Phase Ib Study of Oncolytic Polio/Rhinovirus Recombinant Against Recurrent Malignant Glioma in Children
Phase: PHASE1
Conditions: Malignant Glioma, Anaplastic Astrocytoma, Anaplastic Oligoastrocytoma, Anaplastic Oligodendroglioma, Glioblastoma, Gliosarcoma, Atypical Teratoid/Rhabdoid Tumor of Brain, Medulloblastoma, Ependymoma, Pleomorphic Xanthoastrocytoma of Brain, Embryonal Tumor of Brain
Sponsor: Istari Oncology, Inc.

 
 
Phase Ib study of Oncolytic Polio/Rhinovirus 
Recombinant Against Recurrent Malignant Glioma in 
Children 
 
 
Sponsor: 
Istari Oncology, Inc. 
 
Funding Source: 
Solving Kids Cancer in 
collaboration with The Andrew 
McDonough B+ Foundation 
 
Protocol Source: 
Duke Cancer Institute 
 
Duke IRB#: 
WIRB#: 
Pro00071228 
20171431 
 
IND# 
14,735 
Principal Investigator 
Eric Thompson, MD 
Duke University Medical Center 
DUMC Box 3272 
2301 Erwin Rd. 
Durham, NC 27710 
Ph. (919) 684-5013 
Eric.thompson@duke.edu 
 
Co-Principal Investigator 
Daniel Landi, MD 
The Preston Robert Tisch Brain 
Tumor Center at Duke 
Hanes House, 330 Trent Drive 
DUMC Box 102382 
Durham, NC 27710 
Ph. (919) 681-4047 
Fax. (919) 668-2485  
Daniel.landi @duke.edu 
 
Sponsor Contacts: 
Garrett Nichols, MD 
Chief Medical Officer, Istari 
Oncology, Inc. 
Gnichols@istarioncology.com 
 
Andrea True Kelly, PhD 
VP of Medical Affairs/Clin Ops, Istari 
Oncology, Inc. 
atkelly@istarioncology.com 
 
Sub-Investigator(s) 
Bridget Archambault, RN, MSN, CPNP 
Bridget.archambault@duke.edu 
 
Darell Bigner, MD, PhD (Istari signatory for 
IND) Darell.bigner@duke.edu 
 
Annick Desjardins, MD, FRCPC 
Annick.desjardins@duke.edu 
 
Allan Friedman, MD 
Allan.friedman@duke.edu 
 
Matthias Gromeier, MD 
Matthias.gromeier@duke.edu 
 
Mustafa Khasraw, MBChB, MD, FRCP, 
FRACP 
Mustafa.Khasraw@duke.edu 
 
John Sampson, MD, PhD 
John.sampson@duke.edu 
 
Kristin Schroeder, MD, MPH 
Kristin.schroeder@duke.edu 
 
David Van Mater, MD, PhD 
David.vanmater@duke.edu 
 
David Ashley, MBBS, FRACP, PhD 
David. ashley@duke.edu 
Statistician 
James E. Herndon II, PhD 
James.herndon@duke.edu 
 
Lead Study Coordinator 
Stevie Threatt 
Stevie.threatt@duke.edu 
 
Regulatory Coordinator 
Elizabeth Miller 
Elizabeth.s.Miller@duke.edu 
 
Data Manager 
Eric Lipp 
Eric.lipp@duke.edu 
 
 
 

Revision History 
Original version: 
02/24/2016 
 
Amended version 1: 
10/24/2016 
Revisions post RAC review 
Amended version 2: 
12/23/2016 
Revisions post CPC review 
Amended version 3: 
03/09/2017 
Change in Dose Level 
Amended version 4: 
05/16/2017 
Change in IND Sponsor 
Amended version 4.1: 
08/04/2017 
Revisions post WIRB review 
Amended version 5: 
10/10/2017 
 
Added David Ashley contact information.  
Added WIRB protocol number to title page.  
Added three additional core biopsies for full 
genome and full exome sequencing 
(Sections 9.1.1, 9.1.4, 12.3). Edited dose 
error in Section 9.2. Removed stool sampling 
at 8 weeks based on the IBC decision that it 
was no longer necessary (Section 9.1.6, 12, 
and 12.4Error! Reference source not 
found.).  Removed reference to Therataxis 
as they will no longer be used to review MRIs 
(Section 9.1.3).  Clarified that the study will 
include 18 year old patients (Section 7.3 and 
11.1).  Added patient and guardian in 
addition to parents as able to sign consent in 
the Inclusion Criteria and changed 
SGOT/SGPT to AST/ALT (Section 11.1).  
SGOT=AST and SGPT=ALT, but we 
commonly use the term AST/ALT in our 
clinic, so the wording has been updated.  
Monitoring rules updated to monitoring 
occurring after each of the first 3 patients, 
followed by annual monitoring of 1-3 patients 
(Section 14.1). 
Amended version 6: 
12/11/2017 
Minor corrections or revisions (Sections 5.2, 
9.1.2, 9.1.6).  Updates to the study drug 
information (Sections 10.3, 10.4).  Remove 
“unsupported” from 2 laboratory inclusion 
criteria (Section 11.1).  Changes or 
corrections made to schedule of procedures 
in Table 2 and throughout Section 12.  
Changes to blood volumes in Table 3 and 
throughout Section 12. 
Amended version 7: 
03/06/2018 
Withdrawn/not implemented 
Amended version 8: 
07/30/2018 
Revisions to inclusion and exclusion criteria 
in Sections 11.1 and 11.2 to allow for a 
greater number of potentially eligible 
patients.  Rationale provided in Section 7.3.  
Additional information regarding use of 
bevacizumab in the pediatric population has 
been added to Section 9.1.6. 
Amended version 8.1: 
09/18/2018 
Revisions requested by FDA to v.8.0 
inclusion criteria #1 and #2 in Section 11.1. 
Amended version 9: 
11/21/2018 
Remove rationale for including patients with 
lesions in the posterior fossa in Section 7.3. 
Revisions to eligibility criteria to add 
‘supratentorial’ to inclusion criterion #1 in 
Section 11.1 and add ‘cerebellum’ to 
exclusion criterion #10 in Section 11.2.  
Additional instructions for safety reporting 
(SAE reporting within 24h of PI/site learning 
of event) to the Sponsor provided in Section 
13.2.1. 
Amended version 10: 
02/20/2019 
Change Co-PI to Daniel Landi, MD; add 
David Ashley, MBBS, FRACP, PhD as sub-
investigator 
 
Amended version 11: 
10/31/2019 
Updated sub-investigators and changed 
Lead Study Coordinator on p.1; Rationale for 

expanding the eligible age to a younger 
patient population added to Section 7.3. Re-
wording in Section 7.2.4. Clinical experience 
background information updated in Section 
7.2.6. Magnevist® co-infusion removed from 
study, along with corresponding immediate 
post-infusion MRI affecting Sections 5.3, 
9.1.2, 9.1.3, 11.2, and 12.3 and Table 2. 
Sentence removed from Section 9.1.3. 
Sentence describing risks of sedation added 
in Section 9.1.6; Age range changed in 
Section 9.2 and in inclusion criteria in Section 
11.1.  Inclusion criterion regarding ability to 
undergo brain MRI without anesthesia 
removed in Section 11.1. Clarified assent in 
inclusion criteria in Section 11.1; Exclusion 
criterion regarding previous PV infection re-
worded in Section 11.2; Exclusion criterion 
regarding gadolinium allergy changed in 
Section 11.2; Remove KPS on Day 1 in 
Section 12, Table 2 and Section 12.4; Add a 
4 mL blood draw for immune analysis on Day 
1 in Section 12, Table 2 and Section 12.4; 
Baseline MRI within 3 days changed to within 
5 days in Table 2 and Section 12.1; Text in 
Section 13.1.1 changed to N/A.  
Amendment Version 
12 
08/19/2020 
Cover page: updated CMO for Istari. 
Updated language to remove 14-day hold 
between treated patients based on interim 
safety review in Sections 5.5, 7.3, and 
15.4.1. Clarified Sponsor monitoring in 
Section 14.1. 

 
 
1 TABLE OF CONTENTS 
1 
TABLE OF CONTENTS ........................................................................................... 3 
2 
LIST OF TABLES ..................................................................................................... 7 
3 
LIST OF FIGURES ................................................................................................... 7 
4 
LIST OF ABBREVIATIONS ...................................................................................... 8 
5 
PROTOCOL SYNOPSIS AND RESEARCH SUMMARY ....................................... 10 
5.1 
Purpose ............................................................................................................ 10 
5.2 
Background and Significance ........................................................................... 10 
5.3 
Design and Procedure ..................................................................................... 11 
5.4 
Selection of Subjects ........................................................................................ 11 
5.5 
Data Analysis and Statistical Considerations ................................................... 11 
6 
STUDY SCHEMA ................................................................................................... 12 
7 
BACKGROUND AND SIGNIFICANCE .................................................................. 13 
7.1 
Study Disease .................................................................................................. 13 
7.2 
Study Agent...................................................................................................... 13 
7.2.1 
PVSRIPO ................................................................................................... 13 
7.2.2 
PVSRIPO Oncolysis .................................................................................. 13 
7.2.3 
PVSRIPO Tumor Tropism ......................................................................... 14 
7.2.4 
PVSRIPO Tumor-Specific Cell Killing ........................................................ 14 
7.2.5 
Pre-clinical Experience .............................................................................. 14 
7.2.6 
Clinical Experience .................................................................................... 15 
7.3 
Study Purpose/Rationale ................................................................................. 16 
8 
OBJECTIVES AND ENDPOINTS .......................................................................... 17 
9 
INVESTIGATIONAL PLAN ..................................................................................... 18 
9.1 
Study Design .................................................................................................... 18 
9.1.1 
Catheter Implantation ................................................................................ 18 
9.1.2 
Agent Infusion ............................................................................................ 18 
9.1.3 
Gadolinium Distribution Quantitation (applies only to patients treated prior 
to October 31, 2019) .............................................................................................. 19 
9.1.4 
Biopsy Sampling and Analyses ................................................................. 20 
9.1.5 
Definition of Unacceptable Toxicity ............................................................ 20 
9.1.6 
Safety Considerations................................................................................ 21 
9.1.7 
Missed Doses ............................................................................................ 24 
9.1.8 
Concomitant Medications .......................................................................... 24 

 
 
9.1.9 
Study Drug Blinding ................................................................................... 25 
9.2 
Rationale for Selection of Dose, Regimen, and Treatment Duration ................ 25 
9.3 
Rationale for Correlative Studies ...................................................................... 25 
9.4 
Definition of Evaluable Subjects ....................................................................... 26 
9.5 
Early Study Termination ................................................................................... 26 
10 
STUDY DRUG ..................................................................................................... 26 
10.1 Names, Classification, and Mechanism of Action ............................................. 26 
10.2 Packaging and Labeling ................................................................................... 26 
10.3 Supply, Receipt, and Storage........................................................................... 26 
10.4 Dispensing and Preparation ............................................................................. 27 
10.5 Compliance and Accountability ........................................................................ 27 
10.6 Disposal and Destruction ................................................................................. 27 
11 
SUBJECT ELIGIBILITY ....................................................................................... 27 
11.1 Inclusion Criteria .............................................................................................. 27 
11.2 Exclusion Criteria ............................................................................................. 28 
12 
SCREENING AND ON-STUDY TESTS AND PROCEDURES ............................ 30 
12.1 Screening Examination .................................................................................... 31 
12.2 Run-In Period ................................................................................................... 31 
12.3 Treatment Period ............................................................................................. 31 
12.4 Follow-up Period .............................................................................................. 31 
12.5 End of Study..................................................................................................... 33 
12.6 Early Withdrawal of Subject(s) ......................................................................... 33 
12.6.1 
Criteria for Early Withdrawal ................................................................... 33 
12.6.2 
Follow-up Requirements for Early Withdrawal........................................ 33 
12.6.3 
Replacement of Early Withdrawal(s) ...................................................... 33 
12.7 Study Assessments .......................................................................................... 33 
12.7.1 
Medical History ....................................................................................... 33 
12.7.2 
Physical Exam ........................................................................................ 33 
12.7.3 
Radiographic Review ............................................................................. 33 
12.7.4 
Laboratory Evaluations ........................................................................... 34 
12.7.5 
Correlative Assessments ........................................................................ 34 
13 
SAFETY MONITORING AND REPORTING ....................................................... 35 
13.1 Adverse Events ................................................................................................ 35 
13.1.1 
AEs of Special Interest ........................................................................... 36 

 
 
13.1.2 
Reporting of AEs .................................................................................... 36 
13.2 Serious Adverse Events ................................................................................... 36 
13.2.1 
Reporting of SAEs .................................................................................. 36 
13.3 Emergency Unblinding of Investigational Treatment ........................................ 37 
13.4 Other Reportable Information ........................................................................... 37 
13.5 Special Warnings and Precautions .................................................................. 37 
13.6 Safety Oversight Committee (SOC) ................................................................. 37 
13.7 External Data and Safety Monitoring Board (DSMB) ....................................... 37 
14 
QUALITY CONTROL AND QUALITY ASSURANCE........................................... 37 
14.1 Monitoring ........................................................................................................ 37 
14.2 Audits ............................................................................................................... 38 
14.3 Data Management and Processing .................................................................. 38 
14.3.1 
Case Report Forms (CRFs) ................................................................... 38 
14.3.2 
Data Management Procedures and Data Verification ............................ 38 
14.3.3 
Study Closure ......................................................................................... 39 
15 
STATISTICAL METHODS AND DATA ANALYSIS ............................................. 39 
15.1 Analysis Sets .................................................................................................... 39 
15.2 Patient Demographics and Other Baseline Characteristics .............................. 39 
15.3 Treatments ....................................................................................................... 39 
15.4 Primary Objective ............................................................................................. 39 
15.4.1 
Overview of Study Design and Monitoring ............................................. 39 
15.4.2 
Primary Objective Analyses ................................................................... 40 
15.5 Secondary Objective ........................................................................................ 41 
15.6 Exploratory Objective ....................................................................................... 41 
15.7 Other Exploratory Objectives ........................................................................... 41 
15.8 Interim Analysis ................................................................................................ 41 
15.9 Sample Size Calculation .................................................................................. 41 
16 
ADMINISTRATIVE AND ETHICAL CONSIDERATIONS..................................... 42 
16.1 Regulatory and Ethical Compliance ................................................................. 42 
16.2 DUHS Institutional Review Board and DCI Cancer Protocol Committee .......... 42 
16.3 Informed Consent ............................................................................................. 42 
16.4 Privacy, Confidentiality, and Data Storage ....................................................... 42 
16.5 Data and Safety Monitoring .............................................................................. 43 
16.6 Protocol Amendments ...................................................................................... 43 

 
 
16.7 Records Retention ........................................................................................... 43 
17 
REFERENCES .................................................................................................... 43 
 
 
 

 
 
2 LIST OF TABLES 
Table 1.  Objectives and Endpoints ............................................................................... 17 
Table 2.  Schedule of Study Tests and Procedures ...................................................... 30 
Table 3.  Schedule of Blood Draw Volumes .................................................................. 35 
Table 4.  Acute Toxicity Monitoring Rules. .................................................................... 40 
Table 5.  Probability of study suspension. ..................................................................... 40 
3 LIST OF FIGURES 
Figure 1.  The genetic structure of PVSRIPO (C) and its precursors type 1 (Mahoney) 
(A), and PV1S (B).......................................................................................................... 13 
Figure 2.  Pathways involved in PVSRIPO oncolytic immunotherapy. .......................... 14 
Figure 3.  Genetic structure of PVSRIPO. ..................................................................... 26 
 
 
 

 
 
4 LIST OF ABBREVIATIONS 
aPTT 
Activated Partial Thromboplastin Time 
AE 
Adverse Event 
ALK 
Alkaline Phosphatase 
ALT 
Alanine Transaminase 
AST 
Aspartate aminotransferase 
ATRT 
Atypical Teratoid/Rhabdoid Tumor 
CBC 
Complete Blood Count 
CED 
Convection-enhanced Delivery 
CMP 
Comprehensive Metabolic Panel 
CNS 
Central Nervous System 
CPC 
Cancer Protocol Committee 
CR 
Complete Response 
CRM 
Continual Reassessment Method 
CT 
Computed Tomography 
CTCAE 
Common Terminology Criteria for Adverse Events 
DLT 
Dose Limiting Toxicity 
DMEM 
Dulbecco’s Minimal Essential Medium 
DSMB 
Data and Safety Monitoring Board 
DVT 
Deep Vein Thrombosis 
EGFRvIII 
Epidermal Growth Factor Receptor (variant III) 
eIF 
Eukaryotic Initiation Factor 
FDA 
Food and Drug Administration 
FLAIR 
Fluid-Attenuated Inversion Recovery 
GBM 
Glioblastoma 
GCS 
Glascow Coma Scale 
Gd-DTPA Gadolinium Diethylene Triamine Pentaacetic Acid 
H&P 
History & Physical Exam 
HRPP 
Human Research Protections Program 
HRV2 
Human Rhinovirus Type 2 
HAS 
Human Serum Albumin 
IgG 
Immunoglobulin G 
ICS 
Investigational Chemotherapy Service 
IND 
Investigational New Drug 
IRB 
Institutional Review Board 
IRES 
Internal Ribosomal Entry Site 
KPS 
Karnofsky Performance Status 
ICH 
Intracerebral Hemorrhage 
IV (or iv) 
Intravenously 
LSQ 
Lymphocyte Subset Quantitation 
MG 
Malignant Glioma 
MMSE 
Mini-Mental State Exam 
NCI 
National Cancer Institute 
Necl-5 
Nectin-like Molecule 5 
NED 
No Evidence of Disease 
NHP 
Non-human Primate 
NINDS 
National Institute of Neurological Disease & Stroke 
NSICU 
Neuro-Surgical Intensive Care Unit 
OARC 
Office of Audit, Risk, and Compliance 
ORR 
Overall Response Rate 
OS 
Overall Survival 
PD 
Progressive Disease 
PET 
Positron Emission Tomography 
PFS 
Progression Free Survival 
PKC 
Protein Kinase C 

 
 
p.o. 
per os/by mouth/orally 
PR 
Partial Response 
PRTBTC 
Preston Robert Tisch Brain Tumor Center 
PT 
Prothrombin Time 
PV 
Poliovirus 
PV1S 
Poliovirus Serotype 1 (Sabin) 
PVSRIPO Polio/Rhinovirus Recombinant 
RAC 
Recombinant DNA Advisory Committee 
SAE 
Serious Adverse Event 
SD 
Stable Disease 
SIADH 
Syndrome of Inappropriate Antidiuretic Hormone 
TCID 
Tissue Culture Infectious Dose 
UTR 
Untranslated Region 
WHO 
World Health Organization 
 
 

 
 
5 PROTOCOL SYNOPSIS AND RESEARCH SUMMARY 
5.1 Purpose 
The purpose of the study is to confirm the safety of the selected dose and potential toxicity of 
oncolytic poliovirus (PV) immunotherapy with PVSRIPO for pediatric patients with recurrent WHO 
grade III or IV malignant glioma, but evidence for efficacy will also be sought. 
 
Primary Objective: 
1. Confirm the safety of the selected dose of PVSRIPO when delivered intracerebrally by 
convection-enhanced delivery (CED) in children with recurrent WHO Grade III malignant glioma 
(anaplastic astrocytoma, anaplastic oligoastrocytoma, anaplastic oligodendroglioma, anaplastic 
pleomorphic xanthoastrocytoma, ependymoma) or WHO Grade IV malignant glioma (glioblastoma, 
gliosarcoma)  
 
Secondary Objective: 
1. Estimate Overall Survival (OS) in children with recurrent WHO grade III and IV malignant glioma 
treated with a single dose of PVSRIPO  
 
Exploratory Objectives: 
1. Describe changes visualized on imaging due to intratumoral inoculation of PVSRIPO 
2. Describe changes in tumor volume visualized on imaging after intratumoral inoculation of 
PVSRIPO 
3. Assess immunologic responses in peripheral blood and in serum  
4. Identify genetic predictors of response or failure of response to treatment with PVSRIPO 
 
Hypothesis: 
Intracerebral injection of PVSRIPO will be feasible and safe in children with WHO grade III or IV 
malignant glioma. 
5.2 Background and Significance 
Pediatric brain tumors are the most common solid tumor in children with approximately 4,000 cases 
every year in the United States.  While significant improvements have been made in the treatment of 
these children, the outcome for those with recurrent brain tumors remains grim. This is particularly so 
for those with recurrent malignant glioma. Treatment failure is frequently due to the presence of a 
blood-brain barrier resulting in poor penetration of cytotoxic drugs into areas where this barrier is 
intact and poor tumor control. Various approaches have been used successfully to circumvent the 
blood-brain barrier, including CED, a process by which large molecules (>400 daltons) are directly 
infused under pressure into a tumor through a catheter.  CED results in adequate distribution of such 
molecules into the tumor over large areas via inherent interstitial fluid pathways.  
 
Oncolytic virus immunotherapy for brain tumors is a unique approach with several advantages over 
more conventional drugs. Certain oncolytic viruses are capable of selective tumor cell killing with a 
range of inflammatory and immune-stimulatory effects on the tumor itself, tumor stromal component 
and the host immune system at large. The objective of oncolytic immunotherapy is to recruit effector 
adaptive immune responses against tumor-associated antigens that can produce lasting immunologic 
control of cancers.  
 
PVSRIPO is a genetically recombinant, non-pathogenic PV:rhinovirus chimera with a tumor-specific 
conditional replication phenotype. It consists of the genome of the live attenuated PV serotype 1 
(Sabin) vaccine (PV1S) with its cognate IRES element replaced with that of HRV2. PVSRIPO tumor 
tropism is mediated by the PV receptor, CD155. CD155, an onco-fetal cell adhesion molecule 
ectopically upregulated in ectodermal/ neuroectodermal cancers, is broadly expressed on cancerous 
cells, cancer ‘stem-cell-like cells’ and tumor-associated proliferating vasculature. Infection with 
PVSRIPO results in swift destruction of tumor cells. PV’s inherent neuropathogenicity was removed 
by IRES exchange; this ablated the virus’ ability to propagate in cells of neuronal lineage and to 

 
 
cause poliomyelitis. However, PVSRIPO replicates efficiently in cancerous cells and exhibits potent 
anti-neoplastic effects in animal tumor models. Tumor cell-specificity mediated by the foreign IRES in 
PVSRIPO relies (i) on a ‘non-productive’ ribonucleoprotein complex forming at the foreign HRV2 
IRES in neurons; (ii) constitutive signal transduction via PKC-Ras-ERK1/2 to translation machinery, 
which stimulates viral, cap-independent translation via the HRV2 IRES in cancerous cells.  
5.3 Design and Procedure 
This protocol is designed primarily to confirm the safety of the selected dose of a novel oncolytic 
agent with MG-specific viral translation and cytotoxicity. PVSRIPO will be delivered intratumorally by 
CED using an intracerebral catheter placed within the enhancing portion of the tumor. Based on a 
Phase I study of the agent in adult patients with recurrent glioblastoma, the amount to be delivered 
will be 5 x 107 tissue culture infectious dose (TCID50). A total of 3 mLs of the agent in physiologic 
saline stabilized with 0.2% human serum albumin will be delivered over 6 hours 30 minutes, 
corresponding to a flow-rate of 0.5 mL/hr. The agent is stable at room temperature during the 
instillation period and there is no adsorptive loss of PVSRIPO in the intended delivery apparatus.  
5.4 Selection of Subjects 
All inclusion/exclusion criteria may be found in Section 11. 
5.5 Data Analysis and Statistical Considerations 
A maximum of 12 patients will be treated with PVSRIPO, and then carefully monitored for safety for at 
least a year after treatment.  Of particular interest will be the incidence of adverse events that occur 
during the first 14 days after PVSRIPO treatment and the inflammatory events that occur during the 
first year after PVSRIPO treatment.  Our experience with PVSRIPO treatment of adult patients with 
recurrent WHO grade IV malignant glioma suggests that a patient remains at risk, during the first 12 
months post-PVSRIPO treatment, of demonstrating neurologic or radiographic signs suggestive of an 
inflammatory reaction secondary to the immune response triggered by PVSRIPO that requires a dose 
of dexamethasone above 4 mg a day.  As described in Section 9.1.6, such patients will be treated 
with bevacizumab 7.5 mg/kg IV every 3 weeks if possible, or alternatively corticosteroids or surgery.  
 
The accrual of pediatric patients to this study was staggered for the first 5 patients enrolled so that no 
more than 1 patient was treated every 14 days.  A slow accrual was planned to minimize the number 
of pediatric patients exposed to PVSRIPO before adequate information about the sequelae of the 
inflammatory process within a pediatric patient is known. Based on a review of the available safety 
data and acceptable tolerability in the first 5 patients treated with PVSRIPO (no dose-limiting toxicities 
or suspected unexpected serious adverse reactions observed), the protocol was amended to allow for 
accrual of subsequent patients without the 14-day waiting period in Protocol Amendment 12. 
 
Two sets of safety monitoring guidelines are proposed in Section 15.4.1, with one focused on acute 
toxicity (i.e. occurring during the first 14 days after PVSRIPO administration), and one focused on 
long-term issues. 
 
The Kaplan-Meier estimator will describe the survival of patients treated with PVSRIPO.  Survival 
(OS) will be calculated as the time between PVSRIPO infusion and death, or last follow-up if the 
patient remains alive.  Analyses will be stratified by grade. 
 
 

 
 
6 STUDY SCHEMA 
 
 
 
 
HGG Recurrence 
Consent 
Biopsy 
Confirm Tumor 
Catheter Placement 
Confirm Catheter Placement (CT) 
Start Infusion 
Catheter Removal 
Stop Infusion 
~30 min 
~390 min 

 
 
7 BACKGROUND AND SIGNIFICANCE 
7.1 Study Disease 
The brain is the most frequent site for the occurrence of primary solid tumors in children; mortality 
from this disease has overtaken leukemias and is now the most frequent cause of cancer death in 
this population 1. Malignant glioma (including WHO grades III and IV) is the third most common 
primary CNS malignancy in children, but far less frequent in this age group as compared to adults. 
The incidence is 0.59 per 100,000 children in the United States with ~200-300 new cases per year 2. 
The standard of care treatment for those with newly diagnosed disease includes adequate surgical 
resection followed by focal radiotherapy and chemotherapy 1. The use of oral temozolomide 
concurrently with radiotherapy followed by maintenance chemotherapy using the same agent did not 
show improved survival compared to historical controls 3. In a recent Children’s Oncology Group 
(COG) phase II randomized study, the addition of novel agents including bevacizumab or a histone 
deacetylase (HDAC) inhibitor (Vorinostat; Zolinza, Merck Corporation, USA) did not show any 
improvement in survival compared to temozolomide alone resulting in early termination of the study1. 
Once tumor recurs, salvage therapies are often unsuccessful. Treatment failure is frequently due to 
the presence of a blood-tumor barrier and poor penetration of cytotoxic drugs into tumor 4. The blood-
tumor barrier can be bypassed by CED 4-6. This therapeutic approach has been widely tested in 
adults and has also been found to be feasible and safe in children in both cerebral hemispheres and 
brain stem locations 7.  
7.2 Study Agent 
7.2.1 PVSRIPO 
PVSRIPO is based on the prototype neurovirulent poliovirus (PV) serotype 1 (Mahoney) 
(Figure 1A), which is derived from a stool isolate obtained from a non-symptomatic carrier in 
Cleveland in the 1940s. Throughout the 1940s, PV type 1 (Mahoney) was subjected to serial 
passage, either in non-human primates or in tissue culture cells derived from various non-
human primate tissues, which yielded the type 1 live-attenuated (Sabin) vaccine strain (PV1S; 
Figure 1B). PV1S is the preferred agent for routine, infant vaccination in the world. The PV1S 
vaccine was modified by exchange of its internal ribosomal entry site (IRES) with its counterpart 
from human rhinovirus type 2 (HRV2), generating PVSRIPO (Figure 1C). PVSRIPO is 
characterized by loss of the inherent neurovirulence of poliovirus 8-11. 
 
Figure 1.  The genetic structure of PVSRIPO (C) and its precursors type 1 (Mahoney) (A), and PV1S (B). 
7.2.2 PVSRIPO Oncolysis 
PVSRIPO’s anti-neoplastic potential is due to a series of (i) direct lytic effects on tumor cells; (ii) 
presentation of tumor-associated antigens in a highly adjuvanted context; (iii) pro-inflammatory 
and danger signals stemming from tumor destruction and activation of an antiviral type 1 
interferon response; (iv) infection and pro-inflammatory activation of dendritic cells and tumor 
associated macrophages; (v) durable anti-tumor immunity evoked by effector cytotoxic T 
lymphocyte responses 12 (Figure 2). The principal elements determining PVSRIPO tumor 
tropism, tumor-specific cell killing, neuronal incompetence/safety, and immunogenicity are well 
established empirically 13-15. 

 
 
 
Figure 2.  Pathways involved in PVSRIPO oncolytic immunotherapy. PVSRIPO oncolytic immunotherapy is 
based on combined cytotoxic and immunogenic effects on tumor cells (left), pro-inflammatory effects on antigen-
presenting cells (middle) and the generation of adaptive anti-tumor immunity (right). 
7.2.3 PVSRIPO Tumor Tropism 
Every aspect of PVSRIPO oncolytic immunotherapy is dominated by its relationship with its 
host cell receptor, CD155 16. CD155 is broadly expressed in neuroectodermal malignancies, 
e.g. MG. Its expression has been empirically confirmed in laboratory MG cell lines, primary 
patient explant MG cells, primary patient explant MG xenografts maintained in athymic mice, 
and patient MG tissues obtained during surgery 17. CD155 is expressed on tumor cells proper, 
on proliferating tumor vascular cells, and on macrophages and dendritic cells 18,19. 
7.2.4 PVSRIPO Tumor-Specific Cell Killing 
Infection of tumor cells with PVSRIPO results in their swift destruction, due to very early 
translation of highly cytotoxic viral proteins immediately after entry of the viral RNA into infected 
host cells 20. PVSRIPO achieves this through an unorthodox mechanism of translation initiation 
21.  At eukaryotic mRNAs, protein synthesis occurs upon recruitment of a multi-partite protein 
complex at the canonical 5’ ‘cap’ structure.  Poliovirus RNAs are un-capped and, hence, rely on 
cap-independent translation for viral protein synthesis.  This involves direct recruitment of 
ribosomal subunits via binding of the eukaryotic initiation factor (eIF) 4G to viral RNA (Figure 1) 
22.  eIF4G-binding occurs via the IRES, a cis-acting genetic element located in the 5’ 
untranslated region of the viral RNA. In tumor cells, this process is highly efficient, due to un-
hinged signal transduction networks that greatly favor ribosome recruitment to viral RNA 23. 
 
There has been no evidence of PVSRIPO causing infectious polio; PVSRIPO does not infect 
normal brain cells and is incapable of driving viral translation in normal brain cells.  8 This is due 
to profound neuronal incompetence of the foreign HRV2 IRES in its genome. The foreign HRV2 
IRES cannot recruit eIF4G to the viral genome, precluding viral protein synthesis in neuronal 
cells/the normal brain 24. By blocking this crucial, early step in the infectious cycle specifically in 
neurons, PVSRIPO neurovirulence is eliminated; therefore, PVSRIPO has been deemed 
acceptable for intracerebral inoculation. 
7.2.5 Pre-clinical Experience 
In animal tumor models, oncolytic polioviruses elicit efficient anti-neoplastic effects resulting in 
tumor regression and, eventually, destruction 20.  There is histologic evidence for direct, virus-
mediated tumor cell killing and indirect, host-mediated inflammatory responses directed against 
tumor.  PVSRIPO was subjected to extensive dose-range finding, toxicology, biodistribution, 
shedding and neutralizing antibody tests with intrathalamic inoculation of up to 5 x 109 TCID50 

 
 
of PVSRIPO in M. Fascicularis 25. These revealed: (i) absence of morbidity and mortality; (ii) 
absence of neuropathological signs consistent with virus-induced CNS damage; (iii) absence of 
virus dissemination from the brain or viremia; (iv) absence of extraneural replication; (v) 
absence of shedding with saliva, urine or stool; (vi) presence of a neutralizing antibody 
response. 
7.2.6 Clinical Experience 
The first in human oncolytic PV, PVSRIPO, therapy was initiated at the PRTBTC at Duke 
University Medical Center in early 2012 as a Phase I trial in patients with histologically 
confirmed recurrent WHO grade IV malignant glioma (glioblastoma or gliosarcoma) (size of 1-
5.5 cm).  Key inclusion criteria in this ongoing clinical trial include: ≥ 18 years, adequate 
performance status/organ function, and prior PV immunity. Key exclusion criteria include: 
pregnant/breast-feeding females, those who have received radiotherapy less than 12 weeks 
prior, and those with known immune dysfunction or febrile and/or other systemic illnesses. 
Patients are given a booster of inactivated PV vaccine > 1 week prior to PVSRIPO infusion.  
Consented patients first undergo biopsy of the lesion to confirm recurrence and then have a 
catheter placed into the tumor ≥ 1 cm away from the ventricles. The catheter is tunneled 5 cm 
under the scalp and connected to an infusion pump. The agent is then infused at a rate of 0.5 
mL/hour over a period of 6.5 hours. In the IND toxicology studies in non-human primates 
(NHP), no dose-limiting toxicities (DLTs) were observed and the MTD was not reached. The 
starting dose in this adult first-in-human trial was 1 x 108 TCID50, which is 1/10th of the highest 
non-toxic dose in NHPs from the definitive, IND-directed toxicology study and 1/50th of the 
highest non-toxic dose in NHPs from the dose-range finding toxicology study. In the dose 
escalation portion of the Phase I trial, cohorts of patients were dosed at 1 x 108 TCID50 (dose 
level 1), followed by 3.3 x 108 (level 2), 1 x 109 (level 3), 3.3 x 109 (level 4), and 1 x 1010 (level 
5). The rationale and outcome associated with subsequent dose reductions to 3.3 x 108 (dose 
level 2), 5 x 107 (dose level -1), and 1.0 x 107 (dose level -2) are described below. 
 
A total of 61 patients were treated on the Phase I study. The first 15 patients, who were treated 
at dose levels 1 through 5, constituted the dose escalation portion of the study.  Due to 
observed tumor inflammation at the higher dose levels that could potentially have been related 
to treatment or tumor growth, the dose was de-escalated and six additional patients were 
treated at dose level 2. Due to the continuing requirement for steroids in all of the additional 
patients treated at dose level 2 because of tumor inflammation, due to treatment and/or tumor 
growth, a decision was made to de-escalate to dose level -1 (5 x 107 TCID50).  Despite the fact 
that subjects on dose level -1 were able to remain off significant doses of steroids, it was 
believed that the subjects benefiting the most from PVSRIPO had been those who had 
experienced minimal inflammation. 
 
As such,  the dose of PVSRIPO was reduced to dose level -2 (1.0 x 107 TCID50) on 6/30/2016 
with the goal of limiting the occurrence of undesirable burden from the inflammation and its 
treatment on as many subjects (including caregivers) as possible. Based upon additional 
animal studies, dose level -2 (1.0 x 107 TCID50) is considered to be a therapeutic dose. This 
dose reduction was not made due to concerns for the safety of the patients on dose level -1, 
but due to the hypothesis that dose level -2 would cause less inflammatory reaction to the virus 
inoculum and result in better survival and treatment response.   
 
Based upon both pre-clinical tests highlighting the role of virus dose in PVSRIPO:host relations 
and clinical data collected thus far on subjects who were treated with either dose level -1 or 
dose level -2, the anticipated dose going forward is dose level -1.  There does not appear to be 
evidence of pre-clinically or clinically important differences between the 2 dose levels, and most 
importantly no added benefit attributable to the reduced dose level -2.  The need for initial 
treatment of cerebral inflammation either secondary to PVSRIPO or due to tumor growth, and 
the toxicity profile for the two dose levels, is comparable.  As of 10/15/2019, eight patients 

 
 
treated with PVSRIPO at any dose level remain alive with survival from initial PVSRIPO 
infusion ranging from 89 months to 30 months.   
 
In June 2017, the Phase II study of PVSRIPO opened for patients diagnosed with recurrenty 
WHO Grade IV malignant glioma.  Initially, this study was a single-site (Duke), randomized 
study that included 2 arms (PVSRIPO alone vs. PVSRIPO in combination with single dose of 
lomustine 8 weeks after PVSRIPO infusion).  The lomustine arm was removed from the study in 
February 2019 after it was observed that patients randomized to receive lomustine often 
experienced a significant decline in blood counts, as expected. However, this could lead to the 
postponement of Avastin® (bevacizumab) therapy, often utilized for initial control of cerebral 
edema associated with the PVSRIPO anti-tumor response. The study is now a single arm, non-
randomized study.  In addition, the study was amended to add additional sites in October 2017.  
The study is remains open to enrollment at Duke, as well as at five other sites.  As of 
10/25/2019, a total of 89 patients have been treated on study across 4 centers.  
7.3 Study Purpose/Rationale 
Outcome for children with recurrent malignant glioma, especially GBM, is dismal with currently 
available therapeutic modalities. The blood-tumor barrier is a major impediment to therapy and 
approaches that bypass this barrier may be required. Oncolytic immunotherapy is a promising 
biologic approach to treatment that not only induces viral mediated tumor destruction, but also 
harnesses a complex immune response that can serve in disease control. The genetically engineered 
oncolytic PV (PVSRIPO) has tropism for tumor cells by virtue of expression of CD155 in tumor cells. 
To facilitate concentration of the therapeutic agent at the tumor site, while minimizing systemic 
exposure, PVSRIPO will be delivered directly into the tumor. Significant pre-clinical anti-tumor activity 
of PVSRIPO has been observed in several rodent tumor models and in vitro. PVSRIPO was devoid of 
neuropathogenicity when injected into the thalami of non-human primates even at doses as high as 5 
x 109 TCID. Also, the phase I trial of PVSRIPO in adults with recurrent WHO grade IV malignant 
glioma still in the follow-up phase did not yield evidence of viral encephalomyelitis, poliomyelitis or 
meningitis with a proportion of patients enjoying prolonged disease survival up to 58+ months. The 
main toxicity attributable to PVSRIPO has been post-treatment peri-tumoral inflammation that has 
required prolonged steroid therapy and/or low dose bevacizumab to control edema. CD155 
expression in pediatric GBM has been confirmed in 10 tumor samples. Therefore, a phase Ib study 
should be conducted in this high-risk pediatric population to evaluate feasibility, safety, and 
preliminary evidence of efficacy of the selected adult dose (5 x 107 TCID).   
 
The Food and Drug Administration (FDA) recommended that the pediatric trial initially enroll patients 
12 years of age and older and up to and including 18 years of age.  We are including a number of 
WHO grade III and IV malignant brain tumor histologies and tumor locations in this trial.  Specifically, 
eligible patients may include patients with recurrent WHO Grade III gliomas, such as anaplastic 
astrocytoma, anaplastic oligoastrocytoma, anaplastic oligodendroglioma, anaplastic pleomorphic 
xanthoastrocytoma, or ependymoma, as well as patients with WHO Grade IV gliomas, including 
glioblastoma or gliosarcoma.  CD155 is avidly expressed in medulloblastoma, atypical teratoid 
rhabdoid tumor (ATRT), and embryonal tumors 26, so these histologies will be included here as well.   
 
Given that the incidence of embryonal tumors, ependymoma, and glioma peaks at ages lower than 
the initial eligible age for this clinical trial (≥ 12 years old), the study has experienced relative 
challenges to enrollment during its first two years.  The incidence of the aforementioned tumor types 
is higher in younger children approximately 5 to 9 years of age and then continuously falls after 9 
years old to a nadir in early adulthood 27,28  In addition, the biology of brains in children aged 4 to 11 
and the biology of the predominant types of brain tumors seen in this age range (i.e., glioma, 
ependymoma, and medulloblastoma) are similar to that of children who are 12 and over 1. Tumors in 
children younger than 4 years old tend to have different genotypes and thus behave differently than 
tumors in children older than 4 years old 1.  There are currently active pediatric clinical trials in the US 
investigating the use of different oncolytic viruses, such as herpes simplex virus-1 (HSV-1) 
(NCT03911388 and NCT02451845) and a modified measles virus (NCT02962167), in a similar 
pediatric population.  These trials also deliver the oncolytic virus via intratumoral administration and 

 
 
include a population of children ranging in age from 3 to 18 years old in the case of aforementioned 
HSV-1 trial and children and adults ranging in age from 1 to 39 years old in the case of the modified 
measles virus. 
 
As of June 26, 2019, five patients consented to participate in the study with three patients receiving 
PVSRIPO (i.e., 2 patients failed screening).  Of the three treated patients, one was 9 years and 
treated under a planned deviation; one was 14 years old; and, one was 19 years old.  Serious 
adverse events in these three patients that were considered possibly related to PVSRIPO include 
Grade 3 headache in two of the patients.  Other toxicities considered possibly, probably, or definitely 
related to PVSRIPO have included Grade 1 eye disorders (intermittent double vision and difficulty 
seeing right-sided peripheral vision) (N=1), Grade 1 gastrointestinal disorders (nausea and vomiting) 
(N=1), Grade 1 general disorders (edema face) (N=1), Grade 1 general disorders (fatigue) (N=2), 
Grade 1 metabolism/nutrition disorders (anorexia) (N=1), Grade 1 metabolism/nutrition disorders 
(night sweats) (N=1), Grade 1 nervous system disorders (memory impairment) (N=1), and Grade 1 
nervous system disorders (difficulty comprehending) (N=1).  One of the three patients discontinued 
the study due to a secondary malignancy resulting from prior chemotherapy, while another patient 
discontinued due to disease progression; both of these patients are now deceased.  One of the three 
patients discontinued the study due to re-resection of tumor and was being followed for survival as of 
June 26, 2019.   
 
It is therefore reasonable to consider expanding enrollment of the clinical trial of oncolytic PV to 
younger children who have been diagnosed with a recurrence of eligible tumors who are between the 
ages of 4 and 12 years old.  Accrual of all pediatric patients to this study was staggered so that no 
more than one patient was treated every 14 days for the first 5 patients enrolled.  Staggered entry to 
this study was intended to minimize the number of pediatric patients exposed to PVSRIPO before 
adequate information about the sequelae of the inflammatory process within pediatric patients is 
known.  Given limited enrollment over two years, the experience of treating 3 patients on study who 
range from 9 to 19 years of age, and the toxicity profile of PVSRIPO treatment in these patients, we 
believe expansion of the study to children as young as 4 years old will be safe and tolerable and will 
improve the ability to enroll on this pediatric study for patients diagnosed with recurrent supratentorial 
WHO Grade III malignant glioma or WHO Grade IV malignant glioma.   
8 OBJECTIVES AND ENDPOINTS 
Table 1.  Objectives and Endpoints 
 
Objective 
Endpoint 
Analysis 
Primary 
Confirm the safety of the 
selected dose of PVSRIPO 
when delivered 
intracerebrally by convection-
enhanced delivery (CED) in 
children with recurrent Grade 
III or IV MG 
Percentage of patients that 
experience unacceptable 
toxicity during the 14-day period 
post-PVSRIPO treatment. 
 
Proportion of patients who 
require low-dose bevacizumab 
or steroid treatment due to an 
inflammatory reaction 
secondary to an immune 
response to PVSRIPO 
treatment will be calculated. 
See Section 
15.4 
Secondary 
Estimate Overall Survival 
(OS) in children with 
recurrent WHO Grade III and 
IV malignant glioma treated 
with a single dose of 
PVSRIPO 
Proportion of patients alive 2 
years after PVSRIPO treatment 
See Section 
15.5 

 
 
Exploratory 
Describe changes visualized 
on imaging due to 
intratumoral inoculation of 
PVSRIPO 
Description of imaging changes 
See Section 
15.6 
Other 
Exploratory 
Describe changes in tumor 
volume visualized on 
imaging after intratumoral 
inoculation of PVSRIPO 
Change from baseline in tumor 
volume based on imaging 
See Section 
15.7 
Other 
Exploratory 
Assess immunologic 
responses in peripheral 
blood  and in serum 
Change from baseline in 
immune function 
See Section 
15.7 
Other 
Exploratory 
Identify genetic predictors of 
response or failure of 
response to treatment with 
PVSRIPO 
Identification of genetic markers 
as predictors of response 
See Section 
15.7 
 
9 INVESTIGATIONAL PLAN 
9.1 Study Design 
9.1.1 
Catheter Implantation 
Systemic delivery of high molecular weight therapeutic agents to brain tumors is limited by the blood-
brain barrier and increased interstitial pressure within the tumor.  PV CNS invasion after intravenous 
administration via trans-endothelial passage is inefficient 29.  Intratumoral delivery bypasses these 
physiologic barriers and concentrates the therapeutic agent at the tumor site while minimizing 
systemic exposure.  Therefore, PVSRIPO will be delivered directly into the tumor.  A stereotactic 
biopsy will be performed prior to virus administration for frozen section confirmation of viable tumor 
and further analysis (see Section 9.1.4 below).  The biopsy needle will be placed with stereotactic 
guidance by a Cosman-Robert-Wells, MRI-compatible, stereotactic head frame or a similar frameless 
device.  Collection of biopsy tissue for pathologic diagnosis will be performed under traditionally 
accepted conditions according to standard of care.  Up to three additional core biopsies will be 
obtained for molecular genetic testing as described in section 9.1.4. 
 
If this biopsy does not confirm recurrent tumor, the subject will be withdrawn from the study. 
 
Immediately following the stereotactically-guided tumor biopsy, a catheter [Vygon PIC-030 (Sophysa, 
Inc.; Crown Point, IN)] will be implanted in the operating room at a site the same or different from that 
used for the biopsy using sterile techniques under general anesthesia.  Implantation will occur at a 
coordinate selected by the operating surgeon with the assistance of the clinically-approved iPlan Flow 
(BrainLAB).  The catheter will be implanted at least 1 cm away from the ventricles. Based on our 
experience, a tumor ≤ 1cm from the ventricles can safely and feasibly have a catheter placed ≥ 1cm 
from the ventricles while minimizing the possibility of infusate entering the ventricles. The catheter will 
be tunneled beneath the scalp for a distance of at least 5 cm to aid in the prevention of infection. A 
CT scan may be used to confirm catheter placement post-operatively. 
9.1.2 
Agent Infusion 
Refer to the study Investigational Product Handling Plan (IPHP) for additional details regarding the 
infusate preparation and infusion procedure. The entire volume of the prepared infusate will be 
provided in a sterile infusion syringe by the investigational pharmacist and connected to the catheter 
under sterile conditions in the Pediatric Intensive Care Unit (PICU) just prior to beginning of infusion.  
Due to the complexity of scheduling all of the necessary components for the infusion (operating room 
time, pharmacy time, and radiology appointments), a +1 day window has been built into the study for 
the study drug infusion.  This means that the infusion is allowed to start the following day after the 

 
 
biopsy/catheter placement.  This will still be considered “day 0” with regard to the protocol and the 
timing of the subsequent events.  At the time of virus injection, emergency drugs, including 
epinephrine and diphenhydramine will be available and the neurologic status, oxygen saturation, and 
cardiac rhythm will be monitored.  Drug infusion will occur in the PICU so that all other emergency 
facilities will be available.  Patients will be treated with a prophylactic antibiotic prior to biopsy and 
catheter insertion per DUHS neurosurgical standard practice.  
 
Based on our own experience, previously published reports 5 and IRB- and FDA-approved trials using 
similar infusion techniques, patients will be infused at a rate of 0.5mL/hr.  A Medfusion 3500 or 3010a 
infusion pump will be pre-programmed to a delivery rate of 500 µL/hr.  The infusate  will be loaded in 
a syringe into the syringe pump at the initial onset to avoid any interruptions in the infusion.  The total 
amount of the infusate delivered to the patient will be 3 mL.  The infusion pump will be programmed 
for delivery of the 3.0mL at a rate of 0.5ml/hr, over approximately 6.5 hours. The catheter itself (30 cm 
length, 1 mm interior diameter) cannot be preloaded with virus suspension. To account for this 
catheter dead space, the pump will be stopped when the delivered amount is between 3.125 to 3.250 
mL. The infusion will be performed using a Medfusion 3500 or 3010a (Smiths Medical ASK, Inc., 
Minneapolis, MN) syringe infusion pump.    The catheter will be removed in the PICU following the 
post-PVSRIPO infusion MRI. No sedation is required for catheter removal. 
 
The infusion catheter (PIC 030) and infusion tubing (PIT 400) are manufactured by Sophysa, Inc. 
(Crown Point, IN).  The Infusion Catheter Kit is a 30 cm clear, open-ended catheter (1.0 mm ID/2.0 
mm OD) with 1 cm markings for 20 cm.  The catheter comes with a 30 cm stainless steel stylet, a 
barbed female luer lock with cap and a stainless steel trocar.  The Infusion Tubing Kit consists of a 3-
way stopcock connector with air filter, 4 m of microbore tubing with antisiphon valve, a red, vented 
cap and a white luer lock cap.  The catheter products are packaged sterile and non-pyrogenic and are 
intended for single (one-time) use only.   
 
Acute Reaction. Any acute reaction symptoms determined to be an acute reaction to the study drug 
will be managed by the PICU and pediatric neurosurgical teams. 
9.1.3 
Gadolinium Distribution Quantitation (applies only to patients treated 
prior to October 31, 2019) 
This gadolinium tracking MRI procedure has been removed and applies only to patients treated prior 
to October 31, 2019. Magnevist® is no longer being manufactured and, as such, is no longer being 
included in the infusate.  
 
MRI imaging, within 4 hours of the completion of infusion, was registered to define the shape and 
position of the contrast agent distribution relative to the patient’s brain anatomy in patients treated 
prior to October 31, 2019.  Because enrollment criteria for these patients was limited to patients aged 
≥ 12 years old, sedation was not required for the MRI. While gadolinium–diethylene triamine 
pentaacetic acid (Gd-DTPA) is a widely available MRI contrast agent, there has been speculation that 
its small molecular weight (938 Dalton) could limit its ability to predict the distribution of the larger 
molecules typically infused therapeutically with CED [reviewed in Reardon et al., 2011 30].  
Development of large molecule tracers labeled with gadolinium (Gd) has been problematic, but it has 
been stipulated that infusion of low molecular weight Gd-DTPA can predict the distribution of larger 
molecules by systematic post-infusion manipulation of the images based on theoretical differences in 
the predicted distribution of the Gd-DTPA and the therapeutic drug being infused 31.  This has been 
confirmed by simultaneously infusing a patient with a supratentorial recurrent malignant glioma with 
an epidermal growth factor receptor (variant III) (EGFRvIII)-targeted immunotoxin in combination with 
124I-human serum albumin (HSA) [to permit positron emission tomography (PET) imaging] and Gd-
DTPA.  Gd-DTPA co-infusion provided direct information about the distribution of large molecules 
with high resolution 31.  In combination with fluid-attenuated inversion recovery (FLAIR) imaging, Gd-
DTPA co-infusion provides additional information about leak into cerebrospinal fluid spaces and 
resection cavities 32.  

 
 
9.1.4 
Biopsy Sampling and Analyses 
Biopsy material will be obtained from tumor tissue prior to virus administration.  This tissue material 
will be subjected to routine histology to confirm tumor recurrence by the study neuropathologist, Dr. 
R. McLendon or his designate.   
 
Molecular genetic tests will also be conducted on extracts of tumor cells from the protocol-specified 
biopsy prior to PVSRIPO infusion.  After acquiring sufficient tissue for standard clinical pathologic 
testing, up to three additional core biopsies will be obtained, if possible.  These additional core 
biopsies will be frozen in optimal cutting temperature (OCT) fixative and kept at liquid nitrogen 
temperature.  They will be used for genetic analysis, including full genome or full exome sequencing 
as well as other molecular genetic testing. The goal of these molecular genetic tests is to identify 
genetic predictors of response or failure of response to treatment with PVSRIPO.  The molecular 
genetic testing will include, but is not limited to, DNA sequencing, gene amplification, and gene 
expression.   
 
The goal of these molecular genetic tests is to identify genetic predictors of response or failure of 
response to treatment with PVSRIPO.  The amount of tissue required for this testing should be 
sufficient to yield a minimum of one microgram of DNA.  
 
Following PVSRIPO administration and outside the context of this protocol, patients may undergo 
tumor resection or biopsy as medically indicated for reasons not part of the investigative protocol.  
Should subjects have a resection at a later time, we will request samples of this resected tissue for 
tissue analysis.  If subjects agree to this by opting in on the informed consent, portions of resected 
tissue will be delivered to the study neuropathologist, Dr. R. McLendon or his designate, for 
histopathological analyses and to Dr. M. Gromeier or his designate for correlative molecular analyses.  
Tissue will be processed and stored in the Brain Tumor Biorepository at Duke (Duke IRB 
Pro0007434).  Samples are assigned a patient de-identified biorepository number.  The Biorepository 
database is Duke IRB-approved, accessible by Biorepository personnel only, and includes relevant 
data associated with the sample, such as Biorepostiory number, treatment with PVSRIPO, dates 
related to the sample, diagnosis, age, race, gender, pathology, and any other relevant information. 
Samples from tissue collected and stored for the aforementioned purpose may be analyzed within 
Duke at the request of the study investigators or may be sent to outside collaborators for analysis.  
Samples will always be identified by Biorepository number and/or Study ID number. Investigators will 
submit any such requests to the Biorepository using their internal tissue request form.  Additional 
pathology analyses may include, but is not limited to, EGFRvIII and EGFRwt status, TERT, IDH 1 and 
2, and MGMT IHC and MGMT promoter methylation, if a sufficient amount of tissue remains after 
standard clinical pathologic testing. 
 
Tissue that is processed and stored by the Brain Tumor Biorepository for this study will be maintained 
indefinitely in the Biorepository for future testing until such time as the tissue supply designated for 
research is exhausted. 
9.1.5 
Definition of Unacceptable Toxicity 
Toxicities will be graded according to the NCI CTCAE version 4 criteria.  Any Grade 3 or any Grade 4 
toxicity that is not reversible within 2 weeks, any life-threatening event deemed related to the study 
drug, or any treatment-related death will be considered unacceptable.   
 
Any Grade 2 or higher autoimmune toxicities, particularly those affecting vital organs (e.g. cardiac, 
renal, CNS), will be considered an unacceptable toxicity, if it occurs within 2 weeks post-infusion. 
Grade 3 or higher surgical complications from insertion of the catheter are considered an 
unacceptable toxicity.  
 

 
 
Exceptions to these unacceptable toxicities are as follows: 
 
• 
Events associated with the biopsy procedure/catheter placement: Seizures or hemorrhages 
occurring during anesthesia or the biopsy/catheter insertion proper prior to administration of 
the agent if Grade 2 or lower. 
 
• 
Seizures: Due to the nature of the disease under investigation in this protocol, patients may 
have pre-existing seizures or be susceptible to new seizures as a result of the underlying 
disease process.  Although seizures may be defined as Grade 3 or 4 toxicities under NCI 
CTC, and will be reported as such in this protocol, seizures will not be considered an 
unacceptable toxicity if, in the opinion of the site Principal Investigator they have not 
increased in frequency or can be attributed to another recognized cause of increasing seizure 
frequency such as sub-therapeutic anti-convulsant levels or biopsy proven tumor 
progression. 
 
• 
New or worsening neurologic deficits: Due to the nature of the disease under investigation in 
this protocol, patients may have an increase in pre-existing neurologic deficits or may 
develop new neurologic deficits as a result of tumor invasion. A new neurologic deficit, which 
resolves within 2 weeks after initiation of medical therapy, will not be considered an 
unacceptable toxicity. New neurological symptoms will not be an unacceptable toxicity if they 
can be ascribed to tumor progression (e.g. documented with histopathologic analyses of 
biopsy tissue), or they respond to treatment (e.g. oral steroids) within 2 weeks.  
 
• 
Thromboembolism:  While deep vein thrombosis (DVT) is not as common in children as in 
adults with malignant glioma, patients may have undiagnosed pre-existing DVTs or be 
susceptible to the development of DVTs due to the underlying disease process.  Although 
DVT may be defined as Grade 3 or 4 toxicities under NCI CTC, and will be reported as such 
in this protocol, DVT will not be considered an unacceptable toxicity in this protocol. 
 
• 
Syndrome of Inappropriate Antidiuretic Hormone (SIADH): Due to the high incidence of 
SIADH in this patient population, patients may be susceptible to the development of SIADH 
due to the underlying disease process.  Although SIADH may be defined as Grade 3 toxicity 
under NCI CTC, and will be reported as such in this protocol, SIADH will not be considered 
an unacceptable toxicity in this protocol unless it is refractory to medical management. 
 
• 
Muscle Weakness and Weight Gain:  Due to the high incidence of muscle weakness and 
weight gain in patients taking steroids in this patient population, patients may be susceptible 
to the development of muscle weakness or weight gain which is due to steroids alone.  
Although muscle weakness may be defined as Grade 3 or 4 toxicity and weight gain  20% 
may be defined as Grade 3 toxicity under NCI CTC, and will be reported as such in this 
protocol, muscle weakness or weight gain will not be considered an unacceptable toxicity in 
this protocol if the patient has required steroids greater than physiologic doses in the interval 
between the immunization and the development of the toxicity. 
9.1.6 
Safety Considerations 
• 
Surgical Complications: The stereotactic tumor biopsy and catheter implantation procedure 
carries a risk for cerebrospinal fluid leak, infection, hemorrhage, loss of neurologic function, 
non-neurologic complications and death. These risks depend primarily on the preoperative 
condition of the patient, the size and location of the tumor and associated diseases. The 
potential risk for the patient will be discussed in detail with the patient and family. 
 
• 
Anesthesia:  Patients undergoing general anesthesia may be subjected to associated risks 
including pneumothorax, pneumonia, airway injury, hypotension, myocardial infarction, 
stroke, hepatic and renal injury and death. 
 

 
 
• 
Poliomyelitis:  PVSRIPO has been tested in non-human primates according to the WHO 
standardized monkey neurovirulence tests. These tests revealed the absence of 
neuropathogenic properties, evident as failure to induce symptoms of poliomyelitis in non-
human primates after intracerebral inoculation of virus. However, PVSRIPO is a replication-
competent viral agent that, in principle, retains the potential to cause motor neuron damage. 
Possible complications include transient or permanent mono- or paraparesis, paralysis, and 
life-threatening respiratory insufficiency. 
 
• 
Virus Recombination and Mutation:  PVSRIPO retains the ability to alter its genome during 
replication upon administration to patients. Various mechanisms are known to lead to genetic 
adaptation, including spontaneous mutagenesis and recombination that may give rise to viral 
progeny with changed properties. Empirical studies in tissue culture and laboratory animals 
demonstrated that prolonged passaging in GBM cells does not select for genetic changes in 
viral progeny. However, the occurrence of such events cannot be categorically excluded in 
patients receiving intracerebral PVSRIPO. Genetic changes may cause an altered phenotype 
of PVSRIPO, including adaptation to improved virus replication in the normal CNS. 
 
• 
Long-Term Sequelae of Virus Injection:  PVSRIPO does not encode foreign genetic material; 
PVs are unable to insert viral genetic material in the host genome and PVs are incapable of 
inducing latent or chronic CNS infection. Therefore, PVSRIPO is unable to induce long-term 
expression of foreign polypeptides or long-term persistence. Thus, there are no long-term 
neurologic consequences to intracerebral PVSRIPO administration in study subjects. For 
these reasons, no specific measures to analyze the potential for persistence of virus 
replication in the CNS of long-term survivors are indicated. 
 
• 
Gastrointestinal Infection and Virus Excretion:  After oral uptake, PV replicates in the 
gastrointestinal tract and is excreted by infected individuals’ stool. Gastrointestinal viral 
replication usually is asymptomatic, but may cause mild symptoms of gastrointestinal 
discomfort. Tests of PVSRIPO in non-human primates and in subjects from the phase I study 
suggest that excretion of PVSRIPO does not occur after intracerebral inoculation in 
immunized/boosted patients. This is based on thorough testing in patients in our Phase 1 
clinical trial and much historical evidence from investigations in non-human primates.  
 
• 
Cerebral Edema and Increased Intracranial Pressure:  Cerebral edema may be secondary to 
the disease process itself, the surgical procedure, necrosis from previous radiation, or 
inflammation due to immune infiltration of the brain or destruction of tumor cells. Symptoms 
may include, but are not limited to, severe headache, confusion, lethargy, unresponsiveness, 
coma, or focal neurological deficits. Patients will be monitored throughout the course of the 
study and upon any signs or symptoms of cerebral edema, may have their steroid doses 
increased or receive treatment with an osmotic diuretic, or surgical decompression. Edema 
that fails to respond to aggressive therapy may lead to permanent neurological impairment. 
The probability of this risk can be predicted to some degree based upon tumor size, location, 
pre-operative neurological impairment, and post-operative course prior to virus 
administration. 
 
Special Consideration:  In the event that a patient demonstrates neurologic or radiographic 
signs suggestive of an inflammatory reaction secondary to the immune response triggered by 
PVSRIPO that requires a dose of dexamethasone above 4 mg a day after the first 2 weeks 
post PVSRIPO infusion, dexamethasone will not be increased any further. Instead, patients 
will be treated with bevacizumab 7.5 mg/kg IV every 3 weeks and neuroimaging (MRI) after 3 
doses (every 9 weeks +/- 2 weeks) to assess if further treatment with bevacizumab is needed 
to control the cerebral inflammation. Bevacizumab will not be provided by the study. Every 
attempt should be made to discontinue dexamethasone.  Once patients start bevacizumab, 
the follow-up visit schedule will be adjusted to coincide with the timing of the bevacizumab 
instead of the prior PVSRIPO infusion.  Therefore, patients will return approximately every 9 
weeks for an evaluation with MRI.   

 
 
 
Per the packet insert for bevacizumab dated 12/2017, the safety and effectiveness of 
bevacizumab have not been established in the pediatric population 33.  Bevacizumab is 
currently not approved by the FDA for use in patients under the age of 18 years. “In published 
literature reports, cases of non-mandibular osteonecrosis have been observed in patients 
under the age of 18 years who have received Avastin 33.”  In initial Phase I and II clinical 
trials, four potential bevacizumab-associated safety signals were identified: hypertension, 
proteinuria, thromboembolic events, and hemorrhage.  Additional completed Phase II and 
Phase III studies of bevacizumab in adults, as well as spontaneous reports have further 
defined the safety profile of this agent. Bevacizumab-associated adverse events identified in 
Phase III trials in adults include congestive heart failure (CHF) primarily in metastatic breast 
cancer, gastrointestinal perforations, wound-healing complications, and arterial 
thromboembolic events (ATEs). These and other safety signals are described in further detail 
in the bevacizumab package insert. 
 
If there are adverse events or other circumstances prohibiting the use of bevacizumab, we 
will use corticosteroids or surgery, or other interventions deemed more appropriate for the 
patient by the treating physician, if needed, to treat the inflammatory reaction secondary to 
PVSRIPO. 
 
• 
Risk of Infection:  The intracerebral catheter placement and infusion may include the risk of 
infection.  However, similar procedures including stereotactic biopsy and ventriculostomy 
placement are commonly used in the routine clinical care of patients with malignant brain 
tumors with a very low and acceptable rate of infection.  In the most extreme situation, 
however, infection may lead to systemic bacterial/fungal sepsis and possibly death.  The risk 
of infection will be minimized though by performing catheter implantations in the Operating 
Room.  Patients will be monitored throughout the course of the study for any signs and 
symptoms of infection.  If an active infection is suspected, patients will be cultured and 
treated with appropriate antibiotics. 
 
• 
Risk of Phlebotomy:  Drawing blood or inserting an intravenous catheter into an arm vein may 
result in bruising or swelling in the area of the insertion, bleeding at the site of the needle 
puncture, light headedness, fainting and very rarely, local infection, which may be severe. 
These risks are reduced by the fact that the blood will be drawn by a qualified physician, 
nurse or phlebotomist (a professional trained to draw blood). 
 
• 
Risks of MRIs:  Risks and/or discomforts associated with MRI scans include anxiety produced 
from being in a tight, enclosed space (claustrophobia). In addition, the machine operates 
using a large and powerful magnet, which attracts certain metals. Therefore, people with 
these metals in their bodies (specifically pacemakers, infusion pumps, metal aneurysm clips, 
metal prostheses, joints, rods or plates) will be excluded from the study. Patients will also be 
checked to make sure that they do not bring any metal objects into the MRI facility.  Dental 
fillings are less affected by the magnetic fields generated and are therefore permitted.  It will 
be asked that patients let the physicians conducting this study know of any metal in their 
bodies other than dental fillings.  Depending upon the age of the patient, sedation for imaging 
may be needed.  Risks of sedation include an allergic reaction, aspiration, and over-sedation. 
Use of an IV may cause bruising.  Occasionally, an infection develops at the IV site. 
 
• 
Allergic Reactions to Contrast Agents:  During the MRI, patients will be given a contrast 
agent. The agent is given routinely to obtain enhanced MRI scans of the brain. The agent is 
administered through the vein and requires the placement of an IV catheter.  The catheter 
placement is similar to drawing blood except that the catheter remains in the vein during the 
time the agent is actively delivered. The risks of a blood draw and insertion of a catheter are 
similar. There have been a few, rare cases of allergies to the agent used in MRI contrast 
enhanced scans. Patients with any known severe allergies (anaphylaxis) to contrast agents 
will be excluded from the study. Patients with mild allergies (i.e., rash only) will be pretreated 

 
 
with Tylenol and Benadryl prior to injection of the contrast agent. In addition to the gadolinium 
(contrast agent) being given for MRI contrast, it will also be co-infused with the study drug to 
assess infusion distribution.  It is not FDA approved for gadolinium to be administered in this 
way.  The potential risks of intracerebral infusion of gadolinium contrast agents are not 
completely known but are believed to be small. Risks for infusion of gadolinium contrast 
agents intrathecally for procedures such as cistemography have been somewhat better 
studied, and were recently summarized by Selcuk et al (2009) 34. Encephalopathy, coma and 
seizures have been reported as side effects in case reports of accidental administration of 
large amounts of gadolinium contrast agents intrathecally in humans 32,35. When these 
contrast agents are used in appropriately low dose, however, the risk of intrathecal 
administration appears reasonably low. No neurological sequelae attributable to the 
procedure were detected in a series of 85 patients 34, in another series of 95 patients at a one 
year follow-up 36, or in any of 51 patients after over 4 years of mean follow-up in another 
study 37. Although these results cannot be extrapolated to the procedure proposed in the 
current research, they indicate that direct exposure of the brain to small amounts of 
gadolinium contrast agent is generally well tolerated. 
 
A rare but serious adverse reaction has been observed in patients that received a 
gadolinium-based contrast material during MRI examinations, a reaction called nephrogenic 
systemic fibrosis (NSF). Patients with kidney disease are at increased risk of developing 
NSF. NSF may cause skin thickening, joint pain and/or swelling. In rare cases NSF can lead 
to lung and heart problems and cause death. 
 
• 
Risks to Household Contacts of Study Subjects:  Primate toxicology studies showed that 
intracerebral infusion of PVSRIPO does not lead to extraneural dissemination or replication 
and, hence, is not associated with virus shedding. Therefore, and because PV transmission 
occurs via the fecal-oral route, the likelihood of unintended exposure of patient household 
contacts is exceedingly low. If accidental exposure occurred, it would equal the risk of 
exposure to any type 1 Sabin vaccine virus or vaccine virus derivatives. Thus, in essence, 
exposure with PVSRIPO is equal to oral immunization with a safe version of type 1 Sabin. 
Since type 1 Sabin vaccine virus or vaccine virus derivatives have to be considered part of 
the human environment, exposure to PVSRIPO would not represent an added risk beyond 
the possibility for exposure that already exists.  
 
• 
Unknown Risks:  The overall risk classification of this research is unknown.   
9.1.7 
Missed Doses 
This is not applicable, as the PVSRIPO is a single administration agent. 
9.1.8 
Concomitant Medications 
9.1.8.1 Steroids 
Corticosteroids should be used at the lowest dose to control symptoms of edema and mass 
effect, and discontinued, if possible. Use of corticosteroids should be recorded in the electronic 
database. 
9.1.8.2 Anticonvulsants 
Anticonvulsants drugs should be used or continued, if indicated. Use of such anticonvulsants 
should be recorded in the electronic database. 
9.1.8.3 Growth Factors 
Routine use of growth factors (i.e. G-CSF, GM-CSF, and erythropoietin) is not permitted.  
However, therapeutic use of G-CSF in patients with serious neutropenic conditions, such as 
sepsis, may be used at the investigator’s discretion. Use of such growth factors should be 
recorded in the electronic database. 

 
 
9.1.8.4 Anti-emetics 
The use of anti-emetics will be at the investigator’s discretion. Use of anti-emetics should be 
recorded in the electronic database. 
9.1.8.5 Proton Pump Inhibitors 
The use of proton pump inhibitors (PPI) (e.g. rabeprazole, omeprazole, pantoprazole, 
lansoprazole or esomeprazole) is allowed on this study. 
9.1.8.6 Febrile Neutropenia 
Febrile neutropenia should be managed according to the local institutional guidelines.  Measures 
include laboratory testing, blood and urine cultures, and institution of broad spectrum antibiotics. 
9.1.8.7 Pneumocystis jiroveci pneumonia (PJP) prophylaxis 
The use of medication (i.e., Bactrim) for PJP prophylaxis in patients on chronic steroids is 
recommended, but is at the investigator’s discretion. 
9.1.8.8 Neurosurgical Procedures 
If a neurosurgical procedure is required for a reason other than tumor progression (i.e. the onset 
of hydrocephalus), these procedures should be documented, but will not constitute criteria for 
declaring the patient “off therapy.” 
9.1.9 
Study Drug Blinding 
Not applicable 
9.2 
Rationale for Selection of Dose, Regimen, and Treatment 
Duration 
Dose: The dose for this trial was selected based on IND-directed toxicity studies 25 and on 
experience from an ongoing study in adults with recurrent WHO grade IV malignant glioma. Dose-
range finding and toxicity studies in non-human primates documented the absence of viral 
encephalomyelitis, poliomyelitis and meningitis with intracerebral injection of PVSRIPO up to a dose 
of 5 x 109 TCID50 25. In the adult WHO grade IV malignant glioma study, 4 patients were dosed at the 
maximum dose of 1 x 1010 TCID50 without signs of viral encephalomyelitis, poliomyelitis or 
meningitis. Given that the brain size of the target population for this trial (children and young adults 
aged 4-21) is nearly identical to that of adults, and that the human brain is ~10-times the size of the 
primate species used in the IND-directed toxicity studies (M. Fascicularis), the dose of 5 x 107 
TCID50 in this trial is 500-times below the maximum feasible safe dose in M. Fascicularis and 1000-
times below the maximum dose in the adult WHO grade IV malignant glioma study. The proposed 
dose of 5 x 107 TCID50 corresponds to the dose selected for future studies based on the Phase I 
adult WHO grade IV malignant glioma study.  If unacceptable toxicity that requires dose reduction 
occurs, according to Section 15.4.1, we may consider a PVSRIPO dose reduction to 3.3 x 106 
TCID50.   
 
Regimen and Treatment Duration: The treatment regimen for PVSRIPO is single intracerebral 
infusion, as in the ongoing adult WHO grade IV malignant glioma study.  
9.3 Rationale for Correlative Studies 
While tumor-selective PVSRIPO propagation is an important mediator of cytotoxicity, significant intra- 
and peritumoral inflammation likely ensues. This raises the possibility that an immunogenic response 
is being generated, certainly against the virus itself and with great likelihood against the tumor as well 
(Figure 2) 15. While the exact host immune response to PVSRIPO oncolysis is currently unknown (but 
is being investigated), host innate antiviral defenses are likely to trigger a broad immune effector 
cascade that needs to be examined in patients receiving PVSRIPO therapy.  Blood will therefore be 
collected for immune function studies before and at periodic intervals following treatment with 
PVSRIPO.  

 
 
9.4 Definition of Evaluable Subjects 
Any patient who has undergone surgery, placement of the catheter, and any dose of PVSRIPO will be 
evaluable for both toxicity and efficacy of agent. 
9.5 Early Study Termination 
This study can be terminated at any time for any reason by the PI-sponsor.  If this occurs, all subjects 
on study should be notified as soon as possible.  Additional procedures and/or follow up should occur 
in accordance with Section 12.6.2. 
10 STUDY DRUG 
10.1 Names, Classification, and Mechanism of Action 
PVSRIPO is a modified version of the serotype 1 live-attenuated (Sabin) PV vaccine (PV1) and its 
immunogenic properties and potential for long-term sequelae are expected to be similar.  PV1S has 
been safely administered to >10 billion individuals worldwide without untoward long-term sequelae.  
The only known effect of PV1S administration to human subjects is neutralizing immunity to PV.  
 
Figure 3.  Genetic structure of PVSRIPO. 
PVSRIPO is PV1S containing a heterologous IRES of HRV2 (Figure 3). The IRES is a cis-acting, 
non-coding genetic element within the 5’ untranslated region (UTR) of all enteroviruses and is 
essential for translation of the viral genome. PVSRIPO is a non-enveloped, positive-sense ssRNA 
virus with a genome of ~7300 nucleotides (nt) in length (Figure 3). PVSRIPO particles consist of a 
proteinaceous capsid composed of 60 copies of each of 4 capsid proteins (VP1-VP4) arranged in 
icosahedral geometry.  Since the coding regions for the viral polyprotein (giving rise to all viral 
polypeptides) of PVSRIPO and PV1S are the same, the physical structure of the viral capsid and all 
non-structural viral polypeptides of PVSRIPO and PV1S are identical. 
10.2 Packaging and Labeling 
PVSRIPO is formulated in 50 mM sodium phosphate in 0.9% sodium chloride, pH 7.4 with 0.2% 
human serum albumin.  It is provided in sterile, single use containers.  PVSRIPO was manufactured 
at the Biopharmaceutical Development Program/SAIC-Frederick at NCI-Frederick. Prior to October 
31, 2019, for tracking intratumoral/intracerebral distribution of the inoculum, the study agent 
suspension was  supplemented with gadolinium-DTPA diamide (Magnevist®) at a final concentration 
of 1 mM and Magnevist was stored at room temperature.  Magnevist® is no longer being 
manufactured in the US so has been removed from the infusate along with the corresponding post-
infusion MRI within 4h of infusion. 
10.3 Supply, Receipt, and Storage 
Refer to the study Investigational Product Handling Plan (IPHP) for details regarding supply, receipt 
and storage of study materials. The study agent and vehicle will be supplied directly to Investigational 
Chemotherapy Service (ICS) by the Sponsor (or designee).  The study agent will be shipped via 
approved methods in the appropriate packaging on dry ice. 
 

 
 
PVSRIPO should be stored long-term in an ultra-low temperature freezer (e.g., a ≤ -80°C freezer).  
Thawed vials should be kept at 4°C whenever possible. Thawed vials of PVSRIPO are stable at 4°C 
for 48 hours.  Once thawed, it is a clear colorless liquid with no evidence of particulates or foreign 
matter. 
10.4 Dispensing and Preparation 
Refer to the study IPHP for details of PVSRIPO dispensing and preparation. In brief, preparation of 
PVSRIPO will occur in a biosafety cabinet designated for viral vector agents.  For thawing, vials 
containing PVSRIPO should be removed from the -80°C freezer and kept at room temperature. 
PVSRIPO contained in the clinically intended delivery apparatus is stable at room temperature for 18 
hours, but should be used as soon as possible.  The vials contain a clear, aqueous solution, which 
does not require reconstitution.  Do not thaw with heating devices (waterbath, heat-block) as a 
loss of potency may result.  Do not expose vials to UV-light (in biosafety cabinets), as a loss 
of potency will result.  Thawed vials should not be re-frozen/thawed for later use, because potency 
may be lost.  Thawed vials/study syringes containing PVSRIPO, which were kept for >48 hrs at 4°C 
prior to the infusion procedure, should be discarded, as a loss of potency may have occurred.  
Thawed vials/study syringes containing PVSRIPO, which were exposed for >12 hrs at room 
temperature prior to the infusion procedure, shall be discarded as a loss of potency may have 
occurred.  For aliquot preparation, the agent will be thawed slowly on ice (~4C) and kept at that 
temperature.  The vials take approximately 30 minutes to thaw.  The manufacturer will provide the 
study agent’s potency (as tissue culture infectious doses) and will also supply the appropriate vehicle 
for aliquot preparation. Aliquot preparation will occur in the ICS.  All handling of the study agent will 
occur in a biosafety cabinet or a similarly contained environment.  
 
Any materials in contact with the study agent, e.g. syringes, vials, etc., will be disposed of as 
biological waste.  The final desired aliquot of the study agent infusate will be prepared at the intended 
volume sufficient for priming the infusion tubing and for infusion. .   The capped infusion syringe 
containing infusate (and primed infusion tubing if primed in pharmacy) will be transported to the study 
site in a ‘ziplock’ bag placed in a portable cooler or similarly contained transport device with a frozen 
ice block/ice pack, in order to maintain a temperature of approximately 4C.  No need to monitor the 
temperature during transport to the bedside. 
10.5 Compliance and Accountability 
Drug accountability records will be maintained for all clinical trial supplies.  All empty and partially 
used clinical trial supplies will be destroyed in accordance with institutional guidelines.  ICS will 
maintain detailed documentation of the receipt and/or destruction of the study agent.  All materials 
coming in contact with the study agent, the syringe delivered from ICS, tubing, dressings or coverings 
used to protect the trepanation site, will be disposed of as biological waste in the treatment room. 
10.6 Disposal and Destruction 
All surgical materials used in the procedure and (potentially) coming in contact with the study agent 
will be disposable.  These materials will be disposed of as biological waste using established 
procedures.  Used sharps will be disposed in biohazard sharps container and incinerated for final 
disposal. 
11 SUBJECT ELIGIBILITY 
11.1 Inclusion Criteria 
1. Patients must have a recurrent supratentorial WHO Grade III malignant glioma 
(anaplastic astrocytoma, anaplastic oligoastrocytoma, anaplastic oligodendroglioma, 
anaplastic pleomorphic xanthoastrocytoma, ependymoma) or WHO Grade IV malignant 
glioma, medulloblastoma, or atypical teratoid/rhabdoid tumor (ATRT) based on imaging 
studies with measurable disease (≥ 1 cm and ≤ 5.5 cm).   

 
 
a. The prior histopathology must be consistent with a World Health Organization 
(WHO) Grade III or IV malignant tumor confirmed by the study pathologist, Roger 
McLendon, or his designee. 
b. The tip of the infusion catheter can be placed ≥ 1cm from any ventricle.   
2. There is no standard of care treatment for children with Grade III/IV gliomas; however, 
patients must have received some form of definitive treatment, i.e., standard therapy with 
known clinical benefit, for their initial diagnosis prior to their recurrence/progression. 
Definitive treatment includes maximal safe resection (if possible) and radiation therapy 
with or without chemotherapy.  (Please note that patients who are unable to receive 
radiation therapy due to genetic disorders that put them at significant risk for radiation-
induced secondary malignancies (i.e., Gorlin’s syndrome or NF1 mutation) are still 
eligible to participate). 
3. All patients must be ≥ 4 years of age and ≤ 21 years of age at the time of entry into the 
study. 
4. The patient must have a Lansky or Karnofsky Performance Score (KPS) of ≥ 70% at the 
time of entry. 
5. Laboratory Studies: 
a. Platelet count ≥ 125,000/l prior to biopsy.  Platelets ≥ 100,000/l prior to infusion; 
b. Prothrombin and Activated Partial Thromboplastin Times ≤ 1.2 x upper limit of 
normal (ULN) prior to biopsy; 
c. Positive serum anti-poliovirus titer ≥ 1:8 prior to biopsy; 
d. Creatinine ≤ 1.2 x ULN prior to biopsy; 
e. Total bilirubin, AST, ALT, alkaline phosphatase ≤ 2.5 x ULN prior to biopsy; 
f. 
Neutrophil count ≥ 1000 /l prior to biopsy; 
g. Hemoglobin ≥ 9 gm/dl prior to biopsy (can be transfused). 
6. The patient must have received a boost immunization with trivalent inactivated IPOL™ 
(Sanofi-Pasteur) ≥ 1 week prior to administration of the study agent.  
7. At the time of biopsy, prior to administration of virus, the presence of recurrent tumor 
must be confirmed by histopathological analysis. 
8. A signed informed consent form approved by the Duke University Institutional Review 
Board (IRB) will be required for patient enrollment into the study. Patients (if 18 years old) 
or their parent(s) or guardian(s) (if younger than 18 years old) must be able to read and 
understand the informed consent document and must sign the informed consent 
indicating that they are aware of the investigational nature of this study. Children who are 
≥ 12 years old and < 18 years old should provide written assent to participate in the 
study.  Children who are ≤ 6 years old and < 12 years should provide verbal assent to 
participate in the study. 
11.2 Exclusion Criteria 
1. Pregnant or breast-feeding.  Female patients of child-bearing potential or female sexual 
partners (who are of child-bearing potential) of male patients must use at least one of the 
following methods of medically acceptable contraceptives:  approved hormonal 
contraceptives (such as birth control pills, patches, implants or infusions), an intrauterine 
device (IUD), or a barrier method of contraception (such as a condom or diaphragm) 
used with spermicide. Because all patients are required to have a boost immunization of 
trivalent inactivated IPOL™, there should be no risk of transmission of a mother to her 
fetus after receiving intracranial PVSRIPO. As such, patients who become pregnant after 
receiving PVSRIPO will continue to be monitored in the same manner, i.e. per protocol, 
unless the assessment is contra-indicated during pregnancy.  Partners who become 
pregnant will sign a Pregnant Partner Information Form and information regarding the 
pregnancy and its outcome may be collected. 
2. Patients with an impending, life-threatening cerebral herniation syndrome, based on the 
assessment of the study neurosurgeon.  
3. Patients with an active infection requiring intravenous treatment or having an unexplained 
febrile illness (Tmax > 99.5°F). 

 
 
4. Patients with known immunosuppressive disease or known human immunodeficiency 
virus infection. 
5. Patients with unstable or severe intercurrent medical conditions such as severe heart 
(New York Heart Association Class 3 or 4) or lung (FEV1 < 50%) disease, uncontrolled 
diabetes mellitus. 
6. Patients with allergy to human serum albumin. 
7. Patients with a known severe allergy (anaphylaxis) to gadolinium.  Patients with mild 
allergies (e.g., rash only) are eligible and may be pretreated per institutional guidelines 
prior to injection of the contrast agent. 
8. Patients with a previous history of neurological complications due to PV infection.. 
9. Patients who have not recovered from the toxic effects of prior chemo- and/or radiation 
therapy.  Guidelines for this recovery period are dependent upon the specific therapeutic 
agent being used:   
a. Patients who are less than 12 weeks from radiation therapy, unless progressive 
disease outside of the radiation field or 2 consecutive scans with disease 
progression or histopathologic confirmation of recurrent tumor. 
b. Patients who have received chemotherapy or bevacizumab ≤ 4 weeks [except for 
nitrosourea (6 weeks) or metronomic dosed chemotherapy such as daily 
etoposide or cyclophosphamide (1 week)] prior to starting the study drug unless 
patients have recovered from side effects of such therapy. 
c. Patients who have received immunotherapy ≤ 4 weeks prior to starting the study 
drug unless patients have recovered from side effects of such therapy.  
10. Patients with neoplastic lesions in the brainstem, cerebellum, spinal cord, intraventricular 
tumors, pituitary tumors, leptomeningeal disease, or other locations at the discretion of 
the treating neurosurgeon.  
11. Patients with a diagnosis of agammaglobulinemia, that is: 
a. Undetectable anti-tetanus toxoid IgG 
b. Known history of agammaglobulinemia 
12. Patients who are on dexamethasone receiving > 4 mg/day in the two weeks prior to 
admission for intra-cerebral delivery of PVSRIPO or who demonstrate worsening steroid 
myopathy. 
13. Patients with prior, unrelated malignancy requiring current active treatment with the 
exception of cervical carcinoma in situ and adequately treated basal cell or squamous 
cell carcinoma of the skin. 
 

 
 
12 SCREENING AND ON-STUDY TESTS AND PROCEDURES 
Table 2.  Schedule of Study Tests and Procedures 
Description 
Within 6 
months 
Screening: 
Within14 
days prior 
to catheter 
placement 
Screening: 
Within 5 
days prior 
to 
PVSRIPO 
Catheter 
Placement 
biopsy, 
PVSRIPO 
infusion 
After 
Completion 
of Infusion 
Post infusion (follow-up period) a 
Week 
 
 
 
 
 
0 
1 
2 
4 
8 
16, 24, 
32, 40, 
48 
Day 
 
Within 14 
days prior 
to catheter 
placement 
Within 5 
days prior 
to 
PVSRIPO 
0 
0 
1 
7 
14 
28 
56 
112, 168, 
224, 280, 
336 
General 
Evaluations 
 
Physical Exam 
 
X 
X 
 
 
Xb 
X 
X 
X 
X 
X 
Neurologic Exam 
 
X 
X 
 
 
Xb 
X 
X 
X 
X 
X 
Performance Status 
 
X 
X 
 
 
 
X 
X 
X 
X 
X 
Adverse Eventsh 
X 
X 
X 
Continuous 
Laboratory 
Evaluations 
 
Poliovirus 
Immunization 
Booster 
X 
 
 
 
 
 
 
 
 
 
 
CBC w/diff 
 
X 
 
 
 
X 
 
X 
X 
X 
X 
CMP 
 
X 
 
 
 
X 
 
X 
X 
X 
X 
PT, aPTT 
 
X 
 
 
 
 
 
 
 
 
 
Serum Pregnancy 
Test 
 
X 
Xc 
 
 
 
 
 
 
 
 
Serum for LSQ  
and anti-tetanus 
toxoid IgG 
X 
 
 
 
 
 
 
 
 
 
 
Whole blood for 
immunologic 
analysis and/or 
poliovirus titer (up 
to 10 mL, except for 
4 mL on Day 1 and 
6 mL on Day 14) 
X 
X 
 
 
 
X 
X 
X 
X 
X 
X (at 16 
and 24 
wks)d 
Whole blood for 
immunologic 
analysis (not to 
exceed 3 mL/kg in 
a 24 hr period or 7 
mL/kg in a 8 week 
period) 
Xe 
Xe 
 
 
 
 
Xe 
Xe 
Xe 
Xf 
X (at 16 
and 24 
wks)d, f 
Disease 
Evaluations 
 
MRI 
 
 
X 
 
Xg (based 
upon date of 
enrollment)  
 
 
 
X 
X 
X 
CT Scan 
 
 
 
X 
 
 
 
 
 
 
 
Biopsy 
 
 
 
X 
 
 
 
 
 
 
 
Treatment 
 
 
 
 
 
 
 
 
 
 
 
PVSRIPO 
 
 
 
X 
 
 
 
 
 
 
 
a Starting with Day 7 (Week 1), all tests and procedures have a 7-day plus or minus window. 
b Daily after infusion until discharged from hospital 
c Serum pregnancy test, if appropriate, within 48 hours of catheter placement and PVSRIPO 
d At the discretion of the Principal Investigator, an additional blood sample at least 2 years post study drug infusion 
may be obtained.   
e  Whole blood draws for immunologic analysis may be taken (up to 36 mL) at the discretion of the Investigator, but 
should remain consistent with allowable volumes for pediatric patients depending upon the weight of the child (per 
Duke policy, the maximum blood volume drawn for research should not exceed 3 mL/kg in a 24 hr period or 7 mL/kg 
in an 8 week period).   

 
 
f At 8, 16, and 24 weeks, up to 76.5 mL of whole blood may be drawn for immunologic analysis (please refer to Duke 
blood volume policy mentioned above). 
g Prior to October 31, 2019, MRI to track Magnevist® containing infusate should have been obtained within 4 hours 
after completion of infusion.  This MRI after the infusion was without gadolinium enhancement, but this MRI will not 
be performed on patients enrolled after October 31, 2019.  All other MRIs in the study include gadolinium 
enhancement. 
h Adverse events directly related to required screening procedures should be recorded. 
12.1 Screening Examination 
The screening examination will take place within 2 weeks before catheter placement.  An informed 
consent must be signed by the patient and their legal guardian(s) before any study-specific screening 
procedure takes place.  Within 6 months before receiving the PVSRIPO infusion, the patient should 
have received a PV booster, a serum test for LSQ and anti-tetanus toxoid IgG.  The patient should 
also have whole blood drawn (up to 10 mL; red top tube(s)) for immunologic analysis and PV titer and 
an additional amount (up to 36 mL; yellow top ACD tube(s)) for other immunologic analysis during this 
time.  Blood volumes drawn for immunologic analyses should not exceed 3 mL/kg in a 24 hour period 
or 7 mL/kg in an 8 week period, allowable per Duke policy.  At the discretion of the investigator, blood 
volumes for immunologic analysis should be adjusted depending upon the weight of the child, in order 
to be consistent with Duke policy.   
 
Pre-treatment evaluations within 2 weeks before catheter placement to determine eligibility and as a 
baseline will include the following, unless otherwise indicated: 
• 
History and physical exam, including a full neurologic assessment and KPS or Lansky, within 
2 weeks of catheter placement and within 5 days of PVSRIPO infusion  
• 
Laboratory Evaluations: 
o 
CBC with differential 
o 
CMP 
o 
PT, aPTT 
o 
Beta-HCG, if appropriate (within 48 hours of catheter placement and PVSRIPO) 
o 
Anti-tetanus toxoid IgG (within 6 months of catheter placement and PVSRIPO) 
o 
LSQ (within 6 months of catheter placement and PVSRIPO) 
o 
Whole blood (up to 10 mL) for immunologic analysis and poliovirus titer 
o 
Whole blood (up to 36 mL, but not to exceed 3 mL/kg in a 24 hour period or 7 mL/kg 
in an 8-week period) for immunologic analysis (volumes drawn will be adjusted at the 
discretion of the Investigator, but will be based on the weight of the child to be 
consistent with Duke policy) 
• 
Baseline MRI of the brain within 5 days of starting treatment 
 
If a subject does not receive PVSRIPO, minimal records regarding the subject and the reason for 
screen failure will be retained in the study database. 
12.2  Run-In Period 
Not applicable. 
12.3  Treatment Period 
Day 0 
• 
Biopsy and catheter placement 
• 
After obtaining tissue for the standard clinical pathologic testing, up to three additional core 
biopsies will be obtained for molecular genetic tests (Section 9.1.4) 
• 
CT of the brain to confirm catheter placement ≥ 1 cm from the ventricles prior to beginning 
infusion 
• 
PVSRIPO infusion (infusion may be same day or within a +1 day window) 
12.4 Follow-up Period 
Day 1 post infusion 

 
 
• 
History and physical exam, including a full neurologic assessment, to be performed daily until 
discharged from the hospital 
• 
CBC with differential  
• 
CMP 
• 
Whole blood (up to 4 mL) for immunologic analysis 
 
Week 1 (+/- 1 week) 
• 
History and physical exam, including a full neurologic assessment and KPS  
• 
Whole blood (up to 10 mL) for immunologic analysis and PV titer 
• 
Whole blood (up to 36 mL, but not to exceed 3 mL/kg in a 24 hour period or 7 mL/kg in an 8-
week period) for immunologic analysis at the discretion of the Investigator 
 
Week 2 (+/- 1 week) 
• 
History and physical exam, including a full neurologic assessment and KPS 
• 
CBC with differential  
• 
CMP 
• 
Whole blood (up to 6 mL) for immunologic analysis and PV titer 
• 
Whole blood (up to 36 mL, but not to exceed 3 mL/kg in a 24 hour period or 7 mL/kg in an 8 
week period) for immunologic analysis at the discretion of the Investigator 
 
Week 4 (+/- 1 week) 
• 
History and physical exam, including a full neurologic assessment and KPS 
• 
MRI of the brain with gadolinium contrast  
• 
CBC with differential  
• 
CMP 
• 
Whole blood (up to 10 mL) for immunologic analysis and PV titer 
• 
Optional whole blood (up to 36 mL, but not to exceed 3 mL/kg in a 24 hour period or 7 mL/kg 
in an 8 week period) for immunologic analysis at the discretion of the Investigator 
 
Week 8 (+/- 1 week) 
• 
History and physical exam, including a full neurologic assessment and KPS 
• 
MRI of the brain with gadolinium contrast 
• 
CBC with differential 
• 
CMP 
• 
Whole blood (up to 10 mL) for immunologic analysis and PV titer 
• 
Whole blood (up to 76.5 mL, but not to exceed 3 mL/kg in a 24 hour period or 7 mL/kg in an 8 
week period) for immunologic analysis at the discretion of the Investigator 
 
Weeks 16, 24, 32, 40, 48 (+/- 1 week) 
• 
History and physical exam, including a full neurologic assessment and KPS 
• 
MRI of the brain with gadolinium contrast 
• 
CBC with differential 
• 
CMP 
• 
Whole blood (up to 10 mL) for immunologic analysis and PV titer (weeks 16 and 24 only) 
• 
Whole blood (up to 76.5 mL, but not to exceed 3 mL/kg in a 24 hour period or 7 mL/kg in an 8 
week period) for immunologic analysis (weeks 16 and 24 only) at the discretion of the 
Investigator 
 
After week 48, patient will follow-up at the discretion of the treating physician. 
• 
Whole blood (up to 10 mL) for immunologic analysis and PV titer (At the discretion of the 
Investigator, this blood sample, at least 2 years post-study drug infusion, may be obtained.) 
• 
Whole blood (up to 76.5 mL, but not to exceed 3 mL/kg in a 24 hour period or 7 mL/kg in an 8 
week period) for immunologic analysis (At the discretion of the Investigator, this blood 
sample, at least 2 years post-study drug infusion, may be obtained.) 

 
 
 
Patients may not be treated with any other modality (other than bevacizumab per the ‘special 
considerations’ in Section 9.1.6) unless progressive tumor is noted or they are otherwise removed 
from the study.  Patients will be considered off study upon tumor progression or upon treatment of the 
tumor with another modality.  When subjects are considered off study, this indicates that subjects will 
no longer be obligated to undergo study-related tests and procedures, but the data described below 
will still be collected from these subjects as feasible.  Subjects will be followed for serious adverse 
events for 30 days after coming off study.  Collection of the following additional data from off study 
subjects will be performed, if possible, but is not mandatory and will not be considered a deviation if 
the data cannot be obtained. Subjects’ medical records will be reviewed for the remainder of their life, 
in order to collect data on subsequent treatments, disease progression, tumor size/volume, and 
survival.  Follow-up activity will be at the discretion of the treating physician.   
12.5  End of Study 
The study will be considered complete once enrollment has been met, follow-up procedures outlined 
in Section 12.4 have been conducted on all subjects, and data analysis is concluded.  The study may 
also be terminated early for any reason by the PI-sponsor.   
 
Subjects that are lost to follow-up will be documented in the patient record and in the 21 CFR Part 11 
database.  In the compliant database, the subject will be marked as “Patient Status Unknown,” along 
with a corresponding explanation, if any.   
12.6  Early Withdrawal of Subject(s) 
12.6.1  Criteria for Early Withdrawal 
Subjects may voluntarily withdraw from the study at any time.  The Principal Investigator may 
also withdraw a subject from the study at any time based on his/her discretion.  Reasons for PI-
initiated withdrawal may include, but are not limited to the following: 
 
- 
Non-compliance of the subject 
- 
Administrative issues 
- 
Disease progression 
12.6.2  Follow-up Requirements for Early Withdrawal 
Subjects should be seen in clinic or contacted at a minimum of every 3 months for 1 year.  
12.6.3  Replacement of Early Withdrawal(s) 
Subjects that withdraw from the study prior to receiving PVSRIPO infusion, either voluntarily or 
due to ineligibility, will be considered non-evaluable; those subjects will be replaced. 
12.7  Study Assessments 
12.7.1  Medical History 
Standard medical history will be obtained and documented per institutional guidelines. 
12.7.2  Physical Exam 
Standard physical exam and neurological assessment will be conducted and documented per 
institutional and PRTBTC guidelines. 
12.7.3  Radiographic Review 
A CT will be obtained to confirm catheter placement prior to beginning the PVSRIPO infusion.  
A MRI of the brain without gadolinium enhancement will be obtained within 4 hours of 

 
 
completing the PVSRIPO infusion to monitor distribution of the infusion.  A MRI of the brain with 
gadolinium enhancement will be obtained at all subsequent visits.  
 
Given that imaging following immunotherapy differs greatly from what is typically seen following 
chemoradiation treatment or treatment with anti-angiogenic compounds, evaluation of response 
using with Macdonald criteria or RANO criteria is not appropriate in this trial.  Immunotherapy 
can trigger an inflammatory immune response that is observed on imaging.  Distinguishing 
between the inflammatory immune response and progressive disease is difficult.  Therefore, an 
exploratory objective of this study is to describe radiographic imaging post-PVSRIPO treatment.  
Another exploratory objective of this study is to describe changes in tumor volume based upon 
radiographic imaging.  For that reason, subjects who have gone off study may continue to be 
followed for collection of MRI measurements.  For the purpose of this objective, relative tumor 
volume is estimated from the MRI using the metric of multiplying the widest tumor diameter with 
the perpendicular height of the tumor in that same slice. 
12.7.4  Laboratory Evaluations 
The timing of laboratory assessments that will be obtained during the course of the study is 
given above in Table 2.   
12.7.5 Correlative Assessments 
• 
Anti-tetanus toxoid IgG titer within 6 months prior to treatment 
• 
Up to 10 mL of whole blood for tests of immunologic assays and anti-PV antibody titer at 
day 1 and weeks 1, 4, 8, 16, and 24 after treatment.  Up to 6 mL collection of whole blood 
at week 2 (day 14). 
• 
Up to 36 mL of whole blood for tests of anti-tumor immune responses within 6 months 
before receiving PVSRIPO, and at weeks 1, 2, and 4 and up to 76.5 mL at weeks 8, 16, 
and 24 after treatment.  At the discretion of the Principal Investigator, an additional blood 
sample at least 2 years post study drug infusion may be obtained (see Table 2). Because 
of the large volume of the whole blood draws taken for immune monitoring, it will be 
essential to carefully consider the weight of each patient when determining whether a blood 
draw may occur, or how much blood may be drawn at a given time point.  In this pediatric 
population the maximum blood volumes are 3 mL/kg in a 24 hour period or 7 mL/kg in an 8 
week period.  Table 3 contains predicted blood draw volumes that are required for planned 
study assessments.  
   

 
 
Table 3.  Schedule of Blood Draw Volumes 
Maximum blood draw volumes (mL) a 
Screening 
Treatment 
Day 
Within 6 
months 
(≈d -28) 
Within 14 
days prior 
to catheter 
(≈d -14) 
1 
7 
14 
28 
56 
CBC & CMP 
 
6 
6 
 
6 
6 
6 
PT, aPTT 
 
1.8 
 
 
 
 
 
Pregnancy 
 
2 
 
 
 
 
 
LSQ, tetanus 
titer 
6 
 
 
 
 
 
 
Polio titer & 
immunologic 
analysis 
Up to 10 
(red top 
tube) 
Up to 10 
(red top 
tube) 
 
Up to 10 
(red top 
tube) 
Up to 6 
(red top 
tube) 
Up to 10 
(red top 
tube) 
Up to 10 
(red top 
tube) 
Whole blood 
immunologic 
analysis 
Up to 36b 
(yellow 
top ACD 
tube) 
Up to 36b 
(yellow top 
ACD tube) 
 
Up to 36b 
(yellow top 
ACD tube) 
Up to 36b 
(yellow top 
ACD tube) 
Up to 36b 
(yellow 
top ACD 
tube) 
Up to 76.5b 
(yellow top 
ACD tube) 
Daily Total 
52 
55.8 
6 
46 
48 
52 
92.5 
Total 8 week volume (day -28 through +28):  259.8 mL 
a  Volumes for standard lab tests are based on DUHS Clinical Labs volumes for patients age 8-15. For patients in the 
lower age range of this study and/or with a low body weight, lower pediatric volumes (for patients age 0-8) may be 
used. Some volumes may be higher for patients >15 years old. 
b  Volumes of whole blood for immunologic analysis will be reduced as necessary to not exceed 3 mL/kg in a 24 hr 
period or 7 mL/kg in an 8 week period.   
13 SAFETY MONITORING AND REPORTING 
The Principal Investigator is responsible for the identification and documentation of adverse events and 
serious adverse events, as defined below.  At each study visit, the Principal Investigator or designee must 
assess, through non-suggestive inquiries of the subject or evaluation of study assessments, whether an 
adverse event (AE) or serious adverse event (SAE) has occurred. 
13.1  Adverse Events 
An AE is any untoward medical occurrence in a subject receiving study drug and which does not 
necessarily have a causal relationship with this treatment.  For this protocol, the definition of AE also 
includes worsening of any pre-existing medical condition.  An AE can therefore be any unfavorable 
and unintended or worsening sign (including an abnormal laboratory finding), symptom, or disease 
temporally associated with the use of PVSRIPO, whether or not related to use of the PVSRIPO.  
Abnormal laboratory findings with or without clinical significance are collected according to CTCAE 
v.4. 
 
From the time the subject has their first study-related procedure until their last study-related 
procedure, all AEs must be recorded in the subject medical record and adverse events case report 
form.  Thus, AE directly related to study-specific and required screening procedures should be 
reported.  
 
AEs will be assessed according to the CTCAE version 4.  If CTCAE grading does not exist for an AE, 
the severity of the AE will be graded as mild (1), moderate (2), severe (3), life-threatening (4), or fatal 
(5). 
 

 
 
Attribution of AEs will be indicated as follows: 
- 
Definite:  The AE is clearly related to the study drug 
- 
Probably:  The AE is likely related to the study drug 
- 
Possible:  The AE may be related to the study drug 
- 
Unlikely:  The AE is doubtfully related to the study drug 
- 
Unrelated:  The AE is clearly NOT related to the study drug 
13.1.1  AEs of Special Interest 
Not applicable. 
13.1.2  Reporting of AEs 
A database of all adverse events (not just those considered related to the study drug) will be 
maintained in 21 CFR Part 11 Compliant database. The event will be categorized by organ 
system, relationship to treatment, its grade of severity, and resolution. The Principal 
Investigator and study statistician will periodically review the collective adverse events with the 
intention of identifying any trends or patterns in toxicity. If any such trends are identified, 
depending on their severity and frequency, a protocol amendment will be considered. 
13.2  Serious Adverse Events 
An AE is considered “serious” if in the opinion of the investigator it is one of the following outcomes: 
 
- 
Fatal 
- 
Life-threatening 
- 
Constitutes a congenital anomaly or birth defect 
- 
A medically significant condition (defined as an event that compromises subject safety or 
may require medical or surgical intervention to prevent one of the three outcomes above). 
- 
Requires inpatient hospitalization or prolongation of existing hospitalization 
- 
Results in persistent or significant incapacity or substantial disruption to conduct normal life 
functions. 
13.2.1  Reporting of SAEs 
All serious and unexpected AEs should be reported immediately to the Principal Investigator or 
designee.  Only AEs that the Principal Investigator determines to be serious, unanticipated, and 
related or possibly/probably (i.e. more likely than not) related to the research must be reported 
to the Duke IRB.  Those AEs will be submitted in the electronic IRB system, according to the 
following guidelines: 
• 
Report within 24 hours of learning about any subject’s death that was unanticipated and 
more likely related to the research than unrelated; 
• 
Report within 5 business days of learning about any serious, unanticipated, and related 
or possibly/probably related AEs; 
• 
Report within 10 business day of learning about any other unanticipated problem or event 
that was more likely related to the research than unrelated. 
 
SAEs must be reported to the Sponsor or their designee within 24 h of learning of the event 
whether or not the SAE is considered to be related to the study drug.  Applicable forms 
provided by the Sponsor or their designee must be transmitted via secure email to 
pv_istari@klserv.com.  The Sponsor must report to the FDA, in an IND safety report, any 
suspected adverse reaction that is both serious and unexpected. Before submitting this report, 
the sponsor needs to ensure that the event meets all three of the definitions contained in the 
requirement:  
 
• 
Suspected adverse reaction (i.e. there is a reasonable possibility that the drug caused the 
AE) 

 
 
• 
Serious  
• 
Unexpected  
 
If the AE does not meet all three of the definitions, it should not be submitted as an expedited 
IND safety report. 
 
The Sponsor is required to report to the FDA all IND Safety reports in writing within 15 days (7 
days for unexpected fatal or life-threatening suspected adverse reaction).  The FDA Form 
3500A can be found on the FDA website, www.fda.gov.  All other AEs will be reported to the 
FDA in the Annual Report. 
13.3  Emergency Unblinding of Investigational Treatment 
Not applicable. 
13.4  Other Reportable Information 
Not Applicable. 
13.5  Special Warnings and Precautions 
Not Applicable. 
13.6   Safety Oversight Committee (SOC) 
The Duke Cancer Institute SOC is responsible for annual data and safety monitoring of DUHS 
sponsor-investigator Phase I and II, therapeutic interventional studies that do not have an 
independent Data Safety Monitoring Board (DSMB).  The primary focus of the SOC is review of 
safety data, toxicities and new information that may affect subject safety or efficacy. Annual safety 
reviews include but may not be limited to review of safety data, enrollment status, stopping rules if 
applicable, accrual, toxicities, reference literature, and interim analyses as provided by the Principal 
Investigator.  The SOC in concert with the DCI Monitoring Team (see Section 14.1 for Monitoring 
Team description) oversees the conduct of DUHS cancer-related, sponsor-investigator greater-than-
minimal-risk intervention studies that do not have an external monitoring plan, ensuring subject safety 
and that the protocol is conducted, recorded and reported in accordance with the protocol, standing 
operating procedures (SOPs), Good Clinical Practice (GCP), and applicable regulatory requirements. 
13.7  External Data and Safety Monitoring Board (DSMB) 
Due to a potential conflict of interest, an external DSMB-Plus was created for the Phase I clinical trial 
performed at Duke University Medical Center.  The current study will continue to be monitored by the 
same DSMB-Plus.  The external DSMB-Plus will be responsible for safeguarding the interests of trial 
subjects and assessing the safety of the interventions during the trial. The DSMB-Plus will provide 
recommendations about stopping or continuing enrollment in the trial.  To contribute to enhancing the 
integrity of the trial, the DSMB-Plus may also formulate recommendations relating to the selection, 
recruitment, and retention of subjects and their management.  Additional details regarding the 
responsibility of the DSMB-Plus and its chair may be found in the charter document. 
14 QUALITY CONTROL AND QUALITY ASSURANCE 
14.1  Monitoring 
The Duke Cancer Institute (DCI) Monitoring Team will conduct monitoring visits to ensure subject 
safety and to ensure that the protocol is conducted, recorded, and reported in accordance with the 
protocol, standard operating procedures, good clinical practice, and applicable regulatory 
requirements.  The DCI Monitoring Team has agreed to conduct monitoring of the first three subjects 
enrolled following completion of their 4-week follow up visit.  This monitoring of eligibility is in 
conjunction with annual monitoring of accrual, regulatory, consenting, eligibility, conduct, safety, data 
quality, investigation product accountability, and biologic samples. This sequential patient monitoring 
for the first three patients will be followed by annual monitoring of 1-3 subjects until the study is 

 
 
closed to enrollment and subjects are no longer receiving study interventions that are more than 
minimal risk.  Results of the DCI monitoring may be shared with the Sponsor, Istari Oncology, Inc. In 
addition, the Sponsor may perform risk-based monitoring oversight visits for the study.   
 
Additional monitoring may be prompted by findings from monitoring visits, unexpected frequency of 
serious and/or unexpected toxicities, or other concerns and may be initiated upon request of DUHS 
and DCI leadership, the DCI Cancer Protocol Committee, the Safety Oversight Committee (SOC), the 
Sponsor, the Principal Investigator, or the IRB.  All study documents must be made available upon 
request to the DCI Monitoring Team and other authorized regulatory authorities, including but not 
limited to the National Institute of Health, National Cancer Institute, and the FDA.  Every reasonable 
effort will be made to maintain confidentiality during study monitoring. 
14.2  Audits 
The Duke Office of Audit, Risk, & Compliance (OARC) office may conduct confidential audits to 
evaluate compliance with the protocol and the principles of GCP.  The Principal Investigator agrees to 
allow the OARC auditor(s) direct access to all relevant documents and to allocate his/her time and the 
time of the study team to the OARC auditor(s) in order to discuss findings and any relevant issues. 
 
OARC audits are designed to protect the rights and well-being of human research subjects. CTQA 
audits may be routine or directed (for cause). Routine audits are selected based upon risk metrics 
generally geared towards high subject enrollment, studies with limited oversight or monitoring, 
Investigator initiated Investigational Drugs or Devices, federally-funded studies, high degree of risk 
(based upon adverse events, type of study, or vulnerable populations), Phase I studies, or studies 
that involve Medicare populations. Directed audits occur at the directive of the IRB or an authorized 
Institutional Official. 
 
OARC audits examine research studies/clinical trials methodology, processes and systems to assess 
whether the research is conducted according to the protocol approved by the DUHS IRB. The primary 
purpose of the audit/review is to verify that the standards for safety of human subjects in clinical trials 
and the quality of data produced by the clinical trial research are met. The audit/review will serve as a 
quality assurance measure, internal to the institution. Additional goals of such audits are to detect 
both random and systemic errors occurring during the conduct of clinical research and to emphasize 
“best practices” in the research/clinical trials environment. 
14.3  Data Management and Processing 
14.3.1  Case Report Forms (CRFs) 
The electronic CRF (eCRF) will be the primary data collection document for the study and is 
developed in conjunction with statistical oversight.  The CRFs will be updated in a timely 
manner following acquisition of new source data.  Only the Principal Investigator, the study 
coordinator, the data management team, and the clinical trials manager are permitted to make 
entries, changes, or corrections in the eCRF. 
 
An audit trail will be maintained automatically by the electronic CRF management system.  All 
users of this system will complete user training, as required or appropriate per DCI 
requirements and other regulations. 
14.3.2  Data Management Procedures and Data Verification 
Access to electronic databases will be managed by the PRTBTC Data Manager. 
 
Completeness of entered data will be checked automatically by the eCRF system, and users will 
be alerted to the presence of data inconsistencies.  Additionally, the data management team and 
the statistical team will cross-reference the data to verify accuracy.  Missing or implausible data 
will be brought to the attention of the Principal Investigator requiring appropriate responses (i.e. 

 
 
confirmation of data, correction of data, completion or confirmation that data is not available, 
etc.). 
 
The database will be reviewed and discussed prior to database closure, and will be closed only 
after resolution of all remaining queries.  An audit trail will be kept of all subsequent changes to 
the data. 
14.3.3  Study Closure 
Following completion of the study, the Principal Investigator will be responsible for ensuring the 
following activities: 
 
- 
Data clarification and/or resolution 
- 
Accounting, reconciliation, and destruction/return of used and unused study drugs 
- 
Review of site study records for completeness 
- 
Shipment of all remaining laboratory samples to the designated laboratories 
15 STATISTICAL METHODS AND DATA ANALYSIS 
All statistical analysis will be performed under the direction of the statistician designated in key personnel.  
Any data analysis carried out independently by any investigator must be approved by the statistician and 
Sponsor before publication or presentation, in keeping with the executed clinical trial agreement(s). 
15.1  Analysis Sets 
All patients who receive PVSRIPO treatment will be included in all safety and efficacy analyses. 
15.2  Patient Demographics and Other Baseline Characteristics 
Socio-demographic and clinical characteristics of patients enrolled and treated on this study will be 
summarized.  For categorical variables, frequencies and percentages will be provided.  Means with 
standard deviations and medians/percentiles will summarize non-categorical variables. 
15.3  Treatments 
The number of patients that receive catheter placement will be calculated, as well as the number of 
patients who subsequently receive PVSRIPO infusion. 
15.4  Primary Objective 
The primary objective of this study is to confirm the safety of the selected dose of PVSRIPO 
treatment among pediatric patients with recurrent WHO Grade III malignant glioma (anaplastic 
astrocytoma, anaplastic oligoastrocytoma, anaplastic oligodendroglioma, anaplastic pleomorphic 
xanthoastrocytoma, ependymoma) or WHO Grade IV malignant glioma (glioblastoma, gliosarcoma).   
15.4.1  Overview of Study Design and Monitoring 
Patients will be treated with PVSRIPO, then carefully monitored for safety for at least 1 year after 
treatment.  Of particular interest will be the incidence of AEs that occur during the first 14 days 
after PVSRIPO treatment and the inflammatory events that occur during the first year after 
PVSRIPO treatment.  The experience with PVSRIPO treatment of adult patients with recurrent 
WHO grade IV malignant glioma suggests that a patient remains at risk, during the first 12 
months post-PVSRIPO treatment, of demonstrating neurologic or radiographic signs suggestive 
of an inflammatory reaction secondary to the immune response triggered by PVSRIPO that 
requires a dose of dexamethasone above 4 mg a day.  As described in Section 9.1.6, such 
patients will be treated with bevacizumab 7.5 mg/kg IV every 3 weeks if possible, or alternatively 
corticosteroids or surgery.  
 

 
 
Two sets of safety monitoring guidelines are proposed, with one focused on acute toxicity (i.e. 
occurring during the first 14 days after PVSRIPO administration), and one focused on long-term 
issues. 
 
Acute Toxicity Monitoring Guidelines:  Monitoring rules based upon the occurrence of an 
unacceptable toxicity, as defined in Section 9.1.5, during the first 14 days after PVSRIPO 
administration are shown below in Table 4.  
 
If the number of patients with an unacceptable toxicity surpasses the noted thresholds, accrual 
will be suspended.  Data will be carefully reviewed to determine if accrual should be terminated, 
continued without modification, or continued with appropriate modification (e.g. PVSRIPO dose 
reduction). 
 
Table 4.  Acute Toxicity Monitoring Rules. 
# of Patients Accrued 
Accrual Suspension Threshold 
0 – 6 
≥2 patients with unacceptable toxicity 
7-9 
≥3 patients with unacceptable toxicity 
10-12 
≥4 patients with unacceptable toxicity 
 
The probability of study suspension and the expected number of treated patients before study 
suspension is tabulated below in Table 5 as a function of the true underlying probability of an 
unacceptable toxicity. 
 
Table 5.  Probability of study suspension. 
True Underlying Probability of a 
Unacceptable Toxicity within 14 days of 
PVSRIPO administration 
Probability of Accrual 
Suspension 
Expected Number of 
Patients to be Accrued 
0.05 
0.030 
11.8 
0.10 
0.124 
11.1 
0.15 
0.262 
10.2 
0.20 
0.411 
9.2 
0.25 
0.560 
8.1 
0.30 
0.681 
7.2 
0.35 
0.792 
6.2 
0.40 
0.873 
5.4 
 
Long-term Monitoring:  If any patient experiences an inflammatory reaction that does not 
improve within 18 weeks of commencing bevacizumab or steroid treatment, the inflammatory 
reaction will be considered “irreversible.”  If a patient experiences “irreversible” inflammatory 
reaction, further enrollment of patients on the PVSRIPO trial will be temporarily interrupted and 
data on all previously treated patients will be carefully reviewed to determine if the study needs 
modification.  Options may include dose reductions in future patients (next PVSRIPO dose would 
be 3.3 x 106 TCID50), modification of the approach to treating a patient with an inflammatory 
reaction, study termination, or accrual continuation without modification. 
 
The FDA and/or the DSMB for this study may be part of this decision-making in the event that 
there are unacceptable toxicities or inflammatory issues. 
15.4.2 Primary Objective Analyses 
Percentage of patients that experience unacceptable toxicity during the 14-day period post-
PVSRIPO treatment will be calculated. 
 

 
 
The proportion of patients who require low-dose bevacizumab or steroid treatment due to an 
inflammatory reaction secondary to an immune response to PVSRIPO treatment will be 
calculated.  In addition, the time to initiation of such treatment will be calculated and summarized 
using Kaplan-Meier methodology. 
 
Adverse events experienced by protocol subjects from the time of catheter placement will be 
summarized in several forms to satisfy scientific and monitoring needs, as well as various 
regulatory reporting needs (e.g. FDA, DCI SOC, and ClinicalTrials.gov).  These include a 
summary of the frequency of adverse events that are possibly, probably, or definitely related to 
protocol treatment tabulated by the maximum grade for each type of adverse event, as well as a 
summary of the frequency of adverse events regardless of attribution tabulated by the maximum 
grade for each type of adverse event. 
15.5  Secondary Objective 
The secondary objective is the estimation of overall survival (OS), which will be calculated as the time 
between PVSRIPO infusion and death, or last follow-up if the patient remains alive. The Kaplan-Meier 
estimator will be used to describe the survival of patients treated with PVSRIPO.  Median OS will be 
estimated, as well as 6-, 12-, and 24-month OS.  Analyses will be stratified by histologic grade. 
15.6  Exploratory Objective 
An exploratory objective of this study is to describe changes visualized on imaging due to intratumoral 
inoculation of PVSRIPO.  Two components of the inflammatory reaction will be characterized:  the 
enhancing lesion and the edema (FLAIR) area. Given that imaging following immunotherapy differs 
greatly from what is typically seen following chemoradiation treatment or treatment with anti-
angiogenic compounds, evaluation of response using with Macdonald criteria or RANO criteria is not 
appropriate in this trial.  Immunotherapy can trigger an inflammatory immune response that is 
observed on imaging.  Distinguishing between the inflammatory immune response and progressive 
disease is difficult.  Therefore, an exploratory objective of this study is to describe radiographic 
imaging post-PVSRIPO treatment.  
15.7  Other Exploratory Objectives 
An additional exploratory objective is to describe changes in tumor volume based upon imaging.  
Subjects’ MRIs will be followed for changes in tumor volume post-PVSRIPO treatment.  Another 
exploratory objective is the assessment of immunologic changes stemming from the administration of 
PVSRIPO treatment. Peripheral blood will be collected at defined intervals for correlative immune 
monitoring studies in serum and in peripheral blood monocytes. These include (i) analyses of innate 
and inflammatory immune events (HMGB1, inflammatory cytokines, NK and NKT-cells and regulatory 
immune subsets (Tregs & MDSCs)); (ii) analyses of adaptive immune responses (lineage, maturation, 
induction and activation/functional status of tumor antigen-specific T-cells).  A final exploratory 
objective is to identify genetic predictors of response or failure of response to treatment with 
PVSRIPO. The Cox model will explore the impact of genetic markers of survival time.  With logistic 
regression, the association between genetic markers and being a long-term survivor will be explored.    
15.8  Interim Analysis 
See Section 15.4.1. 
15.9  Sample Size Calculation 
A maximum of 12 patients will be treated on this study. 

 
 
16 ADMINISTRATIVE AND ETHICAL CONSIDERATIONS 
16.1  Regulatory and Ethical Compliance 
This protocol was designed and will be conducted and reported in accordance with the International 
Conference on Harmonization (ICH) Harmonized Tripartite Guidelines for Good Clinical Practice, the 
Declaration of Helsinki, and applicable federal, state, and local regulations. 
16.2  DUHS Institutional Review Board and DCI Cancer Protocol 
Committee 
The protocol, informed consent form, advertising material, and additional protocol-related documents 
must be submitted to the DUHS Institutional Review Board (IRB) and DCI Cancer Protocol 
Committee (CPC) for review.  The study may be initiated only after the Principal Investigator has 
received written and dated approval from the CPC and IRB. 
 
The Principal Investigator must submit and obtain approval from the IRB for all subsequent protocol 
amendments and changes to the informed consent form.  The CPC should be informed about any 
protocol amendments that potentially affect research design or data analysis (i.e. amendments 
affecting subject population, inclusion/exclusion criteria, agent administration, statistical analysis, 
etc.). 
 
The Principal Investigator must obtain protocol re-approval from the IRB within 1 year of the most 
recent IRB approval.  The Principal Investigator must also obtain protocol re-approval from the CPC 
within 1 year of the most recent IRB approval, for as long as the protocol remains open to subject 
enrollment. 
16.3 Informed Consent 
The informed consent form must be written in a manner that is understandable to the subject 
population and their legal guardian(s).  Prior to its use, the informed consent form must be approved 
by the IRB. 
 
The Principal Investigator or authorized key personnel will discuss with the potential subject and their 
legal guardian(s) the purpose of the research, methods, potential risks and benefits, subject 
concerns, and other study-related matters.  This discussion will occur in a location that ensures 
subject privacy and in a manner that minimizes the possibility of coercion.  Appropriate 
accommodations will be made available for potential subjects and their legal guardian(s) who cannot 
read or understand English or are visually impaired.  Potential subjects and their legal guardian(s) will 
have the opportunity to contact the Principal investigator or authorized key personnel with questions, 
and will be given as much time as needed to make an informed decision about participation in the 
study. 
 
Before conducting any study-specific procedures, the Principal Investigator must obtain written 
informed consent from the subject and/or their legal guardian(s).  The original informed consent form 
will be stored with the subject’s study records, and a copy of the informed consent form will be 
provided to the subject and their legal guardian(s).  The Principal Investigator is responsible for 
asking the subject and their legal guardian(s) whether the subject wishes to notify his/her primary 
care physician about participation in the study.  If the subject agrees to such notification, the Principal 
Investigator will inform the subject’s primary care physician about the subject’s participation in the 
clinical study. 
16.4  Privacy, Confidentiality, and Data Storage 
The Principal Investigator will ensure that subject privacy and confidentiality of the subject’s data will 
be maintained.  Research Data Security Plans (RDSPs) will be approved by the appropriate 
institutional Site Based Research group. 
 

 
 
To protect privacy, every reasonable effort will be made to prevent undue access to subjects during 
the course of the study.  Prospective participants will be consented in an exam room where it is just 
the research staff, the patient and his family, if desired. For all future visits, interactions with research 
staff (study doctor and study coordinators) regarding research activities will take place in a private 
exam room. All research related interactions with the participant will be conducted by qualified 
research staff who are directly involved in the conduct of the research study. 
 
To protect confidentiality, subject files in paper format will be stored in secure cabinets under lock and 
key accessible only by the research staff. Subjects will be identified only by a unique study number 
and subject initials.  Electronic records of subject data will be maintained using a dedicated 21 CFR 
Part 11 compliant database, which is housed in an encrypted and password-protected file on a 
secure network drive.  Access to electronic databases will be managed by the PRTBTC Data 
Manager.  The security and viability of the IT infrastructure will be managed by the DCI and/or Duke 
Medicine.   
 
Upon completion of the study, research records will be archived and handled per DUHS HRPP policy.  
Subject names or identifiers will not be used in reports, presentations at scientific meetings, or 
publications in scientific journals. 
16.5  Data and Safety Monitoring 
Data and Safety Monitoring will be performed in accordance with the DCI Data and Safety Monitoring 
Plan.   
16.6  Protocol Amendments 
All protocol amendments must be initiated by the Sponsor and approved by the IRB prior to 
implementation.  IRB approval is not required for protocol changes that occur to protect the safety of 
a subject from an immediate hazard.  However, the Sponsor and/or Principal Investigator must inform 
the IRB and all other applicable regulatory agencies of such action immediately. 
16.7  Records Retention 
The Principal Investigator will maintain study-related records until the youngest child on study is 21 
years old, or for at least six years after study completion, whichever is longer (Duke policy). 
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