A Phase II Trial of ADI-PEG 20 in Combination With Gemcitabine and Docetaxel for the Treatment of Soft Tissue Sarcoma Osteosarcoma, Ewing's Sarcoma, and Small Cell Lung Cancer
Phase: PHASE2
Conditions: Soft Tissue Sarcoma
Sponsor: Washington University School of Medicine

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A Phase II Trial of ADI-PEG 20 in Combination with Gemcitabine and Docetaxel for the 
Treatment of Soft Tissue Sarcoma, Osteosarcoma, Ewing’s Sarcoma, and Small Cell Lung 
Cancer 
 
Washington University School of Medicine 
Division of Oncology 
660 South Euclid Avenue, Campus Box 8056 
St. Louis, MO  63110 
 
Protocol #: 201912062 (previously 201803202) 
Version Date: 11/04/2025 
 
Coordinating Center: 
Washington University School of Medicine 
 
Principal Investigator: 
Mia Weiss, M.D. 
Phone: (314) 362-5737 
E-mail:  m.c.weiss@wustl.edu  
 
Sub-Investigators 
 
Institution 
 
 
Modality 
Doug Adkins, M.D.  
Washington University 
Medical Oncology 
Angie Hirbe, M.D. 
 
Washington University 
Medical Oncology 
Nam Bui, M.D. 
 
Stanford Cancer Institute 
Medical Oncology 
Steven Wong, M.D.  
Sarcoma Oncology Center 
Medical Oncology 
Jingqin (Rosy) Luo, Ph.D. 
Washington University 
Biostatistics 
 
Pharmacy Co-Chair: 
Sasha Haarberg, PharmD 
 
 
 
 
Phone: (314) 273-2918 
 
 
 
 
E-mail: sasha.haarberg@wustl.edu  
 
Study Drugs:  
 
ADI-PEG 20 (pegylated arginine deiminase) 
Gemcitabine (Gemzar) 
Docetaxel (Taxotere) 
 
IND #: 
138701 
ClinicalTrials.gov #: 
NCT03449901 
 
 
CONFIDENTIAL 
The information contained in this document is regarded as confidential and, except to the extent necessary to 
obtain informed consent, may not be disclosed to another party unless law or regulations require such 
disclosure.  Persons to whom the information is disclosed must be informed that the information is 
confidential and may not be further disclosed by them 

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A Phase II Trial of ADI-PEG 20 in Combination with Gemcitabine and Docetaxel for the 
Treatment of Soft Tissue Sarcoma, Osteosarcoma, Ewing’s Sarcoma, and Small Cell Lung 
Cancer 
 
Protocol Revision History 
 
Initial Approval Version 
 
 
 
 
 
 
03/28/2018 
Amendment #1 Version 
 
 
 
 
 
 
05/07/2018 
Amendment #2 Version 
 
 
 
 
 
 
05/14/2018 
Amendment #3 Version 
 
 
 
 
 
 
06/20/2018 
Amendment #4 Version 
 
 
 
 
 
 
08/24/2018 
Amendment #5 Version 
 
 
 
 
 
 
03/15/2019 
Amendment #6 Version 
 
 
 
 
 
 
06/06/2019 
Amendment #7 Version 
 
 
 
 
 
 
08/25/2019 
Amendment #8 Version 
 
 
 
 
 
 
07/10/2020 
Amendment #9 Version 
 
 
 
 
 
 
08/10/2020 
Amendment #10 Version 
 
 
 
 
 
 
11/04/2025 
 
 

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A Phase II Trial of ADI-PEG 20 in Combination with Gemcitabine and Docetaxel for the 
Treatment of Soft Tissue Sarcoma, Osteosarcoma, Ewing’s Sarcoma, and Small Cell Lung 
Cancer 
 
Principal Investigator Signature Page 
 
 
Principal Investigator 
(printed): 
      
Name of Institution:       
  
 PI Signature 
 
 
 
 
 
Date 
 
By my signature, I agree to personally supervise the conduct of this study 
and to ensure its conduct in compliance with the protocol, informed 
consent, IRB/HRPO procedures, the Declaration of Helsinki, ICH Good 
Clinical Practices guidelines, and the applicable parts of the United 
States Code of Federal Regulations or local regulations governing the 
conduct of clinical studies. 
 
 
 

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A Phase II Trial of ADI-PEG 20 in Combination with Gemcitabine and Docetaxel for the 
Treatment of Soft Tissue Sarcoma, Osteosarcoma, Ewing’s Sarcoma, and Small Cell Lung 
Cancer 
 
 
SCHEMA 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
*ADI-PEG 20 is to be given weekly regardless to Day 1 or Day 8 dose delays. This could 
cause cycles that extend beyond 21 days. 
 
 
 
 
 
 
Treatment Plan 
Patients will receive ADI-PEG 20 on Day -7 of Cycle 
1 and Days 1, 8, and 15 of each 21-day cycle.*  They 
will also receive gemcitabine on Days 1 and 8 of each 
cycle and docetaxel on Day 8 of each cycle. 
Eligible Patients 
Cohort 1: Histologically or cytologically confirmed grade 2 or 3 soft 
tissue sarcoma that is unresectable or metastatic that would be 
standardly treated with gemcitabine or gemcitabine and docetaxel 
 
Cohort 2: Histologically or cytologically confirmed osteosarcoma, 
Ewing’s sarcoma, or small cell lung cancer that is unresectable or 
metastatic that have either failed standard of care therapy or would be 
standardly treated with gemcitabine or gemcitabine and docetaxel 

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Table of Contents 
SCHEMA ........................................................................................................................................ 4 
1.0 
BACKGROUND AND RATIONALE ................................................................................ 7 
1.1 
Sarcoma ............................................................................................................................ 7 
1.2 
Treatment of Metastatic Sarcoma .................................................................................... 7 
1.3 
ADI-PEG 20 ..................................................................................................................... 7 
1.4 
Gemcitabine ..................................................................................................................... 9 
1.5 
Docetaxel ........................................................................................................................ 10 
1.6 
Gemcitabine and Docetaxel Combination ...................................................................... 10 
1.7 
Study Rationale .............................................................................................................. 10 
1.8 
Correlative Studies Background..................................................................................... 13 
1.9 
Rationale for Cohort 2 .................................................................................................... 14 
2.0 
OBJECTIVES .................................................................................................................... 14 
2.1 
Primary Objective .......................................................................................................... 14 
2.2 
Secondary Objectives ..................................................................................................... 14 
2.3 
Exploratory Objectives ................................................................................................... 14 
3.0 
PATIENT SELECTION .................................................................................................... 14 
3.1 
Inclusion Criteria ............................................................................................................ 15 
3.2 
Exclusion Criteria ........................................................................................................... 16 
3.3 
Inclusion of Women and Minorities ............................................................................... 17 
4.0 
REGISTRATION PROCEDURES ................................................................................... 17 
4.1 
Confirmation of Patient Eligibility ................................................................................. 17 
4.2 
Patient Registration in the Siteman Cancer Center OnCore Database ........................... 18 
4.3 
Assignment of UPN ....................................................................................................... 18 
5.0 
TREATMENT PLAN ........................................................................................................ 18 
5.1 
Premedication Administration ........................................................................................ 18 
5.2 
Agent Administration ..................................................................................................... 19 
5.3 
Evaluability .................................................................................................................... 19 
5.4 
General Concomitant Medication and Supportive Care Guidelines .............................. 19 
5.5 
Women of Childbearing Potential .................................................................................. 20 
5.6 
Duration of Therapy ....................................................................................................... 20 
5.7 
Duration of Follow-up .................................................................................................... 21 
5.8 
Study Stopping Rule....................................................................................................... 21 
6.0 
DOSE DELAYS/DOSE MODIFICATIONS .................................................................... 21 
6.1 
Dose Modifications for ADI-PEG 20............................................................................. 21 
6.2 
Dose Modifications for Gemcitabine ............................................................................. 21 
6.3 
Dose Modifications for Docetaxel ................................................................................. 22 
7.0 
REGULATORY AND REPORTING REQUIREMENTS ............................................... 23 
7.1 
Sponsor-Investigator Reporting Requirements .............................................................. 23 
7.2 
Secondary Site Reporting Requirements ........................................................................ 25 
7.3 
Exceptions to Expedited Reporting ................................................................................ 26 
8.0 
PHARMACEUTICAL INFORMATION .......................................................................... 26 
8.1 
ADI-PEG 20 ................................................................................................................... 26 
8.2 
Gemcitabine (Gemzar) ................................................................................................... 28 
8.3 
Docetaxel (Taxotere) ...................................................................................................... 29 
9.0 
CORRELATIVE STUDIES .............................................................................................. 31 

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9.1 
Tumor Biopsy ................................................................................................................. 31 
9.2 
Archival Tissue .............................................................................................................. 32 
9.3 
Mandatory Peripheral Blood .......................................................................................... 32 
10.0 STUDY CALENDAR ....................................................................................................... 33 
11.0 DATA SUBMISSION SCHEDULE ................................................................................. 34 
11.1 Adverse Event Collection in the Case Report Forms ..................................................... 34 
12.0 MEASUREMENT OF EFFECT........................................................................................ 34 
12.1 RECIST 1.1 .................................................................................................................... 35 
12.2 Choi Response Criteria ................................................................................................... 41 
13.0 DATA AND SAFETY MONITORING ............................................................................ 41 
14.0 AUDITING ........................................................................................................................ 42 
15.0 STATISTICAL CONSIDERATIONS............................................................................... 43 
15.1 Study Design .................................................................................................................. 43 
15.2 Endpoints ........................................................................................................................ 43 
15.3 Sample Size Justification ............................................................................................... 44 
15.4 Accrual ........................................................................................................................... 44 
15.5 Analysis Plan .................................................................................................................. 44 
15.6 Study Stopping Rules ..................................................................................................... 44 
15.7 Population for Analysis .................................................................................................. 45 
16.0 MULTICENTER REGULATORY REQUIREMENTS ................................................... 46 
17.0 REFERENCES .................................................................................................................. 47 
APPENDIX A: ECOG Performance Status Scale ........................................................................ 49 
APPENDIX B: Definitions for Adverse Event Reporting ............................................................ 50 
APPENDIX C: Reporting Timelines ............................................................................................ 52 
APPENDIX D: Washington University Unanticipated Problem Reporting Cover Sheet ............ 55 
 
 

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1.0 
BACKGROUND AND RATIONALE 
 
1.1 
Sarcoma 
 
Sarcomas are a heterogeneous group of rare cancers that share a common mesenchymal 
origin.  Mesenchymal cells are derived from pluripotent embryonic connective tissue that 
differentiates into one or several lineages such as muscle, adipose, cartilage, nerves, 
vascular tissue, etc.  They are broadly divided into bone sarcomas and soft tissue sarcomas.  
The incidence of bone and soft tissue sarcomas in 2016 was 12,110 and 3,010, respectively 
1.  There are more than 50 histologic subtypes of soft tissue sarcomas, each with a 
distinctive clinical profile, response to treatment, and prognosis. 
 
1.2 
Treatment of Metastatic Sarcoma 
 
Generally, localized sarcoma is treated with surgery when feasible and adjuvant radiation 
with or without chemotherapy.  Like any other cancer, sarcomas have the ability for 
hematogenous spread to develop distant metastases.  For the majority of patients with 
metastatic soft tissue sarcoma, their cancer is incurable. Therefore, the goal of treatment is 
to reduce cancer bulk, minimize cancer related symptoms, improve quality of life, and 
increase overall survival (OS).  Despite our best efforts, the median survival of patients 
with distant metastases is 11 to 15 months with approximately 20-25% of patients being 
alive at three years 2.   
 
Most cases of advanced soft tissue sarcoma are treated with conventional chemotherapy 
for palliation.  Common first line treatment regimens are based on the agent doxorubicin.  
There have been many attempts to improve upon this treatment approach with dose 
intensification and combination therapies.  A randomized trial by EORTC Soft Tissue and 
Bone Sarcoma Group compared doxorubicin alone, doxorubicin plus ifosfamide, and a 
four-drug regimen of cyclophosphamide, vincristine, doxorubicin, and dacarbazine.  The 
combination treatment did not significantly improve OS, progression-free survival (PFS), 
or response rate (RR), and these combination regimens were significantly more toxic3.  
Another randomized phase III trial evaluated single agent doxorubicin compared to 
doxorubicin plus ifosfamide in advanced or metastatic soft tissue sarcoma.  There was no 
significant difference in OS between the two treatment groups (median OS 12.8 months 
vs. 14.3 months, p=0.076).  The combination chemotherapy group had a significantly 
higher PFS (7.4 months vs. 4.6 months, p 0.003) and there was a higher overall response 
rate in the combination treatment arm (26% vs. 14%)3. Olaratumab has demonstrated an 
overall survival benefit in a randomized phase II trial4. The combination of gemcitabine 
(900 mg/m2 every day 1 and 8 of a 21 day cycle) with docetaxel (day 8) is also 
recommended as a second line treatment option in soft tissue sarcoma (NCCN Guidelines 
2016) 5. This is the treatment that will be tested in this protocol in combination with ADI-
PEG 20. 
  
1.3 
ADI-PEG 20 
 
ADI-PEG 20 is arginine deiminase (ADI) conjugated to polyethylene glycol (PEG) of 

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20,000 molecular weight (pegargiminase [United States Adopted Names Council Name]).  
It is a novel biological therapeutic agent currently being developed to treat unresectable 
hepatocellular carcinoma (HCC), metastatic melanoma and other cancers. 
 
ADI is a microbial enzyme that degrades arginine, a crucial amino acid central to 
metabolism and biosynthesis and required for neoplastic growth and development.  While 
normal human cells are able to synthesize arginine from metabolic precursors, some cancer 
cells such as melanoma and HCC are deficient in the necessary enzymatic pathway to 
make arginine due to loss of expression of argininosuccinate synthetase (ASS1) in the urea 
cycle and must instead obtain arginine from the blood for growth and survival.  As such, 
these cancers are auxotrophic for arginine. Therefore, depleting arginine from the blood 
can control tumor growth and even eliminate arginine requiring cancers without damage to 
normal cells. 
 
There is precedence for the use of an amino acid degrading enzyme as an effective 
treatment for specific forms of cancer which are auxotrophic for non-essential amino acids.  
Conjugation to PEG (20,000 mw) increases the circulating half-life and decreases the 
antigenicity of ADI.  Similar pegylation technology has been used with a number of 
microbial derived therapeutic proteins for the same reasons.  For example, a pegylated 
form of the microbial derived, asparagine-degrading enzyme asparaginase has been used 
as front-line therapy against childhood leukemia for over 30 years with few serious side 
effects.  
 
ADI-PEG 20 has shown efficacy in mice implanted with tumors that require arginine for 
growth.  ADI-PEG 20 also has been effective in treating dogs with spontaneous melanoma.  
A number of tumor cell lines from a variety of human tumors are auxotrophic for arginine, 
including melanomas and HCC.  ADI-PEG 20 has been tested in Phase 1 & 2 trials in 
the USA and in Europe in metastatic melanoma and in unresectable HCC in the USA, 
Europe and Taiwan, and was tested in a global Phase 3 study in HCC. Results from 
several completed and ongoing clinical studies have shown that ADI-PEG 20 is well-
tolerated in humans.   
 
Since humans do not appear to require arginine for survival and growth, ADI-PEG 
20 may have utility in the treatment of cancers that are auxotrophic for arginine.  Arginine 
depletion with ADI-PEG 20 in cancers has recently been reviewed 6 , 7 . 
 
More specifically, ADI-PEG 20 is also being evaluated in multiple phase 1 combination 
studies with single and double chemotherapy agents, including docetaxel (Tomlinson 
2015), and gemcitabine plus nab-paclitaxel in pancreatic carcinoma (ClinicalTrials.gov # 
NCT02101580). This combination has been well tolerated and has moved into the post-
dose escalation cohort with the standard doses of gemcitabine and nab-paclitaxel used in 
pancreatic cancer (1000 mg/m2 and 125 mg/m2, respectively, on days 1, 8, and 15 every 4 
weeks) and the highest dose of ADI-PEG 20 (36 mg/m2) tested (Lowery 2017 Cancer-in 
press). Thus, in that study, patients received 9000 mg/m2 of gemcitabine total every 12 
weeks, in addition to nab-paclitaxel and weekly ADI-PEG 20 36 mg/m2. All planned 
subjects have been accrued and treated. The triplet has been well tolerated and has resulted 

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in numerous objective responses (Lowery 2017 Cancer-in press). In this study, patients 
receive of gemcitabine every 12 weeks and weekly ADI-PEG 20 36 mg/m2 along with nab-
paclitaxel. As such, there is precedent for the combination of weekly ADI-PEG 20 36 
mg/m2 and gemcitabine days 1 and 8 of 21 and a taxane as proposed in this phase 2 study. 
 
Thus this metabolic targeting of tumor cell metabolism as an anticancer strategy may 
potentially attack cancer’s Achilles’ heel. Therefore, anticancer PEG enzymes are a current 
approach to therapy of cancer 8. 
 
Recent trials of ADI-PEG 20 looked at toxicity in combination with taxanes and 
gemcitabine and saw that ADI-PEG 20 did not increase toxicity.  First, in a phase I trial of 
ADI-PEG 20, doses of 18 mg/m2, 27 mg/m2, or 36 mg/m2 were given together with 
pemetrexed 500 mg/m2 and cisplatin 75 mg/m2; no dose-limiting toxicities of ADI-PEG20 
were reported.9 Second, in a phase I trial, patients received ADI-PEG 20 weekly via 
intramuscular injection at doses ranging from 4.5 to 36 mg/m2 and up to 10 doses of 
docetaxel (75 mg/m2) every 3 weeks; ADI-PEG 20 demonstrated reasonable toxicity in 
combination with docetaxel 10.  Finally, in a trial most similar to ours, Phase 1/1B Trial of 
ADI-PEG 20 Plus Nab-Paclitaxel and Gemcitabine in Patients with Advanced Pancreatic 
Adenocarcinoma, the RP2D for ADI-PEG 20 was 36 mg/m2 weekly in combination with 
standard-dose gemcitabine and nab-paclitaxel. Overall, they concluded that ADI-PEG 20 
was well tolerated in combination with gemcitabine and nab-paclitaxel 11.  This data 
provides the rationale for not performing a phase I safety lead in. 
 
1.4 
Gemcitabine 
 
Gemcitabine is a nucleoside metabolic inhibitor indicated in combination with carboplatin 
for the treatment of advanced ovarian cancer that has relapsed at least 6 months after 
completion of platinum-based therapy; in combination with paclitaxel for first-line 
treatment of metastatic breast cancer after failure of prior anthracycline-containing 
adjuvant chemotherapy (unless anthracyclines were clinically contraindicated); in 
combination with cisplatin for the treatment of non-small cell lung cancer; and as a single 
agent for the treatment of pancreatic cancer.  It is also used as a single agent or in 
combination with docetaxel for the treatment of soft tissue sarcoma 12, 5. 
 
The most common (≥20%) adverse reactions of single-agent gemcitabine are 
nausea/vomiting, anemia, increased ALT, increased AST, neutropenia, increased alkaline 
phosphatase, proteinuria, fever, hematuria, rash, thrombocytopenia, dyspnea, and edema. 
The most common (≥5%) Grade 3 or 4 adverse reactions were neutropenia, 
nausea/vomiting; increased ALT, increase alkaline phosphatase, anemia, increased AST, 
and thrombocytopenia. Approximately 10% of the 979 patients discontinued gemcitabine 
due to adverse reactions. Adverse reactions resulting in discontinuation of gemcitabine in 
2% of 979 patients were cardiovascular adverse events (myocardial infarction, 
cerebrovascular accident, arrhythmia, and hypertension) and adverse reactions resulting in 
discontinuation of gemcitabine in less than 1% of the 979 patients were anemia, 
thrombocytopenia, hepatic dysfunction, renal dysfunction, nausea/vomiting, fever, rash, 
dyspnea, hemorrhage, infection, stomatitis, somnolence, flu-like syndrome, and edema. 

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1.5 
Docetaxel 
 
Docetaxel is a microtubule inhibitor indicated for locally advanced or metastatic breast 
cancer after chemotherapy failure, operable node-positive breast cancer, locally advanced 
or metastatic non-small cell lung cancer after platinum therapy failure, unresectable locally 
advanced or metastatic untreated non-small cell lung cancer, hormone-refractory prostate 
cancer, untreated advanced gastric adenocarcinoma, and locally advanced squamous cell 
head and neck cancer. 
 
The most common adverse reactions are infections, neutropenia, anemia, febrile 
neutropenia, hypersensitivity, thrombocytopenia, neuropathy, dysgeusia, dyspnea, 
constipation, anorexia, nail disorders, fluid retention, asthenia, pain, nausea, diarrhea, 
vomiting, mucositis, alopecia, skin reactions, and myalgia. 
 
1.6 
Gemcitabine and Docetaxel Combination  
 
In 2007, Maki et al published SARC02, a randomized Phase II clinical trial comparing 
gemcitabine versus the combination of gemcitabine with docetaxel 12.  One hundred 
twenty-two patients were randomized using adaptive randomization; 73 received 
gemcitabine and docetaxel and 49 received gemcitabine alone. The progression-free 
survival was 6.2 months for gemcitabine and docetaxel versus 3.0 months for gemcitabine 
alone. The overall survival was 17.9 months for gemcitabine-docetaxel and 11.5 months 
for gemcitabine.   The RECIST Response Rate was 16% (gemcitabine-docetaxel) and 8% 
(gemcitabine). In addition, median overall survival was 17.9 months for gemcitabine-
docetaxel and 11.5 months for gemcitabine.  This established the regimen of gemcitabine 
and docetaxel as a standard second-line therapy for most high grade soft tissue sarcomas 5.  
More recently, the control arm of the Morphoteck clinical trial of gemcitabine, docetaxel, 
and Morb009 demonstrated a PFS of 5.6 months, which was not measurably different than 
the Maki trial.  This left gemcitabine and docetaxel as a standard second-line therapy for 
most high grade soft tissue sarcomas. 
 
1.7 
Study Rationale 
 
In a search for common metabolic alterations that could provide treatment opportunities 
for a broad range of sarcomas, we investigated the expression of metabolic genes that are 
commonly altered in other chemorefractory tumors. We immediately noted that ASS1 
expression is commonly lost in multiple such tumors including hepatocellular carcinoma, 
renal cell carcinoma, and platinum refractory ovarian cancer13. Given the chemorefractory 
nature of sarcoma, we decided to perform an extensive analysis across 45 of the most 
common subtypes in 701 unique tumors (Figure 1A). In collaboration with a sarcoma 
pathologist, we initially planned to apply an intensity score for ASS1 expression (intensity 
score 0-3) similar to that used for other biomarkers such as ER and AR, and similar to that 
used for ASS1 in other tumors where heterogeneous expression is noted. Surprisingly, most 
patients 653/701 (93.2%) diagnosed with bone 34/39 (87.2%) or soft tissue sarcomas 
619/662 (93.5%) tumors showed absolutely no signal for ASS1 expression by IHC. Thus 

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sarcomas harbor a metabolic defect, ASS1 deficiency, which would be anticipated to make 
them uniquely susceptible to arginine depletion therapy with ADI-PEG 20. 
 
 
 
1.7.1 ADI-PEG 20 induces Autophagy 
 
We analyzed markers of autophagy, including LC3 cleavage and p62 accumulation 
by immunoblotting. ADI-PEG 20-treated ASS1low sarcoma cells demonstrated 
increased LC3 cleavage and increased p62 abundance, indicating the induction of 
autophagy after arginine deprivation (Fig. 2A). We then used lentiviral expression 
GFP-LC314 in leiomyosarcoma cell line SKLMS1 and treated with ADI-PEG20 for 
three days (Fig. 2B) and observed a dramatic increase in the number of GFP-LC3 
puncta upon treatment. Western blots show this increase in the formation of LC3 
 
Figure 1 Top. In a panel of primary sarcoma tumors with anti-ASS 
immunohistochemistry, ASS1 lacks strong expression in 619/662 (93.5%) 
primary soft tissue and 34/39 (87.2%) bone sarcoma tumors representing 
45 soft tissue and 5 bone sarcoma histological subtypes. Bottom. 
Immunohistochemistry of MPNST ASS1+ and ASS1- tumor samples 
stained with Hematoxylin & Eosin, and anti-ASS1. 
       
 
Figure 2A. Arginine depletion with ADI-PEG20 induces autophagy by day 
2 as seen by increased LC3 cleavage and p62 alterations in ASS1 
deficient MNNG/HOS and SKLMS-1 cell lines. B. Lentiviral expression of 
GFP-LC3 demonstrates an increase in the number of puncta after ADI-
PEG20 treatment. 

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cleavage, clearly implicating autophagy is a pro-survival mechanism for arginine 
depletion. 
 
1.7.2 ADI-PEG 20 inhibits phospho-PKM2 expression 
 
As part of a metabolic characterization of autophagic changes induced by arginine 
starvation, we treated the sarcoma cell lines SKLMS1 and SKUT1 with ADI-
PEG20 for three days and immunoblotted for PKM2, phospho-PKM2, and PMK1 
(Figure 3). A significant decrease in Phospho-PKM2 was seen using western blot. 
A decrease in phospho-PKM2 is important as it is the phosphorylated form of 
PKM2 that is responsible for the metabolic reprograming of cancer. The decrease 
of phospho-PKM2 would alter the flux of glucose, supporting the idea that 
prolonged ADI-PEG20 treatment decreases the predominant use of glycolysis for 
energy generation. 
 
 
1.7.3 Loss of Phospho-PKM2 diverts glucose to serine biosynthesis for 
nucleotide biosynthesis 
 
Finally, as we had discovered that loss of PKM2 led to glutamine anaplerosis for 
TCA substrates, we asked the question what the fate of glucose is in amino acid 
induced autophagy.   Surprisingly, using Carbon 13 (C13) labeling of glucose at all 
6 positions, we identified that glucose was not fluxing towards lactate, citrate or 
adenosine synthesis, but into serine biosynthesis (Figure 4A).  Serine biosynthesis 
is part of the pyrimidine biosynthetic pathway.  We then measured 
phosphoglycerate dehydrogenase (PHGDH), the first enzyme in the serine 
biosythetic pathway and found a dramatic upregulation.  As PKM2 is known to 
downregulate serine biosynthesis as there is a feedback loop 15, this allows for an 
additional theraputic option.  Indeed, gemcitibine was found to have a lower IC50 
when combined with ADI-PEG20 (data not shown). As nucleotides are derived 
from glucose,  targetting with nuceotide deivitive can also be considered a 
metabolic therapy.  
 
Figure 3: The sarcoma cell lines SKLMs1 and 
SKUT1 were treated for three days with ADI-
PEG20.  Loss of phospho-PKM2 espression is 
demonstated.

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1.7.4 Summary 
 
1) ASS1 is a common metabolic defect in most sarcomas across histologies 
making it the most common defect.  
2) ADI-PEG 20 induces autophagy in ASS1 deficient sarcomas. 
3) Glucose is diverted to the serine biosynthetic pathway for pyrimidine synthesis 
due to inhibition of PKM2 by ADI-PEG20. 
4) Combination therapy driven by a biomarker using metabolism can lead to 
synthetic lethal combinations. 
 
In summary, the compelling preliminary data presented in this proposal provide a 
strong rationale for the clinical development of ADI-PEG 20 with gemcitabine in 
sarcoma patients. Our further understanding of the alterations in metabolism is 
critical for developing a biomarker based multi-agent metabolic therapy 
 
1.8 
Correlative Studies Background 
 
Biopsy will be obtained at Day -7 and Day -1 for metabolomic analysis. There is no day 0 
associated with this protocol. Serum will be collected at Days -7 and -1 in addition to Days 
1 and 8 of each cycle for pharmacodynamics and immunogenicity.  We will be evaluating 
the metabolomics of ADI response in human tissue and performing IHC analyses for ASS1 
A 
 
B.  
           
 
Figure 4: A. Carbon 13 tracing in SKLMS1 cells 
after treatment with ADI-PEG20 demonstrate that 
the carbons for glucose are diverted to the serine 
biosynthetic 
pathway. 
B. 
Phosphoglycerate 
dehydrogenase (PHGDH), the first enzyme in the 
serine biosyntetic pathway involved in the early 
steps of L-serine synthesis that are used to make 
pyrimidines, is upregulated in  autophagy.  

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and phospho-PKM2. 
 
1.9 
Rationale for Cohort 2 
 
Recently published data17 shows that priming ASS1-deficient tumors with ADI-PEG 20 
and docetaxel improves the effect of gemcitabine. Therefore, a cohort of patients consisting 
of ten patients diagnosed with either osteosarcoma or Ewing’s sarcoma (ideally five of 
each), and five patients diagnosed with small cell lung cancer will be included as an 
exploratory cohort. Enrollment to Cohort 2 will occur concurrently with Cohort 1.  
 
 
2.0 
OBJECTIVES 
 
2.1 
Primary Objective 
 
To determine progression-free survival in patients with soft tissue sarcoma who are treated 
with ADI-PEG 20 in combination with gemcitabine and docetaxel. 
 
2.2 
Secondary Objectives 
 
1. To determine overall survival in patients with soft tissue sarcoma who are treated with 
ADI-PEG 20 in combination with gemcitabine and docetaxel. 
2. To determine the clinical benefit rate (CBR) of patients with soft tissue sarcoma 
receiving treatment with ADI-PEG 20 in combination with gemcitabine and docetaxel 
(CBR = the percentage of patients who have experienced CR+PR+SD). 
3. To evaluate the safety and tolerability of the combination of ADI-PEG 20, gemcitabine, 
and docetaxel (by CTCAE v 5.0) in soft tissue sarcoma, osteosarcoma, Ewing’s 
sarcoma and small cell lung cancer. 
4. To determine cancer-related mortality in patients with soft tissue sarcoma who are 
treated with ADI-PEG 20 in combination with gemcitabine and docetaxel. 
 
2.3 
Exploratory Objectives 
 
1. To determine the effects of treatment with ADI-PEG 20, gemcitabine, and docetaxel 
on arginine, citrulline, and glutamine plasma levels. 
2. To determine the rate of immunogenicity against ADI-PEG 20. 
3. To evaluate the metabolomics of paired biopsies taken before and after receipt of ADI-
PEG 20. 
4. To correlate response (CBR) to ADI-PEG 20 with phospho-PKM2 status. 
5. To correlate response (CBR) to ADI-PEG 20 with ASS1 negativity in clinical samples. 
6. To correlate response (CBR) to ADI-PEG 20 by Choi criteria16. 
 
 
3.0 
PATIENT SELECTION 
 
Each of the criteria in the checklist that follows must be met in order for a patient to be considered 

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eligible for this study.  Use the checklist to confirm a patient’s eligibility.  For each patient, this 
checklist must be photocopied, completed and maintained in the patient’s chart. 
 
Patient’s Initials (F,M,L)_________________________ 
 
NOTE:  All questions regarding eligibility should be directed to the principal investigator, Mia 
Weiss, M.D. or the primary coordinator, Jacqui Toeniskoetter. 
 
NOTE: Institutions may use the eligibility checklist as source documentation if it has been 
reviewed, signed, and dated prior to registration by the treating physician. 
 
3.1 
Inclusion Criteria 
 
_____  1. Cohort 1: Histologically or cytologically confirmed grade 2 or 3 soft tissue 
sarcoma that is unresectable or metastatic that would be standardly treated with 
gemcitabine or gemcitabine and docetaxel.  For all others, please contact the 
principal investigator. Prior surgery for primary or metastatic disease after 
chemotherapy following a response is allowed.   
 
 
Cohort 2: Histologically or cytologically confirmed osteosarcoma, Ewing’s   
sarcoma, or small cell lung cancer that is unresectable or metastatic that have 
either failed standard of care therapy or would be standardly treated with 
gemcitabine or gemcitabine and docetaxel. 
 
_____  2. Measurable disease defined as lesions that can be accurately measured in at   
Least one  dimension (longest diameter to be recorded) as ≥ 10 mm with CT 
scan, as ≥ 20 mm by chest x-ray, or ≥ 10 mm with calipers by clinical exam. 
 
_____  3. Treated with at least one previous line of systemic therapy. The allowable 
window between treatments is  21 days for chemotherapy or a TKI or 5 half-
lives for a TKI (whichever is shorter), 21 days and progression by CT for 
immunotherapy, 21 days for RT, 21 days for surgery, or 28 days for an 
investigational agent. 
 
_____  4.  Cohort 1: At least 16 years of age. 
 
Cohort 2: Patients with osteosarcoma or Ewing’s sarcoma must be at least 10 
years of age. Patients with small cell lung cancer must be at least 18 years of age. 
 
_____  5.  Cohort 2 (SCLC group ONLY): Must be amenable to biopsy 
 
_____  6.  ECOG performance status ≤ 2 (see Appendix A) 
 
_____  7.  Normal bone marrow and organ function as defined below: 
_____ a. Leukocytes ≥ 3,000/mcL 
_____ b. Absolute neutrophil count ≥ 1,500/mcl 

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_____ c. Platelets ≥ 100,000/mcl 
_____ d. Total bilirubin ≤ 2 x IULN 
_____ e. AST(SGOT)/ALT(SGPT) ≤ 3 x IULN (or ≤ 5 x IULN if liver 
metastases are present) 
_____ f. Creatinine ≤ 1.5 x IULN OR Creatinine clearance ≥ 60 mL/min/1.73 
m2 for patients with creatinine levels above institutional normal 
_____ g. Serum uric acid ≤ 8 mg/dL (with or without medication control) 
 
_____ 8. Women of childbearing potential and men must agree to use adequate  
contraception (hormonal or barrier method of birth control, abstinence) prior to 
study entry and for the duration of study participation.  Should a woman become 
pregnant or suspect she is pregnant while participating in this study, she must 
inform her treating physician immediately. 
 
_____  9.  Ability to understand and willingness to sign an IRB approved written informed 
consent document (or that of legally authorized representative, if applicable). 
 
3.2 
Exclusion Criteria 
 
_____  1.    A history of other high grade malignancy ≤ 5 years previous. Exceptions include 
basal cell or squamous cell carcinoma of the skin which were treated with local 
resection only or carcinoma in situ of the cervix, or other tumors discussed with 
the study PI. 
 
_____  2.  Currently receiving any other investigational agents. 
 
_____  3.  Prior treatment with ADI-PEG 20, gemcitabine, or docetaxel. Patients treated > 
one year ago in the adjuvant/neoadjuvant setting with gemcitabine or docetaxel 
would be allowed to be enrolled on trial. 
 
_____  4.  Known brain metastases.  Patients with known brain metastases must be 
excluded from this clinical trial (except for patients with SCLC, see below) 
because of their poor prognosis and because they often develop progressive 
neurologic dysfunction that would confound the evaluation of neurologic and 
other adverse events. 
 
Patients with SCLC are allowed to enroll with brain metastases provided they 
are stable and they are at least 3 months post-treatment for brain metastases. 
 
_____  5.  A history of allergic reactions attributed to compounds of similar chemical or 
biologic composition to ADI-PEG 20, gemcitabine, pegylated compounds, or 
other agents used in the study. 
 
_____  6.  Uncontrolled intercurrent illness including, but not limited to, ongoing or active 
infection, symptomatic congestive heart failure, unstable angina pectoris, 

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cardiac\arrhythmia, or psychiatric illness/social situations that would limit 
compliance with study requirements. 
 
_____  7.  History of seizure disorder not related to underlying cancer. 
 
_____  8.  Patients with Grade 2 or higher neuropathy 
 
_____  9.  Pregnant and/or breastfeeding.  Women of childbearing potential must have a 
negative pregnancy test within 14 days of study entry. 
 
_____ 10. Known HIV-positivity on combination antiretroviral therapy because of the 
potential for pharmacokinetic interactions with the study treatment.  In addition, 
these patients are at increased risk of lethal infections when treated with 
marrow-suppressive therapy.  Appropriate studies will be undertaken in patients 
receiving combination antiretroviral therapy when indicated. 
 
 
___________________________________________ 
   Physician Name 
 
 
___________________________________________ 
______________________ 
 
 
            Physician Signature                                                          Date 
 
3.3 
 Inclusion of Women and Minorities 
 
Both men and women and members of all races and ethnic groups are eligible for this trial. 
 
 
4.0 
REGISTRATION PROCEDURES 
 
Patients must not start any protocol intervention prior to registration through the Siteman 
Cancer Center. 
 
The following steps must be taken before registering patients to this study: 
 
1. Confirmation of patient eligibility by Washington University 
2. Registration of patient in the Siteman Cancer Center database 
3. Assignment of unique patient number (UPN) 
 
Once the patient has been entered in the Siteman Cancer Center OnCore database, the WUSM 
coordinator will forward verification of enrollment and the UPN via email. 
 
4.1 
Confirmation of Patient Eligibility 
 
Confirm patient eligibility by collecting the information listed below and scanning and 
emailing it to the research coordinator listed in the Siteman Cancer Center Clinical Trials 

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Core Protocol Procedures for Secondary Sites packet at least one business day prior to 
registering patient: 
 
1. Name and contact information (telephone number, fax number, and email address) 
2. Site PI’s name, the registering MD’s name, and the institution name 
3. Patient’s race, sex, and DOB 
4. Three letters (or two letters and a dash) for the patient’s initials 
5. Currently approved protocol version date 
6. Copy of signed consent form (patient name may be blacked out) 
7. Planned date of enrollment 
8. Completed eligibility checklist, signed and dated by a member of the study team 
9. Copy of appropriate source documentation confirming patient eligibility 
 
4.2 
Patient Registration in the Siteman Cancer Center OnCore Database 
 
Registrations may be submitted Monday through Friday between 8am and 5pm CT. Urgent 
late afternoon or early morning enrollments should be planned in advance and coordinated 
with the Washington University research coordinator.  Registration will be confirmed by 
the research coordinator or his/her delegate by email within one business day. Verification 
of eligibility and registration should be kept in the patient chart. 
 
All patients at all sites must be registered through the Siteman Cancer Center OnCore 
database at Washington University. 
 
4.3 
Assignment of UPN 
 
Each patient will be identified with a unique patient number (UPN) for this study.  Patients 
will also be identified by first, middle, and last initials.  If the patient has no middle initial, 
a dash will be used on the case report forms (CRFs).  All data will be recorded with this 
identification number on the appropriate CRFs. 
 
 
5.0 
TREATMENT PLAN 
 
5.1 
Premedication Administration 
 
It is recommended that patients be pre-medicated for docetaxel with dexamethasone 8 mg 
PO BID (or per institutional standards) the day before docetaxel administration and the day 
after docetaxel administration.  It is also recommended that patients receive IV 
dexamethasone (10 mg recommended or per institutional standards) on day of docetaxel 
administration in order to reduce the incidence and severity of fluid retention as well as the 
severity of hypersensitivity reactions for both gemcitabine and docetaxel.  Modifications 
to the recommended premedication regimen may be made after discussion with the PI.  
 
All patients should be pre-medicated for gemcitabine on Days 1 and 8 for anti-emesis per 
institutional practice.  

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There is no recommended premedication for ADI-PEG 20 except for Diphenhydramine 
50mg PO on Days 1, 8, +15, if prior reaction. 
 
5.2 
Agent Administration 
 
ADI-PEG 20 will be given on Day -7 of Cycle 1 and then on Days 1, 8, and 15 of each 
subsequent cycle.  Cycles are 21 days.   ADI-PEG 20 is to be given weekly regardless to 
Day 1 or Day 8 dose delays. This could cause cycles that extend beyond 21 days. 
 
ADI-PEG 20 will be given on an outpatient basis at a dose of 36 mg/m2 via intramuscular 
injection into either the deltoid or gluteal muscle.  
 
Gemcitabine will be given intravenously at a dose of 600 mg/m2 over 90 (± 10 minutes) 
minutes on Days 1 and 8 of each cycle.  Docetaxel will be given intravenously at a dose of 
60 mg/m2 over 60 minutes (± 10 minutes) on Day 8 of each cycle. 
 
Patients started on gemcitabine at a dose of 900 mg/m2 or 750 mg/m2 or docetaxel at a dose 
of 75 mg/m2 per previous protocol version will be allowed to continue at that dose level 
provided they are not experiencing unacceptable toxicity and the treating physician feels it 
is in their best interests to continue at that dose level.  
 
On days of doublet or triplet dosing, ADI-PEG 20 is to be given first, followed by 
gemcitabine and then, if indicated, by docetaxel. 
 
Growth factor support to be initiated on Day 9 is recommended with either pegfilgrastim, 
Neupogen or equivalent. 
 
After Cycle 8, patients may continue on ADI-PEG 20 alone (without gemcitabine and 
docetaxel) upon request. 
 
5.3 
Evaluability 
 
All patients who receive any study treatment are evaluable for toxicity.  Patients are 
evaluated from first receiving study treatment until a 30-day follow up after the conclusion 
of treatment or death. 
 
All Cohort 1 patients who receive gemcitabine are evaluable for disease response unless 
they discontinue treatment due to treatment related adverse events(s) prior to completion 
of Cycle 2 and have not had any disease assessment. Any Cohort 1 patient who does not 
receive gemcitabine will be replaced.  Cohort 2 patients will not be replaced. 
 
5.4 
General Concomitant Medication and Supportive Care Guidelines 
 

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It is advised that uric acid levels should be routinely tested and that prompt treatment, 
including the administration of fluids, allopurinol, and/or urate oxidase, should be provided 
as clinically indicated to treat suspected tumor lysis syndrome. 
 
5.5 
Women of Childbearing Potential 
 
Women of childbearing potential (defined as women with regular menses, women with 
amenorrhea, women with irregular cycles, women using a contraceptive method that 
precludes withdrawal bleeding, and women who have had a tubal ligation) are required to 
have a negative pregnancy test within 14 days prior to the first dose of ADI-PEG 20.   
 
Female and male patients (along with their female partners) are required to use two forms 
of acceptable contraception, including one barrier method, during participation in the study 
and for 1 month following the last dose of any study agent.  
 
If a patient is suspected to be pregnant, treatment should be immediately discontinued.  In 
addition, a positive urine test must be confirmed by a serum pregnancy test.  If it is 
confirmed that the patient is not pregnant, the patient may resume dosing. 
 
If a female patient or female partner of a male patient becomes pregnant during therapy or 
within 1 months after the last dose of any study agent, the investigator must be notified in 
order to facilitate outcome follow-up, and the investigator will be responsible for reporting 
to Polaris per Section 7.6. 
 
5.6 
Duration of Therapy 
 
If at any time the constraints of this protocol are considered to be detrimental to the 
patient’s health and/or the patient no longer wishes to continue protocol therapy, the 
protocol therapy should be discontinued and the reason(s) for discontinuation documented 
in the case report forms. 
 
In the absence of treatment delays due to adverse events, treatment may continue for up to 
34 cycles or 103 weeks or until one of the following criteria applies: 
 
• Documented and confirmed disease progression (by RECIST 1.1) 
• Death 
• Adverse event(s) that, in the judgment of the investigator, may cause severe or 
permanent harm or which rule out continuation of study drug 
• General or specific changes in the patient’s condition render the patient unable to 
receive further treatment in the judgment of the investigator 
• Suspected pregnancy 
• Serious non-compliance with the study protocol 
• Lost to follow-up 
• Patient withdraws consent 
• Investigator removes the patient from study 
• The Siteman Cancer Center decides to close the study 

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• At the request of Polaris Pharmaceuticals  
 
Patients who prematurely discontinue treatment for any reason will be followed as 
indicated in the study calendar for survival. 
 
5.7 
Duration of Follow-up 
 
Patients will be followed as per standard of care. Patients removed from study for 
unacceptable adverse events will be followed until resolution or stabilization of the adverse 
event.  Data will be collected on progression, at the time of reporting, and survival at 5 
years after the off treatment date by review of the medical record. 
 
5.8 
Study Stopping Rule 
 
Accrual will be paused and the event will be reviewed by the Data and Safety Monitoring 
Board if a grade 5 non-hematological toxicity is observed that is deemed probably or 
definitely related to ADI-PEG 20, gemcitabine, docetaxel, or combination.  Note that sepsis 
is considered a hematological toxicity. 
 
 
6.0 
DOSE DELAYS/DOSE MODIFICATIONS 
 
Dosing adjustments beyond what is specified below may be made to the gemcitabine and docetaxel 
chemotherapy regimen for tolerance after discussion with the PI. 
 
6.1 
Dose Modifications for ADI-PEG 20 
 
There will be no dose modifications for ADI-PEG 20.  ADI-PEG 20 is to be given weekly 
regardless to day 1 or day 8 dose delays. In the event of an allergic reaction or anaphylaxis, 
treat with antihistamines and glucocorticoids.  Treatment with ADI-PEG 20 should be 
immediately discontinued for grade 3 or greater allergic reaction.  Patients who discontinue 
ADI-PEG 20 may continue to receive gem/tax on protocol. 
 
6.2 
Dose Modifications for Gemcitabine 
 
Recommended dose modifications for myelosuppression are described in the table below. 
 
Absolute granulocyte count 
(x 106/L) 
 
Platelet count 
(x 106/L) 
% of full dose 
≥ 1000 
And 
≥ 100,000 
100 
500-999 
Or 
50,000-99,999 
Reduce by 1 
dose level 
< 500 
Or 
< 50,000 
Hold 
 
For patients continuing on gemcitabine 900 mg/m2, if toxicity develops, reduce dose 
to 750 mg/m2 and then follow current protocol dose modification guidelines. For 

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patients continuing on gemcitabine 750 mg/m2, if toxicity develops, reduce dose to 600 
mg/m2 and then follow current protocol dose modification guidelines. 
 
For intolerance or prolonged cytopenias requiring dosing delays of more than 2 weeks, 
reduce gemcitabine to 500 mg/m2 the first time, to 400 mg/m2 the second time, and to 300 
mg/m2 the third time. 
 
Permanently discontinue gemcitabine for any of the following: 
• Unexplained dyspnea or other evidence of severe pulmonary toxicity 
• Severe hepatic toxicity 
• Hemolytic-uremic syndrome 
• Capillary leak syndrome 
• Posterior reversible encephalopathy syndrome 
 
Withhold gemcitabine or reduce dose to 500 mg/m2 for other severe (Grade 3 or 4) non-
hematologic adverse drug reaction until resolved.  No dose modifications are 
recommended for alopecia nausea, or vomiting. At the discretion of the investigator, the 
dose may be increased for future cycles. 
 
If gemcitabine or docetaxel is held on Day 8, dose reduce gemcitabine one dose level at 
next treatment. If dose reduction would result in the patient coming off study, it should be 
done at the principal investigator’s discretion.  
 
6.3 
Dose Modifications for Docetaxel 
 
For patients continuing on docetaxel 75 mg/m2, if toxicity develops, reduce dose to 60 
mg/m2, and then follow current protocol dose modification guidelines. Patients who are 
dosed initially at 60 mg/m2 and who experience either febrile neutropenia, neutrophils < 
500 cells/mm3 for more than 1 week, or severe or cumulative cutaneous reactions during 
docetaxel should have the dosage adjusted from 60 mg/m2 to 50 mg/m2.  If the patient 
continues to experience these reactions, the dosage should either be decreased from 50 
mg/m2 to 40 mg/m2 or their treatment should be discontinued.  Docetaxel should not be 
given if neutrophil counts are < 1500 cells/mm3. 
 
Docetaxel should not be given to patients with bilirubin > ULN or to patients with AST 
and/or ALT > 1.5 x ULN concomitant with alkaline phosphatase > 2.5 x ULN. 
 
Severe hypersensitivity and anaphylaxis require immediate discontinuation of docetaxel. 
 
If grade ≥ 3 peripheral neuropathy occurs, hold docetaxel until symptoms ≤ grade 1, then 
restart at a reduced dose level (if 75 mg/m2, reduce to 60 mg/m2; if 60 mg/m2, reduce to 50 
mg/m2; if 50 mg/m2, reduce to 40 mg/m2; if 40 mg/m2, discontinue docetaxel).  Docetaxel 
can be discontinued independently of gemcitabine for neuropathy. 
 
Docetaxel should not be given to patients with platelets less than 100 K/mm3 
 

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Docetaxel should not be given to patients with hemoglobin < 7.0g/dL 
 
Docetaxel should be dose reduced in discussion with the principal investigator when 
gemcitabine is reduced. After discussion with the principal investigator, reduce docetaxel 
to 50 mg/m2 the first time, 40 mg/m2 the second time, and 30 mg/m2 the third time. 
 
Any other dose modifications or delays should be discussed with the principal 
investigator before continuing treatment.  
 
 
7.0 
REGULATORY AND REPORTING REQUIREMENTS 
 
The entities providing oversight of safety and compliance with the protocol require reporting as 
outlined below. Please refer to Appendix B for definitions and Appendix C for a grid of reporting 
timelines. 
 
Adverse events will be tracked from start of treatment through 30 days following the last day of 
study treatment.  All adverse events must be recorded on the toxicity tracking case report form.  
 
Refer to the data submission schedule in Section 11 for instructions on the collection of AEs in the 
EDC. 
 
Reporting requirements for Washington University study team may be found in Section 7.1.  
Reporting requirements for secondary site study teams participating in Washington University-
coordinated research may be found in Section 7.2. 
 
7.1 
Sponsor-Investigator Reporting Requirements 
 
7.1.1 Reporting to the Human Research Protection Office (HRPO) at 
Washington University 
 
Reporting will be conducted in accordance with Washington University IRB 
Policies. 
 
Pre-approval of all protocol exceptions must be obtained prior to implementing the 
change. 
 
7.1.2 Reporting to the Quality Assurance and Safety Monitoring 
Committee (QASMC) at Washington University 
 
The Washington University Sponsor-Investigator (or designee) is required to notify 
the QASMC of any unanticipated problems involving risks to participants or others 
occurring at WU or any BJH or SLCH institution that has been reported to and 
acknowledged by HRPO.  (Unanticipated problems reported to HRPO and 
withdrawn during the review process need not be reported to QASMC.) 
 

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QASMC must be notified within 10 days of receipt of IRB acknowledgment via 
email to a qasmc@wustl.edu.  Submission to QASMC must include the myIRB 
form and any supporting documentation sent with the form. 
 
For events that occur at secondary sites, the Washington University Sponsor 
Investigator (or designee) is required to notify the QASMC within 10 days of 
Washington University notification via email to qasmc@wustl.edu.  Submission to 
QASMC must include either the myIRB form and supporting documentation or (if 
not submitted to myIRB) the date of occurrence, description of the event, whether 
the event is described in the currently IRB approved materials, the event outcome, 
determination of relatedness, whether currently enrolled participants will be 
notified, and whether the informed consent document and/or any study procedures 
will be modified as a result of this event. 
 
7.1.3 Reporting to Polaris Pharmaceuticals 
 
Polaris will be notified via email of all initial and follow up serious unexpected 
suspect adverse reactions (SUSARs) in real time (within 24 hours of study PI 
notification) and include copy of MedWatch(s).   
 
Polaris will be notified of all AEs and SAEs monthly via AE notification or cross-
reporting reports/documents per usual institutional practice. Report or document 
item requirements/format to be discussed. The reports should be sent via email to 
agreed upon personnel and or Polaris safety email – safety@polarispharma.com.   
 
Polaris will be notified of any study drug overdoses and pregnancies that occur on 
study, as they occur. This will include initial reporting as well as follow up 
(resolution) data. 
 
7.1.4 Reporting to the FDA  
 
The conduct of the study will comply with all FDA safety reporting requirements.  
PLEASE NOTE THAT REPORTING REQUIREMENTS FOR THE FDA 
DIFFER FROM REPORTING REQUIREMENTS FOR HRPO/QASMC.  It 
is the responsibility of the Washington University Sponsor-Investigator to report to 
the FDA as follows: 
 
• Report any unexpected fatal or life-threatening suspected adverse reaction 
(refer to Appendix B for definitions) no later than 7 calendar days after 
initial receipt of the information.   
• Report a suspected adverse reaction that is both serious and unexpected 
(SUSAR, refer to Appendix B) no later than 15 calendar days after it is 
determined that the information qualifies for reporting.  Report an adverse 
event (refer to Appendix B) as a suspected adverse reaction only if there is 
evidence to suggest a causal relationship between the drug and the adverse 
event, such as: 

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o A single occurrence of an event that is uncommon and known to be 
strongly associated with drug exposure 
o One or more occurrences of an event that is not commonly associated 
with drug exposure but is otherwise uncommon in the population 
exposed to the drug 
o An aggregate analysis of specific events observed in a clinical trial that 
indicates those events occur more frequently in the drug treatment 
group than in a concurrent or historical control group 
• Report any findings from epidemiological studies, pooled analysis of 
multiple studies, or clinical studies that suggest a significant risk in humans 
exposed to the drug no later than 15 calendar days after it is determined 
that the information qualifies for reporting. 
• Report any findings from animal or in vitro testing that suggest significant 
risk in humans exposed to the drug no later than 15 calendar days after it 
is determined that the information qualifies for reporting.  
• Report any clinically important increase in the rate of a serious suspected 
adverse reaction of that listed in the protocol or IB within 15 calendar days 
after it is determined that the information qualifies for reporting.    
 
Submit each report as an IND safety report in a narrative format or on FDA Form 
3500A or in an electronic format that FDA can process, review, and archive.  Study 
teams must notify the Siteman Cancer Center Protocol Development team of each 
potentially reportable event within 1 business day after initial receipt of the 
information, and must bring the signed 1571 and FDA Form 3500A to the Siteman 
Cancer Center Protocol Development team no later than 1 business day prior to the 
due date for reporting to the FDA. 
 
Each notification to FDA must bear prominent identification of its contents (“IND 
Safety Report”) and must be transmitted to the review division in the Center for 
Drug Evaluation and Research (CDER) or in the Center for Biologics Evaluation 
and Research (CBER) that has responsibility for review of the IND.  Relevant 
follow-up information to an IND safety report must be submitted as soon as the 
information is available and must be identified as such (“Follow-up IND Safety 
Report”). 
 
7.1.5 Reporting to Secondary Sites 
 
The Washington University Sponsor-Investigator (or designee) will notify the 
research team at each secondary site of all unanticipated problems involving risks 
to participants or others that have occurred at other sites within 10 working days 
of the occurrence of the event or notification of the Sponsor-Investigator (or 
designee) of the event.  This includes events that take place both at Washington 
University and at other secondary sites, if applicable.  Refer to Section 16.0 
(Multicenter Management) for more information 
 
7.2 
Secondary Site Reporting Requirements 

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The research team at each secondary site is required to promptly notify the Washington 
University Sponsor-Investigator and designee of all serious adverse events (refer to 
Appendix B, Section D) within 1 working day of the occurrence of the event or notification 
of the secondary site’s PI of the event.  This notification may take place via email if there 
is not yet enough information for a formal written report (using FDA Form 3500a 
(MedWatch) and Washington University’s cover sheet (Appendix D)).  A formal written 
report must be sent to the Washington University Sponsor-Investigator and designee within 
4 calendar days (for fatal or life-threatening suspected adverse reactions) or 11 calendar 
days (for serious unexpected adverse reactions) of the occurrence of the event or 
notification of the secondary site’s PI of the event. 
 
The research team at a secondary site is responsible for following its site’s guidelines for 
reporting applicable events to its site’s IRB according to its own institutional guidelines.  
The research team at Washington University is responsible for reporting all applicable 
events to the FDA and Polaris Pharmaceuticals as needed. 
 
Washington University pre-approval of all protocol exceptions must be obtained prior to 
implementing the change.  Local IRB approval must be obtained as per local guidelines.  
Washington University IRB approval is not required for protocol exceptions occurring at 
secondary sites. 
 
7.3 
Exceptions to Expedited Reporting 
 
Events that do not require expedited reporting as described in Section 1.1 include: 
• planned hospitalizations 
• hospitalizations < 24 hours 
• respite care 
• events related to disease progression 
 
Events that do not require expedited reporting must still be captured in the EDC. 
 
 
8.0 
PHARMACEUTICAL INFORMATION 
 
8.1 
ADI-PEG 20 
 
8.1.1 ADI-PEG 20 Description 
 
Arginine deiminase (ADI) is a recombinant protein cloned from M. hominis, 
produced in E. coli, and conjugated with PEG of 20,000 mw using a succinimidyl 
succinate linker.  Thus ADI-PEG 20 is an arginine degrading enzyme, ADI, coupled 
to PEG. 
 
8.1.2 Clinical Pharmacology 
 

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The primary mechanism of action of ADI is arginine depletion. 
 
ADI catalyzes the hydrolysis of L-arginine into L-citrulline and ammonia.  For 
tumors that are auxotrophic for arginine due to lack of ASS, the depletion of 
arginine via ADI treatment may lead to inhibition of protein synthesis and eventual 
cell death.  Conversely, normal human cells will be able to utilize L-citrulline to 
synthesize L-arginine via the Krebs urea cycle and therefore should not be affected 
by the ADI treatment. 
 
8.1.3 Pharmacokinetics and Drug Metabolism 
 
Analyses for pharmacodynamics and pharmacokinetics after multiple dosing of 
ADI-PEG 20 were performed in multiple studies.  The pharmacodynamic data 
demonstrate that in most humans, repetitive dosing of ADI-PEG 20 results in 
sustained decreases in peripheral blood arginine, with a concomitant rise in 
citrulline in humans with metastatic melanoma or HCC. 
 
8.1.4 Supplier(s) 
 
ADI-PEG 20 will be provided free of charge by Polaris Pharmaceuticals, Inc. 
 
8.1.5 Dosage Form and Preparation 
 
ADI-PEG 20 is ready for injection without reconstitution.  All vials should be used 
only once and any remaining material present in the vial should be discarded.  
 
The preparation does not need a laminar flow unit.  
 
The drug may be drawn up into a syringe by the pharmacy or the vials may be 
dispensed to a nurse who can draw up the required amount “at the bedside.” 
 
8.1.6 Storage and Stability 
 
ADI-PEG 20 is shipped on dry ice. Histidine-buffered ADI-PEG 20 must be stored 
frozen at -20°C.  The drug can be thawed by one of two methods.   
 
Immediately prior to use each study medication vial may be taken out of the freezer 
and left at room temperature to thaw. This may take approximately 1-2 hours. Once 
thawed and brought to room temperature ADI-PEG 20 is ready for use. If there is 
delay in administering the study medication, the thawed vial may be stored in a 
refrigerator until ready for use. If stored in a refrigerator allow the vial to stand at 
room temperature for at least 10 minutes prior to administration.  
 
Alternatively, each vial may be taken out of the freezer and thawed overnight in the 
refrigerator at 2-8°C prior to the day of administration. Allow the vial to come to 
room temperature for ~10 minutes prior to administration. Once thawed and 

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brought to room temperature ADI-PEG 20 is ready for use. 
 
The study medication vial must be used within 24 hours after removal from the 
freezer.  The study medication is stable at 2-8°C for up to 24 hours. 
 
8.1.7 Administration 
 
ADI-PEG 20 will be given on an outpatient basis as a single intramuscular injection 
into either the deltoid or gluteal muscle. 
 
8.1.8 Special Handling Instructions 
 
No special handling instructions. 
 
8.2 
Gemcitabine (Gemzar) 
 
8.2.1 Gemcitabine Description 
 
Gemcitabine is a nucleoside metabolic inhibitor that exhibits antitumor activity.  
Gemcitabine HCl is 2’-deoxy-2’,2’-difluorocytidine monohydrochloride (β-
isomer).  The structural formula is as follows: 
 
 
 
The empirical formula for gemcitabine HCl is C9H11F2N3O4·HCl.  It has a 
molecular weight of 299.66. 
 
8.2.2 Clinical Pharmacology 
 
Gemcitabine kills cells undergoing DNA synthesis and blocks the progression of 
cells through the G1/S-phase boundary.  Gemcitabine is metabolized by nucleoside 
kinases to diphosphate (dFdCDP) and triphosphate (dFdCTP) nucleosides. 
Gemcitabine diphosphate inhibits ribonucleotide reductase, an enzyme responsible 
for catalyzing the reactions that generate deoxynucleoside triphosphates for DNA 
synthesis, resulting in reductions in deoxynucleotide concentrations, including 
dCTP. Gemcitabine triphosphate competes with dCTP for incorporation into DNA. 
The reduction in the intracellular concentration of dCTP by the action of the 
diphosphate enhances the incorporation of gemcitabine triphosphate into DNA 
(self-potentiation). After the gemcitabine nucleotide is incorporated into DNA, 
only one additional nucleotide is added to the growing DNA strands, which 
eventually results in the initiation of apoptotic cell death. 

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8.2.3 Pharmacokinetics and Drug Metabolism 
 
Gemcitabine disposition was studied in 5 patients who received a single 1000 
mg/m2/30-minute infusion of radiolabeled drug. Within one (1) week, 92% to 98% 
of the dose was recovered, almost entirely in the urine. Gemcitabine (<10%) and 
the inactive uracil metabolite, 2’-deoxy-2’,2’-difluorouridine (dFdU), accounted 
for 99% of the excreted dose. The metabolite dFdU is also found in plasma. 
  
The active metabolite, gemcitabine triphosphate, can be extracted from peripheral 
blood mononuclear cells. The half-life of the terminal phase for gemcitabine 
triphosphate from mononuclear cells ranges from 1.7 to 19.4 hours. 
 
8.2.4 Supplier(s) 
 
Gemcitabine will be given as per routine care from commercial supply. 
 
8.2.5 Dosage Form and Preparation 
 
Gemcitabine for injection, USP, is available in sterile single-use vials individually 
packaged in a carton containing: 200 mg white to off-white lyophilized powder in 
a 10-mL size sterile single use vial or 1 g white to off-white lyophilized powder in 
a 50-mL size sterile single use vial. 
 
8.2.6 Storage and Stability 
 
Unopened vials of gemcitabine are stable until the expiration date indicated on the 
package when stored at controlled room temperature 20°C to 25°C and that allows 
for excursions between 15°C and 30°C. 
 
8.2.7 Administration 
 
Gemcitabine should be given as an intravenous infusion over the course of 90 
minutes (± 10 minutes). 
 
8.2.8 Special Handling Instructions 
 
No special handling instructions. 
 
8.3 
Docetaxel (Taxotere) 
 
8.3.1 Docetaxel Description 
 
Docetaxel is an antineoplastic agent belonging to the taxoid family.  It is prepared 
by semisynthesis beginning with a precursor extracted from the renewable needle 
biomass of yew plants.  The chemical name for docetaxel is (2R,3S)-N-carboxy-3-

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Page 30 of 55 
phenyllisoserine,N-tert-butyl ester, 13-ester with 5β-20-epoxy-1,2α,4,7β,10β,13α-
hexahydroxytax-11-en-9-one 4-acetate 2-benzoate, trihydrate.  Docetaxel has the 
following structural formula: 
 
 
 
Docetaxel is a white to almost-white powder with an empirical formula of 
C43H53NO14·3H2O and a molecular weight of 861.9. 
 
8.3.2 Clinical Pharmacology 
 
Docetaxel is an antineoplastic agent that acts by disrupting the microtubular 
network in cells that is essential for mitotic and interphase cellular functions.  
Docetaxel binds to free tubulin and promotes the assembly of tubulin into stable 
microtubules while simultaneously inhibiting their disassembly.  This leads to the 
production of microtubule bundles without normal function and to the stabilization 
of microtubules, which results in the inhibition of mitosis in cells.  Docetaxel’s 
binding to microtubules does not alter the number of protofilaments in the bound 
microtubules, a feature which differs from most spindle poisons currently in use. 
 
8.3.3 Pharmacokinetics and Drug Metabolism 
 
The AUC is dose proportional following doses of 70 mg/m2 to 115 mg/m2 with 
infusions times of 1 to 2 hours.  Docetaxel’s pharmacokinetic profile is consistent 
with a three-compartment pharmacokinetic model, with half-lives for the α, β, and 
γ phases of 4 min, 36 min, and 11.1 hr, respectively.  Mean total body clearance 
was 21 L/h/m2. 
 
It is metabolized by the CYP3A4 isoenzyme.  It is eliminated in both the urine and 
feces following oxidative metabolism of the tert-butyl ester group, but fecal 
excretion was the main elimination route. 
 
8.3.4 Supplier(s) 
 
Docetaxel will be given as per routine care from commercial supply. 
 

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8.3.5 Dosage Form and Preparation 
 
Docetaxel is supplied in a single use vial as a sterile, pyrogen-free, non-aqueous 
solution.  One-vial (injection concentrate) docetaxel requires NO prior dilution with 
a diluent and is ready to add to the infusion solution.  Use only a 21 gauge needle 
to withdraw from the vial because larger bore needles may result in stopper coring 
and rubber particulates.  Inject via a single injection into a 250 mL infusion bag or 
bottle of either 0.9% Sodium Chloride solution or 5% Dextrose solution to produce 
a final concentration of 0.3 mg/mL to 0.74 mg/mL.  Thoroughly mix the infusion 
by gentle manual rotation. 
 
8.3.6 Storage and Stability 
 
Store between 2°C and 25°C.  Retain in the original package to protect from light. 
 
8.3.7 Administration 
 
Docetaxel should be given as an intravenous infusion over the course of 60 minutes 
(± 10 minutes). 
 
8.3.8 Special Handling Instructions 
 
No special handling instructions. 
 
 
9.0 
CORRELATIVE STUDIES 
 
9.1 
Tumor Biopsy 
 
9.1.1 Collection of Specimens 
 
Tumor biopsies (10 cores by needle biopsies) will be performed up to 21 days prior 
to initiation of ADI-PEG 20 and Day -1 (± 1 day as long as performed prior to 
initiation of gemcitabine).  Tumor biopsies are mandatory for the first 20 patients 
amenable to biopsy enrolled at Washington University (completed as of 05/14/19) 
and for all patients enrolled to the SCLC cohort. 
 
9.1.2 Handling of Specimens 
 
Samples should be snap frozen in liquid nitrogen or an equivalent process such as 
placing in All Protect Tissue Reagent and placed on dry ice. 
 
9.1.3 Specimen Storage 
 
Samples will be stored in the Van Tine laboratory at -80°C. 
 

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9.2 
Archival Tissue 
 
Five to 10 slides 5 to 10 microns thick of archival tissue will be collected on all patients.  
Ship to: 
 
 
Dr. Brian Van Tine – Research Lab 
4515 McKinley, 3rd Floor 
4523 Clayton Avenue 
St. Louis, MO 63110 
 
9.3 
Mandatory Peripheral Blood 
 
9.3.1 Collection of Specimens 
 
Eight mL of blood will be drawn into 2 red top SST (Tiger) tubes at the following 
time points: 
• Day -7 (pre-treatment) 
• Day -1 
• Days 1 and 8 of each cycle 
 
9.3.2 Handling of Specimens 
 
Invert the tubes gently 10 times.  Let the tubes stand at room temperature to 
coagulate for 30-60 minutes.  Centrifuge within one hour of collection at 1600 G 
for 15 minutes.  Draw off serum from one red top SST tube and fill two 1.8ml. 
cryovials labeled “AAA” (amino acid analysis), ADI-PEG, subject ID, time point 
and date.  Draw off serum from the second red top SST tube and fill two 1.8ml 
cryovials labeled “immun” immunogenicity), ADI-PEG, subject ID, time point, 
and date.  Store tubes at -70°C or cooler until shipment. Please complete 
requisition and batch ship all frozen specimens per subject at the end of their 
treatment. Ship on dry ice to: 
 
Dr. Brian Van Tine – Research Lab 
4515 McKinley, 3rd Floor 
4523 Clayton Avenue 
St. Louis, MO 63110 
 
9.3.3 Specimen Storage 
 
Samples will be stored in the Van Tine laboratory at -80°C. 
 
 
 

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10.0 
STUDY CALENDAR 
 
Screening/baseline evaluations are to be conducted within 14 days prior to start of protocol 
therapy.  Scans and x-rays must be done no more than 28 days prior to the start of the protocol 
therapy.  There is a +/- 2 day window for all trial-related activities including treatment. 
 
 
Screening 
D -7 
D -1 
D1 of 
each 
cycle 
D8 of 
each 
cycle 
D15 of 
cycle  
Every 6 weeks       
+/- 1 week during 
treatment 
30-Day 
F/U 
5-Year 
F/U 
Informed consent 
X 
 
 
 
 
 
 
 
 
H&P, ECOG PS 
X 
X 
 
X 
 
 
 
 
 
CBC 
X 
 
 
X 
X 
X5 
 
 
 
CMP + uric acid 
X 
 
 
X 
 
 
 
 
 
β-hCG1 
X 
 
 
 
 
 
 
 
 
CT C/A/P and 
tumor assessments 
X 
 
 
 
 
 
X 
 
X4 
ECG 
X 
 
 
 
 
 
 
 
 
ADI-PEG 20 
 
X 
 
X 
X 
X 
 
 
 
Gemcitabine 
 
 
 
X 
X 
 
 
 
 
Docetaxel 
 
 
 
 
X 
 
 
 
 
Archival tissue 
X 
 
 
 
 
 
 
 
 
Tumor biopsy6 
 
X 
X 
 
 
 
 
 
 
Blood for amino 
acid analysis2 
 
X 
X 
X 
X 
 
 
 
 
Blood for 
immunogenicity2 
 
X 
X 
X 
X 
 
 
 
 
Adverse events 
 
Monitor AEs continuously and during follow-up for 30 days after the last dose 
of study treatment. 
 
Review of medical 
record3 
 
 
 
 
 
 
 
 
X 
1. 
Women of childbearing potential only. 
2. 
Obtained before drug treatment on days of drug treatment, and before tumor biopsy on days of tumor biopsy. 
3. 
For progression and survival information 
4. 
After discontinuation of treatment, scans will be performed every 6-12 weeks (for patients who discontinue for reasons other than 
progression).  There will be a +/- 7 day window for the first year and a  +/- 7 week window for Years 2-5.  Scans will continue until 
progression. Patients who discontinue due to progression will not have any further scans mandated by this protocol. 
5. 
Obtain CBC on day 15 of each cycle if patient is taking a blood thinner. 
6. 
Tumor biopsies are mandatory for the first 20 patients amenable to biopsy enrolled at Washington University (completed as of 
05/14/19) and for all patients enrolled to the SCLC cohort.  Day -7 biopsy may occur up to 21 days prior to initiation of ADI-PEG 
20.  Day -1 biopsy may occur on Day -2 or Day 1 as long as it is performed prior to initiation of gemcitabine on Day 1. 
 
 
 

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11.0 
DATA SUBMISSION SCHEDULE 
 
Case report forms with appropriate source documentation will be completed according to the 
schedule listed in this section. 
 
Case Report Form 
Submission Schedule 
Original Consent Form 
Prior to registration 
On-Study Form 
Cohort 2 On Study Form 
Archival Tissue Form 
Prior to starting treatment 
Treatment Form 
Every cycle 
Peripheral Blood Form 
Day -7, Day -1, Day 1 of each cycle, Day 8 of each cycle 
Tumor Biopsy Form 
Day -7, Day -1 
AE Form 
Continuous 
Treatment Summary Form 
Completion of treatment 
Follow Up Form 
Day 30 after end of treatment, 5 years after end of treatment 
RECIST Form 
Choi Response Form 
Baseline, every 6 weeks while on treatment, every 6-12 weeks 
after treatment ends for 5 years or until time of progression. 
Death Form 
Time of death 
MedWatch Form 
See Section 7.0 for reporting requirements 
 
Any queries generated by Washington University must be responded to within 28 days of receipt 
by the participating site.  The Washington University research team will conduct a regular review 
of data status at all secondary sites, with appropriate corrective action to be requested as needed. 
 
11.1 
Adverse Event Collection in the Case Report Forms 
 
All adverse events that occur beginning with start of treatment (minus exceptions defined 
in Section 7.0) must be captured in the Toxicity Form.  Baseline AEs should be captured 
on the Toxicity Form. 
 
Participant death due to disease progression should be reported on the Toxicity Form as 
grade 5 disease progression.  If death is due to an AE (e.g. cardiac disorders: cardiac arrest), 
report as a grade 5 event under that AE.  Participant death must also be recorded on the 
Death Form. 
 
 
12.0 
MEASUREMENT OF EFFECT 
 
For the purposes of this study, patients should be re-evaluated for response every 6 weeks.  In 
addition to a baseline scan, confirmatory scans should also be obtained 6 weeks following initial 
documentation of objective response. 
 
Response and progression will be evaluated in this study using the new international criteria 
proposed by the revised Response Evaluation Criteria in Solid Tumors (RECIST) guideline 
(version 1.1) [Eur J Ca 45:228-247, 2009].  Changes in the largest diameter (unidimensional 

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measurement) of the tumor lesions and the shortest diameter in the case of malignant lymph nodes 
are used in the RECIST criteria.  Please refer to Section 12.1. 
 
Choi response criteria will also be used to evaluate response in this study.  Please refer to Section 
12.2. 
 
12.1 
RECIST 1.1 
 
12.1.1 Disease Parameters 
 
Measurable disease:  Measurable lesions are defined as those that can be 
accurately measured in at least one dimension (longest diameter to be recorded) as 
>20 mm by chest x-ray, as >10 mm with CT scan, or >10 mm with calipers by 
clinical exam.  All tumor measurements must be recorded in millimeters (or 
decimal fractions of centimeters). 
 
Malignant lymph nodes:  To be considered pathologically enlarged and 
measurable, a lymph node must be >15 mm in short axis when assessed by CT scan 
(CT scan slice thickness recommended to be no greater than 5 mm).  At baseline 
and in follow-up, only the short axis will be measured and followed. 
 
Non-measurable disease:  All other lesions (or sites of disease), including small 
lesions (longest diameter <10 mm or pathological lymph nodes with ≥10 to <15 
mm short axis), are considered non-measurable disease.  Bone lesions, 
leptomeningeal disease, ascites, pleural/pericardial effusions, lymphangitis 
cutis/pulmonitis, inflammatory breast disease, and abdominal masses (not followed 
by CT or MRI), are considered as non-measurable. 
 
Note:  Cystic lesions that meet the criteria for radiographically defined simple cysts 
should not be considered as malignant lesions (neither measurable nor non-
measurable) since they are, by definition, simple cysts. 
 
‘Cystic lesions’ thought to represent cystic metastases can be considered as 
measurable lesions, if they meet the definition of measurability described above. 
However, if non-cystic lesions are present in the same patient, these are preferred 
for selection as target lesions. 
 
Target lesions:  All measurable lesions up to a maximum of 2 lesions per organ 
and 5 lesions in total, representative of all involved organs, should be identified as 
target lesions and recorded and measured at baseline.  Target lesions should be 
selected on the basis of their size (lesions with the longest diameter), be 
representative of all involved organs, but in addition should be those that lend 
themselves to reproducible repeated measurements.  It may be the case that, on 
occasion, the largest lesion does not lend itself to reproducible measurement in 
which circumstance the next largest lesion which can be measured reproducibly 
should be selected.  A sum of the diameters (longest for non-nodal lesions, short 

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axis for nodal lesions) for all target lesions will be calculated and reported as the 
baseline sum diameters.  If lymph nodes are to be included in the sum, then only 
the short axis is added into the sum.  The baseline sum diameters will be used as 
reference to further characterize any objective tumor regression in the measurable 
dimension of the disease. 
 
Non-target lesions:  All other lesions (or sites of disease) including any measurable 
lesions over and above the 5 target lesions should be identified as non-target lesions 
and should also be recorded at baseline.  Measurements of these lesions are not 
required, but the presence, absence, or in rare cases unequivocal progression of 
each should be noted throughout follow-up.  
 
12.1.2 Methods for Evaluation of Measurable Disease 
 
All measurements should be taken and recorded in metric notation using a ruler or 
calipers.  All baseline evaluations should be performed as closely as possible to the 
beginning of treatment and never more than 4 weeks before the beginning of the 
treatment. 
 
The same method of assessment and the same technique should be used to 
characterize each identified and reported lesion at baseline and during follow-up. 
Imaging-based evaluation is preferred to evaluation by clinical examination unless 
the lesion(s) being followed cannot be imaged but are assessable by clinical exam. 
 
Clinical lesions:  Clinical lesions will only be considered measurable when they 
are superficial (e.g., skin nodules and palpable lymph nodes) and ≥10 mm diameter 
as assessed using calipers (e.g., skin nodules).  In the case of skin lesions, 
documentation by color photography, including a ruler to estimate the size of the 
lesion, is recommended.  
 
Chest x-ray:  Lesions on chest x-ray are acceptable as measurable lesions when 
they are clearly defined and surrounded by aerated lung.  However, CT is 
preferable.  
 
Conventional CT and MRI:  This guideline has defined measurability of lesions 
on CT scan based on the assumption that CT slice thickness is 5 mm or less.  If CT 
scans have slice thickness greater than 5 mm, the minimum size for a measurable 
lesion should be twice the slice thickness.  MRI is also acceptable in certain 
situations (e.g. for body scans).   
 
Use of MRI remains a complex issue.  MRI has excellent contrast, spatial, and 
temporal resolution; however, there are many image acquisition variables involved 
in MRI, which greatly impact image quality, lesion conspicuity, and measurement.  
Furthermore, the availability of MRI is variable globally.  As with CT, if an MRI 
is performed, the technical specifications of the scanning sequences used should be 
optimized for the evaluation of the type and site of disease.  Furthermore, as with 

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Page 37 of 55 
CT, the modality used at follow-up should be the same as was used at baseline and 
the lesions should be measured/assessed on the same pulse sequence.  It is beyond 
the scope of the RECIST guidelines to prescribe specific MRI pulse sequence 
parameters for all scanners, body parts, and diseases.  Ideally, the same type of 
scanner should be used and the image acquisition protocol should be followed as 
closely as possible to prior scans.  Body scans should be performed with breath-
hold scanning techniques, if possible. 
 
PET-CT:  At present, the low dose or attenuation correction CT portion of a 
combined PET-CT is not always of optimal diagnostic CT quality for use with 
RECIST measurements.  However, if the site can document that the CT performed 
as part of a PET-CT is of identical diagnostic quality to a diagnostic CT (with IV 
and oral contrast), then the CT portion of the PET-CT can be used for RECIST 
measurements and can be used interchangeably with conventional CT in accurately 
measuring cancer lesions over time.  Note, however, that the PET portion of the CT 
introduces additional data which may bias an investigator if it is not routinely or 
serially performed.   
 
Ultrasound:  Ultrasound is not useful in assessment of lesion size and should not 
be used as a method of measurement.  Ultrasound examinations cannot be 
reproduced in their entirety for independent review at a later date and, because they 
are operator dependent, it cannot be guaranteed that the same technique and 
measurements will be taken from one assessment to the next.  If new lesions are 
identified by ultrasound in the course of the study, confirmation by CT or MRI is 
advised.  If there is concern about radiation exposure at CT, MRI may be used 
instead of CT in selected instances. 
 
Endoscopy, Laparoscopy:  The utilization of these techniques for objective tumor 
evaluation is not advised.  However, such techniques may be useful to confirm 
complete pathological response when biopsies are obtained or to determine relapse 
in trials where recurrence following complete response (CR) or surgical resection 
is an endpoint. 
 
Tumor markers:  Tumor markers alone cannot be used to assess response.  If 
markers are initially above the upper normal limit, they must normalize for a patient 
to be considered in complete clinical response.  Specific guidelines for both CA-
125 response (in recurrent ovarian cancer) and PSA response (in recurrent prostate 
cancer) have been published [JNCI 96:487-488, 2004; J Clin Oncol 17, 3461-3467, 
1999; J Clin Oncol 26:1148-1159, 2008].  In addition, the Gynecologic Cancer 
Intergroup has developed CA-125 progression criteria which are to be integrated 
with objective tumor assessment for use in first-line trials in ovarian cancer [JNCI 
92:1534-1535, 2000]. 
 
Cytology, Histology:  These techniques can be used to differentiate between partial 
responses (PR) and complete responses (CR) in rare cases (e.g., residual lesions in 

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tumor types, such as germ cell tumors, where known residual benign tumors can 
remain). 
 
The cytological confirmation of the neoplastic origin of any effusion that appears 
or worsens during treatment when the measurable tumor has met criteria for 
response or stable disease is mandatory to differentiate between response or stable 
disease (an effusion may be a side effect of the treatment) and progressive disease. 
 
FDG-PET:  While FDG-PET response assessments need additional study, it is 
sometimes reasonable to incorporate the use of FDG-PET scanning to complement 
CT scanning in assessment of progression (particularly possible 'new' disease).  
New lesions on the basis of FDG-PET imaging can be identified according to the 
following algorithm:  
 
• Negative FDG-PET at baseline, with a positive FDG-PET at follow-up is a 
sign of PD based on a new lesion. 
• No FDG-PET at baseline and a positive FDG-PET at follow-up:  If the 
positive FDG-PET at follow-up corresponds to a new site of disease 
confirmed by CT, this is PD.  If the positive FDG-PET at follow-up is not 
confirmed as a new site of disease on CT, additional follow-up CT scans 
are needed to determine if there is truly progression occurring at that site (if 
so, the date of PD will be the date of the initial abnormal FDG-PET scan).  
If the positive FDG-PET at follow-up corresponds to a pre-existing site of 
disease on CT that is not progressing on the basis of the anatomic images, 
this is not PD. 
• FDG-PET may be used to upgrade a response to a CR in a manner similar 
to a biopsy in cases where a residual radiographic abnormality is thought to 
represent fibrosis or scarring.  The use of FDG-PET in this circumstance 
should be prospectively described in the protocol and supported by disease-
specific medical literature for the indication.  However, it must be 
acknowledged that both approaches may lead to false positive CR due to 
limitations of FDG-PET and biopsy resolution/sensitivity. 
  
Note:  A ‘positive’ FDG-PET scan lesion means one which is FDG avid with an 
uptake greater than twice that of the surrounding tissue on the attenuation 
corrected image. 
 
12.1.3 Response Criteria 
 
12.1.3.1 
Evaluation of Target Lesions 
 
Complete Response (CR):  Disappearance of all target lesions.  Any 
pathological lymph nodes (whether target or non-target) must have 
reduction in short axis to <10 mm. 
 

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Partial Response (PR):  At least a 30% decrease in the sum of the 
diameters of target lesions, taking as reference the baseline sum diameters. 
 
Progressive Disease (PD):  At least a 20% increase in the sum of the 
diameters of target lesions, taking as reference the smallest sum on study 
(this includes the baseline sum if that is the smallest on study).  In addition 
to the relative increase of 20%, the sum must also demonstrate an absolute 
increase of at least 5 mm.  (Note:  the appearance of one or more new lesions 
is also considered progressions). 
 
Stable Disease (SD):  Neither sufficient shrinkage to qualify for PR nor 
sufficient increase to qualify for PD, taking as reference the smallest sum 
diameters while on study. 
 
12.1.3.2 
Evaluation of Non-Target Lesions 
 
Complete Response (CR):  Disappearance of all non-target lesions and 
normalization of tumor marker level.  All lymph nodes must be non-
pathological in size (<10 mm short axis). 
 
Note:  If tumor markers are initially above the upper normal limit, they must 
normalize for a patient to be considered in complete clinical response. 
 
Non-CR/Non-PD:  Persistence of one or more non-target lesion(s) and/or 
maintenance of tumor marker level above the normal limits. 
 
Progressive Disease (PD):  Appearance of one or more new lesions and/or 
unequivocal progression of existing non-target lesions.  Unequivocal 
progression should not normally trump target lesion status.  It must be 
representative of overall disease status change, not a single lesion increase.     
 
Although a clear progression of “non-target” lesions only is exceptional, the 
opinion of the treating physician should prevail in such circumstances, and 
the progression status should be confirmed at a later time by the review 
panel (or Principal Investigator). 
 
12.1.3.3 
Evaluation of Best Overall Response 
 
The best overall response is the best response recorded from the start of the 
treatment until disease progression/recurrence (taking as reference for 
progressive disease the smallest measurements recorded since the treatment 
started).  The patient's best response assignment will depend on the 
achievement of both measurement and confirmation criteria. 
   

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For Patients with Measurable Disease (i.e., Target Disease) 
Target 
Lesions 
Non-Target 
Lesions 
New 
Lesions 
Overall 
Response 
Best Overall Response 
when Confirmation is 
Required* 
CR 
CR 
No 
CR 
>4 wks. Confirmation** 
CR 
Non-CR/Non-
PD 
No 
PR 
>4 wks. Confirmation** 
CR 
Not evaluated 
No 
PR 
PR 
Non-CR/Non-
PD/not 
evaluated 
No 
PR 
SD 
Non-CR/Non-
PD/not 
evaluated 
No 
SD 
Documented at least once 
>4 wks. from baseline** 
PD 
Any 
Yes or 
No 
PD 
no prior SD, PR or CR 
Any 
PD*** 
Yes or 
No 
PD 
Any 
Any 
Yes 
PD 
* 
See RECIST 1.1 manuscript for further details on what is evidence of a new lesion. 
** 
Only for non-randomized trials with response as primary endpoint. 
*** 
In exceptional circumstances, unequivocal progression in non-target lesions may be 
accepted as disease progression. 
Note: 
Patients with a global deterioration of health status requiring discontinuation of treatment 
without objective evidence of disease progression at that time should be reported as “symptomatic 
deterioration.”  Every effort should be made to document the objective progression even after 
discontinuation of treatment. 
                           
    For Patients with Non-Measurable Disease (i.e., Non-Target Disease) 
Non-Target Lesions 
New Lesions 
Overall Response 
CR 
No 
CR 
Non-CR/non-PD 
No 
Non-CR/non-PD* 
Not all evaluated 
No 
not evaluated 
Unequivocal PD 
Yes or No 
PD 
Any 
Yes 
PD 
*  ‘Non-CR/non-PD’ is preferred over ‘stable disease’ for non-target disease since SD is 
increasingly used as an endpoint for assessment of efficacy in some trials so to assign this 
category when no lesions can be measured is not advised 
 
 
12.1.3.4 
Duration of Response 
 
Duration of overall response:  The duration of overall response is 
measured from the time measurement criteria are met for CR or PR 
(whichever is first recorded) until the first date that recurrent or progressive 
disease is objectively documented (taking as reference for progressive 
disease the smallest measurements recorded since the treatment started). 
 
The duration of overall CR is measured from the time measurement criteria 

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Page 41 of 55 
are first met for CR until the first date that progressive disease is objectively 
documented.  
 
Duration of stable disease:  Stable disease is measured from the start of 
the treatment until the criteria for progression are met, taking as reference 
the smallest measurements recorded since the treatment started, including 
the baseline measurements.  
 
12.2 
Choi Response Criteria 
 
Using disease parameters and response assessment methods as outlined in Section 12.1, 
Choi response criteria are as follows16: 
 
Complete Response (CR): Disappearance of all target lesions 
 
Partial Response (PR): Decrease in tumor size ≥ 10% on CT or decrease in tumor density 
≥ 15% on CT, no new lesions 
 
Stable Disease (SD): Does not meet the criteria for CR, PR, or PD 
 
Progressive Disease (PD): Increase in tumor size ≥ 10% (sum of longest diameters of 
lesions) and does not meet PR criteria by tumor density, or new intratumoral nodules or 
disease or an increase in the size of the existing intratumoral nodules 
 
 
13.0 
DATA AND SAFETY MONITORING 
 
In compliance with the Washington University Institutional Data and Safety Monitoring Plan, an 
independent Data and Safety Monitoring Board (DSMB) will be specifically convened for this 
trial to review toxicity data.  A DSMB will consist of no fewer than 3 members including 2 clinical 
investigators and a biostatistician.  DSMB members must be employed by Washington University, 
Barnes-Jewish  Hospital, or St. Louis Children’s Hospital.  Like investigators, DSMB members 
are subject to the Washington University School of Medicine policies regarding standards of 
conduct. Individuals invited to serve on the DSMB will disclose any potential conflicts of interest 
to the trial principal investigator and/or appropriate university officials, in accordance with 
institution policies. Potential conflicts that develop during a trial or a member’s tenure on a DSMB 
must also be disclosed.  
 
Until such a time as the first secondary site enrolls its first patient, a semi-annual DSM report to 
be prepared by the study team will be submitted to the Quality Assurance and Safety Monitoring 
Committee (QASMC) semi-annually beginning 6 months after study activation at Washington 
University (if at least one patient has been enrolled) or one year after study activation (if no patients 
have been enrolled at the 6-month mark). 
 
The DSM report for the DSMB will be prepared by the study team with assistance from the study 
statistician, will be reviewed by the DSMB, and will be submitted to the QASM Committee.  The 

Protocol Version: 11/04/2025, Amendment 10 
 
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DSMB must meet beginning six months after enrollment of the first patient at a secondary site, no 
more than one month prior to the due date of the DSM report to QASMC.  This report will include: 
 
• HRPO protocol number, protocol title, Principal Investigator name, data coordinator name, 
regulatory coordinator name, and statistician 
• Date of initial HRPO approval, date of most recent consent HRPO approval/revision, date 
of HRPO expiration, date of most recent QA audit, study status, and phase of study 
• History of study including summary of substantive amendments; summary of accrual 
suspensions including start/stop dates and reason; and summary of protocol exceptions, 
error, or breach of confidentiality including start/stop dates and reason 
• Study-wide target accrual and study-wide actual accrual including numbers from 
participating sites 
• Protocol activation date at each participating site 
• Average rate of accrual observed in year 1, year 2, and subsequent years at each 
participating site 
• Expected accrual end date and accrual by site 
• Objectives of protocol with supporting data and list the number of participants who have 
met each objective 
• Measures of efficacy 
• Early stopping rules with supporting data and list the number of participants who have met 
the early stopping rules 
• Summary of toxicities at all participating sites 
• Abstract submissions/publications 
• Summary of any recent literature that may affect the safety or ethics of the study 
 
Further DSMB responsibilities are described in the DSMB charter. 
 
The study principal investigator and coordinator will monitor for serious toxicities on an ongoing 
basis. Once the principal investigator or coordinator becomes aware of an adverse event, the AE 
will be reported to the HRPO and QASMC according to institutional guidelines (please refer to 
Section 7.0). 
 
Refer to the Washington University Quality Assurance and Data Safety Monitoring Committee 
Policies and Procedures for full details on the responsibilities of the DSMB.  This is located on the 
QASMC website at https://siteman.wustl.edu/research/clinical-research-resources/protocol-
office-prmcqasmc/. 
 
 
14.0 
AUDITING 
 
As coordinating center of this trial, Washington University (via the Quality Assurance and Safety 
Monitoring Committee (QASMC) will monitor each participating site to ensure that all protocol 
requirements are being met; that applicable federal regulations are being followed; and that best 
practices for patient safety and data collection are being followed per protocol.  Participating sites 
will be asked to send copies of all audit materials, including source documentation.  The audit 
notification will be sent to the Washington University Research Patient Coordinator, who will 

Protocol Version: 11/04/2025, Amendment 10 
 
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obtain the audit materials from the participating institution. 
 
Notification of an upcoming audit will be sent to the research team one month ahead of the audit. 
Once accrual numbers are confirmed, and approximately 30 days prior to the audit, a list of the 
cases selected for review (up to 10 for each site) will be sent to the research team. However, if 
during the audit the need arises to review cases not initially selected, the research team will be 
asked to provide the additional charts within two working days. 
 
Items to be evaluated include: 
• Subject screening and enrollment 
• Reporting of adverse events 
• Maintenance of HIPAA compliance 
• Completeness of regulatory documentation 
• Completeness of participant documentation 
• Acquisition of informed consent 
• IRB documentation 
• Issues of protocol adherence 
 
Additional details regarding the auditing policies and procedures can be found at 
https://siteman.wustl.edu/research/clinical-research-resources/protocol-office-prmcqasmc/. 
 
 
15.0 
STATISTICAL CONSIDERATIONS 
 
15.1 
Study Design 
 
For Cohort 1, a single arm phase II trial will be conducted to evaluate effiacy of the 
combination regimen, where eligible grade 2 or 3 soft tissue sarcoma patients with 
unresectable or metastatic tumors will be treated by ADI-PEG20 in combination with 
gemcitabine and docetaxel.  
 
For Cohort 2, 15 patients (10 with osteosarcoma or Ewing’s sarcoma (ideally 5 of each) 
and 5 with SCLC) will be enrolled to evaluate safety and tolerability of the combination 
regimen. 
 
15.2 
Endpoints 
 
The primary endpoint is progression-free survival (PFS) defined as time on study to time 
patients progressed on the drug combination or death or latest follow-up if 
progression/death is not observed yet. 
 
The secondary endpoints include clinical benefit rate (CBR) defined as the proportion of 
patients with complete response (CR), partial response (PR), or stable disease (SD) lasting 
24 weeks or longer by the RECIST standard, overall survival (OS) defined as time on study 
to time of death due to any reasons or latest follow up (whichever is earlier), and cancer-
mortality rate. 

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15.3 
Sample Size Justification 
 
The median PFS of metastatic sarcoma patients receiving the standard gemcitabine and 
docetaxel treatment was estimated to be 6.2 months in a randomized phase II study (Maki 
et. al., 2007, JCO).  With the addition of ADI-PEG 20, we target to improve the median 
PFS to 9 months, a 2.8 months or 12 weeks, or 45.2% improvement on patients treated on 
gemcitabine + docetaxel + ADI-PEG 20 against null hypothesis median PFS of 6.2 months.  
Assuming uniform accrual within a 12-month enrollment period and a minimum of 6-
month follow up, the study will enroll 75 patients to Cohort 1 (with a per month accrual 
rate ~6.2), expecting 44 events (progression or death), to achieve 80% power to detect the 
improvement in PFS at a 5% alpha level. 
 
Fifteen patients will be enrolled to Cohort 2: 10 with either osteosarcoma or Ewing’s 
sarcoma (ideally 5 of each) and 5 with non-small cell lung cancer. 
 
15.4 
Accrual 
 
The study accrual will continue until 44 events (progression or death) are observed, 
expected in 75 patients in Cohort 1. By the accrual rate of 6 to 8 patients per month, the 
study will take 10 to 13 months to enroll a total of 75 patients in Cohort 1. With a targeted 
minimum follow up of 6 months after enrollment period, the study will take 16 to 19 
months to complete. Patient will be followed for 5 years for overall  suvivial. Only patients 
who receive at least one dose of gemcitabine will be counted towards accrual and analysis. 
 
15.5 
Analysis Plan 
 
Survival endpoints for Cohort 1 will be analyzed by the Kaplan-Meier product limit method 
and illustrated by the KM curves with 95% CI, overal and by groups of interest while 
survival probability will be estimated. Log-rank test will be applied to test difference in 
survival curves between groups of interest. The CBR and cancer-mortality rate will be 
estimated with 95% confidence interval (CI). Toxicity profiles will be decribed by 
descriptive statistics overall and by patient characterisics (age, gender etc) and response 
categories, by the two cohorts separately and in combination. For explorative analyses, 
arginine plasma level, phospho-PKM2 and metabolomics marker measured in baseline and 
a after-treatment sample will be compared by paired t-test or Wilcoxon signed rank test as 
proper while the measurements at each time point and the change after treatment relative 
to baseline will be comapred between patients showing CBR versus those not by two 
sample t-test or Wilcoxon rank sum test as approporate. 
 
15.6 
Study Stopping Rules 
 
Non-hematological toxicity and plans for data and safety monitoring: 
Early stopping of the Phase II phase of the trial will be based on unacceptable non-
hematological toxicity. Approximately less than 25% of patients are expected to experience 
grade 3 and up non-hematological toxicity and a non-hematological toxicity rate of 40% 

Protocol Version: 11/04/2025, Amendment 10 
 
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or more would definitely be unacceptable. Based on the sequential probability ratio test 
(SPRT) with 80% power and a 0.05 significance level, the study will be halted if 6 of the 
first 6, or 7 of the first 9, or 8 of the first 12, or 9 of the first 16, or 10 of the first 19, or 11 
of the first 22, or 12 of the first 25, or 13/28, or 14/31, or 15/34, 16/37, 17/40, 18/43, 19/47, 
20/50, 21/53, 22/56, 23/59, 24/62, 25/65, 26/68, 27/71, if the 28th non-hematological 
toxicity is observed before the last patient has completed the trial.  
 
Accrual will be paused and the event will be reviewed by the Data and Safety Monitoring 
Board if a grade 5 non-hematological toxicity is observed that is deemed probably or 
definitely related to ADI-PEG 20, gemcitabine, docetaxel, or combination. Note that sepsis 
is considered a hematological toxicity. 
 
For Cohort 2 with 15 patients in total, we similarly monitor its stopping due to over 
toxicities. Based on the sequential probability ratio test (SPRT) with 80% power and a 0.05 
significance level, the enrollment to Cohort 2 will be halted if 6 of the first 6, or 7 of the 
first 9, or 8 of the first 12, or 9 before the last patient enrolled are observed with grade 4 or 
higher non-hematological toxicities not considered unrelated to study treatment and not 
considered related to disease progression. 
 
Updated stopping rule as of Amendment 7: 
Respecting the gemcitabine dosage reductions (from 900 mg/m2 to 750 mg/m2 to 600 
mg/m2), the stopping rule will be evaluated by dose cohort and sequential toxicity 
monitoring will be ensured for future trial conduct. Based on the redefined treatment-
emergent serious adverse events (TESAEs, defined as any type of grade 4 or higher non-
hematological treatment-emergent toxicity that is not considered unrelated to study 
treatment and is not considered related to disease progression) and evaluating TESAEs 
within each of the 3 starting dose cohorts separately, the safety boundary was crossed with 
gemcitabine at 900 mg/m2 but stopped immediately after the boundary was crossed and the 
dose was reduced to 750 mg/m2.  The safety boundary was not crossed and should be safe 
by definition with gemcitabine at 750 mg/m2 or 600 mg/m2. Thus, the dose level of 600 
mg/m2 should be a safe starting dose level for future patients and the same stopping rule 
based on the sequential probability ratio test (SPRT) with 80% power and a 0.05 
significance level will be restarted for evaluation on the trial patients enrolled to receive 
gemcitabine at 600 mg/m2: the study will be halted if 6 of the first 6, or 7 of the first 9, or 
8 of the first 12, or 9 of the first 16, or 10 of the first 19, or 11 of the first 22, or 12 of the 
first 25, or 13/28, or 14/31, or 15/34, 16/37, 17/40, 18/43, 19/47, 20/50, 21/53, 22/56, 23/59, 
24/62, 25/65, 26/68, 27/71, or if the 28th non-hematological toxicity is observed before the 
last patient has completed the trial.  
 
15.7 
Population for Analysis 
 
Safety Population: All subjects who receive at least one dose of ADI-PEG20 will be used 
for safety analyses. Cohort 1 and Cohort 2 will be analyzed separately and together for 
toxicities. 
 
Efficacy Evaluable Population: All Cohort 1 subjects who complete the first 2 cycles of 

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treatments and receive at least 6 doses of ADI-PEG20 and have at least 1 post-baseline 
RECIST 1.1 assessment, or who discontinue treatment/study due to disease progression or 
death after having received at least 1 dose of the study drug ADI-PEG20. 
 
16.0 
MULTICENTER REGULATORY REQUIREMENTS 
 
Washington University requires that each participating site sends its informed consent document 
to be reviewed and approved by the Washington University Regulatory Coordinator (or designee) 
prior to IRB/IEC submission.    
 
Site activation is defined as when the secondary site has received official written documentation 
from the coordinating center that the site has been approved to begin enrollment.  At a minimum, 
each participating institution must have the following documents on file at Washington University 
prior to study activation: 
• Documentation of IRB approval of the study in the form of a letter or other official 
document from the participating institution’s IRB.  This documentation must show which 
version of the protocol was approved by the IRB. 
• Documentation of IRB approval of an informed consent form. The consent must include a 
statement that data will be shared with Washington University, including the Quality 
Assurance and Safety Monitoring Committee (QASMC), the DSMC (if applicable), and 
the Washington University study team. 
• Documentation of FWA, signed FDA Form 1572 (if applicable), and the CVs of all 
participating investigators. 
• Protocol signature page signed and dated by the investigator at each participating site. 
 
The coordinating center Principal Investigator (or designee) is responsible for disseminating to the 
participating sites all study updates, amendments, reportable adverse events, etc.  Protocol/consent 
modifications and IB updates will be forwarded electronically to the secondary sites within 4 
weeks of obtaining Washington University IRB approval.  Activated secondary sites are expected 
to submit protocol/consent/IB modifications to their local IRBs within 4 weeks of receipt unless 
otherwise noted.   
 
Upon the secondary sites obtaining local IRB approval, documentation of such shall be sent to the 
Washington University study team within 2 weeks of receipt of approval. 
 
Documentation of participating sites’ IRB approval of annual continuing reviews, protocol 
amendments or revisions, all SAE reports, and all protocol violations/deviations/exceptions must 
be kept on file at Washington University. 
 
The investigator or a designee from each institution must participate in a regular conference call 
to update and inform regarding the progress of the trial. 
 
      
 
 
 

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17.0 
REFERENCES 
 
1 
Siegel, R. L., Miller, K. D. & Jemal, A. Cancer statistics, 2016. CA Cancer J Clin 66, 7-
30, doi:10.3322/caac.21332 (2016). 
2 
Van Glabbeke, M., Verweij, J., Judson, I. & Nielsen, O. S. Progression-free rate as the 
principal end-point for phase II trials in soft-tissue sarcomas. European journal of cancer 
38, 543-549 (2002). 
3 
Santoro, A. et al. Doxorubicin versus CYVADIC versus doxorubicin plus ifosfamide in 
first-line treatment of advanced soft tissue sarcomas: a randomized study of the European 
Organization for Research and Treatment of Cancer Soft Tissue and Bone Sarcoma 
Group. Journal of Clinical Oncology 13, 1537-1545, doi:10.1200/jco.1995.13.7.1537 
(1995). 
4 
Tap D. W., L. J. L. R., Chmielowski B., et al. A randomized phase Ib/II study evaluating 
the safety and efficacy of olaratumab (IMC-3G3), a human anti-platelet-derived growth 
factor α (PDGFRα) monoclonal antibody, with or without doxorubicin (Dox), in 
advanced soft tissue sarcoma (STS). J Clin Oncol 33, 2015 (suppl; abstr 10501) (2015). 
5 
von Mehren, M. et al. Soft Tissue Sarcoma, Version 2.2016, NCCN Clinical Practice 
Guidelines in Oncology. J Natl Compr Canc Netw 14, 758-786 (2016). 
6 
Phillips, M. M., Sheaff, M. T. & Szlosarek, P. W. Targeting Arginine-Dependent Cancers 
with Arginine-Degrading Enzymes: Opportunities and Challenges. Cancer research and 
treatment : official journal of Korean Cancer Association 45, 251-262, 
doi:10.4143/crt.2013.45.4.251 (2013). 
7 
Patil, M. D., Shinde, K. D., Patel, G., Chisti, Y. & Banerjee, U. C. Use of response 
surface method for maximizing the production of arginine deiminase by Pseudomonas 
putida. Biotechnology Reports 10, 29-37, doi:10.1016/j.btre.2016.03.002 (2016). 
8 
Pasut, G., Sergi, M. & Veronese, F. M. Anti-cancer PEG-enzymes: 30 years old, but still 
a current approach. Advanced Drug Delivery Reviews 60, 69-78, 
doi:https://doi.org/10.1016/j.addr.2007.04.018 (2008). 
9 
Beddowes, E. et al. Phase 1 Dose-Escalation Study of Pegylated Arginine Deiminase, 
Cisplatin, and Pemetrexed in Patients With Argininosuccinate Synthetase 1-Deficient 
Thoracic Cancers. Journal of clinical oncology : official journal of the American Society 
of Clinical Oncology 35, 1778-1785, doi:10.1200/JCO.2016.71.3230 (2017). 
10 
Tomlinson, B. K. et al. Phase I Trial of Arginine Deprivation Therapy with ADI-PEG 20 
Plus Docetaxel in Patients with Advanced Malignant Solid Tumors. Clinical cancer 
research : an official journal of the American Association for Cancer Research 21, 2480-
2486, doi:10.1158/1078-0432.CCR-14-2610 (2015). 
11 
Lowery, M. A. et al. A phase 1/1B trial of ADI-PEG 20 plus nab-paclitaxel and 
gemcitabine in patients with advanced pancreatic adenocarcinoma. Cancer 123, 4556-
4565, doi:10.1002/cncr.30897 (2017). 
12 
Maki, R. G. Gemcitabine and docetaxel in metastatic sarcoma: past, present, and future. 
Oncologist 12, 999-1006, doi:12/8/999 [pii] 
10.1634/theoncologist.12-8-999 (2007). 
13 
Delage, B. et al. Arginine deprivation and argininosuccinate synthetase expression in the 
treatment of cancer. Int J Cancer 126, 2762-2772, doi:10.1002/ijc.25202 (2010). 

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Page 48 of 55 
14 
Ju, J. S. et al. Valosin-containing protein (VCP) is required for autophagy and is 
disrupted in VCP disease. The Journal of cell biology 187, 875-888, 
doi:10.1083/jcb.200908115 (2009). 
15 
Chaneton, B. et al. Serine is a natural ligand and allosteric activator of pyruvate kinase 
M2. Nature 491, 458-462, doi:nature11540 [pii] 
10.1038/nature11540. 
16 
Tirkes, T. et al. Response Criteria in Oncologic Imaging: Review of Traditional and New 
Criteria. RadioGraphics 33, 1323-1341, doi:10.1148/rg.335125214 (2013). 
17. 
Prudner BC, Rathore R, Robinson AM, et al.  Arginine Starvation and Docetaxel Induce 
c-Myc-Driven hENT1 Surface Expression to Overcome Gemcitabine Resistance in ASS1-
Negative Tumors. Clin Cancer Res. 2019 Aug 15;25(16):5122-5134. doi: 10.1158/1078-
0432.CCR-19-0206. 
 
We would like to thank the Alvin J. Siteman Cancer Center at Washington University School of 
Medicine and Barnes-Jewish Hospital in St. Louis, Missouri, for the use of the Clinical Trials Core 
which provided protocol development and study coordination services. The Siteman Cancer 
Center is supported in part by an NCI Cancer Center Support Grant #P30 CA91842. 
 
 
 
 
 
 
 

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APPENDIX A: ECOG Performance Status Scale 
 
 
Grade 
 
 
Description 
0 
Normal activity.  Fully active, able to carry on all pre-disease 
performance without restriction. 
1 
Symptoms, but ambulatory.  Restricted in physically strenuous 
activity, but ambulatory and able to carry out work of a light or 
sedentary nature (e.g., light housework, office work). 
2 
In bed <50% of the time.  Ambulatory and capable of all self-care, but 
unable to carry out any work activities.  Up and about more than 50% 
of waking hours. 
3 
 
In bed >50% of the time.  Capable of only limited self-care, confined 
to bed or chair more than 50% of waking hours. 
 
4 
100% bedridden.  Completely disabled.  Cannot carry on any self-care.  
Totally confined to bed or chair. 
5 
 
Dead. 
 
 
 
 
 

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APPENDIX B: Definitions for Adverse Event Reporting 
 
A. Adverse Events (AEs) 
 
As defined in 21 CFR 312.32: 
 
Definition: any untoward medical occurrence associated with the use of a drug in humans, 
whether or not considered drug-related. 
 
Grading: the descriptions and grading scales found in the revised NCI Common Terminology 
Criteria for Adverse Events (CTCAE) version 5.0 will be utilized for all toxicity reporting.  A 
copy of the CTCAE version 5.0 can be downloaded from the CTEP website. 
 
Attribution (relatedness), Expectedness, and Seriousness: the definitions for the terms 
listed that should be used are those provided by the Department of Health and Human Services’ 
Office for Human Research Protections (OHRP).  A copy of this guidance can be found on 
OHRP’s website: 
http://www.hhs.gov/ohrp/policy/advevntguid.html 
 
B. Suspected Adverse Reaction (SAR) 
 
As defined in 21 CFR 312.32: 
 
Definition: any adverse event for which there is a reasonable possibility that the drug caused 
the adverse event.  “Reasonable possibility” means there is evidence to suggest a causal 
relationship between the drug and the adverse event.  “Suspected adverse reaction” implies a 
lesser degree of certainty about causality than adverse reaction, which means any adverse event 
caused by a drug. 
 
C. Life-Threatening Adverse Event / Life Threatening Suspected Adverse Reaction  
 
As defined in 21 CFR 312.32: 
 
Definition: any adverse drug event or suspected adverse reaction is considered “life-
threatening” if, in the view of the investigator, its occurrence places the patient at immediate 
risk of death. It does not include an adverse event or suspected adverse reaction that, had it 
occurred in a more severe form, might have caused death. 
 
D.  Serious Adverse Event (SAE) or Serious Suspected Adverse Reaction 
 
As defined in 21 CFR 312.32: 
 
Definition:  an adverse event or suspected adverse reaction is considered “serious” if, in the 
view of the investigator, it results in any of the following outcomes: 
o Death 
o A life-threatening adverse event 

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o Inpatient hospitalization or prolongation of existing hospitalization 
o A persistent or significant incapacity or substantial disruption of the ability to conduct 
normal life functions 
o A congenital anomaly/birth defect 
o Any other important medical event that does not fit the criteria above but, based upon 
appropriate medical judgment, may jeopardize the subject and may require medical or 
surgical intervention to prevent one of the outcomes listed above 
 
E. Protocol Exceptions 
 
Definition: A planned change in the conduct of the research for one participant. 
 
F. Deviation 
 
Definition: Any alteration or modification to the IRB-approved research without prospective 
IRB approval.  The term “research” encompasses all IRB-approved materials and documents 
including the detailed protocol, IRB application, consent form, recruitment materials, 
questionnaires/data collection forms, and any other information relating to the research study. 
 
A minor or administrative deviation is one that does not have the potential to negatively impact 
the rights, safety, or welfare of participants or others or the scientific validity of the study. 
 
A major deviation is one that does have the potential to negatively impact the rights, safety, or 
welfare of participants or others or the scientific validity of the study. 
 

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APPENDIX C: Reporting Timelines 
 
Expedited Reporting Timelines 
Event 
HRPO 
QASMC 
FDA 
Polaris 
Serious AND 
unexpected suspected 
adverse reaction 
 
 
Report no later than 15 calendar 
days after it is determined that the 
information qualifies for 
reporting 
Report to Polaris within 24 hours of 
Sponsor-Investigator notification. 
Unexpected fatal or 
life-threatening 
suspected adverse 
reaction 
 
 
Report no later than 7 calendar 
days after initial receipt of the 
information 
 
Unanticipated 
problem involving risk 
to participants or 
others 
Report within 10 working days.  If the 
event results in the death of a 
participant enrolled at WU/BJH/SLCH, 
report within 1 working day. 
Report via email 
after IRB 
acknowledgment 
 
 
Pregnancy 
 
 
 
Notify Polaris as they occur. 
Overdose 
 
 
 
Notify Polaris as they occur. 
Major deviation 
Report within 10 working days.  If the 
event results in the death of a 
participant enrolled at WU/BJH/SLCH, 
report within 1 working day. 
 
 
 
A series of minor 
deviations that are 
being reported as a 
continuing 
noncompliance 
Report within 10 working days.   
 
 
 
Protocol exception 
Approval must be obtained prior to 
implementing the change 
 
 
 
Clinically important 
increase in the rate of 
a serious suspected 
adverse reaction of 
that list in the protocol 
or IB 
 
 
Report no later than 15 calendar 
days after it is determined that the 
information qualifies for 
reporting 
 

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Expedited Reporting Timelines 
Event 
HRPO 
QASMC 
FDA 
Polaris 
Complaints 
If the complaint reveals an 
unanticipated problem involving risks 
to participants or others OR 
noncompliance, report within 10 
working days.  If the event results in the 
death of a participant enrolled at 
WU/BJH/SLCH, report within 1 
working day.  Otherwise, report at the 
time of continuing review. 
 
 
 
Breach of 
confidentiality 
Within 10 working days. 
 
 
 
Incarceration 
If withdrawing the participant poses a 
safety issue, report within 10 working 
days.   
 
If withdrawing the participant does not 
represent a safety issue and the patient 
will be withdrawn, report at continuing 
review. 
 
 
 
 
Routine Reporting Timelines 
Event 
HRPO 
QASMC 
FDA 
Polaris 
Adverse event or SAE 
that does not require 
expedited reporting 
If they do not meet the definition of an 
unanticipated problem involving risks to 
participants or others, report summary 
information at the time of continuing review 
Adverse events will be 
reported in the toxicity 
table in the DSM report 
which is typically due 
every 6 months. 
The most current 
toxicity table from the 
DSM report is provided 
to the FDA with the 
IND’s annual report. 
Notify monthly via AE 
notification or cross-reporting 
reports/documents. 
Minor deviation 
Report summary information at the time of 
continuing review. 
 
 
 
Complaints 
If the complaint reveals an unanticipated problem 
involving risks to participants or others OR 
noncompliance, report within 10 working days.  If 
the event results in the death of a participant 
enrolled at WU/BJH/SLCH, report within 1 
 
 
 

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Routine Reporting Timelines 
Event 
HRPO 
QASMC 
FDA 
Polaris 
working day.  Otherwise, report at the time of 
continuing review. 
Incarceration 
If withdrawing the participant poses a safety 
issue, report within 10 working days.   
 
If withdrawing the participant does not represent a 
safety issue and the patient will be withdrawn, 
report at continuing review. 
 
 
 
 
Expedited Reporting Timelines for Secondary Sites 
Event 
WU (Coordinating Center) 
Local IRB 
FDA 
Polaris 
Serious AND unexpected 
suspected adverse reaction 
Report no later than 11 calendar days 
after it is determined that the information 
qualifies for reporting. 
Report all applicable 
events to local IRB 
according to local 
institutional guidelines. 
The research team at 
Washington University is 
responsible for reporting all 
applicable events to the 
FDA as needed. 
The research team at 
Washington University is 
responsible for reporting all 
applicable events to Polaris 
as needed. 
Unexpected fatal or life-
threatening suspected 
adverse reaction 
Report no later than 4 calendar days after 
initial receipt of the information. 
Unanticipated problem 
involving risk to participants 
or others 
Report no later than 4 calendar days after 
initial receipt of the information. 
Adverse event or SAE that 
does not require expedited 
reporting 
As per routine data entry expectations 
Protocol exception 
Approval must be obtained prior to 
implementing the change. 
 
 

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APPENDIX D: Washington University Unanticipated Problem Reporting Cover Sheet 
 
SAE COVER SHEET- Secondary Site Assessment 
 
Washington University HRPO#:  
Sponsor-Investigator:  
Subject Initials:   
Subject ID:  
Treating MD:  
Treating Site:  
EVENT TERM:  
Admission Date:  
EVENT GRADE: 
Date of site’s first notification: 
 
Treating MD Event Assessment:  
 
 
Is this event possibly, probably, or definitely related study treatment? 
 
 yes  
 
 no 
 
If yes, please list which drug (if more than one)______________________________ 
 
       
Explain ______________________________________________________________________ 
 
______________________________________________________________________________ 
 
 
 
_________________________           ___________________________      _________________  
Physician’s Name 
 
 
Physician’s Signature 
 
Date