Clinical trial NCT02788201
Conditions: Urothelial Carcinoma Bladder Cancer Urinary Bladder Neoplasms
Keywords: COXEN Bladder Cancer Molecular Profiles
Sponsor: National Cancer Institute (NCI)
Start date: 2017-03-27; Completion date: 2019-10-23
Enrollment: 8 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
Background:

Advanced urothelial cancer has no cure. But only a few chemotherapy drugs have been tested for it. The Co-eXpression ExtrapolatioN (COXEN) model predicts if cells respond to treatment. It may also help determine which drugs fight urothelial cancer based on the characteristics of a tumor. Researchers want to test if this model can choose the best therapy for advanced urothelial cancer within 3 weeks and how tumors respond to the next best therapy.

Objective:

To test if the COXEN model can choose the best therapy for advanced urothelial cancer within 3 weeks.

Eligibility:

People ages 18 and older whose urothelial cancer has spread after at least 1 line of chemotherapy

Design:

Participants will be screened with medical history, physical exam, blood and urine tests, and tumor scans.

Participants will provide a tumor sample from a previous surgery and a new biopsy. A needle will remove a small piece of tumor.

Participants will repeat screening tests, plus have an electrocardiogram (EKG) and scan. For the scan, they will get an injection of radioactive drug. They will lie in a machine that takes pictures.

Participants will take the drugs assigned by the COXEN model. They will have visits every 2-3 weeks. These will include blood and urine tests.

Participants will have tumor scans every 8-9 weeks.

Participants may have another biopsy.

Participants will take the drugs until they can't tolerate the side effects or their cancer worsens. They may be assigned to a second COXEN therapy.

Participants will have a follow-up visit 4-5 weeks after their last drug dose.

Participants will be contacted by phone every few months until death.

Detailed description:
Background:

* Patients presenting de novo with metastatic bladder cancer, or developing visceral metastatic disease after local treatment, are incurable with currently available therapeutic modalities.
* Only a small number of chemotherapeutic agents have been tested and very few have some single agent activity in the treatment of metastatic urothelial carcinoma. However most (\>100) Food and Drug Administration (FDA) approved anticancer agents have yet to be tested in this disease.
* Novel approaches to the development of genomic predictors of chemosensitivity that do not require clinical trials for their identification are urgently needed in order to identify agents that are clinically effective when either repurposed or discovered de novo specifically for urothelial carcinoma. Such repurposing of an FDA approved anticancer agent in order to advance therapy from one cancer to another would require only minimal clinical development, saving billions of dollars and reducing the time required to reach routine clinical practice.
* Our established extramural-intramural National Cancer Institute (NCI) collaboration pulls together significant expertise in biomarker development and clinical trial design in bladder cancer. The innovation of this group lies not only in the novel scientific approaches i.e. CoeXpression ExtrapolatioN (COXEN) under investigation, but also in the successful creation of a cohesive multi-institutional research collaboration dedicated to improved clinical outcomes in bladder cancer patients.
* COXEN uses molecular profiles as a Rosetta Stone for translating drug sensitivities of one set of cancers into predictions for another completely independent set of cell lines or human tumors. The COXEN methodology has been scrutinized and deemed methodologically sound by peer review. The ability of COXEN to predict drug effectiveness in patients a priori, from purely in vitro assays, is unique as no other tool currently either in practice or in development provides similar results.

Objectives:

\- To determine the feasibility of using the Co-eXpression ExtrapolatioN (COXEN) model in making a real-time treatment decision (within 3 weeks) in patients with advanced urothelial carcinoma.

Eligibility:

* Patients must have a histologically confirmed diagnosis of metastatic, progressive urothelial carcinoma of the bladder, urethra, ureter, or renal pelvis.
* Patients must have progressive metastatic disease defined as new or progressive lesions on cross-sectional imaging.
* Patients must have at least:

  * One measurable site of disease (according to Response Evaluation Criteria in Solid Tumors (RECIST) criteria)
  * Or, appearance of one new bone lesion
* Patients must have been previously treated, as defined by treatment with at least one prior cytotoxic chemotherapy regimen or agent. Patients may have received any number of prior cytotoxic agents.
* Archival tumor tissue must be available for enrollment.
* Tumor amenable to biopsy will be mandatory for this study.
* 18 years of age or older
* Eastern Cooperative Oncology Group (ECOG) performance status \<2 (Karnofsky \>60%)

Design:

* This will be a pilot single-arm, open-label study using the COXEN score to select the best next therapy from a list of 75 FDA approved anti-neoplastic drugs, in patients with metastatic bladder cancer who have progressed despite treatment with cytotoxic chemotherapy. Combinations of the listed agents may also be utilized provided that phase 1 data are available.
* The COXEN algorithm requires a multi-step process (pathology, tissue processing, messenger ribonucleic acid (mRNA) profiling, bioinformatics, etc.) and is potentially labor intensive and time intensive.
* Given the disease state of patients eligible for this protocol, using this algorithm to select a treatment would only be a worthwhile process to undertake if it can be demonstrated that a very high fraction of patients are likely to obtain the benefit from the procedure.

Arms: arm 1: Treatment regimen selected by CO eXpression ExtrapolatioN (COXEN) model
Interventions: intervention 1: 75 approved agents intervention 2: COXEN
Intervention descriptions: intervention 1: One or combination of agents: Abiraterone, Arsenic Trioxide, Asparaginase Escherichia coli source, Axitinib, Azacitidine, Bendamustine, Bleomycin, Bortezomib, Busulfan, Carboplatin, Carfilzomib, Carmustine, Chlorambucil, Cisplatin, Cladribine, Clofarabine, Crizotinib, Cytarabine, Dacarbazine, Dactinomycin, Dasatinib, Daunorubicin, Decitabine, Docetaxel, Doxorubicin, Epirubicin, Eribulin, Erlotinib, Estramustine, Etoposide, Exemestane, Floxuridine, Fludarabine, Fluorouracil, Gefitinib, Gemcitabine, Hydroxyurea, Idarubicin, Ifosfamide, Imatinib, Irinotecan, Ixabepilone, Lapatinib, Lomustine, Mechlor, Melphalan, Mercapto, Methotrexate, Mitomycin, Mitotane, Mitoxantrone, Nilotinib, Oxaliplatin, Paclitaxel, Pazopanib, Pentostatin, Romidepsin, Ruxolitinib, Sorafenib, Streptozocin, Sunitinib, Tamoxifen, Temsirolimus, Teniposide, Thioguanine, Thiotepa, Topotecan, Toremifene, Tretinoin, Vandetanib, Vemurafenib, Vinblastine, Vincristine, Vismodegib, and/or Vorinostat intervention 2: The CO eXpression ExtrapolatioN (COXEN) algorithm will be used to determine the next best therapy from among 75 Food and Drug Administration (FDA) approved agents (single agent or combination) in patients that have progressed on at least one chemotherapy regimen.

Protocol documents:
Prot_SAP_000.pdf: 
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Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
Abbreviated Title: COXEN study 
CC Protocol #: 16-C-0121 C 
Version Date: March 21st, 2018  
NCT Number: NCT02788201 
 
Title: A Pilot Clinical Trial of Genomic Based Assignment of Therapy in Advanced Urothelial 
Carcinoma 
NCI Principal Investigator: 
Andrea B. Apolo, M.D. 
 Genitourinary Malignancies Branch (GMB) 
National Cancer Institute 
 Building 10, Rm 13N240 
10 Center Drive 
Bethesda, MD 20892 
Phone: 301-480-0536 
E-mail: andrea.apolo@nih.gov 
 
 
 
 
  
 
Investigational agents: none 
Commercial agents: COXEN Drug Index 


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Version Date: March 21, 2019 
 
PRÉCIS  
Background: 
• 
Patients presenting de novo with metastatic bladder cancer, or developing visceral 
metastatic disease after local treatment, are incurable with currently available therapeutic 
modalities. 
• 
Only a small number of chemotherapeutic agents have been tested and very few have some 
single agent activity in the treatment of metastatic urothelial carcinoma. However most 
(>100) FDA approved anticancer agents have yet to be tested in this disease. 
• 
Novel approaches to the development of genomic predictors of chemosensitivity that do 
not require clinical trials for their identification are urgently needed in order to identify 
agents that are clinically effective when either repurposed or discovered “de novo” 
specifically for urothelial carcinoma. Such “repurposing” of an FDA approved anticancer 
agent in order to advance therapy from one cancer to another would require only minimal 
clinical development, saving billions of dollars and reducing the time required to reach 
routine clinical practice. 
• 
Our established extramural-intramural NCI collaboration pulls together significant 
expertise in biomarker development and clinical trial design in bladder cancer. The 
innovation of this group lies not only in the novel scientific approaches i.e. Co-eXpression 
ExtrapolatioN (COXEN) under investigation, but also in the successful creation of a 
cohesive multi-institutional research collaboration dedicated to improved clinical 
outcomes in bladder cancer patients. 
• 
COXEN uses molecular profiles as a “Rosetta Stone” for translating drug sensitivities of 
one set of cancers into predictions for another completely independent set of cell lines or 
human tumors. The COXEN methodology has been scrutinized and deemed 
methodologically sound by peer review. The ability of COXEN to predict drug 
effectiveness in patients a priori, from purely in vitro assays, is unique as no other tool 
currently either in practice or in development provides similar results.  
Objectives: 
• 
To determine the feasibility of using the “Co-eXpression ExtrapolatioN” (COXEN) model 
in making a real-time treatment decision (within 3 weeks) in patients with advanced 
urothelial carcinoma. 
Eligibility: 
• 
Patients must have a histologically confirmed diagnosis of metastatic, progressive 
urothelial carcinoma of the bladder, urethra, ureter, or renal pelvis. 
• 
Patients must have progressive metastatic disease defined as new or progressive lesions on 
cross-sectional imaging.  
• 
Patients must have at least: 
o One measurable site of disease (according to RECIST criteria)  
o Or, appearance of one new bone lesion 


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• 
Patients must have been previously treated, as defined by treatment with at least one prior 
cytotoxic chemotherapy regimen or agent. Patients may have received any number of prior 
cytotoxic agents. 
• 
Archival tumor tissue must be available for enrollment.   
• 
Tumor amenable to biopsy will be mandatory for this study. 
• 
18 years of age or older 
• 
ECOG performance status <2 (Karnofsky >60%) 
Design: 
• 
This will be a pilot single-arm, open-label study using the COXEN score to select the “best 
next therapy” from a list of 75 FDA-approved anti-neoplastic drugs, in patients with 
metastatic bladder cancer who have progressed despite treatment with cytotoxic 
chemotherapy. Combinations of the listed agents may also be utilized provided that phase 
1 data are available. 
• 
The COXEN algorithm requires a multi-step process (pathology, tissue processing, mRNA 
profiling, bioinformatics, etc.) and is potentially labor intensive and time intensive.  
• 
Given the disease state of patients eligible for this protocol, using this algorithm to select 
a treatment would only be a worthwhile process to undertake if it can be demonstrated that 
a very high fraction of patients is likely to obtain the benefit from the procedure.  


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TABLE OF CONTENTS  
PRÉCIS ...........................................................................................................................................2 
TABLE OF CONTENTS .................................................................................................................4 
1 
INTRODUCTION ..................................................................................................................6 
1.1 
Study Objectives ..................................................................................................... 6 
1.2 
Background and Rationale ...................................................................................... 6 
2 
ELIGIBILITY ASSESSMENT AND ENROLLMENT .......................................................14 
2.1 
Eligibility Criteria ................................................................................................. 14 
2.2 
Screening Evaluation ............................................................................................ 15 
2.3 
Registration Procedures ........................................................................................ 16 
2.4 
Baseline and On Study Evaluations ...................................................................... 17 
3 
STUDY IMPLEMENTATION ............................................................................................17 
3.1 
Study Design ......................................................................................................... 17 
3.2 
Drug Administration ............................................................................................. 21 
3.3 
Dose Modifications ............................................................................................... 21 
3.4 
Study Calendars .................................................................................................... 22 
3.5 
Criteria for Removal from Protocol Therapy and Off Study Criteria ................... 25 
4 
CONCOMITANT MEDICATIONS/MEASURES ..............................................................25 
5 
CORRELATIVE STUDIES .................................................................................................26 
5.1 
Biospecimen Collection ........................................................................................ 26 
5.2 
FDG-PET MRI...................................................................................................... 34 
5.3 
Sample Storage, Tracking and Disposition ........................................................... 34 
5.4 
Samples for Genetic/Genomic Analysis ............................................................... 36 
6 
DATA COLLECTION AND EVALUATION .....................................................................37 
6.1 
Data Collection ..................................................................................................... 37 
6.2 
Data Sharing Plans ................................................................................................ 38 
6.3 
Response Criteria .................................................................................................. 38 
6.4 
Toxicity Criteria .................................................................................................... 44 
7 
SAFETY REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING 
PLAN ....................................................................................................................................44 
7.1 
Definitions............................................................................................................. 44 
7.2 
NIH Intramural IRB and Clinical Director Reporting .......................................... 46 


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7.3 
Data and Safety Monitoring Plan .......................................................................... 46 
8 
STATISTICAL CONSIDERATIONS..................................................................................47 
9 
HUMAN SUBJECTS PROTECTIONS ...............................................................................47 
9.1 
Rationale For Subject Selection ............................................................................ 47 
9.2 
Participation Of Children ...................................................................................... 48 
9.3 
Participation of Subjects Unable to Give Consent ................................................ 48 
9.4 
Evaluation of Benefits and Risks/Discomforts ..................................................... 48 
9.5 
Risks/Benefits Analysis ........................................................................................ 49 
9.6 
Consent Process and Documentation .................................................................... 50 
10 
PHARMACEUTICAL INFORMATION .............................................................................50 
11 
REFERENCES .....................................................................................................................51 
12 
APPENDICES ......................................................................................................................55 
12.1 
Appendix A: Performance Status Criteria ............................................................ 55 
12.2 
Appendix B: COXEN Drug Index with FDA-Approved Dose and Schedule ...... 56 
12.3 
Appendix C:  Sample Collection for Correlative Studies ..................................... 67 
 


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1 
INTRODUCTION 
1.1 
STUDY OBJECTIVES 
1.1.1 Primary Objective 
To determine the feasibility of using the “Co-eXpression ExtrapolatioN” (COXEN) model in 
making a real-time treatment decision (within 3 weeks) in patients with advanced urothelial 
carcinoma. 
1.1.2 Secondary Objectives 
1.1.2.1  To assess the progression-free survival using treatment suggested by the COXEN model 
in patients with advanced urothelial carcinoma. 
1.1.2.2 To assess the response rate using treatment suggested by the COXEN model in patients 
with advanced urothelial carcinoma. 
1.1.2.3 To assess the overall survival using treatment suggested by the COXEN model in patients 
with advanced urothelial carcinoma. 
1.1.2.4 To evaluate toxicity with “best next therapy”. 
1.1.3 Exploratory Objectives 
1.1.3.1 To characterize changes in mutation frequency due either to metastatic progression or 
initial therapy by exome sequencing FFPE metastatic tumor samples from patients prior 
to “best next therapy” compared to archival primary tissue.  
1.1.3.2 To identify the sensitivity of cellular circulating tumor DNA mutation detection and 
changes in the mutational landscape as patients undergo COXEN directed treatment. 
1.1.3.3 To assess feasibility of PET-MRI in patients with metastatic bladder cancer.  
1.1.3.4 To determine sensitivity and specificity of whole body diffusion weighted MRI imaging 
in detection of distant metastasis.  
1.1.3.5 To evaluate role of PET-MRI in evaluation of response to therapy.  
1.2 
BACKGROUND AND RATIONALE 
1.2.1 The Burden and Natural History of Bladder Cancer 
Bladder cancer is the fifth most common cancer in the United States, with >70,000 new cases and 
> 14,000 deaths each year [1]. While the majority of patients present with non-muscle-invasive 
tumors, approximately 30-40% of patients have muscle-invasive tumors and a few harbor 
metastatic disease at the time of diagnosis. Surgical resection of the bladder (radical cystectomy) 
cures approximately 50% of these patients, while the remainder of patients develops lethal 
metastatic recurrence.  
Summary: Patients presenting de novo with metastatic bladder cancer, or developing visceral 
metastatic disease after local treatment, are incurable with currently available therapeutic 
modalities. 
1.2.2 Chemotherapy for Metastatic Bladder Cancer 
The development of the MVAC (methotrexate, vinblastine, doxorubicin, cisplatin) regimen in the 
1980’s was an advance in the treatment of metastatic bladder cancer, with multiple trials 


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demonstrating objective response rates of <50% [2-5]. However, durable responses were rare and 
treatment-related toxicities were problematic. A landmark phase III trial comparing MVAC with 
gemcitabine plus cisplatin (GC) demonstrated similar overall survival outcomes (~14 months) 
between the regimens with better tolerability with GC [6] resulting in GC becoming the first-line 
treatment regimen of choice.  
There is currently no standard chemotherapy for patients with metastatic bladder cancer who have 
progressed on first-line treatment. A variety of cytotoxic agents used in this “second-line” or 
“salvage” setting have demonstrated response rates of approximately 5-20% [7-14]. A randomized 
phase III trial of second-line vinflunine versus placebo in patients with metastatic disease failed to 
demonstrate a statistically significant improvement in survival [15]. These findings can be 
summarized as follows: 1) only a subset of patients with bladder cancer benefit (i.e. has some level 
of objective response) from cytotoxic agents; 2) only a small number of the >120 FDA approved 
anticancer agents have ever been tested and only a few of these have clinically significant single 
agent activity in urothelial cancer. The latter fact is emblematic of urothelial cancer being one of 
the most nationally underfunded cancers despite its clinical impact on the American population.  
Combination second-line therapy may have more clinical activity then just single-agent therapy. 
The COXEN waterfall plot reports on the high activity of several agents that may be used in 
combination if previously tested in at least a phase I study and found to be safe. 
Summary: Only a small number of chemotherapeutic agents have been tested and very few have 
some single agent activity in the treatment of metastatic urothelial carcinoma. However most 
(>100) FDA approved anticancer agents have yet to be tested in this disease. 
1.2.3 Current Genomic Predictors of Chemosensitivity 
The therapeutic response to a particular anticancer agent is modulated by multiple signaling 
pathways. As a result, effective prediction of a given patient’s response requires analysis of 
multiple biomarkers across different gene networks [16]. Multi-gene expression-based prediction 
strategies have already demonstrated utility in predicting clinical outcomes [17-21]. Some of these 
tools, such as Oncotype DX™ and Mamma Print™, have become incorporated into the clinical 
care of patients with breast cancer. Despite the tremendous potential utility of multi-gene 
predictors, the current development of such predictors of drug sensitivity has critical limitations. 
Foremost, the majority of these genomic models have been developed using human patient data 
from clinical trials exploring treatment with a specific regimen. As a result, the predictive models 
are relevant only to that specific treatment and new models must be developed for each new 
treatment regimen. Furthermore, as indicated above, since only a small fraction of all FDA 
approved anticancer agents has been used in bladder cancer, single agent and combination trials 
would be required in order to generate these biomarkers. This lengthy and costly approach is 
clearly not feasible and cannot keep pace with the current arsenal of chemotherapeutic or targeted 
agents in the drug pipeline.  
Summary: Novel approaches to the development of genomic predictors of chemosensitivity that 
do not require clinical trials for their identification are urgently needed in order to identify agents 
that are clinically effective when either repurposed or discovered “de novo” specifically for 
urothelial carcinoma. Such “repurposing” of an FDA approved anticancer agent as single agent 
therapy or in combination in order to advance therapy from one cancer to another would require 


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only minimal clinical development, saving billions of dollars and reducing the time required to 
reach routine clinical practice. 
1.2.4 UCCC-NCI Bladder Cancer Collaboration 
Progress in clinical drug development in advanced bladder cancer is hampered by a relative lack 
of industry interest and the inability of most single centers to conduct even large Phase II trials in 
a timely manner. Over the past 3 years, we have formed a collaboration between the University of 
Colorado Comprehensive Cancer Center (UCCC), the National Cancer Institute (NCI) and the 
NIH-Clinical Center (CC). This group is comprised of basic, translational and clinical investigators 
focused on the development of novel therapeutic approaches for the treatment of bladder cancer. 
In this manner, the group leverages the bladder cancer expertise of its participating researchers and 
NCI funded infrastructure (UCCC P30, NCI and NIH Clinical Center). 
Summary: Our established extramural-intramural NCI collaboration pulls together significant 
expertise in biomarker development and clinical trial design in bladder cancer. The innovation of 
this group lies not only in the novel scientific approaches (i.e. COXEN) under investigation, but 
also in the successful creation of a cohesive multi-institutional research collaboration dedicated to 
improved clinical outcomes in bladder cancer patients.  
1.2.5 COXEN for Individualized Chemosensitivity Prediction 
The “Co-eXpression ExtrapolatioN” (COXEN) algorithm utilizes molecular profiles as a “Rosetta 
Stone” for translating drug sensitivity signatures of one set of cancers into that of another [22]. 
The COXEN Principle has been validated using the NCI Developmental Therapeutics Program 
NCI-60 cell lines [23-25]. The NCI-60 panel is comprised of cell lines from diverse human cancers 
that have been profiled at the DNA, RNA, protein, and functional levels. The molecular 
characteristics of each of the NCI-60 cell lines and their relationship to patterns of anticancer 
activity in over 100,000 chemically defined compounds and natural product extracts has been 
established [23, 26-28]. Unfortunately, it was not feasible to include all important tumor types in 
the NCI-60 such as bladder cancer. Furthermore, even if other cancer cell types were added to the 
panel now, all compounds screened in the past 20 years would have to be tested again in the 
updated panel to gain the full predictive power of the database for all legacy compounds. Several 
years ago, these limitations raised two practical questions for our research team: Can drug 
sensitivity data on the NCI-60 panel be extrapolated to predict sensitivity of cell lines not included? 
More ambitiously, can the screening data be used to obtain predictive power for clinical responses 
of human cancers? To address these questions, we developed the COXEN algorithm and have 
subsequently demonstrated accurate predictions of chemotherapy sensitivity in both human 
bladder cancer cells lines and in bladder cancer patients [22, 29]. COXEN was retrospectively 
tested in seven independent cohorts of patients with breast (n = 275), bladder (n = 59), and ovarian 
(n= 143) cancer treated with multiagent chemotherapy, of which 233 patients were from 
prospectively enrolled clinical trials. In all studies, COXEN effectively stratified tumor response 
and patient survival independent of established clinical and pathologic tumor variables. In bladder 
cancer patients treated with neoadjuvant methotrexate, vinblastine, adriamycin, and cisplatin, the 
3-year overall survival for those with favorable COXEN gene expression models’ score was 81% 
versus 33% for those with less favorable scores (P = 0.002). COXEN for breast cancer patients 
treated with 5-fluorouracil, doxorubicin, and cyclophosphamide and ovarian cancer patients 
treated with platinum-containing regimens also stratified patient survival [5-year overall survival 


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100% versus 74% (P = 0.05) and 3-year overall survival 68% versus 43% (P = 0.008), 
respectively]. The COXEN algorithm is currently being studied in a randomized phase II study of 
muscle-invasive bladder cancer being run by the Southwest Oncology Group (SWOG) 
(NCT02177695). Muscle-invasive (pT2-4N0) patients are randomized to receive gemcitabine and 
cisplatin (GC) or methotrexate, vinblastine, adriamycin and cisplatin (MVAC), the patient then 
undergoes cystectomy and tumor is analyzed by the COXEN and the COXEN score for the GC vs 
MVAC combination is compared to test whether COXEN predicted a pathologic response for the 
combination (pT0). This approach has also been intensely scrutinized by well-known statisticians 
(MD Anderson Statistics Group, K. Baggerly; CTEP-Statistical Group, Lisa Meier-McShane; 
SWOG-Statistical Group, Cathy Tangen) and evaluated in other tumor types and by other 
investigators with similarly promising predictive results. This approach remains to be examined in 
a prospective clinical study. 
Summary: COXEN uses molecular profiles as a “Rosetta Stone” for translating drug sensitivities 
of one set of cancers into predictions for another completely independent set of cell lines or human 
tumors. The COXEN methodology has been scrutinized and deemed methodologically sound by 
peer review. The ability of COXEN to predict drug effectiveness in patients a priori, from purely 
in vitro assays, is unique as no other tool currently either in practice or in development provides 
similar results. 
1.2.6 Preliminary Data 
1.2.6.1 Use of COXEN to rank predicted sensitivity of bladder tumors to 75 FDA approved 
anticancer agents  
1.2.6.1.1 Drug Selection 
Approximately 120 FDA approved anticancer agents have been tested in vitro by the 
Developmental Therapeutics Program (DTP) at the National Cancer Institute using the NCI-60 
cell line panel (http://dtp.nci.nih.gov). The 75 FDA approved anticancer agents that will be used 
in Aim 1 (Table 1) of this proposal were those drugs that had 1) complete data sets and 2) a 
variation in response between “sensitive” and “resistant” cell lines that was determined by 
performing a linear regression on scaled rankings of cell lines vs. scaled negative logGI50 values 
for each drug following the removing cell lines in the sensitive and resistant groups. This analysis 
verified that there was a graded drug response within the NCI-60 panel that could be utilized for 
developing and testing gene expression-based COXEN models. 
 
Table 1. Selected 75 FDA Approved Anticancer Agents 
Abiraterone 
Cisplatin 
Eribulin 
Imatinib 
Oxaliplatin 
Thiotepa 
Arsenic Trioxide 
Cladribine 
Erlotinib 
Irinotecan 
Paclitaxel 
Topotecan 
Asparaginase 
Clofarabine 
Estramustine 
Ixabepilone 
Pazopanib 
Toremifene 
Axitinib 
Crizotinib 
Etoposide 
Lapatinib 
Pentostatin 
Tretinoin 
Azacitidine 
Cytarabine 
Exemestane 
Lomustine 
Romidepsin 
Vandetanib 


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Bendamustine 
Dacarbazine 
Floxuridine 
Mechlorethamin 
Ruxolitinib 
Vemurafenib 
Bleomycin 
Dactinomycin 
Fludarabine 
Melphalan 
Sorafenib 
Vinblastine 
Bortezomib 
Dasatinib 
Fluorouracil 
Mercaptopurine 
Streptozocin 
Vincristine  
Busulfan 
Daunorubicin 
Gefitinib 
Methotrexate 
Sunitinib 
Vismodegib 
Carboplatin 
Decitabine 
Gemcitabine 
Mitomycin 
Tamoxifen 
Vorinostat 
Carfilzomib 
Docetaxel 
Hydroxyurea 
Mitotane 
Temsirolimus 
 
Carmustine 
Doxorubicin 
Idarubicin 
Mitoxantrone 
Teniposide 
 
Chlorambucil 
Epirubicin 
Ifosfamide 
Nilotinib 
Thioguanine 
 
1.2.6.1.2 COXEN Score Determination and Patient Drug Assignment 
Microarray expression data from the GeneChip® Human Genome U133 Plus 2.0 Array will be 
processed and normalized using frozen robust multiarray analysis (fRMA) [30] that allows for the 
individual analysis of microarray data and subsequent utilization with other microarray data for 
data analysis such as the COXEN algorithm [22]. Gene expression models (GEM) for the 
prediction of chemosensitivity have been constructed for all the 75 FDA approved anticancer 
agents listed in Table 1 using the COXEN method [22] with biomarker identification done using 
drug sensitive and resistant gene expression from the NCI60 cell panel and co-expression done 
using gene expression microarray data from a bladder tumor cohort [31]. The modeling process is 
completed in three distinct steps; preprocessing, feature extraction, and finally, model construction 
and validation. Preprocessing involves the application of frozen robust multiarray analysis (frma) 
to perform background correction, normalization, and summarization allowing for direct 
comparison among samples [30]. fRMA was applied using the fRMA package included in the 
Bioconductor suite in R. Second, for each drug, a set of biomarkers was found by implementing 
COXEN (coexpression extrapolation) [22] to 189 bladder tumor samples from the Lehmann tumor 
panel in conjunction with the National Cancer Institute 60 cell anticancer drug screening panel 
(NCI60). Finally, model building has been approached from two different methods. Using the 
biomarkers selected by COXEN, a linear discriminant analysis model (LDA) was employed on a 
smaller subset of genetic markers identified by Misclassification Penalized Posterior Classification 
(MiPP) to the top twelve most resistant and most sensitive cell lines in the NCI60 for every selected 
drug. This approach requires a new model to be constructed whenever a new sample is to be 
predicted and often leads to unstable outcomes. For this reason, a second approach was developed. 
The second method employed logistic regression with an L2 penalty using dual optimization 
included in the Scikit-learn machine learning suite in Python [32]. While MiPP systematically adds 
features until addition of features does not increase model performance beyond a user defined 
threshold the L2 penalty allows all COXEN selected features to be included in the model by 
mathematically weighting features that contribute most to model performance. These weights are 
controlled via a parameter that was selected by performing cross validation over all possible 
0.75/0.25 testing validation splits of the top 12 most resistant and sensitive cell lines in the NCI60. 
After the L2 parameter was determined via cross-validation, a final model was constructed using 


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all 12 most resistant and sensitive cell lines. Once complete the final model is stationary, meaning 
the model is not reconstructed for each new sample allowing robust prediction among new 
samples. Drug sensitivity and resistance predictions will range from -1 to +1 with -1 representing 
the most drug-resistant and +1 representing the most drug-sensitive. The resulting data will be 
displayed for the treating clinician as shown in Figure 1 with the agents ranked in groups of 5 in 
descending order from the most sensitive on one of the tumors shown in Figure 2. The robust 
nature of these predictions is shown in Figure 2 which plots the correlation of predicted sensitivity 
to these 75 drugs in 2 separate biopsies taken from 3 urothelial bladder tumors. While we will 
evaluate this intratumoral variation in the proposal by obtaining 2-3 core biopsies of the same site, 
we do not expect this to metrically alter the predictions. 
 
1.2.7 Overview and Hypothesis 
Multiple chemotherapeutic agents have demonstrated response rates of 5-20% when utilized as 
second-line chemotherapy in patients with metastatic bladder cancer. However, no 
chemotherapeutic agents have been shown to improve survival in this setting and therefore, there 
is no FDA-approved second line drug for patients with metastatic bladder cancer. Furthermore, the 
ability to select patients most likely to respond to a given anti-neoplastic agent is critical to 
improving outcomes in this clinical state. Hence, our ultimate objective is to build on the prior 
Figure 1: Waterfall-plot of predicted drug sensitivity of a bladder tumor (one of the samples in Figure 2) to the 75 
FDA anti-neoplastic drugs based on microarray gene expression and COXEN analysis. COXEN scores are 
normalized from -1 to +1 with -1 representing the most drug resistant and +1 representing the most drug 
sensitive. Agents ranked in groups of 5 in descending order from the most sensitive. 
Floxuridine
Gemcitabine
Fluorouracil
Busulfan
Toremifene
Streptozocin
Vorinostat
Vismodegib
Crizotinib
Clofarabine
Dasatinib
Bleomycin
Imatinib
Gefitinib
Chlorambucil
Docetaxel
Valrubicin
Vincristine
Mercaptopurine
Paclitaxel
Cisplatin
Pentostatin
Temsirolimus
Cytarabine
Epirubicin
Thioguanine
Exemestane
Mitomycin
Decitabine
Oxaliplatin
Bortezomib
Doxorubicin
Pazopanib
Ixabepilone
Ruxolitinib
Tamoxifen
Ifosfamide
Irinotecan
Nilotinib
Arsenic Trioxide
Abiraterone
Eribulin
Carmustine
Methotrexate
Mitoxantrone
Daunorubicin
Romidepsin
Cladribine
Hydroxyurea
Topotecan
Teniposide
Vinblastine
Idarubicin
Erlotinib
Vandetanib
Carfilzomib
Mitotane
Thiotepa
Vemurafenib
Carboplatin
Dactinomycin
Dacarbazine
Sunitinib
Estramustine
Fludarabine
Asparaginase
Lapatinib
Axitinib
Mechlorethamine
Tretinoin
Sorafenib
Melphalan
Bendamustine
Etoposid
Lomu
Aza
-1
0
1
Resistant - Sensitive
Drugs 11-15
Dasatinib
Bleomycin
Imatinib
Gefitinib
Chlorambucil
Drugs 6-10
Streptozocin
Vorinostat
Vismodegib
Crizotinib
Clofarabine
Drugs 1-5
Floxuridine
Gemcitabine
Fluorouracil
Busulfan
Toremifene


12 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
retrospective experience with COXEN and prospectively test the hypothesis that use of tumor 
COXEN scores to select “best next therapy” for individual patients with chemotherapy-refractory 
metastatic bladder cancer will provide superior response rates, progression free and overall 
survival compared to historical controls. However, given the complexity of this approach, before 
embarking on such a trial, we propose to carry out a pilot trial evaluating feasibility of COXEN 
assigned “best next therapy”. 


13 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
 
 
 
Figure 2: COXEN Scores for 75 FDA anti-
neoplastic drugs on two independent core biopsies 
(M and Q) in tumors from 3 different patients. 
Correlations coefficient is shown. 
-1.00
-0.75
-0.50
-0.25
0.00
0.25
0.50
0.75
1.00
-1.00
-0.75
-0.50
-0.25
0.00
0.25
0.50
0.75
1.00
Q Sample COXEN Score
r=0.8590
-1.00
-0.75
-0.50
-0.25
0.00
0.25
0.50
0.75
1.00
-1.00
-0.75
-0.50
-0.25
0.00
0.25
0.50
0.75
1.00
M Sample COXEN Score
Q Sample COXEN Score
r=0.8630
-1.00
-0.75
-0.50
-0.25
0.00
0.25
0.50
0.75
1.00
-1.00
-0.75
-0.50
-0.25
0.00
0.25
0.50
0.75
Q Sample COXEN Score
r=0.9232
5
0
5
0
5
0
5
r=0 8590
0
5
0
5
0
5
0
5
r=0.9232
5
0
5
0
5
0
5
r=0.8630
R=0.86
R=0.92
R=0.86


14 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
2 
ELIGIBILITY ASSESSMENT AND ENROLLMENT  
2.1 
ELIGIBILITY CRITERIA 
2.1.1 Inclusion Criteria 
2.1.1.1 Patients must have a histologically confirmed diagnosis of metastatic, progressive 
urothelial carcinoma of the bladder, urethra, ureter, or renal pelvis. 
2.1.1.2 Patients must have progressive metastatic disease defined as new or progressive lesions 
on cross-sectional imaging.  
2.1.1.3 Patients must have at least: 
2.1.1.3.1 One measurable site of disease (according to RECIST criteria), defined as at least one 
lesion that can be accurately measured in at least one dimension (longest diameter to be 
recorded for non-nodal lesions and short axis for nodal lesions) as >20 mm with 
conventional techniques or as >10 mm with spiral CT scan.  See Section 6.3 for the 
evaluation of measurable disease. 
2.1.1.3.2 Or, appearance of one new bone lesion 
2.1.1.4 Patients must have been previously treated with at least one prior cytotoxic chemotherapy 
regimen or agent. Patients may have received any number of prior cytotoxic agents. 
2.1.1.5 Archival tumor tissue must be available for enrollment.   
2.1.1.6 Tumor amenable to biopsy will be mandatory for this study. 
2.1.1.7  Age >18 years. ECOG performance status <2 (Karnofsky >60%, see Appendix A). 
2.1.1.8 Patients must have normal organ and marrow function as defined below:  
 hemoglobin  
    
 
 
≥ 9 g/dL 
 leukocytes  
 
 
 
 
>3,000/mcL 
 absolute neutrophil count  
>1,200/mcL 
 platelets  
 
 
 
 
 
>75,000/mcL 
 total bilirubin 
 
 
 
 
within normal institutional limits 
 AST(SGOT)/ALT(SGPT)  
<2.5 X institutional upper limit of normal 
 creatinine 
 
 
 
 
 
1.5 x the normal institutional limits 
 
OR 
 creatinine clearance  
 
 
>40 mL/min/1.73 m2 
2.1.1.9 Because many of the therapeutic agents used in this trial are known to be teratogenic, 
women of child-bearing 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 she or her partner is participating in this study, she should inform her 
treating physician immediately. 
2.1.1.10 HIV-positive patients on combination antiretroviral therapy may be eligible if there are 
no pharmacokinetic interactions with the agents used on the study, stable on CART 
therapy and CD4 is >200 and viral load is undetectable.  
2.1.1.11 Ability of subject to understand and the willingness to sign a written informed consent 
document. 


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Abbreviated Title: COXEN study 
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2.1.2 Exclusion Criteria 
2.1.2.1 The patient has received cytotoxic chemotherapy (including investigational cytotoxic 
chemotherapy) within 3 weeks or biologic agents (e.g., cytokines or antibodies) within 4 
weeks prior to study enrollment. 
2.1.2.2 Patients who are receiving any investigational agents. 
2.1.2.3 Patients with known brain metastases should be excluded from this clinical trial because 
of their poor prognosis and because they often develop progressive neurologic 
dysfunction that would confound the evaluation of neurologic and other adverse events. 
Patients with brain metastases that are stable after ≥ 1 year after primary surgery or 
radiation will not be excluded.   
2.1.2.4 The subject has not recovered to baseline or CTCAE ≤ Grade 1 from toxicity due to all 
prior therapies except alopecia and other non-clinically significant AEs. 
2.1.2.5 Uncontrolled intercurrent illness including, but not limited to, ongoing or active infection, 
symptomatic congestive heart failure, unstable angina pectoris, cardiac arrhythmia, or 
psychiatric illness/social situations that would limit compliance with study requirements. 
2.1.2.6 Patients who are Hepatitis B or C positive.  
2.1.2.7 Pregnant women are excluded from this study because the agents used in the study have 
the potential for teratogenic or abortifacient effects. Because there is an unknown but 
potential risk for adverse events in nursing infants secondary to treatment of the mother 
with these agents, breastfeeding should be discontinued if the mother is treated with these 
agents. These potential risks may also apply to other agents used in this study. 
2.2 
SCREENING EVALUATION 
Study eligibility is based on meeting all of the study inclusion criteria and none of the exclusion 
criteria at screening before study treatment administration. Screening evaluations may be 
performed as part of an NIH Screening protocol. This does not include the baseline correlative 
studies that will only be performed after the patient has signed the consent form. 
The following assessments will be conducted no more than 28 days prior to subjects receiving 
the first dose of study drug on this protocol: 
x 
History including performance status assessment and medication history 
x 
Physical examination including height, weight, vital signs 
x 
Laboratory assessments: CBC with differential, acute care panel (sodium, potassium, 
chloride, total CO2 Bicarbonate, creatinine, glucose, urea nitrogen), mineral panel 
(albumin, calcium total, magnesium total, phosphorus), hepatic panel (alkaline 
phosphatase, ALT/GPT, AST/GOT, total bilirubin, direct bilirubin), ionized calcium, 
amylase, lipase, LDH, total protein, GGT, Hepatitis B and C. 
x 
24-hour urine collection for creatinine clearance if needed  
x 
Urinalysis 
x 
Urine or serum HCG (in women of childbearing potential) 
x 
CT scan of chest, abdomen and pelvis (or all known sites of disease) (MRI may be 
performed in subjects unable to tolerate contrast for CT). 


16 
Abbreviated Title: COXEN study 
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x 
CD4 levels and HIV viral load (for patients with known HIV infection on combination 
antiretroviral therapy) 
x 
Pathology evaluation/confirmation of urothelial carcinoma as documented in a pathology 
report confirmed by the referring institution’s pathology department 
x 
Research biopsy (see section 5.1). This biopsy may be performed while patients are 
receiving other therapies, if they have signed this consent and been registered for screening 
as described in Section 2.3.  
2.3 
REGISTRATION PROCEDURES 
Registration will be a two-part process as patients are screened on this protocol. Authorized staff 
must register an eligible candidate with NCI Central Registration Office (CRO) within 24 hours 
of signing consent. To initially register a subject after the participant has signed the consent, 
complete the top portion the registration Eligibility Checklist from the web site 
(http://home.ccr.cancer.gov/intra/eligibility/welcome.htm) indicating that the patient is being 
registered for screening and sent via encrypted email to: NCI Central Registration Office 
ncicentralregistration-l@mail.nih.gov. Once eligibility is confirmed after completion of screening 
studies, complete the remainder of the form which is the eligibility checklist, indicating that the 
patient is being registered for treatment and email the completed registration checklist to the CRO 
at NCI Central Registration Office ncicentralregistration-l@mail.nih.gov. After confirmation of 
eligibility at Central Registration Office, CRO staff will call pharmacy to advise them of the 
acceptance of the patient on the protocol prior to the release of any investigational agents. 
Verification of Registration will be forwarded electronically via e-mail to the research team. A 
recorder is available during non-working hours.  
Subjects that do not meet screening criteria should be removed from the study following the 
procedure in section 3.5.4.  
2.3.1 Treatment Assignment Procedures 
Cohorts 
Number Name 
Description 
1 
Cohort 1 
Patients with metastatic, progressive urothelial 
carcinoma of the bladder, urethra, ureter, or renal pelvis  
 
Arms 
Number Name 
Description 
1 
Arm 1 
Treatment regimen as selected by COXEN model 
 
2.3.2 Arm Assignment 
Patients in Cohort 1 will be directly assigned to Arm 1. 


17 
Abbreviated Title: COXEN study 
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2.4 
BASELINE AND ON STUDY EVALUATIONS 
The baseline assessments do not need to be repeated if they have been performed at screening 
within the appropriate 16 day timeframe. 
x 12-lead ECG 
x Vital Signs 
x Laboratory Assessments 
 
CBC with differential 
Serum chemistries: 
 
Hepatic panel (alkaline phosphatase, ALT/GPT, AST/GOT, total bilirubin, direct 
bilirubin) 
 
Acute care panel (sodium, potassium, chloride, total CO2 (Bicarbonate), creatinine, 
glucose, urea nitrogen) 
 
Mineral panel (albumin, calcium total, magnesium total, phosphorus) 
 
Ionized calcium 
 
Amylase 
 
Lipase 
 
Lactate dehydrogenase (LDH) 
 
Total protein 
 
J-glutamyl transferase (GGT) (baseline only) 
x 
Urinalysis  
x 
TSH (baseline only) 
x 
PT/INR or PTTT  
x 
Urine or serum HCG (in women of childbearing potential) within 48 hours prior to 
receiving study medication. 
x 
Research blood (see section 5.1) 
x 
Research FDG-PET MRI (see section 5.1.10) 
On study assessments for other procedures performed during the study are found in the study 
calendar section 3.4. 
3 
STUDY IMPLEMENTATION 
3.1 
STUDY DESIGN 
This will be a Pilot single-arm, open-label study using the COXEN score to select the “best next 
therapy” from a list of 75 FDA anti-neoplastic drugs (Table 1), in patients with metastatic bladder 
cancer who have progressed despite treatment with cytotoxic chemotherapy. Agents on the list 
may be given as single agent therapy or as combination therapy provided there are at least phase 
1 data to support the use of the combination. Eligible patients will be treated with one or a 
combination of drugs based on the 75-drug COXEN algorithm analysis. Each participant will be 
given an option to be assigned to a second COXEN-based regimen at the time of disease 


18 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
progression. The COXEN analysis for retreatment will be performed on the biopsy collected at the 
time of progression; however, if the patient refuses the biopsy or the recurrent disease is not 
amenable to biopsy, the baseline COXEN waterfall plot will be used to make the decision. No 
more than 2 COXEN-based assignments will be allowed on study. The primary objective of this 
pilot study is to determine if a sufficiently high fraction of patients is able to have results produced 
by the COXEN algorithm within 3 weeks of initiating the process. The COXEN algorithm requires 
a multi-step process (pathology, tissue processing, mRNA profiling, bioinformatics etc.) and is 
potentially labor intensive and time intensive. Given the disease state of patients eligible for this 
protocol, using this algorithm to select a treatment would only be a worthwhile process to 
undertake if it can be demonstrated that a very high fraction of patients is likely to obtain the 
benefit from the procedure. The secondary objectives include: assessment of toxicity, response 
rate and progression-free survival by Response Evaluation Criteria in Solid Tumors (RECIST) 
version 1.1 and overall survival of the COXEN selected therapeutic agent(s).  
 
 
Figure 3 
3.1.1 Schema  
(See Figure 3) 
1. 
Consultation in the medical oncology clinic at the NIH Clinical Center 
2. 
Provide tumor block of the primary tumor 
3. 
Undergo a CT of the chest, abdomen and pelvis (MRI may be performed in subjects unable 
to tolerate contrast for CT)  


19 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
5. 
Mandatory core biopsy of a metastatic site of disease (Sample 2, Figure 4). (Note: samples 
1 and 2, Figure 4 will be batched and analyzed together once sample 2 has been collected.) 
6. 
Tissue will be processed and whole genome Affymetrix microarray (Hu133A or similar) 
will be performed in Dr. Meltzer’s CLIA certified lab at the NIH Clinical Center. 
7. 
The genomic data will be transferred from the NIH to University of Colorado Cancer 
Center (UCCC) as described in section 6.2.1 and COXEN analysis performed. 
8. 
UCCC team will return a rank report with COXEN scores for the 75 anti-neoplastic FDA 
approved drugs (Figure 1).  
9. 
Molecular Tumor Board: Drug selection will be based on COXEN predicted potency and 
tailored to the patient’s case (renal function, allergies prior use etc.) as determined by the 
advisement from the members of a molecular tumor board. 
10. 
Drug(s) will be ordered from the NIH pharmacy. 
11. 
COXEN tailored treatment will be started within 3 weeks of initial biopsy.  
12. 
Depending on restaging schedule, restaging CT at 8 weeks or 9 weeks (± 1 week).  
13. 
Optional core biopsy of a metastatic site will be scheduled at time of disease progression.  
COXEN analysis will be performed along with mutational analysis as described in section 
5.1.1. 
 
3.1.2 Pathology 
Tumor profiling will be carried out on FFPE tissues from the primary site obtained from the 
referring hospital (Sample 1, Figure 4) and on FFPE biopsy tissue (Samples 2 and 5, Figure 4) 
obtained at the NIH Clinical Center with study enrollment. The RNA extraction and methodology 
will be as described in our previous work [31]. We will profile the metastasis biopsy and use this 
data to generate the COXEN Score that will rank the 75 FDA approved anti-neoplastic drugs. 
3.1.3 COXEN Score Determination 
This will be carried out in two steps.  
 
 
Figure 4: Tissue and blood samples available for gene expression and mutational evaluation in relation to prior or planned 
treatment 
Primary
Primary 
Course of 
Systemic 
Therapy
Metastasis
Blood
Blood
3
1
4
2
Second 
Course of 
Systemic 
Therapy
“Best 
Next 
Therapy”
Aim 1
FFPE Tumor Tissue
Outside Hospital
FFPE Tumor Tissue,
Fresh Blood
NIH Clinical Center
Fresh Blood
NIH Clinical Center
Any number 
of additional 
systemic 
therapy 
Metastasis
4
5


20 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
3.1.3.1 Quality Control Assessment of Microarray Data 
Microarray data from individual patient tumors will be generated using the GeneChip® Human 
Genome U133 Plus 2.0 Array (Affymetrix, Santa Clara, CA) and will be assessed for quality using 
the following accepted criteria: 1) inspection of array images and observe for “spatial artifacts” 
and obvious problems; 2) analysis of box and histogram plots of log2(PM) values for each 
individual array with comparison for consistency across a set of fixed bladder tumor arrays; 3) 
proportion present plots will be generated using probe pairs (PM, MM) testing gene “presence” or 
“absence” and analyzing for consistency compared to a fixed set of bladder tumor arrays; 4) 
hybridization controls (bioB, bioC, bioD, and cre) will be plotted for verification of increasing 
concentrations within the array; 5) QC stats plots will be created measuring % present, average 
background intensity, bioB, bioC, bioD and creX presence, and 3’:5’ ratios of actin and GAPDH; 
and 6) evaluation of RNA degradation plots. If an outlier is identified in more than 2 of these tests, 
the sample will be flagged and the data deemed unreliable for COXEN analysis.  
3.1.3.2 COXEN analysis 
COXEN analysis will be done by the Cancer Pharmacology Shared Resource for the University of 
Colorado Cancer Center by Dr. Daniel L. Gustafson and Dr. Ryan J. Hansen as a backup under 
the supervision of Dr. Dan Theodorescu. 
Sample Processing & Reporting: 
• 
Coded gene expression data will be sent to Dr. Gustafson or Hansen at the Cancer 
Pharmacology Shared Resource with a unique identifier (UI). 
• 
Data will be processed for COXEN analysis with the UI carried and validated at all steps 
using a defined SOP. 
• 
All data processing and predictive steps will be tracked with the use of a “sample checklist” 
as outlined in the SOP verifying each step. 
• 
Sample output (drug sensitivity/resistance prediction) will be sent to the proper individual 
responsible for collecting the data along with an electronic copy of the “sample checklist” 
using the UI to identify the sample.  
Sample Analysis: 
1. Receive and log in sample data and begin “sample checklist” 
2. Perform QC Analysis for MicroArray Data. 
3. Frozen RMA process the gene expression data. 
4. Run the COXEN algorithm for prediction of drug sensitivity and resistance.  
5. Complete the data analysis and presentation (as specified in the SOP) and deliver. 
QC Analysis Steps for Individual MicroArrays 
1. Inspect array images 
– Look for “spatial artifacts”, obvious problems  
2. Boxplot and histogram plot of log2 (PM) values for each array 


21 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
– Looking for consistency across arrays, identifying outliers 
3. Proportion present plot 
– Using probe pairs (PM,MM) to test whether gene is “Present” of “Absent” 
– Looking for consistency across arrays, identifying outliers 
4. Plot of Hybrid Controls 
– bioB, bioC, bioD, and cre are present in increasing concentrations on each array 
5. QC stats plot 
– The “qc” function returns an object containing scale-factors, % present, average 
background intensities (values greater than 100 indicates problem) and bioB, bioC, 
bioD, and creX present calls.  It also plots the 3’/5’ ratios of actin and GAPDH (3-
fold or greater discrepancies between arrays indicates a problem) 
6. RNA Degradation plot 
– It is expected that 5’ end will have more degradation than 3’ end. We look for 
consistency across arrays, identifying outliers 
*If an outlier is identified in more than 2 of these tests, the sample should be thrown out. 
3.2 
DRUG ADMINISTRATION 
Agents will be administered per package insert. Please see Appendix B for a summary of FDA 
approved doses. Alternate dosing or frequencies of the listed agents may also be utilized provided 
that phase 1 data are available. Agents will be administered until whichever occurs first among 
the following: disease progression or toxicity which requires discontinuation of study drug. 
Subjects who have a response to COXEN-based therapy lasting 6 months or greater have the option 
to be re-assigned to a second COXEN-based agent. If there is not an established combination (with 
at least phase 1 data for safety), only one drug would be given. No more than 2 COXEN-based 
assignments will be allowed on study. 
3.3 
DOSE MODIFICATIONS 
Dose modifications will be guided by package inserts and consultation between the investigator 
and NIH CC Pharmacy. 
If a drug must be permanently discontinued due to toxicity, an alternative COXEN based regimen 
may be selected per PI discretion. Only one change in regimen will be made. At the second 
occurrence of intolerable toxicity in a patient, the patient must be removed from study therapy.    


22 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
3.4 
STUDY CALENDARS 
All drugs have different schedules; we will try (as much as possible) to establish an every 2 week (28 day cycle) or every 3 week (21-
day cycles) clinic visits and infusions when appropriate. 
Unless otherwise specified, screening evaluations may be performed within 28 days prior to initiation of study therapy, baseline 
assessments may be performed within 16 days prior to initiation of study therapy, day 1 assessments and beyond may be performed 
within 1 week prior to indicated time.  
3.4.1 For subjects seen every 2 weeks (28 day cycle) 
Procedure 
Screening/ 
Baseline 
Cycle 1  
(28 days) 
Subsequent Cycles 
Post Therapy Follow-up 
Day 1 
Day 15 
Day 1 
Day 15 
Safety Visit (4-5 
weeks post 
treatment) 
Long-
term 
follow up 
History and PE 
X 
X 
X 
X 
X 
X 
 
Vital signs 
X 
X 
X 
X 
X 
 
 
Performance Score 
X 
X 
X 
X 
X 
 
 
Laboratory 
assessments1  
X 
X 
X 
X 
X 
X 
 
Urinalysis 
X 
 
 
 
 
 
 
TSH, PT/INR, PTT 
X 
 
 
 
 
 
 
24 hour urine 
collection if needed 
X 
 
 
 
 
 
 
Anti HIV 1/2, HIV 
viral load, CD4 
X 
 
 
 
 
 
 
Urine or serum HCG in 
women of childbearing 
potential 
X 
 
 
 
 
 
 
Confirmation of 
diagnosis 
X 
 
 
 
 
 
 
Biopsies2 
X 
 
 
X 
 
 
 
COXEN assessment 
X 
 
 
 
 
 
 


23 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
Procedure 
Screening/ 
Baseline 
Cycle 1  
(28 days) 
Subsequent Cycles 
Post Therapy Follow-up 
Day 1 
Day 15 
Day 1 
Day 15 
Safety Visit (4-5 
weeks post 
treatment) 
Long-
term 
follow up 
Correlative studies3 
X 
Refer to Section 5.1 and Appendix C 
 
 
CT CAP or MRI4 
X 
Every 2 cycles 
 
 
FDG-PET MRI4 
X 
 
 
C3 only 
 
 
 
EKG 
X 
 
 
 
 
 
 
Adverse Events 
 
X 
 
 
 
 
 
Concomitant 
Medications 
 
X 
 
 
 
 
 
Phone calls to assess 
survival status5 
 
 
 
 
 
 
X 
1 Laboratory evaluations include: For screening see section 2.2; for baseline and on study evaluations, see section 2.4. For follow up evaluations, see section 
3.5.2.1 
2 Screening/Baseline biopsy is mandatory. Optional biopsy scheduled with restaging scans, but only performed at the first occurrence of progressive disease. 
3 Refer to Section 5.1 and Appendix C for further information.  
4 Imaging assessments may be performed within +/- 1 week of indicated time. 
5 Patients will be followed for survival with telephone calls every 2 months until death. 


24 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
3.4.2 For subjects seen every 3 weeks (21 day cycle) 
Procedure 
Screening/ 
Baseline 
Cycle 1  
(21 
days) 
Subsequent 
Cycles 
Post Therapy Follow-up 
Day 1 
Day 1 
Safety Visit (4 – 5 
weeks post rx) 
Long-term 
follow up 
History and PE 
X 
X 
X 
X 
 
Vital signs 
X 
X 
X 
 
 
Performance Score 
X 
X 
X 
 
 
Laboratory assessements1  
X 
X 
X 
X 
 
Urinalysis 
X 
 
 
 
 
TSH, PT/INR, PTT 
X 
 
 
 
 
24 hour urine collection if needed 
X 
 
 
 
 
Anti HIV 1/2, HIV viral load, CD4 
X 
 
 
 
 
Urine or serum HCG in women of 
childbearing potential 
X 
 
 
 
 
Confirmation of diagnosis 
X 
 
 
 
 
Biopsies2 
X 
 
X 
 
 
COXEN assessment 
X 
 
 
 
 
Correlative studies3 
 
Refer to Section 5.1 and 
Appendix C 
 
 
CT CAP or MRI4 
X 
Every 3 cycles 
 
 
FDG-PET MRI4 
X 
 
C4 only  
 
 
EKG 
X 
 
 
 
 
Adverse Events 
 
X 
 
 
 
Concomitant Medications 
 
X 
 
 
 
Phone call to assess survival status5 
 
 
 
 
X 
1 Laboratory evaluations include: For screening see section 2.2; for baseline and on study evaluations, see section 2.4. For follow up evaluations, see section 3.5.2.1  
2 Screening/Baseline biopsy is mandatory. Optional biopsy scheduled with restaging scans, but only performed at the first occurrence of progressive disease. 
3 Refer to Section 5.1 and Appendix C for further information. 
4 Imaging assessments may be performed within +/- 1 week of indicated time. 
5 Patients will be followed for survival with telephone calls every 2 months until death.


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Abbreviated Title: COXEN study 
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3.5 
CRITERIA FOR REMOVAL FROM PROTOCOL THERAPY AND OFF STUDY CRITERIA 
Prior to removal from study, effort must be made to have all subjects complete a safety visit 
approximately 30 days following the last dose of study therapy. 
3.5.1 Criteria for removal from protocol therapy 
x Progressive disease after 2 COXEN based regimens 
x Participant requests to be withdrawn from active therapy 
x Unacceptable Toxicity as defined in section 3.3 
x Investigator discretion 
3.5.2 Follow Up 
3.5.2.1 Safety visit 
Patients will be asked to return to NIH, 4-5 weeks after receiving the last dose of study medication 
for a safety assessment that will include a history, physical examination, CBC, acute care panel, 
mineral panel and LFTS. Those patients that cannot return to the NIH will be contacted by 
telephone. Patients removed from study therapy for unacceptable adverse event(s) will be followed 
until resolution or stabilization of the adverse event. 
3.5.2.2 Long term follow up 
Patients will be followed until death. The Investigator or designees will make every possible 
attempt every 2 months (±7 days) after the final safety visit to contact the patient or family to 
obtain the survival information of the patient and start date of additional anticancer treatment. 
3.5.3 Off-Study Criteria 
x Participant requests to be withdrawn from study 
x Death 
3.5.4 Off Protocol Therapy and Off-Study Procedure 
Authorized staff must notify Central Registration Office (CRO) when a subject is taken off 
protocol therapy and when a subject is taken off-study. A Participant Status Updates Form from 
the web site (http://home.ccr.cancer.gov/intra/eligibility/welcome.htm) main page must be 
completed and sent via encrypted email to: NCI Central Registration Office ncicentralregistration-
l@mail.nih.gov. 
4 
CONCOMITANT MEDICATIONS/MEASURES 
Supportive care will be administered per NIH CC Pharmacy guidelines.  
All concomitant medications will be recorded. Medications will be prohibited if contraindicated in 
the package insert. Other medications may be administered at the discretion of the investigators 
with the guidance of the package insert. 


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5 
CORRELATIVE STUDIES 
5.1 
BIOSPECIMEN COLLECTION 
5.1.1 Mutational landscape of bladder cancer before and after initial therapy 
5.1.1.1 Overview and Hypothesis 
Exome sequencing has opened the door for cost efficient, high-throughput, and accurate mutation 
calling by limiting the search space to the small fraction of the genome (~2%) that encodes for 
proteins. With 340 treatment naïve bladder cancer tumors already sequenced, we have a 
compendium of catalogued mutations for which we can compare new samples. Here we will 
evaluate the catalogued mutations on exome sequences from treatment naïve FFPE patient samples 
to both verify if common mutations occur in the patient and also determine if the detection of 
mutations is different between exome sequenced fresh frozen and FFPE tumor tissue samples. In 
addition, we will also compare the same patient’s treatment naïve FFPE primary tumor samples to 
an FFPE biopsy taken after initial therapy but before “best next therapy” to test the hypothesis that 
certain mutations will be found at a higher frequency in treated patients suggesting their possible 
importance in drug resistance.  
5.1.1.2 Approach 
5.1.1.2.1 Tissue Samples 
Patients will undergo a percutaneous core needle biopsy of the tumor under local anesthesia. These 
percutaneous biopsies will be performed by interventional radiology (CT scan or ultrasound 
guidance). If needed, patients will be offered conscious sedation for the biopsy procedure. Sample 
collection will be performed according to standard operating procedures. 2-3 core needle biopsies 
will be obtained and 2-32 samples will be formalin fixed and sent to the Laboratory of Pathology 
in the NCI. The remaining samples will be immediately placed in cryovials, snap frozen on liquid 
nitrogen or dry ice and transported to Dr. Donald Bottaro’s laboratory (See Appendix C). 
Archival samples will be labeled and reported as sample 1 – formalin fixed paraffin embedded 
(FFPE) sample; fresh tissue biopsies will be labeled and reported as sample 2 – fresh baseline 
biopsy or sample 5 – biopsy at time of progression (optional). 
Mutation Detection: Here we will perform either directed exome sequencing (DES) from 
formalin fixed paraffin embedded (FFPE) tissues for 476 mutated genes described above or whole 
exome sequencing (WES) using Illumina HiSeq machines and other technologies as needed.  
Tissues: Tissues from patients will be collected for the specific analysis as described below in 
detail. Matched normal tissues (Buffy coat/WBC) will also be collected for SNP verification.  
5.1.1.2.2 Application of Next Generation Sequencing (NGS) on FFPE Tissues 
Recent prior work has demonstrated the feasibility and robustness of FFPE exome sequencing. 
One study describes massively parallel DNA sequencing that characterizes base substitutions, 
short insertions and deletions (indels), copy number alterations and selected fusions across 287 
cancer-related genes from FFPE clinical specimens [33]. Validation of this approach using pooled 
cell lines that model key determinants of accuracy, including mutant allele frequency, indel length 
and amplitude of copy change indicated a test sensitivity of 95-99% across alteration types with 


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>99% positive predictive value. The ability to carry out reliable exome sequencing in FFPE was 
confirmed by others [34]. There is less clarity on the issue of whether the same mutations are found 
in identical tissue samples preserved by FFPE and frozen methodologies for the purposes of 
discovery but this appears to be the case. A small study compared the validity of whole exome 
sequencing in one primary fresh-frozen sample, corresponding FFPE prostate cancer sample and 
matched adjacent normal prostatic tissue. The sequenced reads were mapped and compared and 
showed comparable (but not identical) exome sequencing results between FFPE and corresponding 
fresh-frozen cancer tissues in one sample [35]. 
5.1.1.2.3 DNA extraction from FFPE Tissues 
There are several challenges that are present when preparing FFPE versus frozen genomic DNA, 
all of which can be overcome to provide high quality sequence information. The first step will be 
to extract the sample from its paraffin embedding. For the FFPE samples from the metastasis 
biopsy volume of 17.7mm3, we expect between 3.5-5μg of isolated DNA. The second challenge 
with FFPE samples is the general poor quality of the DNA extracted due to cross-linking of protein 
to DNA, separating the DNA from paraffin and DNA degradation from the embedding process, 
which includes time to embedding, temperature, and buffering of the paraffin itself. These issues 
have been worked out with gel separation techniques and proper buffering of DNA extraction 
solutions. Standard protocols are now available from peer-reviewed publications such as the two 
cited above [33, 34]. Furthermore, we have successfully used the FFPE DNA Extraction Kit 
(Diagenode) and will use this kit for all FFPE gDNA extractions.  
5.1.1.2.4 Library Prep and Exome Sequencing 
Isolated DNA from FFPE samples will be sent to the Meltzer lab for library preparation and exome 
sequencing (pmeltzer@mail.nih.gov – 240-760-7376). We will be following the SureSelect 
Human All Exon Capture Kit (V5, Agilent Technologies) protocols for library preparation, 
hybridizations, and exome capture. Sequencing will be done using the Illumina HiSeq 2500, 
providing roughly 46Gbp per lane of a flow cell run in the 2X125 mode. The exome represents 
roughly 30Mbp, thus by multiplexing 8 samples per lane, we can produce roughly 150X coverage 
of the exome.  
Sequence reads will be aligned using GSNAP [36] to produce .bam files (binarized sequence 
alignment/map files). We will use the GATK platform [37] to make SNP and indel calls, thus 
producing .vcf files (variant call format). We will finally use AnnoVar [38] to annotate the variant 
calls. FFPE samples require two additional corrections to be directly compared. First, we must 
correct for the allelic fraction to account for the purity or the admixture of the sample. By plotting 
the frequency of alleles between the two samples, we can model the difference allele frequencies 
by fitting a simple regression and thus correct of the any bias between samples. Secondly, we must 
correct for the clonality of the cells within the population. That is, does the mutation occur in all 
of the cells, thus clonal, or does a mutation arise in a subpopulation of cells, thus being subclonal. 
We will make this correction using the program ABSOLUTE [39]. After these two corrections 
have been made we can directly compare the mutations between the FFPE samples. 
5.1.1.3 Sample Comparisons 
Several analyses will assess relationships between mutation frequency and preservation method of 
the specimen, tumor heterogeneity, tumor progression and prior treatment.  


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Frozen vs. FFPE (DES): The 476 mutated genes will be evaluated on the 20 patients enrolled in 
the study on their treatment naïve tumor sample (original specimen-Specimen 1, Figure 4). The 
frequency of the mutations will be compared in a stage and grade matched fashion (to eliminate 
the effect of stage/grade on tumor mutation frequency which we know exist [40, 41]) to determine 
if there are any differences in mutation frequency (i.e. detection) as a function of preservation 
method and if this is gene/mutation dependent. 
Tumor Heterogeneity (WES): Three replicate biopsy samples will be collected (Samples 2 and 
5, Figure 4) and 3 cores will be taken from the FFPE block (as described by us [31]) of the original 
specimen (Specimen 1, Figure 4). For each specimen source, an analysis will be carried to 
determine the changes in mutation frequency as a function of tumor heterogeneity. 
Tumor Progression and Initial Treatment (WES): We assess the relationship of mutation 
frequency as a function of progression (primary vs metastasis) and initial treatment. Only one 
comparison will be made: baseline untreated from the primary tumor -Specimen 1, Figure 4) vs. 
metastatic site biopsy (pre “best next treatment”, Specimen 2, Figure 4 and between pre and post 
treatment biopsies (Specimens 2 and 5 in Figure 4 respectively), which will encompass both 
variables (primary vs metastasis and pre vs post treatment). This is a recognized limitation of the 
study but it is nevertheless a clinical reality since we know almost all patients that present will 
NOT have a biopsy of a treatment naïve metastasis, while most will have a treatment naïve tissue 
specimen from the initial specimen. 
5.1.2 Saliva samples for genetic/genomic analysis  
Tumor samples will be used as a source of DNA for genetic/genomic studies. Saliva samples will 
be used as a source of genomic (germline DNA) from the patients to compare with tumor DNA 
for somatic mutations. Whole genome and whole exome studies may be conducted using the 
material collected. Studies for the analysis of chromosomal abnormalities may also be performed. 
We plan to do genomic analysis of the saliva samples by both whole-genome and whole-exome 
sequencing. We will confirm recurrent mutations in genes previously identified as being mutated 
in urothelial carcinoma and we will identify additional altered genes and pathways that have been 
implicated in this disease and compare the responders and non-responders. 
See Appendix C for details regarding saliva collection and handling  
5.1.3 Mutational landscape of circulating tumor DNA before and after “best next therapy” 
5.1.3.1 Overview and Hypothesis 
There are currently no peripheral blood-based markers available to provide an early indication of 
response to systemic therapy in patients with metastatic bladder cancer. We will test the hypothesis 
that circulating tumor DNA (ctDNA) with cancer specific mutations will be present in all patients 
before “best next therapy”. We will also examine 1) the molecular similarity of the “liquid biopsy” 
(ctDNA) to the contemporaneous tumor biopsy; 2) The change in mutation profile of the liquid 
biopsy following the “best next therapy” described in Aim 1 which may provide clues to the 
mechanisms responsible for the observed clinical response. 
5.1.3.2 Sample Collection and Handling 
Patients enrolled in the trial will undergo collection of a blood sample for evaluation of changes 
in mutation profile, as detailed in Appendix C.  


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5.1.3.3 Approach 
The ctDNA will be evaluated using well described approaches that we [42] and other have recently 
described [43, 44]. The studies will be performed in the laboratory of Dr. Paul Meltzer. 
5.1.3.3.1 ctDNA isolation   
ctDNA will be extracted using the QIAmp Circulating Nucleic Acid Kit (Qiagen). It is estimated 
that there is between 1-100ng/ml ctDNA in blood plasma. Given the 6-10ml of plasma samples, 
we estimate the total ctDNA yield to be 6ng-1μg. To determine if somatic mutations or copy 
number alterations have arisen in patient tumors, we will also extract gDNA from white blood 
cells, which can be done using the DNAeasy Blood & Tissue Kit (Qiagen).  
5.1.3.3.2 Exome Sequencing 
Isolated DNA samples will be sent to the Meltzer Lab for library preparation and exome 
sequencing. We will be following the SureSelect Human All Exon Capture Kit (V5, Agilent 
Technologies) protocols for library preparation, hybridizations, and exome capture. Sequencing 
will be done using the Illumina HiSeq 2500, providing roughly 46Gbp per lane of a flow cell run 
in the 2X125 mode. We expect to find only a small fraction of the full exome within ctDNA but 
use the full 30Mbp for calculating coverage. The exome represents roughly 30Mbp, thus by 
multiplexing 8 samples per lane, we can produce roughly 150X coverage of the exome. To avoid 
technical differences in sequencing, we will match multiplexed ctDNA with exome white blood 
cell exomes on the same flow cell. We will follow the same sequence analysis methods as outlined 
in Aim 2. Sequence reads will be aligned using GSNAP to produce bam files (binarized sequence 
alignment/map files). We will use the GATK platform to make SNP and indel calls, thus producing 
vcf files (variant call format). We will finally use AnnoVar to annotate the variant calls.  
5.1.3.3.3 Sample Comparisons and Patient Outcome Analyses 
Two analyses will assess relationships between mutation frequency and prior treatment and 
mutation frequency and patient outcome. 
Figure 5: Blood sample processing  


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Sensitivity of in ctDNA Tumor Mutation to Detect Metastasis (DES): ctDNA will be obtained 
at baseline. Since all these patients have metastatic disease detectable by imaging, we expect the 
ctDNA to harbor detectable mutations. This will be carried out on a smaller subset (to enhance the 
feasibility of detection by using multiple methods) [42] of the 476 mutated genes mentioned above 
with the addition of any new genes found in Aim 2 as a function of progression and treatment. 
Tumor Mutation in ctDNA vs. Mutation in Biopsy (DES): ctDNA and contemporaneous biopsy 
mutations will be compared (Specimen 2 vs. 3, Figure 4). This will be carried out on a smaller 
subset (to enhance the feasibility of detection by using multiple methods) [42] of the 476 mutated 
genes mentioned above with the addition of any new genes found in Aim 2 as a function of 
progression and treatment. 
Tumor Mutation vs “best next therapy” (WES): We assess the relationship of mutation 
frequency as a function of treatment. Only one comparison will be made: baseline pretrial blood 
(Specimen 3, Figure 4) vs. post-treatment (Specimen 4, Figure 4). We realize that it is possible 
that all 20 patients will be treated with a different drug. However, we will still be able to answer 
the question if some treatments alter the genomic composition of the ctDNA, a question that has 
not yet been answered in bladder cancer and a stepping stone to larger and more directed studies.  
Tumor Mutation vs Patient Outcome (WES): As above, the ctDNA will be obtained at baseline 
and 8 or 9 weeks post “best next therapy”. Cox proportional hazards model will be used to 
determine the association of overall survival and progression-free survival with various ctDNA 
findings (presence or absence of detectable mutations, type of mutations/gene families in baseline, 
type of mutations/gene families in post treatment sample, presence of new mutations as a function 
of treatment vs no new mutations arising after treatment). For all survival analyses, the time to 
death due to bladder cancer will be defined as the time between the date when the baseline sample 
of blood was obtained and the date of death. Survival curves will be compared using log-rank 
testing. This will be an exploratory analysis based on the limited number of patients that will be 
enrolled on the study.  
5.1.4 Immune Subset Analysis  
Peripheral blood mononuclear cells (PBMC) will be assessed using multiparameter flow cytometry 
for immune subsets including but not necessarily limited to CD8+ T-cells, CD4+Foxp3- T-cells, 
Tregs, monocyte subsets, MDSC subsets. Assessment will include functional markers, i.e. PD-1, 
Tim-3, CTLA-4, PD-L1, HLA-DR and/or CD40.  
See Appendix C for details regarding blood collection and handling.  
Members of the Trepel lab will enter the samples in a secure patient database, process the samples 
for viable cell storage, label each sample with a unique 2D barcode, and viably store the samples. 
They will prepare the samples for staining, stain and run the samples by multiparametric flow 
cytometry (MACSQuant, Miltenyi Biotec, Bergisch Gladbach, DE), and analyze the data using 
FlowJo (FlowJo LLC, Ashland, OR) software.   
5.1.5 Peripheral Blood Immune Gene Expression  
Peripheral blood immune gene expression will be evaluated by the Trepel Lab using the 
NanoString nCounter® platform (NanoString Technologies, Seattle, WA). RNA will be isolated 
using the PAXgene Blood RNA Kit according to the manufacturer’s instructions. Peripheral 
immune gene expression will be evaluated using the PanCancer Immune Profiling Panel of 730 


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immune genes and 40 control genes at baseline and post-therapy to look for correlates of clinical 
response with innate or adaptive immunity. 
Contact the Trepel Lab, as detailed in Section 5.1.4 and in Appendix C:  Sample Collection for 
Correlative Studies.  
5.1.6 Pro-inflammatory cytokines  
Plasma levels of a comprehensive panel of pro-inflammatry molecules will be tested, including 
IFN-γ, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, and TNF-α (inflammatory cytokines 
panel III). Many of the markers can be produced by activated T cells. The analysis will be 
conducted with MSD V-PLEX technology that provides the best assays for cytokine analysis.  
See Appendix C:  Sample Collection for Correlative Studies for details regarding collection 
and handling of specimens 
For samples collected by outside institutions: 
Please notify the lab by email (NCIBloodcore@mail.nih.gov ) when the sample is being shipped 
(Include the Fed Ex tracking number in the email) 
Send samples by FedEx Priority Overnight Shipping. The NIH is not open on weekends or 
government holidays. Be careful not to ship any samples to arrive on a Saturday, Sunday or 
government holiday. We will send the site a government holiday calendar. If you are not certain 
whether the government will be open, please send an email to the addresses above at any time and 
we will respond as quickly as possible. 
Ship Monday-Thursday only to: 
William Figg Laboratory 
9000 Rockville Pike 
Bldg. 10/Room 4B11 
Bethesda, MD 20892 
Phone: 240-760-6180 
5.1.6.1 Laboratory test and data analysis  
All plasma samples will be transferred to Dr. Liang Cao’s laboratory (at Bldg. 37/Rm6134, phone: 
301-435-9039) for the analysis of angiogenesis markers and pro-inflammatory cytokines. 
5.1.7 Plasma HGF and MET  
Plasma will be collected to determine whether urinary hepatocyte growth factor (HGF), urinary 
soluble MET receptor (sMet), plasma HGF and plasma Met levels are biomarkers of bladder 
cancer (transitional cell carcinoma; TCC) and/or response to systemic treatment with the 
combination of cabozantinib and nivolumab. Blood samples for correlatives may be drawn ± 2 
days of specified collection time point. 
Electrochemiluminescence immunoassays for c-Met and HGF. We use Streptavidin-coated 96-
well plates (MSD, Gaithersburg, MD) designed specifically for use in a Meso Scale Discovery 
Sector 2400 Imager. For both assays, a biotin-tagged, affinity-purified specific capture antibody 
either R&D BAF 358 or R&D MAB 694 is added to each well and incubated for 1 hour at room 
temperature (RT) with shaking. Sample wells are then washed with PBS before adding standards 


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and patient plasma / urine samples; diluted standards are used for c-Met (R&D 358-MT-100) and 
HGF (R&D 294-HG recombinant human HGF) to generate a curve. Plates are then incubated for 
1 hour at RT while shaking. Wells are then washed with PBS before adding detection antibody 
(R&D AF 276 with MSD Sulfotag for c-Met), (R&D AF-294 labeled with MSD Sulfotag for HGF) 
for 1 hour at RT with shaking. Finally, plate wells are washed four times with PBS before adding 
MSD Read Buffer T and then read immediately in a MSD Sector 2400 Imager. All samples and 
standards are measured in duplicate. Mean values from a standard curve are constructed by plotting 
signal intensity against standard concentration. A nonlinear regression curve fitting algorithm 
using GraphPad Prism is used to generate an equation from which sample values for c-Met or HGF 
concentration are calculated from mean signal intensity values. 
x Blood samples should be collected prior to any surgery or drug treatment in the presence 
of citrate or EDTA as anticoagulant. Assuming 50% of the whole blood volume will be 
obtained as plasma, the blood volume required for sMet analysis is 1 ml per sample; the 
blood volume required for HGF analysis is 4 mL. 
x Blood samples should be centrifuged within 30 min at 1,300 rcf at room temperature for 
10 minutes. Do not use braking to stop the centrifuge. This will give three layers: (from 
top to bottom) plasma, leucocytes (buffy coat) and erythrocytes.  
x Carefully aspirate the supernatant (plasma) at room temperature and transfer to a new 
centrifuge tube. Take care not to disrupt the cell layer or transfer any cells.  
x Inspect the plasma for turbidity. Turbid samples should be centrifuged and the clarified 
supernatant should be transferred to cryovials and stored at –80°c.  Ensure that the cryovials 
are properly labeled. 
Plasma samples will be stored by Dr. Figg’s laboratory and processed and analyzed by Dr. 
Bottaro’s laboratory. See Appendix C:  Sample Collection for Correlative Studies for details 
regarding collection and handling of specimens. 
5.1.8 Circulating tumor cells (CTC)  
Peripheral blood will be collected to correlate changes in circulating tumor cells with clinical 
response.  CTCs will be assessed using ferrofluidic enrichment and multi-parameter flow 
cytometric detection. CTCs are identified by positive expression of epithelial markers and a 
viability marker and negative expression of hematopoietic markers.  
See Section 5.1.4 and Appendix C:  Sample Collection for Correlative Studies for details 
regarding collection and handling of specimens. 
5.1.9 Circulating tumor cells (Epic Science)  
For details regarding collection and handling of blood samples to be sent to Epic Science, see 
Appendix C:  Sample Collection for Correlative Studies. 
IMPORTANT: The first 5 mL of blood collected from the fresh venipuncture cannot be used for 
the collection into the Streck tubes due to possibility of contaminating epithelial cells during 
venipuncture. Please ensure that at least one blood tube of 5 mL or more is collected prior to 
collection of the CTC sample to avoid adversely affecting the test results.  
Prevention of Backflow: Since Streck Cell-Free DNA BCT tubes contain chemical additives, it is 
important to avoid possible backflow from the tube. To guard against backflow, observe the 
following precautions:  


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x Keep patient’s arm in the downward position during the collection procedure.  
x Hold the tube with the stopper uppermost.  
x Release tourniquet once the blood starts to flow into the tube, or within 2 minutes of 
application.  
x Tube contents should not touch stopper or the end of the needle during the collection 
procedure.  
5.1.9.1 Specimen Shipment Logistics  
All shipments must include requisition forms that contain Patient ID, Collection Date and time, 
Collection Site (including address), Time Point (if applicable), and the appropriate trial code 
(Partner Protocol # / Epic Internal ID). Requisition forms for samples being shipped to LabCorp 
must also contain the appropriate LSN (Accessioning) code listed in the table below. Partner will 
provide a copy of the requisition form to Epic for review and approval prior to trial initiation. 
Clinical sites should provide email notification of sample shipment to Epic Sciences on the day of 
collection, using the applicable email address specified in the table below. The email should 
contain:  
x Trial codes (Partner Protocol # / Epic Internal ID)  
x Patient ID  
x Collection date and time*  
x Time point/Visit  
x Tracking information  
If possible, include a scanned copy of the completed sample requisition form. Partner will be 
responsible for all blood collection supplies, shipping materials, and shipping expenses.  
* When collection time is not provided, Epic Sciences will assume the sample was collected at 
8:00AM (local time) on the date of collection. 
 
Sent notification to: partners@epicsciences.com  
 
Ship to: Epic Science 
 
Attn: (Partner Protocol #/Epic internal ID) 
 
9381 Judicial Dr. Suite 200 
 
San Diego, CA 92121 
 
Phone: 1-858-356-6610 
Holiday Outages  
Epic Science will promptly notify the study team via email of any holiday closures . 
5.1.10 Pharmacogenetic Studies  
Blood samples will be collected for pharmacogenetic studies to analyze the genomic DNA and 
assess genotype of the most relevant drug metabolizing enzymes and transporters (DMET). DNA 
will be analyzed on a DMET Plus (Affymetrix) genotyping platform that tests for 1,936 genetic 
variations in 225 drug disposition genes, including 47 CYP (phase I metabolism) genes, 13 non-
CYP (phase I metabolism) genes, 78 phase II metabolizing genes (including UGTs), 63 
transporters, 4 genes involved in facilitation of drug transporters, 9 genes involved in global 
regulation of drug metabolizing/transporting proteins, 4 drug binding proteins, and 4 drug 
targets. 


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Of specific interest to etoposide are polymorphisms in CYP3A4, ABCB1 (P-glycoprotein), and 
UGT1A1, all of which are included in the DMET analysis. 
See Appendix C:  Sample Collection for Correlative Studies for details regarding collection 
and handling of specimens. 
5.2 
FDG-PET MRI 
Whole body Magnetic Resonance Imaging (MRI) combined with diffusion-weighted imaging is 
becoming an established method in staging of cancers and has been proven to be a feasible method 
for detection of small foci of metastases (1). In addition, MRI provides superb soft tissue 
characterization, which is helpful in evaluation of local spread and recurrence of pelvic 
malignancies – such as bladder cancer - where Computed Tomography (CT) can be limited due to 
attenuation and beam hardening, caused by bony frame of the pelvis.  
Positron Emission Tomography (PET), on the other hand, provides sensitive functional data for N 
and M staging of malignancies with poor anatomical details and soft tissue characterization. This 
pitfall of PET imaging led researchers to propose and ultimately develop a combined technique 
comprising of PET and MRI (2). PET-MRI scanners became available relatively recently, and 
several applications of this powerful tool have been investigated in central nervous system and 
body malignancies (3). However, metastatic bladder cancer has not been a focus of evaluation with 
this technique and combined PET-MRI data for staging of bladder cancer is scarce.  
Patients will be scanned at baseline on a combined 3T Siemens Verio MRI –PET system (Erlangen, 
Germany) in the Laboratory of Diagnostic Radiology Research, Clinical Center. The first scan will 
be performed at baseline before chemotherapy, followed by a second scan after the first restaging 
scans (8 or 9 weeks) to assess the treatment response. Patient will arrive at the nuclear medicine 
department and will receive an injection of radiotracer –18F Fludeoxyglucose (FDG) – via an 
intravenous catheter. Dose of FDG will be calculated based on patient’s body weight. Patient will 
be encouraged to be hydrated during the uptake phase of 60 mins. Afterwards, patient will be 
brought to MRI-PET suite to undergo whole body MRI-PET, which will be performed using 
several predetermined pulse sequences including; diffusion weighted imaging, conventional T1 
and T2 weighted sequences, and dynamic post contrast.  Patient will receive a single injection of 
0.1 mmol/kg of gadolinium-based contrast via a previously placed intravenous line. We expect the 
entire image acquisition - from neck down to mid thigh - to be accomplished in less than 1 hour.  
5.3 
SAMPLE STORAGE, TRACKING AND DISPOSITION 
Samples will be ordered in CRIS and tracked through a Clinical Trial Data Management system. 
Should a CRIS screen not be available, the CRIS downtime procedures will be followed.  Samples 
will not be sent outside NIH without IRB notification and an executed MTA.  
5.3.1 Laboratory of Paul Meltzer 
All samples will be stored in monitored freezers/refrigerators in the Meltzer laboratory at specified 
temperatures with alarm systems in place. All samples will be tracked by unique sample identifiers 
with bar code labeling, using the Labmatrix software and the Genologic Clarity LIMS system. 
Secure computer database systems (Labmatrix and Clarity LIMS) will be used to track all samples 
collected on this protocol.  The system will contain data that includes, but is not limited to the 
unique sample identifiers, storage locations and conditions, biologic study results, clinical 


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information, and corresponding records of all derivatives generated from samples/tissues collected 
on this protocol.  The system will employ mechanisms for restricting users to viewing only the 
level of data appropriate for each individual user, will provide the capability to audit any data 
modification, and will be maintained and backed up according to established standards. 
5.3.2 Laboratory of Jane Trepel  
Samples will be processed immediately by the Trepel laboratory. Biospecimens will be collected 
and processed using validated SOPs that will ensure both specimen quality and patient 
confidentiality. Using a computerized inventory system and a backup hardcopy process, all 
specimen collection and processing steps will be documented and the specific location of each 
specimen will be tracked. Each new specimen collected will be assigned a unique barcode 
identifier that can be linked to the original specimen collected and other relevant information 
within the inventory system. Specimen labels will indicate: protocol number, order in which the 
patient enrolled on the trial, type of sample, collection time, and total volume collected, as 
appropriate. The inventory process contains other security provisions sufficient to safeguard 
patient privacy and confidentiality. Access to the inventory system and associated documents will 
be restricted to appropriate individuals. Requests to use specimens stored in the repository must 
be approved. SOPs ensure that any changes in informed consent made by a patient and relayed to 
the PI will be reflected in the inventory system to ensure that specimens are destroyed as 
appropriate. All laboratory personnel will be trained to adhere to SOPs and will be monitored for 
high-quality performance. 
5.3.3 Clinical Pharmacology Program 
All samples sent to the Blood Processing Core (BPC) will be bar-coded, with data entered and 
stored in the LABrador (aka LabSamples) utilized by the CPP. This is a secure program, with 
access to LABrador limited to defined CPP personnel, who are issued individual user accounts. 
Installation of LABrador is limited to computers specified by Dr. Figg. These computers all have 
a password restricted login screen. All Figg lab personnel with access to patient information 
annually complete the NIH online Protection of Human Subjects course.  
The program creates a unique barcode ID for every sample and sample box, which cannot be traced 
back to patients without LABrador access. The data recorded for each sample includes the patient 
ID, name, trial name/protocol number, time drawn, cycle time point, dose, material type, as well 
as box and freezer location. Patient demographics associated with the clinical center patient 
number are provided in the system. For each sample, there are notes associated with the processing 
method (e.g. delay in sample processing, storage conditions on the ward, etc.). 
Bar-coded samples are stored in bar-coded boxes in locked freezers at either -20 C or -80 C 
according to stability requirements. These freezers are located onsite in the CPP and offsite at NCI 
Frederick Central Repository Services in Frederick, MD. Visitors to the laboratory are required to 
be accompanied by laboratory staff at all times. 
Access to stored clinical samples is restricted. Samples will be stored until requested by a 
researcher named on the protocol. All requests are monitored and tracked in LABrador. All 
researchers are required to sign a form stating that the samples are only to be used for research 
purposes associated with this trial (as per IRB approved protocol) and that any unused samples 
must be returned to the CPP. It is the responsibility of the NCI Principal Investigator to ensure that 
the samples requested are being used in a manner consistent with IRB approval. 


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Following completion of this study, samples will remain in storage as detailed above. Access to 
these samples will only be granted following IRB approval of an additional protocol, granting the 
rights to use the material. 
Sample bar-codes are linked to patient demographics and limited clinical information. This 
information will only be provided to investigators listed on this protocol, via registered use of the 
LABrador. It is critical that the sample remains linked to patient information such as race, age, 
dates of diagnosis and death, and histological information about the tumor, in order to correlate 
genotype with these variables. 
5.3.4 Sample Destruction, End of Protocol Procedures 
All specimens obtained in the protocol are used as defined in the protocol. Any specimens that are 
remaining at the completion of the protocol will be stored in the conditions described below. The 
study will remain open so long as sample or data analysis continues. Samples from consenting 
subjects will be stored until they are no longer of scientific value or if a subject withdraws consent 
for their continued use, at which time they will be destroyed. The PI will report any loss or 
destruction of samples to the NIH Intramural IRB as soon as he is made aware of such loss. 
If the patient withdraws consent the participants data will be excluded from future distributions, 
but data that have already been distributed for approved research use will not be able to be 
retrieved. 
The PI will report destroyed samples to the IRB if samples become unsalvageable because of 
environmental factors (ex. broken freezer or lack of dry ice in a shipping container) or if a patient 
withdraws consent. Samples will also be reported as lost if they are lost in transit between facilities 
or misplaced by a researcher. Freezer problems, lost samples or other problems associated with 
samples will also be reported to the IRB, the NCI Clinical Director, and the office of the CCR, 
NCI. 
5.4 
SAMPLES FOR GENETIC/GENOMIC ANALYSIS 
5.4.1 Description of the scope of genetic/genomic analysis 
The genetic studies are discussed in detail in sections 5.1.1 and 5.1.2. They will be performed on 
tumor tissue as well as blood and will include directed exome sequencing of 476 genes and/or 
whole exome sequencing.  
5.4.2 Description of how privacy and confidentiality of medical information/biological 
specimens will be maximized 
Confidentiality for genetic samples will be maintained as described in section 5.3.1.  
5.4.3 Management of Results 
Subjects will be contacted if a clinically actionable gene variant is discovered. Clinically 
actionable findings for the purpose of this study are defined as disorders appearing in the American 
College of Medical Genetics and Genomics recommendations for the return of incidental findings 
that is current at the time of primary analysis. A list of current guidelines is maintained on the CCR 
intranet:  
https://ccrod.cancer.gov/confluence/display/CCRCRO/Incidental+Findings+Lists). Subjects who 
still remain on the study will be contacted at this time with a request to provide a blood sample to 


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be sent to a CLIA certified laboratory within the NIH or as a send-out test to Mayo Clinic. If the 
research findings are verified in the CLIA certified lab, the subject will be referred to a genetic 
healthcare provider within the NIH for the disclosure of the results.  
This is the only time during the course of the study that incidental findings will be returned. No 
interrogations regarding clinically actionable findings will be made after the primary analysis. 
6 
DATA COLLECTION AND EVALUATION  
6.1 
DATA COLLECTION 
C3D and LabMatrix will be used for data collection purposes on this study. All data will be kept 
secure. The PI will be responsible for overseeing entry of data into an in-house password protected 
electronic system and ensuring data accuracy, consistency and timeliness. The principal 
investigator, associate investigators/research nurses and/or a contracted data manager will assist 
with the data management efforts. All data obtained during the conduct of the protocol will be kept 
in secure network drives or in approved alternative sites that comply with NIH security standards. 
Primary and final analyzed data will have identifiers so that research data can be attributed to an 
individual human subject participant. 
An abnormal laboratory value will be considered an AE if the laboratory abnormality is 
characterized by any of the following: 
x 
Results in discontinuation from the study 
x 
Is associated with clinical signs or symptoms  
x 
Requires treatment or any other therapeutic intervention 
x 
Is associated with death or another serious adverse event, including hospitalization.  
x 
Is judged by the Investigator to be of significant clinical impact 
x 
If any abnormal laboratory result is considered clinically significant, the investigator will 
provide details about the action taken with respect to the test drug and about the patient’s 
outcome. 
End of study procedures: Data will be stored according to HHS, FDA regulations and NIH 
Intramural Records Retention Schedule as applicable.  
Loss or destruction of data: Should we become aware that a major breach in our plan to protect 
subject confidentiality and trial data has occurred, the IRB will be notified.  
As all agents on the study are FDA approved, only the following events will be recorded: 
x All Grade 2 unexpected events that are possibly, probably or definitely related to the 
research;  
x All Grade 3 and 4 events that are possibly, probably or definitely related to the research;  
x All Grade 5 events regardless of attribution;  
x All Serious Events regardless of attribution.  


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6.2 
DATA SHARING PLANS 
6.2.1 Human Data Sharing Plan 
What data will be shared? 
I will share human data generated in this research for future research as follows: 
_X_ Coded/linked data in an NIH-funded or approved public repository.   
_X_ Coded/linked data in BTRIS  
_X_ Identified or coded, linked data with approved outside collaborators under appropriate 
agreements. 
How and where will the data be shared?  
Data will be shared through: 
_X_ An NIH-funded or approved public repository. Insert name or names:_dbGaP, clinical 
trials.gov. 
_X_ BTRIS 
_X_ Approved outside collaborators* under appropriate individual agreements. 
_X_ Publication and/or public presentations. 
* Genomic data will be transferred from Dr. Meltzer’s lab to UCCC via a password protected CEL 
file identified by an accession number and sent through Fileman. 
When will the data be shared? 
_X_ Before publication. 
_X_ At the time of publication or shortly thereafter. 
6.2.2 Genomic Data Sharing Plan 
Unlinked genomic data will be deposited in public genomic databases such as dbGaP in 
compliance with the NIH Genomic Data Sharing Policy. 
6.3 
RESPONSE CRITERIA 
For the purposes of this study, patients should be re-evaluated for response every 8 or 9 weeks 
depending on visit schedule. In addition to a baseline scan, confirmatory scans should also be 
obtained not less than 4 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) [45]. Changes in the largest diameter (unidimensional measurement) of the tumor 
lesions and the shortest diameter in the case of malignant lymph nodes are used in the RECIST 
criteria. 
6.3.1 Definitions 
Evaluable for toxicity: All patients will be evaluable for toxicity from the time of their first 
treatment with any of the agents. 


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Evaluable for objective response: Only those patients who have measurable disease present at 
baseline, have received at least one cycle of therapy, and have had their disease re-evaluated will 
be considered evaluable for response. These patients will have their response classified according 
to the definitions stated below. (Note: Patients who exhibit objective disease progression prior to 
the end of cycle 1 will also be considered evaluable.) 
Evaluable Non-Target Disease Response: Patients who have lesions present at baseline that are 
evaluable but do not meet the definitions of measurable disease, have received at least one cycle 
of therapy, and have had their disease re-evaluated will be considered evaluable for non-target 
disease. The response assessment is based on the presence, absence, or unequivocal progression 
of the lesions.  
6.3.2 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: 
x By chest x-ray: >20 mm 
x By CT scan: 
o Scan slice thickness 5 mm or under: as >10 mm 
x With calipers on clinical exam: >10 mm 
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 


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(longest for non-nodal lesions, short axis for nodal lesions) for all target lesions will be calculated 
and reported as the baseline sum diameters.  If lymph nodes are to be included in the sum, 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.  
6.3.3 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 t10 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 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 


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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 [46-48]. 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 [49]. 
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 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:  
a. Negative FDG-PET at baseline, with a positive FDG-PET at follow-up is a sign of PD 
based on a new lesion. 
b. 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. 
c. 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 


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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. 
6.3.4 Response Criteria 
6.3.4.1 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. 
Partial Response (PR): At least a 30% decrease in the sum of the diameters of target lesions, taking 
as reference the baseline sum of 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 of diameters while on study. 
6.3.4.2 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). 
6.3.4.3 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. 
For Patients with Measurable Disease (i.e., Target Disease) 


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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 
6.3.5 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). 


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The duration of overall CR is measured from the time measurement criteria 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.  
6.3.6 Progression-Free Survival 
PFS is defined as the duration of time from start of treatment to time of progression or death, 
whichever occurs first. 
6.4 
TOXICITY CRITERIA 
The following adverse event management guidelines are intended to ensure the safety of each 
patient while on the study. The descriptions and grading scales found in the revised NCI Common 
Terminology Criteria for Adverse Events (CTCAE) version 4.0 will be utilized for AE reporting. 
All appropriate treatment areas should have access to a copy of the CTCAE version 4.0. A copy 
of 
the 
CTCAE 
version 
4.0 
can 
be 
downloaded 
from 
the 
CTEP 
web 
site 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm#ctc_40).  
7 
SAFETY REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING 
PLAN 
7.1 
DEFINITIONS 
7.1.1 Adverse Event 
Any untoward medical occurrence in a human subject, including any abnormal sign (for example, 
abnormal physical exam or laboratory finding), symptom, or disease, temporally associated with 
the subject’s participation in research, whether or not considered related to the subject’s 
participation in the research.  
7.1.2 Suspected adverse reaction 
Suspected adverse reaction means any adverse event for which there is a reasonable possibility 
that the drug caused the adverse event. For the purposes of IND safety reporting, ‘reasonable 
possibility’ means there is evidence to suggest a causal relationship between the drug and the 
adverse event. A suspected adverse reaction implies a lesser degree of certainty about causality 
than adverse reaction, which means any adverse event caused by a drug. 
7.1.3 Unexpected adverse reaction 
An adverse event or suspected adverse reaction is considered “unexpected” if it is not listed in the 
investigator brochure or is not listed at the specificity or severity that has been observed; or, if an 
investigator brochure is not required or available, is not consistent with the risk information 
described in the general investigational plan or elsewhere in the current application. "Unexpected” 
also refers to adverse events or suspected adverse reactions that are mentioned in the investigator 
brochure as occurring with a class of drugs or as anticipated from the pharmacological properties 
of the drug but are not specifically mentioned as occurring with the particular drug under 
investigation. 


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7.1.4 Serious  
An Unanticipated Problem or Protocol Deviation is serious if it meets the definition of a Serious 
Adverse Event or if it compromises the safety, welfare or rights of subjects or others. 
7.1.5 Serious Adverse Event 
An adverse event or suspected adverse reaction is considered serious if in the view of the 
investigator or the sponsor, it results in any of the following: 
x 
Death, 
x 
A life-threatening adverse drug experience 
x 
Inpatient hospitalization or prolongation of existing hospitalization 
x 
Persistent or significant incapacity or substantial disruption of the ability to conduct normal 
life functions 
x 
A congenital anomaly/birth defect. 
x 
Important medical events that may not result in death, be life-threatening, or require 
hospitalization may be considered a serious adverse drug experience when, based upon 
appropriate medical judgment, they may jeopardize the patient or subject and may require 
medical or surgical intervention to prevent one of the outcomes listed in this definition. 
7.1.6 Disability 
A substantial disruption of a person’s ability to conduct normal life functions. 
7.1.7 Life-threatening adverse drug experience 
Any adverse event or suspected adverse reaction that places the patient or subject, in the view of 
the investigator or sponsor, at immediate risk of death from the reaction as it occurred, i.e., it does 
not include a reaction that had it occurred in a more severe form, might have caused death. 
7.1.8 Protocol Deviation (NIH Definition) 
Any change, divergence, or departure from the IRB-approved research protocol.   
7.1.9 Non-compliance (NIH Definition) 
The failure to comply with applicable NIH Human Research Protections Program (HRPP) policies, 
IRB requirements, or regulatory requirements for the protection of human research subjects. 
7.1.10 Unanticipated Problem 
Any incident, experience, or outcome that: 
x  Is unexpected in terms of nature, severity, or frequency in relation to  
(a) the research risks that are described in the IRB-approved research protocol and 
informed consent document; Investigator’s Brochure or other study documents, and 
(b) the characteristics of the subject population being studied; AND 
x Is related or possibly related to participation in the research; AND 


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x Suggests that the research places subjects or others at a greater risk of harm (including 
physical, psychological, economic, or social harm) than was previously known or 
recognized. 
7.2 
NIH INTRAMURAL IRB AND CLINICAL DIRECTOR REPORTING 
7.2.1 NIH Intramural IRB and NCI Clinical Director Expedited Reporting of Unanticipated 
Problems and Deaths 
The Protocol PI will report in the NIH Problem form to the NIH Intramural IRB and NCI Clinical 
Director: 
x All deaths, except deaths due to progressive disease  
x All Protocol Deviations 
x All Unanticipated Problems 
x All non-compliance 
Reports must be received within 7 days of PI awareness via iRIS. 
7.2.2 NIH Intramural IRB Requirements for PI Reporting at Continuing Review 
The protocol PI will report to the NIH Intramural IRB:  
1. A summary of all protocol deviations in a tabular format to include the date the deviation 
occurred, a brief description of the deviation and any corrective action. 
2. A summary of any instances of non-compliance 
3. A tabular summary of the following adverse events: 
x All Grade 2 unexpected events that are possibly, probably or definitely related to the 
research;  
x All Grade 3 and 4 events that are possibly, probably or definitely related to the research;  
x All Grade 5 events regardless of attribution;  
x All Serious Events regardless of attribution.  
NOTE: Grade 1 events are not required to be reported.  
7.3 
DATA AND SAFETY MONITORING PLAN 
7.3.1 Principal Investigator/Research Team  
The clinical research team will meet on a regular basis when patients are being actively treated on 
the trial to discuss each patient. Decisions about dose level enrollment and dose escalation if 
applicable will be made based on the toxicity data from prior patients.  
All data will be collected in a timely manner and reviewed by the principal investigator or a lead 
associate investigator. Adverse events will be reported as required above. Any safety concerns, 
new information that might affect either the ethical and or scientific conduct of the trial, or protocol 
deviations will be immediately reported to the IRB using iRIS.  
The principal investigator will review adverse event and response data on each patient to ensure 
safety and data accuracy. The principal investigator will personally conduct or supervise the 


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investigation and provide appropriate delegation of responsibilities to other members of the 
research staff.  
8 
STATISTICAL CONSIDERATIONS   
It would be desirable if a fraction consistent with 90% of patients has a successful drug assignment 
by COXEN. To evaluate this in a pilot fashion, a total of 20 patients will undergo the processes 
necessary to undertake the algorithm and the time required to obtain the final treatment assignment 
will be classified as being less than or equal to 21 days or greater than 21 days. A success would 
be a result in 21 days or less. With 20 patients, if 16 or greater are able to have a success, the 
probability that this would occur would be 95.7% if the true probability of a success for a given 
patient were 90% and it would be 5.1% if it were only 60%. Thus, observing 16 or more out of 20 
patients with success would make it much more likely that the process would be associated with 
90% success for each patient than 60%. Should only 15 patients be included in this trial, if 12 or 
greater are able to have a success, then the probability that this would occur would be 94.4% if the 
true probability of a success for a given patient were 90% and it would be 9.1% if it were only 
60%. Thus, while 15 patients would yield useful information, 20 patients as opposed to 15 patients 
would reduce the probability of accepting the results as positive when they may not be, based on 
a low true rate of 60% success, from 9.4% to 5.1%.  
The secondary objectives for this study are as follows: to assess the progression-free survival using 
treatment suggested by the COXEN model in patients with advanced urothelial carcinoma; to 
assess the response rate using treatment suggested by the COXEN model in patients with advanced 
urothelial carcinoma; to assess the overall survival using treatment suggested by the COXEN 
model in patients with advanced urothelial carcinoma; to evaluate toxicity with “best next 
therapy”. The Kaplan Meier method will be used to estimate PFS and OS.  Responses will be 
estimated along with a 95% confidence interval. Toxicity will be evaluated using descriptive 
methods. 
We expect that accrual of 20 patients can be completed within 12 months given our previous rates 
of accrual for similar patients. 
9 
HUMAN SUBJECTS PROTECTIONS  
9.1 
RATIONALE FOR SUBJECT SELECTION 
Subjects with a diagnosis of urothelial carcinoma of the bladder, urethra, ureter, or renal pelvis 
will be eligible for participation in this study. Individuals of any race or ethnic group will be 
eligible for this study.  Eligibility assessment will be based solely on the patient’s medical status.  
Recruitment of patients onto this study will be through standard CCR mechanisms.  No special 
recruitment efforts will be conducted. 
9.1.1 Selection Based on Gender, Ethnicity, and Race 
Subjects from all racial/ethnic groups and both genders are eligible for this study if they meet the 
eligibility criteria. To date, there is no information that suggests that differences in drug 
metabolism, immune response or disease response would be expected in one group compared with 
another. Efforts will be made to extend accrual to a representative population, but in this 
preliminary study, a balance must be struck between patient safety considerations and limitations 
on the number of individuals exposed to potentially toxic and/or ineffective treatments on one hand 


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and the need to explore gender and ethnic aspects of clinical research on the other hand. If 
differences in outcome that correlate with ethnic identity are noted, accrual may be expanded or a 
follow-up study may be written to investigate those differences more fully.  
9.1.2 Justification for Exclusions 
Pregnant women are excluded due to potentially increased risks of serious side effects from the 
drugs administered on this study. Many of the therapeutic agents used in this trial are known to be 
teratogenic. 
9.2 
PARTICIPATION OF CHILDREN 
Individuals under the age of 18 will not be eligible for participation in this study based on the fact 
that patients under 18 are unlikely to have this disease and there are unknown toxicities in pediatric 
patients. 
9.3 
PARTICIPATION OF SUBJECTS UNABLE TO GIVE CONSENT 
Adults unable to give consent are excluded from enrolling in the protocol.  However, re-consent 
may be necessary and there is a possibility, though unlikely, that subjects could become 
decisionally impaired. For this reason and because there is a prospect of direct benefit from 
research participation (section 9.4.1), all subjects ≥ age 18 will be offered the opportunity to fill in 
their wishes for research and care, and assign a substitute decision maker on the “NIH Advance 
Directive for Health Care and Medical Research Participation” form so that another person can 
make decisions about their medical care in the event that they become incapacitated or cognitively 
impaired during the course of the study. Note: The PI or AI will contact the NIH Ability to Consent 
Assessment Team (ACAT) for evaluation as needed for the following: an independent assessment 
of whether an individual has the capacity to provide consent; assistance in identifying and 
assessing an appropriate surrogate when indicated; and/or an assessment of the capacity to appoint 
a surrogate. For those subjects that become incapacitated and do not have pre-determined 
substitute decision maker, the procedures described in MAS Policy 87-4 and NIH HRPP SOP 14E 
for appointing a surrogate decision maker for adult subjects who are (a) decisionally impaired, and 
(b) who do not have a legal guardian or durable power of attorney, will be followed.  
9.4 
EVALUATION OF BENEFITS AND RISKS/DISCOMFORTS 
9.4.1 Benefits 
The potential benefit to a patient who enters study is a reduction in the bulk of his/her tumor, which 
may or may not have a favorable impact on symptoms and/or outcome. 
9.4.2 Risks 
9.4.2.1 Study Agents 
The risks of the study are as described in the various package inserts. 
9.4.2.2 Biopsy risks 
All care will be taken to minimize risks that may be incurred by tumor sampling. However, there 
are procedure-related risks (such as bleeding, infection and visceral injury) that will be explained 
fully during informed consent. If patients suffer any physical injury as a result of the biopsies, 
immediate medical treatment is available at the NCI’s Clinical Center in Bethesda, MD.  Although 


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no compensation is available, any injury will be fully evaluated and treated in keeping with the 
benefits or care to which patients are entitled under applicable regulations. 
9.4.2.3 Radiation Risks 
Some biopsies may be performed with CT guidance.  If that is the case, then this research study 
involves exposure to radiation from up to 2 CT scans with a combined effective dose of 1.6 rem 
and 2 injections of 18F-FDG of 7 mCi each for 2 FDG-PET MRI scans with a combined effective 
dose of 0.94 rem for a total dose of 2.5 rem.  This is below the guideline of 5.0 rem per year 
allowed for research subjects by the NIH Radiation Safety Committee. 
9.4.2.4 Non-Physical Risks of Genetic Research 
9.4.2.4.1 Risk of receiving unwanted information 
Anxiety and stress may arise as a result of the anticipation that unwanted information regarding 
disease related DNA sequencing or disease tendencies, or misattributed paternity. Patients will be 
clearly informed that the data related to DNA sequencing and genetic analysis is coded, 
investigational and will not be shared with patients, family members or health care providers. In 
addition, a Certificate of Confidentiality will be obtained for the study. 
9.4.2.4.2 Risk related to possibility that information may be released 
This includes the risk that data related to genotype, DNA sequencing or risk for disease tendency 
or trait can be released to members of the public, insurers, employers, or law enforcement agencies. 
Although there are no plans to release results to the patients, family members or health care 
providers, this risk will be included in the informed consent document. 
9.4.2.4.3 Risk to family or relatives 
Family members or relatives may or may not want to be aware of familial tendencies or genetic 
risks of disease which may cause anxiety about possible future health problems.  
9.4.2.5 Other Risks 
Additional risks include the possible occurrence of any of a range of side effects which are listed 
in the Consent Document. Frequent monitoring for adverse effects will help to minimize the risks 
associated with administration of the study agents. 
9.5 
RISKS/BENEFITS ANALYSIS 
Although urothelial cancer is a chemosensitive malignancy with response proportions of over 50% 
with conventional cytotoxic regimens, the response durations are short and the median survival of 
patients with metastatic disease is approximately 14 months. There is no FDA-approved second 
line drug for metastatic urothelial cancer.   
Given the efforts to minimize risk with the administration of these agents, this protocol involves 
greater than minimal risk, but presents the potential for direct benefit to individual subjects. The 
ongoing risks and benefits of participation for adults who become unable to consent are no 
different than those described for less vulnerable patients. 


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9.6 
CONSENT PROCESS AND DOCUMENTATION 
The principal investigator or a designee will obtain written informed consent from each subject 
participating in this study after adequate explanation of the aims, methods, anticipated benefits, 
and potential hazards of the study. Potential hazards of the selected agent(s) will also be provided 
as an information sheet in conjunction with the consent as it would be impractical to include all 75 
agents in the consent. Consent for the optional biopsies on this study will be obtained at the time 
of the procedure, using the procedure consent. If the patient refuses the optional biopsy at that 
time, the refusal will be documented in the medical record and in the research record. 
If new safety information results in significant changes in the risk/ benefit assessment, the consent 
form will be reviewed and updated as necessary. All subjects (including those already being 
treated) will be informed of the new information, be given a copy of the revised form, and be asked 
to give their consent to continue in the study. 
It will be documented in the medical record that informed consent has been obtained. 
9.6.1 Telephone Re-consent Procedure 
Re-consent on this study may be obtained via telephone according to the following procedure: the 
informed consent document will be sent to the subject.  An explanation of the study will be 
provided over the telephone after the subject has had the opportunity to read the consent form.  
The subject will sign and date the informed consent. A witness to the subject signature will sign 
and date the consent. The original informed consent document will be mailed back to the 
consenting investigator who will sign and date the consent form with the date the consent was 
obtained via telephone.  A fully executed copy will be returned via mail for the subject’s records.  
The informed consent process will be documented on a progress note by the consenting 
investigator. 
10 PHARMACEUTICAL INFORMATION   
For this study, all drugs are commercially available; therefore, Investigator Brochures are not 
applicable to this/these drug(s). Information about commercial drugs is publicly available in the 
package insert and other resources. Refer to Appendix B for the COXEN drug index FDA 
approved dose and schedule. 
All of the agents are not approved for use in urothelial cancer; however, the purpose of the 
investigation is to determine the feasibility of the use of the COXEN algorithm. The study is 
exempt from IND application because it meets all of the following requirements: the investigation 
is not intended to support a new indication for use or any other significant changes to labeling; the 
drugs are already approved and marketed and the investigation is not intended to support a 
significant change in advertising; and the investigation does not involve a route of administration 
or dosage level in use in a patient population or other factor that significantly increases the risks 
(or decreases the acceptability of the risks) associated with the use of the drug products.  
 
 
 


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92(18): p. 1534-5. 
 


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12 APPENDICES 
12.1 APPENDIX A: PERFORMANCE STATUS CRITERIA 
ECOG Performance Status Scale 
 
Karnofsky Performance Scale 
Grade 
Descriptions 
Percent 
Description 
0 
Normal activity.  Fully active, 
able to carry on all pre-disease 
performance without restriction. 
100 
Normal, no complaints, no 
evidence of disease. 
90 
Able to carry on normal activity; 
minor signs or symptoms of 
disease. 
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). 
80 
Normal activity with effort; 
some signs or symptoms of 
disease. 
70 
Cares for self, unable to carry on 
normal activity or to do active 
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. 
60 
Requires occasional assistance, 
but is able to care for most of 
his/her needs. 
50 
Requires considerable assistance 
and frequent medical care. 
3 
In bed >50% of the time.  
Capable of only limited self-care, 
confined to bed or chair more 
than 50% of waking hours. 
40 
Disabled, requires special care 
and assistance. 
30 
Severely disabled, 
hospitalization indicated.  Death 
not imminent. 
4 
100% bedridden.  Completely 
disabled.  Cannot carry on any 
self-care.  Totally confined to bed 
or chair. 
20 
Very sick, hospitalization 
indicated. Death not imminent. 
10 
Moribund, fatal processes 
progressing rapidly. 
5 
Dead. 
0 
Dead. 


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12.2 APPENDIX B: COXEN DRUG INDEX WITH FDA-APPROVED DOSE AND SCHEDULE 
# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
1 
Abiraterone  
(PO) 
N 
mCRPC 
1000mg QDay w/ prednisone 
2 
Arsenic Trioxide 
(IV) 
N 
APL, relapsed or 
refractory 
Induction: 0.15 mg/kg/day until bone 
marrow remission, max 60 induction 
doses 
Consolidation: 0.15 mg/kg/day 
starting 3-6 weeks after completion 
of induction, max 25 consolidation 
doses 
3 
Asparaginase, 
Escherichia coli 
source 
(IV, IM) 
Y 
ALL 
6000 units/m2/dose TIW  
4 
Axitinib 
(PO) 
N 
RCC, advanced 
Initial: 5mg BID  
Dose Increases: If tolerance criteria 
are met (see package insert), dose 
may be increased to 7mg BID then to 
10mg BID 
5 
Azacitidine 
(IV, SubQ) 
Y 
MDS 
Initial Cycle: 75mg/m2/day x 7days 
Subsequent Cycles: 75 mg/m2/day x 
7 days Q4weeks 
Dose Increases: may increase to 
100mg/m2/day after 2 cycles 
6 
Bendamustine 
(IV) 
Y 
CLL  
100mg/m2 on days 1 and 2 of a 28-
day cycle, max 6 cycles 
NHL 
120mg/m2 on days 1 and 2 of a 21-
day cycle, max 8 cycles 
7 
Bleomycin 
(IV, IM, SubQ) 
Y 
Squamous Cell 
0.25-0.5units/kg Qweek or BIW 
Note: Lymphoma patients should be 
treated with ≤ 2 units for the first two 
doses 
Lymphomas 
Testicular 
8 
Bortezomib 
Y 
MM,  
1.3mg/m2 days 1, 4, 8, 11, 22, 25, 29, 
and 32 of a 42-day cycle x 4 cycles 


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# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
(IV, SubQ) 
first-line 
THEN 1.3mg/m2 days 1, 8, 22 and 
29 of a 42-day cycle x 5 cycles 
w/ melphalan and prednisone  
MM, relapsed   
1.3mg/m2 days 1, 4, 8, and 11 of a 
21-day cycle  
Mantle Cell 
Lymphoma 
9 
Busulfan 
(PO) 
N 
 
CML, palliation 
60mcg/kg/day or 1.8 mg/m2/day 
 
Busulfan 
(IV) 
Y 
HSCT conditioning 
regimen 
0.8mg/kg Q6hours x 4 days 
10 
Carboplatin 
(IV) 
Y 
Ovarian, advanced 
AUC 4-6 Q4weeks 
Bladder 
(unlabeled use) 
AUC 5 Q3weeks w/gemcitabine and 
paclitaxel OR 
AUC 5 Q3weeks w/ gemcitabine OR 
AUC 6 Q3weeks w/ paclitaxel 
11 
Carfilzomib 
(IV) 
N 
MM, relapsed or 
refractory 
Cycle 1: 20mg/m2 on days 1, 2, 8, 9, 
15, and 16 of a 28-day cycle 
Subsequent Cycles: 27mg/m2 on 
days 1, 2, 8, 9, 15, and 16 of a 28-day 
cycle 
12 
Carmustine 
(IV) 
Y 
Brain Tumors 
150-200mg/m2 Q6weeks OR 
75-100mg/m2/day x 2 days Q6weeks  
Hodgkin’s 
Lymphoma 
MM 
NHL 
13 
Chlorambucil 
(PO) 
Y 
CLL 
0.1mg/kg/day x 3-6 weeks 
NHL 
Hodgkin’s 
Lymphoma 
0.2mg/kg/day x 3-6 weeks  


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# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
14 
Cisplatin 
(IV) 
Y 
Bladder, advanced 
50-70mg/m2 Q3-4weeks  
Ovarian,  
metastatic 
Single Agent: 100mg/m2 Q4weeks 
Combination Therapy: 75-100 mg/m2 
Q4weeks 
Testicular, 
metastatic 
20mg/m2/day x 5 days Q3weeks 
15 
Cladribine 
(IV) 
Y 
Hairy Cell Leukemia 
0.09mg/kg/day CIVI x 7 days x 1 
cycle 
16 
Clofarabine 
(IV) 
N 
ALL, 
adults ≤ 21 years 
52mg/m2/day days 1-5 Q2-6weeks 
17 
Crizotinib 
(PO) 
N 
NSCLC, metastatic, 
ALK-positive 
250mg BID 
18 
Cytarabine- 
Conventional 
(IV) 
Y 
AML, remission 
induction 
Combination Therapy: 
100mg/m2/day CIVI x 7 days OR 
100mg/m2 Q12hours x 7 days 
19 
Dacarbazine 
(IV) 
Y 
Hodgkin’s Disease  
Combination Therapy: 
375mg/m2/dose days 1 & 15 
Q4weeks  
Melanoma, 
metastatic 
250mg/m2/dose days 1-5 Q3weeks  
20 
Dactinomycin 
(IV) 
N 
Testicular, metastatic 
Combination Therapy: 1000mcg/m2 
on day 1 
Gestational 
Trophoblastic 
Neoplasm 
Single Agent: 12mcg/kg/day x 5 days  
Combination Therapy: 500mcg/dose 
days 1 and 2  
Wilms’ Tumor, 
Ewing’s Sarcoma, 
Rhabdomyo-sarcoma 
Combination Therapy: 15mcg/kg/day 
x 5 days 
21 
Dasatinib 
Y 
CML, Ph+ 
Chronic Phase: 100mg Qday (may 
increase to 140mg Qday)  


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# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
(PO) 
Accelerated or Blast Phase: 140mg 
Qday (may increase to 180mg Qday) 
ALL, Ph+ 
Initial: 140mg Qday 
Dose Increases: may increase to 
180mg Qday  
22 
Daunorubicin- 
Conventional 
(IV) 
Y 
ALL  
 
Combination Therapy: 45mg/m2/day 
x 3 days Q3-4weeks  
AML, induction, 
adults <60 years 
Combination Therapy 
Cycle 1: 45mg/m2/day x 3 days  
Subsequent Cycles: 45mg/m2/day x 2 
days Q3-4weeks  
AML, induction, 
elderly ≥ 60 years 
Combination Therapy 
Cycle 1: 30mg/m2/day x 3 days  
Subsequent Cycles: 30mg/m2/day x 2 
days Q3-4weeks  
23 
Decitabine 
(IV) 
N 
MDS 
15mg/m2 Q8H x 3 days Q6weeks OR 
20mg/m2 Qday x 5 days Q4weeks  
24 
Docetaxel 
(IV) 
Y 
Breast  
60-100mg/m2 Q3weeks  
NSCLC 
75mg/m2 Q3weeks  
Prostate 
75mg/m2 Q3weeks w/prednisone 
Gastric Adeno-
carcinoma 
75mg/m2 Q3weeks w/cisplatin and 
fluorouracil 
Head and Neck 
75mg/m2 Q3weeks w/cisplatin and 
fluorouracil x 3-4 weeks, followed 
by RT 
Bladder 
(unlabeled use) 
Single Agent: 100mg/m2 Q3weeks 
Combination Therapy: 35mg/m2 day 
1 and 8 Q21 days w/ gemcitabine and 
cisplatin 
25 
Doxorubicin- 
Y 
Breast 
60mg/m2 on day 1 of a 21-day cycle 
x 4 cycles w/ cyclophosphamide 


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# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
Conventional 
(IV) 
Metastatic Solid 
Tumors, Leukemia, 
or Lymphoma 
Single Agent: 60-75mg/m2 Q21days 
Combination Therapy: 40-75mg/m2 
Q21-28days 
26 
Epirubicin 
(IV) 
N 
Breast, adjuvant 
60mg/m2 day 1 and 8 Q28days x 6 
cycles w/ cyclophosphamide and 
fluorouracil  OR 
100mg/m2 day 1 Q21days x 6 cycles 
w/ cyclophosphamide and 
fluorouracil  
27 
Eribulin 
(IV) 
N 
Breast, metastatic 
1.4mg/m2/dose days 1 and 8 
Q21days 
28 
Erlotinib 
(PO) 
Y 
NSCLC,  
first-line EGFR 
mutations 
150mg Qday  
NSCLC, refractory or 
maintenance 
Pancreatic 
100mg Qday  
29 
Estramustine 
(PO) 
Y 
Prostate, progressive 
or metastatic 
14mg/kg/day (3 to 4 divided doses) 
30 
Etoposide 
(IV) 
N 
SCLC 
35mg/m2/day x 4 days, up to 
50mg/m2/day x 5 days Q3-4weeks  
Testicular 
50-100mg/m2/day x 5 days OR 
100mg/m2/day days 1, 3, and 5 Q3-
4weeks 
 
Etoposide  
(PO) 
Y 
SCLC 
PO twice IV dose 
31 
Exemestane 
(PO) 
N 
Breast 
25mg Qday  
32 
Floxuridine 
(intra-arterial) 
N 
CRC,  
hepatic mets 
0.1-0.6mg/kg/day continuous 
infusion  


61 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
33 
Fludarabine 
(IV) 
Y 
CLL 
25mg/m2/day x 5 days Q28days 
34 
Fluorouracil 
(IV) 
Y 
Breast 
Initial Course: 12mg/kg Qday x 4 
days then 6mg/kg days 6, 8, 10 and 
12 
Maintenance Therapy: repeat dosage 
of initial course every 30 days after 
the last day of previous course OR 
10-15mg/kg Qweek  
CRC 
Pancreatic 
Gastric 
35 
Gefitinib 
(PO) 
N 
NSCLC, 
first-line EGFR 
mutations 
250mg Qday  
36 
Gemcitabine 
(IV) 
Y 
Breast, 
metastatic 
1250mg/m2 days 1 and 8 Q21days w/ 
paclitaxel  
NSCLC, locally 
advanced or 
metastatic 
1000mg/m2 days 1, 8, and 15 
Q28days w/ cisplatin OR 
1250mg/m2 days 1 and 8 Q21days 
w/cisplatin 
Ovarian, 
advanced 
1000mg/m2 days 1 and 8 Q21days w/ 
carboplatin 
Pancreatic, locally 
advanced or 
metastatic 
1000mg/m2 Qweek x 7 weeks 
followed by 1 week rest THEN 
Qweek x 3 weeks every 4 weeks 
Bladder 
(unlabeled use) 
1000mg/m2 days 1, 8, and 15 Q28 
days w/ cisplatin OR 
1000mg/m2 days 1 and 8 Q21 days 
w/ carboplatin 
37 
Hydroxyurea 
(PO) 
Y 
CML, resistant 
20-30mg/kg Qday 
Solid Tumors (Head 
and Neck, 
Melanoma, Ovarian) 
Intermittent: 80mg/kg Q3days  
Continuous: 20-30mg/kg Qday  
38 
Idarubicin 
(IV) 
Y 
AML 
Induction: 12mg/m2/day x 3 days 


62 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
Consolidation: 10-12mg/m2/day x 2 
days 
39 
Ifosfamide 
(IV) 
Y 
Testicular 
1200mg/m2/day x 5 days Q3weeks, 
combo therapy & w/mesna 
40 
Imatinib 
(PO) 
Y 
CML, Ph+ 
Chronic Phase: 400mg Qday, may 
increase to 600mg Qday 
Accelerated Phase or Blast Crisis: 
600mg Qday, may increase to 400mg 
BID 
GIST 
400mg Qday, may increase to 400mg 
BID 
ALL, Ph+ 
600mg Qday  
41 
Irinotecan 
(IV) 
Y 
CRC, metastatic 
125mg/m2 days 1, 8, 15, and 22 of a 
6 week cycle OR 
350mg/m2 Q3weeks 
42 
Ixabepilone 
(IV) 
N 
Breast, metastatic or 
locally advanced 
40mg/m2/dose Q3 weeks, max 88mg 
43 
Lapatinib 
(PO) 
N 
Breast, HER2+ 
1250mg Qday w/ capecitabine OR 
1500mg Qday w/ letrozole 
44 
Lomustine 
(PO) 
N 
Brain Tumors 
130mg/m2 Q6weeks  
Hodgkin’s 
Lymphoma 
45 
Mechlor- 
Ethamine 
(IV) 
Y 
Hodgkin’s 
Lymphoma, 
palliative 
0.4mg/kg 
46 
Melphalan  
(IV) 
Y 
MM, palliative 
16mg/m2 Q2weeks x 4 doses then 
Q4weeks 
 
Melphalan  
(PO) 
Y 
MM, palliative  
6mg Qday x 2-3 weeks, followed by 
4 weeks rest then 2mg Qday 
Ovarian 
0.2mg/kg/day x 5 days Q4-5weeks 


63 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
47 
Mercapto- 
Purine 
(PO) 
Y 
ALL 
1.5-2.5mg/kg Qday  
48 
Methotrexate 
(intrathecal, IV, IM, 
SubQ) 
Y 
Numerous Oncology-
Related Indications 
Intrathecal: 15mg day 1, 8, 15, 22, 
and 29; 12mg Q2-7days 
IV (large range of approved doses): 
40mg/m2 Qweek; 12gm/m2 (max 
20gm) weeks 3, 4, 8, and 9 
 
Methotrexate 
(PO) 
Y 
Numerous Oncology-
Related Indications 
20mg/m2 Qweek 
49 
Mitomycin 
(IV) 
Y 
Stomach or Pancreas 
Adeno-carcinoma 
20mg/m2 Q6-8weeks 
50 
Mitotane 
(PO) 
Y 
ACC 
2-6gm Qday (divided) then increase 
incrementally to 9-10gm Qday 
(divided)  
51 
Mitoxantrone 
(IV) 
N 
ANLL 
Induction: 12mg/m2 Qday x 3 days 
w/ cytarabine 
Consolidation (6 weeks after 
induction): 12mg/m2 Qday x 2 days, 
repeat in 4 weeks 
Prostate, advanced, 
hormone-refractory 
12-14mg/m2 Q3weeks w/ steroids 
52 
Nilotinib 
(PO) 
Y 
CML, Ph+ 
Newly-Diagnosed in Chronic Phase: 
300mg BID  
Resistant, Chronic or Accelerated 
Phase: 
400mg BID  
53 
Oxaliplatin 
(IV) 
Y 
CRC 
85mg/m2 Q2weeks 
54 
Paclitaxel 
(IV) 
Y 
Ovarian 
135-175mg/m2 Q3weeks OR 
50-80mg/m2 Qweek OR 


64 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
1.4-4mg/m2/day CIVI x 14 days 
Q4weeks 
Breast, metastatic  
175-250mg/m2 Q3weeks OR 
50-80mg/m2 Qweek OR 
1.4-4mg/m2/day CIVI x 14 days 
Q4weeks 
NSCLC 
135mg/m2 Q3weeks 
AIDS-Related 
Kaposi’s Sarcoma 
135mg/m2 Q3weeks OR 
100mg/m2 Q2weeks 
55 
Pazopanib 
(PO) 
N 
RCC 
800mg Qday  
Soft Tissue Sarcoma, 
advanced refractory 
56 
Pentostatin 
(IV) 
Y 
Hairy Cell Leukemia 
4mg/m2 Q2weeks  
57 
Romidepsin 
(IV) 
N 
Cutaneous T-cell 
Lymphoma 
14mg/m2 days 1, 8 and 15 of a 28-
day cycle  
Peripheral T-cell 
Lymphoma 
58 
Ruxolitinib 
(PO) 
N 
Myelofibrosis 
Platelets >200,000/mm3: 20mg BID 
Platelets 100,000-200,000/mm3: 
15mg BID  
Platelets 50,000-<100,000/mm3: 5mg 
BID 
59 
Sorafenib 
(PO) 
Y 
HCC 
400mg BID 
RCC, advanced 
Thyroid, 
differentiated 
60 
Streptozocin 
(IV) 
Y 
Pancreatic Islet Cell  
500mg/m2/day x 5 days Q6 weeks 
OR 
1000mg/m2 Qweek 


65 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
61 
Sunitinib 
(PO) 
Y 
GIST 
50mg Qday x 4 weeks Q6weeks  
RCC, advanced 
PNET, advanced 
37.5mg Qday 
62 
Tamoxifen 
(PO) 
Y 
Breast 
20mg Qday  
63 
Temsirolimus 
(IV) 
N 
RCC 
25mg Qweek  
64 
Teniposide 
(IV) 
N 
ALL, pediatric 
165-250 mg/m2 twice weekly or 
weekly x 8-9 doses. 
65 
Thioguanine 
(PO) 
Y 
AML 
2mg/kg Qday 
66 
Thiotepa 
(intravesical, IV) 
N 
Bladder 
60mg Qweek x 4 weeks 
(intravesical) 
Ovarian 
0.3-0.4mg/kg Q1-4weeks (IV) 
Breast 
67 
Topotecan  
(IV) 
Y 
Cervical 
0.75mg/m2/day x 3 days Q21days w/ 
cisplatin 
Ovarian 
1.5mg/m2/day x 5 days Q21days  
SCLC 
 
Topotecan  
(PO) 
N 
SCLC 
2.3mg/m2/day x 5 days Q21days 
68 
Toremifene 
(PO) 
N 
Breast, metastatic 
60mg Qday 
69 
Tretinoin  
(PO) 
Y 
APL 
45mg/m2/day (divided) 


66 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
# 
Drug Name and 
route of 
administration 
Formulary 
Status 
(Y, yes; N, 
no) 
FDA-Approved 
Indication 
FDA-Approved Dose & 
Schedule 
70 
Vandetanib 
(PO) 
N 
MTC, locally 
advanced or 
metastatic 
300mg Qday 
71 
Vemurafenib 
(PO) 
N 
Melanoma, 
metastatic or 
unresectable, 
BRAFV600E 
mutation 
960mg BID  
72 
Vinblastine 
(IV) 
Y 
Hodgkin’s, NHL, 
Testicular, Breast, 
Kaposi’s Sarcoma 
Typical dosages: 3.7mg/m2(initial), 
up to 5.5mg/m2 (dose 2), up to 
7.4mg/m2 (dose 3), up to 9.25mg/m2 
(dose 4), and up to 11.1mg/m2 (dose 
5); administered Q7days 
Usual Range: 5.5-7.4mg/m2 Q7days 
73 
Vincristine 
(IV) 
Y 
Numerous Oncology-
Related Indications 
1.4mg/m2/dose, max 2mg/dose, 
various frequencies 
74 
Vismodegib 
(PO) 
N 
Basal Cell, metastatic 
or locally advanced 
150mg Qday 
75 
Vorinostat 
(PO) 
N 
Cutaneous  
T-cell Lymphoma 
400mg Qday  
 
 


67 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
12.3 APPENDIX C:  SAMPLE COLLECTION FOR CORRELATIVE STUDIES 
The date and exact time of each blood draw should be recorded on each tube. 
Test description 
Amount/Type/
Tube/Handling 
Collection Timing 
Contact 
Special Instructions 
Archival Tissue 
 
1-2 Tumor 
blocks or 25 
unstained slides 
and 2 H&E 
slides  
(Sample 1) 
Baseline  
(Pre treatment) 
Marissa Mallek, 
marissa.mallek@nih.gov, 
240-760-7498 
Tumor blocks will be submitted to Maria Merino 
for cutting into slides. 
Unstained slides will be distributed by research 
nurse to: 
Laboratory of pathology, Dr. Merino:5 unstained 
slides and 1 H&E 
Meltzer’s lab for NGS: 10 unstained slides and 1 
H&E 
Trepel’s lab for Immune gene signature: 10 
unstained slides and 1 H&E 
Fresh Tumor 
Biopsy  
 
2-3 cores  
(Sample 2 and 
Sample 5) 
Baseline (mandatory) 
and at progression 
(optional)   
Marissa Mallek, 
marissa.mallek@nih.gov, 
Ph: 240-760-7498   
Dr. Bottaro, 
don.bottaro@nih.gov,  
Ph: 301-402-6499  
Rene Costello, 
rene.costello@nih.gov, 
Ph: 301-443-6915 to 
arrange for pick-up  
Fresh tumor will be fixed and submitted to Maria 
Merino for processing. 
Unstained Slides will be distributed by research 
nurse to: 
Meltzer’s lab for NGS: 10 unstained slides and 1 
H&E 
Trepel’s lab for Immune gene signature: 10 
unstained slides and 1 H&E 


68 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
Test description 
Amount/Type/
Tube/Handling 
Collection Timing 
Contact 
Special Instructions 
Saliva 
3 mL; Oragene 
saliva collection 
kit (DNA 
Genotek) 
Baseline  
(at any time after 
consent is signed, but 
preferably at baseline) 
Marissa Mallek, 
marissa.mallek@nih.gov,  
24 hours prior to 
anticipated collection.  
Call 240-760-7498 for 
immediate pick-up of 
sample   
Patient should not eat or drink anything 30 
minutes prior to collection. 
After the sample collection, when the top of the 
device is closed a preservative is released. 
Remove the top and close the tube with a cap. 
Store at room temperature. 
 ctDNA isolation 
10 mL of blood; 
Streck tube 
(Streck, Omaha, 
NE) 
(Sample 3 and 
Sample 4) 
Baseline and C3D1 
Meltzer lab, 
pmeltzer@mail.nih.gov. 
Email 24 hr prior to 
collection. 
 
Invert Streck tube 2-4 times, keep at room 
temperature (15- 30 degrees C) and transferr to 
the lab for processing . 
Immune Subset 
analysis 
8 mL of blood; 
two BD 
Vacutainer Cell 
Preparation 
Tubes (CPT) 
citrate 
Baseline (just prior to 
beginning therapy), 
C2D1 and C3D1 
Developmental 
Therapeutics Branch:  
Jane Trepel, 
trepelj@mail.nih.gov  
Yusuke Tomita, 
yusuke.tomita@nih.gov 
Sunmin Lee, 
lees@pop.nci.gov 
Mi-Jung 
leemin@,mail.nih.gov  
Call 240-760-6330 
immediately after blood is 
drawn 
Invert several times and keep at room temp  


69 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
Test description 
Amount/Type/
Tube/Handling 
Collection Timing 
Contact 
Special Instructions 
Peripheral 
Blood Immune 
Gene 
Expression 
2.5 ml of blood; 
one PAXgene 
RNA tube 
(PreAnalytix) 
Baseline (just prior to 
beginning therapy) 
and C2D1 
Developmental 
Therapeutics Branch : 
Phone: 240-760-6330  
Jane Trepel, 
trepelj@mail.nih.gov  
Yusuke Tomita, 
yusuke.tomita@nih.gov 
Sunmin Lee, 
lees@pop.nci.gov 
Mi-Jung, 
leemin@mail.nih.gov  
Invert several times and keep at room temp  
Pro-
Inflammatory 
Cytokines 
10 mL of blood; 
one EDTA tube 
Baseline, C2D1,  
and C3D1 
Figg lab. 
NCIBloodcore@mail.nih.
gov.  
Contact 24 hr prior or for 
questions regarding 
sample processing. 
Page 102-11964 for 
sample pick-up.  
For immediate help, call 
240-760-6180 (main blood 
processing core number) 
or, 240-760-6190 (main 
clinical pharmacology lab 
number).  
Invert EDTA tube 2-4 times, place on wet ice and 
store at 4°C until processing for a maximum of 
30-60 min. 
Upon arrival in the BPC, blood samples will be 
centrifuged for 5 minutes at 1200 x g at 4qC. 
Plasma will be aliquoted into 2 cryovials and 
stored at -80qC until the time of analysis. 


70 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
Test description 
Amount/Type/
Tube/Handling 
Collection Timing 
Contact 
Special Instructions 
Plasma HGF 
and MET 
4 mL of blood; 
one EDTA tube 
Baseline  
and day 1 of each 
treatment cycle 
Figg lab. 
NCIBloodcore@mail.nih.
gov. Contact 24 hr prior or 
for questions regarding 
sample processing.  
Page 102-11964 for 
sample pick-up.  
For immediate help, call 
240-760-6180 (main blood 
processing core number) 
or, 240-760-6190 (main 
clinical pharmacology lab 
number).  
Invert EDTA tube 2-4 times, place on wet ice and 
store at 4°C in the refrigerator until processing for 
a maximum of 30-60 min. 
Upon arrival in the BPC, samples will be 
centrifuged and the plasma transferred into 
cryovials for storage at -80 C until the time of 
analysis. Samples will be barcoded as described 
in Section 5.3. 
DMET 
6 mL of blood; 
one EDTA tube 
Baseline 
Figg lab. 
NCIBloodcore@mail.nih.
gov. Contact 24 hr prior or 
for questions regarding 
sample processing.  
Page 102-11964 for 
sample pick-up.  
For immediate help, call 
240-760-6180 (main blood 
processing core number) 
or, 240-760-6190 (main 
clinical pharmacology lab 
number).  
Invert EDTA tube 2-4 times, place on wet ice and 
store at 4°C in the refrigerator until processing for 
a maximum of 30-60 min. The date and exact 
time of each blood draw should be recorded on 
the sample tube and on the PK sheet. 
Upon arrival in the Clinical Pharmacology Lab, 
samples will be centrifuged and the plasma 
transferred into cryovials for storage at -80°C 
until the time of analysis. Samples will be 
barcoded as described in Section 5.3 


71 
Abbreviated Title: COXEN study 
Version Date: March 21, 2019 
 
Test description 
Amount/Type/
Tube/Handling 
Collection Timing 
Contact 
Special Instructions 
Circulating 
tumor cells 
(Trepel Lab) 
10 ml of blood; 
Cell save tube  
Baseline, (just prior to 
beginning therapy)and 
C2D1 
Jane Trepel, 
trepelj@mail.nih.gov  
Yusuke Tomita, 
yusuke.tomita@nih.gov  
Sunmin Lee, 
lees@pop.nci.gov  
Call 240-760-6330 for 
immediate pick-up 
Invert several times and place at room temp or 
refrigerate 
Circulating 
tumor cells  
(Epic Science) 
4-10 mL of 
blood; Streck 
Cell-Free DNA 
BCT 
Baseline and  
First Restaging 
Send to Epic Science as 
detailed in Section 5.1.9.1  
 
Confirm blood tube is not expired.  
Collect minimum of 4 mL blood per sample, but a 
full 10 mL tube of blood should be provided when 
possible (*see note regarding prevention of 
backflow in Section 5.1.9).  
Remove tube from adapter and immediately mix 
by gentle inversion 8 to 10 times to prevent 
clotting. Inadequate or delayed mixing may result 
in inaccurate test results.  
Label the tube with subject’s identification and 
date and time of blood draw. Unlabeled blood 
tubes may not be processed.  
Keep sample at room temperature and ship on day 
of collection in shipper with ambient gel packs. 
  
 

ICF_001.pdf: 
MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
INSTITUTE: 
National Cancer Institute 
STUDY NUMBER: 
16-C-0121 
PRINCIPAL INVESTIGATOR: 
Andrea Apolo, M.D. 
STUDY TITLE:  
A Pilot Clinical Trial of Genomic Based Assignment of Therapy in Advanced Urothelial 
Carcinoma 
 
Continuing Review Approved by the IRB on 02/11/19 
Amendment Approved by the IRB on 02/12/19 (B) 
 
Date posted to web: 02/21/19 
Standard 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent (1) 
 
 
INTRODUCTION 
We invite you to take part in a research study at the National Institutes of Health (NIH). 
First, we want you to know that: 
Taking part in NIH research is entirely voluntary.  
You may choose not to take part, or you may withdraw from the study at any time.  In 
either case, you will not lose any benefits to which you are otherwise entitled.  However, 
to receive care at the NIH, you must be taking part in a study or be under evaluation for 
study participation. 
You may receive no benefit from taking part.  The research may give us knowledge that 
may help people in the future. 
Second, some people have personal, religious or ethical beliefs that may limit the kinds of 
medical or research treatments they would want to receive (such as blood transfusions).  If you 
have such beliefs, please discuss them with your NIH doctors or research team before you agree 
to the study. 
Now we will describe this research study.  Before you decide to take part, please take as much 
time as you need to ask any questions and discuss this study with anyone at NIH, or with family, 
friends or your personal physician or other health professional.   
Why is this study being done?  
Urothelial cancer that has spread is currently incurable, yet there are only a small number of 
chemotherapy drugs that have been tested in this disease. Conducting standard trials of already 
FDA approved drugs in urothelial cancer would take years and be very costly.  


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 2 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
In collaboration with the University of Colorado Comprehensive Cancer Center, we have 
developed an approach, the COXEN (Co-eXpression ExtrapolatioN) model which is the 
mathematical method that predicts whether cells respond to treatment, and that we think will 
help us determine, which, if any currently FDA approved anticancer drugs would be effective in 
your urothelial cancer based on the mutations (changes in genes) that are present in your tumor.  
The study is being done in order to test whether it is possible to use the COXEN model to choose 
the next best therapy for your urothelial cancer (after it has gotten worse on standard therapy) in 
three weeks, which is about the usual amount of time a doctor would have to make treatment 
decisions outside of a clinical trial.   
We will also look at how well your tumor responds to the next best therapy.  
While all of the drugs used in this study are FDA approved, they are not necessarily approved for 
the treatment of urothelial cancer. 
Why are you being asked to take part in this study? 
You are being asked to participate in this study because you have been urothelial cancer that has 
spread after you have received at least one line of chemotherapy. 
How many people will take part in this study?  
Up to 20 subjects will take part in this study. 
Description of Research Study 
What will happen if you take part in this research study?   
Before you begin the study 
You will undergo the following tests and/or procedures to find out if you can be in the study: 
• Medical history 
• Physical examination 
• Routine blood and urine tests 
• CT scan or MRI of your tumor 
• Tests for certain viruses  
• Pregnancy test if you are a woman who can have children (you will not be allowed to 
participate if you are pregnant) 
We will also need a sample of your tumor from a previous surgery and a fresh mandatory biopsy 
to test using the COXEN model so that we may determine which of 75 approved drugs or 
combinations of these drugs should work best on your tumor. You will be given an information 
sheet about the drug(s) selected by the model. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 3 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
During the study 
You will undergo additional tests/procedures before you begin any study therapy. Some of the 
tests are the same as those done at screening and be repeated if they were performed within the 
appropriate timeframe.  Additional tests/procedures include an EKG. 
You will begin taking the assigned medication based on the study doctor’s instructions and the 
information sheet specific to the drug(s) assigned to you by the COXEN model.  Depending on 
the dosing schedule of the medication(s) you are assigned, you will be seen every 2 or 3 weeks. 
At each visit you will undergo routine blood and urine tests. You will continue taking the 
assigned drug(s) until you have intolerable side effects or until your disease worsens.  If your 
disease worsens after beginning your first COXEN selected therapy, you will be given the option 
to then be assigned to a second COXEN selected therapy which will continue until intolerable 
side effect occur or your disease worsens. A second COXEN based regimen may also be selected 
if you must be removed from the first one due to side effects and the study doctor feels that it 
would be to your benefit.   
If you are being seen every 2 weeks, you will have scans of your tumor every 8 weeks so we can 
determine whether your cancer is responding.  If you are being seen every 3 weeks, you will 
have your scans every 9 weeks.  You will undergo a second tumor biopsy, this one optional, at 
the time your disease gets worse the first time. The biopsy to be performed is exclusively for 
research purposes and will not benefit you.  It might help other people in the future.  You can 
decide whether or not you’d like to participate in the optional biopsy at the time of the biopsy.  
Research Procedures 
In addition to the biopsy, you will also be asked for additional blood and undergo scans for 
research testing.   
Sampling 
Blood and tumor tissue used for research studies will undergo genetic testing. Please note that 
the genetic analyses that we perform in our laboratory are for research purposes only; they are 
not nearly as sensitive as the tests that are performed in a laboratory that is certified to perform 
genetic testing. Changes that we observe unrelated to our research may or may not be valid. 
Therefore, we do not plan to inform you of the results of testing on your specimens that is 
performed in our research lab. However, in the unlikely event that we discover a finding that is 
believed to be clinically important based on medical standards at the time we first analyze your 
results, we will contact you. This could be many years in the future. We will ask you to have an 
additional tube of blood drawn to verify the findings we have seen in our lab. If the results are 
verified, you will be re-contacted and offered a referral to a genetic healthcare provider to 
discuss the results. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 4 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Scans 
There are two additional FDG-PET MRIs scans which are being performed for research.  These 
will be done before you start treatment and after 8 or 9 weeks, depending on the first treatment to 
which you are assigned. PET is a nuclear medicine medical imaging technique that produces a 3-
D image of functional processes in the body. An FDG PET scan uses a small amount of a 
radioactive drug, 18F-FDG (fluorodeoxyglucose) to show differences between healthy tissue and 
diseased tissue. Before the PET scan, a small amount of FDG is injected into you. Because 
cancer grows at a faster rate than healthy tissue, cancer cells absorb more of the 18F-FDG. The 
PET scanner detects the radiation given off by the 18F-FDG and produces color-coded images of 
the body that show both normal and cancerous tissue.  
The PET scan will be done in combination with a convention MRI scan so that images of both 
anatomy (MRI) and function (PET) can be taken during the same examination.  
On the day of your exam, you will be encouraged to drink water.  You will be given an injection 
of 18F-FDG. Then you will wait ~ 60 minutes for the drug to travel through your body after 
which you will be asked to urinate and then lie on a partially enclosed scanning table. The table 
will slide into the machine. You will be asked to remain still during the scan. You will hear 
buzzing or clicking sounds during the scan. You will need to lie still for about 20-60 minutes 
before coming off of the scanning table. If you feel any anxiety over being in enclosed spaces, let 
your study doctor know.    
When you are finished taking the drugs  
Four to five weeks after you have had your last dose of study drug, you will be asked to return to 
the NIH for a safety assessment that will include a medical history, a physical examination and 
routine blood tests. If you are unable to return to NIH, we will contact you by phone for an 
assessment. 
We will remain in contact, generally by telephone, approximately every two months to follow 
the progress of your disease, to find out about any new cancer drugs you are taking and your 
survival status. 
Treatment 
Treatments covered under this study may include a single medication or a combination of 
medications, surgery or radiation to treat your cancer. These treatments will not be experimental. 
Your doctors will describe your treatment plan to you in detail before asking you to sign this 
consent form. You may be asked to sign a separate consent form for any treatment procedures 
not outlined in this consent. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 5 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Birth Control 
If you are a woman who is breast feeding or pregnant, you may not take part in the study because 
the medicine may affect your baby or your unborn child. If you are a woman who can become 
pregnant, or are the partner of a woman who can become pregnant, you will need to practice an 
effective form of birth control before starting study treatment and during study treatment. Please 
refer to your information sheet for how long after your last dose of study therapy you should 
continue practicing contraception. If you think that you or your partner is pregnant, you should 
tell your study doctor or nurse at once.   
Effective forms of birth control include:  
• abstinence  
• intrauterine device (IUD) 
• hormonal [birth control pills, injections, or implants] 
• tubal ligation 
• vasectomy 
Please refer to your information sheet. It is possible that some of the methods listed above may 
not be recommended for use with your assigned drug(s). 
Risks or Discomforts of Participation 
Risks of study drugs 
The drugs used in this study may affect how different parts of your body work such as your liver, 
kidneys, heart, and blood. The study doctor will be testing your blood and will let you know if 
changes occur that may affect your health. 
There is also a risk that you could have side effects from the study drug(s)/study approach. 
Here are important points about side effects: 
• The study doctors do not know who will or will not have side effects. 
• Some side effects may go away soon, some may last a long time, or some may never go 
away. 
• Some side effects may interfere with your ability to have children. 
• Some side effects may be serious and may even result in death. 
Here are important points about how you and the study doctor can make side effects less of a 
problem: 
• Tell the study doctor if you notice or feel anything different so they can see if you are 
having a side effect. 
• The study doctor may be able to treat some side effects. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 6 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
• The study doctor may adjust the study drugs to try to reduce side effects. 
Please refer to the information sheet for your assigned drug(s) for the specific side effects. 
Risks of research blood sampling 
Side effects of blood draws include pain and bruising in the area where the needle was placed, 
lightheadedness, and rarely, fainting. When large amounts of blood are collected, low red blood 
cell count (anemia) can develop. 
Risks of biopsy  
As indicated above, you will undergo 2 research biopsies (1 optional and neither of any medical 
benefit to you) to obtain tumor tissue during your participation in this study.  One biopsy will be 
done before you have taken any study medication and the second biopsy is optional that will be 
done at the time your disease gets worse.  The specific risks will depend upon the location of 
your tumor and the method by which it is biopsied.  The doctor will review these specific risks 
with you and have you sign a separate consent for the biopsy.  In general the potential risks of 
biopsy include bleeding and infection. A CT scan may also be used to guide the biopsy which 
brings the additional risk of exposure to radiation as discussed below. 
Research radiation risks 
In this study, you may be exposed to up to 2 CT scans to guide your biopsies and 2 injections of 
18F-FDG of 7 mCi each for imaging studies. This radiation exposure is not required for your 
medical care and is for research purposes only. The amount of radiation you will receive in this 
study is 2.5 rem which is below the guideline of 5 rem per year allowed for research subjects by 
the NIH Radiation Safety Committee. The average person in the United States receives a 
radiation exposure of 0.3 rem per year from natural sources, such as the sun, outer space, and the 
earth's air and soil. If you would like more information about radiation, please ask the 
investigator for a copy of the pamphlet, An Introduction to Radiation for NIH Research Subjects. 
While there is no direct evidence that the amount of exposure received from participating in this 
study is harmful, there is indirect evidence it may not be completely safe. There may be a very 
slight increase in the risk of cancer.  
Please tell your doctor if you have had any radiation exposure in the past year, either from other 
research studies or from medical tests or care, so we can make sure that you will not receive too 
much radiation. Radiation exposure includes x-rays taken in radiology departments, cardiac 
catheterization, and fluoroscopy as well as nuclear medicine scans in which radioactive materials 
were injected into your body.  
If you are pregnant you will not be permitted to participate in this research study. If you are 
breast feeding and the protocol involves injection of radioactive material, you will not be 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 7 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
permitted to participate. It is best to avoid radiation exposure to unborn or nursing infants since 
they are more sensitive to radiation than adults. 
Risks of genetic research 
Psychological or Social Risks Associated with Return of Incidental or Secondary Findings 
Additional risk from genetic testing in the face of a diagnosis of cancer is less likely.  
Although your genomic information is unique to you, you share some genomic similarities with 
your children, parents, brothers, sisters, and other blood relatives. Therefore, learning your 
research results could mean something about your family members and might cause you or your 
family distress. Before joining the study, it may be beneficial to talk with your family members 
about whether and how they want you to share your results with them.  
Privacy Risks Associated with Return of Incidental or Secondary Findings 
Your privacy is very important to us and we will use many safety measures to protect your 
privacy. However, in spite of all of the safety measures that we will use, we cannot guarantee 
that your identity will never become known. While neither the public nor the controlled-access 
databases developed for this project will have information such as your name, address, telephone 
number, or social security number, it may be possible to identify you based on the information in 
these databases and other public information (including information you tell people or post about 
yourself).  The risk of this happening is currently very low.  
Although your genetic information is unique to you, you do share some genetic information with 
your children, parents, brothers, sisters, and other blood relatives. Consequently, it may be 
possible that genetic information from them could be used to help identify you.  Similarly, it may 
be possible that genetic information from you could be used to help identify them.  Patterns of 
genetic variation also can be used by law enforcement agencies to identify a person or his/her 
blood relatives.   
It is possible also that someone could get unauthorized access or break into the system that stores 
information about you.  Every precaution will be taken to minimize this risk.  There also may be 
other privacy risks that we have not foreseen. 
Protections against misuse of genetic information 
Since some genetic variations can help to predict future health problems for you and your 
relatives, this information might be of interest to health care providers, life insurance companies, 
and others. However, Federal and State laws provide some protections against discrimination 
based on genetic information.  For example, the Genetic Information Nondiscrimination Act 
(GINA) makes it illegal for health insurance companies, group health plans, and most employers 
to discriminate against you based on your genetic information. However, GINA does not prevent 
companies that sell life insurance, disability insurance, or long-term care insurance from using 
genetic information as a reason to deny coverage or set premiums. GINA also does not apply to 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 8 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
members of the United States military, individuals covered by the Indian Health Service, or 
veterans obtaining health care through the Veteran’s Administration.  Lastly, GINA does not 
forbid insurance medical underwriting based on your current health status though the Affordable 
Care Act limits consideration of pre-existing conditions by insurers. 
Potential Benefits of Participation 
Are there benefits to taking part in this study?  
The aim of this study is to see if this experimental method of assigning treatment can be done 
quickly enough (within 3 weeks) to make it an effective tool in clinical practice. We do not know 
if you will receive personal, medical benefit from taking part in this study. These potential 
benefits could include shrinking of your tumor or lessening of your symptoms, such as pain, that 
are caused by the cancer. Because there is not much information about the drug’s effect on your 
cancer, we do not know if you will benefit from taking part in this study, although the knowledge 
gained from this study may help others in the future who have cancer.   
Alternative Approaches or Treatments 
What other choices do I have if I do not take part in this study? 
Instead of being in this study, you have these options: 
• Getting treatment or care for your cancer without being in a study 
• Taking part in another study 
• Getting comfort care, also called palliative care. This type of care helps reduce pain, 
tiredness, appetite problems and other problems caused by the cancer. It does not treat the 
cancer directly. Instead, it tries to improve how you feel. Comfort care tries to keep you 
as active and comfortable as possible. 
Please talk to your doctor about these and other options. 
Stopping Therapy  
Your doctor may decide to stop your therapy for the following reasons: 
• if he/she believes that it is in your best interest 
• if your disease comes back after your second COXEN selected treatment 
• if you become pregnant 
• if you have side effects from the treatment that your doctor thinks are too severe 
• if new information shows that another treatment would be better for you 
• In this case, you will be informed of the reason therapy is being stopped. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 9 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
You can stop taking part in the study at any time. However, if you decide to stop taking part in 
the study, we would like you to talk to the study doctor and your regular doctor first. 
If you decide at any time to withdraw your consent to participate in the trial, we will not collect 
any additional medical information about you. However, according to FDA guidelines, 
information collected on you up to that point may still be provided to collaborators or designated 
representatives. If you withdraw your consent and leave the trial, any samples of yours that have 
been obtained for the study and stored at the NCI can be destroyed upon request. However, any 
samples and data generated from the samples that have already been distributed to other 
researchers or placed in the research databases cannot be recalled and destroyed. 
Research Subject’s Rights 
What are the costs of taking part in this study?  
If you choose to take part in the study, the following will apply, in keeping with the NIH policy: 
• 
You will receive study treatment at no charge to you. This may include 
surgery, medicines, laboratory testing, x-rays or scans done at the 
Clinical Center, National Institutes of Health (NIH), or arranged for you 
by the research team to be done outside the Clinical Center, NIH if the 
study related treatment is not available at the NIH.  
• 
There are limited funds available to cover the cost of some tests and 
procedures performed outside the Clinical Center, NIH. You may have to 
pay for these costs if they are not covered by your insurance company. 
• 
Medicines that are not part of the study treatment will not be provided or 
paid for by the Clinical Center, NIH. 
• 
Once you have completed taking part in the study, medical care will no 
longer be provided by the Clinical Center, NIH. 
Will your medical information be kept private?  
We will do our best to make sure that the personal information in your medical record will be 
kept private. However, we cannot guarantee total privacy. Organizations that may look at and/or 
copy your medical records for research, quality assurance, and data analysis include: 
• The National Cancer Institute (NCI) and other government agencies, like the Food and 
Drug Administration (FDA), which are involved in keeping research safe for people. 
• National Institutes of Health Intramural Institutional Review Board 
• Collaborators at the University of Colorado Comprehensive Cancer Center where the 
COXEN assay will be performed 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 10 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Your individual genomic data and health information will be put in a controlled-access database.  
This means that only researchers who apply for and get permission to use the information for a 
specific research project will be able to access the information.  Your genomic data and health 
information will not be labeled with your name or other information that could be used to 
identify you.  Researchers approved to access information in the database have agreed not to 
attempt to identify you. 
Certificate of Confidentiality 
To help us protect your privacy, we have obtained a Certificate of Confidentiality.  The 
researchers can use this Certificate to legally refuse to disclose information that may identify you 
in any federal, state, or local civil, criminal, administrative, legislative, or other proceedings, for 
example, if there is a court subpoena. The researchers will use the Certificate to resist any 
demands for information that would identify you, except as explained below.  
You should also know that there are several circumstances in which the Certificate does not 
provide coverage. These include when information:  
• will be used for auditing or program evaluation internally by the NIH; or  
• must be disclosed to meet the legal requirements of the federal Food and Drug 
Administration (FDA).  
• is necessary for your medical treatment and you have consented to this disclosure;  
• is for other research.  
In addition, identifiable, sensitive information protected by this Certificate cannot be admissible 
as evidence or used for any purpose in any action, suit, or proceeding without your consent. 
You should understand, however, that a Certificate of Confidentiality does not prevent you or a 
member of your family from voluntarily releasing information about yourself or your 
involvement in this research.  If an insurer, employer, or other person obtains your written 
consent to receive research information, then the researchers will not use the Certificate to 
withhold that information. 
The Certificate of Confidentiality will not protect against the required reporting by hospital staff 
of information on suspected child abuse, reportable communicable diseases, and/or possible 
threat of harm to self or others. 
Conflict of Interest 
The National Institutes of Health (NIH) reviews NIH staff researchers at least yearly for conflicts 
of interest. This process is detailed in a Protocol Review Guide. You may ask your research team 
for a copy of the Protocol Review Guide or for more information. Members of the research team 
who do not work for NIH are expected to follow these guidelines but they do not need to report 
their personal finances to the NIH. 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 11 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Members of the research team working on this study may have up to $15,000 of stock in the 
companies that make products used in this study.  This is allowed under federal rules and is not a 
conflict of interest.  
Use of Specimens and Data for Future Research 
To advance science, it is helpful for researchers to share information they get from studying 
human samples.  They do this by putting it into one or more scientific databases, where it is 
stored along with information from other studies. A researcher who wants to study the 
information must apply to the database and be approved. Researchers use specimens and data 
stored in scientific databases to advance science and learn about health and disease.   
We plan to keep some of your specimens and data that we collect and use them for future 
research and share them with other researchers. We will not contact you to ask about each of 
these future uses.  These specimens and data will be stripped of identifiers such as name, address 
or account number, so that it may be used for future research on any topic and shared broadly for 
research purposes.  Your specimens and data will be used for research purposes only and will not 
benefit you.  It is also possible that the stored specimens and data may never be used.  Results of 
research done on your specimens and data will not be available to you or your doctor.  It might 
help people who have cancer and other diseases in the future.   
If you do not want your stored specimens and data used for future research, please contact us in 
writing and let us know that you do not want us to use your specimens and/or data.  Then any 
specimens that have not already been used or shared will be destroyed and your data will not be 
used for future research. However, it may not be possible to withdraw or delete materials or data 
once they have been shared with other researchers. 


MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 12 of 13 pages 
 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH 
STUDY (Continuation Sheet) 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.:  09-25-0099 
 
File in Section 4:  Protocol Consent 
 
 
 
OTHER PERTINENT INFORMATION 
1. 
Confidentiality. When results of an NIH research study are reported in medical journals 
or at scientific meetings, the people who take part are not named and identified.  In most cases, 
the NIH will not release any information about your research involvement without your written 
permission. However, if you sign a release of information form, for example, for an insurance 
company, the NIH will give the insurance company information from your medical record. This 
information might affect (either favorably or unfavorably) the willingness of the insurance 
company to sell you insurance. 
The Federal Privacy Act protects the confidentiality of your NIH medical records. However, you 
should know that the Act allows release of some information from your medical record without 
your permission, for example, if it is required by the Food and Drug Administration (FDA), 
members of Congress, law enforcement officials, or authorized hospital accreditation 
organizations. 
2. 
Policy Regarding Research-Related Injuries. The Clinical Center will provide short-
term medical care for any injury resulting from your participation in research here.  In general, 
no long-term medical care or financial compensation for research-related injuries will be 
provided by the National Institutes of Health, the Clinical Center, or the Federal Government.  
However, you have the right to pursue legal remedy if you believe that your injury justifies such 
action. 
3. 
Payments. The amount of payment to research volunteers is guided by the National 
Institutes of Health policies. In general, patients are not paid for taking part in research studies at 
the National Institutes of Health. Reimbursement of travel and subsistence will be offered 
consistent with NIH guidelines. 
4. 
Problems or Questions. If you have any problems or questions about this study, or about 
your rights as a research participant, or about any research-related injury, contact the Principal 
Investigator, Andrea Apolo, M.D., Building 10, Room 13N240, Telephone:  301-480-0536. You 
may also call the Clinical Center Patient Representative at 301-496-2626. If you have any 
questions about the use of your specimens or data for future research studies, you may also 
contact the Office of the Clinical Director, Telephone: 240-760-6070. 
5. 
Consent Document. Please keep a copy of this document in case you want to read it 
again. 


MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
STUDY NUMBER: 16-C-0121 
CONTINUATION: page 13 of 13 pages 
 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH 
STUDY (Continuation Sheet) 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.:  09-25-0099 
 
File in Section 4:  Protocol Consent 
 
 
 
 
COMPLETE APPROPRIATE ITEM(S) BELOW: 
A. Adult Patient’s Consent 
B. Parent’s Permission for Minor Patient. 
I have read the explanation about this study 
and have been given the opportunity to discuss 
it and to ask questions.  I hereby consent to 
take part in this study. 
I have read the explanation about this study 
and have been given the opportunity to discuss 
it and to ask questions.  I hereby give 
permission for my child to take part in this 
study.   
(Attach NIH 2514-2, Minor’s Assent, if 
applicable.) 
 
  
  
 
 
 
Signature of Adult Patient/ 
Legal Representative 
Date 
Signature of Parent(s)/ Guardian Date 
 
 
Print Name 
 
 
Print Name 
C. Child’s Verbal Assent (If Applicable) 
The information in the above consent was described to my child and my child agrees to 
participate in the study. 
 
  
  
 
Signature of Parent(s)/Guardian 
Date 
Print Name 
THIS CONSENT DOCUMENT HAS BEEN APPROVED FOR USE  
FROM FEBRUARY 11, 2019 THROUGH FEBRUARY 25, 2020.  
 
 
 
 
 
 
 
 
Signature of Investigator 
Date 
Signature of Witness 
Date 
 
 
Print Name 
 
 
Print Name