Phase II Study of Nivolumab (Anti-PD1), Tadalafil and Oral Vancomycin in Patients With Refractory Primary Hepatocellular Carcinoma or Liver Dominant Metastatic Cancer From Colorectal or Pancreatic Cancers
Phase: PHASE2
Conditions: Hepatocellular Carcinoma, Hepatocellular Cancer, Metastatic Pancreatic Cancer, Metastatic Colorectal Cancer, Liver Metastasis
Sponsor: National Cancer Institute (NCI)

Abbreviated Title: Nivo, tadalafil, vancomycin
Version Date: 01/19/2022
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Abbreviated Title: Nivo, tadalafil, vancomycin
NIH Protocol: 19C0033 
Version Date: 01/19/2022
NCT: NCT03785210
Title: Phase II Study of Nivolumab (anti-PD1), Tadalafil and Oral Vancomycin in Patients with 
Refractory Primary Hepatocellular Carcinoma or Liver Dominant Metastatic Cancer from 
Colorectal or Pancreatic Cancers
NCI Principal Investigator:   
       Tim F. Greten, MD 
         Thoracic & GI Malignancies Branch
         National Cancer Institute
         Building 10, Room 2B38B
         9000 Rockville Pike
         Bethesda, MD 20892
         240-760-6114 
         gretentf@mail.nih.gov
 
Investigational Agents:
Drug Name:
Nivolumab
Tadalafil
Vancomycin
IND Number:
133973
133973
133973
Sponsor:
NCI CCR
NCI CCR
NCI CCR
Manufacturer:
Bristol Myers Squibb 
Generic
Generic
Supplier:
Bristol Myers Squibb
NIH Clinical Center 
Pharmacy
NIH Clinical Center 
Pharmacy

Abbreviated Title: Nivo, tadalafil, vancomycin
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PRÉCIS
Background:

Current treatment options for patients with liver cancers, including hepatocellular carcinoma 
(HCC) and advanced liver cancers are limited and take no account of the known biological and 
genetic heterogeneity in these diseases. Median survival for advanced disease remains poor at 
approximately 1 year. 

Nivolumab is a fully human monoclonal immunoglobulin G4 (IgG4) antibody that is specific 
for human programmed death-1 (PD-1, cluster of differentiation 279 [CD279]) cell surface 
membrane receptor. Nivolumab has been approved by FDA for the treatment of HCC and other 
solid tumors.

Tadalafil is a phosphodiesterase type 5 (PDE5) inhibitor which have been approved by the 
FDA for the treatment of pulmonary arterial hypertension, benign prostatic hyperplasia and 
erectile dysfunction, with a relative safe clinical profile. PDE5 inhibitors have been examined 
in multiple malignancies and cancer cell lines for their direct anticancer activities, for their 
efficacy as chemo-sensitizers and for cancer chemoprevention. 

Oral vancomycin is antibiotic that has effect on altering gut commensal bacteria subsequently 
inducing a liver-selective anti-tumor effect.

The aim of the study is to evaluate whether the immunomodulatory effect induced by PDE5 
inhibitor and oral vancomycin can be enhanced by immune checkpoint inhibition in advanced 
liver cancer. 
Objective:

To determine the Best Overall Response (BOR) according to Response Evaluation Criteria 
(RECIST 1.1) to combined treatment of nivolumab, oral vancomycin and tadalafil in patients 
with refractory primary HCC or liver dominant metastatic cancer from colorectal cancer (CRC) 
or pancreatic adenocarcinoma (PDAC).
Eligibility:

Histologically confirmed, hepatocellular carcinoma (HCC) 
Or 

Histologically confirmed carcinoma highly suggestive of a diagnosis of HCC
Or

Histologically confirmed advanced colorectal or pancreatic malignancy with liver involvement 
as dominant site of metastasis

Measurable lesion, accessible for biopsy. 

Age ≥ 18 years

ECOG ≤ 1

Acceptable renal, bone marrow and liver function. 

Willingness to undergo two mandatory tumor biopsies. 
Design:

The proposed study is a phase II study of combined nivolumab, oral vancomycin and tadalafil 
treatment in patients with HCC or liver dominant metastatic cancer from colorectal or 
pancreatic cancers.

Abbreviated Title: Nivo, tadalafil, vancomycin
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
Treatment will be delivered in cycles consisting of 4 weeks (+/- 3 days) until progression or 
unacceptable toxicity. 

Patients will be seen in Clinical Center on monthly basis with disease status evaluation every 
8 (+/-1) weeks after start of study therapy.

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TABLE OF CONTENTS
PRÉCIS...........................................................................................................................................2
TABLE OF CONTENTS ..............................................................................................................4
STATEMENT OF COMPLIANCE.............................................................................................9
1
INTRODUCTION .................................................................................................................9
1.1
Study Objectives..............................................................................................................9
1.1.1
Primary Objective:.......................................................................................................9
1.1.2
Secondary Objectives: .................................................................................................9
1.1.3
Exploratory Objectives:...............................................................................................9
1.2
Background and Rationale.............................................................................................10
1.2.1
HCC And Immune Checkpoint Inhibitors (ICIs) ......................................................10
1.2.2
Liver Metastases and Current Treatment Options.....................................................12
1.2.3
Tumor Microenvironment (TME) .............................................................................14
1.2.4
PDE5 Inhibitors Augment Immunotherapy by Targeting on TME...........................16
1.2.5
Gut Microbiome and Liver Cancer Control...............................................................23
1.2.6
Nivolumab .................................................................................................................25
1.2.7
PDE5 Inhibitor and Oral Vancomycin in Combination with Nivolumab .................27
2
ELIBILITY ASSESSMENT AND ENROLLMENT .......................................................28
2.1
Eligibility Criteria..........................................................................................................28
2.1.1
Inclusion Criteria .......................................................................................................28
2.1.2
Exclusion Criteria ......................................................................................................30
2.1.3
Recruitment Strategies...............................................................................................31
2.2
Screening Evaluation.....................................................................................................31
2.2.1
Screening activities performed prior to obtaining informed consent ........................31
2.2.2
Screening activities performed after a consent for screening has been signed..........32
2.3
Participant Registration and Status Update Procedures.................................................33
2.3.1
Treatment Assignment Procedures (for registration purposes only) .........................33
2.4
Baseline Evaluation .......................................................................................................33
3
STUDY IMPLEMENTATION...........................................................................................33
3.1
Study Design..................................................................................................................33
3.2
Drug Administration......................................................................................................34
3.2.1
Nivolumab Administration ........................................................................................34
3.2.2
Tadalafil Administration............................................................................................34
3.2.3
Vancomycin Administration......................................................................................35
3.2.4
Sequence and Monitoring of Dose Administration ...................................................35
3.3
Dose Delay or Modifications.........................................................................................35
3.3.1
General Guidance ......................................................................................................35
3.3.2
Nivolumab .................................................................................................................36
3.3.3
Tadalafil.....................................................................................................................41
3.3.4
Oral Vancomycin.......................................................................................................41
3.4
Study Calendar...............................................................................................................43
3.5
Costs and Compensation................................................................................................46

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3.5.1
Costs ..........................................................................................................................46
3.5.2
Compensation ............................................................................................................46
3.5.3
Reimbursement..........................................................................................................46
3.6
Criteria for Removal from Protocol Therapy and Off Study Criteria ...........................46
3.6.1
Criteria for Removal from Protocol Therapy ............................................................46
3.6.2
Off Study Criteria ......................................................................................................46
3.6.3
Lost to Follow-up ......................................................................................................47
4
CONCOMITANT MEDICATIONS/MEASURES...........................................................47
4.1
Excluded Concomitant Medications..............................................................................47
4.2
Methods of Contraception .............................................................................................48
4.2.1
Female Patient of Child-bearing Potential.................................................................48
4.2.2
Male Patients with a Female Partner of Childbearing Potential................................48
4.2.3
Highly Effective Methods of Contraception..............................................................48
4.3
Blood Donation..............................................................................................................49
5
CORRELATIVE STUDIES FOR RESEARCH...............................................................49
5.1
Biospecimen Collection.................................................................................................49
5.2
Correlative Studies of Blood Samples...........................................................................51
5.2.1
Immune Monitoring...................................................................................................51
5.2.2
Serum Cytokines........................................................................................................51
5.2.3
Tadalafil Pharmacokinetic Analysis..........................................................................51
5.2.4
Vancomycin Pharmacokinetic Analysis....................................................................51
5.2.5
Bile Acid Analysis.....................................................................................................51
5.2.6
Food Diary for Mixed Meal Study ............................................................................51
5.3
Tumor Studies (Non-genetic Analysis) .........................................................................52
5.3.1
Tumor Tissue Analysis..............................................................................................52
5.3.2
Stool Collection and Processing ................................................................................52
5.4
Sample Storage, Tracking and Disposition ...................................................................52
5.4.1
Samples Managed by Dr. Figg’s Blood Processing Core (BPC) ..............................53
5.4.2
Samples Managed by the Laboratory of Dr. Tim Greten ..........................................53
5.4.3
Protocol Completion/Sample Destruction .................................................................53
5.5
Samples for Genetic/Genomic Analysis........................................................................54
5.5.1
TCR Beta Sequencing................................................................................................54
5.5.2
Whole exome/Panel (tumor/normal) and RNAseq (tumor).......................................54
5.5.3
Microbial DNA Sequence..........................................................................................54
5.5.4
Management of Results .............................................................................................54
5.5.5
Genetic Counseling....................................................................................................54
6
DATA COLLECTION AND EVALUATION..................................................................55
6.1
Data Collection ..............................................................................................................55
6.2
Data Sharing Plans.........................................................................................................55
6.2.1
Human Data Sharing Plan .........................................................................................55
6.2.2
Genomic Data Sharing Plan.......................................................................................56
6.3
Response Criteria...........................................................................................................56
6.3.1
Disease Parameters ....................................................................................................56
6.3.2
Methods for Evaluation of Measurable Disease........................................................57

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6.3.3
Response Criteria.......................................................................................................58
6.4
Toxicity Criteria.............................................................................................................60
7
NIH REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING PLAN
60
7.1
Definitions .....................................................................................................................60
7.2
OHSRP Office of Compliance and Training / IRB Reporting ......................................60
7.2.1
Expedited Reporting ..................................................................................................60
7.2.2
IRB Requirements for PI Reporting at Continuing Review ......................................61
7.3
NCI Clinical Director Reporting ...................................................................................61
7.4
NIH Required Data and Safety Monitoring Plan...........................................................61
8
SPONSOR SAFETY REPORTING ..................................................................................61
8.1
Definitions .....................................................................................................................61
8.1.1
Adverse Event............................................................................................................61
8.1.2
Serious Adverse Event (SAE) ...................................................................................61
8.1.3
Life-threatening .........................................................................................................62
8.1.4
Severity......................................................................................................................62
8.1.5
Relationship to Study Product ...................................................................................62
8.2
Assessment of Safety Events.........................................................................................62
8.3
Reporting of Serious Adverse Events............................................................................63
8.4
Waiver of expedited reporting to CCR..........................................................................63
8.5
Safety Reporting to the Manufacturer, Bristol Myers Squibb.......................................63
8.6
Reporting Pregnancy .....................................................................................................65
8.6.1
Maternal exposure ....................................................................................................65
8.6.2
Paternal exposure.......................................................................................................66
8.7
Regulatory Reporting for Studies Conducted Under CCR-Sponsored IND .................66
9
CLINICAL MONITORING...............................................................................................66
10
STATISTICAL CONSIDERATIONS...............................................................................66
10.1
Study Objective .............................................................................................................66
10.1.1
Primary Objective..................................................................................................66
10.1.2
Secondary Objectives ............................................................................................67
10.2
Sample Size Determination ...........................................................................................67
10.3
Populations for Analyses ...............................................................................................68
10.4
Statistical Analyses........................................................................................................68
10.4.1
General Approach..................................................................................................68
10.4.2
Analysis of the Primary Endpoints........................................................................68
10.4.3
Analysis of the Secondary Endpoint......................................................................68
10.4.4
Safety Analyses .....................................................................................................68
10.4.5
Baseline Descriptive Statistics...............................................................................68
10.4.6
Planned Interim Analyses......................................................................................68
10.4.7
Exploratory Analyses.............................................................................................68
11
COLLABORATIVE AGREEMENT.................................................................................69
11.1
Cooperative Research and Development Agreement (CRADA) ..................................69

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12
HUMAN SUBJECTS PROTECTIONS ............................................................................69
12.1
Rationale for Subject Selection .....................................................................................69
12.2
Participation of Children................................................................................................69
12.3
Participation of Subjects Unable to Give Consent ........................................................69
12.4
Evaluation of Benefits and Risks/Discomforts for All Participants ..............................70
12.4.1
Benefits..................................................................................................................70
12.4.2
Risks ......................................................................................................................70
12.5
Risks/Benefits Analysis for All Participants .................................................................71
12.6
Consent Process and Documentation.............................................................................72
12.6.1
Consent Process for Adults Who Lack Capacity to Consent to Research 
Participation...........................................................................................................................72
13
REGULATORY AND OPERATIONAL CONSIDERATIONS.....................................72
13.1
Study Discontinuation and Closure ...............................................................................72
13.2
Quality Assurance and Quality Control.........................................................................73
13.3
Conflict of Interest Policy..............................................................................................73
13.4
Confidentiality and Privacy ...........................................................................................73
14
PHARMACEUTICAL INFORMATION .........................................................................74
14.1
Nivolumab (133973)......................................................................................................74
14.1.1
Source: ...................................................................................................................74
14.1.2
Toxicity:.................................................................................................................74
14.1.3
Formulation and Preparation .................................................................................74
14.1.4
Stability and Storage..............................................................................................74
14.1.5
Administration Procedures ....................................................................................75
14.2
Tadalafil.........................................................................................................................75
14.2.1
Source ....................................................................................................................75
14.2.2
Description.............................................................................................................75
14.2.3
How Supplied ........................................................................................................75
14.2.4
Toxicity..................................................................................................................75
14.2.5
Administration:......................................................................................................76
14.3
Oral Vancomycin...........................................................................................................76
14.3.1
Source ....................................................................................................................76
14.3.2
Mode of Action......................................................................................................76
14.3.3
Description.............................................................................................................76
14.3.4
How Supplied ........................................................................................................76
14.3.5
Administration:......................................................................................................77
14.3.6
Storage ...................................................................................................................77
15
REFERENCES ....................................................................................................................78
16
APPENDICES......................................................................................................................84
16.1
Appendix A: Performance Status Criteria .....................................................................84
16.2
Appendix B: Patient’s Medication Diary ________________________ .....................85
16.3
Appendix C: Management Algorithms For Endocrinopathy, Gastrointestinal, Hepatic, 
Neurological, Pulmonary, Renal, And Skin Adverse Events ....................................................88
16.4
Appendix D: Infusion-related Reaction Management Algorithm .................................96

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16.5
Appendix E: Modified immune-related response criteria (irRC)..................................98
16.6
Appendix F: Child-Pugh Classification System..........................................................100
16.7
Appendix G: Pregnancy Form.....................................................................................101
16.8
Appendix H: Food Diary for Mixed Meal Study.........................................................105
16.9
Appendix I : Mixed Meal PK Sheet ............................................................................106

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STATEMENT OF COMPLIANCE
The trial will be carried out in accordance with International Conference on Harmonisation Good 
Clinical Practice (ICH GCP) and the following: 
United States (US) Code of Federal Regulations (CFR) applicable to clinical studies (45 
CFR Part 46, 21 CFR Part 50, 21 CFR Part 56, 21 CFR Part 312, and/or 21 CFR Part 812) 
National Institutes of Health (NIH)-funded investigators and clinical trial site staff who are 
responsible for the conduct, management, or oversight of NIH-funded clinical trials have 
completed Human Subjects Protection and ICH GCP Training.
The protocol, informed consent form(s), recruitment materials, and all participant materials will 
be submitted to the Institutional Review Board (IRB) for review and approval.  Approval of both 
the protocol and the consent form must be obtained before any participant is enrolled.  Any 
amendment to the protocol will require review and approval by the IRB before the changes are 
implemented to the study.  In addition, all changes to the consent form will be IRB-approved; an 
IRB determination will be made regarding whether a new consent needs to be obtained from 
participants who provided consent, using a previously approved consent form
1
INTRODUCTION
1.1
Study Objectives
1.1.1
Primary Objective:

To determine the Best Overall Response (BOR) according to Response Evaluation Criteria 
(RECIST 1.1) to combined treatment of nivolumab, oral vancomycin and tadalafil in patients 
with refractory primary HCC or liver dominant metastatic cancer from colorectal cancer (CRC) 
or pancreatic adenocarcinoma (PDAC).
1.1.2
Secondary Objectives:

To assess the safety and tolerability of nivolumab in combination with oral vancomycin and 
tadalafil in patients with refractory primary HCC or liver dominant metastatic cancer from 
CRC or PDAC.

To assess overall survival (OS) of nivolumab combined with oral vancomycin and tadalafil in 
patients with refractory HCC or liver dominant metastatic cancer from CRC or PDAC.
1.1.3
Exploratory Objectives:

To measure changes in immune cell population in the peripheral blood and tumors of patients 
treated on this study, including CD4+, CD8+, MDSCs, MAITS NKT cells, IFNgamma 
signaling pathway, perforin and granzyme. 

To measure secreted cytokines in the peripheral blood to monitor the relative level change with 
the treatment

To measure changes in bile acid in the peripheral blood of patients treated in this study.

To evaluate tadalafil and oral vancomycin pharmacokinetic level to monitor compliance and 
association between the level and response rate or immune parameters

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
To evaluate TCR beta sequencing to analyze the T cell response from PBMC and tumor 
samples 

To assess gene and protein expression and metabolite profile from tumor to evaluate the 
association of the change immune cell signature, signaling pathways and metabolite with 
clinical outcome

To measure changes in gut microbiome in the stool of patients treated on this study.

To evaluate the incidence of vancomycin-resistant enterococci (VRE) development during the 
trial.

To evaluate the relationship between the response rate, overall survival and immune 
parameters, gut microbiome, and bile acid component in patients treated on this study.
1.2
Background and Rationale
1.2.1
HCC And Immune Checkpoint Inhibitors (ICIs)
Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide and the second 
leading cause of cancer associated mortality with an average life expectancy of 6-9 months[1]. In 
the United States, there were 39,230 new cases and 27,170 deaths related to liver cancer in 2016[2]. 
By 2030, these numbers are expected to rise, primarily due to increase rates of hepatitis C virus 
(HCV) related cirrhosis[3]. In early stage disease, treatment has traditionally comprised surgery 
(partial resection or transplantation) or locoregional therapies like ablation or chemoembolization. 
Survival rates are varied between the different modalities. However, most patients relapse with 
recurrent and often, metastatic disease. 
In advanced disease, systemic therapy with sorafenib or lenvatinib is the standard first line 
treatment. Sorafenib is a potent oral multikinase inhibitor that prevents tumor cell growth and 
angiogenesis. It is approved for inoperable or metastatic HCC based on two randomized phase III 
clinical trials. In the SHARP trial, sorafenib improved median overall survival (OS) by 3 months 
compared to best supportive care (BSC); 10.7 months in the sorafenib arm and 7.9 months in the 
BSC arm (HR 0.69;95% CI 0.55-0.87, p<0.001)[4]. In a similarly designed study by Cheng, et al, 
Asian patients were enrolled to receive sorafenib versus placebo. Median OS improved by 
approximately 2 months, 6.5 months in the sorafenib arm vs 4.2 months with placebo (HR 0.68, 
95% CI 0.50-0.93, p=0.014)[5]. Recent FDA-approved lenvatinib for advanced HCC based on 
REFLECT trial showed non-inferior but not statistically superior to sorafenib for OS (HR 0.92; 95% 
CI: 0.79, 1.06). Median OS in the lenvatinib arm was 13.6 months and 12.3 months in the sorafenib 
arm. REFLECT also demonstrated a statistically significant improvement in progression-free 
survival (PFS) with lenvatinib as compared to sorafenib. Median PFS was 7.3 months in the 
lenvatinib arm and 3.6 months in the sorafenib arm (HR 0.64; 95% CI: 0.55, 0.75; p<0.001). The 
overall response rate was higher for the lenvatinib arm as compared to sorafenib (19% vs. 7% per 
RECIST 1.1).[6]
For patients who progress following first line treatment, regorafenib and recently nivolumab, are 
approved as second line agents. Regorafenib is a multikinase inhibitor targeting tumor growth and 
angiogenesis. In a study comparing regorafenib versus placebo in patients with advanced HCC 
who progressed through sorafenib, regorafenib improved OS by approximately 4 months (HR 0.63, 
95% CI 0.50-0.79, p<0.001), and progression free survival (HR 0.46, 95% CI 0.37-0.56, p<0.001) 
[7].

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To date, there have been 4 published immunotherapy trials in HCC (Table 1) [8-11]. The first was 
a single-arm phase II trial evaluating anti-CTLA-4 tremelimumab in 21 patients with HCC in the 
setting of second line treatment[10]. Tremelimumab showed promising antitumor activity and an 
acceptable safety profile. The partial response rate was 17.6% (3/17), and the disease control rate 
was 76% (13/17). Duration of response was 3.6, 9.2, and 15.7 months in the 3 responders, 
respectively. More recently, tremelimumab in combination with loco-reginal therapy has 
demonstrated activity and feasibility [8]. In this pilot study, 32 patients with HCC received 
tremelimumab at 2 dose levels (3.5 mg/kg in 6 patients and 10 mg/kg in 26 patients) every 4 weeks 
for 6 doses, then every 3 months. In conjunction with this, 7 patients with Barcelona Clinic Liver 
Cancer (BCLC) stage B disease underwent Transarterial chemoembolization, 10 patients with 
BCLC stage C disease underwent radiofrequency ablation, and 11 patients with BCLC stage C 
disease underwent cryoablation. Rapid disease progression was observed in 4 patients who were 
then excluded. Response assessment was performed in the 19 patients with evaluable lesions 
outside of the ablation zone. The partial response rate was 26% (5/19) and the disease control rate 
was 84% (16/19). CheckMate-040 was the largest study to evaluate immunotherapy in HCC thus 
far[9]. This was a multicenter phase I/II dose escalation and expansion study investigating 
nivolumab in patients with advanced HCC. A total of 214 patients with advanced HCC and Child-
Pugh score of 6 or less received nivolumab 3 mg/kg every 2 weeks on the dose expansion phase. 
The study included patients with and without chronic viral hepatitis, including active HBC (31%) 
and HCV (21%) but not those with active co-infection with HBV and HCV or with hepatitis D 
virus infection. Most patients had advanced progressive disease, with 68% having extrahepatic 
spread and 76% having progressed despite prior systemic therapy. Partial responses were observed 
in 15% and 20% of patients in the dose-escalation and dose-expansion phases, respectively. 
Disease control rates were 64% and 58%, respectively. Responses were durable, averaging 17 
months and 10 months, respectively. On the basis of CheckMate-040, FDA granted nivolumab 
accelerated approval in September 2017 for patients with advanced HCC previously treated with 
sorafenib. Following the aforementioned strong objective activity of nivolumab in the second-line 
scenario, phase III trials are testing the effect of nivolumab against sorafenib for patients naive to 
systemic therapy (NCT02576509). Recently, pembrolizumab (another PD-1–blocking agent) 
showed clinical activity for sorafenib-exposed patients (NCT02702401, KEYNOTE-240)[11]. In 
this study, 104 eligible patients were enrolled and treated, and objective response was 17% 
(18/104). The best overall responses were one (1%) complete and 17 (16%) partial responses; 
meanwhile, 46 (44%) patients had stable disease, 34 (33%) had progressive disease. Overall, the 
ORR is still suboptimal from these strategies. 

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Table 1: Key immunotherapy trials in Hepatocellular Carcinoma (HCC) 
Drug(s)
Target
Population
No.
RR
Reference
Tremelimumab
CTLA-4
Advanced HCC
21
17.6%
Sangro et 
al.[10]
Tremelimumab + 
Ablation
CTLA-4
Advanced HCC
32
26.3%
Duffy et al. [8]
Advanced HCC (naïve to 
Sorafenib)
80
22.5%
Nivolumab
PD-1
Advanced HCC (Intolerant or 
progressed to Sorafenib)
182
18.6%
EI-Khoueiry et 
al. [9]
Pembrolizumab
PD-1
Advanced HCC (Intolerant or 
progressed to Sorafenib)
104
17.3%
Zhu et al. [11]
1.2.2
Liver Metastases and Current Treatment Options
Tumors metastatic to the liver are more common than primary tumors. The most common sites of 
primary tumor are breast, lung, and colorectal cancer. Some authors have reported hepatic 
metastases in as many as 40 to 50% of adult patients with extrahepatic primary tumors[12]. Due 
to the central role in the portal circulation, the liver is a frequent site for metastatic tumor, 
especially for tumors of the digestive tract such as colon and pancreas [13-16]. The incidence of 
liver metastases is different among the primary tumor types. Accordingly, liver metastasis affects 
more than 50% of colorectal cancer (CRC) patients during their lifespan[17] and 60% metastatic 
pancreatic adenocarcinoma (PDAC) patients[13]. Liver metastases also occur at high 
frequencies in other solid tumors but are usually detected in the late stage of tumor progression. 
About 8% of prostate cancer patients develop clinically evident liver metastases during their 
treatment period [18] and a much higher percentage (25–45%) of prostate cancer patients show 
liver involvement at autopsy[19, 20]. A similar pattern is also observed in breast cancer patients 
where less than 5% of the patients have clinically evident liver metastases at the time of diagnosis 
[21, 22]. 
Liver metastases are associated with a poor prognosis and remain a major cause of morbidity and 
mortality in patients with malignant tumors of different origins. The presence of liver metastases 
still remains a clinically signiﬁcant and therapeutically frustrating problem in clinical oncology 
secondary to the sanctuary effect of the liver in relation to systemic chemotherapy. For example, 
15 to 25% of patients with CRC present with liver metastases, and another 25 to 50% develop liver 
metastasis within 3 years following resection of the primary tumor [23, 24]. 
Table 2: Key immunotherapy trials in CRC
Drug(s)
Target
Population
No.
Liver 
metastases
RR
Refer
ence
dMMR CRC
Pembrolizumab
PD-1
Refractory CRC
11
55%
40%
Le et 
al. 
[25]
Nivolumab
PD-1
Refractory CRC
74
N/A
32%
Over
man 

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Table 2: Key immunotherapy trials in CRC
Drug(s)
Target
Population
No.
Liver 
metastases
RR
Refer
ence
et al. 
[26]
Nivolumab + Ipilimumab
PD-1 + 
CTLA-4
Refractory CRC
119
N/A
55%
Over
man 
et al. 
[27]
pMMR CRC
Pembrolizumab
PD-1
Refractory CRC
21
52%
0%
Le et 
al. 
[25]
Unselected CRC
Tremelimumab
CTLA-4
Refractory CRC
47
N/A
2%
Chun
g et 
al. 
[28]
Nivolumab
PD-1
Refractory CRC
19
N/A
0%
Topal
ian et 
al. 
[29] 
[30]
BMS-936559
PD-L1
Refractory CRC
18
N/A
0%
Brah
mer 
et al. 
[30]
Atezolizumab + 
Bevacizumab
Refractory CRC
14
N/A
7%
Atezolizumab + 
FOLFOX/bev
PD-L1
Metastatic CRC
(70% first line)
30
N/A
40% 
Bend
ell et 
al. 
[31]
Atezolizumab + 
Cobimetinib
PD-L1 + 
MEK
Refractory CRC 
23
N/A
17%
Bend
ell et 
al. 
[32]
Conventional treatment of liver metastases include surgery (hepatic resection), intervention 
(embolization and transcatheter arterial chemoembolization), systemic chemotherapy, and 
radiofrequency ablation. Among them, only hepatic resection for metastatic disease has gained 
general acceptance as a potentially curative option. The potentially curative treatment of liver 
metastasis of CRC with surgical resection has been reported for many years, with 5-year survival 
rates now of 50%-60% [33, 34]. Moreover, if patients are well selected, up to 20% can achieve 
long-term disease-free survival after metastectomy [33-35]. The role of surgery for metastases 
from neuroendocrine tumors on long-term outcome is also well-documented [36, 37]. Surgical 
resection of hepatic metastases from gastric adenocarcinoma was associated with a significantly 
improved overall survival [38]. It is also noted that surgical resection of hepatic metastases was 
associated with a significantly improved overall 1-year and 3-year survival with median survival 
of 9.9 months for all PDAC patients with liver metastases [39]. Surgical resection allows for the 
maximum response rates, but unfortunately, the number of patients that are eligible for resection 
and/or ablation based on a ﬁnite number of liver metastasis and liver-only disease is small. Only 
10%-20% of liver metastases from CRC are amenable to resection. Nevertheless, recurrences 

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occur in two-thirds of patients after resection for liver metastases within the first 2 years. One 
multi-center study of 1669 patients found that more than half developed recurrences, with median 
disease-free survival duration of 16.3 months[40].
Table 3: Key immunotherapy trials in Pancreatic adenocarcinoma (PDAC)
Drug(s)
Target
Population
No.
Liver 
metastases
RR
Refer
ence
Ipilimumab
CTLA-4
Advanced/Metastatic PDAC
27
N/A
3.7%
Royal 
et al. 
[41]
Ipilimumab + 
Gemcitabine
CTLA-4
Advanced/Metastatic PDAC
13
N/A
15%
Anjali 
et al. 
[42] 
Ipilimumab + GVAX
CTLA-4
Advanced/Metastatic PDAC
30
N/A
0%
Le et 
al. 
[43]
BMS-936559
PD-L1 
Advanced/Metastatic PDAC
14
N/A
0%
Brah
mer 
et 
al.[30
]
Pembrolizumab + 
Gem/Nab
PD-1
Advanced/Metastatic PDAC
11
N/A
18%
Weiss 
et al. 
[44]
Pembrolizumab + 
Gem/Nab
PD-1
Advanced/Metastatic PDAC
15
N/A
20%
Weiss 
et al. 
[45]
MEDI4736
PD-L1
Advanced/Metastatic PDAC
10-
20
N/A
?%
Segal 
et al. 
[46]
CP-870,893
CD40
Advanced/Metastatic PDAC
22
82%
18%
Beatt
y et 
al. 
[47]
Despite the positive and encouraging results in HCC, clinical results appear different in patients 
with liver metastases. No single trial specifically targets patients with liver metastatic disease. 
However, the majority of trials testing ICIs included patients with liver metastasis from CRC 
(Table 2) and PDAC (Table 3). 
1.2.3
Tumor Microenvironment (TME)
It is critical to understand the mechanism of immune evasion during tumorigenesis in order to 
develop innovative and effective therapeutics based on cancer immunology. There is emerging 
data suggesting that the tumor microenvironment (TME) controls anti-tumor immunity by 
accumulation of regulatory T cells (Tregs), natural killer T cells (NKT), tumor associated 
macrophages (TAMs), indoleamine 2,3-dioxygenase (IDO) and myeloid derived suppressor cells 
(MDSCs) [48]. MDSCs form a critical link between the innate and adaptive immune system 
through their ability to influence the fate of Tregs in antigen specific T cell tolerance through 
diverse mechanisms [49]. MDSCs are a heterogeneous population of immature and 
immunosuppressive myeloid cells, which display a variety of pro-tumoral effects. They promote 
tumor angiogenesis through VEGF production[50] and subvert both innate and adaptive antitumor 
immunity[51]. MDSCs impair CD4+ and CD8+ T-cell responses via increased arginase activity, 

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leading to arginine depletion[52], and through the production of reactive oxygen and nitrogen 
species that disrupt TCR signaling[53]. MDSCs abrogate hepatic NK-cell activity via membrane-
bound TGF-β[54] and facilitate the expansion of Tregs cells and the induction of Tregs cells 
through IL-10 and TGF-β production. Increased numbers of CD14+ HLA-DR−/low MDSCs have 
been found in both tumor tissue and peripheral blood from patients with HCC, and elevated cell 
counts were related to tumor progression[50, 52]. Nevertheless, MDSCs affect T-cell function, 
survival and trafficking. MDSCs express galectin-9 that binds to TIM-3 on T cells, inducing T cell 
apoptosis [55]. MDSCs also express ADAM17 that down-regulates L-selectin (CD62L) levels on 
T cells, limiting their homing to lymph nodes and tumors[56]. Additionally, MDSCs interact with 
liver macrophages and causes their up-regulation of B7-H1, further strengthening the 
immunosuppressive phenotype[57]. MDSCs can also impair NK cell function. In human HCC, 
MDSCs (CD14+HLA-DR−/low) inhibit NK cell cytotoxicity and cytokine release, which is 
mediated by the NKp30 receptor[52]. Tumor-derived IL-1β induces Ly6C negative MDSCs which 
also inhibit NK cell development and function[58]. Together, MDSCs contribute to 
immunosuppressive network through multiple mechanisms and are potential immunotherapy 
targets for liver cancer. Therefore, the abundance of these immunosuppressive factors in the HCC 
or metastatic liver cancer TME calls for a multi-targeted approach based on solid rationale for 
synergy.
NKT cells represent another important cell type in the liver cancer TME. The liver has the largest 
number of NKT cells, which play critical roles in the pathogenesis of liver diseases and liver cancer 
[59]. NKT cells are a heterogeneous group of nonconventional T lymphocytes expressing both NK 
and T cell markers that share phenotypic and functional characteristics with NK cells. NKT cells 
not only recognize malignant cells directly and protect against tumor progression in a CD1d-
dependent manner[60], but also kill CD1d-expressing tumor-associated macrophages to suppress 
tumor growth[61]. We recently described that NKT cell activity is regulated by the ratio of primary 
bile acid to secondary bile acid, which can be modified by the gut microbiome[62]. Nevertheless, 
clinical evidence has showed high intratumoral NKT cells, high intratumoral IFN-g or combination 
of both high was associated with prolonged overall and recurrence-free survival in patients with 
HCC after curative resection[63]. Overall, it is generally believed that NKT cells play anti-tumor 
roles through the production of IFN-g promoting NK cell activation and killing tumor cells or 
tumor-associated macrophages. 
Therefore, we hypothesize that antitumor effect of ICIs will be boosted by augmenting NKT cells 
and suppressing MDSCs would be a promising synergistic strategy to treat liver cancer (Figure 
1).

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Figure 1: Tadalafil suppressed tumor growth and reduced tumor MDSC number. (A) RIL-175 
hepatoma cells (5x105/ 20μL) were orthotopically implanted into the livers of male B6(Cg)-Tyrc-2J/J mice. 
The establishment and growth of tumors were blindly monitored by BLI with the Xenogen IVIS. BLI 
represents proliferation rate through luciferase total flux signals. They were followed up for 21 days. 
Tadalafil (2mg/kg), a PDE5 inhibitor, was daily administered intraperitoneally in tumor bearing mice. (B) 
Analysis of BLI images was carried out by Living Image 2.50 software (PerkinElmer, Waltham, MA, 
USA). Calculated ROIs are graphed. (C) Accumulation of intrahepatic tumoral MDSCs was determined by 
flow cytometry.
1.2.4
PDE5 Inhibitors Augment Immunotherapy by Targeting on TME
Phosphodiesterase type 5 (PDE5) inhibitor, e.g. sildenafil and tadalafil, have been approved by the 
FDA for the treatment of pulmonary arterial hypertension, benign prostatic hyperplasia and erectile 
dysfunction, with a relative safe clinical profile. In the last decade, a significant number of studies 
have reported an increased expression of PDE5 in several human cancers compared to normal or 
surrounding non-neoplastic tissues[64-67]. Concomitantly, PDE5 inhibitors have been examined 
in multiple malignancies and cancer cell lines for their direct anticancer activities, for their efficacy 
as chemo-sensitizers and for cancer chemoprevention [68, 69]. Interestingly, the suppression of 
MDSCs by PDE5 inhibitor was well documented recently. In preclinical mouse models, it has 
been demonstrated that PDE5 inhibitor, sildenafil, and tadalafil significantly inhibited the MDSC 
functions by the downregulation of iNOS and ARG1 activities, leading to the activation of 
antitumor immunity and the prolongation of survival of tumor-bearing mice [70-72]. Recently, we 
found the treatment with a PDE5 inhibitor, tadalafil, caused suppression of tumor growth by 
reversing MDSC suppressor function (Figure 1). In 2015, two randomized, prospective, double 
blinded, placebo controlled, phase II clinical trials to determine the activity of PDE5 inhibitors on 
immune function in head and neck squamous cell carcinoma (HNSCC) patients were conducted[70, 
73] (Figure 2). Results showed that tadalafil can reverse tumor-specific immune suppression in 
these patients by lowering MDSCs and Tregs and increasing tumor-specific CD8+ T cells in a 
dose-dependent fashion, and augments general and tumor-specific immunity in HNSCC patients. 
In an open-labeled pilot study, the biologic effects, safety and efficacy of palliative treatment with 
tadalafil was assessed in patients with advanced metastatic melanoma [74]. Stable disease was 
achieved in 3/12 patients (25%). Median progression-free survival was 4.6 months, median overall 
survival 8.5 months. Stable patients displayed significantly higher numbers of CD8+ tumor 
infiltrating lymphocytes (TILs) in the center of metastases before treatment as compared with 
progressive patients. Upon therapy, they showed increased expression of ζ-chain (used as a marker 
of T cell activation) in CD8+and CD4+TILs and CD8+T cells in the peripheral blood as compared 
with baseline. Tadalafil improved clinical outcome of advanced melanoma patients by enhancing 
antitumor immunity. It was also demonstrated that the addition of tadalafil in a patient with end-
stage relapsed/refractory multiple myeloma demonstrated reduced MDSC function and generated 
a dramatic and durable anti-myeloma immune and clinical response[75]. All of these clinical 
studies elaborate the decreased amounts of MDSC by PDE5 inhibitor and their 
immunosuppressive pattern correlated with an increased T cell reactivity and improved clinical 
outcome of advanced cancer patients. Currently, there are two clinical trials ongoing to evaluate 
the efficacy of combination of PDE5 inhibitor with regorafenib on advanced solid tumor 
(NCT02466802) and with vaccine on HNSCC (NCT02544880), respectively. No clinical trial is 
evaluating the therapeutic effects of PDE5 inhibitor on liver cancer or liver metastases with ICIs. 

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Based on the available pre-clinical and clinical trial data, it is highly suggested PDE5 inhibitor 
likely has synergistic antitumor efficacy when combined with ICIs (Figure 1).
Figure 2: Tadalafil suppressed tumor growth by modulating tumor microenvironment. (A-C): 
tadalafil modulates tumor microenvironment. CD33/IL4Rα (A), CD4/FoxP3 (B), or CD8/CD69 (C) 
intratumoral concentration was evaluated by immune-fluorescence microscopy in HNSCC. D. 
Representative image of PET-CT scans from metastatic melanoma patient prior (1), after 4 cycles of 
ipilimumab (2), and 6 weeks later to exclude pseudo-progression (3); here progressive disease was 
confirmed, and the patient included into the TaMe trial; after 8 weeks of treatment with tadalafil 5 mg daily 
PO (4) and after 5 months (5). The scans show progressive disease under ipilimumab (1–3) and stable 
disease under tadalafil (3–5) – with a good regression of cutaneous metastases in the leg and a progressing 
left inguinal lymph node.
1.2.4.1
Tadalafil toxicity
Because clinical trials are conducted under widely varying conditions, adverse reaction rates 
observed in the clinical trials of a drug cannot be directly compared to rates in the clinical trials of 
another drug and may not reflect the rates observed in practice. Tadalafil was administered to over 
9000 men during clinical trials worldwide. In trials of tadalafil for once daily use, a total of 1434, 
905, and 115 were treated for at least 6 months, 1 year, and 2 years, respectively. For tadalafil for 
use as needed, over 1300 and 1000 subjects were treated for at least 6 months and 1 year, 
respectively. In addition, as discussed above, tadalafil has been administrated to study the efficacy 
to improve antitumor immunity with daily dose for short period times. 
1.2.4.1.1 Tadalafil for Use as Needed for Erectile Dysfunction (ED) 
In eight primary placebo-controlled clinical studies of 12 weeks duration, mean age was 59 years 
(range 22 to 88) and the discontinuation rate due to adverse events in patients treated with tadalafil 
10 or 20 mg was 3.1%, compared to 1.4% in placebo treated patients. 

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When taken as recommended in the placebo-controlled clinical trials, the following adverse 
reactions were reported (Table 4) for tadalafil for use as needed: 
Table 4: Treatment-Emergent Adverse Reactions Reported by ≥2% of Patients Treated with 
Tadalafil (10 or 20 mg) and More Frequent on Drug than Placebo in the Eight Primary Placebo-
Controlled Clinical Studies (Including a Study in Patients with Diabetes) for tadalafil for Use as 
Needed for ED
Adverse 
Reaction 
Placebo 
(N=476) 
Tadalafil 
5 
mg 
(N=151) 
Tadalafil 10 mg 
(N=394) 
Tadalafil 20 mg 
(N=635) 
Headache 
5% 
11% 
11% 
15% 
Dyspepsia 
1% 
4% 
8% 
10% 
Back pain 
3% 
3% 
5% 
6% 
Myalgia 
1% 
1% 
4% 
3% 
Nasal congestion 1% 
2% 
3% 
3% 
Flushing a 
1% 
2% 
3% 
3% 
Pain in limb 
1% 
1% 
3% 
3% 
a The term flushing includes: facial flushing and flushing 
1.2.4.1.2 Tadalafil for Once Daily Use for ED 
In three placebo-controlled clinical trials of 12 or 24 weeks duration, mean age was 58 years (range 
21 to 82) and the discontinuation rate due to adverse events in patients treated with tadalafil was 
4.1%, compared to 2.8% in placebo-treated patients. 
The following adverse reactions were reported (Table 5) in clinical trials of 12 weeks duration: 

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Table 5: Treatment-Emergent Adverse Reactions Reported by ≥2% of Patients Treated with 
tadalafil for Once Daily Use (2.5 or 5 mg) and More Frequent on Drug than Placebo in the Three 
Primary Placebo-Controlled Phase 3 Studies of 12 weeks Treatment Duration (Including a Study in 
Patients with Diabetes) for tadalafil for Once Daily Use for ED
Adverse Reaction 
Placebo 
(N=248) 
Tadalafil 
2.5 
mg 
(N=196) 
Tadalafil 
5 
mg 
(N=304) 
Headache 
5% 
3% 
6% 
Dyspepsia 
2% 
4% 
5% 
Nasopharyngitis 
4% 
4% 
3% 
Back pain 
1% 
3% 
3% 
Upper respiratory tract infection 1% 
3% 
3% 
Flushing 
1% 
1% 
3% 
Myalgia 
1% 
2% 
2% 
Cough 
0% 
4% 
2% 
Diarrhea 
0% 
1% 
2% 
Nasal congestion 
0% 
2% 
2% 
Pain in extremity 
0% 
1% 
2% 
Urinary tract infection 
0% 
2% 
0% 
Gastroesophageal reflux disease 0% 
2% 
1% 
Abdominal pain 
0% 
2% 
1% 
The following adverse reactions were reported (Table 6) over 24 weeks treatment duration in one 
placebo-controlled clinical study: 

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Table 6: Treatment-Emergent Adverse Reactions Reported by ≥2% of Patients Treated with 
tadalafil for Once Daily Use (2.5 or 5 mg) and More Frequent on Drug than Placebo in One Placebo-
Controlled Clinical Study of 24 Weeks Treatment Duration for tadalafil for Once Daily Use for ED
Adverse Reaction 
Placebo 
(N=94) 
Tadalafil 
2.5 
mg 
(N=96) 
Tadalafil 
5 
mg 
(N=97) 
Nasopharyngitis 
5% 
6% 
6% 
Gastroenteritis 
2% 
3% 
5% 
Back pain 
3% 
5% 
2% 
Upper respiratory tract infection 0% 
3% 
4% 
Dyspepsia 
1% 
4% 
1% 
Gastroesophageal reflux disease 0% 
3% 
2% 
Myalgia 
2% 
4% 
1% 
Hypertension 
0% 
1% 
3% 
Nasal congestion 
0% 
0% 
4% 
1.2.4.1.3 Tadalafil for Once Daily Use for Benign Prostate Hyperplasia (BPH) and for ED and 
BPH 
In three placebo-controlled clinical trials of 12 weeks duration, two in patients with BPH and one 
in patients with ED and BPH, the mean age was 63 years (range 44 to 93) and the discontinuation 
rate due to adverse events in patients treated with tadalafil was 3.6% compared to 1.6% in placebo-
treated patients. Adverse reactions leading to discontinuation reported by at least 2 patients treated 
with tadalafil included headache, upper abdominal pain, and myalgia. The following adverse 
reactions were reported (Table 7). 
Table 7: Treatment-Emergent Adverse Reactions Reported by ≥1% of Patients Treated with 
tadalafil for Once Daily Use (5 mg) and More Frequent on Drug than Placebo in Three Placebo-
Controlled Clinical Studies of 12 Weeks Treatment Duration, including Two Studies for Tadalafil 
for Once Daily Use for BPH and One Study for ED and BPH
Adverse Reaction 
Placebo (N=576) 
Tadalafil 5 mg (N=581) 
Headache 
2.3% 
4.1% 
Dyspepsia 
0.2% 
2.4% 
Back pain 
1.4% 
2.4% 
Nasopharyngitis 
1.6% 
2.1% 
Diarrhea 
1.0% 
1.4% 
Pain in extremity 
0.0% 
1.4% 

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Table 7: Treatment-Emergent Adverse Reactions Reported by ≥1% of Patients Treated with 
tadalafil for Once Daily Use (5 mg) and More Frequent on Drug than Placebo in Three Placebo-
Controlled Clinical Studies of 12 Weeks Treatment Duration, including Two Studies for Tadalafil 
for Once Daily Use for BPH and One Study for ED and BPH
Adverse Reaction 
Placebo (N=576) 
Tadalafil 5 mg (N=581) 
Myalgia 
0.3% 
1.2% 
Dizziness 
0.5% 
1.0% 
1.2.4.1.4 Tadalafil for Once Daily Use for HNSCC 
In a study of HNSCC with tadalafil to modify antitumor immunity, the following AEs were 
reported (Table 8) [73].
Table 8: Treatment-Emergent Adverse Reactions Reported with Tadalafil for Once Daily for at 
least 20 days
Possible
Probable
Symptoms
Placebo
10
20
Placebo
10
20
Back pain
2
Constipation
1
Cough
1
Diarrhea
1
1
Ear, nose and throat examination 
abnormal
1
Headache
1
2
1
Neck pain
1
2
Pain in extremity
1
Pulmonary
1
1
Sexual 
1
TOTAL AE grade 1
3
2
9
2
0
2
Grade 1
Total patients with AE grade 1
2
2
4
1
0
2
Back pain
1
Headache
1
Neck pain
1
Pain in extremity
1
1
Grade 2
Total AE grade 2
1
3
1

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Table 8: Treatment-Emergent Adverse Reactions Reported with Tadalafil for Once Daily for at 
least 20 days
Possible
Probable
Symptoms
Placebo
10
20
Placebo
10
20
Total patients with AE grade 2
1
1
1
Back pain
1
1
Myalgia
1
Total AE grade 3
1
2
Grade 3
Total patients with AE grade 3
1
2
Note: placebo (n=5); 10 mg arm (n=14); 10 mg arm (n=15). AE= Adverse Events 
Additional, less frequent adverse reactions (<1%) reported in the controlled clinical trials of 
tadalafil for BPH or ED and BPH included: gastroesophageal reflux disease, upper abdominal 
pain, nausea, vomiting, arthralgia, and muscle spasm. 
Back pain or myalgia was reported at incidence rates described in Table 4, Table 5, Table 6, 
Table 7, Table 8. In tadalafil clinical pharmacology trials, back pain or myalgia generally occurred 
12 to 24 hours after dosing and typically resolved within 48 hours. The back pain/myalgia 
associated with tadalafil treatment was characterized by diffuse bilateral lower lumbar, gluteal, 
thigh, or thoracolumbar muscular discomfort and was exacerbated by recumbency. In general, pain 
was reported as mild or moderate in severity and resolved without medical treatment, but severe 
back pain was reported with a low frequency (<5% of all reports). When medical treatment was 
necessary, acetaminophen or non-steroidal anti-inflammatory drugs were generally effective; 
however, in a small percentage of subjects who required treatment, a mild narcotic (e.g., codeine) 
was used. Overall, approximately 0.5% of all subjects treated with tadalafil for on demand use 
discontinued treatment as a consequence of back pain/myalgia. In the 1-year open label extension 
study, back pain and myalgia were reported in 5.5% and 1.3% of patients, respectively. Diagnostic 
testing, including measures for inflammation, muscle injury, or renal damage revealed no evidence 
of medically significant underlying pathology. In studies of tadalafil for once daily use, adverse 
reactions of back pain and myalgia were generally mild or moderate with a discontinuation rate of 
<1% across all indications. 
Across all studies with any tadalafil dose, reports of changes in color vision were rare (<0.1% of 
patients). 
The following adverse reactions have been identified during post approval use of tadalafil. Because 
these reactions are reported voluntarily from a population of uncertain size, it is not always possible 
to reliably estimate their frequency or establish a causal relationship to drug exposure. These 
events have been chosen for inclusion either due to their seriousness, reporting frequency, lack of 
clear alternative causation, or a combination of these factors. 
Cardiovascular and Cerebrovascular — Serious cardiovascular events, including myocardial 
infarction, sudden cardiac death, stroke, chest pain, palpitations, and tachycardia, have been 
reported post marketing in temporal association with the use of tadalafil. Most, but not all, of these 
patients had preexisting cardiovascular risk factors. Many of these events were reported to occur 

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during or shortly after sexual activity, and a few were reported to occur shortly after the use of 
tadalafil without sexual activity. Others were reported to have occurred hours to days after the use 
of tadalafil and sexual activity. It is not possible to determine whether these events are related 
directly to tadalafil, to sexual activity, to the patient’s underlying cardiovascular disease, to a 
combination of these factors, or to other factors.
Body as a Whole — hypersensitivity reactions including urticaria, Stevens-Johnson syndrome, 
and exfoliative dermatitis Nervous — migraine, seizure and seizure recurrence, transient global 
amnesia
Ophthalmologic — visual field defect, retinal vein occlusion, retinal artery occlusion
Non-arteritic anterior ischemic optic neuropathy (NAION), a cause of decreased vision including 
permanent loss of vision, has been reported rarely post marketing in temporal association with the 
use of phosphodiesterase type 5 (PDE5) inhibitors, including tadalafil. Most, but not all, of these 
patients had underlying anatomic or vascular risk factors for development of NAION, including 
but not necessarily limited to: low cup to disc ratio (“crowded disc”), age over 50, diabetes, 
hypertension, coronary artery disease, hyperlipidemia, and smoking. It is not possible to determine 
whether these events are related directly to the use of PDE5 inhibitors, to the patient’s underlying 
vascular risk factors or anatomical defects, to a combination of these factors, or to other factors. 
Otologic — Cases of sudden decrease or loss of hearing have been reported post marketing in 
temporal association with the use of PDE5 inhibitors, including tadalafil. In some of the cases, 
medical conditions and other factors were reported that may have also played a role in the otologic 
adverse events. In many cases, medical follow-up information was limited. It is not possible to 
determine whether these reported events are related directly to the use of tadalafil, to the patient’s 
underlying risk factors for hearing loss, a combination of these factors, or to other factors 
Urogenital — priapism
1.2.5
Gut Microbiome and Liver Cancer Control
Various bacteria populating in mammal gastrointestinal tract are an indispensable part in intestine 
ecosystem and play a pivotal role in gut barrier[76]. Commensal bacteria have been showed to 
regulate host immune system through the crosstalk with host intestinal epithelial cells and 
lymphatic cells[77]. In addition, the bacterial metabolism is another factor influencing the host 
immune homeostasis[77]. It is well known that oral antibiotics results in short- and long-term 
changes of the intestinal microbiome in humans[78, 79]. For instance, oral vancomycin is noted to 
reduce fecal microbial diversity with a decrease of gram-positive bacteria (mainly Firmicutes) and 
a compensatory increase in gram-negative bacteria (mainly Proteobacteria). Concurrently, 
vancomycin increases plasma primary bile acids with decreased fecal secondary bile acids that is 
predominantly associated with altered Firmicutes[80]. In line, this change might have implication 
on host immunity. 
The gut microbiome has emerged as a critical factor regulating anti-tumor immunity controlling 
the efficacy of chemo- and immunotherapies[81-85]. Most recently, it was noted that there were 
significant differences in the diversity and composition of the patient gut microbiome of 
responders versus non-responders to anti-PD-1 treatment in melanoma [84]. This study further 
found patient fecal microbiome samples (n = 43, 30 responders, 13 non-responders) significantly 
higher alpha diversity and relative abundance of bacteria of the Ruminococcaceae family in 
responding patients. The intestinal microbiome change by oral antibiotics may change the clinical 

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outcome to ICIs. It was found that antibiotics inhibited the clinical benefit of anti-PD-1 or anti-
PD-L1 in patients with advanced cancer. Fecal microbiota transplantation from responder into 
germ-free or antibiotic-treated mice ameliorated the antitumor effects of PD-1 blockade, whereas 
fecal microbiota transplantation from non-responder failed to do so. Surprisingly, analysis of 
patient stool samples at diagnosis revealed correlations between clinical responses to PD-1 
blockade and the relative abundance of Akkermansia muciniphila. Oral supplementation with A. 
muciniphila after Fecal microbiota transplantation with no responder feces restored the efficacy of 
PD-1 blockade in an interleukin-12–dependent manner by increasing the recruitment of 
CCR9+CXCR3+CD4+ T lymphocytes into mouse tumor beds[85]. These data have important 
implications for the treatment of cancer patients with ICIs.
Figure 3: Wildtype mice, were treated with ABX or H2O for 3 weeks (A). Then mice were given intra 
splenic B16 tumor cell injection. Liver metastases were measured. Representative images and liver surface 
metastatic tumor counts are shown. Liver metastasis was measured. Representative images, cumulative 
liver metastasis counts are shown. N=16 for H2O, 14 for ABX. p<0.05, one-way ANOVA.
Profound effects of the gut microbiome on HCC development have been described[86, 87]. 
Recently, we demonstrated that altering gut commensal bacteria by oral vancomycin induced a 
liver-selective anti-tumor effect[62] (Figure 3). An increase of hepatic CXCR6+ NKT cells was 
observed, independent of mouse strain, gender or presence of liver tumor. Hepatic NKT cells 
showed an activated phenotype and produced more IFNg upon antigen stimulation. In 
vivo functional studies confirmed that NKT cells mediated the liver-selective tumor inhibition. The 
data also showed that NKT cell accumulation was regulated by CXCL16 expression of liver 
sinusoidal endothelial cells, which was controlled by gut microbiome-mediated primary-to-
secondary bile acid conversion. Interestingly, previous study on healthy volunteer, treated with 
oral 
vancomycin, 
showed 
decreased 
fecal 
secondary 
bile 
acids 
with 
a 
simultaneous postprandial increase in primary bile acids in plasma (Figure 4)[80]. This finding 
indicates there are certain similarity between human and mice regarding the metabolic change of 
bile acid to oral vancomycin. Feeding secondary bile acids or colonization of bile acid-
metabolizing bacteria reversed both NKT cell accumulation and inhibition of liver tumor growth 
in mice with altered gut commensal bacteria. This study suggests a new link between gut bacteria-
controlled bile acid metabolism and liver anti-tumor immune-surveillance.

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Figure 4: Effect of antibiotics on plasma bile acid concentration. After vancomycin treatment the 
postprandial rise in plasma concentrations of primary bile acids is increased (A), whereas the concentration 
of secondary bile acids is decreased (B). Amoxicillin treatment did not have an effect on either postprandial 
primary (C) or secondary (D) plasma bile acids. Data are expressed as medians (n = 10 per group), ∗p <0.05.
1.2.6
Nivolumab 
PD-1 is a negative regulatory molecule that is expressed transiently following T-cell activation 
and on chronically stimulated T cells characterized by an “exhausted” phenotype. Subsequently, 
PD-1 positive T cells lose function and proliferative capacity while enhancing a suppressive tumor 
microenvironment. PD-1 may act together with other T-cell modulating molecules, including 
CTLA-4, TIM-3, lymphocyte-activation gene 3 (LAG-3) as well as indoleamine-pyrrole 2,3-
dioxygenase 1 (IDO-1), cytokines, and transforming growth factor beta. Two ligands specific for 
PD-1 have been identified: PD-ligand 1 (PD-L1, also known as B7-H1 or CD274, expressed on 
tumor, antigen-presenting cells [APCs], and dendritic cells [DCs]) and PD-L2 (also known as B7-
DC or CD273, expressed on endothelial cells). The interaction of PD- 1 with PD-L1 and PD-L2 
results in negative regulatory stimuli that down-modulate the activated T-cell immune response 
through SHP-1 phosphatase. PD-L1 expression is found on a number of tumors, and is associated 
with poor prognoses based on OS in many tumors[88]. 
Nivolumab (OPDIVO®, BMS-936558, MDX-1106, and ONO-4538) is a fully human monoclonal 
immunoglobulin G4 (IgG4) antibody (HuMAb) that is specific for human programmed death-1 
(PD-1, cluster of differentiation 279 [CD279]) cell surface membrane receptor[88]. Nivolumab 
binds to cynomolgus monkey PD-1 but not mouse, rat, or rabbit molecules. Clinical activity of 

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nivolumab has been evidenced by the recent FDA approval in patients with melanoma, non-small 
cell lung cancer (NSCLC), SCLC, renal cell carcinoma (RCC), HCC, dMMR CRC, HNSCC and 
urothelial cancer. Specifically, on the basis of CheckMate-040, FDA granted nivolumab 
accelerated approval in September 2017 for patients with advanced HCC previously treated with 
sorafenib. 
1.2.6.1
Pharmacokinetics 
Pharmacokinetics (PK) of nivolumab was linear in the range of 0.3 to 10 mg/kg, with dose- 
proportional increases in maximum serum concentration (Cmax) and area under the concentration- 
time curve from time zero to infinity (AUC0-∞), with low to moderate inter-subject variability 
observed at each dose level[88]. Clearance of nivolumab is independent of dose in the dose range 
(0.1 to 10 mg/kg) and tumor types studied. Body weight normalized dosing showed approximately 
constant trough concentrations over a wide range of body weights. The mean terminal elimination 
half-life of BMS-936558 is 17 to 25 days consistent with the half-life of endogenous IgG4. 
1.2.6.2
Efficacy 
In the multicenter phase I/II dose escalation and expansion study, CheckMate-040, total 214 
patients with advanced HCC and Child-Pugh score of 6 or less received nivolumab 3 mg/kg every 
2 weeks on the dose expansion phase[9]. Partial responses were observed in 15% and 20% of 
patients in the dose-escalation and dose-expansion phases, respectively. Disease control rates were 
64% and 58%, respectively. Responses were durable, averaging 17 months and 10 months, 
respectively.
In the monotherapy cohort of CheckMate-142, nivolumab provided durable responses with 31% 
investigator-assessed objective response rate (ORR) (median duration of response not yet reached 
with median follow-up of 12.0 months), 69% sustained disease control, 54% (9 months) and 50% 
(12 months) PFS, and 78% (9 months) and 73% (12 months) OS in previously treated patients with 
dMMR/MSI-H mCRC [26]. In the combination therapy cohort of CheckMate-142, at median 
follow-up of 13.4 months, nivolumab plus ipilimumab demonstrated 55% investigator-assessed 
ORR (95% CI, 45.2 to 63.8), and 80% disease control rate in the patients with dMMR/MSI-H 
mCRC. Median duration of response was not reached and most responses (94%) were ongoing at 
data cutoff. PFS rates were 76% (9 months) and 71% (12 months); respective OS rates were 87% 
and 85%. Statistically significant and clinically meaningful improvements were observed in 
patient-reported outcomes, including functioning, symptoms, and quality of life [27]. 
1.2.6.3
Toxicology 
A maximum tolerated dose (MTD) of nivolumab was not defined[29]. Serious adverse events 
(SAEs) occurred in 32 of 296 patients (11%) similar to the immune-related inflammatory events 
seen with ipilimumab: pneumonitis, vitiligo, colitis, hepatitis, hypophysitis, and thyroiditis (with 
noted pulmonary toxicity resulting in 3 deaths. Renal failure, symptomatic pancreatic and DM, 
neurologic events, vasculitis, insomnia and hematologic toxicity have also been reported). In 
combination with ipilimumab in the concurrent-regimen, grade 3 to 4 treatment-related adverse 
events (AEs) occurred in 32% of patients and were manageable. Patients (13%) who discontinued 
treatment because of study drug-related AEs had an ORR (63%) consistent with that of the overall 
population[27].

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1.2.6.4
Pharmacodynamics/Biomarkers 
Tumor-cell expression (melanoma) of PD-L1 was characterized in combination with ipilimumab 
with the use of IHC staining and pharmacodynamics changes in the peripheral-blood absolute 
lymphocyte count[89]. With PD-L1 positivity defined as expression in at least 5% of tumor cells, 
biopsy specimens from 21 of 56 patients (38%) were PD-L1–positive. Among patients treated with 
the concurrent regimen of nivolumab and ipilimumab, ORs were observed in patients with either 
PD-L1–positive tumor samples (6 of 13 patients) or PD-L1–negative tumor samples (9 of 22). In 
the sequenced regimen cohorts, a higher number of overall responses was seen among patients 
with PD-L1–positive tumor samples (4 of 8 patients) than among patients with PD-L1– negative 
tumor samples (1 of 13) suggesting the possibility that these tumors have higher response rates to 
the combination. The relationship between PDL-1 expression and responses may not be present in 
patients treated with the combination. Tissue expression of PDL-2, interferon-γ (IFN- γ), IDO, and 
T cell CD8+ are of current interest. Until more reliable data based on standardized procedures for 
tissue collection and assays are available, PD-L1 status cannot be used to select patients for 
treatment at this time. 
1.2.7
PDE5 Inhibitor and Oral Vancomycin in Combination with Nivolumab
Because the mechanisms of action of PDE5 inhibitor, oral vancomycin and PD-1 are non-
redundant targeting at MDSCs, NKT cells and PD-1 pathways as abovementioned, the 
combination may have additive or synergistic activity (Figure 5). There has no such treatment 
strategy developing or developed. 
Figure 5: Concept of the protocol.
1.2.7.1
PDE5 inhibitor and oral vancomycin in combination with nivolumab regimen dose 
justification
Per the FDA approval for HCC treatment with nivolumab, patients can be treated with 240 mg 
every 2 weeks or 480 mg every 4 weeks for HCC. 
As discussed above, patients with HNSCC received tadalafil 20 mg once a day (20 mg daily) for 
at least 10 days demonstrated augmentation of general and tumor-specific immunity[70]. However, 

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in the other study, it seemed that tadalafil immunomodulatory activity was maximized at an 
intermediate dose (10 mg daily) but not at higher doses (20 mg daily)[73]. The potential 
mechanism is a possible off-target effect on PDE11 at high dosages that, by increasing intracellular 
cAMP, may negatively affect antitumor immunity.
Vancomycin is poorly absorbed after oral administration. During multiple dosing of 250 mg every 
8 hours for 7 doses, fecal concentrations of vancomycin in volunteers exceeded 100 mg/kg in the 
majority of samples. No blood concentrations were detected, and urinary recovery did not exceed 
0.76%. Oral vancomycin remains the mainstay of therapy for severe infections produced 
by Clostridium difficile, the most prevalent cause of healthcare-associated infectious diarrhea in 
developed countries. Its short- and long-term effects on the human intestinal microbiota remain 
largely unknown, however, 125 mg 4 times daily for 10 days has been used as initial standard dose 
for non-severe Clostridium difficile infection. 
Since there are no previous reports regarding this combination therapy in cancer patients, we will 
choose the FDA approved standard dose and previous reported single agent dose as initial one. 
The dose may need titrate for the future study after the complicate analysis after this trial. 
2
ELIBILITY ASSESSMENT AND ENROLLMENT
2.1
Eligibility Criteria
2.1.1
Inclusion Criteria

Patients must have 
histopathological confirmation of HCC (Cohort 1)
OR
histopathological confirmation of carcinoma in the setting of clinical and radiological 
characteristics which, together with the pathology, are highly suggestive of a diagnosis 
of HCC (Cohort 1)
OR
histopathological confirmation of advanced colorectal or pancreatic malignancy with 
liver involvement as dominant site of metastasis (Per multidiscipline tumor board 
review and approval) (Cohort 2).

Patients must have disease that is not amenable to potentially curative resection, 
transplantation or ablation. 

Patients must have progressed on, been intolerant to, or refused prior sorafenib/lenvatinib 
and/or atezolizumab/bevacizumab therapy (Cohort 1 only). 

Subjects must have progressed on or after standard systemic chemotherapy (at least one 
line of chemotherapy for patients with liver metastasis from PDAC, at least two lines of 
chemotherapy for patients with liver metastasis from CRC) (Cohort 2 only).

Patients must have evaluable or measurable disease per RECIST 1.1 (See Section 6.3.1). 

Patients must have lesion accessible for biopsy and be willing to undergo pre- and post-
treatment biopsies.

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
ECOG performance status of 0 to 1 (Appendix A)

If liver cirrhosis is present, patient must have a Child-Pugh score 7 (Appendix F)

Active chronic HBV infected subjects must be on antivirals and have HBV DNA 
<100IU/mL. HCV infected subjects can be enrolled with close HCV RNA level 
monitoring. 

Age > 18 years. Because no dosing or adverse event data are currently available on the 
use of nivolumab in combination with tadalafil and vancomycin in patients <18 years of 
age, children are excluded from this study, but will be eligible for future pediatric trials

Adequate hematological function defined by:
white blood cell (WBC) count ≥ 3 × 109/L 
absolute neutrophil count (ANC) ≥ 1.5 ×109/L, 
lymphocyte count ≥ 0.5 × 109/L, 
platelet count ≥ 60 × 109/L, and 
Hgb ≥9 g/ dL (more than 48 hours post-completion of blood transfusion))

Adequate hepatic function defined by:
a total bilirubin level ≤ 1.5 × ULN, 
an AST level <5×ULN, 
an ALT level <5 ×ULN. 

Adequate renal function defined by:
Creatinine clearance (CrCl) >50 mL/min/1.73 m2 by 24 hours urine collection or as 
predicted by the Cockcroft-Gault formula:
                 CrCl = (140−age (y)) x (weight in kg) x (0.85, if female) x1.73 m2
                                     72 x Serum Creatinine (mg/dL) x pt.’s BSA (m2)

The effects of nivolumab on the developing human fetus are unknown. For this reason, 
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 and up to 5 months (women) and 7 months (men) after the 
last dose of the drug. 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.

Patients with a history of cardiovascular disease may be enrolled per cardiology 
consultation and approval with echocardiogram and troponin level in normal range at the 
time of enrollment.

Patient must be able to understand and willing to sign a written informed consent 
document.

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2.1.2
Exclusion Criteria

Patients who have had standard-of-care anti-cancer therapy or therapy with investigational 
agents (e.g. chemotherapy, immunotherapy, endocrine therapy, targeted therapy, biologic 
therapy, tumor embolization, monoclonal antibodies or other investigation agents), large 
field radiotherapy, or major surgery within 4 weeks prior to enrollment. 

Therapy with antibiotics within 30 days prior to enrollment. 

Therapy with nitrates, alpha-blockers, or cytochrome P450 (CYP3A4) inhibitors within 7-
days prior to enrollment and for whom stopping is unsafe and/or a safe substitute is not 
medically recommended. Use of PDE5 inhibitors such as vardenafil (Levitra®), tadalafil 
(Cialis®), and sildenafil citrate (Viagra®) ≤15-days prior to enrollment.

The patient must not be currently on a corticosteroid dose greater than physiologic 
replacement dosing defined as 10 mg of cortisone per day or its equivalent.

For PDAC patients with liver metastases, primary PDAC has not been resected (unless the 
primary is in the tail of the pancreas).

Patients with known brain metastases will 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.

Have signs of liver failure, e.g. clinically significant ascites, encephalopathy, or variceal 
bleeding within 6 months prior to enrollment.

Prior major liver resection: remnant liver <50% of the initial liver volume. Patients with a 
biliary stent can be included. 

Patients with active autoimmune disease or history of autoimmune disease that might recur, 
which may affect vital organ function or require immune suppressive treatment including 
systemic corticosteroids. These include but are not limited to patients with a history of 
immune related neurologic disease, multiple sclerosis, autoimmune (demyelinating) 
neuropathy, Guillain-Barre syndrome or CIDP, myasthenia gravis; systemic autoimmune 
disease such as SLE, connective tissue diseases, scleroderma, inflammatory bowel disease 
(IBD), Crohn’s, ulcerative colitis, hepatitis; and patients with a history of toxic epidermal 
necrolysis (TEN), Stevens-Johnson syndrome, or phospholipid syndrome. Such diseases 
should be excluded because of the risk of recurrence or exacerbation of disease. 
Of note, patients with vitiligo, endocrine deficiencies including thyroiditis managed 
with replacement hormones including physiologic corticosteroids are eligible. Patients 
with rheumatoid arthritis and other arthropathies, Sjogren’s syndrome and psoriasis 
controlled with topical medication and patients with positive serology, such as 
antinuclear antibodies (ANA), anti-thyroid antibodies should be evaluated for the 
presence of target organ involvement and potential need for systemic treatment but 
should otherwise be eligible. 

Have history of idiopathic pulmonary fibrosis (including bronchiolitis obliterans with 
organizing pneumonia) or evidence of active pneumonitis on screening chest CT scan.

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
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. 

Patients with myocardial infarction or myocarditis within 12 months prior to enrollment.

History of severe or unstable cerebrovascular disease.

Sustained hypotension (<90/50 mmHg) or uncontrolled hypertension (>160/100 mmHg)

Stroke within 6 months prior to enrollment.

HIV-positive patients are excluded because HIV causes complicated immune deficiency 
and study treatment can possess more risks for these patients. 

Have had prior transplant of any kind. 

Have ascites.

History of allergic reactions attributed to compounds of similar chemical or biologic 
composition to nivolumab, tadalafil or vancomycin.

History of severe hypersensitivity reaction to any monoclonal antibody.

Prior invasive malignancy (except non-melanomatous skin cancer) unless disease free for 
a minimum of 3 years prior to enrollment.

Pregnant women are excluded from this study because nivolumab’s potential for 
teratogenic or abortifacient effects is unknown. Because there is an unknown but potential 
risk for adverse events in nursing infants secondary to treatment of the mother with 
nivolumab, breastfeeding should be discontinued if the mother is treated with nivolumab 
2.1.3
Recruitment Strategies
The study will be posted on the CCR website and on clinicaltrials.gov. NIH social media platforms 
may also be used to notify potential subjects of the study.  Outside providers and colleagues may 
directly refer patients for screening into this study.
2.2
Screening Evaluation
2.2.1
Screening activities performed after a consent for screening has been signed
The following activities will be performed only after the subject has signed the consent for study 
#01-C-0129 on which screening activities will be performed. Assessments performed at outside 
facilities or on another NIH protocol within the timeframes below may also be used to determine 
eligibility once a patient has signed the consent.
Studies should be done within 28 days prior to enrollment unless otherwise noted below.

Complete Medical History and Physical Evaluation (including height, weight, vital signs, 
and ECOG performance status).

Laboratory Evaluation
o
Hematological Profile: CBC with differential and platelet count.

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o
Biochemical Profile: electrolytes, BUN, creatinine, AST, ALT, total and direct 
bilirubin, calcium, phosphorus, albumin, magnesium. 
o
PT, INR, PTT, fibrinogen (if clinically indicated)
o
24-hour urine collection (if creatinine clearance is tested this way)
o
ACTH 
o
morning cortisol
o
Thyroid tests TSH, T3, T4
o
Uric acid, amylase and lipase
o
Tumor marker CEA, AFP, CA19-9
o
Serum or urine pregnancy test for female participants of childbearing potential (within 
2 weeks prior to enrollment). 
o
HIV, Hepatitis B and C serology and/or viral load 
o
TB testing (if clinically indicated)
o
Troponin I (if clinically indicated)
o
Urinalysis

Cardiology Consultation (if clinically indicated)

Echocardiogram (if clinically indicated)

EKG

CT scan of chest, abdomen and pelvis (or MRI of abdomen if clinically indicated)

Documentation of histologic cancer confirmation (at any time point prior to enrollment; 
required from each participant to confirm diagnosis). If there is no available documentation, 
biopsy will be performed to confirm the diagnosis.
2.3
Participant Registration and Status Update Procedures
Registration and status updates (e.g. when a participant is taken off protocol therapy and when a 
participant is taken off-study) will take place per CCR SOP ADCR-2, CCR Participant 
Registration & Status Updates found here.
2.3.1
Treatment Assignment Procedures (for registration purposes only)
Cohorts
Number
Name
Description
1
Cohort 1
Patients with HCC or patients with carcinoma highly suggestive of a 
diagnosis of HCC
2
Cohort 2
Patients with liver metastases from CRC or PDAC
Arms

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Number
Name
Description
1
Arm 1
Nivolumab, tadalafil and oral vancomycin
Treatment Assignment
Patients in Cohort 1 and 2 will be directly assigned to treatment in Arm 1
2.4
Baseline Evaluation
Tests performed during screening do not need to be repeated if done in designated time frame prior 
to start of study treatment.
Within 28 days prior to first dose:

CT scan of chest, abdomen and pelvis (or MRI of abdomen if clinically indicated) 

HLA subtype- HLA-A, B, C and sequenced based HLA-A-02* (any time prior to treatment)

Baseline research biopsy (does not need to be repeated if biopsy was done during screening)
Within 3 days prior to first dose:

Concomitant medications

Baseline signs and symptoms evaluation

Stool collection or rectal swab for bile acid

Research blood for mixed meal test for bile acid and for whole exome panel
3
STUDY IMPLEMENTATION
3.1
Study Design
The proposed study is an open label, single-arm study of nivolumab, oral vancomycin and tadalafil 
in patients with refractory primary HCC or liver dominant metastatic cancer from CRC or PDAC, 
see Table 9 and Schema 1. 
Treatment will be delivered in cycles consisting of 4 weeks (+/- 3 days). 
Treatment will continue until off treatment criteria are met (see Section 3.6.1).
If one or two study drugs permanently discontinued because of toxicity, treatment with the other 
drug/s will continue.
Patients will be monitored every 8 (+/- 1) weeks with imaging per Study Calendar.
Patients will have two mandatory research biopsies: at baseline and anytime during week 3 of 
Cycle 2 while patient is still on oral vancomycin. (If patient’s disease progresses before week 3 of 
cycle 2, the post-treatment biopsy may be performed per PI discretion at the time of progression).
Table 9: Treatment regimen
Nivolumab
Tadalafil
Oral Vancomycin

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Table 9: Treatment regimen
480 mg intravenously 
every 4 weeks
10 mg PO daily
 125 mg every 6-hour PO from week 1 to week 3, week 4 
off, 500 mg total daily
Schema 1: Treatment and monitoring schedule
3.2
Drug Administration 
3.2.1
Nivolumab Administration
Nivolumab will be given on Day 1 of every cycle at a flat dose of 480 mg. Nivolumab will be 
administered over approximately 30 to 60-minutes via IV infusion (See section 14.1.5). 
3.2.2
Tadalafil Administration
Tadalafil will be given orally at 10 mg dose daily. 
Patients will begin tadalafil dosing on the Day 1 of Cycle 1 and will continue to receive tadalafil 
on every day of every cycle.
In case of missing dose or vomiting after taking tadalafil, patients will be instructed not to increase 
next day dosage and not to repeat taking tadalafil on this day.
Patients will complete and return Patient’s Diary (Appendix B).
Missing of 20% of the doses for any given cycle is allowed.
3.2.3
Vancomycin Administration
Vancomycin will be given orally 125 mg every 6 hours at 500 mg total daily dose.
Patients will begin vancomycin dosing on the Day 1 of Cycle 1 and will continue to receive 
vancomycin from week 1 to week 3 with week 4 off treatment on every cycle. 
If vancomycin administered earlier than scheduled or missed, dose can be taken in the time period 
of +/- 2 hours of scheduled time. In case of vomiting after taking vancomycin, next administration 
of vancomycin should proceed as scheduled.  

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Patients will complete and return Patient’s Diary (Appendix B).
Missing of 20% of the doses for any given cycle is allowed.
3.2.4
Sequence and Monitoring of Dose Administration 
On day 1 of every cycle tadalafil and vancomycin can be administered at any time prior to or after 
nivolumab injection. 
Vital signs will be collected within 1 hour before nivolumab infusions, at least once during the 
infusions, and within 30 minutes after the completion of the infusion. 
In the event of a ≤Grade 2 infusion-related reaction, the infusion rate of study drug may be 
decreased by 50% or interrupted until resolution of the event and re-initiated at 50% of the initial 
rate until completion of the infusion. For patients with a ≤Grade 2 infusion related reaction, 
subsequent infusions must be administered at 50% of the initial rate. Acetaminophen and/or an 
antihistamine (e.g. diphenhydramine) or equivalent medications per institutional standard may be 
administered at the discretion of the investigator. If the infusion related reaction is ≥Grade 3 or 
higher in severity, study drug will be discontinued. See also Appendix D
As with any antibody, allergic reactions to dose administration are possible. Therefore, appropriate 
drugs and medical equipment to treat acute anaphylactic reactions must be immediately available, 
and study personnel must be trained to recognize and treat anaphylaxis, as per local institutional 
guidelines.
3.3
Dose Delay or Modifications
3.3.1
General Guidance 
When, at the beginning of a treatment cycle, treatment delay related to either tadalafil or 
vancomycin treatment alone is indicated, treatment with nivolumab may not be delayed. If 
treatment delay is due to nivolumab only, tadalafil and vancomycin treatment may be continued 
without delay.
If, in the opinion of the investigator, a toxicity is considered to be due solely to one drug (e.g. 
flushing by tadalafil, dysgeusia by oral vancomycin), the dose of the other drugs does not require 
modification.
3.3.2
Nivolumab
Dose modifications of nivolumab may be required in the event of treatment-related toxicity. 
General guidelines regarding dose modification are provided in Appendix C. 
3.3.2.1
Adverse Events of Special Interest for Nivolumab
An adverse event of special interest (AESI) is one of scientific and medical interest specific to 
understanding of the Investigational Product and may require close monitoring. An AESI may be 
serious or non-serious. 
AESIs for nivolumab include but are not limited to events with a potential inflammatory or 
immune-mediated mechanism and which may require more frequent monitoring and/or 
interventions such as steroids, immunosuppressants and/or hormone replacement therapy. These 
AESIs are being closely monitored in clinical studies with nivolumab. An immune-mediate 
adverse event (imAE) is defined as an adverse event that is associated with drug exposure and is 
consistent with an immune-mediated mechanism of action and where there is no clear alternate 

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aetiology. Serologic, immunologic, and histologic (biopsy) data, as appropriate, should be used to 
support an imAE diagnosis. Appropriate efforts should be made to rule out neoplastic, infectious, 
metabolic, toxin, or other etiologic causes of the imAE. 
AESIs observed with nivolumab include: 

Diarrhea / Colitis

Pneumonitis / ILD

ALT/AST increases / hepatitis / hepatotoxicity

Neuropathy / neuromuscular toxicity (e.g. Guillain-Barré, and myasthenia gravis)

Endocrinopathies (i.e. events of hypophysitis, hypopituitarism adrenal insufficiency, 
diabetes insipidus, hyper- and hypothyroidism and type I diabetes mellitus)

Rash / Dermatitis

Nephritis / Blood creatinine increases

Pancreatitis (or labs suggestive of pancreatitis - increased serum lipase, increased serum 
amylase) 

Other inflammatory responses that are rare with a potential immune-mediated etiology 
include, but are not limited to, myocarditis, pericarditis, and uveitis.
In addition, infusion-related reactions and hypersensitivity/anaphylactic reactions with a different 
underlying pharmacological etiology are also considered AESIs.
More specific guidelines for AESIs evaluation and treatment are described in detail in the Dosing 
Modification and Toxicity Management Guidelines (see Appendix C).
If new or worsening pulmonary symptoms (e.g. dyspnea) or radiological abnormality suggestive 
of pneumonitis/interstitial lung disease is observed, toxicity management as described in detail in 
the Dosing Modification and Toxicity Management Guidelines (see Appendix C) will be applied. 
It is strongly recommended to perform a full diagnostic workup, to exclude alternative causes such 
as lymphangitic carcinomatosis, infection, allergy, cardiogenic edema, or pulmonary hemorrhage. 
In the presence of confirmatory HRCT scans where other causes of respiratory symptoms have 
been excluded, a diagnosis of pneumonitis (ILD) should be considered and the Dosing 
Modification and Toxicity Management Guidelines should be followed. 
For nivolumab, AESIs will comprise the following: 

Pneumonitis: AEs of pneumonitis are also of interest, as pneumonitis has been observed 
with use of anti-PD-1 mAbs (but not with anti-PD-L1 mAbs). Initial work-up should 
include a high-resolution CT scan, ruling out infection, and pulse oximetry. Pulmonary 
consultation is highly recommended. Guidelines for the management of patients with 
immune-mediated AEs (imAEs) including pneumonitis are provided in Appendix C. 

Pneumonitis (ILD): The following assessments, and additional assessments if required, 
will be performed to enhance the investigation and diagnosis of potential cases of 
pneumonitis. The results of the assessment will be collected.

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Physical examination: Signs and symptoms (cough, shortness of breath and pyrexia, 
etc.) including auscultation for lung field will be assessed.
SpO2: Saturation of peripheral oxygen (SpO2)
Other items: When pneumonitis (ILD) is suspected during study treatment, the 
following markers should be measured where possible: 
o
ILD Markers (KL-6, SP-D) and β-D-glucan

Tumor markers: Particular tumor markers which are related to disease progression. 

Additional Clinical chemistry: CRP, LDH

Infusion reactions: AEs of infusion reactions (also termed infusion-related reactions) are 
of special interest to BMS and are defined, for the purpose of this protocol, as all AEs 
occurring from the start of nivolumab infusion up to 48 hours after the infusion start time. 
For all infusion reactions, SAEs should be reported to BMS Patient safety.

Hypersensitivity reactions: Hypersensitivity reactions as well as infusion-related reactions 
have been reported with anti PD-L1 and anti-PD-1 therapy [90]. As with the administration 
of any foreign protein and/or other biologic agents, reactions following the infusion of 
mAbs can be caused by various mechanisms, including acute anaphylactic (IgE-mediated) 
and anaphylactoid reactions against the mAbs and serum sickness. Acute allergic reactions 
may occur, may be severe, and may result in death. Acute allergic reactions may include 
hypotension, dyspnea, cyanosis, respiratory failure, urticaria, pruritus, angioedema, 
hypotonia, arthralgia, bronchospasm, wheeze, cough, dizziness, fatigue, headache, 
hypertension, myalgia, vomiting, and unresponsiveness. Guidelines for the management of 
patients with hypersensitivity (including anaphylactic reaction) and infusion-related 
reactions are provided in Appendix D. 

Hepatic function abnormalities (hepatotoxicity): nivolumab induced liver injury is defined 
as concurrent ALT 10 × ULN AND total bilirubin 2 × ULN or baseline value (if elevated 
total bilirubin at study entry), AND no other immediately apparent possible causes of ALT 
elevation and hyperbilirubinemia, including, but not limited to, tumor progression, acute 
viral hepatitis, cholestasis, pre-existing hepatic disease or the administration of other 
drug(s), herbal medications and substance known to be hepatotoxic. Guidelines for 
management of patients with hepatic function abnormality are provided in Appendix C. 

Gastrointestinal disorders: Diarrhea/colitis is one of the most commonly observed 
treatment emergent SAE when nivolumab is used. In rare cases, colon perforation may 
occur that requires surgery (colectomy) or can lead to a fatal outcome if not properly 
managed. Guidelines on management of diarrhea and colitis in patients receiving 
nivolumab are provided in Appendix C.

Endocrine disorders: Immune-mediated endocrinopathies include hypophysitis, adrenal 
insufficiency, and hyper- and hypothyroidism. Guidelines for the management of patients 
with immune-mediated endocrine events are provided in Appendix C.

Pancreatic disorders: Immune-mediated pancreatitis includes autoimmune pancreatitis, and 
lipase and amylase elevation. Guidelines for the management of patients with immune-
mediated pancreatic disorders are provided in Appendix C.

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
Neurotoxicity: Immune-mediated nervous system events include encephalitis, peripheral 
motor and sensory neuropathies, Guillain-Barré, and myasthenia gravis. Guidelines for the 
management of patients with immune-mediated neurotoxic events are provided in 
Appendix C.

Nephritis: Consult with Nephrologist. Monitor for signs and symptoms that may be related 
to changes in renal function (e.g. routine urinalysis, elevated serum BUN and creatinine, 
decreased creatinine clearance, electrolyte imbalance, decrease in urine output, proteinuria, 
etc.) 

Patients should be thoroughly evaluated to rule out any alternative etiology (e.g. disease 
progression, infections etc.)

Steroids should be considered in the absence of clear alternative etiology even for low 
grade events (Grade 2), in order to prevent potential progression to higher grade event. 
Guidelines for the management of patients with immune-mediated neurotoxic events are 
provided in Appendix C.
3.3.2.2
Nivolumab Dose Delay Criteria
Dose delay criteria apply for all nivolumab-related adverse events. Treatment delay up to 8 weeks 
from the last dose is allowed. If treatment is not resumed during 8 weeks, patients will be taken 
off nivolumab treatment.
After nivolumab discontinuation, the patient can continue with tadalafil and vancomycin treatment.
Nivolumab administration should be delayed for the following:

Any grade ≥ 2 non-skin, drug-related adverse event, with the following exceptions: Grade 
2 drug-related fatigue or laboratory abnormalities do not require a treatment delay 

Any grade 3 skin, drug-related adverse event, any grade 3 drug-related laboratory 
abnormality, with the following exceptions for asymptomatic amylase or lipase, ALT, or 
AST: 
Grade 3 amylase or lipase abnormalities that are not associated with symptoms or 
clinical manifestations of pancreatitis do not require a dose delay. 
If a subject has a baseline AST or ALT that is within normal limits, delay dosing 
for drug-related Grade ≥ 2 toxicity (2-grade shift). 
If a subject has baseline AST or ALT within the grade 1 toxicity range, delay dosing 
for drug-related Grade ≥ 3 toxicity (2-grade shift). 
If a subject has baseline AST or ALT within the Grade 2 toxicity range, delay 
dosing for drug-related increase in AST or ALT at 2x baseline value or when AST 
or ALT is 8x ULN (whichever is lower). 
Any adverse event, laboratory abnormality, or intercurrent illness which, in the 
judgment of the investigator, warrants interrupting the dose of study medication. 
subjects who require delay of nivolumab should be re-evaluated weekly or more 
frequently if clinically indicated. It is recommended to monitor elevations in AST 
or ALT approximately every 3 days till levels peak or begin to decline. Nivolumab 

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dosing can be resumed when re-treatment criteria are met. Tumor assessments for 
all subjects should continue as per protocol even if study drug dosing is delayed.
Resumption after delay is discussed in section 3.3.2.4 below.
3.3.2.3
Nivolumab Dose Discontinuation Criteria
Nivolumab administration should be permanently discontinued if at least one of the following 
drug-related adverse event(s) occurs: 

Any ≥ Grade 2 drug-related uveitis, eye pain, or reduction of visual acuity that does not 
respond to topical therapy and does not improve to Grade 1 severity within 2 weeks of 
starting therapy OR requires systemic treatment 

Any Grade 3 non-skin, drug-related adverse event lasting > 7 days, with the following 
exceptions: 
Grade 3 drug-related uveitis, pneumonitis, bronchospasm, neurological toxicity, 
hypersensitivity reaction, or infusion reaction of any duration requires 
discontinuation. 
Grade 3 drug-related endocrinopathies adequately controlled with only physiologic 
hormone replacement do not require discontinuation. 
Grade 3 drug-related laboratory abnormalities do not require treatment 
discontinuation except: 
Grade 3 drug-related thrombocytopenia > 7 days or associated with bleeding 
requires discontinuation 
Any drug-related liver function test (LFT) abnormality that meets the following 
criteria require discontinuation: 
o
AST or ALT > 10 x ULN for > 2 weeks, 
o
AST or ALT > 15 x ULN irrespective of duration, 
o
T. bilirubin > 8 x ULN irrespective of duration for subjects with elevated 
bilirubin at study entry or > 5 x ULN for those with normal T bilirubin at 
entry, 
o
Concurrent AST or ALT > 3 x ULN and T. bilirubin > 5 x ULN for subjects 
entering treatment with a normal bilirubin and up to 8 x ULN for subjects 
with elevated bilirubin 

Any drug-related Grade 4 drug-related adverse event or laboratory abnormality, except for 
the following events which do not require discontinuation: 
Grade 4 neutropenia < 7 days 
Grade 4 lymphopenia or leukopenia 
Grade 4 amylase or lipase abnormalities that are not associated with symptoms or 
clinical manifestations of pancreatitis. 

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Isolated Grade 4 electrolyte imbalances/abnormalities that are not associated with 
clinical sequelae and are corrected with supplementation/appropriate management 
within 72 hours of their onset 
For Grade 4 endocrinopathy adverse events such as hyper- or hypothyroidisms, or 
glucose intolerance, which resolve or are adequately controlled with physiologic 
hormone replacement (steroids, thyroid hormones) or glucose controlling agents, 
respectively 

Dosing delays to allow for prolonged steroid tapers to manage drug-related adverse events 
are allowed. Tumor assessments should continue as per protocol even if dosing is delayed. 
Periodic study visits to assess safety and laboratory studies should also continue every 6 
weeks or more frequently if clinically indicated during such dosing delays. 

Dosing delays lasting > 6 weeks from the previous dose that occur for non-drug-related 
reasons may be allowed. Tumor assessments should continue as per protocol even if dosing 
is delayed. Periodic study visits to assess safety and laboratory studies should also continue 
every 6 weeks or more frequently if clinically indicated during such dosing delays. 

Any adverse event, laboratory abnormality or intercurrent illness which, in the judgment 
of the Investigator, presents a substantial clinical risk to the subject with continued 
nivolumab dosing.
3.3.2.4
Nivolumab Dose Resumption Criteria
Subjects may resume treatment with study drug when the drug-related AE(s) resolve to Grade ≤ 1 
or baseline value, with the following exceptions:

Subjects may resume treatment in the presence of Grade 2 fatigue. 

Subjects who have not experienced a Grade 3 drug-related skin AE may resume treatment 
in the presence of Grade 2 skin toxicity. 

Subjects with baseline Grade 1 AST, ALT, or total bilirubin who require dose delays for 
reasons other than a drug-related hepatic event may resume treatment in the presence of 
Grade 2 AST, ALT, or total bilirubin. 

Subjects who require dose delays for drug-related increased AST, ALT, or bilirubin may 
resume treatment when hepatic parameters are at baseline or Grade 1. 

Subjects with AST, ALT or bilirubin values meeting discontinuation parameters (Section 
3.3.2.3) should have treatment permanently discontinued. 

Drug-related pulmonary toxicity, diarrhea, or colitis must have resolved to baseline before 
treatment is resumed. 

Drug-related endocrinopathies adequately controlled with only physiologic hormone 
replacement may resume treatment. 

Subjects who delay study treatment due to any Grade ≥ 3 amylase or lipase abnormality 
that is not associated with symptoms or clinical manifestations of pancreatitis, and that is 
assessed by the investigator  not to be related to nivolumab, may resume nivolumab when 
the amylase or lipase abnormality has resolved to Grade < 3. 

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Resuming treatment after a delay of up to 8 weeks from the previous dose is permitted with the
following exceptions:

Dosing delays to allow for prolonged steroid tapers to manage drug-related adverse events 
are allowed. Tumor imaging assessments should continue as per protocol even if dosing is 
interrupted. 

Dosing delays > 8 weeks from the last dose that occur for non-drug-related reasons may be 
allowed. 
3.3.3
Tadalafil
If patient’s CrCl is reduced to 30 - 50 mL/minute (as measured by the Cockcroft-Gault formula), 
the dose of tadalafil will be reduced to 5 mg daily.
If patient’s CrCl is reduced less than 30 mL/minute, tadalafil will be held until the renal function 
improved:
to 30 - 50 mL/minute, tadalafil will resume at 5 mg daily;
to > 50 mL/minute, tadalafil will resume at 10 mg daily;   
If renal function is not improved after 4 weeks, tadalafil will be permanently discontinued.
If patient experiences priapism, hearing loss, vision change, or hypotension that is associated with 
the use of tadalafil, tadalafil will be permanently discontinued.
If tadalafil is permanently discontinued, treatment with nivolumab and vancomycin will continue.
3.3.4
Oral Vancomycin
Vancomycin will be held temporally for adverse events > grade 1 confirmed by PI as associated 
with oral vancomycin:
nephrotoxicity 
ototoxicity 
uncontrolled nausea, diarrhea or abdominal pain. 
Vancomycin can be resumed at full dose after toxicity resolves to baseline. If toxicity is not 
resolved for 4 weeks, vancomycin will be permanently discontinued.
Vancomycin will be discontinued if patient develops anaphylactoid reactions that is confirmed to 
be associated with oral vancomycin.
If vancomycin is permanently discontinued, treatment with nivolumab and tadalafil will continue.

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3.4
Study Calendar
Screening1
Baseline1
All Cycles
Day 1 1
28 
Days 
Safety 
FU11,13
Long 
Term 
FU12, 13
Nivolumab2
X
Tadalafil3
X
Oral vancomycin4 
X
Medical History
X
Height
X
Cardiology consult 5
X
Echocardiogram 5
X
Histologic confirmation of disease 
X
TB testing 5
X
24-hour urine (if creatinine 
clearance is tested this way)
X
Troponin I 5
X
HIV, Hepatitis B and C serology 
and/or viral load 6 
X
X
EKG
X
X
Physical exam, weight and ECOG
X
X
X
X
Vital Signs
X
X7
X
CBC w/differential,
Platelets 
X
X
X
X
Biochemical profile8
X
X
X
X
Radiologic Evaluation9
X
X
X
HLA
X
Urinalysis
X
Thyroid tests TSH, T3, T4
X
X
X
Uric acid, amylase and lipase
X
X
X
ACTH and morning cortisol
X
PT, INR, PTT, fibrinogen
X5
X
X
Serum or urine pregnancy test
X
X

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Screening1
Baseline1
All Cycles
Day 1 1
28 
Days 
Safety 
FU11,13
Long 
Term 
FU12, 13
Tumor marker CEA, AFP, CA19-9
X
X
X
Concomitant medications
X
X
X
Adverse event evaluation
X
X
X
Baseline signs and symptoms
X
Tumor biopsy10
X
X
Research blood for Immune 
monitoring14
X
Research blood for cytokines 14
X
Research blood for tadalafil PK15 
X
Research blood for mixed meal test 
for bile acid16
X
X
Research blood for vancomycin 
level 17
X
Research blood for TCR beta 
sequencing
X
Research blood for whole exome 
panel
X
Stool collection or rectal swab for 
bile acid19
X
X
Stool collection or rectal swab for 
Vancomycin-resistant enterococci 
(VRE) development 18 
X
Stool collection or rectal swab for 
microbial DNA sequences 18
X
Nutrition consult and completion of 
food diary (Appendix H) 19
X
X
Phone call or e-mail for survival 
every 6 month
X
1 Baseline and C1D1 evaluations do not need to be repeated if performed at screening or baseline 
in designated time frame. All evaluations will be done within 72 hours before treatment initiation 
on Day 1 of every cycle. If treatment does not start within 28 days after enrollment, screening 
evaluations will be repeated. 
2 480 mg of nivolumab via IV infusion on Day 1 of each cycle. 

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3 10 mg of tadalafil PO on every day of every cycle. 
4 125 mg of vancomycin every 6 hours (500 mg daily dose) from Day 1 to day 21 of every cycle. 
5 if clinically indicated.
6 If screening serology results indicate potential HBV or HCV infection, viral load will be tested 
before every nivolumab infusion 
7 For vital signs see Section 3.2.
8 Biochemical Profile: electrolytes, BUN, creatinine, AST, ALT, total and direct bilirubin, 
calcium, phosphorus, albumin, magnesium. 
9 CT scan of chest, abdomen and pelvis (or MRI of abdomen if clinically indicated) every 8 (+/-
1) weeks after start of study therapy.  
10 Mandatory tumor biopsies will be performed at baseline and any day during week 3 of cycle 2 
while patient is taking oral vancomycin. If patient’s disease progresses before C3D1, the post-
treatment biopsy may be performed per PI discretion at the time of progression.
11 +/- 1 week
12 Follow up visits are planned to be performed at 60 (+/- 14 days) and 90 (+/- 14 days) days 
after treatment discontinuation to evaluate patient’s safety. After this visit, subjects will be 
followed every 6 months (± 1 month) for survival by phone call or e-mail.
13 If subjects are not willing to come to NIH for FU visits, they will be contacted by phone call or 
e-mail for survival and adverse events.
14 On Day 1 of cycles 1-4, then every 12 weeks until PD.
15 Before patient takes tadalafil on day 1 of every cycle. 
16 At baseline and C1D10, an IV line will be placed and a mixed meal test (Boost Plus Vanilla) 
will be performed with collection of blood samples at 0 (+/- 10), 30 (+/- 10), 60 (+/- 15), 90 (+/- 
20), 120 (+/- 20), 150 (+/- 20), and 180 (+/- 30) min for bile acid analysis. T=0 is the time patient 
ingests mixed meal.  If patient is still on the protocol treatment after 6 months, patient will repeat 
the mixed meal test on day 10 of whatever cycle the patient is on.
17 Before patient takes vancomycin on day 1 of every cycle.
18 On day 1 of every cycle.
19 At baseline and C1D10. If patient is still on the protocol treatment after 6 months, this will be 
performed on day 10 of whatever cycle patient is on.

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3.5
Costs and Compensation
3.5.1
Costs
NIH does not bill health insurance companies or participants for any research or related clinical 
care that participants receive at the NIH Clinical Center. If some tests and procedures performed 
outside the NIH Clinical Center, participants may have to pay for these costs if they are not covered 
by insurance company. Medicines that are not part of the study treatment will not be provided or 
paid for by the NIH Clinical Center.
3.5.2
Compensation
No compensation is offered on this study.
3.5.3
Reimbursement 
The NCI will cover the costs of some expenses associated with protocol participation. Some of 
these costs may be paid directly by the NIH and some may be reimbursed to the 
participant/guardian as appropriate. The amount and form of these payments are determined by 
the NCI Travel and Lodging Reimbursement Policy.
3.6
Criteria for Removal from Protocol Therapy and Off Study Criteria
Prior to removal from study, efforts must be made to have all subjects complete a safety visit 
approximately 90 days following the last dose of study therapy
3.6.1
Criteria for Removal from Protocol Therapy

Participant requests to be withdrawn from active therapy 

Unacceptable Toxicity as defined in the Section 3.3 and Appendix C

Positive pregnancy test or intent to become pregnant

Investigator discretion 

Initiation of therapy that prevents further administration of study treatment (See Section 4)

Progressive Disease. NOTE: While RECIST PD will be noted and recorded the immune-
related RECIST criteria (Appendix E) will be applied to determine discontinuation of 
study treatment.

Intercurrent illness that prevents further administration of treatment
3.6.2
Off Study Criteria 

Lost to follow-up

Death

Participant requests to be withdrawn from study

Investigator discretion

PI decision to end the study

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3.6.3
Lost to Follow-up
A participant will be considered lost to follow-up if he or she fails to return for the 60 or 90 day 
scheduled safety visits post-treatment or is unable to be contacted by the study site staff. 
The following actions must be taken if a participant fails to return to the clinic for a required 
study visit:

The site will attempt to contact the participant and reschedule the missed visit within 2 
weeks and counsel the participant on the importance of maintaining the assigned visit 
schedule and ascertain if the participant wishes to and/or should continue in the study.

Before a participant is deemed lost to follow-up, the investigator or designee will make 
every effort to regain contact with the participant (where possible, 3 telephone calls and, 
if necessary, an IRB approved certified letter to the participant’s last known mailing 
address or local equivalent methods). These contact attempts should be documented in 
the participant’s medical record or study file. 

Should the participant continue to be unreachable, he or she will be considered to have 
withdrawn from the study with a primary reason of lost to follow-up.
4
CONCOMITANT MEDICATIONS/MEASURES
All routine and appropriate supportive care (including blood products) will be provided during this 
study, as clinically indicated, and in accordance with the standard of care practices. Clinical 
judgment should be utilized in the treatment of any AE experienced by the patient.
Drugs with laxative properties and herbal or natural remedies for constipation should be used with 
caution through to 90 days after the last dose of immune checkpoint inhibitors.
Drugs with significant ototoxicity should be used with caution
4.1
Excluded Concomitant Medications
Table 10: Prohibited concomitant medications
Prohibited medication/class of drug:
Usage:
Any investigational anticancer therapy 
other than those under investigation in this 
study
Should not be given concomitantly whilst the patient is 
on study treatment
mAbs against CTLA-4, PD-1, or PD-L1 
other than those under investigation in this 
study
Should not be given concomitantly whilst the patient is 
on study treatment
Any 
concurrent 
chemotherapy, 
radiotherapy, immunotherapy, or biologic 
or hormonal therapy for cancer treatment 
other than those under investigation in this 
study
Should not be given concomitantly whilst the patient is 
on study treatment. (Concurrent use of hormones for non-
cancer-related conditions [e.g. insulin for diabetes and 
hormone replacement therapy] is acceptable. Local 
treatment of isolated lesions, excluding target lesions, for 
palliative intent is acceptable [e.g. by local surgery or 
radiotherapy])

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Table 10: Prohibited concomitant medications
Prohibited medication/class of drug:
Usage:
Immunosuppressive medications including, 
but not limited to, systemic corticosteroids 
at doses exceeding 10 mg/day of prednisone 
or equivalent, methotrexate, azathioprine, 
and tumor necrosis factor-α blockers
Should 
not 
be 
given 
concomitantly. 
(Use 
of 
immunosuppressive medications for the management of 
IP-related AEs, premedication for patients who had 
infusion reaction, or in patients with contrast allergies is 
acceptable). In addition, use of inhaled, topical, and 
intranasal corticosteroids is permitted.
Viagra
Should not be given concomitantly whilst the patient is 
on study treatment
Microbiome changing drugs, including but 
not limited to probiotics 
Should not be given concomitantly whilst the patient is 
on study treatment
4.2
Methods of Contraception
4.2.1
Female Patient of Child-bearing Potential
Females of childbearing potential who are sexually active with a non-sterilized male partner must 
use effective method of contraception (Table 11) from the time of screening and must agree to 
continue using such precautions for 5 months after the last dose of nivolumab + oral vancomycin 
+ tadalafil combination therapy. Non-sterilized male partners of a female patient must use male 
condom plus spermicide throughout this period. Cessation of birth control after this point should 
be discussed with a responsible physician. Not engaging in sexual activity for the total duration of 
the drug treatment and the drug washout period is an acceptable practice; however, periodic 
abstinence, the rhythm method, and the withdrawal method are not acceptable methods of birth 
control. Female patients should also refrain from breastfeeding throughout this period.
4.2.2
Male Patients with a Female Partner of Childbearing Potential
Non-sterilized males who are sexually active with a female partner of childbearing potential must 
use a male condom plus spermicide from screening through 7 months after the last dose of 
nivolumab + oral vancomycin + tadalafil combination therapy. Engaging in sexual activity is an 
acceptable practice; however, occasional abstinence, the rhythm method, and the withdrawal 
method are not acceptable methods of contraception. Male patients should refrain from sperm 
donation throughout this period.
Female partners (of childbearing potential) of male patients must also use a highly effective 
method of contraception throughout this period (Table 11). 
4.2.3
Highly Effective Methods of Contraception.
Highly effective methods of contraception, defined as one that results in a low failure rate (i.e. less 
than 1% per year) when used consistently and correctly are described in Table 11. Note that some 
contraception methods are not considered highly effective (e.g. male or female condom with or 
without spermicide; female cap, diaphragm, or sponge with or without spermicide; non-copper 
containing intrauterine device; progestogen-only oral hormonal contraceptive pills where 

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inhibition of ovulation is not the primary mode of action [excluding Cerazette/desogestrel which 
is considered highly effective]; and triphasic combined oral contraceptive pills).
Table 11: Highly Effective methods of contraception (<1% failure rate)
Barrier/Intrauterine methods
Hormonal Methods

Copper T intrauterine device

Levonorgesterel-releasing intrauterine 
system (e.g. Mirena®)a

Etonogestrel implants: e.g. Implanon 
or Norplant

Intravaginal 
device: 
e.g. 
ethinylestradiol and etonogestrel

Medroxyprogesterone injection: e.g. 
Depo-Provera 

Normal and low dose combined oral 
contraceptive pill

Norelgestromin/ethinylestradiol 
transdermal system

Cerazette (desogestrel)
a
This is also considered a hormonal method
4.3
Blood Donation
Subjects should not donate blood while participating in this study, and for 5 months (women) and 
7 months (men) after the last dose of nivolumab + oral vancomycin + tadalafil combination therapy. 
5
CORRELATIVE STUDIES FOR RESEARCH
5.1
Biospecimen Collection
A description of correlative studies including a brief statement of rationale and processing 
information is made below.
Test/assay
Sample 
volume 
(approx.)
Type of tubea / 
tissue sample
Collection point 
Location of 
specimen 
analysis1
Immune- 
monitoring by 
FACS
Blood, 120 
mL
EDTA (Purple 
top tubes)
See Study 
Calendar 3.4
Blood 
Processing Core 
(BPC)
Serum cytokines 
by ELISA
2mL
2 green top 
tube (GTT) or 
CPT tubes with 
sodium citrate
See Study 
Calendar 3.4
Blood 
Processing Core 
(BPC)
Tadalafil PK
4 mL
Sodium heparin 
green top tube 
(GTT)
See Study 
Calendar 3.4
Blood 
Processing Core 
(BPC)

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Test/assay
Sample 
volume 
(approx.)
Type of tubea / 
tissue sample
Collection point 
Location of 
specimen 
analysis1
Vancomycin 
Pharmacokinetic 
Analysis
4 mL
Sodium heparin 
green top tube 
(GTT)
See Study 
Calendar 3.4
Blood 
Processing Core 
(BPC)
Mixed meal test 
for bile acid levels 
in blood
5 mL
Sodium heparin 
green top tube 
(GTT)
See Study 
Calendar 3.4
Blood 
Processing Core 
(BPC)
IHC for immune 
cell infiltration
Tumor sample
At time of 
biopsies
Laboratory of 
Pathology
Proteomics
Tumor sample
At time of 
biopsies
Greten Lab
Metabolomics
Tumor sample
At time of 
biopsies
Greten Lab
TCRbeta 
sequencing
10 mL
EDTA (Purple 
top tube)
See Study 
Calendar 3.4
Blood 
Processing Core 
(BPC)
TCRbeta 
sequencing
Tumor sample
At time of 
biopsies
Greten Lab
RNA seq
Tumor sample
At time of 
biopsies
Greten Lab
Whole 
exome/panel
Tumor sample
At time of 
biopsies
Greten Lab
Whole 
exome/panel
Blood, 2.5 mL
1 PAX gene 
DNA tube
See Study 
Calendar 3.4
Greten Lab
Fecal bile acid 
composition
Stool sample 
or rectal swab
Feccontainer 
tubes or rectal 
swabs
See Study 
Calendar 3.4
Greten Lab
Vancomycin-
resistant 
enterococci (VRE) 
development
Stool sample 
or rectal swab
Feccontainer 
tubes or rectal 
swabs
See Study 
Calendar 3.4
NIH 
Department of 
Laboratory 
Medicine
DNA sequencing 
of the microbiome
Stool sample 
or rectal swab
Feccontainer 
tubes or rectal
See Study 
Calendar 3.4
Greten Lab
a.
Please note that tubes and media may be substituted based on availability with the 
permission of the PI or laboratory investigator.
 

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5.2
Correlative Studies of Blood Samples
Blood samples will be collected at time points indicated in the Study Calendar 3.4. Blood samples 
will be initially sent to the Blood Processing Core (BPC) for barcoding and storage. On certain 
occasions, the blood may also be brought to the Greten lab for processing and analysis
5.2.1
Immune Monitoring
We will analyze PBMC for quantitative and functional changes of effector cells by FACS. The 
effect on (i) CD4 T cell number and activity, (ii) CD8 T cell number and activity, (iii) NK cell 
number and activity, (iv) Treg number, (v) MDSC: frequency + functional assay, and (vi) the 
detection of tumor-associated antigens using tetramer assay. These experiments will be done in 
Blood Processing Core (BPC).
5.2.2
Serum Cytokines
We will analyze cytokine profile from sera for cytokines and chemokines by ELISA, that are 
associated with treatment efficacy and outcome. These experiments will be done in Blood 
Processing Core (BPC).
5.2.3
Tadalafil Pharmacokinetic Analysis
Blood samples for the determination of tadalafil plasma trough levels will be obtained from 
participating patients at time points indicated in Study Calendar 3.4. Bioanalytical measurements 
will be conducted on an ultra HPLC-MS/MS system by the Clinical Pharmacology Program (CPP)
5.2.4
Vancomycin Pharmacokinetic Analysis
Blood samples for the determination of vancomycin plasma trough levels will be obtained from 
participating patients at time points indicated in Study Calendar 3.4. Bioanalytical measurements 
will be conducted on an ultra HPLC-MS/MS system by the Clinical Pharmacology Program (CPP) 
5.2.5
Bile Acid Analysis
Fasting blood samples will be drawn. At T = 0 min the participants ingest a liquid meal containing 
25% of total daily energy expenditure (Boost Plus Vanilla; 1.5 kcal/mL; 49% carbohydrates, 35% 
fat and 16% protein). Participants are asked to drink the liquid meal within 10 min. Blood samples 
for determination of plasma bile acid levels will be drawn at time points indicated in Study 
Calendar 3.4 and Appendix I. The blood samples will be collected on baseline and ten days after 
oral vancomycin is taken during the first treatment cycle. Bile acid profile is measured using liquid 
chromatography tandem mass spectrometry (LC-MS/MS) as described[91]. Moreover, fecal bile 
acid composition was determined from rectal swab at baseline and 10 days after oral vancomycin 
treatment, using gas chromatography (GC). The total amount of primary and secondary bile salts 
is calculated as the sum of the individually quantified bile salts. AUC will be calculated. These 
experiments will be done in Dr. Figg lab.
5.2.6
Food Diary for Mixed Meal Study
It has been well demonstrated the diet has great impact on gut microbiota. Therefore, a food diary 
for a mixed meal study (Appendix H) will be used in this study to assess the dietary composition, 
in addition to nutrition consult. A specialist will evaluate the diary in detail. The completion of 
this diary will take 15-30 minutes with the time of consultation included.

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Patients will have nutrition consult and complete the diary on the same days as bile acid analysis 
(See Study Calendar 3.4.) 
Non-English-speaking subjects will complete this instrument with help of an interpreter.
5.3
Tumor Studies (Non-genetic Analysis)
Mandatory tumor biopsies will be performed at baseline and anytime during week 3 of Cycle 2 
while patient is still on oral vancomycin. If patient disease progresses before week 3 of Cycle 2, 
post-treatment biopsy may be performed per PI discretion at the time of progression. It is preferred 
that at least two core biopsies ≥18 gauge in diameter and ≥1 cm in length will be obtained. 
The use of imaging to facilitate biopsies will be decided by members of the Interventional 
Radiology team and may include ultrasound, CT scan, PET scan or MRI. 
Tumor biopsies are mandatory on this study, however, biopsies and local anesthesia will be 
administered only if they are of minimal risk to the participant, as determined by the investigators 
and Interventional Radiology. If these procedures are not considered to be of minimal risk to the 
participant, they will not be done. Patient will continue treatment. 
5.3.1
Tumor Tissue Analysis
5.3.1.1
IHC
IHC will be performed on tumor tissue for assessment of immune cell infiltration (e.g. CD3+ 
CD4/8 cells, PDL1 etc.). Immunohistochemistry will be performed in the Laboratory of Pathology 
under the direction of Dr. David Kleiner.
5.3.1.2
Omics Study
Tumor samples will be collected for proteomics and metabolomics studies to investigate the 
differential expression of protein and metabolite that might be associated with the treatment 
outcome or efficacy. These experiments will be done in Dr. Greten lab.
5.3.2
Stool Collection and Processing
Stool samples or rectal swabs will be collected by the patients at time points indicated in the Study 
Calendar 3.4. 
Specimens will be frozen and kept at -20°C until analysis, which will be done in batches. 
Samples will be collected for bile acid composition (Section 5.2.5), Vancomycin-resistant 
enterococci (VRE) development DNA and sequencing of the microbiome (Section 5.5.3).
5.3.2.1
Vancomycin-resistant Enterococci (VRE) Development
Stool specimen will be collected to evaluate VRE development given prolonged vancomycin 
exposure increase the odds of VRE. The protocol will evaluate VRE incidence as one of safety 
monitor.
5.4
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. All 
samples will be sent to Blood Processing Core (BPC) for processing and storage until they are 

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distributed to the designated place of analysis as described in the protocol. Samples will not be 
sent outside NIH without appropriate approvals and/or agreements, if required.
All samples will be barcoded, with data entered and stored in the secure databases. These databases 
create a unique barcode ID for every sample and sample box, which cannot be traced back to 
patients without database 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 
(delay in sample processing, storage conditions on the ward, etc.).
Barcoded samples are stored in barcoded boxes in a locked freezer at either -20 or -800C according 
to stability requirements. 
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 database. 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 the IRB approved protocol) and that any unused samples 
must be returned. 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. 
5.4.1
Samples Managed by Dr. Figg’s Blood Processing Core (BPC) 
5.4.1.1
BPC processing/contact Information 
Please e-mail  NCIBloodcore@mail.nih.gov at least 24 hours before transporting samples (the 
Friday before is preferred).
For sample pickup, page 102-11964.
For immediate help, call 240-760-6180 (main blood processing core number) or, if no answer, 
240-760-6190 (main clinical pharmacology lab number).
For questions regarding sample processing, contact NCIBloodcore@mail.nih.gov.
5.4.2
Samples Managed by the Laboratory of Dr. Tim Greten
Contact information:
Sophie Wang
Building 10 Rm 3B44
Phone: 240-858-3218
E-mail: sophie.wang@nih.gov
5.4.3
Protocol Completion/Sample Destruction
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 in sections 
above. 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.
If the patient withdraws consent the participant’s 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.

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The PI will record any loss or unanticipated destruction of samples as a deviation. Reporting will 
be per the requirements of section 7.2.
5.5
Samples for Genetic/Genomic Analysis
5.5.1
TCR Beta Sequencing 
PBMC and tumor samples will be collected for TCR beta sequencing performed in Blood 
Processing Core (BPC) and Dr. Greten’ lab to analyze the T cell response.
5.5.2
Whole exome/Panel (tumor/normal) and RNAseq (tumor)
Whole exome will be done on tumor and blood sample. RNAseq will be done on tumor for 
expressed somatic mutations and neoantigen discovery. Tumor material for these analyses will be 
sent to the laboratory of Dr Greten. Specifically, the expression of different signaling pathway, 
including interferon gamma signaling, immune cell functional molecular, e.g. perforin and 
granzyme, will be investigated
5.5.3
Microbial DNA Sequence
Samples sequenced to study microbial communities will contain human DNA sequences. However, 
bioinformatic analyses of the sequences will remove sequences containing significant alignment 
to publicly available human DNA sequences. These data will be coded and stored at the NIH 
Intramural Sequencing Center (NISC) per standard protocols for intramural sequencing. The 
resulting sequences will be analyzed. Publicly available human DNA sequences likely do not 
include all possible DNA sequences and could result in fragments of human DNA sequences being 
incorporated into the microbial DNA analysis. Microbial DNA sequence from this study will be 
published and posted on the web. Coded information obtained from sequencing samples will be 
placed 
into 
an 
open 
access 
(public) 
scientific 
database 
such 
as 
Genbank: 
http://www.ncbi.nlm.nih.gov/Genbank/. For example, we will deposit 16S rRNA gene sequences 
in Genbank. Microbial sequencing is also deposited in the Short Read Trace Archive that is a 
component of Genbank. All of the data will be coded to minimize any risk of identifying an 
individual. Samples will be reported without any unique identifiable features of the subject.
5.5.4
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 will be contacted at that time with a request to provide a blood sample to be sent to a 
CLIA certified laboratory.
5.5.5
Genetic Counseling
If the research findings are verified in the CLIA certified lab, the subject will be offered the 
opportunity to come to NIH (at our expense) to have genetic education and counseling with the 
NCI Genetics Branch to explain this result. If the subject does not want to come to NIH, a referral 
to a local genetic healthcare provider will be provided (at their expense). 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. 

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6
DATA COLLECTION AND EVALUATION 
6.1
Data Collection
The PI will be responsible for overseeing entry of data into a 21 CFR Part 11-compliant data 
capture system provided by the NCI CCR 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. Primary and final analyzed data will have 
identifiers so that research data can be attributed to an individual human subject participant.
All adverse events, including clinically significant abnormal findings on laboratory evaluations, 
regardless of severity, will be followed until return to baseline or stabilization of event. 
Document AEs from the first study intervention, Study Day 1 of cycle 1 through 90 days after the 
study agent (s) was/were administered. Beyond 90 days after the last intervention, only adverse 
events which are serious and related to the study intervention need to be recorded.
An abnormal laboratory value will be recorded in the database as an AE only if the laboratory 
abnormality is characterized by any of the following:
•
Results in discontinuation from the study
•
Is associated with clinical signs or symptoms 
•
Requires treatment or any other therapeutic intervention
•
Is associated with death or another serious adverse event, including hospitalization. 
•
Is judged by the Investigator to be of significant clinical impact
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.
Adverse Events of grade 1 will not be collected.
The results of the full diagnostic workup, blood and sputum culture, hematological parameters etc. 
will be captured in the eCRF.
Information on all concomitant medications, administered blood products, as well as interventions 
occurring during the study must be recorded on the patient’s eCRF in C3D.
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 breech in our plan to protect 
subject confidentiality and trial data has occurred, this will be reported expeditiously per 
requirements in section 7.2.1. 
6.2
Data Sharing Plans
6.2.1
Human Data Sharing Plan
The PI will share coded linked human data generated in this research for future research 

in a NIH-funded or approved public repository clinicaltrials.gov, dbGaP and GenBank

in BTRIS 

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
in publication and/or public presentations
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 weeks (+/- 1 
week). Whilst immune-related RECIST criteria (Appendix E) will be taken into consideration 
regarding continuation of therapy in the event of growth (with a requirement for confirmation of 
PD), standard RECIST criteria will be the primary method used for evaluation of the primary 
endpoint.
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) [92, 93] and Modified Immune-related response criteria (Section 6.3.3.5). 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
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: 

By chest x-ray: >20 mm; 

By CT scan: 
o
Scan slice thickness 5 mm or under as >10 mm with CT scan
o
Scan slice thickness >5 mm: double the slice thickness 

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/pneumonitis, 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.

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‘Cystic lesions’ thought to represent cystic metastases can be considered as measurable lesions, if 
they meet the definition of measurability described above. However, if non-cystic lesions are 
present in the same patient, these are preferred for selection as target lesions.
Target lesions: All measurable lesions up to a maximum of 2 lesions per organ and 5 lesions in 
total, representative of all involved organs, should be identified as target lesions and recorded and 
measured at baseline. Target lesions should be selected on the basis of their size (lesions with the 
longest diameter), be representative of all involved organs, but in addition should be those that 
lend themselves to reproducible repeated measurements. It may be the case that, on occasion, the 
largest lesion does not lend itself to reproducible measurement in which circumstance the next 
largest lesion which can be measured reproducibly should be selected. A sum of the diameters 
(longest for non-nodal lesions, short 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.
6.3.2
Methods for Evaluation of Measurable Disease 
All measurements should be taken and recorded in metric notation using a ruler or calipers. All 
baseline evaluations should be performed as closely as possible to the beginning of treatment and 
never more than 4 weeks before the beginning of the treatment.
The same method of assessment and the same technique should be used to characterize each 
identified and reported lesion at baseline and during follow-up. Imaging-based evaluation is 
preferred to evaluation by clinical examination unless the lesion(s) being followed cannot be 
imaged but are assessable by clinical exam.
Clinical lesions: Clinical lesions will only be considered measurable when they are superficial (e.g., 
skin nodules and palpable lymph nodes) and 10 mm diameter as assessed using calipers (e.g., 
skin nodules). In the case of skin lesions, documentation by color photography, including a ruler 
to estimate the size of the lesion, is recommended. 
Chest x-ray: Lesions on chest x-ray are acceptable as measurable lesions when they are clearly 
defined and surrounded by aerated lung. However, CT is preferable. 
Conventional CT and MRI: This guideline has defined measurability of lesions on CT scan based 
on the assumption that CT slice thickness is 5 mm or less. If CT scans have slice thickness greater 
than 5 mm, the minimum size for a measurable lesion should be twice the slice thickness. MRI is 
also acceptable in certain situations (e.g. for body scans). 
Use of MRI remains a complex issue. MRI has excellent contrast, spatial, and temporal resolution; 
however, there are many image acquisition variables involved in MRI, which greatly impact image 
quality, lesion conspicuity, and measurement. Furthermore, the availability of MRI is variable 
globally. As with CT, if an MRI is performed, the technical specifications of the scanning 
sequences used should be optimized for the evaluation of the type and site of disease. Furthermore, 
as with CT, the modality used at follow-up should be the same as was used at baseline and the 

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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.
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 during 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.
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.
6.3.3
Response Criteria
6.3.3.1
Evaluation of Target Lesions
Complete Response (CR): Disappearance of all target lesions. Any pathological lymph nodes 
(whether target or non-target) must have reduction in short axis to <10 mm.
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.
6.3.3.2
Evaluation of Non-Target Lesions
Complete Response (CR): Disappearance of all non-target lesions and normalization of tumor 
marker level. All lymph nodes must be non-pathological in size (<10 mm short axis).
Note: If tumor markers are initially above the upper normal limit, they must normalize for a patient 
to be considered in complete clinical response.

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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).
6.3.3.3
Evaluation of Best Overall Response
The best overall response is the best response recorded from the start of the treatment until disease 
progression/recurrence (taking as reference for progressive disease the smallest measurements 
recorded since the treatment started). The patient's best response assignment will depend on the 
achievement of both measurement and confirmation criteria.
For Patients with Measurable Disease (i.e., Target Disease)
Target 
Lesions
Non-Target 
Lesions
New 
Lesions
Overall 
Respons
e
Best Overall Response when 
Confirmation is Required*
CR
CR
No
CR
>4 wks. Confirmation**
CR
Non-CR/Non-PD
No
PR
CR
Not evaluated
No
PR
PR
Non-CR/Non-
PD/not evaluated
No
PR
>4 wks. Confirmation**
SD
Non-CR/Non-
PD/not evaluated
No
SD
Documented at least once >4 
wks. from baseline**
PD
Any
Yes or No
PD
Any
PD***
Yes or No
PD
Any
Any
Yes
PD
no prior SD, PR or CR
*
See RECIST 1.1 manuscript for further details.
**
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

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Non-Target Lesions
New Lesions
Overall Response
* ‘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
6.3.3.4
Duration of Response
Duration of overall response: The duration of overall response is measured from the time 
measurement criteria are met for CR or PR (whichever is first recorded) until the first date that 
recurrent or progressive disease is objectively documented (taking as reference for progressive 
disease the smallest measurements recorded since the treatment started).
The duration of overall CR is measured from the time measurement criteria 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. 
6.3.3.5
Modified Immune-mediated response criteria (imRC)
Modified immune-mediated response criteria (imRC) will also be employed in this study. This 
new classification is based on the recent learning from clinical studies with cancer 
immunotherapies that even if some new lesions appear at the beginning of a treatment or if the 
total tumor burden does not increase substantially, tumor regressions or stabilizations might still 
occur later. The irRC were created using bi-dimensional measurements (as previously widely used 
in the World Health Organization criteria). For this trial, the concepts of the irRC are combined 
with RECIST 1.1 to come up with the modified imRC. Please refer to Appendix E for further 
details.
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 5.0 will be utilized for AE reporting. 
All appropriate treatment areas should have access to a copy of the CTCAE version 5.0. A copy 
of 
the 
CTCAE 
version 
5.0 
can 
be 
downloaded 
from 
the 
CTEP 
web 
site 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm#ctc_50).
7
NIH REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING PLAN
7.1
Definitions
Please refer to definitions provided in Policy 801: Reporting Research Events found here.
7.2
OHSRP Office of Compliance and Training / IRB Reporting
7.2.1
Expedited Reporting
Please refer to the reporting requirements in Policy 801: Reporting Research Events and Policy 
802 Non-Compliance Human Subjects Research found here. Note: Only IND Safety Reports that 
meet the definition of an unanticipated problem will need to be reported per these policies.

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7.2.2
IRB Requirements for PI Reporting at Continuing Review
Please refer to the reporting requirements in Policy 801: Reporting Research Events found here.
7.3
NCI Clinical Director Reporting 
Problems expeditiously reported to the OHSRP in iRIS will also be reported to the NCI Clinical 
Director.  A separate submission is not necessary as reports in iRIS will be available to the Clinical 
Director.
In addition to those reports, all deaths that occur within 30 days after receiving a research 
intervention should be reported via email to the Clinical Director unless they are due to progressive 
disease.  
To report these deaths, please send an email describing the circumstances of the death to Dr. Dahut 
at NCICCRQA@mail.nih.gov within one business day of learning of the death. 
7.4
NIH Required Data and Safety Monitoring Plan
The clinical research team will meet weekly when patients are being actively treated on the trial 
to discuss each patient. 
All data will be collected in a timely manner and reviewed by the principal investigator or a lead 
associate investigator. Events meeting requirements for expedited reporting as described in section 
7.2.1 will be submitted within the appropriate timelines. 
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 
investigation and provide appropriate delegation of responsibilities to other members of the 
research staff. 
8
SPONSOR PROTOCOL/SAFETY REPORTING
8.1
Definitions
8.1.1
Adverse Event 
Any untoward medical occurrence in a patient or clinical investigation subject administered a 
pharmaceutical product and which does not necessarily have a causal relationship with this 
treatment. An adverse event (AE) can therefore be any unfavorable and unintended sign (including 
an abnormal laboratory finding), symptom, or disease temporally associated with the use of a 
medicinal (investigational) product, whether or not related to the medicinal (investigational) 
product (ICH E6 (R2))
8.1.2
Serious Adverse Event (SAE)
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:

Death,

A life-threatening adverse event (see section 8.1.3)

Inpatient hospitalization or prolongation of existing hospitalization 
o
A hospitalization/admission that is pre-planned (i.e., elective or scheduled surgery 
arranged prior to the start of the study), a planned hospitalization for pre-existing 

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condition, or a procedure required by the protocol, without a serious deterioration in 
health, is not considered a serious adverse event.
o
A hospitalization/admission that is solely driven by non-medical reasons (e.g., 
hospitalization for patient or subject convenience) is not considered a serious adverse 
event. 
o
Emergency room visits or stays in observation units that do not result in admission to 
the hospital would not be considered a serious adverse event. The reason for seeking 
medical care should be evaluated for meeting one of the other serious criteria.

Persistent or significant incapacity or substantial disruption of the ability to conduct normal 
life functions

A congenital anomaly/birth defect

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.
8.1.3
Life-threatening 
An adverse event or suspected adverse reaction is considered "life-threatening" if, in the view of 
either the investigator or sponsor, its occurrence places the patient or subject at immediate risk of 
death. It does not include an adverse event or suspected adverse reaction that, had it occurred in a 
more severe form, might have caused death. (21CFR312.32)
8.1.4
Severity
The severity of each Adverse Event will be assessed utilizing the CTCAE version 5.0.
8.1.5
Relationship to Study Product
All AEs will have their relationship to study product assessed using the terms:  related or not 
related.  

Related – There is a reasonable possibility that the study product caused the adverse event. 
Reasonable possibility means that there is evidence to suggest a causal relationship 
between the study product and the adverse event.

Not Related – There is not a reasonable possibility that the administration of the study 
product caused the event.
8.2
Assessment of Safety Events
AE information collected will include event description, date of onset, assessment of severity and 
relationship to study product and alternate etiology (if not related to study product), date of 
resolution of the event, seriousness and outcome. The assessment of severity and relationship to 
the study product will be done only by those with the training and authority to make a diagnosis 
and listed on the Form FDA 1572 as the site principal investigator or sub-investigator.  AEs 
occurring during the collection and reporting period will be documented appropriately regardless 
of relationship.  AEs will be followed through resolution.
SAEs will be:

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
Assessed for severity and relationship to study product and alternate etiology (if not related 
to study product) by a licensed study physician listed on the Form FDA 1572 as the site 
principal investigator or sub-investigator.

Recorded on the appropriate SAE report form, the medical record and captured in the 
clinical database.

Followed through resolution by a licensed study physician listed on the Form FDA 1572 
as the site principal investigator or sub-investigator.
For timeframe of recording adverse events, please refer to section 6.1. All serious adverse events 
recorded from the time of first investigational product administration must be reported to the 
sponsor with the exception of any listed in section 8.4. 
8.3
Reporting of Serious Adverse Events
Any AE that meets protocol-defined serious criteria or meets the definition of Adverse Event of 
Special Interest that require expedited reporting must be submitted immediately (within 24 hours 
of awareness) to OSRO Safety using the CCR SAE report form.  Any exceptions to the expedited 
reporting requirements are found in section 8.4.
All SAE reporting must include the elements described in section 8.2.
SAE reports will be submitted to the Center for Cancer Research (CCR) at: 
OSROSafety@mail.nih.gov and to the CCR PI and study coordinator.  CCR SAE report form 
and instructions can be found at: 
https://ccrod.cancer.gov/confluence/display/CCRCRO/Forms+and+Instructions.
Following the assessment of the SAE by OSRO, other supporting documentation of the event may 
be requested by the OSRO Safety and should be provided as soon as possible.
8.4
Waiver of expedited reporting to CCR 
As death due to disease progression is part of the study objectives (OS), and captured as an 
endpoint in this study, death due to disease progression will not be reported in expedited manner 
to the sponsor. However, if there is evidence suggesting a causal relationship between the study 
drug and the event, report the event in an expedited manner according to section 8.3.
Hospitalization that is deemed to be due to disease progression, and not attributable to the 
intervention will not be reported as an SAE. The event, and the assessment that it was caused by 
disease progression will be documented in the medical records. The causality assessment of 
hospitalization will be re‐evaluated any time when new information is received. If the causality 
assessment changes from disease progression to related to the study intervention, SAE report will 
be sent to the Sponsor in an expedited manner according to section 8.3. If there is any uncertainty 
whether the intervention is a contributing factor to the event, the event should be reported as AE 
or SAE as appropriate.
8.5
Safety Reporting to the Manufacturer, Bristol Myers Squibb
All events listed below must be reported in the defined timeline to CCRsafety@mail.nih.gov. The 
CCR Office of Regulatory Affairs will send all reports to the manufacturer as described below.
8.5.1
Death Reporting

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All deaths that occur during the study, or within 90-day post-last dose of nivolumab/oral 
vancomycin/tadalafil safety follow-up period must be reported to BMS as follows:

Death that is clearly the result of disease progression should be documented but should not 
be reported as an SAE.

Where death is not due (or not clearly due) to progression of the disease under study, the 
AE causing the death must be reported to BMS as a SAE within 24 hours of Sponsor 
awareness. The report should contain a comment regarding the co-involvement of 
progression of disease, if appropriate, and should assign main and contributory causes of 
death. 

Deaths with an unknown cause should always be reported as a SAE.
8.5.2
Reporting of Serious Adverse Events
All SAEs will be reported, whether considered causally related to the investigational product, or 
to the study procedure(s). The reporting period for SAEs is the period immediately following the 
time that written informed consent is obtained through 90 days after the last dose of nivolumab/oral 
vancomycin/tadalafil or until the initiation of alternative anticancer therapy. 
Sponsor must inform the FDA, via a MedWatch, of any serious or unexpected adverse events. 
SAEs and pregnancies must be reported to BMS within one business day of Sponsor becoming 
aware of the event. SAEs must be recorded on MedWatch or equivalent form. Pregnancies must 
be reported and submitted to BMS. BMS will perform due diligence follow-up using the BMS 
pregnancy Form (Appendix G) which the investigator must complete. It is the responsibility of 
the sponsor to compile all necessary information and ensure that the FDA receives a report 
according to the FDA reporting requirement timelines and to ensure that these reports are also 
submitted to BMS at the above time points.
* A cover page should accompany the MedWatch form indicating the following:
•
“Notification from an Investigator Sponsored Study”
•
The investigator IND number assigned by the FDA
•
The investigator’s name and address
•
The trial name/title and BMS ISS reference number (CA209-8R3) (BMS only)
* Sponsor must also indicate, either in the SAE report or the cover page, the causality of events in 
relation to all study medications and if the SAE is related to disease progression, as determined by 
the principal investigator.
* Send SAE report and accompanying cover page by way of email to BMS’s designated mailbox: 
Worldwide.Safety@BMS.com. SAE FAX Number: +1 609-818-3804
If a non-serious AE becomes serious, this and another relevant follow-up information must also 
be provided to BMS, and the FDA.
For all infusion reactions, SAEs should be reported to BMS Patient safety.
8.5.3
Drug Induced Liver Injury (DILI)
Wherever possible, timely confirmation of initial liver-related laboratory abnormalities should 
occur prior to the reporting of a potential DILI event. All occurrence of potential DILIs, meeting 

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the defined criteria, must be reported as SAEs (See Section 8.3 for reporting details). The following 
definition takes into account anticipated baseline compromise of liver function in patients.
Potential drug induced liver injury is defined as: 
1) Concurrent ALT > 10 times upper limit of normal (ULN)
AND
2) Total bilirubin > 2 times ULN or baseline value (if elevated bilirubin at study entry), 
AND
3) No other immediately apparent possible causes of AT elevation and hyperbilirubinemia, 
including, but not limited to, viral hepatitis, pre-existing chronic or acute liver disease, or the 
administration of other drug(s) known to be hepatotoxic.
Drug induced liver injury (DILI) in a study subject, with or without associated clinical 
manifestations, is required to be reported as “hepatic function abnormal” within 24 hours of 
Sponsor knowledge of the event to the BMS Patient Safety using the designated Safety e-mailbox 
Worldwide.Safety@BMS.com unless a definitive underlying diagnosis for the abnormality (e.g., 
cholelithiasis or bile duct obstruction) that is unrelated to investigational product has been 
confirmed.
If the definitive underlying diagnosis for the abnormality has been established and is unrelated to 
investigational product, the decision to continue dosing of the study subject will be based on the 
clinical judgment of the investigator.
If no definitive underlying diagnosis for the abnormality is established, dosing of the study subject 
must be interrupted immediately. Follow-up investigations and inquiries must be initiated by the 
investigational site without delay.
Each reported event of hepatic function abnormality will be followed by the investigator and 
evaluated by the sponsor and BMS.
8.6
Reporting Pregnancy 
All required pregnancy reports/follow-up to OSRO will be submitted to: 
OSROSafety@mail.nih.gov and to the CCR PI and study coordinator.  Forms and instructions 
can be found here: 
https://ccrod.cancer.gov/confluence/display/CCRCRO/Forms+and+Instructions 
8.6.1
Maternal exposure
If a patient becomes pregnant during the course of the study, the study treatment should be 
discontinued immediately, and the pregnancy reported to the Sponsor no later than 24 hours of 
when the Investigator becomes aware of it. The Investigator should notify the Sponsor no later 
than 24 hours of when the outcome of the Pregnancy becomes known,
Pregnancy itself is not regarded as an SAE. However, congenital abnormalities or birth defects 
and spontaneous miscarriages that meet serious criteria (section 8.1.2) should be reported as SAEs. 
The outcome of all pregnancies (spontaneous miscarriage, elective termination, ectopic pregnancy, 
normal birth, or congenital abnormality) should be followed up and documented.

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8.6.2
Paternal exposure
Male patients should refrain from fathering a child or donating sperm during the study and for 7 
months after the last dose of study drug (s).
Pregnancy of the patient’s partner is not considered to be an AE. However, the outcome of all 
pregnancies (spontaneous miscarriage, elective termination, ectopic pregnancy, normal birth, or 
congenital abnormality) occurring from the date of the first dose until 7 months after the last dose 
should, if possible, be followed up and documented.
8.7
Regulatory Reporting for Studies Conducted Under CCR-Sponsored IND
Following notification from the investigator, CCR, the IND sponsor, will report any suspected 
adverse reaction that is both serious and unexpected.  CCR will report an AE as a suspected adverse 
reaction only if there is evidence to suggest a causal relationship between the study product and 
the adverse event.  CCR will notify FDA and all participating investigators (i.e., all investigators 
to whom the sponsor is providing drug under its INDs or under any investigator’s IND) in an IND 
safety report of potential serious risks from clinical trials or any other source, as soon as possible, 
in accordance to 21 CFR Part 312.32. 
All serious events will be reported to the FDA at least annually in a summary format.
8.8
Sponsor Protocol Non-Adherence Reporting
Protocol non-adherence is defined as any noncompliance with the clinical trial protocol, GCP, or 
protocol-specific procedural requirements on the part of the participant, the Investigator, or the 
study site staff inclusive of site personnel performing procedures or providing services in support 
of the clinical trial.
It is the responsibility of the study Staff to document any protocol non-adherence identified by the 
Staff or the site Monitor on the OSRO Site Protocol Non-Adherence Log.  The protocol-specific, 
cumulative non-adherence log should be maintained in the site essential documents file and 
provided to OSRO as required over the duration of the study. In addition, any non-adherence to 
the protocol should be documented in the participant’s source records and reported to the local 
IRB per their guidelines. OSRO required protocol non-adherence reporting is consistent with 
E6(R2) GCP: Integrated Addendum to ICH E6(R1): 4.5 Compliance with Protocol; 5.18.3 (a), and 
5.20 Noncompliance; and ICH E3 16.2.2 Protocol deviations.
9
CLINICAL MONITORING 
As a sponsor for clinical trials, FDA regulations require the CCR to maintain a monitoring 
program. The CCR’s program allows for confirmation of: study data, specifically data that could 
affect the interpretation of primary and secondary study endpoints; adherence to the protocol, 
regulations, ICH E6, and SOPs; and human subjects protection. This is done through independent 
verification of study data with source documentation focusing on:

Informed consent process

Eligibility confirmation

Drug administration and accountability

Adverse events monitoring

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
Response assessment.
The monitoring program also extends to multi-site research when the CCR is the coordinating 
center.
This trial will be monitored by personnel employed by a CCR contractor. Monitors are qualified 
by training and experience to monitor the progress of clinical trials. Personnel monitoring this 
study will not be affiliated in any way with the trial conduct.
Clinical site monitoring is conducted to ensure that the rights of the participants are protected, that 
the study is implemented per the approved protocol, Good Clinical Practice and standard operating 
procedures, and that the quality and integrity of study data and data collection methods are 
maintained.  Monitoring for this study will be performed by NCI CCR Office of Sponsor and 
Regulatory Oversight (OSRO) Monitoring based on OSRO standards, FDA Guidance E6(R2) 
Good Clinical Practice: Integrated Addendum to ICH E6(R1) March 2018, and applicable 
regulatory requirements.
Details of clinical site monitoring will be documented in a Clinical Monitoring Plan (CMP) 
developed by OSRO. CMPs will be protocol-specific, risk-based and tailored to address human 
subject protections and integrity of the study data. The intensity and frequency of monitoring will 
be based on several factors, including study type, phase, risk, complexity, expected enrollment rate, 
and any unique attributes of the study and the site. OSRO Monitoring visits and related activities 
will be conducted throughout the life cycle of each protocol, with the first activity being before 
study start to conduct a Site Assessment Visit (SAV) (as warranted), followed by a Site Initiation 
Visit (SIV), Interim Monitoring Visit(s) (IMVs), and a study Close-Out Visit (COV).
Some monitoring activities may be performed remotely, while others will take place at the study 
site(s).  Monitoring visit reports will describe visit activities, observations, findings of protocol 
non-adherence and associated action items or follow-up required for resolution of findings.  
Monitoring reports will be distributed to the study PI, NCI CCR QA, coordinating center (if 
applicable) and the OSRO regulatory file.
If protocol non-adherence is identified by the Monitor (i.e., any noncompliance with the clinical 
trial protocol, GCP, or protocol-specific procedural requirements on the part of the participant, the 
Investigator, or the site Staff) the Monitor will note the observation, review with site Staff and if 
unresolved, request that the Staff document the non-adherence on the protocol-specific OSRO Site 
Protocol Non-Adherence Log (see Section 8.8).
10 STATISTICAL CONSIDERATIONS
10.1 Study Objective
10.1.1 Primary Objective

The primary objective of this pilot trial is to preliminarily determine the best overall response 
according to RECIST 1.1 to combined treatment of nivolumab, oral vancomycin and tadalafil 
in the treatment of patients with HCC or non-HCC liver dominant metastases from CRC and 
PDAC.

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10.1.2 Secondary Objectives

To assess the safety and tolerability of nivolumab in combination with oral vancomycin and 
tadalafil in patients with refractory primary HCC or liver dominant metastatic cancer from 
CRC or PDAC.

To assess overall survival (OS) of nivolumab combined with oral vancomycin and tadalafil in 
patients with refractory HCC or liver dominant metastatic cancer from CRC or PDAC.
10.2 Sample Size Determination
Separately in HCC and Non-HCC patients, a small one-stage phase II design will be utilized:
In HCC cohort 12 and in Non-HCC cohort 13 evaluable patients will be enrolled. With 12/13 
patients, an exact binomial test with a one-sided 0.10 significance level will have 84% power to 
detect the difference between an unacceptably low 4% response rate (PR + CR) and a desirable, 
targeted response rate (PR + CR) of 25%. In practice, if there are at least 2 responses among the 
12/13 patients, then the lower one-sided 90% confidence bound is at least 4.5%. Simultaneously, 
the upper one-sided 90% confidence bound is at least 38.6%. Thus, attaining 2 or more responses 
would be sufficient to demonstrate ruling out a 4% response rate and also be consistent with a 
desirable 25% rate or greater.
If the response rate (PR + CR) in a cohort is identified as being adequate, consideration will be 
given to enrolling replicate cohorts as appropriate by amendment to evaluate additional patients 
independently from the initial 12/13 patients.
It is expected that up to 5-6 HCC patients and 5-6 non-HCC can be enrolled within one year; thus, 
2-3 years may be required to accrue 12 evaluable HCC patients and 13 evaluable non-HCC patients 
(25 total evaluable patients). To allow for a small number of inevaluable patients, the accrual 
ceiling will be set at 27 patients.
As of Amendment version date 12/22/2021:
The trial will no longer enroll any patients with HCC but will enroll only additional patients 
without HCC. There were 5 evaluable patients with HCC enrolled prior to December 2021, and 
up to 20 evaluable non-HCC patients will be enrolled based on the following:
With 20 patients, an exact binomial test with a one-sided 0.10 significance level will have 82.3% 
power to detect the difference between an unacceptably low 5% response rate (PR + CR) and a 
desirable, targeted response rate (PR + CR) of 21%. In practice if there are at least 3 responses 
among the 20 patients, then the lower two-sided 90% exact mid-p Clopper-Pearson confidence 
bound is at least 5.2% (and the lower one sided 90% confidence bound is at least 6.9%). 
Simultaneously, the upper two-sided 90% confidence bound is at least 32.0% (and the upper one-
sided 90% confidence bound is 28.0%). Thus, attaining 3 or more responses would be sufficient 
to demonstrate ruling out a clinically not interesting 5% (that is, will not be worse than 5%) 
response rate and be consistent with a desirable 21% rate or greater.
As of December 2021, there are 12 non-HCC patients enrolled. It is expected that the remaining 5 
non-HCC patients can be enrolled in less than one year. With 5 HCC patients and 20 non-HCC 
patients, the total evaluable patients will be 25. To allow for a small number of inevaluable patients, 
the accrual ceiling will remain at 27.

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10.3 Populations for Analyses
All patients received treatments, regardless of drug types included in this study and had at least 
one re-staging will be evaluated for response (PR + CR).
10.4 Statistical Analyses
10.4.1 General Approach
The fraction of patients who experience a response will be reported along with confidence intervals.
10.4.2 Analysis of the Primary Endpoints
In each cohort, the fraction of patients who experience a response (PR + CR) will be reported along 
with 80% and 95% two-sided confidence intervals.
10.4.3 Analysis of the Secondary Endpoint
The safety of the treatment will be monitored, and any toxicities identified will be reported by type 
and grade.
Overall survival will be calculated from the on-study date using the Kaplan-Meier method.
10.4.4 Safety Analyses
Patients will be assessed for toxicity by reporting the grades of toxicity and the type of toxicity 
observed for all patients. 
10.4.5 Baseline Descriptive Statistics
Baseline demographic characteristics will be reported.
10.4.6 Planned Interim Analyses
There are no planned interim analyses for efficacy.
10.4.7 Exploratory Analyses
The following objectives will result in descriptive or comparative analyses when adequate data 
exist to perform them:
-
To measure changes in immune cell population in the peripheral blood and tumors of 
patients treated on this study, including CD4+, CD8+, MDSCs, MAITS NKT cells, 
IFNgamma signaling pathway, perforin and granzyme. 
-
To measure secreted cytokines in the peripheral blood to monitor the relative level change 
with the treatment
-
To measure changes in bile acid in the peripheral blood of patients treated in this study.
-
To evaluate tadalafil and oral vancomycin pharmacokinetic level to monitor compliance 
and association between the level and response rate or immune parameters
-
To evaluate TCR beta sequencing to analyze the T cell response from PBMC and tumor 
samples 
-
To assess gene and protein expression and metabolite profile from tumor to evaluate the 
association of the change immune cell signature, signaling pathways and metabolite with 
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-
To measure changes in gut microbiome in the stool of patients treated on this study.
-
To evaluate the incidence of vancomycin-resistant enterococci (VRE) development during 
the trial.
-
To evaluate the relationship between the response rate, overall survival and immune 
parameters, gut microbiome, and bile acid component in patients treated on this study.
Any exploratory evaluations which generate quantitative measures will be done using descriptive 
statistics including confidence intervals when appropriate. Any statistical tests performed for 
evaluation of exploratory objectives will be done without formal adjustment for multiple 
comparisons, but in the context of the number of tests performed.
11 COLLABORATIVE AGREEMENT
11.1 Cooperative Research and Development Agreement (CRADA)
The CRADA for this protocol (03259) has been executed between Thoracic & GI Malignancies 
Branch, NCI, NIH and Bristol-Myers Squibb, the manufacturer of nivolumab. 
12 HUMAN SUBJECTS PROTECTIONS 
12.1 Rationale for Subject Selection
This study was designed to include women and minorities but was not designed to measure 
differences of intervention effects. Males and females will be recruited with no preference to 
gender. No exclusion to this study will be based on race. Minorities will actively be recruited to 
participate.
12.2 Participation of Children
Children (younger than 18 years) will not be included in this protocol due to the limited data on 
nivolumab in children and the different biology of childhood malignancy.
12.3 Participation of Subjects Unable to Give Consent
Adults unable to consent are excluded from enrolling in the protocol. However, it is possible that 
subjects enrolled in the protocol may permanently lose the capacity to consent for themselves 
during the course of this study.  If a subject loses the capacity to consent during the course of the 
study for reasons unrelated to protocol participation and is receiving direct benefit from the study 
therapy in the opinion of the PI, the subject may be retained on the study therapy with the 
permission of the alternate decision maker. If the permanent loss of capacity is due to toxicity from 
the study therapy or disease progression, the participant will be removed from study therapy as 
required per protocol, but may be retained on study to be followed for safety reasons and/or overall 
survival as described in the Study Calendar 3.4 The follow-up procedures involve non-invasive, 
low risk studies. The benefit of a safety follow up would outweigh the risks of such procedures. 
All subjects 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 to 
assess ongoing capacity of the subjects and to identify a LAR, as needed.
Please see section 12.6.1 for consent procedure.

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12.4 Evaluation of Benefits and Risks/Discomforts for All Participants
12.4.1 Benefits
The study drug may help to control the disease. The results may help the investigators learn more 
about the disease and develop new treatments for patients with this disease.
12.4.2 Risks
The primary risk to patients participating in this research study is from the toxicity of nivolumab, 
oral vancomycin and tadalafil.
All care will be taken to minimize study treatment side effects, but they can be unpredictable in 
nature and severity. Patients will be examined and evaluated prior to enrollment. All evaluations 
to monitor the treatment of patients will be recorded in the patient chart. If patients suffer any 
physical injury as a result of the participation in this study, immediate medical treatment is 
available at the Clinical Center, National Cancer Institute, Bethesda, Maryland.
Although no compensation is available, any injury will be evaluated and treated in keeping with 
the benefits or care to which patients are entitled under applicable regulations. In all publications 
and presentations resulting from this trial, patients’ anonymity will be protected to the maximum 
extent possible. Authorized personnel from the National Cancer Institute (NCI) and Food and Drug 
Administration (FDA) or other regulatory authorities may have access to research files in order to 
verify that patients’ rights have been safeguarded. In addition, patient names will be given to the 
Central Registration to register and verify patients’ eligibility.
12.4.2.1 Risk of Biopsy
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.
12.4.2.2 Risks of Sedation
Biopsies will be done under sedation. Potential side effects of sedation include headache, nausea 
and drowsiness. These side effects usually go away quickly.
12.4.2.3 Risks of exposure to Ionizing Radiation 
The study will involve radiation from the following sources:
•
Up to 6 CT scans per year for disease assessment
•
2 CT scans for the collection of 2 mandatory biopsies
Subjects in this study may have up to 6 CT scans and 2 CT guided biopsies and be exposed to 
approximately 8.2 rem per year. This amount is more than would be expected from everyday 
background radiation. Being exposed to excess radiation can increase the risk of cancer. The risk 
of getting cancer from the radiation exposure in this study is 0.8 out of 100 (0.8%) and of getting 
a fatal cancer is 0.4 out of 100 (0.4%). 
12.4.2.4 Risks of CT Scans
In addition to the radiation risks discussed above, CT scans may include the risks of an allergic 
reaction to the contrast. Participants might experience hives, itching, headache, difficulty breathing, 
increased heartrate and swelling.

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12.4.2.5 Blood Collection Risks
Risks of blood draws include pain and bruising in the area where the needle is placed, 
lightheadedness, and rarely, fainting. When large amounts of blood are collected, low red blood 
cell count (anemia) can develop. 
12.4.2.6 Risks of EKG
Risks include some minor skin irritation from the electrodes
12.4.2.7 Stool Collection or Rectal Swabs
There is no physical risk involved with stool or rectal swab collection.
12.4.2.8 Questionnaires Risk
Questionnaires may contain questions that are sensitive in nature. The participants are asked to 
only answer questions they are comfortable with
12.4.2.9 Other Risks
Risks include the possible occurrence of any of a range of side effects which are listed in the 
Consent Document or this protocol document. Frequent monitoring for adverse effects will help 
to minimize the risks associated with administration of the study agents.
12.4.2.10
Risk of Losing Data 
This includes the risk that data obtained during this study, including 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
12.4.2.11
Non-Physical Risks of Genetic Research
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.
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
12.5 Risks/Benefits Analysis for All Participants
It is possible that treatment on this protocol may reduce tumor burden or lessen symptoms caused 
by the cancer. While treatment on this protocol may not individually benefit subjects, the 
knowledge gained from this study may help others in the future who have gastric cancer. Potential 
risks include the possible occurrence of any of a range of side effects listed. The risks and benefits 
of participation for adults who become unable to consent, are no different than those described for 
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12.6 Consent Process and Documentation
The informed consent document will be provided as a physical or electronic document to the 
participant or consent designee(s) as applicable for review prior to consenting. A designated study 
investigator will carefully explain the procedures and tests involved in this study, and the 
associated risks, discomforts and benefits. In order to minimize potential coercion, as much time 
as is needed to review the document will be given, including an opportunity to discuss it with 
friends, family members and/or other advisors, and to ask questions of any designated study 
investigator. A signed informed consent document will be obtained prior to entry onto the study.
The initial consent process as well as re-consent, when required, may take place in person or 
remotely (e.g., via telephone or other NIH approved remote platforms used in compliance with 
policy, including HRPP Policy 303) per discretion of the designated study investigator and with 
the agreement of the participant. Whether in person or remote, the privacy of the subject will be 
maintained. Consenting investigators (and participant, when in person) will be located in a private 
area (e.g., clinic consult room). When consent is conducted remotely, the participant will be 
informed of the private nature of the discussion and will be encouraged to relocate to a more private 
setting if needed.
Consent will be documented with required signatures on the physical document (which includes 
the printout of an electronic document sent to participant) or as described below, with a manual 
(non-electronic) signature on the electronic document.  When required, witness signature will be 
obtained similarly as described for the investigator and participant.
Manual (non-electronic) signature on electronic document:
When a manual signature on an electronic document is used for the documentation of consent at 
the NIH Clinical Center, this study will use the following to obtain the required signatures:

Adobe platform (which is not 21 CFR Part 11 compliant); or,  

iMedConsent platform (which is 21 CFR Part 11 compliant)
During the consent process, participants and investigators will view individual copies of the 
approved consent document on screens at their respective locations (if remote consent); the same 
screen may be used when in the same location, but is not required.  
Both the investigator and the participant will sign the document using a finger, stylus or mouse. 
Note: Refer to the CCR SOP PM-2, Obtaining and Documenting the Informed Consent Process 
for additional information (e.g., verification of participant identity when obtaining consent 
remotely) found here.
12.6.1 Consent Process for Adults Who Lack Capacity to Consent to Research 
Participation
For participants addressed in section 12.3, an LAR will be identified consistent with Policy 403 
and informed consent obtained from the LAR, as described in Section 12.6.
13 REGULATORY AND OPERATIONAL CONSIDERATIONS
13.1 Study Discontinuation and Closure
This study may be temporarily suspended or prematurely terminated if there is sufficient 
reasonable cause. Written notification, documenting the reason for study suspension or termination, 

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will be provided by the suspending or terminating party to study participants. If the study is 
prematurely terminated or suspended, the Principal Investigator (PI) will promptly inform study 
participants, the Institutional Review Board (IRB), and sponsor and will provide the reason(s) for 
the termination or suspension. Study participants will be contacted, as applicable, and be informed 
of changes to study visit schedule.
 Circumstances that may warrant termination or suspension include, but are not limited to:

Determination of unexpected, significant, or unacceptable risk to participants

Demonstration of efficacy that would warrant stopping 

Insufficient compliance to protocol requirements

Data that are not sufficiently complete and/or evaluable

Determination that the primary endpoint has been met

Determination of futility
Study may resume once concerns about safety, protocol compliance, and data quality are addressed, 
and satisfy the sponsor, IRB and as applicable, Food and Drug Administration (FDA).
13.2 Quality Assurance and Quality Control
The clinical site will perform internal quality management of study conduct, data and biological 
specimen collection, documentation and completion. An individualized quality management plan 
will be developed to describe a site’s quality management.
Quality control (QC) procedures will be implemented beginning with the data entry system and 
data QC checks that will be run on the database will be generated. Any missing data or data 
anomalies will be communicated to the site(s) for clarification/resolution.
Following written Standard Operating Procedures (SOPs), the monitors will verify that the clinical 
trial is conducted and data are generated and biological specimens are collected, documented 
(recorded), and reported in compliance with the protocol, International Conference on 
Harmonisation Good Clinical Practice (ICH GCP), and applicable regulatory requirements (e.g., 
Good Laboratory Practices (GLP), Good Manufacturing Practices (GMP)). 
The investigational site will provide direct access to all trial related sites, source data/documents, 
and reports for the purpose of monitoring and auditing by the sponsor, and inspection by local and 
regulatory authorities.
13.3 Conflict of Interest Policy
The independence of this study from any actual or perceived influence, such as by the 
pharmaceutical industry, is critical. Therefore, any actual conflict of interest of persons who have 
a role in the design, conduct, analysis, publication, or any aspect of this trial will be disclosed and 
managed. Furthermore, persons who have a perceived conflict of interest will be required to have 
such conflicts managed in a way that is appropriate to their participation in the design and conduct 
of this trial. The study leadership in conjunction with the NCI has established policies and 
procedures for all study group members to disclose all conflicts of interest and will establish a 
mechanism for the management of all reported dualities of interest.

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13.4 Confidentiality and Privacy
Participant confidentiality and privacy is strictly held in trust by the participating investigators, 
their staff, and the sponsor. This confidentiality is extended to cover testing of biological samples 
and genetic tests in addition to the clinical information relating to participants. Therefore, the study 
protocol, documentation, data, and all other information generated will be held in strict confidence. 
No information concerning the study or the data will be released to any unauthorized third party 
without prior written approval of the sponsor. 
All research activities will be conducted in as private a setting as possible.
The study monitor, other authorized representatives of the sponsor, representatives of the 
Institutional Review Board (IRB), and/or regulatory agencies may inspect all documents and 
records required to be maintained by the investigator, including but not limited to, medical records 
(office, clinic, or hospital) and pharmacy records for the participants in this study. The clinical 
study site will permit access to such records.
The study participant’s contact information will be securely stored at each clinical site for internal 
use during the study. At the end of the study, all records will continue to be kept in a secure location 
for as long a period as dictated by the reviewing IRB, Institutional policies, or sponsor 
requirements.
Study participant research data, which is for purposes of statistical analysis and scientific reporting, 
will be transmitted to and stored at the NCI CCR. This will not include the participant’s contact or 
identifying information. Rather, individual participants and their research data will be identified 
by a unique study identification number. The study data entry and study management systems used 
by the clinical site(s) and by NCI CCR research staff will be secured and password protected. At 
the end of the study, all study databases will be archived at the NIH.
To further protect the privacy of study participants, a Certificate of Confidentiality has been issued 
by the National Institutes of Health (NIH). This certificate protects identifiable research 
information from forced disclosure. It allows the investigator and others who have access to 
research records to refuse to disclose identifying information on research participation in any civil, 
criminal, administrative, legislative, or other proceeding, whether at the federal, state, or local level. 
By protecting researchers and institutions from being compelled to disclose information that would 
identify research participants, Certificates of Confidentiality help achieve the research objectives 
and promote participation in studies by helping assure confidentiality and privacy to participants.
14 PHARMACEUTICAL INFORMATION
14.1 Nivolumab (133973)
14.1.1 Source:
Bristol Myers Squibb will supply investigational nivolumab. Only 100mg, 10mg/mL nivolumab 
vials will be provided for this study. There are 5 vials per carton.
14.1.2 Toxicity: 
Nivolumab is most commonly associated with immune-related adverse reactions. Most of these, 
including severe reactions, resolved following initiation of appropriate medical therapy or 
withdrawal of nivolumab.
Refer to investigator brochure for detailed toxicity information.

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14.1.3 Formulation and Preparation
Nivolumab Injection is a clear to opalescent, colorless to pale yellow liquid; light (few) particulates 
may be present. The drug product is a sterile, nonpyrogenic, single-use, isotonic aqueous solution 
formulated in sodium citrate, sodium chloride, mannitol, diethylenetriamine pentacetic acid 
(pentetic acid) and polysorbate 80 (Tween® 80), pH 6.0.
Nivolumab injection can be infused undiluted (10 mg/mL) or diluted with 0.9% Sodium Chloride 
Injection, USP or 5% Dextrose, USP to concentrations no less than 1 mg/mL. 
14.1.4 Stability and Storage 
Store intact vials at 2°C to 8°C (36°F to 46°F); do not freeze. Protect from light. Do not shake. 
Shelf-life surveillance of the intact vials is ongoing.
Store the prepared infusion solution at room temperature or refrigerated. Do not freeze solutions 
prepared for infusion. Refer to investigator brochure for detailed stability information.
14.1.5 Administration Procedures
Intravenous infusion 30 to 60 minutes. Do not administer as an IV push or bolus injection.
Administer through a 0.2 micron to 1.2-micron pore size, low-protein binding in-line filter. 
14.2 Tadalafil
14.2.1 Source
Tadalafil tablets will be provided by the NIH Clinical Center Pharmacy according to standard 
pharmacy procedures.
14.2.2 Description
Tadalafil is a selective inhibitor of cyclic guanosine monophosphate (cGMP)-specific 
phosphodiesterase type 5 (PDE5). Tadalafil has the empirical formula C22H19N3O4 representing 
a molecular weight of 389.41. The chemical designation is pyrazino [1′,2′:1,6] pyrido[3,4-b] 
indole-1,4-dione, 6-(1,3-benzodioxol-5-yl)-2,3,6,7,12,12a hexahydro-2-methyl-, (6R,12aR)-. It is 
a crystalline solid that is practically insoluble in water and very slightly soluble in ethanol. 
14.2.3 How Supplied 
Tadalafil tablets are available in 2.5 mg, 5 mg, 10 mg and 20 mg. Excipient information may vary 
depending on manufacturer (particularly for generics). Consult specific product labeling for 
detailed information.
Appearance of the generic drug product may differ depending on the manufacturer and patients 
will be notified of this difference.
14.2.4 Toxicity
Very common (in 100 people receiving tadalafil, 11 may have):

headaches
Common risks (in 100 people receiving tadalafil, more than 2 and up to 10 may have):

stomach upset,

pain in belly,

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
diarrhea,

nausea,

reflux disease,

muscle or back pain, pain in arms and legs,

flushing (redness or warmth of the face, neck, or chest),

flu-like symptoms (such as stuffy nose, sneezing, or sore throat),

infection,

cough,
Rare risks (in 100 people receiving tadalafil, 1 or fewer may have):

fatigue,

face swelling,

chest pain,

heart problems,

low blood pressure,

elevated liver enzyme in the blood

dry mouth,

hemorrhoids,

dizziness,

luck of sleep,

rash,

blurred vision, changes in color vision, eye pain, swelling around eyes

abnormal ejaculation

a painful or prolonged erection lasting 4 or more hours;

sudden decreased vision (including permanent blindness, in one or both eyes);

a sudden decrease or loss of hearing, sometimes with ringing in the ears and dizziness
 
14.2.5 Administration: 
Please see section 3.2.2. 
14.3 Oral Vancomycin
14.3.1 Source
Vancomycin capsules will be provided by the NIH Clinical Center Pharmacy according to standard 
pharmacy procedures
14.3.2 Mode of Action
Vancomycin is an antibacterial drug that inhibits bacterial cell wall synthesis by blocking 
glycopeptide polymerization through binding tightly to D-alanyl-D-alanine portion of cell wall 
precursor.

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14.3.3 Description
Vancomycin CAPSULES for oral administration contain chromatographically purified 
vancomycin hydrochloride, a tricyclic glycopeptide antibiotic derived from Amycolatopsis 
orientalis 
(formerly 
Nocardia 
orientalis), 
which 
has 
the 
chemical 
formula 
C66H75Cl2N9O24•HCl. The molecular weight of vancomycin hydrochloride is 1485.73; 500 mg 
of the base is equivalent to 0.34 mmol. The capsules contain vancomycin hydrochloride equivalent 
to 125 mg (0.08 mmol) or 250 mg (0.17 mmol) vancomycin. The capsules also contain F D & C 
Blue No. 2, gelatin, iron oxide, polyethylene glycol, titanium dioxide, and other inactive 
ingredients. 
14.3.4 How Supplied
Vancomycin capsules are available in 125 mg and 250 mg strength. Excipient information may 
vary depending on the manufacturer (particularly for generics). Consult specific product labeling 
for detailed information.
Appearance of the generic drug product may differ depending on the manufacturer and patients 
will be notified of this difference.  
14.3.5 Administration: 
Please see section 3.2.3.
14.3.6 Storage 
Store at controlled room temperature, 59° to 86°F (15° to 30°C).

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

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16.2 Appendix B: Patient’s Medication Diary ________________________
Cycle___________________       Patient’s ID ___________________
INSTRUCTIONS TO THE PATIENT:
1.
Complete one form for each cycle of treatment
2.
You will take vancomycin 125 mg every 6 hours a day for 3 weeks on and 1 week off
3.
You will take tadalafil 10 mg tadalafil daily.
4.
Record the date, the number of tablets that you took, and when you took them.
5.
If you have any comments or notice any side effects, please record them in the comments column.
6.
Please bring this form and your bottles (even it is empty) when you come for your clinic visits.
Vancomycin, time
Day
Date
Tadalafil,
time
Comments
1
2
3
4
5
6
7
8
9
10

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Vancomycin, time
Day
Date
Tadalafil,
time
Comments
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26

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Vancomycin, time
Day
Date
Tadalafil,
time
Comments
27
28
Patient’s signature:  ___________________________________________________

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16.3 Appendix C: Management Algorithms For Endocrinopathy, Gastrointestinal, Hepatic, Neurological, Pulmonary, Renal, 
And Skin Adverse Events

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16.4 Appendix D: Infusion-related Reaction Management Algorithm
Infusion-related Reactions
Severity Grade of 
the Event (CTCAE 
5.0
Dose Modifications
Toxicity Management
Any Grade
General Guidance
For Any Grade:
−
Manage per institutional standard at the discretion of 
investigator.
−
Monitor patients for signs and symptoms of infusion-
related reactions (e.g., fever and/or shaking chills, 
flushing and/or itching, alterations in heart rate and 
blood pressure, dyspnoea or chest discomfort, or skin 
rashes) and anaphylaxis (e.g., generalized urticaria, 
angioedema, wheezing, hypotension, or tachycardia).
Grade 1 or 2
For Grade 1:

The infusion rate of study drug/study 
regimen may be decreased by 50% or 
temporarily interrupted until resolution of 
the event.
For Grade 2:

The infusion rate of study drug/study 
regimen may be decreased 50% or 
temporarily interrupted until resolution of 
the event.

Subsequent infusions must be given at 
50% of the initial infusion rate.
For Grade 1 or 2:
−
Acetaminophen and/or antihistamines may be 
administered per institutional standard at the discretion 
of the investigator.
−
Consider premedication per institutional standard prior 
to subsequent doses.
−
Steroids should not be used for routine premedication of 
Grade ≤2 infusion reactions.
Grade 3 or 4
For Grade 3 or 4:

Permanently discontinue study 
drug/study regimen.
For Grade 3 or 4:
−
Manage severe infusion-related reactions per 
institutional standards (e.g., IM epinephrine, followed 
by IV diphenhydramine and ranitidine, and IV 
glucocorticoid).

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CTCAE - Common Terminology Criteria for Adverse Events; IM - Intramuscular; IV Intravenous; NCI - National Cancer Institute.
Non-immune-mediated Reactions
Severity Grade of the 
Event (CTCAE 5.0)
Dose Modifications
Toxicity 
Management
Any Grade

Note: Dose modifications are not required for AEs not deemed to 
be related to study treatment (i.e., events due to underlying 
disease) or for laboratory abnormalities not deemed to be 
clinically significant.
Grade 1

No dose modifications.
Grade 2

Hold study drug/study regimen until resolution to ≤Grade 1 or 
baseline.
Grade 3

Hold study drug/study regimen until resolution to ≤Grade 1 or 
baseline.

For AEs that downgrade to ≤Grade 2 within 7 days or resolve to 
≤Grade 1 or baseline within 14 days, resume study drug/study 
regimen administration. Otherwise, discontinue study drug/study 
regimen.
Grade 4

Discontinue study drug/study regimen (Note: For Grade 4 labs, 
decision to discontinue should be based on accompanying clinical 
signs/symptoms, the Investigator’s clinical judgment, and 
consultation with the Sponsor.).
Treat accordingly, 
as per institutional 
standard.
AE - Adverse event; CTCAE - Common Terminology Criteria for Adverse Events; NCI - National Cancer Institute.

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16.5 Appendix E: Modified immune-related response criteria (irRC)
This new classification is based on the recent learning from clinical studies with cancer 
immunotherapies that even if some new lesions appear at the beginning of a treatment or if the 
total tumor burden does not increase substantially, tumor regressions or stabilizations might still 
occur later. The irRC were created using bi-dimensional measurements (as previously widely used 
in the World Health Organization criteria). For this trial, the concepts of the irRC are combined 
with RECIST 1.1 to come up with the modified irRC. 
For modified irRC, only target and measurable lesions are taken into account. In contrast to the 
RECIST 1.1 criteria, the modified irRC criteria (a) require confirmation of both progression and 
response by imaging at 6 weeks after initial imaging and (b) do not necessarily score the 
appearance of new lesions as progressive disease if the sum of lesion diameters of target lesions 
(minimum of 10 mm per lesion, maximum of 5 target lesions, maximum of 2 per organ) and 
measurable new lesions does not increase by ≥ 20%. 
The same method of assessment and the same technique should be used to characterize each 
identified and reported target lesion(s) at baseline, during the trial, and at the end of trial visit. All 
measurements should be recorded in metric notation. The modified irRC based on RECIST 1.1 are 
displayed below. 
Overall Responses Derived from Changes in Index, Non-Index, and New Lesions

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16.6 Appendix F: Child-Pugh Classification System

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16.7 Appendix G: Pregnancy Form

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16.8 Appendix H: Food Diary for Mixed Meal Study

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16.9 Appendix I : Mixed Meal PK Sheet