.79 x 105 TCID50/mouse. 
In the i.v. toxicity studies representing a worst-case scenario of complete systemic 
exposure to the virus, mice dosed with the high level of HF10 (1.51 x 108 
TCID50/mouse) showed a number of degenerative and/or inflammatory changes in 
the adrenals, pituitary, ovaries, uterus, spleen, and bone marrow. 
At middle dose levels of HF10 (6.76 x 104 and 5.89 x 103 TCID50/mouse), 
mortality and adverse treatment-related effects were observed. In the toxicity 
study, one female receiving 6.76 x 104 TCID50 was sacrificed and presented with 
significant pathology in the gastrointestinal tract, adrenal gland, and liver. 
Unscheduled deaths also occurred in the biodistribution study between Day 8 and 
10 and considered to be related to treatment. Marked levels of HF10 DNA were 
detected in those pathologically affected organs (e.g., adrenals, liver) and ovaries. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 23 of 61 
 
At lower dose levels of 15.1 and 63.1 TCID50/mouse, HF10 did not elicit any 
adverse effect in any of the parameters investigated. There was no mortality, 
clinical signs or any adverse pathology or distribution associated with HF10 
administration. Thus, a NOAEL was established for HF10 at a dose of 63.1 
TCID50. A dose of 63.1 TCID50/mouse equates to a dose of nearly 2.5 x 103 
TCID50/kg based on body weight, or 1.77 x 105 TCID50/70 kg patient. 
It should be noted that HF10 is susceptible to acyclovir (ACV) to a similar extent 
as wild-type HSV-1 in vitro, and therefore anti-herpes drugs can be used as rescue 
therapy when the virus needs to be quickly controlled. 
 Nonclinical Toxicology Studies of Nivolumab 
Programmed death receptor-1 (PD-1, CD279), a 55 kD type I transmembrane 
protein, is 
a member of the CD28 family of T-cell costimulary receptors that also includes 
CD28, CTLA-4, ICOS, and BTLA.5 PD-1 contains an intracellular membrane 
proximal immunoreceptor tyrosine inhibitory motif (ITIM) and a membrane distal 
immunoreceptor tyrosine-based switch motif (ITSM). Two ligands specific for 
PD-1 have been identified: PD-L1 (B7-H1/CD274) and PD-L2 (B7-DC/CD273). 
PD-L1 and PD-L2 have been shown to down-regulate T-cell activation upon 
binding to PD-1 in both murine and human systems.25,26 PD-1 delivers a negative 
signal by the recruitment of a protein tyrosine phosphatase SHP-2 to the 
phosphorylated tyrosine residue in the ITSM in its cytoplasmic region.27,28 PD-1 
is primarily expressed on activated T cells, B cells and myeloid cells.29 
Further evidence for a negative regulatory role of PD-1 comes from studies of 
PD-1-deficient mice. PD-1-deficient mice develop various autoimmune 
phenotypes, including dilated cardiomyopathy, a lupus-like syndrome with 
arthritis and nephritis, and accelerated diabetes mellitus.30-32 The emergence of 
these autoimmune phenotypes is dependent upon the genetic background of the 
mouse strain and many of these phenotypes emerge at different times and show 
variable penetrance. In addition to the phenotypes of null mutations, PD-1 
inhibition by antibody-mediated blockade in several murine models has been 
found to play a role in the development of autoimmune diseases such as 
encephalomyelitis, graft-versus-host disease, and type I diabetes.33,34 Taken 
together, these results suggest that PD-1 blockade has the potential to activate 
anti-self T-cell responses, but these responses are variable and dependent upon 
various host genetic factors. Thus, PD-1 deficiency or inhibition is not 
accompanied by a universal loss of tolerance to self-antigens. 
 
Preclinical animal models of tumors have shown that blockade by PD-1 by 
monoclonal 
antibodies (mAbs) can enhance the anti-tumor immune response and result in 
tumor rejection. Antitumor activity by PD-1 blockade functions in PD-L1+ 
tumors as well as in tumors that are negative for the expression of PD-L1.33,35-39 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 24 of 61 
 
This suggests that host mechanisms (i.e., expression of PD-L1 in antigen-
presenting cells) limit the antitumor response. Consequently, both PD-L1 positive 
and negative tumors may be targeted using this approach. In humans, constitutive 
PD-L1 expression is normally limited to macrophage-lineage cells, although 
expression of PD-L1 can be induced on other hematologic cells as well, including 
activated T cells. However, aberrant expression of PD-L1 by tumor cells has been 
reported in a number of human malignancies.40-46 PD-L1 expressed by tumor cells 
has been shown to enhance apoptosis of activated tumor-specific T cells in vitro.30 
Moreover, the expression of PD-L1 may protect the tumor cells from the 
induction of apoptosis by effector T cells.47 Retrospective analyses of several 
human tumor types suggest that tumor over-expression (as measured by 
immunohistochemistry (IHC)) of PD-L1 may permit immune evasion by tumors. 
In renal cell carcinoma, high surface expression levels of PD-L1 on tumor cells 
are related to tumor aggressiveness.42,45 Subjects with high tumor and/or 
lymphocyte PD-L1 levels are 4.5 times more likely to die from their cancer than 
subjects exhibiting low levels of PD-L1 expression. In addition, in multivariate 
analysis, high expression of PD-L1 is correlated to have a worse overall survival 
rate compared to low expression levels of PD-L1.48 
Nivolumab is a fully human, IgG4 (kappa) isotype, mAb that binds PD-1. 
Blockade of the PD-1 pathway by nivolumab was studied using the mixed 
lymphocyte reaction (MLR). PD-1 blockade resulted in a reproducible 
enhancement of both proliferation and IFN- 49 The effect of nivolumab on 
antigen-specific recall response was investigated using a cytomegalovirus (CMV) 
-restimulation assay with human peripheral blood mononuclear cells (PBMCs), 
and was evaluated by an enzyme-linked immunosorbent assay (ELISA). 
These data indicated that nivolumab, versus an isotype-matched control antibody, 
augmented IFN-. Nivolumab is therefore considered a promising 
immunotherapeutic option. 
 
Clinical Studies with HF10 
Completed Phase I Studies of HF10 in Japan  
HF10 was studied previously in patients with breast cancer (N=6)50, head and neck 
cancer (N=3)51, and unresectable pancreatic cancer (N=8)52 in three investigator-
initiated open-label studies conducted in Japan. The only adverse event reported was 
low-grade fever in two of the three treated patients with head and neck cancer. This 
adverse event is consistent with adverse events reported previously in other clinical 
studies of oncolytic viruses (e.g. flu-like symptoms such as low grade fever, chills, 
fatigue, nausea/vomiting, headache, diarrhea, and hypotension).53 
Phase I Study of HF10 in Patients with Refractory Head and Neck Cancer or 
Solid Tumors with Cutaneous and/or Superficial Lesion in the US (Protocol No. 
M06-10083, under BB-IND 13342) 
This was an open label, non-randomized, multicenter, two-stage, dose escalation 
Phase I study evaluating single and repeated intratumoral injections of the oncolytic 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 25 of 61 
 
virus, HF10, in patients with refractory head and neck cancer, or solid tumors with 
cutaneous and/or superficial lesions (e.g., squamous cell carcinoma of the skin, 
carcinoma of the breast, and malignant melanoma). 
Stage 1 of the study investigated dose escalation of a single intratumoral injection of 
HF10 over the following dose levels: 1 x 105 TCID50, 3 x 105 TCID50, 1 x 106 TCID50, 
and 1 x 107 TCID50. In Stage 1, 3 patients were to be enrolled per single dose cohort. 
Within each single dose cohort, accrual would temporarily be suspended after the first 
patient was entered and the patient was followed for safety and for viral distribution 
and elimination. In the single dose cohorts, after the first patient in a cohort showed 
no significant increase of virus DNA in comparison to baseline in two consecutive 
weekly samples of each specimen (whole blood, saliva, and urine), safety 
considerations permitting, entry of two additional patients would occur concurrently. 
Escalation to the next dose level required confirmation of no significant change of 
virus DNA, as described for the first patient in each cohort. All patients in a cohort 
completed the first cycle of treatment (with a 28-day safety observation period), 
before escalation to the next cohort was initiated. The patients in Stage 1 were 
required to be seropositive for HSV-1. Stage 1 was completed. 
Stage 2 evaluated repeated intratumoral injections of HF10 at dose levels of 1 x 106 
TCID50/dose and 1 x 107 TCID50/dose. Three patients were enrolled in each of the 
repeated dose cohorts. 
In Stage 2, the first patient treated in each repeated dose cohort was required to be 
seropositive for HSV-1. Then, if no safety issue had occurred as a result of HF10 
treatment of the first patient in a cohort, the cohort was to be opened to enrollment of 
HSV-1 seropositive and seronegative patients. Accrual on the cohort was temporarily 
suspended for seronegative patients only, i.e., after enrollment of the first 
seronegative patient in a cohort, a second seronegative patient could not be enrolled 
in the cohort until after the first seronegative patient completed the safety observation 
period for 2 cycles of treatment. If all of three patients were seropositive, one 
seronegative patient could additionally be enrolled. 
Twenty-eight patients were enrolled in the study and a total of 26 patients were 
treated in the study (15 patients in Stage 1 and 11 patients in Stage 2). Two patients 
were not evaluable for safety because they were not treated with HF10; one withdrew 
consent before HF10 treatment and one was not treated with HF10 due to 
hospitalization for pneumonia. Another patient, enrolled in Stage 2 (repeated HF10 
administration portion of the study), was not evaluable for dose limiting toxicities 
(DLTs) per protocol because the patient received only one HF10 injection; Stage 2 
patients were required to receive two HF10 injections to be considered evaluable for 
DLTs. 
Twenty-four (92.3%) of the 26 patients treated in this study experienced adverse 
events. 
Treatment-emergent adverse events (TEAEs) were reported for ≥ 20% of patients in 
the following system organ classes, in descending order: general disorders and 
administration site condition (13 patients; 50.0%), gastrointestinal disorders (8; 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 26 of 61 
 
30.8%), and metabolism and nutrition disorders (8; 30.8%). The most frequently 
reported TEAEs were chills, fatigue, and nausea, each reported for 3 patients 
(11.5%). 
Nine patients (34.6%) reported HF10-related TEAEs. The most frequent related 
TEAEs were chills, experienced by 3 patients (11.5%), and fatigue, which was 
reported for 2 patients (7.7%). Regarding the repeat dosing cohorts, fewer patients 
(33.3%) in the lower dose cohort (1 x 106 TCID50/mL/dose) experienced HF10-
related TEAEs compared to patients (50.0%) in the 1 x 107 TCID50/mL/dose cohort, 
suggesting a dose-response. It is notable that the frequent HF10-related TEAEs (chills 
and fatigue) were generally mild and are symptoms reported commonly and easily 
managed in viral infections and treatment with other oncolytic viruses.53  All other 
HF10-related TEAEs were each reported for one patient. Overall, 8 patients (30.8%) 
reported severe (≥ Grade 3) TEAEs. No patients reported an event that was both 
HF10-related and Grade 3 or greater in severity. No HF10-related allergic events were 
reported. Also, there were no differences in toxicity between patients who were HSV-
1-positive and HSV-1-negative at enrollment. No HF10-related serious adverse 
events were reported in this study. 
Viral clearance was investigated following intratumoral injection of HF10. Viral 
clearance was assessed in blood, saliva, and urine samples by quantitative polymerase 
chain reaction (qPCR). Patients showed rapid clearance of virus in blood, saliva, and 
urine. In both stages of the study, HF10 DNA was not detected in any of the 
biological samples in the majority of patients. For the few patients in whom HF10 
DNA was detected, it was almost exclusively restricted to saliva samples and was 
only transiently detected in samples. Two patients in Stage 1 had transient virus 
detection in saliva only. In Stage 2, 6 patients had transient virus detection in saliva 
only. Two patients in Stage 2 had transient virus detection in blood, and one of these 
patients also had transient virus detection in urine. 
All 26 patients are off-study. The reasons for off-study are as follows: thirteen 
(50.0%) patients total, including 7 (46.7%) patients in Stage 1 and 6 (54.5%) patients 
in Stage 2, were off-study due to progressive disease per RECIST evaluation. Ten 
(38.5%) patients completed the study per protocol (including 5 patients (33.3%) in 
Stage 1 and 5 patients (45.5%) in Stage 2 [3 patients in the 1 x 106 TCID50/mL cohort 
and 2 in the 1 x 107 TCID50/mL cohort]). Two (7.7%) patients were off-study due to 
unrelated death; both patients were treated in Stage 1. One (3.8%) patient in Stage 1 
was removed from the study due to symptomatic deterioration. 
Clinical Studies with HF10 + Ipilimumab in unresectable melanoma 
Phase 2 multicenter trial to evaluate efficacy and safety of HF10 oncolytic virus 
immunotherapy and ipilimumab in patients with unresectable stage IIIB-IV 
melanoma in the US (Protocol T14-10682, IND No 13,342) 
An ongoing phase 2 trial of HF10 and ipilimumab in unresectable metastatic 
melanoma patients is assessing whether the antitumor effect of HF10 is enhanced by 
concurrent ipilimumab treatment. Ipilimumab naïve adults with Stage IIIB, IIIC or IV 
unresectable melanoma with measurable non-visceral lesion(s) received HF10 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 27 of 61 
 
injections into single or multiple tumors (1x107 TCID50/mL, up to 5mL/dose); 4 
injections every week; then up to 15 injections every 3 weeks. Ipilimumab infusions 
(3 mg/kg) were given every 3 weeks for 4 doses. Tumor responses were assessed at 
12, 18, 24, 36, and 48 wks. Best Overall Response Rate (BORR) was determined at 
24 wks. Of 46 pts treated, 20% were stage IIIB, 43% stage IIIC, and 37% stage IV. 
Most HF10-related AEs were ≤ Grade 2, similar to HF10 monotherapy. No DLTs 
were reported; 3 Grade 4 AEs reported, all not treatment related. 30.4% had Grade 3 
AEs. HF10-related Grade 3 AEs (n=3) were left groin pain, thromboembolic event 
and lymphedema, hypoglycemia, and diarrhea. Twenty-one (46%) patients had 
received at least 1 prior therapy for metastatic melanoma. Of 43 efficacy evaluable 
patients, preliminary BORR at 24 weeks per irRC was 41.8% (11.6% CR, 30.2% PR), 
disease stability rate 67.4% (25.6% SD). Eight responders (53%) were Stage IV. 
Overall study BORR, including those after 24 weeks, by irRC was 47.7% (15.9% CR, 
31.8% PR), disease stability was 65.9% (18.2% SD). In summary, HF10 in 
combination with ipilimumab treatment does not appear to exacerbate ipilimumab 
toxicity, is safe, well tolerated and has both local and systemic antitumor activity. The 
efficacy appears greater than that seen with ipilimumab monotherapy. Since the 
clinical trial is on-going, the final results of the trial are still pending. 
Clinical Studies with Nivolumab  
One Phase 1 study (CA209003) has contributed much to the clinical experience with 
nivolumab monotherapy in subjects with melanoma and other solid malignancies.14 
CA209003 is an ongoing Phase 1 open label, multiple dose escalation study in 304 
subjects with select previously treated advanced solid tumors, including melanoma, 
renal cell carcinoma (RCC), non-small cell lung cancer (NSCLC), colorectal cancer, 
and hormone-refractory prostate cancer. Subjects received nivolumab at doses of 0.1, 
0.3, 1, 3 or 10 mg/kg intravenously every 2 weeks, up to a maximum of 2 years of 
total therapy. As of 03-Jul-2012, a total of 107 melanoma subjects were treated with 
nivolumab in the dose range of 0.1 - 10 mg/kg. 
No maximal tolerated dose was identified in CA209003. The incidence, severity and 
relationship of AEs were generally similar across dose levels and tumor types. 
Nivolumab related AEs of any grade occurred in 72.4% of subjects. The most 
frequent nivolumab related AEs occurring in 5% of subjects included: fatigue 
(25.7%), rash (13.5%), diarrhea (11.8%), pruritus (10.2%), nausea (7.9%), decreased 
appetite (7.9%), hemoglobin decreased (5.9%) and pyrexia (5.3%). The majority of 
events were low grade, with Grade 3 - 4 drug related AEs observed in 14.8% of 
subjects. The most common Grade 3 - 4 drug-related AEs occurring in 1% of subjects 
were: fatigue (1.6%), lymphopenia (1.3%), abdominal pain (1%), diarrhea (1%), 
hypophosphatemia. 
3 
POTENTIAL RISKS AND BENEFITS  
 
HF10 
Potential risks of investigational intratumoral injections include the following 
reported adverse events: 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 28 of 61 
 
Occurring in more than 10% of subjects – chills 
Occurring in 1%-10% of subjects - injection site disorders, fatigue, malaise, pyrexia, 
nausea, dehydration, pruritus, and/or hypotension 
One patient with anal and scrotal tumors injected with HF10 experienced swelling in 
the genital area, skin breakage and scab formation in the area on/near the scrotum. 
The results of previous and ongoing clinical trials of HF10 in Japan, and the Phase I 
clinical trial of HF10 conducted under BB-IND 13,342, suggest that HF10 is safe and 
tolerable. Few HF10- related adverse events have been reported in any of the trials 
and are similar across the trials. They include: flu-like symptoms such as fever, chills, 
fatigue, nausea/vomiting, and hypotension. Such events are consistent both with 
symptoms of viral infections and adverse events reported in trials of other 
investigational oncolytic viruses. No HF10-related serious adverse events have been 
reported in these trials. 
In the event that HF10-treated patients experience unexpected viral toxicity, patients 
may be treated by the anti-herpes drugs acyclovir, famciclovir, or valacyclovir, to 
which HF10 is sensitive. 
The Investigator Brochure may be updated during the course of this study with 
additional risks and benefits. Please see the current Investigator Brochure for further 
details about the potential risks and benefits associated with this study. 
Treatment with HF10 may offer potential benefits. Preliminary evidence suggests that 
intratumoral injection with HF10 may result in an antitumor response. 
 
Nivolumab  
Overall, the safety profile of nivolumab monotherapy is manageable and generally 
consistent across completed and ongoing clinical trials with no MTD reached at any 
dose tested up to 10 mg/kg. There was no pattern in the incidence, severity, or 
causality of AEs to nivolumab dose level. Most AEs were low-grade (Grade 1-2) with 
relatively few drug-related high-grade (Grade 3-4) AEs. The most common adverse 
events (>20%) in patients with melanoma were fatigue, rash, musculoskeletal pain, 
pruritus, diarrhea and nausea. The safety profile of nivolumab combination therapy 
varies with the agent combined with nivolumab, but is generally consistent with the 
safety profiles observed with either agent alone and, in some cases, both frequency 
and severity of AEs were greater than that observed with either agent alone.  
The Investigator Brochure may be updated during the course of this study with 
additional risks and benefits. Please see the current Investigator Brochure for further 
details about the potential risks and benefits associated with this study. 
Treatment with nivolumab offers patients potential benefits. In a significant 
proportion of nivolumab-treated patients, durable tumor responses were achieved, as 
well as a significant increase in overall survival. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 29 of 61 
 
4 
STUDY DESIGN 
 
Description 
This is a single-arm, open label, Phase II study evaluating the safety and efficacy of 
neoadjuvant Nivolumab and HF10 in resectable stage IIIB, IIIC, and IVM1a 
melanoma.  
 
Number of Patients 
 The total number of subjects for all sites is initially planned as 20 patients. The 
primary endpoint of pathologic response rate, including pCR, is expected to be seen 
in 20% of patients, hence 4 of the planned 20 patients are expected to have a 
pathologic response including pCR. Only patients who receive the combination neo-
adjuvant treatment, nivolumab and HF10, and have a surgical resection according to 
the study calendar will be considered evaluable. Patients who discontinue the study 
before surgery then will be considered non-evaluable and will be replaced per 
principal investigator discretion.  
 
Number of Study Centers 
 This study will open at the Huntsman Cancer Institute. If additional recruitment is 
required this will become a multi-center trial.  
 
Study Duration  
1 year and 3 months for enrollment, 3 years for study completion. 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 30 of 61 
 
5 
ELIGIBILITY CRITERIA 
 
This eligibility checklist is used to determine patient eligibility and filed with the 
enrolling investigators signature in the patient research chart.  
 
Patient No. ______________________ 
Patient’s Initials: (L,F,M) _________________ 
 
Inclusion Criteria 
Yes/No (Response of “no” = patient ineligible) 
 _____ Patients must be >18 years or older. 
 _____ Patients must have stage IIIB, IIIC, or IVM1a (equivalent staging at time 
of enrollment via AJCC 7th edition) metastatic melanoma which is eligible for 
complete surgical resection. 
 _____ Prior systemic, regional and radiation anticancer therapies must have been 
completed at least three months prior to enrollment. Prior therapies (including 
anti-PD-1 inhibitors) are allowed provided three months have elapsed from last 
dose. 
 _____ Patients must be a candidate for intralesional therapy. 
 At least 1 injectable cutaneous, subcutaneous, or nodal melanoma lesion > 10 
mm in longest diameter 
OR 
 Multiple injectable melanoma lesions which in aggregate have a longest 
diameter of > 10 mm 
AND 
 Must have no known bleeding diathesis or coagulopathy that would make 
intratumoral injection unsafe. 
 _____ Patients must have an Eastern Cooperative Oncology Group (ECOG) 
performance status of 0 or 1. 
 _____ Serum LDH level < 1.5 upper limit of normal (ULN) within 28 days prior 
to enrollment. 
 _____ Patients have adequate organ function within 28 days prior to enrollment, 
as defined in table 1 listed below.  
Table 1: Adequate Organ Function Laboratory Values: 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 31 of 61 
 
System 
Laboratory Value 
 Hematological:  
 
Absolute Neutrophil Count ANC > 1,500/uL 
Platelets > 100,000/uL 
Hemoglobin > 9 g/dL 
 Renal 
 
Serum Creatinine OR measured 
calculated creatinine clearance 
(GFR may also be used in place 
of creatinine or CrCl)  
< 1.5 upper limit of normal 
(ULN) OR  
> 60 mL/min for subject 
with creatinine levels > 1.5 
X institutional ULN 
 Hepatic 
 
 
Serum total bilirubin < 1.5 X ULN OR  
Direct bilirubin < ULN for 
subjects with total bilirubin 
> 1.5 ULN 
 
AST or ALT  < 2.5 X ULN OR 
< 5 X ULN for subjects 
with liver metastases 
 _____ Men and women of childbearing potential must agree to use adequate 
contraception from the time of consent through 7 months after final nivolumab 
study treatment. 
 _____ Females of childbearing potential must have a negative urine or serum 
pregnancy test within 1 week prior to the start of treatment. 
 _____ Patients must be able to provide informed consent and willing to sign an 
approved consent form that conforms to federal and institutional guidelines. 
 
Exclusion Criteria 
Yes/No (Response of “yes” = patient ineligible) 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 32 of 61 
 
 _____ Patients with active visceral, central nervous system, or any bone 
metastases melanoma (Stage IVM1b or IVM1c). 
 _____ Patients whose primary diagnosis was ocular melanoma 
 _____ Patients receiving anti-herpes medication (i.e., acyclovir, famciclovir, or 
valacyclovir) within 1 week prior to initiating HF10 treatment. Patients may not 
require intermittent or chronic systemic (intravenous or oral) treatment with an 
antiherpetic drug other than intermittent topical use. 
 _____ Patients who have an active herpetic skin lesion(s) or prior complications 
of HSV-1 infection. 
 _____ Uncontrolled intercurrent illness including, but not limited to, ongoing or 
active infection, symptomatic congestive heart failure, unstable angina pectoris, 
cardiac arrhythmia, or psychiatric illness/social situations that would limit 
compliance with study requirements, as determined by the investigator. 
 _____ Medical history of autoimmune disease (e.g. Crohn’s disease, ulcerative 
colitis) or other disease requiring systemic glucocorticoid or immunosuppressive 
therapy. Subjects who receive daily steroid replacement therapy serve as an 
exception to this rule. Daily prednisone equivalent at doses up to 10 mg would 
qualify. 
 _____ Patients with clinically evident Human Immunodeficiency Virus (HIV), 
Hepatitis B Virus (HBV), Hepatitis C Virus (HCV), or Epstein-Barr Virus (EBV) 
infection are excluded 
 _____ Pregnant or breast feeding women; women desiring to become pregnant 
within the timeframe of the study are also excluded. 
 
I certify that this patient meets all inclusion and exclusion criteria for enrollment 
onto this study.  
______________________________ 
 
_________ 
_______ 
Investigator Signature  
 
 
 
Date  
Time 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 33 of 61 
 
6 
TREATMENT PLAN 
 
Administration Schedule 
The order in which the patient receives Nivolumab and HF10 is not specified and will 
be determined based on clinic scheduling.  
 Neo-adjuvant treatment 
Nivolumab 
Nivolumab 240 mg, IV will be administered over 60 minutes (±15 minutes) as 
tolerated by the patient, every 14 days starting on day 0. The rate of infusion may 
be increased to 30 minutes as tolerated. A total of 7 infusions will be administered 
prior to surgery. 
HF10 
HF10 1 x 107 TCID50/mL will be administered by intratumoral injection to a 
single or multiple eligible tumors. HF10 will be administered on days 0, 7, 14, 21, 
28, 42, 56, 70, and 84 for a total of 9 injections. The volume to be injected will be 
determined by lesion size (see table below for injection volume by tumor mass) 
with a maximum volume of 5 mL to be administered on a single treatment day. 
All eligible tumors except one will be treated with HF10 up to the maximum 
volume allowed. The untreated tumor will be used as an untreated control lesion.  
Lesion Size# 
Injection Volume†(Dose*) 
>5.0 cm 
≤ 5.0 mL (5x107 TCID50) 
>2.5 cm to 5.0 cm 
≤ 2.0 mL (2x107 TCID50) 
>1.5 cm to 2.5 cm 
≤ 1.0 mL (1x107 TCID50) 
> 0.5 cm to 1.5 cm 
≤ 0.5 mL (5x106 TCID50) 
≤ 0.5 cm 
0.1 mL (1x106 TCID50) 
#Per longer perpendicular diameter 
†If the dose level is decreased to 1x106 TCID50/mL, the recommended injection volumes stated in this table 
are not to be changed. 
*Administration of maximum volume at 1x107 TCID50/mL. 
 Surgery 
Surgery will be performed within 28 days of completing neo-adjuvant therapy with 
nivolumab and HF10. Patients who cannot proceed to surgery will not be evaluable 
and will be replaced. Patients may be allowed to remain on study treatment at the 
discretion of the principal investigator.  


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 34 of 61 
 
 Adjuvant treatment 
Nivolumab 
Adjuvant nivolumab should be initiated within 90 days after definitive surgery. 
Nivolumab will be administered at a flat dose of 480 mg IV every 28 days (every 
4 weeks) for one year. We have chosen the 480mg IV dose every 4 weeks in 
accordance with the application made by Bristol-Myers Squibb to the FDA on 
July 24, 2017. This dose will be similar to the current FDA approved dose of 240 
mg IV every 2 weeks with fewer infusions for the patient. 
 
Nivolumab  
 Dosage Form 
Nivolumab Infusion, 100 mg/10 mL (10 mg/mL) is a clear to opalescent, colorless 
to pale yellow liquid, which may contain light (few) particulates. The drug 
product is a sterile, non-pyrogenic, single-use, isotonic aqueous solution 
formulated at 10 mg/mL in sodium citrate, sodium chloride, mannitol, 
diethylenetriaminepentacetic acid (pentetic acid), and polysorbate 80 (Tween 80), 
pH 6.0 and includes an overfill to account for vial, needle, and syringe holdup. It 
is supplied in 10-cc Type I flint glass vials, stoppered with butyl rubber stoppers 
and sealed with aluminum seals. 
 Preparation and Administration 
Nivolumab is to be administered as an IV infusion through a 0.2-micron to 1.2-
micronpore size, low-protein binding (polyethersulfone membrane) in-line filter at 
the protocol specified doses and infusion times. It is not to be administered as an 
IV push or bolus injection. Nivolumab can be infused undiluted (10 mg/mL) or 
diluted with 0.9% Sodium Chloride Injection, USP or 5% Dextrose Injection, USP 
to protein concentrations as low as 0.35 mg/mL. When the dose is fixed (e.g., 240 
mg, 360 mg, or 480 mg flat dose), nivolumab can be infused undiluted or diluted 
so as not to exceed a total infusion volume of 120 mL. 
During drug product preparation and handling, vigorous mixing or shaking is to 
be avoided. Dilution and infusion of nivolumab will be conducted per the package 
insert for the adjuvant treatment of melanoma. Care must be taken to assure 
sterility of the prepared solution as the product does not contain any antimicrobial 
preservative or bacteriostatic agent. Nivolumab infusions are compatible with 
polyvinyl chloride (PVC) or polyolefin containers and infusion sets, and glass 
bottles. 
 Storage and Use Conditions  
The administration of nivolumab infusion must be completed within 24 hours of 
preparation. If not used immediately, the infusion solution may be stored under 
refrigeration conditions (2°C to 8°C, 36°F to 46°F) for up to 24 hours, and a 
maximum of 8 hours of the total 24 hours can be at room temperature (20oC to 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 35 of 61 
 
25oC, 68oF to 77oF) and room light. The maximum of 8-hour period under room 
temperature and room light conditions includes the product administration period. 
 
HF10 
 Preparation  
Preparations of the HF10 product will be provided in vials for injection in 
aqueous buffer comprised of 20 mmol/L Tris, pH 7.0-7.5, and 10% glycerol. 
HF10 will be stored at < -70ºC. The HF10 product will be diluted with 
physiological saline just before use to meet the target dose level. Specific 
instructions will be provided in the Pharmacy Manual. 
 
Prohibited Concomitant Medications  
No investigational agents other than the study therapy may be administered. No 
anticancer therapy other than the study therapy may be administered. Any medication 
that is considered necessary for the patient’s welfare, and that will not interfere with 
the study medication, may be given at the discretion of the Investigator.  
It is recommended that patients who routinely receive warfarin anticoagulant therapy 
should receive modified anticoagulant treatment with low-molecular-weight heparin 
(LMWH) during study treatment. Such patients will undergo continued monitoring of 
coagulation function by PT/INR for the duration of study treatment. 
Following are recommendations for providing alternative anticoagulant therapy in 
warfarin receiving patients during HF10 treatment. However, actual management of 
anticoagulant therapy in each patient will be at the discretion of the study Investigator 
and the practice of the treatment center and treating physician. 
Patients should be switched from warfarin to low-molecular-weight heparin (LMWH) 
at least 4 to 5 days prior to HF10 injection. 
LMWH treatment should be stopped 24 hours prior to HF10 injection. 
LMWH treatment should be reinitiated 24 hours after HF10 injection. 
Warfarin should be reinitiated 24 hours after HF10 injection in conjunction with 
LMWH treatment. 
LMWH treatment would then be stopped approximately 3 days later (according to 
INR monitoring), allowing warfarin approximately 3 days to achieve a therapeutic 
level of anticoagulation. 
Continued INR monitoring is necessary for the duration of study treatment.  
Patients should receive anti-viral medication (e.g., acyclovir, famciclovir, or 
valacyclovir) only in the event of viremia caused by HF10 treatment. 
 
Immunosuppressive agents and immunosuppressive disease of system corticosteroids 
are also prohibited. Inhaled or topical steroids, and adrenal replacement steroid doses 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 36 of 61 
 
> 10 mg daily prednisone equivalent, are permitted in the absence of autoimmune 
disease.  
 
Duration of Therapy 
Subjects must be withdrawn from the study treatment for the following reasons:  
    Subject withdraws consent from the study treatment and/or study procedures. 
A subject must be removed from the trial at his/her own request or at the request 
of his/her legally acceptable representative. At any time during the trial and 
without giving reasons, a subject may decline to participate further. The subject 
will not suffer any disadvantage as a result. 
    Adverse events incurred from intra-tumoral administration of the study agent 
including but not limited to grade 3 or greater persistent infection, delayed 
healing, necrosis or ulcer formation. 
    Subject is lost to follow-up. 
    Death. 
Subjects may be withdrawn from the study for the following reasons:  
 The subject is non-compliant with study drug, trial procedures, or both; 
including the use of anti-cancer therapy not prescribed by the study protocol. 
 If, in the investigator's opinion, continuation of the trial would be harmful to 
the subject's well-being. 
 The development of a second cancer.  
 Development of an intercurrent illness or situation which would, in the 
judgment of the investigator, significantly affect assessments of clinical status 
and trial endpoints. 
 Deterioration of ECOG performance status to 4. 
7 
TOXICITIES AND DOSEAGE MODIFICATION 
This study will utilize the CTCAE (NCI Common Terminology Criteria for Adverse 
Events) Version 4.0 for adverse event and serious adverse event reporting. A copy of the 
CTCAE Version 4.0 can be downloaded: (http://safetyprofiler-
ctep.nci.nih.gov/CTC/CTC.aspx). 
 
Dose Modifications 
 Nivolumab  
Recommendations for nivolumab dose modifications are listed in the table 1 below.  
 
Table 1: Recommended Dose Modification for Nivolumab  


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 37 of 61 
 
Adverse Reaction 
Severity* 
Dose Modification  
Colitis  
Grade 3 diarrhea or colitis 
Withhold dosea 
Grade 4 diarrhea or colitis 
Permanently discontinue 
Pneumonitis  
Grade 2 pneumonitis 
Withhold dosea 
Grade 3 or 4 pneumonitis 
Permanently discontinue 
Hepatitis  
Grade 2 AST or ALT or 
total bilirubin increased 
Withhold dosea 
Grade 3 or 4 AST or ALT 
or total bilirubin  
Permanently discontinue 
Hypophysitis  
Grade 2 or 3 hypophysitis 
Withhold dosea 
Grade 4 hypophysitis 
Permanently discontinue 
Adrenal Insufficiency 
Grade 2 adrenal 
insufficiency 
Withhold dosea 
Grade 3 or 4 adrenal 
insufficiency 
Permanently discontinue 
Type 1 Diabetes Mellitus 
Grade 3 hyperglycemia 
Withhold dosea 
Grade 4 hyperglycemia 
Permanently discontinue 
Nephritis and Renal 
Dysfunction 
Grade 2 or 3 Serum 
creatinine 
Withhold dosea 
Grade 4 Serum creatinine 
Permanently discontinue 
Skin  
Grade 3 rash or suspected 
Stevens-Johnson syndrome 
(SJS) or toxic epidermal 
necrolysis (TEN) 
Withhold dosea 
Grade 4 rash or confirmed 
SJSor TEN 
Permanently discontinue 
Encephalitis  
New-onset moderate or 
severe 
Withhold dosea 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 38 of 61 
 
neurologic signs or 
symptoms 
Immune-mediated 
encephalitis 
Permanently discontinue 
Other  
Other Grade 3 adverse 
reaction 
First occurrence 
Recurrence of same 
Grade 3 adverse reactions 
Withhold dosea 
Permanently discontinue 
Life-threatening or Grade 4 
adverse reaction 
Permanently discontinue 
Requirement for 10 mg per 
day or greater prednisone 
or equivalent for more than 
12 weeks 
Permanently discontinue 
Persistent Grade 2 or 3 
adverse reactions lasting 12 
weeks or longer 
Permanently discontinue 
* Toxicity was graded per National Cancer Institute Common Terminology Criteria for 
Adverse Events. Version 4.0 (NCI CTCAE v4). 
a Resume treatment when adverse reaction returns to Grade 0 or 1. 
If nivolumab is withheld for more than 12 weeks, the treatment will be permanently 
discontinued.  
In the event of a nivolumab dose delay, the concurrent HF10 dose will be similarly 
delayed; the HF10 dose will then be administered on the same day that nivolumab is 
administered.  
 HF10 
In the event that the patient is unable to tolerate HF10 (grade 2 intolerable or 
higher chills, injection site disorders, fatigue, malaise, pyrexia, nausea, 
dehydration, pruritus, and/or hypotension which do not respond to optimal 
management) at the discretion of the treating provider, the dose will be reduced to 
1 x 106 TCID50/mL.  The HF10 product will be diluted with physiological saline 
just before use to meet the target dose level. Specific instructions will be provided 
in the Pharmacy Manual. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 39 of 61 
 
 If the patient is unable to tolerate the reduced dose the patient will be withdrawn 
from treatment.  
 
Subject Compliance 
Both Nivolumab and HF10 will be dispensed from the Huntsman Cancer Institute 
Investigational Pharmacy and administered by a study investigator, thus ensuring 
compliance with therapy parameters.  
 
Patient Precautions for Minimizing Virus Exposure to Others 
The HSV-1 virus, from which the study drug HF10 is derived, is very widespread in 
the environment, and many people in the United States have been exposed to HSV-1. 
HF10 is a weakened strain of HSV-1 and is believed to pose minimal health risk. No 
association of HF10 with any human illness has been established. However, given the 
investigational nature of HF10 therapy, it is appropriate that HF10-treated patients 
follow some precautions for minimizing HF10 exposure to persons with whom they 
have close contact. Appendix I provides guidance for the patient for minimizing 
exposure of HF10 to others. 
 
Healthcare Provider Precautions 
HF10 is a replication-competent virus. Although it is believed that the risk of 
inadvertent infection to study staff is extremely low, infectious disease precautions 
should be implemented to minimize viral exposure to healthcare providers. In the 
event of an accidental spill of HF10 containing solution, standard biohazard waste 
procedures should be employed, including use of gloves, gown, and mask, by 
healthcare workers coming in contact with or disposing of the study drug product. 
Contaminated surfaces should be first washed with a 10% sodium hypochlorite 
solution, followed by rinsing with water, and finally with 70% ethanol. All tubing and 
other disposables that may have been in contact with HF10 must be discarded as 
biohazardous waste. 
In the event of an accidental needle stick, the healthcare provider should report the 
incident and follow the institutional inadvertent needle stick exposure policy 
implemented at the study site. In the event of inadvertent contact with HF10 (e.g., 
accidental spill, body fluid contact, or needle stick), exposed individuals may be 
treated, if warranted, with acyclovir, famciclovir, or valacyclovir. HF10 has been 
demonstrated to be sensitive to these anti-herpes agents. Treatment recommendations 
would follow those for acute wild type HSV infection. 
An additional precaution recommended in this clinical trial is that pregnant healthcare 
providers be excluded from participating in the administration of HF10 or the care of 
HF10-treated patients. Although preclinical data indicate that HF10 does not undergo 
germline genomic integration, potential risks of HF10 to the developing human fetus 
have not been determined. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 40 of 61 
 
 
Supportive Care 
All supportive measures consistent with optimal patient care will be given throughout 
the study. 
Management of immune mediated events (irAEs) will follow the package inserts for 
the Nivolumab products. 
 


 
Protocol name: Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
 
8 
STUDY CALENDAR 
Examination 
Pre-
study1 
D0 
D7±4 
and 
D21±4  
D14±4 
and 
D28±4 
D42±4  
D56±4 
and 
D70±4 
D84±4 
Surgery8 
Every 4 wks ±2 
wks for one year9 
EOT10 or 
Progression 
Follow-
up11 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
 
Medical history 
X 
 
 
 
 
 
 
 
 
 
 
Concomitant Medications 
X 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
Eligibility criteria 
X 
 
 
 
 
 
 
 
 
 
 
Vital signs3 
X 
X 
X 
X 
X 
X 
X 
 
X 
X 
 
Height  
X 
 
 
 
 
 
 
 
 
 
 
Weight  
X 
X 
 
X 
X 
X 
X 
 
X 
X 
 
Physical examination 
X 
X2 
X 
X 
X 
X 
X 
 
X 
X 
 
Lesion Photography18 
X 
 
 
X 
X 
 
X 
 
 
 
 
CBC/differential platelet count 
X 
X2 
X 
X 
X 
X 
X 
 
X 
X 
 
Chemistry4 
X 
X2 
X 
X 
X 
X 
X 
 
X 
X 
 
Pregnancy Test (serum or urine) 
X15 
X2 
 
 
 
 
X 
 
X15 
 
 
Thyroid function tests17 
X 
 
X 
 
X 
 
X 
 
 
 
 
PT/INR in patients receiving warfarin 
anticoagulation therapy 
X 
X 
X 
X 
X 
X 
X 
 
 
 
 
LDH 
X 
 
 
 
 
 
X 
 
 
X 
 
Genomic testing16 
X 
 
 
 
 
 
 
 
 
 
 
Adverse Event Assessment7 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Radiologic Evaluation5 
X 
 
 
 
 
 
X 
 
X12 
X 
 
irRC19 
X 
 
 
 
 
 
X 
 
 
 
 
RECIST 1.119 
X 
 
 
 
 
 
X 
 
 
 
 
Biopsy for correlative studies6 
 
X 
 
 
X 
 
 
X 
 
X 
 
Blood for correlative studies14  
 
X 
X 
X 
X 
X 
X 
 
X 
 
 
Nivolumab  
 
X 
 
X 
X 
X 
X 
 
X 
 
 
HF10 injections 
 
X 
X 
X 
X 
X 
X 
 
 
 
 
Surgery 
 
 
 
 
 
 
 
X13 
 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 42 of 61 
 
Study visits during neoadjuvant nivolumab are scheduled in days. Windows are ± business days.  
1 ALL Pre-study/Screening procedures should be completed within 28 days of study enrollment  
2. Does not need to be done if occurred 7 days before D0.  
3. Includes temperature, blood pressure, and pulse rate.  
4. Chemistry includes: Albumin, Alkaline Phosphatase, Aspartate Aminotransferase (AST), Alanine Aminotransferase (ALT), Total Bilirubin, Calcium, 
Carbon Dioxide, Creatinine, Chloride, Glucose, Potassium, Protein, Sodium, Urea Nitrogen. Fasting Glucose will be tested as clinically indicated.  
5. A scan of the chest abdomen and pelvis with extension to the extremities if applicable. The same method (CT scans or possibly PET/CT scan) should be 
used for all assessments.  
6. Biopsies are mandatory at day 0 and surgery.  Biopsies are optional at day 42 and progression.  See section 13.1 for details about the collection. 
7. Adverse events will be followed until resolution while the patient remains on treatment. Events will be followed for 90 post treatment for SAEs and events 
thought to be related to the study medication will be followed until resolution or stabilization of the adverse event, or until the patient starts a new treatment 
regimen.  
8. Surgery should occur no later than 28 days after D84.  
9. Adjuvant nivolumab should be initiated within 90 days after definitive surgery. 
10. EOT visit should occur 28 days (+-14 days) after the completion of therapy. 
11. Follow-up should occur every three months (+-or 14 days) for 1 years post EOT visit.  
12. Surgery will reset the scan schedule and should occur approximately every 12 weeks after surgery as standard of care.  
13. Pathology at surgery will be used to determine pathological response.  
14. Collection of blood samples for correlatives is optional and will be collected prior to HF10 injection during the neo-adjuvant (days 0,7, 14, 21, 28, 42, 56, 
70, 84) and prior to nivolumab infusion during the adjuvant nivolumab administration.  
15. Men and women of childbearing potential must agree to use adequate contraception from the time of consent through 7 months after nivolumab study 
treatment (neoadjuvant and adjuvant). A pregnancy test will be conducted w/in 72 hours prior to beginning adjuvant nivolumab. 
16. Next generation sequcencing is required at screening as per standard of care. Testing will include, but is not limited to BRAF mutational analysis.  
17. Thyroid Tests will include TSH and free T4. In the event of hypo/hyper thyroidism, appropriate support will be provided in accordance with institutional 
guidelines.  
18. Lesion photography should be completed at each time point AND when a new lesion is identified. 
19. irRC and RECIST should be done prior to surgery. Only RECIST will be used as clinical response criteria and used to make treatment decisions


 
Protocol name: Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable 
Stage IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
 
 
 
9 
CRITERIA FOR EVALUATION AND ENDPOINT 
 
Pathological response 
The primary objective of this study will be to identify the pathologic response rate at 
surgery, including complete response, after 12 weeks of neoadjuvant treatment with 
nivolumab and HF10. Pathologic response rate will be defined based on the quantity of 
residual viable tumor in the surgical resection specimen at the time of definitive surgery. 
A percent viable tumor will be assessed semi-quantitatively in the pre treatment biopsy, 
on study biopsy at day 42 and the definitive surgical resection specimen by estimating the 
proportion of residual tumor in relation to the total tumor area and reported as percentage 
viability (Ribero et al 2007) as per institutional practice. A pathologic complete response 
will be defined as no viable residual melanoma cells in the surgical specimen. A major 
pathologic response will be defined as <50% viable tumor cells. A minor pathologic 
response will be defined as 50% or greater viable tumor cells, including specimens that 
have 100% viability at surgery. Pathologic response rate (pRR) will be defined as 
frequency of either pCR or a pathologic major response. This will be compared to the 
viablitiy values of the tumor at baseline. Biopsies will be required at Day 0 and at surgery 
to perform this analysis. A patient must complete the neoadjuvant phase of the trial and 
complete surgery to be considered evaluable for pathological response.  
 
Safety 
Routine safety and tolerability will be evaluated from the results of reported signs and 
symptoms, scheduled physical examinations, vital sign measurements, and clinical 
laboratory test results. More frequent safety evaluations may be performed if 
clinically indicated or at the discretion of the investigator.  
Physical Examination 
Complete and symptom-directed physical examinations will be performed by a 
licensed physician (or physician’s assistant or nurse practitioner). 
Vital Signs 
Vital signs will include (blood pressure, respiratory rate, pulse rate and temperature).  
Safety Laboratory Determinations 
Laboratory evaluations will be performed as noted in the study calendar.  
 
Recurrence Free Survival 
Secondary objectives for this study include recurrence free survival, which is defined 
as the time from complete surgical resection to the date of the first confirmation of 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 44 of 61 
 
disease reoccurance or death (whichever comes first). Progressive disease will be 
based on irRC response criteria. 
 
Neo-Adjuvant Therapy Tumor Response 
Clincal (radiographic or physical exam) Response to neo-adjuvant therapy will be 
assessed by RECIST 1.1. The following definitions and criteria should be used for the 
baseline evaluations of existing disease, and for the ongoing evaluation of tumor 
responses prior to surgery. 
Measurable lesions - lesions that can be accurately measured in at least one 
dimension with longest diameter (LD) ≥ 10 mm using CT, MRI, or caliper or ruler 
measurements or ≥20 mm with x-ray. A lymph node must be ≥15 mm in short axis 
when assessed by CT scan 
Non-measurable lesions - all other lesions including small lesions (LD < 10 mm 
with CT, MRI, or caliper measurements or <20 mm with x-ray). 
Documentation of “Target” and “Non-Target” Lesions 
 All measurable lesions up to a maximum of two lesions per organ and five 
lesions in total, representative of all involved organs should be identified as 
target lesions and recorded and measured at baseline. 
 Target lesions should be selected on the basis of their size (lesions with the 
longest diameter) and their suitability for accurate repeated measurements 
(either by imaging techniques or clinical assessments). 
 A sum of the LD for all target lesions will be calculated and reported as the 
baseline sum LD. The baseline sum LD will be used as the reference by which 
to characterize the objective tumor response. 
 All other lesions (or sites of disease) should be identified as non-target lesions 
and should also be recorded at baseline. Measurements of these lesions are not 
required, but the presence or absence of each should be noted throughout 
follow-up. 
Response Criteria: 
 
Evaluation of target lesions 
Complete Response (CR) 
Disappearance of all target lesions  
(Must persist for a minimum of four weeks) 
Partial Response (PR) 
At least a 30% decrease in the sum of the LD of target lesions, 
taking as reference the baseline sum LD  
(Must persist for a minimum of four weeks) 
Progressive Disease (PD) 
At least a 20% increase in the sum of the LD of target lesions, 
taking as reference the smallest sum LD recorded since the 
treatment started or the appearance of one or more new lesions 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 45 of 61 
 
Stable Disease (SD) 
 
Neither sufficient shrinkage to qualify for PR nor sufficient 
increase to qualify for PD, taking as reference the smallest sum 
LD since the treatment started 
 
 
Evaluation of non-target lesions 
Complete Response (CR) 
Disappearance of all non-target lesions 
Stable Disease (SD) 
Persistence of one or more non-target lesion(s) 
Progressive Disease (PD) 
 
Appearance of one or more new lesions and/or unequivocal 
progression of existing non-target lesions 
Evaluation of Best Overall Response 
The best overall response is the best response observed until progression/recurrence 
and is determined as indicated in the table below: 
Target Lesions 
Non-Target 
Lesions 
 
Evaluation of New 
Lesions 
Best Overall 
Response 
CR 
CR 
No 
CR 
CR 
SD 
No 
PR 
PR 
Non-PD 
No 
PR 
SD 
Non-PD 
No 
SD 
PD 
Any 
Yes or No 
PD 
Any 
PD 
Yes or No 
PD 
Any 
Any 
Yes 
PD 
 
irRC and RECIST 1.1 Concordance 
Exploratory objectives for this study include review of the concordance between irRC 
and RECIST 1.1 response assessments. Both irRC and RECIST 1.1 response 
assessments will be performed at all timepoints prior to surgery.  
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 46 of 61 
 
Immune-Related Response Criteria (irRC) 
Clinical response assessments will be performed according to irRC (Wolchok, 2009) 
Table 1: Overall Response using irRC: 
Response Assessment 
Description 
irCR 
Complete disappearance of all lesions (whether measurable or not, 
and no new lesions) 
Confirmation by repeat, consecutive assessment ≥4 weeks 
after first documentation. 
irPR 
Decrease in tumor burden ≥50% relative to baseline 
Confirmed by a consecutive assessment ≥4 weeks after 
first documentation. 
irSD 
Not meeting criteria for irCR or irPR, in absence of irPD. A 
change in tumor burden of neither 50% decrease from baseline nor 
25% increase from nadir can be determined. 
irPD 
Increase in tumor burden ≥25% relative to nadir (minimum 
recorded tumor burden) 
Confirmation by a repeat, consecutive assessment ≥4 
weeks after the first documentation. 
Abbreviations: CR= Complete Response; irRC= Immune-Related Response Criteria; PD= Progressive 
Disease; PR=Partial Response; SD=Stable Disease 
Table 2: Derivation of irRC overall responses 
Measurable Response 
Non-measurable response 
Overall irRC 
Response 
Target and new, 
measurable lesions % 
change in tumor burden 
Non-target lesions 
New non-
measurable 
lesions 
↓ 1001 
Absent 
Absent 
irCR3 
↓ 1001 
Stable 
Any 
irPR3 
↓ 1001 
Unequivocal progression 
Any 
irPR3 
↓ ≥501 
Absent or Stable 
Any 
irPR3 
↓ ≥501 
Unequivocal progression 
Any  
irPR3 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 47 of 61 
 
↓ ≥50 to ↑ <251 
Absent or Stable 
Any 
irSD 
↓ ≥50 to ↑ <251 
Unequivocal Progression 
Any 
irSD 
↑ ≥252 
Any 
Any 
irPD3 
1 Decrease relative to baseline 
2 Increase relative to nadir 
3 confirmed by consecutive scan ≥4 weeks after first documentation 
 
Overall Survival 
Patient survival will be collected for up to one year following the EOT visit. Survival 
will be calculated by the Kaplan-Meier method.  
 
Correlative Biomarker Analysis  
Tumor and blood samples will be assessed for response and immune activation using: 
histopathology including immunohistochemistry, flow cytometry, NanoString, 
Multispectral analysis. Analysis will include: tumor response/viability, cytokine 
profile changes, T-cell population changes, and RNA level changes.  
 
Complete Surgical Resection 
Patients will be assessed at surgery to determine if complete surgical resection was 
achievable after neo-adjuvant treatment with nivolumab and HF10. Completeness of 
surgical resection will be defined by standard definitions as follows: R0 indicates all 
gross clinical disease resected with microscopic pathologic negative margins; R1 
indicates all gross clinical disease resected, but microscopic pathologic margins are 
positive for residual disease; R2 indicates inability to completely excise all clinical 
disease (i.e. gross residual tumor unable to be removed at surgery).  
 
Pathology 
Archival Tissue: Archival tissue from tumor of lymph node will be used to confirm 
disease pathology and next generation sequencing per standard of care practice. 
 Stopping Rules 
If two subjects experience a grade 4 or 5 toxicity thought to be at least possibly 
related to the study products the study will be paused. Enrollment will be closed until 
the safety issue is discussed with the medical monitor and DSMC.  Enrollment cannot 
reopen without written approval from the DSMC chair.   
10 STATISTICAL CONSIDERATIONS 
The primary endpoint of pathologic response including pCR is expected to be seen in 
30% of patients, hence 6 of the planned 20 patients are expected to have a pathologic 
response, including pCR. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 48 of 61 
 
Pathological response will be determined by assessing the tumor viability at baseline and 
at surgery. Samples will be scored as a percentage viability (0-100%). A pathologic 
complete response (pCR) will be defined as no viable residual melanoma cells in the 
surgical specimen. A major pathologic response will be defined as <50% viable tumor 
cells. A minor pathologic response will be defined as 50% or greater viable tumor cells, 
including specimens that have 100% viability at surgery. Pathologic response rate (pRR) 
will be defined as frequency of either pCR or a major pathologic response. Analysis will 
also be performed of baseline tumor biopsies and on treatment biopsies if available for 
comparisons with the surgical resection specimen. 
The null hypothesis is that the pathologic response rate for patients receiving the 
treatment is 5% or less. The alternative hypothesis is that  the pathologic response  rate is 
30%. With  20 evaluable patients  there  will  be 59% power using  an exact binomial test 
at the one-side 0.05  significance level if the true  response  rate is 20%, and 89% power 
at one-sided  alpha = 0.05  if the true  response  rate is  30%. Four or more pathologic 
responses in up to 20 patients will be sufficient to reject the null hypothesis. In additional 
to the statistical test, a 95% exact binomial confidence interval for the pathologic 
response rate will be calculated.  
Kaplan-Meier methods and associated confidence intervals will be used to analyze RFS 
and OS. All subjects assessed for relapse will contribute to this analysis.  
The statistical analysis of the safety data will be descriptive and tabular in nature.  
The statistical analysis of pathologic and correlative exploratory endpoints will be 
descriptive. No hypothesis tests will be performed for exploratory endpoints. 
11 REGISTRATION GUIDELINES 
Patients must meet all of the eligibility requirements listed in Section 5 prior to 
registration.  
Study related screening procedures can only begin once the patient has signed a 
consent form. Patients must not begin protocol treatment prior to registration. 
Treatment should start within five working days after registration. 
To register eligible patients on study, complete a Clinical Trials Office Patient Registration 
Form and submit to: CTORegistrations@hci.utah.edu.  
12 DATA SUBMISSION SCHEDULE 
The Case Report Forms (CRFs) are a set of (electronic or paper) forms for each patient 
that provides a record of the data generated according to the protocol. CRF’s should be 
created prior to the study being initiated and updated (if applicable) when amendments to 
the protocol are IRB approved. Data capture should be restricted to endpoints and 
relevant patient information required for planned manuscripts. These forms will be 
completed on an on-going basis during the study. The medical records will be source of 
verification of the data. During the study, the CRFs will be monitored for completeness, 
accuracy, legibility and attention to detail by a member of the Research Compliance 
Office. The CRFs will be completed by the Investigator or a member of the study team as 
listed on the Delegation of Duties Log. The data will be reviewed no less than annually 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 49 of 61 
 
by the Data and Safety Monitoring Committee. The Investigator will allow the Data and 
Safety Monitoring Committee or Research Compliance Office personnel access to the 
patient source documents, clinical supplies dispensing and storage area, and study 
documentation for the above-mentioned purpose. The Investigator further agrees to assist 
the site visitors in their activities. 
13 SPECIAL INSTRUCTIONS – CORRELATIVE STUDIES 
 Tissue Samples 
Fresh core tumor biopsies are mandatory at day 0 and surgery, and optional at day 42 
and progression.  On day 0, pre-HF10 injection, collect 2-4 core biopsies from one 
tumor lesion that will be injected (target lesion).  If there is a second lesion available 
collect 2-4 core biopsies from one tumor lesion that will not be injected (non-target 
lesion).  On day 42, surgery and progression collect 2-4 core biopsies from the same 
lesions as days 0 if the lesions are still present. Tumor samples analysis may include, 
but is not limited to:  
 Tumor response/viability 
 Immunohistochemistry  
 Multispectral analysis  
Details of tissue collection, processing and storage will be found in the Lab Manual. 
 Blood Samples 
24 to 28 mLs of blood will be collected prior to HF10 injection at Day 0, 7, 14, 21, 
28, 42, 56, 70 and 84 of the neoadjuvant treatment period and prior to nivolumab on 
infusion days during the adjuvant period. 
Blood sample analysis may include, but is not limited to:  
 Flow cytometry including; CD3+ / CD8, CD3+ / CD4+, CD56+, 
CD3+/CD8+/CD69+, CD3+/CD8+/(TIM3 or LAG3), CD19+ 
 NanoString  
 Multispectral analysis 
 Cytokine/chemokines including; MIP-1 alpha, IL-20, LIF, IL-1 beta, IL-2, IL-
4, IL-5, IP-10, IL-6, IL-7, IL-8, IL-10, PECAM, Eotaxin, IL-12p70, IL-13, IL-
17A, MCP-2, IL-1RA, SCF, RANTES, IFN-gamma, GM-CSF, TNF-alpha, 
HGF, MIP-1 beta, IFN-alpha, IL-9, VEGF-D, IL-16, P-Selectin, TNF-RII, 
BDNF, Gro-alpha/KC, IL-1 alpha, IL-15, IL-21, tPA, OPG, TSLP, PDGF-BB, 
VEGF-A 
Details of blood sample collection, processing and storage will be found in the Lab 
Manual 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 50 of 61 
 
14 ETHICAL AND REGULATORY CONSIDERATIONS 
 Informed consent  
Informed consent will be obtained from all research participants prior to performing 
any study procedures using the most recent IRB approved version.  
 Institutional Review 
Study will be approved by the Institutional Review Board of University of Utah. 
  Data and Safety Monitoring Plan 
A Data and Safety Monitoring Committee (DSMC) is established at Huntsman 
Cancer Institute (HCI) and approved by the NCI to assure the well-being of patients 
enrolled on Investigator Initiated Trials that do not have an outside monitoring 
review. Roles and responsibilities of the DSMC are set forth in the NCI approved 
plan. The activities of this committee include a quarterly review of adverse events 
including SAEs, important medical events, significant revisions or amendments to the 
protocol, and approval of cohort/dose escalations. If the DSMC and/or the PI have 
concerns about unexpected safety issues, the study will be stopped and will not be 
resumed until the issues are resolved. The DSMC also reviews and approves audit 
reports generated by the Research Compliance Office.  
All phase II studies are reviewed by the full committee at each quarterly DSMC 
meeting. This includes a review of all serious adverse events (SAEs) occurring in 
patients treated at HCI or its affiliates as well as all grade 3 or greater toxicities for 
patients on treatment and within 30-day follow-up window (only if possibly, probably 
or definitely related).  
 Adverse Events / Serious Adverse Events 
This study will utilize the CTCAE (NCI Common Terminology Criteria for Adverse 
Events) Version 4.0 for AE and SAE reporting. An electronic copy of the CTCAE 
Version 4.0 can be downloaded from:  
http://safetyprofiler-ctep.nci.nih.gov/CTC/CTC.aspx 
 Adverse Events (AE) 
An adverse event is the appearance or worsening of any undesirable sign, 
symptom, or medical condition occurring after starting the study drug even if the 
event is not considered to be related to study drug. For the purposes of this study, 
the terms toxicity and adverse event are used interchangeably. Medical 
conditions/diseases present before starting study drug are only considered adverse 
events if they worsen after starting study drug. Abnormal laboratory values or test 
results constitute adverse events only if they induce clinical signs or symptoms, 
are considered clinically significant, or require therapy. 
The collection of AEs will begin after the first dose of study treatment and end 30 
days post the last dose of adjuvant study treatment.  


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 51 of 61 
 
Information about all adverse events, whether volunteered by the subject, 
discovered by investigator questioning, or detected through physical examination, 
laboratory test or other means, will be collected and recorded and followed as 
appropriate. 
 
The occurrence of adverse events should be sought by non-directive questioning 
of the patient at each visit or phone contact during the study. Adverse events also 
may be detected when they are volunteered by the patient during or between visits 
or through physical examination, laboratory test, or other assessments. As far as 
possible, each adverse event should be evaluated to determine: 
1. the severity grade based on CTCAE v.4 (grade 1-5) 
2. its relationship to the study drug(s) (definite, probable, possible, unlikely, 
not related) Relationship to both nivolumab and HF10 will both be 
assessed.  
3. its duration (start and end dates or if continuing at final exam) 
4. action taken (no action taken; study drug dosage adjusted/temporarily 
interrupted; study drug permanently discontinued due to this adverse 
event; concomitant medication taken; non-drug therapy given; 
hospitalization/prolonged hospitalization) 
5. whether it constitutes an SAE 
 
All adverse events will be treated appropriately. Such treatment may include 
changes in study drug treatment as listed in the dose modification section of this 
protocol (see section 7 for guidance). Once an adverse event is detected, it should 
be followed until its resolution, and assessment should be made at each visit (or 
more frequently, if necessary) of any changes in severity, the suspected 
relationship to the study drug, the interventions required to treat it, and the 
outcome. 
Information about common side effects already known about both Nivolumab and 
HF10 is described in the Drug Information (section 3) and the most recent 
investigator brochure. This information will be included in the patient informed 
consent and will be discussed with the patient during the study as needed. 
All adverse events will be immediately recorded in the patient research chart.  
 Serious Adverse Event (SAE) 
Information about all serious adverse events will be collected and recorded. A 
serious adverse event is an undesirable sign, symptom or medical condition 
which: 
 is fatal or life-threatening 
 results in persistent or significant disability/incapacity 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 52 of 61 
 
 is medically significant, i.e., defined as an event that jeopardizes the 
patient or may require medical or surgical intervention to prevent one of 
the outcomes listed above 
 causes congenital anomaly or birth defect 
 requires inpatient hospitalization or prolongation of existing 
hospitalization, unless hospitalization is for: 
o routine treatment or monitoring of the studied indication, not 
associated with any deterioration in condition (procedures such as 
central line placements, paracentesis, pain control) 
o elective or pre-planned treatment for a pre-existing condition that is 
unrelated to the indication under study and has not worsened since the 
start of study drug 
o treatment on an emergency outpatient basis for an event not fulfilling 
any of the definitions of a SAE given above and not resulting in 
hospital admission 
o social reasons and respite care in the absence of any deterioration in 
the patient’s general condition 
 
Following the subject’s written consent to participate, all SAEs must be collected. 
SAEs will be followed after consent for 90 days post adjuvant nivolumab 
treatment, or until the patient starts a new a treatment.  
Any death from any cause while a patient is receiving treatment on this protocol 
or up to 30 days after the last dose of protocol treatment, or any death which 
occurs more than 30 days after protocol treatment has ended but which is felt to 
be treatment related, must be reported. 
Toxicities which fall within the definitions listed above must be reported as an SAE 
regardless if they are felt to be treatment related or not. Toxicities unrelated to 
treatment that do NOT fall within the definitions above, must simply be 
documented as AEs in the patient research chart.  
 SAE Reporting Requirements 
SAEs must be reported to the DSMC, the FDA, the IRB, BMS and Takara, according 
to the requirements described below: 
A 
MedWatch 
3500A 
form 
must 
be 
completed 
and 
submitted 
to 
compliance@hci.utah.edu within 1 business day of first knowledge or notification of 
event. 
DSMC Notifications: 
 An HCI Research Compliance Officer (RCO) will process and submit the 
MedWatch form to the proper DSMC member as necessary for each individual 
study.  
 The RCO will summarize and present all reported SAEs according to the Data 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 53 of 61 
 
and Safety Monitoring Plan at the quarterly DSMC meeting. 
For multisite studies the HCI DSMC will notify all participating sites of all unexpected 
and related SAEs via the Research Compliance Office. The RCO will also notify all 
investigators at remote clinical sites participating in a multisite trial of any other safety 
update, including external safety reports, manufacturer’s reports and updates to the 
investigator’s brochure. 
FDA Notifications: 
 Adverse events occurring during the course of a clinical study that meet the 
following criteria will be promptly reported to the FDA: 
 Serious 
 Unexpected 
 Definitely, Probably or Possibly Related to the investigational drug 
 Fatal or life-threatening events that meet the criteria above will be reported 
within 7 calendar days after first knowledge of the event by the investigator; 
followed by as complete a report as possible within 8 additional calendar 
days. 
 All other events that meet the criteria above will be reported within 15 
calendar days after first knowledge of the event by the investigator. 
 The RCO will review the MedWatch report for completeness, accuracy and 
applicability to the regulatory reporting requirements. 
 The RCO will ensure the complete, accurate and timely reporting of the event 
to the FDA. 
 The Regulatory Coordinator will submit the report as an amendment to the 
IND application. 
 All other adverse events and safety information not requiring expedited 
reporting that occur or are collected during the course of the study will be 
summarized and reported to the FDA through the IND Annual Report. 
IRB Notification: 
 Events meeting the University of Utah IRB reporting requirements 
(http://www.research.utah.edu/irb/) will be submitted through the IRB’s 
electronic reporting system within 10 working days. 
 External sites should abide by local IRB requirements for submission of SAEs  
SAEs, whether related or not related to study drug, and pregnancies must be reported to 
BMS within 24 hours. SAEs must be recorded on BMS or an approved form; pregnancies 
must be reported on a Pregnancy Surveillance Form. 
SAE Email Address:  Worldwide.Safety@BMS.com 
SAE Facsimile Number:  609-818-3804 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 54 of 61 
 
If only limited information is initially available, follow-up reports are required. If an 
ongoing SAE changes in its intensity or relationship to study drug or if new information 
becomes available, a follow-up SAE report should be sent within 24 hours to BMS (or 
designee) using the same procedure used for transmitting the initial SAE report.  
Adverse events classified as serious require expeditious handling and reporting to Takara 
Bio Inc. (Takara) to comply with regulatory requirements. 
 For any serious adverse event (SAE) that occurs while a patient is on-study; within 30 days 
of the last study drug administration, regardless of any opinion as to the relationship of the 
SAE to the study drug; or if any SAE that the Investigator feels is related to the study drug 
occurs later than 30 days after the last study drug administration, the investigator must 
notify Takara’s Safety Team immediately (within 24 hours of becoming aware of the 
event) by fax or email. Notification by email is preferred. The fax number and the email 
address are listed below. 
All SAEs require that a Serious Adverse Event Report Form be completed and forwarded 
either via fax or as a PDF via email to Takara at the fax number or email listed below within 
24 hours of becoming aware of the event.  
 SAEs will be reported to:  Takara Safety Team.                                                   
Email: trial1ae1info@takara-bio.co.jp 
FAX: 077-567-9265 
All SAEs should be followed to resolution or stabilization. 
 Reporting of Pregnancy  
Although pregnancy is not considered an adverse event, it is the responsibility of 
investigators or their designees to report any pregnancy or lactation in a subject, 
including the pregnancy of a male subjects’ female partner as an SAE. Pregnancies or 
lactation that occurs during the course of the trial or with 30 days of completing the 
trial or starting another new anticancer therapy, whichever is earlier, must be reported 
to the DSMC, IRB, FDA, and the sponsor as applicable. All subjects and female 
partners who become pregnant must be followed to the completion/termination of the 
pregnancy. Pregnancy outcomes of spontaneous abortion, missed abortion, fetal 
death, intrauterine death, miscarriage and stillbirth must be reported as serious events.  
 Protocol Amendments 
Any amendments or administrative changes in the research protocol during the 
period, for which the IRB approval has already been given, will not be initiated 
without submission of an amendment for IRB review and approval.   
These requirements for approval will in no way prevent any immediate action 
from being taken by the investigator in the interests of preserving the safety of 
all patients included in the trial.  


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 55 of 61 
 
Any amendments to the protocol that significantly affect the safety of subjects, 
scope of the investigation, or the scientific quality of the study are required to 
submit the amendment for FDA review. 
 Protocol Deviations 
A protocol deviation (or violation) is any departure from the defined procedures and 
treatment plans as outlined in the protocol version submitted and previously approved 
by the IRB. Protocol deviations have the potential to place participants at risk and can 
also undermine the scientific integrity of the study thus jeopardizing the justification 
for the research. Protocol deviations are unplanned and unintentional events.  
Because some protocol deviations pose no conceivable threat to participant safety or 
scientific integrity, reporting is left to the discretion of the PI within the context of the 
guidelines below. The IRB requires the prompt reporting of protocol deviations 
which are: 
 
Exceptions to eligibility criteria. 
 
Intended to eliminate apparent immediate hazard to a research participant 
or  
 
Harmful (caused harm to participants or others, or place them at 
increased risk of harm - including physical, psychological, economic, or 
social harm), or  
 
Possible serious or continued noncompliance  
 FDA Annual Reporting  
An annual progress report will be submitted to the FDA within 60 days of the 
anniversary of the date that the IND went into effect. (21 CFR 312.33).  
  Clinical Trials Data Bank  
The study will be registered on http://clinicaltrials.gov and the NCI CTRP (Clinical 
Trials Reporting Program) by the Clinical Trials Office. 
  Record Keeping 
Per 21 CFR 312.57, Investigator records shall be maintained for a period of 2 years 
following the date a marketing application is approved; or, if no application is filed or 
the application is not approved, until 2 years after the investigation is discontinued 
and the FDA is notified. 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 56 of 61 
 
15 BIBLIOGRAPHY 
1. 
Leiter U, Buettner PG, Eigentler TK, et al. Hazard rates for recurrent and 
secondary cutaneous melanoma: an analysis of 33,384 patients in the German Central 
Malignant Melanoma Registry. Journal of the American Academy of Dermatology 
2012;66:37-45. 
2. 
Romano E, Scordo M, Dusza SW, Coit DG, Chapman PB. Site and timing of first 
relapse in stage III melanoma patients: implications for follow-up guidelines. J Clin 
Oncol 2010;28:3042-7. 
3. 
Svedman FC, Pillas D, Taylor A, Kaur M, Linder R, Hansson J. Stage-specific 
survival and recurrence in patients with cutaneous malignant melanoma in Europe - a 
systematic review of the literature. Clin Epidemiol 2016;8:109-22. 
4. 
Eggermont AM, Chiarion-Sileni V, Grob JJ. Correction to Lancet Oncol 2015; 
16: 522-30. Adjuvant ipilimumab versus placebo after complete resection of high-risk 
stage III melanoma (EORTC 18071): a randomised, double-blind, phase 3 trial. The 
lancet oncology 2015;16:e262. 
5. 
Eggermont AM, Chiarion-Sileni V, Grob JJ, et al. Prolonged Survival in Stage III 
Melanoma with Ipilimumab Adjuvant Therapy. The New England journal of medicine 
2016;375:1845-55. 
6. 
Eggermont AM, Chiarion-Sileni V, Grob JJ, et al. Adjuvant ipilimumab versus 
placebo after complete resection of high-risk stage III melanoma (EORTC 18071): a 
randomised, double-blind, phase 3 trial. The lancet oncology 2015;16:522-30. 
7. 
Kirkwood JM, Ibrahim JG, Sondak VK, et al. High- and low-dose interferon alfa-
2b in high-risk melanoma: first analysis of intergroup trial E1690/S9111/C9190. J Clin 
Oncol 2000;18:2444-58. 
8. 
Kirkwood JM, Ibrahim JG, Sosman JA, et al. High-dose interferon alfa-2b 
significantly prolongs relapse-free and overall survival compared with the GM2-
KLH/QS-21 vaccine in patients with resected stage IIB-III melanoma: results of 
intergroup trial E1694/S9512/C509801. J Clin Oncol 2001;19:2370-80. 
9. 
Kirkwood JM, Strawderman MH, Ernstoff MS, Smith TJ, Borden EC, Blum RH. 
Interferon alfa-2b adjuvant therapy of high-risk resected cutaneous melanoma: the 
Eastern Cooperative Oncology Group Trial EST 1684. J Clin Oncol 1996;14:7-17. 
10. 
Larkin J, Chiarion-Sileni V, Gonzalez R, et al. Combined Nivolumab and 
Ipilimumab or Monotherapy in Untreated Melanoma. The New England journal of 
medicine 2015;373:23-34. 
11. 
Larkin J, Hodi FS, Wolchok JD. Combined Nivolumab and Ipilimumab or 
Monotherapy in Untreated Melanoma. The New England journal of medicine 
2015;373:1270-1. 
12. 
Robert C, Long GV, Brady B, et al. Nivolumab in previously untreated melanoma 
without BRAF mutation. The New England journal of medicine 2015;372:320-30. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 57 of 61 
 
13. 
Andtbacka RHI, Ross MI, Agarwala SS, et al. Preliminary results from a phase II 
study of combination treatment with HF10, a replication-competent HSV-1 oncolytic 
virus, and ipilimumab in patients with stage IIIB, IIIC, or IV unresectable or metastatic 
malignant melanoma. American Society of Clinical Oncology, Annual Meeting. Chicago, 
IL2016. 
14. 
Puzanov I, Milhem MM, Minor D, et al. Talimogene Laherparepvec in 
Combination With Ipilimumab in Previously Untreated, Unresectable Stage IIIB-IV 
Melanoma. J Clin Oncol 2016;34:2619-26. 
15. 
Ushijima Y, Luo C, Goshima F, Yamauchi Y, Kimura H, Nishiyama Y. 
Determination and analysis of the DNA sequence of highly attenuated herpes simplex 
virus type 1 mutant HF10, a potential oncolytic virus. Microbes Infect 2007;9:142-9. 
16. 
Berkowitz C, Moyal M, Rosen-Wolff A, Darai G, Becker Y. Herpes simplex 
virus type 1 (HSV-1) UL56 gene is involved in viral intraperitoneal pathogenicity to 
immunocompetent mice. Arch Virol 1994;134:73-83. 
17. 
Koshizuka T, Goshima F, Takakuwa H, et al. Identification and characterization 
of the UL56 gene product of herpes simplex virus type 2. J Virol 2002;76:6718-28. 
18. 
Koshizuka T, Kawaguchi Y, Nishiyama Y. Herpes simplex virus type 2 
membrane protein UL56 associates with the kinesin motor protein KIF1A. J Gen Virol 
2005;86:527-33. 
19. 
Jiang YM, Daikoku T, Yamamoto M, Morishima T, Nishiyama Y. Growth and 
cytopathogenicity of herpes simplex virus in a macrophage cell line, RAW264: A good 
indicator of intraperitoneal pathogenicity. Microbiol Immunol 1995;39:905-9. 
20. 
Sugiura S, Goshima F, Takakuwa H, Sata T, Nakashima T, Nishiyama Y. 
Treatment of solid sarcomas in immunocompetent mice with novel, oncolytic herpes 
simplex viruses. Otolaryngol Head Neck Surg 2004;130:470-8. 
21. 
Takakuwa H, Goshima F, Nozawa N, et al. Oncolytic viral therapy using a 
spontaneously generated herpes simplex virus type 1 variant for disseminated peritoneal 
tumor in immunocompetent mice. Arch Virol 2003;148:813-25. 
22. 
Kimata H, Takakuwa H, Goshima F, et al. Effective treatment of disseminated 
peritoneal colon cancer with new replication-competent herpes simplex viruses. 
Hepatogastroenterology 2003;50:961-6. 
23. 
Teshigahara O, Goshima F, Takao K, et al. Oncolytic viral therapy for breast 
cancer with herpes simplex virus type 1 mutant HF 10. Journal of surgical oncology 
2004;85:42-7. 
24. 
Kohno S, Luo C, Goshima F, Nishiyama Y, Sata T, Ono Y. Herpes simplex virus 
type 1 mutant HF10 oncolytic viral therapy for bladder cancer. Urology 2005;66:1116-
21. 
25. 
Carter L, Fouser LA, Jussif J, et al. PD-1:PD-L inhibitory pathway affects both 
CD4(+) and CD8(+) T cells and is overcome by IL-2. European journal of immunology 
2002;32:634-43. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 58 of 61 
 
26. 
Latchman Y, Wood CR, Chernova T, et al. PD-L2 is a second ligand for PD-1 
and inhibits T cell activation. Nature immunology 2001;2:261-8. 
27. 
Chemnitz JM, Parry RV, Nichols KE, June CH, Riley JL. SHP-1 and SHP-2 
associate with immunoreceptor tyrosine-based switch motif of programmed death 1 upon 
primary human T cell stimulation, but only receptor ligation prevents T cell activation. 
Journal of immunology (Baltimore, Md : 1950) 2004;173:945-54. 
28. 
Sheppard KA, Fitz LJ, Lee JM, et al. PD-1 inhibits T-cell receptor induced 
phosphorylation of the ZAP70/CD3zeta signalosome and downstream signaling to 
PKCtheta. FEBS letters 2004;574:37-41. 
29. 
Nishimura H, Honjo T. PD-1: an inhibitory immunoreceptor involved in 
peripheral tolerance. Trends in immunology 2001;22:265-8. 
30. 
Nishimura H, Nose M, Hiai H, Minato N, Honjo T. Development of lupus-like 
autoimmune diseases by disruption of the PD-1 gene encoding an ITIM motif-carrying 
immunoreceptor. Immunity 1999;11:141-51. 
31. 
Nishimura H, Okazaki T, Tanaka Y, et al. Autoimmune dilated cardiomyopathy 
in PD-1 receptor-deficient mice. Science (New York, NY 2001;291:319-22. 
32. 
Okazaki T, Tanaka Y, Nishio R, et al. Autoantibodies against cardiac troponin I 
are responsible for dilated cardiomyopathy in PD-1-deficient mice. Nature medicine 
2003;9:1477-83. 
33. 
Iwai Y, Ishida M, Tanaka Y, Okazaki T, Honjo T, Minato N. Involvement of PD-
L1 on tumor cells in the escape from host immune system and tumor immunotherapy by 
PD-L1 blockade. Proceedings of the National Academy of Sciences of the United States 
of America 2002;99:12293-7. 
34. 
Salama AD, Chitnis T, Imitola J, et al. Critical role of the programmed death-1 
(PD-1) pathway in regulation of experimental autoimmune encephalomyelitis. The 
Journal of experimental medicine 2003;198:71-8. 
35. 
Blank C, Brown I, Peterson AC, et al. PD-L1/B7H-1 inhibits the effector phase of 
tumor rejection by T cell receptor (TCR) transgenic CD8+ T cells. Cancer Res 
2004;64:1140-5. 
36. 
Dong H, Strome SE, Salomao DR, et al. Tumor-associated B7-H1 promotes T-
cell apoptosis: a potential mechanism of immune evasion. Nature medicine 2002;8:793-
800. 
37. 
Hirano F, Kaneko K, Tamura H, et al. Blockade of B7-H1 and PD-1 by 
monoclonal antibodies potentiates cancer therapeutic immunity. Cancer Res 
2005;65:1089-96. 
38. 
Iwai Y, Terawaki S, Honjo T. PD-1 blockade inhibits hematogenous spread of 
poorly immunogenic tumor cells by enhanced recruitment of effector T cells. 
International immunology 2005;17:133-44. 
39. 
Strome SE, Dong H, Tamura H, et al. B7-H1 blockade augments adoptive T-cell 
immunotherapy for squamous cell carcinoma. Cancer Res 2003;63:6501-5. 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 59 of 61 
 
40. 
Dong H, Chen L. B7-H1 pathway and its role in the evasion of tumor immunity. 
Journal of molecular medicine (Berlin, Germany) 2003;81:281-7. 
41. 
Konishi J, Yamazaki K, Azuma M, Kinoshita I, Dosaka-Akita H, Nishimura M. 
B7-H1 expression on non-small cell lung cancer cells and its relationship with tumor-
infiltrating lymphocytes and their PD-1 expression. Clin Cancer Res 2004;10:5094-100. 
42. 
Ohigashi Y, Sho M, Yamada Y, et al. Clinical significance of programmed death-
1 ligand-1 and programmed death-1 ligand-2 expression in human esophageal cancer. 
Clin Cancer Res 2005;11:2947-53. 
43. 
Thompson RH, Gillett MD, Cheville JC, et al. Costimulatory molecule B7-H1 in 
primary and metastatic clear cell renal cell carcinoma. Cancer 2005;104:2084-91. 
44. 
Thompson RH, Gillett MD, Cheville JC, et al. Costimulatory B7-H1 in renal cell 
carcinoma patients: Indicator of tumor aggressiveness and potential therapeutic target. 
Proceedings of the National Academy of Sciences of the United States of America 
2004;101:17174-9. 
45. 
Thompson RH, Webster WS, Cheville JC, et al. B7-H1 glycoprotein blockade: a 
novel strategy to enhance immunotherapy in patients with renal cell carcinoma. Urology 
2005;66:10-4. 
46. 
Tsushima F, Tanaka K, Otsuki N, et al. Predominant expression of B7-H1 and its 
immunoregulatory roles in oral squamous cell carcinoma. Oral oncology 2006;42:268-74. 
47. 
Azuma T, Yao S, Zhu G, Flies AS, Flies SJ, Chen L. B7-H1 is a ubiquitous 
antiapoptotic receptor on cancer cells. Blood 2008;111:3635-43. 
48. 
Hino R, Kabashima K, Kato Y, et al. Tumor cell expression of programmed cell 
death-1 ligand 1 is a prognostic factor for malignant melanoma. Cancer 2010;116:1757-
66. 
49. 
Effect of MDX-1106 on CD4+ T cells during an allogeneic mixed lymphocyte 
reaction (MLR) (Medarex Study No. MDX-1106-026-R). Bristol-Myers Squibb 
Pharmaceutical Research Institute 2006;Document Control No. 930046581. 
50. 
Kimata H, Imai T, Kikumori T, et al. Pilot study of oncolytic viral therapy using 
mutant herpes simplex virus (HF10) against recurrent metastatic breast cancer. Annals of 
surgical oncology 2006;13:1078-84. 
51. 
Fujimoto Y, Mizuno T, Sugiura S, et al. Intratumoral injection of herpes simplex 
virus HF10 in recurrent head and neck squamous cell carcinoma. Acta oto-laryngologica 
2006;126:1115-7. 
52. 
Nakao A, Kasuya H, Sahin TT, et al. A phase I dose-escalation clinical trial of 
intraoperative direct intratumoral injection of HF10 oncolytic virus in non-resectable 
patients with advanced pancreatic cancer. Cancer gene therapy 2011;18:167-75. 
53. 
Andtbacka RH, Kaufman HL, Collichio F, et al. Talimogene Laherparepvec 
Improves Durable Response Rate in Patients With Advanced Melanoma. J Clin Oncol 
2015;33:2780-8. 
 
 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 60 of 61 
 
16  APPENDIX 
APPENDIX I: PATIENT INFORMATION SHEET: MINIMIZING VIRUS EXPOSURE 
TO OTHERS 
HF10 is an antitumor medicine in which the active component is a strain of a live virus 
called Herpes Simplex Virus Type 1 (HSV-1). HSV-1 is very widespread in the 
environment. In the United States, many people have been exposed to HSV. The HF10 
virus used in the study drug is a weakened strain of the “live virus”, much like many 
vaccines children receive. Therefore, exposure to the virus may pose a risk of infection to 
other people, particularly those with decreased immunity (see list below). If an infection 
occurs, the most common signs are: 
• Fever 
• Flu-like symptoms (e.g., chills, fatigue, or headache) 
• Nausea, vomiting, or diarrhea 
This information sheet addresses questions that some patients may have regarding risks 
that the virus may pose to people with whom they come in contact. Previous clinical 
studies with HF10 suggested that the virus poses minimal health risk. HF10 has been 
studied in three Phase I trials in Japan, in patients with recurrent breast cancer, head and 
neck cancer, and inoperable pancreatic cancer. The only side effect reported was low-
grade fever in two patients in the head and neck cancer study. 
The association of HF10 with any human illness has not been established. However, 
given the investigational nature of this treatment, your physicians believe that it is 
appropriate to take some precautions to minimize the exposure to HF10 in persons with 
whom you have close contact. After the administration of HF10, the virus may be present 
in your saliva and/or other body fluids (e.g., blood or urine). This can begin within a few 
hours of the treatment and may persist for several days. It is currently not known if, or for 
how long, HF10 may be found in these fluids following your treatment. 
Therefore, your physicians believe it is wise for you and your family to take steps to 
reduce their exposure to HF10, and especially, to minimize exposure to persons beyond 
your immediate family. For one week after each injection, you should carry out the 
following precautions to minimize exposure of others to HF10: 
• Stay at home as much as possible for the first five days after receiving your HF10 
injection. When you are around others, including members of your family, please wear a 
mask, provided by the clinic. 
• Avoid close contact with possible exposure to body fluids (e.g., kissing and sexual 
activity) to minimize exposure to your partner. It is also important that patients (or their 
partners) avoid becoming pregnant during this time. 
• If possible, arrange to sleep in a room separate from others. 
• Do not share towels or eating utensils with others. 
• Isolate your toothbrush from those of others. 
• Wash your hands frequently, especially after coughing, blowing your nose, or using a 


Protocol name:  Phase II Neoadjuvant trial of Nivolumab in Combination with HF10 Oncolytic Viral Therapy in Resectable Stage 
IIIB, IIIc, IVM1a Melanoma (Neo-NivoHF10) 
Version Date: 22JUL2019 
Principal Investigator: John Hyngtrom, MD 
Page 61 of 61 
 
restroom. 
• If a surface comes in contact with body fluids, the area should be cleaned using chlorine 
bleach. 
• As much as possible, avoid public transportation and crowds; be sure to wear a mask 
when out in public. 
• Avoid contact with persons who may have decreased immunity, such as: 
 Children 6 years old or younger 
 Elderly persons 
 Anyone who has had an organ or tissue transplant 
 Anyone with AIDS or HIV infection 
 Other cancer patients. 
• Be sure to wear a mask when visiting the treatment clinic. 
You may need to continue to carry out these precautions based on the results of 
laboratory testing of your saliva, blood, and urine for the detection of virus DNA. If you 
have any questions or concerns about any of the precautions listed above, please ask your 
healthcare providers.