A Phase 2 Study of the Safety, Efficacy, and Immune Response of CRS-207, Pembrolizumab, Ipilimumab, and Tadalafil in Patients With Previously Treated Metastatic Pancreatic Adenocarcinoma
Phase: PHASE2
Conditions: Pancreatic Cancer
Sponsor: Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins

 
 
 
 
TITLE: A Phase 2 Study of the Safety, Efficacy, and Immune Response of CRS-207, 
Pembrolizumab, Ipilimumab, and Tadalafil in Patients with Previously Treated Metastatic 
Pancreatic Adenocarcinoma 
 
Johns Hopkins Protocol #:  
 
J2180, IRB00291762 
 
NCT Registration Number:             05014776 
 
Principal Investigator:  
Dr. Katherine Bever 
Sidney Kimmel Comprehensive Cancer Center, 
 
Johns Hopkins University 
 
 
 
 
 
Co-Principal Investigator: 
Dr. Dung Le  
 
Sidney Kimmel Comprehensive Cancer Center, 
 
Johns Hopkins University 
 
 
 
  
 
IND Sponsor: 
Dr. Elizabeth Jaffee 
 
 
Sidney Kimmel Comprehensive Cancer Center, 
 
 
Johns Hopkins University 
 
 
 
 
 
 
 
 
 
   
 
 
 
IND:  
 
 
 
 
 
 
      BB IND 16147 
 
Commercial Agents: 
 
KEYTRUDA® (Pembrolizumab, MK-3475) 
  
 
YERVOY® (Ipilimumab, BMS-734016) 
Tadalafil (generic) 
 
Johns Hopkins University Supplied Agent:  CRS-207 (Lm ΔactA/ΔinlB/hMesothelin) 
 
Date of Issue: 
Version 1/ June 28, 2021 
Version 1.1/ September 10, 2021 
Version 2.0/ March 18, 2022 
Version 3.0/ November 14, 2022 
Version 4.0/ January 13, 2023 
Version 5.0/ January 5, 2024 

J2180 / Version 5.0/ January 5, 2024 
 
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TABLE OF CONTENTS 
 
1. 
OBJECTIVES ....................................................................................................................4 
1.1 
Primary Objective ....................................................................................................4 
1.2 
Secondary Objective ................................................................................................4 
1.3 
Exploratory Objectives ............................................................................................4 
1.4 
Study Design ............................................................................................................5 
2. 
BACKGROUND ................................................................................................................7 
2.1 
Study Disease ...........................................................................................................7 
2.2 
Rationale ..................................................................................................................8 
3. 
PATIENT SELECTION .................................................................................................13 
3.1 
Inclusion Criteria ...................................................................................................13 
3.2 
Exclusion Criteria ..................................................................................................14 
3.3 
Inclusion of Women and Minorities ......................................................................17 
4. 
TREATMENT PLAN ......................................................................................................17 
4.1 
Agent Administration.............................................................................................17 
4.2 
General Concomitant Medication and Supportive Care Guidelines ......................20 
4.3 
Prohibited and/or Restricted Medications and Devices .........................................23 
4.4 
Other Restrictions and Precautions ........................................................................24 
4.5 
Antibiotic Administration ......................................................................................24 
4.6 
Definition of an Overdose for this Protocol ...........................................................26 
4.7 
Unacceptable Toxicity ...........................................................................................27 
4.8 
WOCBP, Contraception, Use in Pregnancy, Use in Nursing ................................27 
4.9 
Duration of Therapy ...............................................................................................30 
4.10 
Criteria for Removal from Treatment ....................................................................30 
4.11 
End of Treatment (EOT) Visit ...............................................................................34 
4.12 
Duration of Follow-Up ..........................................................................................34 
4.13 
Blood Cultures for CRS-207 Surveillance .............................................................35 
5. 
DOSING DELAYS/DOSE MODIFICATIONS ............................................................36 
5.1  
Dose Modifications ................................................................................................36 
5.2  
Dosing Delays ........................................................................................................36 
6. 
ADVERSE EVENTS:  LIST AND REPORTING REQUIREMENTS ......................38 
6.1 
Definitions..............................................................................................................38 
6.2 
Assessment of Causality ........................................................................................40 
6.3 
Expectedness ..........................................................................................................41 
6.4 
Handling of Expedited Safety Reports ..................................................................41 
6.5 
Reporting................................................................................................................41 
7. 
PHARMACEUTICAL INFORMATION ......................................................................44 
7.1  
Tadalafil .................................................................................................................44 
7.2  
CRS-207 .................................................................................................................46 

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7.3  
Pembrolizumab (KEYTRUDA®) .........................................................................49 
7.4  
Ipilimumab (YERVOY®) .....................................................................................51 
8. 
CORRELATIVE/SPECIAL STUDIES .........................................................................53 
8.1 
Tumor Tissue Studies ............................................................................................53 
8.2 
Peripheral Blood Mononuclear Cells (PBMCs) ....................................................54 
8.3 
Serum and Plasma Marker Studies ........................................................................54 
8.4 
Diagnostic Tissue Samples ....................................................................................54 
8.5 
Genomic Analysis ..................................................................................................54 
9. 
STUDY CALENDAR ......................................................................................................56 
10.  
STUDY ENDPOINTS ......................................................................................................60 
10.1 
Primary Endpoint ...................................................................................................60 
10.2 
Secondary Endpoint ...............................................................................................60 
10.3 
Exploratory Endpoints ...........................................................................................60 
11. 
 DATA REPORTING / REGULATORY REQUIREMENTS ....................................61 
11.1 
Data Collection and Processing .............................................................................61 
11.2 
Safety Meetings .....................................................................................................61 
11.3 
Monitoring .............................................................................................................62 
11.4 
Study Documentation.............................................................................................62 
12. 
STATISTICAL CONSIDERATIONS ...........................................................................64 
12.1 
Study Design/Endpoints.........................................................................................64 
12.2 
Safety Analysis ......................................................................................................65 
REFERENCES .............................................................................................................................66 
APPENDIX A: Performance Status Criteria ............................................................................68 
APPENDIX B: Response Evaluation Criteria in Solid Tumors (RECIST) 1.1 
Criteria for Evaluating Response in Solid Tumors ......................................................69 
APPENDIX C: Description of the iRECIST Process For Assessment of Disease 
Progression .......................................................................................................................72 
 

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1. OBJECTIVES 
 
1.1 
Primary Objective 
 
To determine the objective response rate (ORR) using Response Evaluation Criteria for 
Solid Tumors (RECIST 1.1) in subjects with previously treated metastatic pancreatic 
cancer treated with tadalafil, pembrolizumab, ipilimumab, and CRS-207. 
 
1.2 
Secondary Objective 
 
1.2.1 To assess safety and characterize toxicities when combining tadalafil, 
pembrolizumab, ipilimumab, and CRS-207 in subjects with metastatic pancreatic 
adenocarcinoma. 
 
1.3 Exploratory Objectives 
 
1.3.1 To assess the overall survival (OS).  
 
1.3.2  To assess progression free survival (PFS), disease control rate (DCR), duration of 
response (DOR), and time to objective response (TOR) by RECIST 1.1. 
 
1.3.3  To assess ORR, PFS, DCR, DOR, and TOR by immune Response Evaluation 
Criteria for Solid Tumors (iRECIST).  
 
1.3.4  To measure tumor marker kinetics (CA 19-9) in subjects receiving treatment and 
correlate with OS, PFS, and best overall response. 
 
1.3.5  To collect peripheral blood mononuclear cells (PBMC), plasma, and serum to 
identify potential therapeutic targets, biomarkers and predictors of response (OS, 
PFS and best overall response) and autoimmune toxicity.  
 
• 
Measure pre- and post-treatment changes in PBMCs including effector, 
helper, and regulatory T cells, NK cells, monocytes, and macrophages 
through cell phenotyping analysis and gene expression profiling.  
 
• 
Correlate induction of Listeria monocytogenes (Lm)- and mesothelin 
antigen-specific T cell responses and changes to the T cell epitope repertoire 
with OS, PFS, and best overall response. 
 
• 
Correlate telomere length of lymphocytes to help predict response (OS, 
PFS, and best overall response).  
 
• 
Correlate the induction of anti-thyroglobulin and anti-galectin-3 antibody 
responses with response (OS, PFS, and best overall response).   
 
• 
Proteomic approaches will be used on pre- and post-treatment sera to 
identify targets and biomarkers of response (OS, PFS, and best overall 
response) or toxicity. 

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1.3.6  To collect archived tissue and pre- and post-treatment biopsies to test for predictors 
of response (OS, PFS, and best overall response) and future targets for 
combinatorial therapy.   
 Immunohistochemistry (IHC) and/or gene expression profiling will be used 
to compare the nature of tumors and immune infiltrates for responders 
versus non-responders.   
 Next-generation sequencing of T cell receptor (TCR) genes may be used to 
compare the tumor infiltrating T cell repertoire in responders and non-
responders 
 Up-regulation of immune inhibitory molecules (such as programmed death-
ligand 1 [PD-L1]) will be evaluated in the pre- and post-treatment samples. 
 
 Proteomic approaches to quantify protein expression and activation of 
specific signaling pathways in tumors from responders versus non-
responders. 
 
1.4 
Study Design 
 
This is an open-label, phase 2 study to evaluate the safety and clinical activity of tadalafil, 
pembrolizumab, ipilimumab, and CRS-207 in subjects with metastatic pancreatic adenocarcinoma 
who have progressed after at least 1 prior chemotherapy regimen.  
 
The primary endpoint of this study is objective response rate (ORR) using RECIST. The treatment 
will be considered inactive and of no interest for further evaluation if the ORR is 5% or less and 
considered active if the ORR is 30% or greater.  
 
A total of 17 patients will be enrolled on the study. If a total of 3 or more responses are observed, 
we will conclude that the ORR is higher than 5% and that the regimen warrants further study. This 
design has 92% power to reject the null hypothesis of an ORR of 5% in favor of the alternative 
hypothesis of 30%, with one-sided type 1 error 0.0502 (target type I error of 0.1). Additional 
patients may be enrolled as needed until at least 10 paired biopsies are obtained, but will not be 
included in the primary analysis of efficacy endpoint. 
 
The study will consist of a screening period (within 28 days of first dose), a treatment period per 
the table below, and a follow-up period.  
  
Subjects will receive treatment every 3 weeks for 6 cycles of treatment within a course. A course 
of treatment will be 18 weeks and courses can be repeated. The treatment schedule can be found 
in Table 1. Subjects will come to the clinic for dosing and/or assessments on Days 1 and 2 of each 
cycle and additional days for safety and immune monitoring follow-up per the study schedules in 
Section 9. 
 
Dose reductions are only allowed for tadalafil per Section 5.1. If the investigator assesses a drug-
related toxicity (that requires discontinuation) to be related to an individual component of the 
treatment schedule, dosing for that study drug alone may be discontinued while dosing is delayed 

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until the subject meets criteria to resume treatment of the other study drugs. The relationship to the 
discontinued study drug should be well documented in the source documents and permission from 
the 
Principal 
Investigator 
and 
Co-Principal 
Investigator 
needs 
to be obtained 
prior to continuation with the other study drugs. As of protocol version 5.0, enrollment to this 
study has been completed, and any patients that remain on study after February 22, 2024 will 
discontinue CRS-207 (no further drug supply) and may remain on study to continue to receive 
pembrolizumab, ipilimumab, and tadalafil. 
  
Enrollment for the first three patients will be staggered by three weeks to evaluate patients for 
adverse events before the next patient is enrolled (the fourth patient may be enrolled 3 weeks after 
the third patient). The proportion of treated subjects with unacceptable toxicity will be monitored 
routinely. Complete unacceptable toxicity criteria can be found in Section 4.7.  
 
At the investigator’s discretion, subjects may receive additional courses of the assigned treatment 
regimen if they are clinically stable and meet dosing eligibility criteria. All subjects may continue 
in the treatment period up to a maximum of 2 years, or until discontinuation due to unacceptable 
toxicity, lack of clinical benefit as determined by the investigator, subject withdrawal, or 
termination of the study by IND Sponsor. Subjects that begin a new course prior to the 2 year cut-
off may complete that course prior to coming off study. Subjects may continue on treatment with 
radiographic disease progression if subject is clinically stable and investigator believes the 
treatment is providing benefit. Criteria for removal from treatment are found in Section 4.10. 
Subjects will return to the study site 28 (±7) days after the final administration of study treatment 
for an end-of-treatment (EOT) evaluation. To eliminate any potentially residual CRS-207, subjects 
will initiate a 7-day course of antibiotics 7 days after the subject’s last dose of CRS-207 and prior 
to receiving any subsequent cancer-related therapy (or if the patient will be having a semi-
permanent indwelling device placed while on study) per Section 4.5. Subjects who are still 
receiving treatment at the time of study close may complete the current treatment course and the 
EOT evaluation prior to transitioning to long-term follow-up. Subjects will be considered in the 
treatment period until 28 days after the last dose of study drug. Blood cultures through a peripheral 
vein and also through a central line (if applicable) will be collected to monitor for the presence of 
CRS-207 per Section 4.13. 
 
After completion of treatment and EOT assessments, all subjects, including those who did not 
receive treatment, will continue to be followed every three months (+/- 2 weeks) by telephone, e-
mail, or optional clinic visit until death, withdrawal of consent, or closure of study.  Subjects will 
also be contacted at 90 days (+14 day reporting window) from the last dose of pembrolizumab/ 
ipilimumab or 28 days (+7 day reporting window) from the last dose of tadalafil or CRS-207 if the 
subject is no longer receiving pembrolizumab/ipilimumab due to toxicity, whichever reporting 
period is longer. Information on survival and new cancer therapies will be collected. In addition, 
all subjects that received at least one dose of CRS-207 will be monitored for CRS-207 infection 
for one year per Section 4.13. 
 
All subjects who discontinue study treatment should continue to be monitored for disease status 
by radiologic imaging every approximately every two months per standard of care until: 1) the 
start of a new antineoplastic therapy (information of the new cancer therapy will be collected), 2) 
disease progression, 3) death, 4) withdrawal of consent, or 5) the close of the study, whichever 
occurs first.   

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All subjects will be followed after their last dose of study drug for the development of adverse 
events (AEs) and serious adverse events (SAEs) as described in Section 6.5.1.  
 
The primary analysis will be conducted when all treated subjects have documented response or 
progression by RECIST, or have discontinued the study. Information on survival may continue to 
be gathered for supplementary analyses after the completion of the primary analysis. At the 
conclusion of the study, all remaining subjects will be offered enrollment in a long-term follow-
up study and continue to be followed for survival. 
 
2. BACKGROUND 
 
2.1 
Study Disease 
 
Despite decades of basic and clinical research, effective therapy for the treatment of patients with 
pancreatic ductal adenocarcinoma (PDA) remains one of the greatest unmet clinical needs in 
oncology today. Currently, PDA accounts for approximately 8% of all cancer-related mortality 
and has the lowest 5-year survival rate among all cancer types in the United States. PDA is 
currently the 4th leading cause of death from cancer in the U.S. with estimates in 2021 for 60,430 
people diagnosed and about 48,220 dying from the disease1.  Worldwide it will claim more than 
300,000 lives this year2.  It is projected that by 2030, pancreatic cancer will become the second 
leading cause of cancer-related death in the US3.  
 
Most patients are initially diagnosed with advanced disease that is inoperable with median survival 
of less than 1 year. Patients with advanced disease are usually treated with chemotherapy, with the 
intent of prolonging survival and palliating symptoms (pain, weight loss and decrease in 
performance status). From 1997, gemcitabine was the standard chemotherapy for advanced 
pancreatic cancer after demonstrating a significant improvement in survival compared to 5-
fluorouracil (5-FU)4. Median survival was 5.65 months for gemcitabine-treated patients and 4.41 
months for 5-FU treated patients, while overall tumor response rates were 5.4% and 0%, 
respectively. 
 
Until recently, only erlotinib, an oral epidermal growth factor (EGF) inhibitor, was shown in a 
Phase 3 study to modestly improve median OS in combination with gemcitabine over gemcitabine 
alone (6.24 months for the doublet versus 5.91 months for gemcitabine alone) without a significant 
difference in ORR between the treatments5. In 2011, a Phase 2/3 trial conducted by a French 
consortium study group demonstrated FOLFIRINOX, a combined regimen of oxaliplatin, 
irinotecan, fluorouracil, and leucovorin significantly increased survival in patients with pancreatic 
cancer over gemcitabine alone. Median OS was 11.1 months versus 6.8 months for each treatment, 
respectively (hazard ratio [HR] for death, 0.57; 95% confidence interval [CI], 0.45 to 0.73; p < 
0.001). The ORR was also increased to 31.6% from 9.4% (p < 0.001). Adverse events were 
increased in the FOLFIRINOX group and 5.4% of patients in this group experienced febrile 
neutropenia6. Although FOLFIRINOX represents an efficacious regimen in pancreatic cancer, 
there are still concerns about its potential toxicity and it is being reserved for the most fit patients. 
 
In the MPACT (Metastatic Pancreatic Adenocarcinoma Clinical Trial) study, nab-paclitaxel 
combined with gemcitabine demonstrated a statistically significant and clinically meaningful 
median OS of 8.5 versus 6.7 months (HR 0.72, p < 0.0001 including a 59% increase in one-year 
survival (35% versus 22%, p=0.0002) and demonstrated double the rate of survival at two years 

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(9% versus 4%, p=0.02) as compared to gemcitabine alone in previously untreated patients with 
metastatic pancreatic cancer. nab-/gemcitabine also demonstrated a statistically significant 
improvement in key secondary endpoints compared to gemcitabine alone, including a 31% 
reduction in the risk of progression or death with a median PFS of 5.5 versus 3.7 months (HR 0.69, 
p < 0.0001) and an ORR of 23% compared to 7% (response rate ratio of 3.19, p < 0.0001)7. The 
MPACT study and regimen formed the basis for full FDA approval in September 2013 for the 
first-line treatment of metastatic adenocarcinoma of the pancreas. 
 
Nanoliposomal irinotecan in combination with 5-FU was recently approved for patients with 
previously treated pancreatic cancer.  However, it is unknown if this combination is better than 5-
FU and irinotecan (FOLFIRI) as 5-FU was the comparator and there was no benefit in patients 
who previously received irinotecan so the value of this drug is questionable in FOLFIRINOX 
treated patients8.  Therapies for patients with metastatic pancreatic cancer are urgently needed.  
 
2.2 
Rationale 
 
Therapeutic benefit of immunotherapy in pancreatic cancer remains to be seen. The poor 
antigenicity of neoplastic cells and uniquely immunosuppressive stromal compartment are likely 
significant contributors to resistance, by inhibiting T cell priming and infiltration into the tumor 
microenvironment (TME). Single agent therapy using vaccines to induce antigen-specific T cells 
or targeting immune checkpoints such as programmed death-1 (PD-1) or cytotoxic T-lymphocyte 
associated protein-4 (CTLA-4) does not address these complex immunosuppressive mechanisms 
at play both at the systemic level and in the TME.  
 
Current research evaluating responsiveness and resistance to immunotherapy has focused on tumor 
intrinsic factors and in particular, genetic sequencing data. However, these studies have had few 
revelations since PDA is typically genetically bland. Emerging data suggests that tumor extrinsic 
factors, especially the unique PDA desmoplastic stroma, orchestrates immunotherapy resistance. 
In particular, myeloid cells including tumor associated macrophages (TAMs) and myeloid derived 
suppressor cells (MDSCs) infiltrate PDA stroma and support neoplasia initiation, progression and 
metastasis. Multifunctional pathways such as CXCR4, CCR2/CCR5, and ARG1/NOS2 have been 
implicated in myeloid and stromal driven immune suppression. But it remains poorly understood 
how (i) PDA-targeted vaccine strategies with or without ICIs modulate these cells, and (ii) whether 
therapeutic targeting of these cells will enhance the activity of immunotherapy against PDA. 
 
Effective immunotherapy in pancreatic cancer may require combinations of 3 or more agents to 
overcome this tolerance.   
 
Dual Blockade with Pembrolizumab and Ipilimumab 
 
The PD-1 receptor-ligand interaction is a major pathway hijacked by tumors to suppress immune 
control9.  The normal function of PD-1, expressed on the cell surface of activated T cells under 
healthy conditions, is to down-modulate unwanted or excessive immune responses, including 
autoimmune reactions. The ligands for PD-1 (PD-L1 and PD-L2) are constitutively expressed or 
can be induced in various tumors10-13.  Binding of either PD-1 ligand to PD-1 inhibits T cell 
activation triggered through the T cell receptor.  The observed correlation of clinical prognosis 
with PD-L1 expression in multiple cancers suggests that the PD-1/PD-L1 pathway plays a critical 

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9
role in tumor immune evasion and should be considered as an attractive target for therapeutic 
intervention. 
 
CTLA-4, an activation-induced T-cell surface molecule, is a member of the CD28:B7 
immunoglobulin superfamily that competes with CD28 for B7. CTLA-4 mediated signals are 
inhibitory and turn off T cell-dependent immune responses14. The proposed mechanism of action 
for ipilimumab is interference of the interaction of CTLA-4 with B7 molecules on APCs, with 
subsequent blockade of the inhibitory modulation of T-cell activation promoted by the CTLA 4/B7 
interaction. 
 
While OPDIVO® (nivolumab), in combination with YERVOY® (ipilimumab), is currently FDA 
approved for the treatment of patients across multiple types of cancers, the combination of 
KEYTRUDA® (pembrolizumab) with ipilimumab is still under investigation. In a study 
evaluating low-dose ipilimumab (1 mg/kg) plus pembrolizumab (200 mg) following progression 
on anti-PD-1 immunotherapy in advanced melanoma, significant antitumor activity with durable 
long-term responses were observed15. In addition, the combination with low-dose ipilimumab was 
more tolerable with a grade 3-4 adverse event rate of 27%15 in comparison to the 59% reported 
with higher doses of ipilimumab in combination with nivolumab16. Another study in previously 
treated advanced non-small-cell lung cancer (NSCLC) patients also demonstrated some anti-tumor 
activity but with higher toxicity rates given the higher doses of pembrolizumab and ipilimumab17.  
 
Immunotherapies in Pancreatic Cancer 
 
CRS-207 is a live, attenuated, double-deleted Lm engineered to secrete mesothelin, a tumor-
associated antigen which is overexpressed in most pancreatic cancers. An Lm vaccine construct 
permits access to both major histocompatibility complex (MHC) class I and II antigen processing 
pathways and has the capacity to stimulate both adaptive and innate immunity. A microorganism-
based construct can naturally stimulate innate immunity via “danger signals” and toll-like 
receptors, serving as a natural vaccine adjuvant. Listeria also induces IFNβ expression through a 
stimulator of interferon genes (STING)-dependent pathway. Initial studies using CRS-207 in 
metastatic PDA (mPDA) were used in combination with GVAX pancreas vaccine (allogeneic 
pancreatic cancer cells modified to express GM-CSF) as a heterologous prime boost strategy using 
CY/GVAX Pancreas as a prime and CRS 207 as a boost vaccine. A survival benefit was observed 
compared to CY/GVAX Pancreas alone in an initial phase 2 study18. In a follow up study, 
ECLIPSE, with 3 treatment arms (A:  CY/GVAX Pancreas + CRS-207, B:  CRS-207, C:  single 
agent chemotherapy), the study did not meet its primary endpoint and Arm A did not extend 
survival compared to Arm C. There was however, a partial response in Arm A.  Although the study 
was not designed to compare Arm B to Arm C, these curves overlapped. The median survivals 
were 3.8, 5.4, and 4.6 months, respectively19.  This study suggests that vaccines will likely need to 
be combined with immune checkpoint inhibition, but also supports studying CRS-207 alone as an 
interesting vaccine platform.  
 
Based on the observation that PDAs upregulate immune checkpoint expression following vaccine 
treatment, we have undertaken several clinical trials in mPDA combining vaccines with one or 
more immune checkpoint inhibitors (several of which are ongoing: NCT03190265 and 
NCT03006302). In a completed study combining CY/GVAX Pancreas/CRS-207 with or without 
nivolumab, three patients had objective responses, and changes in the TME, including increase in 
CD8+ T cells and a decrease in CD68+ myeloid cells, were observed in long-term survivors of the 

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arm receiving nivolumab20. In addition, T cell receptor (TCR) repertoire analyses on PBL from 
patients treated with either ipilimumab +/- GVAX and GVAX+CRS-207 +/- nivolumab revealed 
that a net diversification of the peripheral TCR repertoire with ipilimumab and an increase in T 
cell clonality with nivolumab that correlated with improved survival21. These data suggested that 
ipilimumab increases the number of T cells expanded and available for vaccine priming and 
nivolumab expands the tumor specific T cells that have been primed by the vaccine, further 
supporting their use in combination. 
 
So far, we have focused on inducing and expanding cancer recognizing T cells. Our reported 
studies and preliminary data demonstrate the ability to induce T cell responses that in some patients 
results in T cell infiltration and durable tumor regressions. However, these modest therapeutic 
effects suggest that there are other immunological partners to be targeted. Multiplexed 
immunohistochemistry (mIHC) imaging of PDAs treated with neoadjuvant GVAX demonstrated 
higher PD-L1 expression in myeloid cells in the TME. Also, analysis of PDA from our prime boost 
vaccine with and without nivolumab in mPDA patients, showed that a higher myeloid inflamed 
status in pretreatment PDAs predicted shorter survival, whereas pre-treatment lymphoid cell 
populations did not predict response (Fig. 1). Moreover, preliminary analysis of the peripheral 
blood samples by mass cytometry (Cytof) demonstrated several features of circulating immune 
cells associated with better clinical outcomes, notably lower abundance of myeloid cells at 
baseline, and trends toward lower CXCR4, CXCR2 and CCR2 expression in myeloid cells, and 
higher HLADR. These results suggest the critical importance of understanding the crosstalk 
between the myeloid and T cell compartments as well as the potential utility of targeting the 
myeloid-stromal signaling to enhance antitumor activity20. 
 
Figure 1: Changes in immune cell 
composition 
are associated 
with 
survival in patients treated with 
vaccine + nivo. FFPE tissue sections 
derived from biopsies obtained at 
baseline and after GVAX/CRS207 with 
(Arm A) or without nivo (Arm B) were 
subjected to immune-detection by 
multiplexed imaging. (A) Immune cell 
densities (cell number per mm2) of 
CD8+ T cell, other lymphoid lineages 
cells, CD68+CSF1R+ tumor associated 
macrophages 
(TAMs), 
CD66b+ 
granulocytes, and other myeloid cell 
lineages are shown, comparing baseline 
and post Cy3 status among short and 
long OS groups. (B, C) Frequency of 
CD8+ T cells (B) and CD68+ myeloid 
cells 
(C) 
are 
exhibited 
as 
cell 
percentages of total CD45+ immune 
cells, comparing baseline and post Cy3 status among short and long OS groups. Cy=cycle; In B, black circles =pre-
treatment and orange squares are Cy3. Arm A: vaccine+nivo; Arm B: vaccine only. 
 
 
We have also evaluated the combination of GVAX and ipilimumab in both treatment refractory 
advanced PDA and as maintenance after induction chemotherapy for newly diagnosed mPDA. 
While GVAX + ipilimumab did not improve survival over continuation of chemotherapy in the 
maintenance treatment of non-biomarker selected mPDA, clear biologic effects on peripheral and 

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11
intratumoral immune cells were observed22. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
 
Phosphodiesterase 5 inhibitors (PDE5i) 
 
In addition to pro-apoptotic tumor effects, PDE5i have been found to have immunomodulatory 
properties including decreasing MDSCs and Tregs and increasing effector T cells in preclinical 
models. The widely available PDE5i tadalafil has also been shown to downregulate the expression 
and activity of ARG1 and NOS2, key MDSC-mediated immunosuppressive effector pathways, in 
intratumoral MDSCs in the CT26 mouse model23. Studies of tadalafil in melanoma and head and 
neck squamous cell cancer (HNSCC) patients have shown biologic activity, with increased T cell 

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activation and infiltration and decreased MDSCs24-27. A preliminary report from a study of nivo 
+/- tadalafil for neoadjuvant treatment of HNSCC showed that tadalafil enhanced T cell and 
myeloid cell infiltration in 47 randomized patients with a biologic effect in the TME27. 
 
 
 
 
 
 
 
 
  
 
Tadalafil has been evaluated extensively for safety and is currently FDA approved for use in 
erectile dysfunction (ED) and benign prostatic hyperplasia (BPH). The recommended dosing for 
ED/BPH is 5 mg for daily dosing and up to 20 mg on demand dosing for ED based on individual 
efficacy and tolerability. For our study, we have chosen a dose of tadalafil based on data from a 
randomized, prospective, double blinded, placebo controlled, phase II clinical trial in HNSCC 
which demonstrated reversal of immunosuppression at a dose of 20 mg/day24. This included 
reduction of MDSC numbers in tadalafil-treated patients, as well as increase in T-cell activation 
and expansion. Our study will allow dose reductions based on tolerability. 
 
A better understanding of predictors of response as well as the mechanisms of resistance to 
immunotherapy remains a critical unmet need, as it will allow better patient selection and 
identification of additional targets for including in combination therapy approaches. Our 
overarching hypothesis will test whether repolarization or depletion of myeloid and stromal cells 
in the PDA TME, will augment activity of immunotherapy combinations and recruit the highest 
quality T cells to the antitumor response. In this study, we will be combining the CRS-207 vaccine 
with the myeloid-stromal targeting phosphodiesterase 5 inhibitor tadalafil and the immune 
checkpoint inhibitors nivolumab and ipilimumab. 
 

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3. PATIENT SELECTION 
 
3.1 
Inclusion Criteria 
 
3.1.1 Age ≥18 years.   
 
3.1.2 Have histologically- or cytologically-proven ductal adenocarcinoma of the 
pancreas.  Patients with mixed histology (>30% non-adenocarcinoma component) 
will be excluded. 
 
3.1.3 Have metastatic disease. 
 
3.1.4 Have documented radiographic disease progression at the time of study enrollment, 
after previous systemic chemotherapy given in a neoadjuvant, adjuvant, locally 
advanced or metastatic setting.  
 
3.1.5 Presence of at least one lesion with measurable disease as defined by 10 mm in 
longest diameter for a soft tissue lesions or 15 mm in short axis for a lymph node 
by RECIST 1.1. 
 
3.1.6 Patient’s acceptance to have a tumor biopsy of an accessible lesion at baseline and 
on treatment if the lesion can be biopsied with acceptable clinical risk (per Section 
8.1 as judged by the investigator). 
 
 
 
3.1.7 
ECOG performance status 0 or 1 (Appendix A). 
 
 
 
3.1.8 
Adequate organ and marrow function as defined below: 
 
 
Leukocytes 
  
 
 
 
≥ 3,000/mcL 
 
Absolute neutrophil count 
≥ 1,500/mcL 
 
Lymphocyte count 
 
 
≥ 800/mcL 
 
Platelets 
 
  
 
 
 
≥ 100 × 103/uL 
 
Hemoglobin    
 
 
 
≥ 8.0 g/dL 
 
Total bilirubin   
 
 
 
≤ upper limit of normal (ULN) except subjects with 
Gilbert Syndrome, who can have total bilirubin < 3.0 
mg/dL 
 
AST(SGOT) and ALT(SGPT)≤2.0 × ULN 
 
Alkaline phosphatase  
 
≤5.0 × ULN  
 
Creatinine  
  
 
 
 
Creatinine clearance (CrCl) ≥ 50 mL/min (if using 
the modified Cockcroft-Gault formula below): 
 
Female CrCl = (140 - age in years) x weight in kg x 0.85 
                              72 x serum creatinine in mg/dL 
    Male CrCl = (140 - age in years) x weight in kg x 1.00  
                     
 
 
 
 
72 x serum creatinine in mg/dL 
 
 
Albumin  
 
  
 
 
≥3.0 g/dL 

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14
 
3.1.9 Women of childbearing potential (WOCBP) must have a negative serum pregnancy 
test (minimum sensitivity 25 IU/L or equivalent units of human chorionic 
gonadotropin [HCG]).  WOCBP is defined in Section 4.8. Patients with a positive 
HCG due to tumor secretion may be permitted to enroll if lack of pregnancy can be 
documented (e.g. transvaginal ultrasound or serial HCG) and with approval by the 
Principal Investigator. 
 
- 
WOCBP must agree to follow instructions for method(s) of contraception from 
the time of enrollment for the duration of treatment with study drug(s) plus 5 
half-lives of study drug(s) plus 4 weeks (duration of ovulatory cycle) for a total 
of 5 months post treatment completion. 
 
- 
Men who are sexually active with WOCBP must agree to follow instructions 
for method(s) of contraception for the duration of treatment with study drug(s) 
plus 5 half-lives of study drug(s) plus 90 days (duration of sperm turnover) for 
a total of 7 months post-treatment completion. 
 
- 
At least one barrier method of contraception must be employed by all sexually 
active patients (male and female), regardless of other methods, to prevent the 
transfer of body fluids. 
 
3.1.10 Ability to understand and willingness to sign a written informed consent document. 
 
3.2 
Exclusion Criteria 
 
 
 
3.2.1 
Patient has a known history or evidence of brain metastases. 
 
3.2.2  Patient who has had chemotherapy, radiation, or biological cancer therapy within 
14 days prior to the first dose of study drug. 
 
3.2.3 Patient has received an investigational agent or used an investigational device 
within 28 days of the first dose of study drug. 
 
3.2.4  Patient is expected to require any other form of systemic or localized antineoplastic 
therapy while on study. 
 
3.2.5 Patients who have had surgery within 28 days of dosing of investigational agent, 
excluding minor procedures (dental work, skin biopsy, etc.), celiac plexus block, 
and biliary stent placement. 
 
3.2.6 Patients who have received any prophylactic vaccine within 14 days of first dose 
of study drug (7 days for the COVID vaccine) or received a live vaccine within 30 
days of planned start of study therapy. 
 
3.2.7  Have used any systemic steroids within 14 days of study treatment. 
 
3.2.8  Use more than 4 g/day of acetaminophen. 

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15
 
3.2.9 Use of organic nitrates. Note: Medicines called nitrates include nitroglycerin that is 
found in tablets, sprays, ointments, pastes, or patches. Nitrates can also be found in 
other medicines such as isosorbide dinitrate or isosorbide mononitrate. Some 
recreational drugs called “poppers” also contain nitrates, such as amyl nitrite and 
butyl nitrite. 
 
3.2.10 Use of alpha blocking antihypertensives (i.e. prazosin, doxazosin, terazosin).  
 
3.2.11  Use of guanylate cyclase (GC) stimulators such as riociguat 
 
3.2.12  Consumption of substantial amounts of alcohol (≥5 units/day) 
 
3.2.13  Use of strong or moderate cytochrome P450 3A4 (CYP3A4) inhibitor or inducer. 
Potential interactions with patient medications should be checked in a clinical 
database 
such 
as 
EPIC 
or 
Lexicomp 
(https://www.uptodate.com/drug-
interactions/?&redirect=true#di-druglist) 
 
3.2.14 Patients on immunosuppressive agents (e.g., TNF pathway inhibitors, PI3 kinase 
inhibitors) within 7 days of study treatment.  
 
3.2.15 Patient has a known allergy to both penicillin and sulfa. 
 
3.2.16  History of severe hypersensitivity reaction to any monoclonal antibody. 
 
3.2.17  History of severe hypersensitivity to tadalafil or any of the excipients of this product 
 
3.2.18 Have current or prior history of infection or clinically significant adverse events 
(AEs) associated with an exogenous implant(s) or device(s) that has not and cannot 
be easily removed.  
 
3.2.19 Subjects who have implanted medical devices that pose high risks for colonization 
and cannot be easily removed (e.g., artificial heart valves, pacemakers, prosthetic 
joints, orthopedic screw(s), metal plate(s)) if infection occurs. Other common 
devices such as venous access devices (e.g., Port-a-Cath or Mediport) may be 
permitted as well as arterial and venous stents and dental and breast implants. 
 
3.2.20 Evidence of clinical ascites. Trace or small amounts of radiographic ascites may be 
approved by the Principal Investigator. 
 
3.2.21  Have clinically significant and/or malignant pleural effusion (pleural effusions that 
are not clinically significant are allowed, defined as no more than 25% fluid level 
of the corresponding hemithorax). 
 
3.2.22 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. 

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16
 
3.2.23 Subjects with active, known or suspected autoimmune disease. Subjects with 
Graves or Hashimoto’s disease, vitiligo, type I diabetes mellitus, psoriasis not 
requiring systemic treatment, or conditions not expected to recur in the absence of 
an external trigger are permitted to enroll.  
 
3.2.24 Presence of any tissue or organ allograft, regardless of need for 
immunosuppression, including corneal allograft. Instances where loss of the graft 
is not a clinical concern (such as dental bone grafts or skin grafts placed only to 
promote skin growth) can be approved by the Principal Investigator. Patients with 
a history of allogeneic hematopoietic stem cell transplant will be excluded. 
 
3.2.25 All toxicities attributed to prior anti-cancer therapy other than alopecia and fatigue 
must have resolved to grade 1 (National Cancer Institute Common Terminology 
Criteria for Adverse Events [CTCAE], version 5) or baseline before administration 
of study drug. Subjects with toxicities attributed to prior anti-cancer therapy which 
are not expected to resolve and result in long-lasting sequelae, such as neuropathy 
after chemotherapy, are permitted to enroll. 
 
3.2.26 Have received a diagnosis of human immunodeficiency virus (HIV), hepatitis B or 
hepatitis C (patients who are hepatitis C antibody positive may be enrolled if they 
are confirmed with negative viral load at screening) 
 
3.2.27 Patient has a pulse oximetry of <92% on room air. 
 
3.2.28 Patient is on supplemental home oxygen. 
 
3.2.29  Patient has an unhealed surgical wound or ulcer, or a bone fracture considered non-
healing. 
 
3.2.30  Patient has clinically significant heart disease. 
• 
Angina requiring treatment with long-acting nitrates 
• 
Positive cardiac stress test without documented evidence of subsequent, 
effective cardiac intervention 
• 
Uncontrolled arrhythmia 
• 
Within 90 days of start of treatment: 
• Angina requiring treatment with short-acting nitrates  
• Unstable angina 
• Myocardial Infarction 
• Coronary artery bypass graft surgery  
• Percutaneous coronary intervention (for example, angioplasty or stent 
placement)  
• 
Any evidence of heart disease (NYHA≥Class III) within 6 months of planned 
tadalafil administration  
• 
Left ventricular outflow obstructions, such as aortic stenosis and idiopathic 
hypertrophic subaortic stenosis 
 

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17
3.2.31  Patient has valvular heart disease that requires antibiotic prophylaxis for prevention 
of endocarditis. 
 
3.2.32  Prior history of non-arterial ischemic optic retinopathy. 
 
3.2.33 History of significant hypotensive episode requiring hospitalization within 6 
months. 
 
3.2.34 History of hypotension and/or blindness and/or sensorineural hearing loss during 
prior treatment with tadalafil or other PDE-5 inhibitors 
 
3.2.35  History of known hereditary degenerative retinal disorders, including retinitis 
pigmentosa 
 
3.2.36  History of stroke within prior 6 months. 
 
3.2.37  Have insufficient peripheral venous access to permit completion of the study dosing 
and compliance with study phlebotomy regimen 
 
3.2.38 Patient is, at the time of signing informed consent, a regular user (including 
“recreational use”) of any illicit drugs (does not include marijuana or its 
derivatives) or other substance abuse (including alcohol) that could potentially 
interfere with adherence to study procedures or requirements. 
 
3.2.39  Patient is unwilling or unable to follow the study schedule for any reason. 
 
3.2.40 Patient is pregnant or breastfeeding. 
 
3.2.41 Have rapidly progressing disease, as judged by the investigator (e.g., rapid 
progression through prior treatment[s]). 
 
3.3 
Inclusion of Women and Minorities 
 
Both men and women of all races and ethnic groups are eligible for this trial.   
 
4. TREATMENT PLAN 
 
4.1 
Agent Administration 
 
Treatment will be administered on an outpatient basis.  Enrollment for the first three patients will 
be staggered by three weeks to evaluate patients for adverse events before the next patient is 
enrolled (the fourth patient may be enrolled 3 weeks after the third patient). Dosing delays are 
described in Section 5. No investigational or commercial agents or therapies other than those 
described below in Table 2 may be administered with the intent to treat the subject’s malignancy.   
 
 
 
 

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18
 
 
Table 1: Treatment Schedule 
TREATMENT SCHEDULE 
Tadalafil 
Pembrolizumab 
Ipilimumab 
CRS-207 
Days 3-21*  
Cycles 1-6
Day 1,  
Cycles 1-6
Day 1,  
Cycles 1, 3, 5
Day 2, 
Cycles 1-6
* Daily starting on Day 1 after discontinuation of CRS-207 
 
Table 2: Regimen Description 
REGIMEN DESCRIPTION 
Agent 
Premedications; Precautions 
Dose 
Route 
Course Length 
Tadalafil 
None 
20mg daily 
Oral 
 
18 weeks (6 
cycles with 21 
day cycles) 
CRS-207 
650 mg acetaminophen; NS 
pre- and post-infusion to total 
1500ml (suggested: 500ml pre 
and 1000ml post). Subjects 
may also be pre-medicated 
with anti-emetics (suggested:  
IV ondansetron and IV 
fosaprepitant).
1 × 109 CFU 
in 100ml NS  
IV infusion 
over 1 
hour**  
Pembrolizumab No prophylactic 
pre-medication will be given 
unless indicated by previous 
experience in an individual 
subject per Section 4.2.3.
200 mg  
IV infusion 
over 30 min* 
Ipilimumab 
No prophylactic 
pre-medication will be given 
unless indicated by previous 
experience in an individual 
subject per Section 4.2.3.
50 mg 
IV infusion 
over 30 min* 
*Infusion times are approximate (-10/+15 min) and may need to be adjusted based on subject 
tolerability. 
**Infusion times are approximate (± 15 min) and may need to be adjusted based on subject 
tolerability. 
 
Please see Section 5.2 for guidance regarding dosing delays. If the investigator assesses a drug-
related toxicity (that requires discontinuation) to be related to an individual component of the 
treatment schedule, dosing for that study drug alone may be discontinued while dosing is delayed 
until the subject meets criteria to resume treatment of the other study drugs. The relationship to the 
discontinued study drug should be well documented in the source documents and permission from 
the Principal Investigator needs to be obtained prior to continuation with the other study drugs.  
 
As of protocol version 5.0, any patients that remain on study after February 22, 2024 will 

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19
discontinue CRS-207 (no further drug supply) and may remain on study to continue to receive 
pembrolizumab, ipilimumab, and tadalafil. 
 
4.1.1 Tadalafil 
 
Tadalafil is primarily metabolized in the liver. Per the package insert, it is not 
recommended to administer tadalafil to patients with severe hepatic impairment. Therefore, 
Tadalafil administration will begin on Day 3 after review of hepatic laboratories and blood 
pressure for patients receiving CRS-207. Tadalafil administration will begin on Day 1 for 
patients who have discontinued CRS-207. 
 
Subjects will self-administer their dose of tadalafil once a day without regard to food.  The 
dose of tadalafil should be taken as close to the regularly scheduled 24-hour dosing interval 
as possible. If a dose is missed, missed doses may be taken on the same day (but not more 
than one dose per day). If a dose is vomited, it will be skipped.  
 
4.1.2 CRS-207 
 

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20
 
4.1.4  Pembrolizumab 
 
Pembrolizumab is to be administered as a 30 minute IV infusion of 200 mg. It is not to be 
administered as an IV push or bolus injection. At the end of the infusion, flush the line with 
a sufficient quantity of normal saline. Subjects should be observed for a minimum of 30 
minutes before administration of ipilimumab. 
 
Antiemetic medications should not be routinely administered prior to dosing of drugs. See 
Section 4.2.3 for subsequent premedication recommendations following a pembrolizumab-
related infusion reaction.  
 
4.1.5  Ipilimumab 
 
Ipilimumab is to be administered as a 30 minute IV infusion of 50 mg. It is not to be 
administered as an IV push or bolus injection. At the end of the infusion, flush the line with 
a sufficient quantity of normal saline. 
 
Antiemetic medications should not be routinely administered prior to dosing of drugs. See 
Section 4.2.3 for subsequent premedication recommendations following an ipilimumab-
related infusion reaction. 
 
4.2 
General Concomitant Medication and Supportive Care Guidelines 
 
4.2.1 Tadalafil 
 
Acute reactions will be managed using standard therapy for acute drug reactions as per 
institutional guidelines. 
 
4.2.2 CRS-207 
 
Guidance on treatment of the common infusion reactions related to CRS-207 dosing is as 
follows: 
 
• 
Fevers: Despite the acetaminophen premedication, subjects can spike fevers up 
to 40°C starting at the end of the CRS-207 infusion generally through the next 
24 hours. Oral ibuprofen (400 to 800 mg) and acetaminophen (650 to 1000 mg) 
may be used in alternate sequence every 4 hours. 
 
• 
Rigors: Rigors (generally once or twice per infusion) have been observed to 
start during or at the end of a CRS-207 infusion through 24 hours. IV narcotics 
such as morphine or meperidine may be administered per institutional policy. 
Oral morphine or non-steroidal anti-inflammatory drugs (NSAIDs) (e.g., 
aspirin, ibuprofen, naproxen) may be used as home treatment. 
 
• 
Blood pressure: Decreases in blood pressure have been observed necessitating 
additional IV fluids during the 4 hour observation period (up to 1 or 2 liters). 
Reasons for this include the development of fever, compartmental shifts of fluid 

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21
resulting from the CRS-207 infusion and the use of narcotics. Some subjects 
have also been slightly hypotensive at 24 hours upon arrival to the clinic after 
CRS-207 administration. Subjects are encouraged to hydrate themselves 
liberally at home with oral fluids. 
 
• 
Nausea and vomiting: Nausea and vomiting have been reported and observed 
within 24 hours after CRS-207 infusion. Subjects may be given anti-emetics as 
needed. 
 
Blood draws for clinical hematology and serum chemistry will be done the day after the 
CRS-207 infusion. Any unexpected grade 3 or greater laboratory abnormalities should be 
repeated within 24-72 hours. Grade 3 or greater creatinine, AST, ALT, and bilirubin should 
be repeated within 24-72 hours as well. 
 
4.2.3 Pembrolizumab and Ipilimumab 
 
Pembrolizumab and ipilimumab are fully human monoclonal immunoglobulin (Ig) 
G4 antibodies. Subjects should be closely monitored for potential AEs during 
antibody infusion and potential AEs throughout the study.  
 
4.2.3.1 Infusion Reactions 
Since pembrolizumab and ipilimumab contain only human immunoglobulin protein 
sequences, it is unlikely to be immunogenic and induce an infusion or 
hypersensitivity reaction. However, if such a reaction were to occur, it might manifest 
with fever, chills, rigors, headache, rash, pruritis, arthralgias, hypo- or hypertension, 
bronchospasm, or other symptoms. All grade 3 or 4 infusion reactions should be 
reported within 24 hours to the Principal Investigator and reported as an SAE if 
criteria are met. Infusion reactions should be graded according to CTCAE (version 
5) guidelines. Treatment recommendations are provided below and may be modified 
based on local treatment standards and guidelines as appropriate: 
 
For grade 1 symptoms (mild reaction; infusion interruption not indicated; 
intervention not indicated): 
 
Remain at bedside and monitor subject until recovery from symptoms. The following 
prophylactic 
premedications 
are 
recommended 
for 
future 
infusions: 
diphenhydramine 50 mg (or equivalent) and/or acetaminophen 325 to 1000 mg at 
least 30 minutes before additional pembrolizumab or ipilimumab administrations. 
 
For grade 2 symptoms (moderate reaction requires therapy or infusion interruption 
but responds promptly to symptomatic treatment [e.g., antihistamines, non-steroidal 
anti-inflammatory drugs, narcotics, corticosteroids, bronchodilators, IV fluids]; 
prophylactic medications indicated for 24 hours): 
 
Stop the pembrolizumab or ipilimumab infusion, begin an IV infusion of normal 
saline, and treat the subject with diphenhydramine 50 mg IV (or equivalent) and/or 
acetaminophen 325 to 1000 mg; remain at bedside and monitor subject until 

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22
resolution of symptoms. Corticosteroid or bronchodilator therapy may also be 
administered as appropriate. If the infusion is interrupted, restart the infusion at 50% 
of the original infusion rate when symptoms resolve; if no further complications 
ensue after 30 minutes, the rate may be increased to 100% of the original infusion 
rate. Monitor subject closely. If symptoms recur then no further pembrolizumab or 
ipilimumab will be administered at that visit. Administer diphenhydramine 50 mg IV, 
and remain at bedside and monitor the subject until resolution of symptoms. The 
amount of study drug infused must be recorded on the case report form (CRF). The 
following prophylactic premedications are recommended for future infusions: 
diphenhydramine 50 mg (or equivalent) and/or acetaminophen 325 to 1000 mg 
should be administered at least 30 minutes before additional pembrolizumab or 
ipilimumab administrations.  
 
For grade 3 or grade 4 symptoms (severe reaction, grade 3: prolonged [i.e., not rapidly 
responsive to symptomatic medication and/or brief interruption of infusion]; 
recurrence of symptoms following initial improvement; hospitalization indicated for 
other clinical sequelae [e.g., renal impairment, pulmonary infiltrates]; grade 4: (life 
threatening; pressor or ventilator support indicated): 
 
Immediately discontinue infusion of pembrolizumab or ipilimumab. Begin an IV 
infusion of normal saline, and treat the subject as follows. Recommend 
bronchodilators, epinephrine 0.2 to 1 mg of a 1:1,000 solution for subcutaneous 
administration or 0.1 to 0.25 mg of a 1:10,000 solution injected slowly for IV 
administration, and/or diphenhydramine 50 mg IV with methylprednisolone 100 mg 
IV (or equivalent), as needed. Subject should be monitored until the investigator is 
comfortable that the symptoms will not recur. Pembrolizumab and ipilimumab will 
be permanently discontinued. Investigators should follow their institutional 
guidelines for the treatment of anaphylaxis. Remain at bedside and monitor subject 
until recovery from symptoms. In the case of late-occurring hypersensitivity 
symptoms (e.g., appearance of a localized or generalized pruritis within 1 week after 
treatment), symptomatic treatment may be given (e.g., oral antihistamine, or 
corticosteroids). 
 
Please refer to Section 5.2 for guidelines regarding ipilimumab, CRS-207, and 
tadalafil treatment delays following a pembrolizumab or ipilimumab infusion-related 
reaction.  
 
4.2.3.2 Pembrolizumab and Ipilimumab-Related Adverse Events  
 
Blocking PD-1 or CTLA-4 function may permit the emergence of auto-reactive T 
cells and 
resultant 
clinical autoimmunity. Rash/pruritus, diarrhea/colitis, 
pneumonitis, hepatitis, and hypothyroidism were drug-related, presumptive 
autoimmune events noted in previous pembrolizumab and ipilimumab studies.  
 
For the purposes of this study, a pembrolizumab or ipilimumab-related AE is defined 
as an AE of unknown etiology, associated with drug exposure and is consistent with 
an immune phenomenon. Efforts should be made to rule out neoplastic, infectious, 
metabolic, toxin or other etiologic causes. Serological, immunological, and 

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23
histological (biopsy) data should be used to support the diagnosis of an immune-
mediated toxicity. Suspected pembrolizumab or ipilimumab-related AEs must be 
documented on an AE or SAE CRF. Identification and treatment of pembrolizumab 
and ipilimumab-related AEs can be found in the NCCN’s guidelines for the 
management of immunotherapy-related toxicities. Additional guidance can be 
found in the pembrolizumab and ipilimumab package inserts. Antibiotics will also be 
administered to subjects who have not yet received antibiotics for CRS-207 and the 
subject requires steroids for a suspected pembrolizumab or ipilimumab-related AE 
(Section 4.5). 
 
Subjects who experience a grade 2 or higher pembrolizumab or ipilimumab-related 
AE should be discussed with the Principal Investigator and IND sponsor 
immediately.  
 
4.3 
Prohibited and/or Restricted Medications and Devices 
 
The following therapies or devices are not permitted during the treatment period (if administered, 
the subject may be removed from the study): 
 
•  Any 
non-study 
anticancer 
chemotherapy 
or 
immunotherapy 
(approved 
or 
investigational) 
• Any major surgery or surgical procedure; if required must be discussed with the Principal 
Investigator to determine if it is appropriate for the subject to continue study treatment 
•  TNF pathway inhibitors or PI3 kinase inhibitors 
•  Another investigational agent 
•  Live vaccines (examples of live vaccines include, but are not limited to: measles, mumps, 
rubella, chicken pox, yellow fever, rabies, BCG, and typhoid [oral] vaccine). Seasonal 
influenza vaccines for injection are generally killed virus vaccines and are allowed. 
However, intranasal influenza vaccines (e.g. Flu-Mist®) are live attenuated vaccines, and 
are not allowed.  
• Where possible, subjects should not receive any dose of the COVID-19 vaccine within 7 
days of pembrolizumab, ipilimumab, and CRS-207.  
• The use of anticoagulants is known to increase the risk of gastrointestinal hemorrhage. 
Since gastrointestinal hemorrhage is an adverse reaction with pembrolizumab and 
ipilimumab, subjects who require concomitant anticoagulant therapy should be 
monitored closely. 
• Implanted medical devices that pose high risks for colonization and cannot be easily 
removed (e.g., artificial heart valves, pacemakers, prosthetic joints, orthopedic screw(s), 
metal plate(s)) if infection occurs are prohibited. Other common devices such as venous 
access devices (e.g., Port-a-Cath or Mediport), arterial and venous stents, and dental and 
breast implants may be permitted if approved by the Principal Investigator. 
• Palliative (limited-field) radiation therapy is permitted, but only for pain control and with 
approval by the Principal Investigator or IND Sponsor. 
• Systemically active steroids can be used but should be reported to the Principal 
Investigator and/or IND Sponsor. Steroid treatment should be completed at least 14 days 
prior to resuming study-related treatments. Patients requiring adrenal replacement steroid 
doses < 10 mg daily prednisone equivalent (in the absence of active autoimmune disease) 
may resume treatment if approved by the Principal Investigator or the IND Sponsor. 

J2180 / Version 5.0 / January 5, 2024 
24
Patients requiring replacement doses of steroids are required to discontinue treatment 
with CRS-207. 
• If steroids or immunosuppressive agents are required during treatment, prophylactic 
antibiotics will be administered as outlined in Section 4.5. 
• Use of organic nitrates. Note: Medicines called nitrates include nitroglycerin that is found 
in tablets, sprays, ointments, pastes, or patches. Nitrates can also be found in other 
medicines such as isosorbide dinitrate or isosorbide mononitrate. Some recreational 
drugs called “poppers” also contain nitrates, such as amyl nitrite and butyl nitrite. 
• Use of guanylate cyclase (GC) stimulators such as riociguat 
• Substantial amounts of alcohol (≥5 units/day) are not recommended while taking tadalafil 
because tadalafil can potentiate the hypotensive effects 
• Alpha-blocking antihypertensives (such as terazosin, doxazosin mesylate, prazosin, 
alfuzosin) are prohibited. Alpha blockers for the treatment of benign prostatic hyperplasia 
should be used with caution. 
• Use of anti-hypertensives should be used with caution 
• Use of strong or moderate CYP3A4 inducers (rifampicin) are not recommended because 
they can decrease tadalafil exposure 
• Use of strong or moderate CYP3A4 inhibitors are not recommended because they can 
increase tadalafil exposure. Patient medications should be checked in a clinical database 
such 
as 
EPIC 
or 
Lexicomp 
(https://www.uptodate.com/drug-
interactions/?&redirect=true#di-druglist) 
 
In addition, the following therapies should not be administered during the treatment period unless 
medically necessary and approval must be obtained from the Principal Investigator for a subject 
to continue dosing if therapy is given concurrently with study participation: 
 
•  General anesthesia  
•  Aspirin >325 mg/day (chronic daily use of aspirin ≤325 mg/day and heparin flushes for 
central lines are allowed except during CRS-207 infusions through 4 days after each 
CRS-207 infusion.) 
•  More than 4 g/day of acetaminophen 
•  Systemic antibiotics 
 
4.4 
Other Restrictions and Precautions 
 
Palliative (limited-field) radiation therapy is permitted, but only for pain control to sites present at 
baseline and with approval by the Principal Investigator or IND Sponsor. 
 
If subjects receive immunosuppressive medications on or after study, prophylactic antibiotics to 
prevent CRS-207 infection are strongly recommended for the duration of the treatment with the 
immunosuppressant (recommended oral 80 mg trimethoprim / 400 mg sulfamethoxazole once 
daily or 160 mg trimethoprim / 800 mg sulfamethoxazole (DS) three days a week). 
 
4.5 
Antibiotic Administration  
 
4.5.1 Antibiotic Regimen for Semi-Permanent Indwelling Device Placement  
 
An antibiotic regimen should be initiated for any patient that is having a semi-permanent 

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25
indwelling device placed such as biliary stents/drains, vascular devices, and pleuryx catheters 
(excluding patients who discontinued CRS-207). If possible, it is recommended that patients 
receive a dose of Ampicillin/Sulbactam (Unasyn, 3g) with Gentamicin (3 mg/kg) pre-operatively. 
This should be followed by 8-hour intervals of 1g Amoxicillin (160 mg trimethoprim/800 mg 
sulfamethoxazole at 12-hour intervals for allergic patients) for 3 days post-operatively. 
 
4.5.2  EOT: 7 Days after Last CRS-207 Dose in Each Treatment Course, or Suspected 
CRS-207 Infection, or Steroid Administration 
 
An antibiotic course will be initiated for each subject 7 days after the subject’s last dose of CRS-
207 for each treatment course, within 7 days of the decision to discontinue treatment if treatment 
is discontinued midcourse to ensure clearance of CRS-207 before additional courses or subsequent 
cancer-related therapy, or 7 days after the subject’s last dose of CRS-207 if CRS-207 is 
discontinued and the patient remains on study to receive the other study drugs. For subjects who 
do not have a central line, a 7-day course of oral amoxicillin (1G at 8-hour intervals) or 
trimethoprim/sulfamethoxazole in penicillin-allergic subjects (160 mg trimethoprim/800 mg 
sulfamethoxazole at 12-hour intervals) will be initiated for each subject. If the patients do not 
tolerate the 1G dose, the antibiotics can be dose reduced to 500mg.  Subjects with a central line 
will receive 2 doses [2g ampicillin 6 hours apart or 3-5 mg/kg trimethoprim/ sulfamethoxazole 8 
hours apart (in penicillin-allergic subjects)] of IV antibiotics through the port, followed by 6 days 
of oral antibiotics [1G at 8-hour intervals amoxicillin or 160 mg trimethoprim/800 mg 
sulfamethoxazole at 12-hour intervals  (in penicillin-allergic subjects)] started within 6 hours of 
completing the IV ampicillin (or within 8 hours from IV trimethoprim/sulfamethoxazole).  If the 
subject is withdrawn from the study more than 7 days after administration of CRS-207, antibiotics 
will be administered as soon as possible after study withdrawal.    
 
Should a patient require emergent implant of a prohibited device (as described in Section 4.3) 
while on therapy, the patient will receive a 14-day IV antibiotic regimen appropriate for the 
coverage of wild-type listeriosis.  
 
Antibiotics will also be administered to subjects who have not yet received antibiotics for CRS-
207 and the subject requires steroids for a suspected pembrolizumab/ipilimumab-related AE or 
non-related AE. After completion of 7-day course of antibiotics for listeria clearance, antibiotic 
prophylaxis should be given for the duration of the treatment with the steroid (recommended oral 
80 mg trimethoprim / 400 mg sulfamethoxazole once daily or 160 mg trimethoprim / 800 mg 
sulfamethoxazole (DS) three days a week).  
 
During or after study, subjects who are confirmed to have a blood culture positive for CRS-207 
after more than 7 days post-infusion, will receive a minimum 14-day course of IV antibiotics (see 
below).  In addition, IV ampicillin (or trimethoprim/sulfamethoxazole in penicillin-allergic 
subjects) plus gentamicin should be initiated earlier for possible infectious complications of CRS-
207 for subjects who are suspected of having CRS-207 infection and meet the specified criteria 
(see below). 
 
Subjects with clinical or laboratory signs or symptoms of persistent infection who require initiation 
of antibiotics other than specified by protocol should have a clinically-relevant evaluation, 
including appropriate bacterial cultures. Culture of cerebrospinal fluid should be obtained for 
subjects with suspected central nervous system infection. In such instances, analysis of 

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cerebrospinal fluid should also include cell count, protein, glucose and Gram stain. IV ampicillin 
(or trimethoprim/sulfamethoxazole in penicillin-allergic subjects) plus gentamicin should be 
initiated for possible infectious complications of CRS-207 for subjects who are suspected of 
having persistent CRS-207 infection and meet the criteria listed below: 
 
• 
Flu-like symptoms Grade 3 or greater lasting for ≥12 hours 
• 
Fever Grade 4 or higher (>40.0ºC for >24 hours) 
• 
Persistent fever >39ºC lasting for ≥48 hours 
• 
Infection Grade 3 or higher (infection with interventional radiology or operative 
intervention indicated) 
• 
Evidence of abscess 
• 
Clinical signs or symptoms (e.g., neurologic signs or symptoms), which, in the 
judgment of the investigator, necessitate starting antibiotics 
 
The preferred antibiotic regimen if persistent CRS-207 infection is suspected or confirmed is IV 
administration of ampicillin (or trimethoprim/sulfamethoxazole in penicillin-allergic subjects) 
plus gentamicin. For this purpose, initial doses of ampicillin should be approximately 12 g/day 
(divided doses every 3 to 4 hours), with gentamicin 3 mg/kg daily in divided doses every 8 hours 
(with adjustments according to renal function and serum levels of gentamicin). In penicillin-
allergic subjects, initial IV doses of trimethoprim/sulfamethoxazole should be 15 to 20 mg/kg/day 
(based on trimethoprim component) divided four times per day, with gentamicin 3 mg/kg/day in 
divided doses every 8 hours (with adjustments according to renal function and serum levels of 
gentamicin). For those individuals who receive IV antibiotics, the course of therapy is anticipated 
to be greater than or equal to 14 days, depending on clinical course. Antibiotic treatment may be 
completed with use of oral antibiotics, if clinically indicated. 
 
If during the course of treatment the subject develops a reaction to antibiotics such that neither 
penicillin derivatives nor trimethoprim/sulfamethoxazole can be safely administered, alternative 
antibiotics such as tetracyclines should be given in all of the above situations. The choice of 
alternative should be made in discussion with the Principal Investigator, and consultation with 
infectious disease should be considered. 
 
Suspected or confirmed infection with CRS-207 and/or Listeria is considered an adverse event of 
special interest (AESI) and should be reported following SAE reporting procedures (Section 6.1.3) 
irrespective of temporal relationship to study drug administration.  This includes scheduled blood 
cultures during surveillance monitoring that are positive for CRS-207 or if a subject presents with 
symptoms suspicious for a Listeria-like infection and/or is tested positive for Listeria at a local 
hospital/clinic. 
 
4.6 
Definition of an Overdose for this Protocol 
 
Overdose of pembrolizumab or ipilimumab are defined as: 
An overdose is defined as the accidental or intentional administration of any dose of a product that 
is considered both excessive and medically important. All occurrences of overdose must be 
reported as SAEs (see Section 6.5.1 for reporting details). Appropriate supportive treatment should 
be provided if clinically indicated. 
 

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All reports of overdose with and without an AE must be reported within 24 hours to the Principal 
Investigator and IND Sponsor (Dr. Elizabeth Jaffee).  
4.7 
Unacceptable Toxicity 
 
Unacceptable toxicities are defined as any pembrolizumab, ipilimumab, CRS-207, or tadalafil-
related toxicities that require treatment discontinuation per Sections 4.10.2, 4.10.3, or 4.10.4. 
 
Exception: While grade 3 and above pembrolizumab and ipilimumab infusion reactions require 
treatment discontinuation of that drug per Section 4.10.2, they will not be considered unacceptable 
toxicities.  
 
Unexpected Grade 3 or greater laboratory abnormalities should be repeated within 24-72 hours if 
clinically indicated and monitored as necessary to determine if event meets toxicity criteria. Grade 
3 or greater creatinine, AST, ALT, and bilirubin should be repeated within 24-72 hours as well. 
 
If the investigator assesses a drug-related toxicity (that requires discontinuation) to be related to 
an individual component of the treatment schedule, dosing for that study drug alone may be 
discontinued while dosing is delayed until the subject meets criteria to resume treatment of the 
other study drugs. The relationship to the discontinued study drug should be well documented in 
the source documents and permission from the Principal Investigator and co-Principal Investigator 
needs to be obtained prior to continuation with the other study drugs. 
 
Starting from the 6th patient, if more than 33% of patients (that is, 2/6, 3/9, 4/12, 5/15, 6/17) are 
observed to experience an unacceptable toxicity within the first cycle of treatment, enrollment will 
be paused and the safety data will be reviewed by the study team and IND Sponsor before the 
study enrolls additional patients. In addition, enrollment will be paused if a patient’s death is 
assessed as possibly related to any of the study drugs while the data is reviewed by the study team 
and IND sponsor.  
 
4.8 
WOCBP, Contraception, Use in Pregnancy, Use in Nursing 
 
A WOCBP is defined as any female who has experienced menarche and who has not undergone 
surgical sterilization (hysterectomy or bilateral oophorectomy) and is not postmenopausal. 
Menopause is defined clinically as 12 months of amenorrhea in a woman over age 45 years in the 
absence of other biological or physiological causes. In addition, women under the age of 62 years 
must have a documented serum follicle stimulating hormone (FSH) level > 40mIU/mL to confirm 
menopause. 
 
Women treated with hormone replacement therapy (HRT) are likely to have artificially suppressed 
FSH levels and may require a washout period in order to obtain a physiologic FSH level. The 
duration of the washout period is a function of the type of HRT used. The duration of the washout 
period below are suggested guidelines and the investigators should use their judgment in checking 
serum FSH levels. If the serum FSH level is >40 mIU/ml at any time during the washout period, 
the woman can be considered postmenopausal: 
 
 1 week minimum for vaginal hormonal products (rings, creams, gels) 
 4 week minimum for transdermal products 

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 8 week minimum for oral products 
 
4.8.1 Contraception 
 
The investigational agents used in this protocol may have adverse effects on a fetus in 
utero. Furthermore, it is not known if the investigational agents have transient adverse 
effects on the composition of sperm.  Non-pregnant, non-breast-feeding women may be 
enrolled if they are considered highly unlikely to conceive. Highly unlikely to conceive is 
defined as 1) surgically sterilized, or 2) postmenopausal (defined as a woman who is > 45 
years of age and has not had menses for greater than 12 months and women under the age 
of 62 must have a documented serum follicle stimulating hormone (FSH) level less than 
40mlU/mL will be considered postmenopausal). or 3) amenorrheaic for < 2 years without 
a hysterectomy and oophorectomy and with a documented FSH value in the 
postmenopausal range, or ) not heterosexually active for the duration of the study, or 5) 
heterosexually active and willing to use 2 methods of birth control (which is also required 
for the female partners of male subjects). The 2 birth control methods can be 2 barrier 
methods or a barrier method plus a hormonal method to prevent pregnancy, used 
throughout the study starting with Visit 1 through 5 months after the last dose of study 
drug.  Male subjects enrolled in this study must also agree to use an adequate method of 
contraception starting with Visit 1 through 7 months after the last dose of study drug. 
 
Investigators shall counsel WOCBP and male subjects who are sexually active with 
WOCBP on the importance of pregnancy prevention and the implications of an unexpected 
pregnancy Investigators shall advise WOCBP and male subjects who are sexually active 
with WOCBP on the use of highly effective methods of contraception. Highly effective 
methods of contraception have a failure rate of < 1% per year when used consistently and 
correctly. 
HIGHLY EFFECTIVE METHODS OF CONTRACEPTION 
• 
Male condoms with spermicide 
• 
Hormonal methods of contraception including combined oral contraceptive pills, 
vaginal ring, injectables, implants, and intrauterine devices (IUDs) such as Mirena 
by WOCBP subject or male subject’s WOCBP partner. 
• 
Nonhormonal IUDs, such as ParaGard 
• 
Tubal ligation 
• 
Vasectomy 
• 
Complete abstinence* 
 
*Complete abstinence is defined as complete avoidance of heterosexual intercourse and is 
an acceptable form of contraception for all study drugs. Abstinence is only acceptable when 
this is in line with the preferred and usual lifestyle of the subject. Periodic abstinence (e.g., 
calendar, ovulation, symptothermal, profession of abstinence for entry into a clinical trial, 
post-ovulation methods) and withdrawal are not acceptable methods of contraception. 
Subjects who choose complete abstinence are not required to use a second method of 
contraception, but female subjects must continue to have pregnancy tests. Acceptable 
alternate methods of highly effective contraception must be discussed in the event that the 
subject chooses to forego complete abstinence.  
 

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LESS EFFECTIVE METHODS OF CONTRACEPTION 
• 
Diaphragm with spermicide 
• 
Cervical cap with spermicide 
• 
Vaginal sponge 
• 
Male condom without spermicide* 
• 
Progestin only pills by WOCBP subject or male subject’s WOCBP partner 
• 
Female condom* 
*A male and female condom must not be used together 
 
Subjects should be informed that taking the study drug may involve unknown risks to the 
fetus (unborn baby) if pregnancy were to occur during the study.  In order to participate in 
the study they must adhere to the contraception requirement (described above) for the 
duration of the study.  If there is any question that a subject will not reliably comply with 
the requirements for contraception, that subject should not be entered into the study. 
 
4.8.2 Use in Pregnancy 
 
The investigational agents used in this protocol may have adverse effects on a fetus; 
therefore, women with a positive pregnancy test at screening will not be eligible for 
enrollment.  If a subject inadvertently becomes pregnant while on treatment, the subject 
will immediately be removed from the study.  The study team will contact the subject at 
least monthly and document the subject’s status until the pregnancy has been completed or 
terminated.   
 
Pregnancy in female subjects throughout the study or within 5 months of completing 
treatment as well as any pregnancy in partners of male subjects throughout the study or 
within 7 months of completing the study should be reported initially as a serious adverse 
event (see SAE reporting procedures in Section 6.5.1 and 6.5.5) by the investigator within 
24 hours of learning of its occurrence. Pregnancy information must be reported on the 
Pregnancy Form. 
 
Protocol required procedures for study discontinuation and follow-up must be performed 
on the subject unless contraindicated by pregnancy (e.g., x-ray studies). Other appropriate 
pregnancy follow-up procedures should be considered if indicated.  
 
Follow-up information regarding the course of the pregnancy, including any voluntary or 
spontaneous termination, perinatal and neonatal outcome and where applicable, offspring 
information must be reported on the Pregnancy Follow-up Form. Pregnancy outcomes 
must also be collected for the female partners of any males in this trial. Consent to report 
information regarding these pregnancy outcomes should be obtained from the female 
partner.  
 
4.8.3 Use in Nursing Women 
 
Since many drugs are excreted in human milk, and because of the potential for serious 
adverse reactions in the nursing infant, subjects who are breast-feeding are not eligible for 
enrollment. 

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4.8.4 All Subjects (Male and Female) 
 
All sexually active patients must use at least a barrier method (i.e., condom) to prevent 
transmission of body fluids.  
 
4.9 
Duration of Therapy 
 
Subjects will receive treatment every 3 weeks for 6 cycles of treatment within a course (18 weeks 
total). At the investigator’s discretion, subjects who are clinically stable and meet dosing 
requirements (per Section 5.2) at the end of the first course may receive additional courses of their 
assigned treatment for up to a maximum of 2 years. Subjects that begin a new course prior to the 
2 year cut-off may complete that course prior to coming off study. The additional course(s) may 
start as early as 3 weeks (+7 days) from last dose of previous course and all assessments will be 
followed per the study schedule in Section 9, with the first dose of the additional course 
corresponding to Day 1, Cycle 1 of the study schedule. The following assessments are not required 
during additional courses: 
 
• HLA-typing 
• Tumor biopsies 
 
4.10 
Criteria for Removal from Treatment 
 
The reason for study removal and the date the subject was removed will be documented in the 
CRF. A subject will be discontinued from the trial for any of the following reasons: 
• The subject or legal representative (such as a parent or legal guardian) withdraws 
consent for participation in the study. 
A subject must be discontinued from treatment (but may continue to be monitored in the post-
treatment follow-up portion of the trial) for any of the following reasons: 
 
 The subject or legal representative (such as a parent or legal guardian) withdraws 
consent for treatment 
 Intercurrent illness that prevents further administration of treatment 
 Unacceptable toxicities that do not resolve to allow for treatment continuation with 
the remainder of the study regimen (see Section 4.7). If the study treatment has 
provided clinical benefit (as defined in Section 4.10.1), the IND sponsor may 
approve trial continuation for patients experiencing toxicities that are not life 
threatening or of major clinical concern (such as transient hypotension). 
 Disease progression as defined in Section 4.10.1 
 Need for >2 dose delays due to the same drug-related toxicity as per the dose delay 
guidelines (see Section 5.2) 
 If, in the opinion of the Investigator, a change or temporal or permanent 
discontinuation of therapy would be in the best interest of the subject, 
 Noncompliance with trial treatment or procedure requirements, 

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 Subject is lost to follow-up 
 Subject becomes pregnant 
 
4.10.1 Disease Progression 
 
CRS-207, pembrolizumab, and ipilimumab are expected to trigger immune-mediated 
responses, which require activation of the immune system prior to the observation of 
clinical responses. Such immune activation may take weeks to months to be evident. Some 
subjects may have objective volume increase of tumor lesions or other disease parameters 
within weeks following the start of immunotherapy. Such subjects may not have had 
sufficient time to develop the required immune activation or, in some subjects, tumor 
volume or other disease parameter increases may represent infiltration of lymphocytes into 
the original tumor. In conventional studies, such tumor volume or relevant laboratory 
parameter increases during the first 2-4 months of the study would constitute disease 
progression and lead to discontinuation of imaging to detect response, thus disregarding 
the potential for subsequent immune-mediated clinical response. This phenomenon was 
observed in approximately 10% of subjects in the Phase 1 study of nivolumab and has also 
been reported for ipilimumab monotherapy28. 
 
Subjects will be permitted to continue with treatment beyond RECIST 1.1 defined PD as 
long as they meet the following criteria: 
 
Investigator-assessed clinical benefit, and 
 
Subject is tolerating study drug. 
 
The assessment of clinical benefit should take into account whether the subject is clinically 
deteriorating and unlikely to receive further benefit from continued treatment. The 
following criteria need to be taken into consideration: 
 
No decline in ECOG performance status. 
 
Absence of rapid progression of disease or of progressive tumor at critical anatomical 
sites (e.g., cord compression) requiring urgent alternative medical intervention. 
 
All decisions to continue treatment beyond PD must be discussed with the Principal 
Investigator or Co-Principal Investigator and documented in the study records. 
 
Tumor assessments will be made using RECIST 1.1 (Appendix B) and iRECIST 
(Appendix C).  
 
4.10.2 Pembrolizumab and Ipilimumab-Related Adverse Events  
 
Permanent discontinuation of pembrolizumab and ipilimumab should be considered for 
any of the following: 
1. Severe or life-threatening related AEs, including, but not limited to, any of the 
following (the IND Sponsor must be notified in the event of these AEs): 
 Any grade 2 treatment-related uveitis, eye pain, or blurred vision 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 

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 Any grade 3 non-skin, drug-related AE lasting > 7 days, with the following 
exceptions: 
 
 Grade 3 treatment-related uveitis, pneumonitis, bronchospasm, colitis, 
neurologic toxicity, hypersensitivity reaction, or infusion reaction (applies 
to pembrolizumab and ipilimumab only) of any duration requires 
discontinuation 
 Diarrhea, nausea, or vomiting that resolves to < grade 3 within 24 hours of 
intervention 
 Grade 3 fatigue does not require discontinuation 
 Grade 3 treatment-related endocrinopathies adequately controlled with 
only physiologic hormone replacement do not require discontinuation 
 Grade 3 treatment-related laboratory abnormalities do not require treatment 
discontinuation except: 
 
o Grade 3 treatment-related thrombocytopenia > 7 days OR that is 
associated with bleeding requires discontinuation 
 
o Any treatment-related liver function test (LFT) abnormality that 
meets the following criteria require discontinuation: 
 
Total bilirubin > 5 × ULN 
 
Concurrent AST or ALT > 3 × ULN and total bilirubin > 2 
× ULN 
 
 Any grade 4 treatment-related AE or laboratory abnormality, except for the 
following events which do not require discontinuation: 
 
 Grade 4 amylase or lipase abnormalities that are not associated with 
symptoms or clinical manifestations, or radiographic signs of pancreatitis. 
It is recommended to consult with the Principal Investigator for grade 4 
amylase or lipase abnormalities. 
 
 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. 
 
 Transient, self-correcting grade 4 AST or ALT that occur after the CRS-207 
infusion and resolves within 2 weeks.  
 
 Grade 4 lymphopenia and leukopenia. 
 
 Grade 4 treatment-related endocrinopathy adverse events, such as adrenal 
insufficiency, ACTH deficiency, hyper- or hypothyroidism, or glucose 
intolerance, which resolve or are adequately controlled with physiologic 
hormone replacement (corticosteroids, thyroid hormones) or glucose-
controlling agents, respectively, may not require discontinuation after 
discussion with and approval from the Principal Investigator. 

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 Any dosing interruption lasting > 6 weeks with the following exceptions: 
 
 Dosing interruptions to allow for prolonged steroid tapers to manage drug-
related adverse events are allowed. Prior to re-initiating treatment in a 
subject with a dosing interruption lasting > 6 weeks, the Principal 
Investigator must be consulted. Tumor assessments should continue as per 
protocol even if dosing is interrupted. 
 
 Dosing interruptions > 6 weeks that occur for non-drug-related reasons may 
be allowed if approved by the Principal Investigator. Prior to re-initiating 
treatment in a subject with a dosing interruption lasting > 6 weeks, the 
Principal Investigator must be consulted. Tumor assessments should 
continue as per protocol even if dosing is interrupted.  
 
Any AE, laboratory abnormality, or intercurrent illness which, in the judgment of the 
Investigator, presents a substantial clinical risk to the subject with continued 
pembrolizumab or pembrolizumab/ipilimumab dosing.  
 
In order to standardize the management of irAEs for all subjects, treatment management 
algorithms can be found in the NCCN’s guidelines for the management of 
immunotherapy-related toxicities. Additional AE treatment management guidance 
included in the package inserts might be considered for individual cases. 
 
Subjects that are required to stop treatment with pembrolizumab/ipilimumab due to toxicity 
may stay on study and receive tadalafil and CRS-207 (assessment schedule per Section 
9.2) once the pembrolizumab/ipilimumab-related toxicity(s) has resolved to a grade 1. 
 
4.10.2 CRS-207-Related Adverse Events 
 
Permanent discontinuation of CRS-207 should be considered for any of the following: 
 
 Any treatment-related ≥ grade 3 AEs.  Exceptions include: fever, chills, rigors, 
hypertension, hypotension, syncope, or hypoxia occurring within 12 hours of CRS-
207 administration 
 A fever of >40°C that lasts for greater than 24 hours and does not respond to 
antipyretics. 
 Clinically significant hypotension unresponsive to IV fluids (e.g., systolic blood 
pressure [BP] <90 mm Hg or mean arterial pressure <55 mm Hg as measured on two 
separate occasions at least 10 minutes apart). 
 Initiation of antibiotic therapy, coincident with simultaneous isolation of CRS-207 
from a normally sterile body site, other than blood (e.g., cerebrospinal fluid, joint 
fluid). 
 
Subjects that are required to stop treatment with CRS-207 due to toxicity may stay on study 
and receive pembrolizumab, ipilimumab, and tadalafil (assessment schedule per Section 
9.2) once the pembrolizumab/ipilimumab-related toxicity(s) has resolved to a grade 1. 
 

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4.10.4 Tadalafil-Related Adverse Events  
 
Permanent discontinuation of tadalafil should be considered for any of the following: 
 Severe or life-threatening tadalafil-related AEs, including, but not limited to, any of the 
following (the IND Sponsor must be notified in the event of these AEs): 
1. Any grade 2 drug-related retinal vascular disorder, eye pain, or blurred 
vision/visual field defect 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 
2. Any grade 3 or above clinically significant drug-related AEs that recurs after 2 
dose reductions or the toxicity does not resolve to < grade 3 within 24 hours of 
intervention 
Subjects that are required to stop treatment with tadalafil due to toxicity may stay on study 
and receive pembrolizumab, ipilimumab, and CRS-207 (assessment schedule per Section 
9.2) once the pembrolizumab/ipilimumab-related toxicity(s) has resolved to a grade 1. 
 
4.11 
End of Treatment (EOT) Visit 
 
All subjects will return to the study site 28 days (± 7 days) after the final study treatment (i.e., 
completion of the final course or upon early discontinuation) for an EOT evaluation. Procedures 
and assessments performed at these visits and beyond should follow the respective guidelines 
described in Sections 4.12 and 9.0 as appropriate.  
 
If the EOT visit occurs early (e.g., 1 week prior to the expected visit as protocol allows) or if the 
patient cannot return due to disease progression, an assessment for AEs should be made by telephone 
or email on day 28 (± 1 day) after last dose of study drug and documented.  
 
To eliminate any potentially residual CRS-207, subjects will be administered a 7-day course of 
antibiotics per Section 4.5. Blood cultures for surveillance of CRS-207 will also be collected per 
Section 4.13. 
 
4.12 
Duration of Follow-Up 
 
Treated subjects will begin the follow-up period after they complete the EOT visit. Subjects will 
be contacted every three months (+/- 1 month) to monitor overall survival until death, withdrawal 
of consent, or study closure. Information of other cancer therapies after discontinuation from the 
study treatment will be collected.  Subjects will also be contacted at 90 days (+14 day reporting 
window) from the last dose of pembrolizumab/ipilimumab if the subject was still receiving 
pembrolizumab/ipilimumab at the time of treatment discontinuation to monitor drug toxicity. In 
addition, all subjects that received at least one dose of CRS-207 will be monitored for CRS-207 
infection for one year per Section 4.13. 
Subjects who discontinued study treatment without documented disease progression should 
continue to be monitored for disease status by radiologic imaging.  Disease monitoring should 
continue to be assessed approximately every two months per standard of care until: 1) start of a 

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new antineoplastic therapy (information of the new cancer therapy will be collected), 2) disease 
progression, 3) death, 4) withdrawal of consent, or 5) study closure, whichever occurs first. 
 
Subjects who are discontinued from the study treatment due to an unacceptable drug-related AE 
will be monitored for safety until the resolution of the AE to ≤ grade 1 or stabilization or until 
initiation of a new therapy for their cancer, whichever occurs first. 
 
All subjects will be followed after their last dose of study drug for the development of AEs and 
SAEs as described in Section 6.5.1   
 
At the conclusion of the study, all remaining subjects who have received at least one dose of study 
treatment will be offered enrollment in a long-term follow-up study and continue to be evaluated 
for survival. Subjects who are still receiving treatment at the time of study close may complete the 
current treatment course (up to 6 cycles) and EOT visit prior to transitioning to participation in the 
separate long-term follow-up study. 
 
4.13 
Blood Cultures for CRS-207 Surveillance 
 
Patients who stop treatment will have blood samples from their peripheral vein drawn at the EOT 
visit to assess clearance of CRS-207. Patients who discontinue CRS-207 but remain on study to 
receive other agents should have blood cultures drawn within 4 weeks of the last dose of CRS-
207. After the first culture, blood will continue to be collected for CRS-207 culture at 3, 6, 9, and 
12 months (+/- 1 week window for each collection) for up to 1 year to monitor for the presence of 
CRS-207. For subjects with a central line, a blood sample will also be taken through the central 
line at time points indicated for CRS-207 testing.  Subjects with samples positive for the presence 
of Listeria will initiate IV antibiotics per Section 4.5 and be re-tested until negative cultures are 
confirmed.   
 
4.13.1  Confirmed Listeria Infection 
 
In the event a subject has a positive Listeria culture at any time during or after study 
participation (except within 7 days after a CRS-207 infusion), the IND Sponsor should be 
notified within 24 hours of the adverse event of special interest (AESI) per Section 6.1.3.  
 
If Listeria has been confirmed at the clinical site or an external laboratory, all efforts should 
be made to obtain a sample of the bacterial isolate from the original positive culture and 
submit to the IND Sponsor or designee for strain confirmation; records on all samples 
cultured during this period must be obtained and provided to the Sponsor. Refer to the 
Central Laboratory Manual for sample collection and shipping instructions. 
 
4.13.2  Suspected Infection with CRS-207 or Listeria 
 
In the case of a suspected persistent CRS-207 or Listeria infection that has not been 
confirmed by culture, collection of blood (peripheral and port for those with indwelling 
ports), urine and stool samples in duplicate is recommended. One set of samples should be 
cultured locally for Listeria per institutional guidelines. Culture of cerebrospinal fluid 
should be obtained for subjects with suspected central nervous system infection. In such 
instances, analysis of cerebrospinal fluid should also include cell count, protein, glucose, 

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and Gram stain. If samples are positive for Listeria, the IND Sponsor must e notified 
immediately. 
 
5. DOSING DELAYS/DOSE MODIFICATIONS 
 
5.1  
Dose Modifications 
 
Dose reduction or dose increase of CRS-207, pembrolizumab, and ipilimumab will not be 
permitted. 
 
Up to two dose reductions for tadalafil are allowed. Patients who experience grade 3 or 4 tadalafil-
related toxicities that are eligible for retreatment will be dose reduced. If the grade 3 or 4 toxicity 
resolves to grades 0-2, then a dose modification of tadalafil is allowed to 10mg/day dosage. Dose 
reduction for persistent grade <3 toxicity may also be considered at the discretion of the 
investigator. A second dose reduction to 5mg/day is allowed. If three dose reductions are required, 
further treatment with tadalafil will be discontinued. Patients may continue to receive other drugs 
if tadalafil is discontinued. 
 
Patients with a creatinine clearance of 30-50 mL/min will be dose reduced to 10mg/day.  A second 
dose reduction to 5mg/day is allowed. 
 
5.2  
Dosing Delays 
 
Dosing of study therapy will be delayed for the following laboratory criteria: 
 
Day 1 of each cycle: 
 
 AST/ALT >3 × ULN   
 Total bilirubin > 1.5 x ULN or direct bilirubin > 2.0 × ULN for subjects with 
Gilbert’s disease 
 Creatinine clearance (CrCl) < 30 mL/min  
 Hemoglobin < 7.5 g/dL 
 ANC < 1000/uL  
 Platelets < 80 × 103/uL 
 
Day 3 of each cycle (N/A for patients no longer receiving CRS-207): 
 
 AST/ALT >5 × ULN   
 Total bilirubin > 1.5 x ULN or direct bilirubin > 2.0 × ULN for subjects with 
Gilbert’s disease 
 Creatinine clearance (CrCl) < 30 mL/min  
 
If a subject’s Day 3 laboratory results require a dosing delay of tadalafil, tadalafil may be dispensed 
to the patient but the patient will be instructed to delay the start of tadalafil until the laboratory 
values come back within dosing range. 
 
All scheduled cycles within a course are to be given approximately 3 weeks apart. If necessary, a 
scheduled cycle may be delayed for up to 1 week. In this case, subsequent cycles should continue 

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so that a subject can still receive all 6 cycles given that the cycles are a minimum of 3 weeks apart 
and they have not experienced an AE necessitating discontinuation. If delayed more than 1 week, 
the Principal Investigator must be contacted for further instructions on continued treatment. 
Additional delays or modifications to the treatment schedule must be approved by the Principal 
Investigator or IND Sponsor.  
 
If a delay occurs between Day 1 and 2 in a cycle: 
 Pembrolizumab and ipilimumab-related infusion reactions must resolve to baseline prior 
to administration of ipilimumab, tadalafil or CRS-207.  
 If a pembrolizumab-related infusion reaction prevents subsequent infusion of ipilimumab 
on the same day, the dose of ipilimumab should be replaced within 72 hours. In such 
instances, at least 18 days must elapse between the replacement dose of ipilimumab and 
the administration of the next dose of pembrolizumab combined with ipilimumab. If the 
dose of ipilimumab cannot be replaced within 72 hours, resume ipilimumab at the next 
scheduled dose per Section 9. 
 Resume Day 2 treatment schedule (CRS-207) and assessments without repeating Day 1 
study treatments (pembrolizumab/ipilimumab) if the delay is within 72 hours. 
 If the delay is longer than 72 hours, repeat Day 1 and Day 2 (if applicable) study 
treatments/assessments with a minimum of 2 weeks from the previous Day 1 treatment.  
This includes steroid treatment requiring at least a 14 day washout prior to resuming study-
related treatments. 
 
Administration of study drug should be delayed for the following: 
 
 Dosing criteria are not met 
 Any grade ≥ 2 non-skin, treatment-related AE, with the following exceptions: 
o Grade 2 drug-related fatigue or laboratory abnormalities do not require a treatment 
delay 
o Grade 2 hypothyroidism or thyroiditis 
 Any grade >3 skin treatment-related AE  
 Any ≥ grade 3 treatment-related laboratory abnormality, with the following exceptions for 
asymptomatic amylase or lipase: 
o Grade 3 or 4 amylase or lipase abnormalities that are not associated with symptoms 
or clinical manifestations, or radiographic signs of pancreatitis do not require a dose 
delay. It is recommended to consult with the Principal Investigator for grade 3 
amylase or lipase abnormalities. 
o Isolated grade 3 or 4 electrolyte imbalances/abnormalities that are not associated 
with clinical sequelae and are corrected with supplementation/appropriate 
management 
 Any AE, laboratory abnormality, or intercurrent illness which, in the judgment of the 
investigator, warrants delaying the dose of study drug. 
 
Guidance for Investigators is provided in the current pembrolizumab and ipilimumab package 
inserts. Additionally, management algorithms have been developed to assist Investigators with 
select toxicities and can be found in the NCCN’s guidelines for the management of 
immunotherapy-related toxicities. 
 

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Subjects may resume treatment with pembrolizumab and ipilimumab when the treatment-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 adverse event 
 Treatment-related pulmonary toxicity, diarrhea, or colitis must have resolved to baseline 
before treatment is resumed. 
 Treatment-related endocrinopathies adequately controlled with only physiologic hormone 
replacement may resume treatment, which include grade 2 hyperglycemia, hypothyroidism 
and thyroiditis. 
 
6. ADVERSE EVENTS:  LIST AND REPORTING REQUIREMENTS 
 
This study will use the descriptions and grading scales found in the revised CTCAE version 5 for 
AE reporting.   
 
Information about all AEs, whether volunteered by the subject, discovered by investigator 
questioning, or detected through physical examination, laboratory test or other means, will be 
collected, recorded, and followed as appropriate.  
 
6.1 
Definitions 
 
6.1.1 Adverse Event  
 
An AE is defined as any undesirable sign, symptom or medical condition occurring after 
starting the study drug (or therapy) even if the event is not considered to be related to the 
study. An undesirable medical condition can be symptoms (e.g., nausea, chest pain), signs 
(e.g., tachycardia, enlarged liver) or the abnormal results of an investigation (e.g., 
laboratory findings, electrocardiogram). Medical conditions/diseases present before 
starting the study treatment are only considered AEs if they worsen after starting the study 
treatment (any procedures specified in the protocol). New medical conditions / diseases 
occurring before starting the study treatment but after signing the informed consent form 
will not be recorded as AEs. Additionally, expected progression of the disease being 
studied will not be recorded as an adverse event.  
 
Laboratory abnormalities: Laboratory abnormalities present at the screening visit will 
be recorded as pre-treatment signs and symptoms. After study treatment administration, all 
grade 3 and 4 clinical laboratory results that represent an increase in severity from baseline 
will be reported as AEs. A grade 1 or 2 clinical laboratory abnormality should be reported 
as an AE only if it is considered clinically significant by the investigator (induce clinical 
signs or symptoms or require therapy). 
 
6.1.2 Serious Adverse Event  
 
A SAE is an undesirable sign, symptom or medical condition which:  
• 
Results in death  

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• 
Is life threatening (defined as an event in which the subject was at risk of death at 
the time of the event; it does not refer to an event which hypothetically might have 
caused death if it were more severe)  
• 
Requires inpatient hospitalization or causes prolongation of existing hospitalization 
(see note below for exceptions) for >24 hours 
• 
Results in persistent or significant disability/incapacity  
• 
Is a congenital anomaly/birth defect (note: reports of congenital anomalies/birth 
defects must also be reported on the Pregnancy Form)  
• 
Is an important medical event (defined as a medical event(s) that may not be 
immediately life-threatening or result in death or hospitalization but, based upon 
appropriate medical and scientific judgment, may jeopardize the subject or may 
require intervention [e.g., medical, surgical] to prevent one of the other serious 
outcomes listed in the definition above.) Examples of such events include, but are 
not limited to, intensive treatment in an emergency room or at home for allergic 
bronchospasm; blood dyscrasias or convulsions that do not result in 
hospitalization.) 
• 
Potential drug induced liver injury (DILI) is also considered an important medical 
event.  
• 
Hemophagocytic lymphohistiocytosis is also considered an important medical 
event. 
• 
Suspected transmission of an infectious agent (eg, pathogenic or nonpathogenic) 
via the study drug is an SAE. 
• 
Is a new cancer (that is not a condition of the study) 
• 
Is associated with an overdose 
• 
Is a pregnancy or pregnancy outcome of spontaneous abortion, missed abortion, 
benign hydatidiform mole, blighted ovum, fetal death, intrauterine death, 
miscarriage, or stillbirth. 
 
Events not considered to be SAEs are hospitalizations for:  
• 
Admissions as per protocol for a planned medical/surgical procedure or to facilitate 
a procedure 
• 
Routine health assessment requiring admission for baseline/trending of health 
status (e.g., routine colonoscopy)  
• 
Medical/surgical admission for purpose other than remedying ill health state and 
was planned prior to entry into the study. Appropriate documentation is required in 
these cases.  
• 
Admission encountered for another life circumstance that carries no bearing on 
health status and requires no medical/surgical intervention (e.g., lack of housing, 
economic inadequacy, care-giver respite, family circumstances, administrative). 
• 
Admissions for monitoring of treatment-related infusion reactions that do not 
otherwise meet the criteria for a SAE. 
 
6.1.3 Adverse Events of Special Interest (AESI) 
 
Suspected infection with CRS-207 and/or Listeria are considered adverse events of special 
interest (AESI) and should be reported following SAE reporting procedures in Section 6.5 
irrespective of temporal relationship to study drug administration. 
 

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In the event a subject has a positive Listeria culture at any time during or after study 
participation, the event should be reported to the IND Sponsor within 24 hours of the event. 
 
All AESIs must be reported for the duration of the study regardless of causality. 
 
6.2 
Assessment of Causality 
 
The relationship of an AE to the administration of the study drug is to be assessed by the 
investigator according to the following definitions:  
 
 No (unrelated, not related, no relation):  The time course between the administration of 
study drug and the occurrence or worsening of the adverse event rules out a causal 
relationship and another cause (concomitant drugs, therapies, complications, etc.) is 
suspected.  
 
 Yes (related): The time course between the administration of study drug and the occurrence 
or worsening of the adverse event is consistent with a causal relationship and no other cause 
(concomitant drugs, therapies, complications, etc.) can be identified.  
 
The following factors should also be considered:  
 The temporal sequence from study drug administration - The event should occur after the 
study drug is given. The length of time from study drug exposure to event should be 
evaluated in the clinical context of the event.  
 Underlying, concomitant, intercurrent diseases - Each report should be evaluated in the 
context of the natural history and course of the disease being treated and any other disease 
the subject may have.  
 Concomitant medication - The other medications the subject is taking or the treatment the 
subject receives should be examined to determine whether any of them might be recognized 
to cause the event in question.  
 Known response pattern for this class of study drug - Clinical and/or preclinical data may 
indicate whether a particular response is likely to be a class effect.  
 Exposure to physical and/or mental stresses - The exposure to stress might induce adverse 
changes in the recipient and provide a logical and better explanation for the event.  
 The pharmacology and pharmacokinetics of the study drug - The known pharmacologic 
properties (absorption, distribution, metabolism, and excretion) of the study drug should 
be considered.  
 
Assessment of Grade: 
 
The investigator will make an assessment of grade for each AE and SAE reported during the study, 
which will be recorded in the CRF. The assessment will be based on the National Cancer Institute’s 
CTCAE (Version 5) and graded as shown below: 
• 
Grade 1: Mild; asymptomatic or mild symptoms; clinical or diagnostic observations only; 
intervention not indicated 
• 
Grade 2: Moderate; minimal, local or noninvasive intervention indicated; limiting age-
appropriate instrumental activities of daily living 

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• 
Grade 3: Severe or medically significant but not immediately life-threatening; 
hospitalization or prolongation of hospitalization indicated; disabling; limiting self-care 
activities of daily living 
• 
Grade 4: Life-threatening consequences; urgent intervention indicated 
• 
Grade 5: Death related to AE 
 
Any AE that changes in grade during its course will be recorded in the CRF at the highest level 
experience by the subject. 
 
6.3 
Expectedness 
 
Unexpected AE: An AE, which varies in nature, intensity or frequency from information on the 
investigational drug/agent provided in the product IB, package insert or safety reports. Any AE 
that is not included in the IB, package insert, safety reports or informed consent is considered 
“unexpected”.  
 
Expected (known) AE: An AE, which has been reported in the IB, package insert or safety reports. 
An AE is considered “expected”, only if it is included in the IB document as a risk. 
 
6.4 
Handling of Expedited Safety Reports 
 
In accordance with local regulations, the IND Sponsor or designee will notify investigators of all 
SAEs that are unexpected (i.e., not previously described in the IB), and related to tadalafil, CRS-
207, pembrolizumab, or ipilimumab. This notification will be in the form of an expedited safety 
report (ESR) that is to be faxed to the investigators and the study coordinators. Upon receiving 
such notices, the investigator must review and retain the notice with the IB and where required by 
local regulations, the investigator will submit the ESR to the appropriate IRB. The investigator 
and IRB will determine if the informed consent requires revision. The investigator should also 
comply with the IRB procedures for reporting any other safety information.  
 
6.5 
Reporting 
 
6.5.1  Adverse Events and Serious Adverse Events 
 
All AEs (both related and unrelated) will be captured on the appropriate study-specific 
CRFs. All AEs experienced by subjects will be collected and reported from the first dose 
of the investigational agent, throughout the study, and will be followed for 28 days after 
last dose of study drug unless related to the investigational agent.   
 
Subjects who experience a grade 2 or higher pembrolizumab or ipilimumab-related AE 
should be discussed with the Principal Investigator.  
 
Report AEs to the IND Sponsor within 24 hours once identified as an unacceptable 
toxicity (defined in Section 4.7).   
Elizabeth Jaffee:  
 
 
Report all AESI to the IND Sponsor within 24 hours once identified (defined in 
Section 6.1.3): 

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Elizabeth Jaffee:  
 
 
All SAEs (including deaths) occurring from the first dose of the study drug through 90 
days (+ 14 day reporting window) after the last dose of pembrolizumab or ipilimumab or  
within 7 days prior to initiation of a new antineoplastic treatment (whichever comes first) 
will be collected and reported. If the subject never received or is no longer receiving 
pembrolizumab or ipilimumab due to toxicity, all SAEs (including deaths) occurring from 
the first dose of the study drug through 28 days from the last dose of tadalafil CRS-207 
(whichever reporting period is longer).  All SAEs (including deaths) that the investigator 
considers related to study drug occurring after the follow-up periods must be reported. 
 
Subjects who have an ongoing AE/SAE related to the study procedures and/or 
medication(s) may continue to be periodically contacted by a member of the study staff 
until the event is resolved or determined to be irreversible by the investigator. 
 
SAEs will be reported promptly to the IND Sponsor within 24 hours of initial 
notification of the SAE. If this falls on a weekend or holiday, an email notification is 
acceptable but must be followed by an SAE reporting form on the next business day. 
 
SAE reports and any other relevant safety information are to be sent to:  
Elizabeth Jaffee:  
 
 
6.5.2 Follow-up of Adverse Events and Serious Adverse Events 
 
After the initial AE or SAE report, the investigator is required to proactively follow each 
subject and provide further information to the safety department in regards to the subject’s 
condition. 
 
All AE(s) and SAE(s) will be followed until: 
• 
Resolution 
• 
The condition stabilizes 
• 
The event is otherwise explained 
• 
The subject is lost to follow-up 
• 
Death 
 
As soon as relevant information is available, a follow-up SAE report will be submitted to 
the IND Sponsor. 
 
6.5.3 Overdose 
 
An overdose is defined as the accidental or intentional administration of any dose of a 
product that is considered both excessive and medically important. All occurrences of 
overdose must be reported as SAEs. 
 
6.5.4 Potential 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 occurrences of potential 

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DILIs, meeting the defined criteria, must be reported as SAEs under the seriousness 
category checked as ‘other medically important event’. Potential drug induced liver injury 
is defined as: 
 
1) ALT or AST elevation > 3 times upper limit of normal (ULN)  
AND 
2) Total bilirubin > 2 times ULN, without initial findings of cholestasis (elevated serum 
alkaline phosphatase) 
AND 
3) No other immediately apparent possible causes of AST/ALT 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. 
 
6.5.5 Pregnancy Reporting 
 
Although pregnancy and lactation are not always serious by regulatory definition, it is the 
responsibility of investigators or their designees to report any pregnancy or lactation in a 
subject (spontaneously reported to them) that occurs during the trial or within 5 months 
of completing the trial as an SAE. This also includes the pregnancy of a male subject's 
female partner who has provided written consent to provide information regarding 
pregnancy, which occurs during the trial or within 7 months of completing the trial.   
 
If a subject or partner of a subject participating in the study becomes pregnant, the 
investigator must report the pregnancy within 24 hours of discovery or knowledge of the 
event. To report a pregnancy, complete the SAE form with the seriousness category 
checked as ‘other important medical event’. When the form is completed, the IND 
Sponsor will be notified of the event.  
 
All subjects or partners of subjects who become pregnant must be followed to the 
completion/termination of the pregnancy.  If the pregnancy ends for any reason before the 
anticipated date, the investigator should notify the IND Sponsor. At the completion of the 
pregnancy, the investigator will document the outcome of the pregnancy. Pregnancy 
outcomes of spontaneous abortion, missed abortion, benign hydatidiform mole, blighted 
ovum, fetal death, intrauterine death, miscarriage and stillbirth must also be reported as 
SAEs (Important Medical Events).  If the pregnancy continues to term, the outcome 
(health of infant) must also be reported to the IND Sponsor.  
 
6.5.6  Institutional Review Board (IRB) and Institutional Biosafety Committee (IBC) 
 
Serious adverse events will be reported to the IRB and IBC per institutional standards. 
Upon receipt, follow-up information will be given to the IRB and IBC (as soon as relevant 
information is available) per institutional standards. 
 
6.5.7  Food and Drug Administration (FDA) 
 
All reporting to the FDA will be completed by the IND Sponsor. 
 
6.5.7.1 Expedited IND Safety Reports 

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7 Calendar-Day Telephone or Fax Report:   
The IND Sponsor is required to notify the FDA of any fatal or life-threatening 
adverse event that is unexpected and assessed by the investigator to be possibly 
related to the investigational agent.  Such reports are to be telephoned or faxed 
(301-827-9796) to the FDA within 7 calendar days of first learning of the event. 
Follow-up information will be submitted to the FDA as soon as relevant 
information is available.  
 
15 Calendar-Day Written Report:   
The IND Sponsor is required to notify the FDA of any SAE that is unexpected 
and related to the investigational agent in a written IND Safety Report.   
 
Written IND Safety Reports should include an Analysis of Similar Events in 
accordance with regulation 21 CFR § 312.32.  All safety reports previously filed 
with the IND concerning similar events should be analyzed.  The new report 
should contain comments on the significance of the new event in light of the 
previous, similar reports.   
 
Written IND safety reports with Analysis of Similar Events are to be submitted 
to the FDA within 15 calendar days of first learning of the event. Follow-up 
information will be submitted to the FDA as soon as relevant information is 
available. 
 
6.5.7.2 IND Annual Reports 
 
In accordance with the regulation 21 CFR § 312.33, the IND Sponsor shall within 
60 days of the anniversary date that the IND went into effect submit a brief report 
of the adverse events and progress of the investigation.  Please refer to Code of 
Federal Regulations, 21 CFR § 312.33 for a list of the elements required for the 
annual report.  All IND annual reports will be submitted to the FDA by the IND 
Sponsor. 
 
7. PHARMACEUTICAL INFORMATION 
 
7.1  
Tadalafil 
 
7.1.1 Agent Accountability 
 
The IND Sponsor or the IND Sponsor’s representative shall take responsibility for and 
shall take all steps to maintain appropriate records and ensure appropriate supply, storage, 
handling, distribution and usage of investigational product in accordance with the protocol 
and any applicable laws and regulations. 
 
7.1.2 Mode of Action 
 
Tadalafil is a selective inhibitor of cyclic guanosine monophosphate (cGMP)-specific 
phosphodiesterase type 5 (PDE5). PDE5 is found in the smooth muscle of the corpus 

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cavernosum, prostate, and bladder as well as in vascular and visceral smooth muscle, 
skeletal muscle, urethra, platelets, kidney, lung, cerebellum, heart, liver, testis, seminal 
vesicle, and pancreas. In addition to pro-apoptotic tumor effects, PDE5 inhibitors have 
been found to have immunomodulatory properties including decreasing MDSCs and Tregs 
and increasing effector T cells in preclinical models. 
 
7.1.3 Description 
 
Tadalafil is available as almond-shaped tablets. Tablets are available in different sizes and 
different shades of yellow. Each tablet contains 2.5, 5, 10, or 20 mg of tadalafil and the 
following inactive ingredients: croscarmellose sodium, hydroxypropyl cellulose, 
hypromellose, iron oxide, lactose monohydrate, magnesium stearate, microcrystalline 
cellulose, sodium lauryl sulfate, talc, titanium dioxide, and triacetin. 
 
7.1.4 Packaging and Labeling Information 
 
Supplies will be labeled in accordance with regulatory requirements. 
 
7.1.5 Storage 
 
Clinical supplies must be stored in a secure, limited-access location under the storage 
conditions specified on the label. Receipt and dispensing of trial medication must be 
recorded by an authorized person at the trial site. Refer to the package insert for storage 
conditions.  
 
Clinical supplies may not be used for any purpose other than that stated in the protocol.. 
 
7.1.6 Stability 
 
Refer to the package insert for stability information. Follow expiration dates listed on 
clinical supply. 
 
7.1.7 Route of Administration 
 
Oral administration 
 
7.1.8 Subject Care Implications 
 
Tadalafil should be taken at about the same time of day each day. Tadalafil may be taken 
with or without meals. If a dose is missed, missed doses may be taken on the same day (but 
not more than one dose per day). If a dose is vomited, it will be skipped. 
 
Tadalafil should not be taken with increased intake of alcohol (for example, 5 glasses of 
wine) as this may result in increased headaches, dizziness, increased heart rate, and 
hypotension. 
 
7.1.9 Returns and Reconciliation 
 

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The investigator is responsible for keeping accurate records of the clinical supplies and the 
amount remaining at the conclusion of the trial. 
 
Upon completion or termination of the study, all unused and/or partially used 
investigational product will be destroyed at the site per institutional policy. It is the 
Investigator’s responsibility to arrange for disposal of all empty containers, provided that 
procedures for proper disposal have been established according to applicable federal, state, 
local and institutional guidelines and procedures, and provided that appropriate records of 
disposal are kept. 
 
7.2  
CRS-207 
 

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7.3  
Pembrolizumab (KEYTRUDA®) 
 
7.3.1 Agent Accountability 
 
The IND sponsor or the IND Sponsor’s representative shall take responsibility for and shall 
take all steps to maintain appropriate records and ensure appropriate supply, storage, 
handling, distribution and usage of investigational product in accordance with the protocol 
and any applicable laws and regulations.  
7.3.2 Mode of Action 
 
Pembrolizumab is a fully human monoclonal immunoglobulin (Ig) G4 antibody that binds 
to the PD-1 cell surface membrane receptor, a negative regulatory molecule expressed by 
activated T and B lymphocytes. Inhibition of the interaction between PD-1 and its ligands 
(PD-L1 and 2) promotes immune responses and antigen-specific T cell responses to both 
foreign antigens as well as self-antigens. 
 
7.3.3 Description 
 
KEYTRUDA (pembrolizumab) injection is a sterile, preservative-free, clear to slightly 
opalescent, colorless to slightly yellow solution for intravenous use. Each vial contains 100 
mg of pembrolizumab in 4 mL of solution. Each 1 mL of solution contains 25 mg of 
pembrolizumab and is formulated in: L-histidine (1.55 mg), polysorbate 80 (0.2 mg), 
sucrose (70 mg), and Water for Injection, USP. 
 
7.3.4 Packaging and Labeling Information 
 
Supplies will be labeled in accordance with regulatory requirements.  
7.3.5 Preparation 
 
Refer to the package insert for preparation instructions. 
 
7.3.6 Storage 
 
Clinical supplies must be stored in a secure, limited-access location under the storage 
conditions specified on the label. Receipt and dispensing of trial medication must be 
recorded by an authorized person at the trial site. Refer to the package insert for storage 
conditions.  
 
Clinical supplies may not be used for any purpose other than that stated in the protocol. 
 
7.3.7 Stability 
 
Refer to the package insert for stability information. Follow expiration dates listed on 
clinical supply. 
 

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7.3.8 Route of Administration 
 
Pembrolizumab is to be administered as a 30 minute IV infusion line containing a sterile, 
non-pyrogenic, low-protein binding 0.2 micron to 5 micron in-line or add-on filter. At the 
end of the infusion, flush the line with a sufficient quantity of normal saline (approximately 
30-50mL).  
 
7.3.9 Subject Care Implications 
 
The most common side effects include fatigue, musculoskeletal pain, decreased appetite, 
pruritus, diarrhea, nausea, rash, pyrexia, cough, dyspnea, constipation, pain, and abdominal 
pain.  
 
KEYTRUDA is a monoclonal antibody that belongs to a class of drugs that bind to either 
the programmed death-receptor 1 (PD-1) or the PD-ligand 1 (PD-L1), blocking the PD-
1/PD-L1 pathway, thereby removing inhibition of the immune response, potentially 
breaking peripheral tolerance and inducing immune-mediated adverse reactions. Important 
immune-mediated adverse reactions listed under the package insert may not include all 
possible severe and fatal immune-mediated adverse reactions. 
Immune-mediated adverse reactions, which may be severe or fatal, can occur in any organ 
system or tissue and can affect more than one body system simultaneously. Immune-
mediated adverse reactions can occur at any time after starting treatment with a PD-1/PD-
L1 blocking antibody. While immune-mediated adverse reactions usually manifest during 
treatment with PD-1/PD-L1 blocking antibodies, immune-mediated adverse reactions can 
also manifest after discontinuation of PD-1/PD-L1 blocking antibodies. 
 
Early identification and management of immune-mediated adverse reactions are essential 
to ensure safe use of PD-1/PD-L1 blocking antibodies. Monitor patients closely for 
symptoms and signs that may be clinical manifestations of underlying immune-mediated 
adverse reactions. Evaluate liver enzymes, creatinine, and thyroid function at baseline and 
periodically during treatment. In cases of suspected immune-mediated adverse reactions, 
initiate appropriate workup to exclude alternative etiologies, including infection. Institute 
medical management promptly, including specialty consultation as appropriate. 
 
Withhold or permanently discontinue KEYTRUDA depending on severity (NCCN’s 
guidelines for the management of immunotherapy-related toxicities). Additional 
guidance can be found in the package insert. In general, if KEYTRUDA requires 
interruption or discontinuation, administer systemic corticosteroid therapy (1 to 2 
mg/kg/day prednisone or equivalent) until improvement to Grade 1 or less. Upon 
improvement to Grade 1 or less, initiate corticosteroid taper and continue to taper over at 
least 1 month. Consider administration of other systemic immunosuppressants in patients 
whose immune-mediated adverse reactions are not controlled with corticosteroid therapy 
 
7.3.10 Returns and Reconciliation 
 
The investigator is responsible for keeping accurate records of the clinical supplies and the 
amount remaining at the conclusion of the trial. 
 

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Upon completion or termination of the study, all unused and/or partially used 
investigational product will be destroyed at the site per institutional policy. It is the 
Investigator’s responsibility to arrange for disposal of all empty containers, provided that 
procedures for proper disposal have been established according to applicable federal, state, 
local and institutional guidelines and procedures, and provided that appropriate records of 
disposal are kept. 
 
7.4  
Ipilimumab (YERVOY®) 
 
7.4.1 Agent Accountability 
 
The IND Sponsor or the IND Sponsor’s representative shall take responsibility for and 
shall take all steps to maintain appropriate records and ensure appropriate supply, storage, 
handling, distribution and usage of investigational product in accordance with the protocol 
and any applicable laws and regulations. 
 
7.4.2 Mode of Action 
 
CTLA-4 is a negative regulator of T-cell activity. Ipilimumab is a monoclonal antibody 
that binds to CTLA-4 and blocks the interaction of CTLA-4 with its ligands, CD80/CD86. 
Blockade of CTLA-4 has been shown to augment T-cell activation and proliferation, 
including the activation and proliferation of tumor infiltrating T-effector cells. Inhibition 
of CTLA-4 signaling can also reduce T-regulatory cell function, which may contribute to 
a general increase in T cell responsiveness, including the anti-tumor immune response. 
 
7.4.3 Description 
 
YERVOY (ipilimumab) injection, for intravenous use is a sterile, preservative-free, clear 
to slightly opalescent, colorless to pale-yellow solution, which may contain a small amount 
of visible translucent-to-white, amorphous ipilimumab particulates. It is supplied in single-
dose vials of 50 mg/10 mL or 200 mg/40 mL. Each milliliter contains 5 mg of ipilimumab 
and the following inactive ingredients: diethylene triamine pentaacetic acid (DTPA) (0.04 
mg), mannitol (10 mg), polysorbate 80 (vegetable origin) (0.1 mg), sodium chloride (5.85 
mg), tris hydrochloride (3.15 mg), and Water for Injection, USP at a pH of 7. 
 
7.4.4 Packaging and Labeling Information 
 
Supplies will be labeled in accordance with regulatory requirements.  
 
7.4.5 Preparation 
 
Refer to the package insert for preparation instructions. 
 
7.4.6 Storage 
 
Clinical supplies must be stored in a secure, limited-access location under the storage 
conditions specified on the label. Receipt and dispensing of trial medication must be 

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recorded by an authorized person at the trial site. Refer to the package insert for storage 
conditions.  
 
Clinical supplies may not be used for any purpose other than that stated in the protocol. 
 
7.4.7 Stability 
 
Refer to the package insert for stability information. Follow expiration dates listed on 
clinical supply.  
 
7.4.8 Route of Administration 
 
Ipilimumab is to be administered as a 30 minute IV through an intravenous line containing 
a sterile, non-pyrogenic, low-protein-binding in-line filter. At the end of the infusion, flush 
the line with a sufficient quantity of normal saline (approximately 30-50mL). 
 
Pembrolizumab is to be administered as a 30 minute IV infusion line containing a sterile, 
non-pyrogenic, low-protein binding 0.2 micron to 5 micron in-line or add-on filter. At the 
end of the infusion, flush the line with a sufficient quantity of normal saline (approximately 
30-50mL).  
 
7.4.9 Subject Care Implications 
 
Most common adverse reactions with YERVOY are fatigue, diarrhea, pruritus, rash, and 
colitis.  
 
YERVOY is a fully human monoclonal antibody that blocks T-cell inhibitory signals 
induced by the CTLA-4 pathway, thereby removing inhibition of the immune response 
with the potential for induction of immune-mediated adverse reactions. Immune-mediated 
adverse reactions listed herein may not be inclusive of all possible severe and fatal 
immune-mediated reactions. 
 
Immune-mediated adverse reactions, which may be severe or fatal, can occur in any organ 
system or tissue. Immune-mediated adverse reactions can occur at any time after starting 
YERVOY. While immune-mediated adverse reactions usually manifest during treatment, 
immune-mediated adverse reactions can also manifest after discontinuation of YERVOY. 
Early identification and management are essential to ensure safe use of YERVOY. Monitor 
for signs and symptoms that may be clinical manifestations of underlying immune-
mediated adverse reactions. Evaluate clinical chemistries including liver enzymes, 
creatinine, adrenocorticotropic hormone (ACTH) level, and thyroid function at baseline 
and before each dose. Institute medical management promptly, including specialty 
consultation as appropriate. 
 
Withhold or permanently discontinue YERVOY depending on severity (NCCN’s 
guidelines for the management of immunotherapy-related toxicities). Additional 
guidance can be found in the package insert.  In general, if YERVOY requires interruption 
or discontinuation, administer systemic corticosteroid therapy (1 to 2 mg/kg/day 
prednisone or equivalent) until improvement to Grade 1 or less. Upon improvement to 

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53
Grade 1 or less, initiate corticosteroid taper and continue to taper over at least 1 month. 
Consider administration of other systemic immunosuppressants in patients whose immune-
mediated adverse reactions are not controlled with corticosteroid therapy. 
 
7.4.10 Returns and Reconciliation 
 
The investigator is responsible for keeping accurate records of the clinical supplies and the 
amount remaining at the conclusion of the trial. 
 
Upon completion or termination of the study, all unused and/or partially used 
investigational product will be destroyed at the site per institutional policy. It is the 
Investigator’s responsibility to arrange for disposal of all empty containers, provided that 
procedures for proper disposal have been established according to applicable federal, state, 
local and institutional guidelines and procedures, and provided that appropriate records of 
disposal are kept. 
 
8. CORRELATIVE/SPECIAL STUDIES 
 
Research samples will be collected at the discretion of the PI based on availability of supplies and 
safety of patient and staff. Sample collection, processing, storage, and shipment instructions will 
be provided in the Laboratory Manual. 
 
8.1 
Tumor Tissue Studies 
 
Tumor biopsies will be collected (if a subject’s tumor is thought to be reasonably safe and easy to 
biopsy) at baseline and at Cycle 3 (4-6 cores per time point). Lesions will be considered acceptable 
for biopsy if palpable or accessible by imaging (CT or US) guidance when necessary, and if not in 
close proximity to a vital cardiovascular structure. If a biopsy was done within 21 days before first 
dose, archived tissue from this biopsy may be used as baseline sample.  Fine needle aspiration will 
not be acceptable. Additional optional biopsies may be obtained later in the course of study 
treatment.  
 
Attempts will be made to obtain archived tissue samples from all subjects.  Archived FNA biopsy 
samples do not contain sufficient tissue and will not be collected.   
 
Detailed instructions for tissue collection, processing, storage, and shipment will be provided in 
the Laboratory Manual. 
 
To explore the association of OS, PD-L1 positivity, and tumor-infiltrating lymphocyte 
characteristics with clinical responses, archived tumor tissue and tumor tissue obtained at baseline 
and during treatment (Cycle 3) will be compared. PD-L1 expression may predict response to anti-
PD-129,30.  However, PD-L1 is also upregulated in response to IFN-γ released by infiltrating T cells 
and could potentially be a predictor of response to any active immunotherapy.  Pre- and post-
treatment tumor biopsies will also be analyzed for PD-1 expression as well as infiltration of 
immune cells (effector T cells, Tregs, B cells, dendritic cells, etc).  Characterization of immune 
checkpoint expression as well as immune infiltrates may be predictive of response to therapy and 
may also give insight into next generation combinatorial approaches.  Preliminary data from a 
pancreatic cancer immunotherapy study suggests that induction of a Th1and Th17 phenotype at the 

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54
tumor itself predicts response.  Furthermore, upregulation of other inhibitory molecules such as 
IL-10 and TGF-β may identify other targets for combinatorial strategies.  Dependent on 
availability of paired tissue samples, additional analysis including determination of gene signatures 
of tumor and immune cells will be performed to look for patterns of response associated with 
immune activation and changes in the tumor. 
 
Attempts will be made to obtain archived tissue samples from all subjects.   
 
8.2 
Peripheral Blood Mononuclear Cells (PBMCs) 
 
Whole blood for isolation of PBMCs will be collected prior to dosing on Day 1 of Cycles 1, 2, 4, 
and 6 only during Course 1. Pre- and post-treatment changes in PBMCs including effector, helper, 
and regulatory T cells, NK cells, and macrophages through cell phenotyping analysis and gene 
expression profiling will be measured. In addition, induction of Listeria and mesothelin antigen-
specific T cell responses and changes to the T cell epitope repertoire will also be evaluated. 
The cellular immune responses directed against Lm and mesothelin will be evaluated by using 
enzyme-linked immunosorbent spot (ELISPOT) and intracellular cytokine staining. Post-
treatment expression of PD-1 and other lymphocyte activation markers will be measured as well. 
These responses will be correlated with OS. PBMCs are isolated and stored frozen (liquid nitrogen) 
until use. Detailed instructions for collection, processing, storage, and shipment are provided in 
the Laboratory Manual. 
 
8.3 
Serum and Plasma Marker Studies 
 
Sera will be collected prior to dosing on Day 1 of Cycles 1-6 only during Course 1. Additionally, 
subjects will have whole blood for serum drawn 20-26 hours post-CRS-207 infusion only during 
Course 1.  Plasma will be collected prior to dosing on Day 1 of Cycles 1, 2, 4, and 6 only during 
Course 1. Humoral immune responses, including anti-Lm, anti-mesothelin, anti-thyroglobulin and 
anti-galectin 3 antibodies will be evaluated by using enzyme-linked immunosorbent assay 
(ELISA). In addition, potential therapeutic targets, biomarkers, and predictors of response and 
autoimmune toxicity will be evaluated. Sera and plasma are isolated and stored frozen (-80oC) 
until use. Detailed instructions for collection, processing, storage, and shipping are provided in the 
Laboratory Manual. 
 
8.4 
Diagnostic Tissue Samples 
 
Tissue, fluid, or blood may be collected from standard of care procedures used to treat or diagnose 
immune-related toxicities. 
 
8.5 
Genomic Analysis 
 
Genomic sequencing library construction, whole genome/exome sequencing, whole transcriptome 
sequencing, microbial sequencing, neoepitope prediction, mutation burden, and bioinformatic 
analysis will be performed either at an on-campus laboratory or at an off-campus sequencing 
service. All the samples will be de-identified before sending to any laboratory for sequencing.  The 
FASTQ files, BAM files and VCF files will be generated and analyzed.  
 
Genomic sequencing data will be stored and computations conducted using a JH IT managed 

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55
subscription of Azure. 
 
Clinical analysis.  Several CLIA-certified laboratories now offer molecular profiling of 
cancer specimens in commercial and noncommercial settings and provide these results to patients 
and their physicians (e.g. Foundation Medicine, PGDx, Michigan Center for Translational 
Pathology, or JHU CLIA Laboratories). It is possible, therefore, that some of our research analyses 
will be conducted in these CLIA-certified environments.  If tissue or cells are evaluated with next 
generation sequencing strategies to provide a molecular profile of individual cancer specimens in 
a CLIA-certified facility, these results will be made available to the patient and their 
physician.  Patient confidentiality will be maintained, and the patient’s identity will not be publicly 
linked to any study results.  Researchers may use the data set generated in the CLIA assay setting 
to study genetic alterations across a large number of genes important in cancer. Germline mutations 
are only identified in punitive cancer genes. Researchers will use the data set for exploratory 
research to study cancer cell heterogeneity. Some of the sequencing data obtained from the NGS 
strategies will be uploaded to government sponsored databases, such as GEO and dbGAP.  The 
results of the research studies may be published but subjects will not be identified in any 
publication. 
 
If a germline alteration of clinical importance (as judged by the Investigator) to the subject and his 
or her family members is identified by a CLIA-certified test in the course of this analysis, attempts 
will be made in writing to contact the subject and/or family members for genetic counseling 
referral.

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9. 
STUDY CALENDAR 
 
Study Procedures 
Pre-Study 
Cycle 124 
Cycle 2 
Cycle 3 
Cycle 4 
Cycle 5 
Cycle 6 
EOT27
D1 
D229
D329 
D929 
D1
D229
D329 
D929 
D1
D229
D329
D929
D1
D229 
D329
D929
D1
D229
D329
D929
D1
D229
D329
D929 
 
Visit Windows (days)1 
-28 to 
D1
- 
- 
- 
±1 -2/ 
+7 
- 
- 
±1 -2/ 
+7 
- 
- 
±1 -2/ 
+7 
- 
- 
±1 -2/ 
+7 
- 
- 
±1 -2/ 
+7 
- 
- 
±1 
+/- 7 
Tadalafil2 
 
X2 
 
Pembrolizumab3 
 
X 
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
 
Ipilimumab3 
 
X 
 
 
 
 
 
 
 
X
 
 
 
 
 
 
 
X
 
 
 
 
 
 
 
 
CRS-207 
 
 
X 
 
 
 
X 
 
 
 
X 
 
 
 
X 
 
 
 
X 
 
 
 
X 
 
 
 
Hydration 
 
 
X 
X 
 
 
X 
X 
 
 
X 
X 
 
 
X 
X 
 
 
X 
X 
 
 
X 
X 
 
 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Inclusion/ exclusion criteria
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Medical, Cancer, & Con 
Med History4
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Con Meds, Adverse Events 
 
X 
X 
X 
X25 
X
X 
X 
X25
X
X 
X 
X25
X
X 
X 
X25
X
X 
X 
X25
X
X 
X 
X25 
X 
Physical Exam, ECOG  PS5
X 
X 
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X 
Vitals, Weight, & Height6 
X 
X 
X 
X 
 
X
X 
X 
 
X
X 
X 
 
X
X 
X 
 
X
X 
X 
 
X
X 
X 
 
X 
Hematology, Chemistry7, 13 
X 
X 
 
X 
X 
X
 
X 
X 
X
 
X 
X 
X
 
X 
X 
X
 
X 
X 
X
 
X 
X 
X 
Endocrine8, 13 
 
X 
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X 
Urinalysis9 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Virology10 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Coagulation panel11 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Pregnancy Test12, 13 
X 
X26
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
 
CA19-9 13 
X 
X 
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
X
 
 
 
 
ECG14 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Scans, RECIST/iRECIST15 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X
 
 
 
 
 
 
 
 
 
 
 
X 

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57
Study Procedures 
Pre-Study 
Cycle 124 
Cycle 2 
Cycle 3 
Cycle 4 
Cycle 5 
Cycle 6 
EOT27
D1 
D229
D329 
D929 
D1
D229
D329 
D929 
D1
D229
D329
D929
D1
D229 
D329
D929
D1
D229
D329
D929
D1
D229
D329
D929 
 
Whole blood for PBMC16, 20 
 
X 
 
 
 
X
 
 
 
 
 
 
 
X
 
 
 
 
 
 
 
X
 
 
 
X 
Whole blood for plasma16, 20 
 
X 
 
 
 
X
 
 
 
 
 
 
 
X
 
 
 
 
 
 
 
X
 
 
 
X 
Serum16, 20 
 
X 
 
X 
 
X
 
X 
 
X
 
X 
 
X
 
X 
 
X
 
X 
 
X
 
X 
 
X 
HLA17, 20 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Archival Tissue18, 20 
X 
Tumor Biopsies19, 20 
X 
 
 
 
 
 
 
 
 
X
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Antibiotics21,22 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X28 
X28 
Blood sample for CRS-207 
testing23
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
In order to minimize the need for research-only in-person visits, telemedicine visits may be substituted for in person clinical trial visits or portions 
of clinical trial visits where determined to be appropriate and where determined by the investigator not to increase the participants risks. Prior to 
initiating telemedicine for study visits the study team will explain to the participant, what a telemedicine visit entails and confirm that the study 
participant is in agreement and able to proceed with this method. Telemedicine acknowledgement will be obtained in accordance with the Guidance 
for Use of Telemedicine in Research. In the event telemedicine is not deemed feasible, the study visit will proceed as an in-person visit. Telemedicine 
visits will be conducted using HIPAA compliant method approved by the Health System and within licensing restrictions. 
 
1:   If necessary, a scheduled cycle may be delayed for up to 1 week. Longer delays to be approved by the IND Sponsor and/or Principal Investigator. 
2:   Tadalafil administration occurs on Days 3-21 (daily starting on Day 1 for patients no longer receiving CRS-207) after review of hepatic laboratories 
and blood pressure on Day 3 (or Day 1 for patients no longer receiving CRS-207). 
3:  Order of administration is Pembrolizumab followed by Ipilimumab. Subjects should be observed for a minimum of 30 minutes between each infusion. 
4: Cancer history includes: primary site of cancer, gross location of primary tumor, secondary sites of cancer, histology, histologic grade, date of initial 
diagnosis, date of metastatic diagnosis, prior cancer therapy regimens. 
5: Complete physical examination and assessment of ECOG PS will be completed at baseline; focused physical examinations and assessment of ECOG 
PS will be conducted thereafter.  Day 1 Physical examination and ECOG status may be done up to 1 day prior to dosing. 
6:   Blood pressure, pulse, and temperature are required as indicated. Weight and pulse oximetry will be obtained at baseline and prior to each cycle.  Height 
will be taken at or prior to screening only. Only weight and blood pressure are required on Day 3 of each cycle.  Pembrolizumab: vitals will be collected 
prior to the infusion. Ipilimumab: vitals will be collected prior to and at the end of the infusion (-5/+ 15 minutes). CRS-207: vital signs will be obtained 
prior to and then every 30 minutes (± 15 minutes) during infusion and every hour (-5/+ 15 minutes) during post-infusion follow-up. Subjects will be 

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58
observed for at least 4 hours after each CRS-207 infusion. Subjects who are not stable enough to be released at 4 hours after infusion should continue 
to be monitored until stable. Presence of fever alone does not indicate subject is not clinically stable. 
7:  Clinical hematology: CBC with differential ANC, ALC, AEC, and platelet count; serum chemistry: sodium, potassium, chloride, bicarbonate, glucose, 
BUN, creatinine, ALT, AST, alkaline phosphatase, total bilirubin, direct bilirubin, amylase, total protein, albumin, calcium, magnesium, and phosphate. 
Required labs on Day 3 only after CRS-207 dosing: Any unexpected Grade 3 or greater laboratory abnormalities should be repeated within 24-72 hours. 
Grade 3 or greater creatinine, AST, ALT, and bilirubin should be repeated within 24-72 hours as well. 
8:   TSH (Total T3 and free T4 if TSH abnormal and clinically indicated). 
9:   Bilirubin, blood, glucose, ketones, leukocytes, nitrite, pH, color, protein, RBC and WBC count, and specific gravity. 
10: Virology screen: HIV antibody, hepatitis B surface antigen and hepatitis C antibody; additional virology may also be evaluated. Subjects who are 
hepatitis C antibody positive and confirmed negative viral load at screening will be allowed to enroll. 
11:  Coagulation panel: D-dimer, fibrinogen, international normalized ratio of prothrombin time, APTT 
12: Pregnancy tests will be administered to WOCBP: serum pregnancy test is required at screening; urine pregnancy tests are required before doses on Day 
1 of dosing weeks.  
13: Labs may be collected within a window of up to 3 days prior to dosing. Blood draws must not be collected from a central line for at least 4 days after 
infusion of CRS-207.  
14: ECG should be performed at baseline.  
15: Spiral CT of thorax, abdomen and pelvis (and other imaging studies as clinically indicated to evaluate suspected sites of metastatic disease). If a subject 
cannot have a CT scan (e.g., allergy to contrast dye), an MRI should be performed. 3-D CT scans and RECIST reads will not be used to determine 
eligibility at baseline. On study radiologic evaluations and tumor measurements (RECIST and iRECIST per Appendix B and Appendix C) will be 
performed every 10 weeks (± 1 week; starting from the date of first treatment) including the EOT evaluation (± 4 weeks). If the EOT visit occurs early, 
scans do not need to be repeated if one has been done within the past 6 weeks. Weeks are in reference to calendar week and should not be adjusted due 
to dosing delays. 
16: Up to 120 mL of whole blood for PBMC isolation may be drawn up to 72 hours prior to dosing and must be processed by sponsor-qualified operators 
within 6 hours of collection and stored in liquid nitrogen. Approximately 20 mL of whole blood for plasma collection will be drawn as indicated. 
Approximately 5.0 mL of blood for serum for immune monitoring will be drawn as indicated. Day 3 blood draws should be taken between 20 and 26 
hours after start of dosing. Collection of whole blood for isolation of PBMCs, plasma, and serum will only be drawn during Course 1. EOT samples 
are optional (collection at the discretion of the PI). 
17: HLA-typing to include HLA class I type A and B, low resolution. HLA typing is only done during the first course of study treatment. If HLA is missed 
during Cycle 1, it may be collected during subsequent cycles. 
18: Attempts to obtain surgical or biopsy archival tumor samples will be made for every subject until the sample is obtained or documentation that the 
sample cannot be obtained. Detailed instructions for tissue collection, processing and shipment are provided in the Laboratory Manual.  
19: Tumor biopsies to be taken (if a subject’s tumor is thought to be reasonably safe and easy to biopsy) at baseline and at Cycle 3 (4-6 cores per timepoint). 
If a biopsy was done within 21 days before first dose, archived tissue from this biopsy may be used as baseline sample.  The Cycle 3 biopsy has a ± 1 
week window. Additional optional biopsies may be obtained later in the course of study treatment. Fine needle aspiration will not be acceptable. 
Biopsies will only be collected during the first course of study treatment. Detailed instructions for tissue collection, processing and shipment are 
provided in the Laboratory Manual.  
20: Research samples will be collected at the discretion of the PI based on availability of supplies and safety of patient and staff.

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21: A 7-day course of antibiotics will be administered 7 days after final dose of CRS-207 in each course (or after final dose if discontinued early or 7 days 
after the subject’s last dose of CRS-207 if CRS-207 is discontinued and the patient remains on study to receive the other study drugs) per Section 4.5. 
Antibiotic regimens must be completed prior to initiation of any other cancer-related therapy. Antibiotics should also be administered to subjects who 
have not received antibiotics after CRS-207 treatment and the subject is administered steroids for the treatment of suspected pembrolizumab or 
ipilimumab-related AE.   
22: Refer to Section 4.5 for the recommended antibiotic regimen for patients having a semi-permanent indwelling device placed while on study. 
23: Blood for CRS-207 culture will be collected at EOT (or within 4 weeks of the last dose of antibiotics if patients do not complete 6 cycles of treatment 
or if patients discontinue CRS-207 but remain on study to receive other agents) to assess clearance of CRS-207. After the first culture, blood will 
continue to be collected for CRS-207 culture at 3, 6, 9, and 12 months from last dose of CRS-207 (+/- 1 week window for each collection). For subjects 
with a central line, blood samples should be collected from both peripheral and central lines. 
24: Cycle 1 Day 1 evaluations do not need to be repeated if they were conducted within 3 days of the pre-study evaluations. The additional course(s) may 
start as early as 3 weeks (+7 days) from last dose of previous course. 
25: Day 9 (± 1 day) adverse event assessments may be conducted by telephone or email. 
26: Course 2 and beyond only 
27: Subjects will return to the study site for an EOT evaluation. EOT follow-up will occur 28 (±7) days after the final dose. NOTE: CT scan assessment at 
EOT will occur 28 days (± 4 weeks) after the final dose. If the EOT visit occurs early and the patient cannot return due to disease progression, an 
assessment for AEs should be made by telephone or email on day 28 (±1) after last study dose in place of the PE/PS/vitals (clinical laboratory values 
should still be collected). Subjects who discontinue from treatment should be contacted every three months (+/- 1 month) to monitor overall survival. 
Information of other cancer therapies after discontinuation from the study treatment will be collected as well. Subjects should be contacted by telephone 
or email at 90 days (+ 14 day reporting window) to assess for treatment related toxicities that occur in the follow-up period. 
28: Site personnel will contact the subject by telephone (prior to and within 3 days after completion) to facilitate compliance with antibiotic treatment and 
document in source. 
29: Not required if patient discontinues CRS-207

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10.  
STUDY ENDPOINTS 
 
10.1 
Primary Endpoint 
 
The primary endpoint is ORR, which is defined as the proportion of subjects with PR or CR 
according to RECIST. Subjects who discontinue due to toxicity or clinical progression prior to 
post-baseline tumor assessments will be considered as non-responders.  Subjects who discontinue 
for other reasons prior to their first post-baseline tumor assessment will be replaced and not 
included in the primary efficacy analysis. 
 
10.2 
Secondary Endpoint 
 
The secondary endpoint is as follows: 
 Safety assessed by the following measures: 
– Number of patients who have grade 3 or higher drug-related toxicities  
– Frequency of drug-related toxicity by grade  
– Frequency of pembrolizumab-related infusion reactions 
– Frequency of ipilimumab-related infusion reactions 
– Frequency of CRS-207-related infusion reactions 
– Frequency of immune-related AEs 
– Frequency of unacceptable toxicities 
– Frequency of treatment-emergent changes from normal to abnormal values in key 
laboratory parameters:  
o Vital signs: BP, pulse, temperature 
o Physical examination 
o Changes in ECG readings  
o Clinical hematology: complete blood count (CBC) with differential 
ANC, ALC, AEC, and platelet count 
o Clinical serum chemistry: sodium, potassium, chloride, bicarbonate, 
glucose, blood urea nitrogen (BUN), creatinine, ALT, AST, alkaline 
phosphatase, amylase, bilirubin (total), total protein, albumin, calcium, 
magnesium and phosphate 
 
10.3 
Exploratory Endpoints 
 
Exploratory endpoints are as follows: 
 Overall survival (OS), progression-free survival (PFS), disease control rate (DCR), 
duration of response (DOR), and time to objective response (TOR) assessed using 
RECIST 1.1. 
 
OS is defined as the number of months from the date of first dose until death or end 
of follow-up (OS will be censored on the date the subject was last known to be alive 
for subjects without documentation of death at the time of analysis). 
 
PFS is defined as the number of months from the date of first dose to disease 
progression (PD or relapse from CR as assessed using RECIST 1.1 criteria) or death 
due to any cause. PFS will be censored at the date of the last scan for subjects 
without documentation of disease progression at the time of analysis. 
 
DCR is defined as the proportion of subjects with SD, PR, or CR according 
to RECIST 1.1.   

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 
DOR is defined as the number of months from the first documentation of a response 
to date of disease progression.  
 
TOR is defined as the number of months from the date of first dose to the date of 
first response (PR or CR as assessed using RECIST 1.1 criteria).  
 Tumor marker kinetics measured by change in serum CA19-9 concentrations from 
baseline  
 ORR, PFS, DCR, DOR, and TOR assessed using iRECIST 
 Humoral and cellular immune responses directed against Lm and mesothelin assessed 
by using the following measures: 
– ELISPOT or intracellular cytokine staining assays of PBMC 
– Induction of proinflammatory cytokines and chemokines in the serum 
– ELISA detection of mesothelin- and Lm-specific antibodies in the serum 
 Immune subset analyses by IHC and gene expression profiling of tumor tissue  
 Immune subset analyses in PBMCs including effector, helper, and regulatory T cells, 
NK cells, and macrophages  
 Telomere length of lymphocytes  
 Thyroglobulin and galectin-3 antibody responses  
 Peripheral blood specimens, intratumoral core biopsy specimens, and resection 
specimens will be studied using a variety of laboratory techniques including but not 
limited to: immunohistochemistry (IHC), flow cytometry, CITE-Seq, RNA-Seq, whole 
exome sequencing, whole genome sequencing, T cell receptor and B cell receptor 
sequencing, ChIP-seq, ATAC-seq, and MBD-seq. 
 
11.  
DATA REPORTING / REGULATORY REQUIREMENTS 
 
AE guidelines and instructions for AE reporting can be found in Section 6.0 (Adverse Events: 
List and Reporting Requirements). 
 
Dr. Elizabeth Jaffee will be holding the IND for this study.  She will comply with all regulated 
reporting requirements to the FDA. 
 
11.1 
Data Collection and Processing 
 
All information will be collected on study-specific CRFs by study staff. These data will be 
reviewed for completeness and accuracy by the Principal Investigator.  
 
CRFs will be used to capture study results and data. The study coordinator or other authorized 
study personnel will transcribe data from source documents onto paper or eCRFs. Before or 
between visits, the Principal Investigator, IND Sponsor, or designee may request copies of the 
CRFs for preliminary medical review. Once the CRFs are complete and source-verified, the 
investigator must sign and date all required pages, verifying the accuracy of all data contained 
within the CRF.  
 
11.2 
Safety Meetings 
 
Scheduled meetings will take place weekly and will include the principal investigator, study 
coordinator(s), nurse(s), sub-investigators (as appropriate), collaborators (as appropriate), and 
biostatisticians (as appropriate) involved with the conduct of the protocol. During these meetings 

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matters related to the following will be discussed: safety of protocol participants, validity and 
integrity of the data, enrollment rate relative to expectation, characteristics of participants, 
retention of participants, adherence to protocol (potential or real protocol violations), data 
completeness, and progress of data for objectives. 
 
11.3 
Monitoring 
 
The SKCCC Compliance Monitoring Program will provide external monitoring for JHU-affiliated 
sites in accordance with SKCCC DSMP (Version 6.0, 02/21/2019).  The SMC Subcommittee will 
determine the level of patient safety risk and level/frequency of monitoring. The PI shall internally 
monitor the progress of the trial, including review and confirmation of all safety/treatment-related 
outcomes, response assessments, safety reports and/or any related source documentation. The 
protocol will be monitored externally by the SKCCC CRO QA Office. Additional data and safety 
monitoring oversight will also be performed by the SKCCC Safety Monitoring Committee (SMC 
- as defined in the DSMP). 
 
11.4 
Study Documentation 
 
11.4.1 Informed Consent and Authorization for use and Disclosure of Protected 
Health Information 
 
Written informed consent and authorization of use and disclosure of protected health 
information (PHI) must be obtained from each subject (or the subject’s legally authorized 
representative) before performing any study-specific screening/baseline period 
evaluations. The ICF and authorization for use and disclosure of PHI, which is prepared by 
the investigator or the site, must be reviewed and approved by the IND Sponsor, the study 
monitor (if applicable), and the site’s IRB before the initiation of the study.  
 
11.4.2 Investigator Study Files 
 
Documentation about the investigator and study staff, the IRB and the institution, is 
required before study site initiation. A list of required documents will be provided by the 
IND Sponsor or designee to each participating investigator. Copies of these documents as 
well as supplemental information, such as the investigator’s obligations, IB, clinical study 
protocol and amendments, safety information, investigational agent information, biological 
samples and laboratory procedures, SRM, study logs and IND Sponsor/investigator/study 
monitor correspondence will be kept on-site in study site-specific binders. 
 
The IND Sponsor or designee will be responsible for maintaining original and backup of 
all CRF data. The investigator is responsible for maintaining backup of all electronic data 
systems used for primary documentation or source documentation. Backup of electronic 
data will be performed periodically as described in the site-specific SOPs. Backup records 
must be stored at a secure location on site and backup and recovery logs must be maintained 
to facilitate data recovery. If an electronic medical records system that is not supported by 
the IND Sponsor or designee (or is discontinued or decommissioned) is used, the 
investigator must maintain a system to retrieve these records or arrange for the transfer of 
these records to an alternate electronic format or to paper. 
 

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Changes to any electronic records require an audit trail, in accordance with 21 CFR 
11.10(e), and should include who made the changes and when and why the changes were 
made. An audit trail is defined as a secure, computer-generated, time-stamped electronic 
record that will allow reconstruction of the course of events relating to the creation, 
modification and deletion of an electronic record. Audit trails must be created 
incrementally, in chronological order and in a manner that does not allow new audit trail 
information to overwrite existing data. Audit trails should be in a readable format and 
readily available at the study site and any other location where electronic study records are 
maintained. 
 
Audit trails are generated automatically for eCRFs. The investigator is responsible for 
maintaining audit trails of all electronic data systems used for primary documentation or 
source documentation. 
 
11.4.3 Case Report Forms and Source Documentation 
 
The investigator must make study data accessible to the site monitor, to other authorized 
representatives of the IND Sponsor (or designee) and to the appropriate regulatory 
authority inspectors. The original CRF for each subject will be checked against source 
documents at the study site by the site monitor. 
 
11.4.4 Retention of Study Documents 
 
According to ICH E6, Section 4.9, all CRFs, as well as supporting paper and electronic 
documentation and administrative records, must be retained for at least 2 years after the 
last approval of a marketing application and until there are no pending or contemplated 
marketing applications, or at least 2 years have elapsed since the formal discontinuation of 
clinical development of an individual product. Longer retention periods may apply. The 
IND Sponsor will notify investigators as to when documents no longer need to be retained. 
No study documents will be destroyed or moved to a new location without prior written 
approval from the IND Sponsor. If the investigator relocates, retires or withdraws from the 
clinical study for any reason, all records required to be maintained for the study should be 
transferred to an agreed-upon designee, such as another investigator at the institution where 
the study was conducted. 
 
Audit trails for electronic documents must be retained for a period at least as long as that 
required for the subject electronic records to which they pertain. The investigator must 
retain either the original or a certified copy of audit trails. 
 
11.4.5 Data Confidentiality and Subject Anonymity 
 
All information about the nature of the proposed investigation provided by the IND 
Sponsor or their representative to the investigator (with the exception of information 
required by law or regulations to be disclosed to the IRB, the subject or the appropriate 
regulatory authority) must be kept in confidence by the investigator. 
 
The anonymity of participating subjects must be maintained. Subjects will be identified by 
their initials and an assigned subject number on CRFs and other documents retrieved from 

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the site or sent to the IND Sponsor, regulatory agencies, or central laboratories/reviewers. 
Documents that identify the subject (e.g., the signed ICF) must be maintained in strict 
confidence by the investigator, except to the extent necessary to allow auditing by the 
appropriate regulatory authority, the study monitor, IND Sponsor or their representative. 
 
12. STATISTICAL CONSIDERATIONS 
 
12.1 
Study Design/Endpoints 
 
Sample Size 
 
This is an open-label, phase 2 study to evaluate the safety and clinical activity of tadalafil, 
pembrolizumab, ipilimumab, and CRS-207 in subjects with metastatic pancreatic adenocarcinoma 
who have progressed after at least 1 prior chemotherapy regimen.  
 
The primary endpoint of this study is objective response rate (ORR) using RECIST. The treatment 
will be considered inactive and of no interest for further evaluation if the ORR is 5% or less and 
considered active if the ORR is 30% or greater.  
 
A total of 17 evaluable patients will be enrolled for ORR. Patients will be considered evaluable 
for the primary endpoint if they receive at least one dose of study drug. If 3 or more responses are 
observed, we will conclude that the ORR is higher than 5% and that the regimen warrants further 
study. This design has 92% power to reject the null hypothesis of an ORR of 5% in favor of the 
alternative hypothesis of 30%, with one-sided target type 1 error of 0.1 (actual type error is 0.0502).  
Additional patients may be enrolled as needed until at least 10 paired biopsies are obtained, but 
will not be included in the primary analysis of efficacy endpoint. 
 
Statistical Analyses 
 
The primary endpoint is objective response, defined as complete response (CR) or partial response 
(PR) according to RECIST criteria. Objective response rate (ORR) will be estimated as the 
proportion of subjects whose best overall response is either a CR or PR per RECIST with 
corresponding 95% exact CI. The primary population for the analysis is all subjects who receive 
at least one dose of study drug, and have at least one post-baseline tumor assessments or 
discontinue due to toxicity or clinical progression prior to post-baseline tumor assessments. 
Subjects who discontinue due to toxicity or clinical progression prior to post-baseline tumor 
assessments will be considered as non-responders. Subjects who discontinue for other reasons 
prior to their first post-baseline tumor assessments will be replaced and not included in the primary 
efficacy analysis.   
 
The secondary and exploratory endpoints are listed in Section 10.2 and 10.3. 
 
PFS, OS, TOR, and DOR will be summarized using Kaplan-Meier methods. DCR and their 
associated 95% exact CI will be calculated.  
 
All biomarker measures, T-cell response, immune phenotype and inflammatory response 
evaluation will be listed, tabulated, and where appropriate plotted. Tumor tissue IHC, functional 
analysis and genomic analysis will be listed, tabulated and where appropriate plotted. For the 

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continuous data, summary statistics and the corresponding changes (or percent changes) from 
baseline will be provided at each time-point of assessment as well as for the changes from baseline 
to the peak value. Categorical data will be summarized using frequency table. As this is an open-
label study without a control treatment, statistical analyses will be done to aid in the understanding 
of the results. The associations of biomarkers with objective response, or time-to-event endpoints 
may be assessed using logistic regression and Cox regression, respectively.  Correlation of baseline 
expression levels or changes in expression levels with response or time-to-event endpoints will 
identify potential responsive (or resistant) subgroups. 
 
12.2 
Safety Analysis 
 
The safety analysis will be performed in all subjects who receive at least one dose of study 
treatment. AE data will be listed individually and incidence of AEs summarized by system organ 
class and preferred terms within a system organ class.  When calculating the incidence of AEs, 
each AE (based on preferred terminology defined by CTCAE version 5.0) will be counted only 
once for a given subject.  In analyses of grade and causality, if the same AE occurs on multiple 
occasions, the highest grade and strongest relationship to study drug will be assumed.  If 2 or more 
AEs are reported as a unit, the individual terms will be reported as separate experiences.   
 
Changes in vital signs, hematology and clinical chemistry parameters from baseline to the end of 
the study will be examined.  Toxicity will be tabulated by type and grade. Treatment-emergent 
changes from normal to abnormal values in key laboratory parameters will be identified. 
 
Toxicity will be monitored throughout the trial. Starting from the 6th patient, if more than 33% of 
patients (that is, 2/6, 3/9, 4/12, 5/15, 6/17) are observed to experience an unacceptable toxicity 
within the first cycle of treatment, enrollment will be paused and the safety data will be reviewed 
by the study team and IND Sponsor before we enroll additional patients. Complete unacceptable 
toxicity criteria can be found in Section 4.7. In addition, enrollment will be paused if a patient’s 
death is assessed as possibly related to any of the study drugs while the data is reviewed by the 
study team and IND sponsor. 
 
 
 
 

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Francisco, L. M., Sage, P. T. & Sharpe, A. H. The PD-1 pathway in tolerance and 
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Olson, D. J. et al. Pembrolizumab Plus Ipilimumab Following Anti-PD-1/L1 Failure in 
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Gubens, M. A. et al. Pembrolizumab in combination with ipilimumab as second-line or 
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APPENDIX A: Performance Status Criteria 
 
ECOG Performance Status Scale 
Karnofsky Performance Scale 
Grade
Descriptions
Percent
Description
0 
Normal activity.  Fully active, able 
to carry on all pre-disease 
performance without restriction. 
100 
Normal, no complaints, no evidence 
of disease.
90 
Able to carry on normal activity; 
minor signs or symptoms of disease.
1 
Symptoms, but ambulatory.  
Restricted in physically strenuous 
activity, but ambulatory and able 
to carry out work of a light or 
sedentary nature (e.g., light 
housework, office work). 
80 
Normal activity with effort; some 
signs or symptoms of disease. 
70 
Cares for self, unable to carry on 
normal activity or to do active work. 
2 
In bed <50% of the time.  
Ambulatory and capable of all 
self-care, but unable to carry out 
any work activities.  Up and about 
more than 50% of waking hours. 
60 
Requires occasional assistance, but 
is able to care for most of his/her 
needs.
50 
Requires considerable assistance and 
frequent medical care.
3 
In bed >50% of the time.  Capable 
of only limited self-care, confined 
to bed or chair more than 50% of 
waking hours. 
40 
Disabled, requires special care and 
assistance.
30 
Severely disabled, hospitalization 
indicated.  Death not imminent.
4 
100% bedridden.  Completely 
disabled.  Cannot carry on any 
self-care.  Totally confined to bed 
or chair. 
20 
Very sick, hospitalization indicated. 
Death not imminent.
10 
Moribund, fatal processes 
progressing rapidly.
5
Dead.
0
Dead.
 
 
 
 
 
 
 
 
 
 
 
 

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APPENDIX B: Response Evaluation Criteria in Solid Tumors (RECIST) 1.1 Criteria for 
Evaluating Response in Solid Tumors 
 
RECIST version 1.1 will be used in this study for assessment of tumor response.  While either CT 
or MRI may be used utilized, as per RECIST 1.1, CT is the preferred imaging technique in this 
study.   
Disease Parameters 
 
Measurable disease:  Measurable lesions are defined as those that can be accurately measured in 
at least one dimension (longest diameter to be recorded) as >20 mm by chest x-ray, as >10 mm 
with CT scan, or >10 mm with calipers by clinical exam.  All tumor measurements must be 
recorded in millimeters (or decimal fractions of centimeters). 
 
Note:  Tumor lesions that are situated in a previously irradiated area might or might not be 
considered measurable unless there is evidence of progression in the irradiated site. Malignant 
lymph nodes.  To be considered pathologically enlarged and measurable, a lymph node must 
be >15 mm in short axis when assessed by CT scan (CT scan slice thickness recommended to 
be no greater than 5 mm).  At baseline and in follow-up, only the short axis will be measured 
and followed. 
 
Non-measurable disease:  All other lesions (or sites of disease), including small lesions (longest 
diameter <10 mm or pathological lymph nodes with ≥10 to <15 mm short axis), are considered 
non-measurable disease.  Bone lesions, leptomeningeal disease, ascites, pleural/pericardial 
effusions, lymphangitis cutis/pulmonitis, inflammatory breast disease, and abdominal masses (not 
followed by CT or MRI), are considered as non-measurable. 
 
Note:  Cystic lesions that meet the criteria for radiographically defined simple cysts should not 
be considered as malignant lesions (neither measurable nor non-measurable) since they are, by 
definition, simple cysts. 
 
‘Cystic lesions’ thought to represent cystic metastases can be considered as measurable lesions, 
if they meet the definition of measurability described above. However, if non-cystic lesions 
are present in the same subject, 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 

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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.  
 
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 diameters. 
 
Progressive Disease (PD): At least a 20% increase in the sum of the diameters of target lesions, 
taking as reference the smallest sum on study (this includes the baseline sum if that is the smallest 
on study).  In addition to the relative increase of 20%, the sum must also demonstrate an absolute 
increase of at least 5 mm.  (Note:  the appearance of one or more new lesions is also considered 
progressions). 
 
Stable Disease (SD): Neither sufficient shrinkage to qualify for PR nor sufficient increase to 
qualify for PD, taking as reference the smallest sum diameters while on study. 
 
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 subject 
to be considered in complete clinical response. 
 
Non-CR/Non-PD: Persistence of one or more non-target lesion(s) and/or maintenance of tumor 
marker level above the normal limits. 
 
Progressive Disease (PD): Appearance of one or more new lesions and/or unequivocal progression 
of existing non-target lesions.  Unequivocal progression should not normally trump target lesion 
status.  It must be representative of overall disease status change, not a single lesion increase.     
 
Although a clear progression of “non-target” lesions only is exceptional, the opinion of the treating 
physician should prevail in such circumstances, and the progression status should be confirmed at 
a later time by the review panel (or Principal Investigator). 
 
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 subject's best response assignment will depend on the 
achievement of both measurement and confirmation criteria. 
 
For Subjects with Measurable Disease (i.e., Target Disease) 

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Target 
Lesions 
Non-Target 
Lesions 
New 
Lesions 
Overall 
Response
Best Overall 
Response when 
Confirmation is 
Required* 
CR 
CR 
No 
CR 
>4 wks. 
Confirmation**
CR 
Non-CR/Non-
PD
No 
PR 
>4 wks. 
Confirmation** 
CR
Not evaluated
No
PR
PR 
Non-CR/Non-
PD/not 
evaluated
No 
PR 
SD 
Non-CR/Non-
PD/not 
evaluated
No 
SD 
Documented at least 
once >4 wks. from 
baseline**
PD 
Any 
Yes or 
No
PD 
no prior SD, PR or CR
Any 
PD*** 
Yes or 
No
PD 
Any
Any
Yes
PD
* 
See RECIST 1.1 manuscript for further details on what is evidence of 
a new lesion. 
** 
Only for non-randomized trials with response as primary endpoint. 
*** 
In exceptional circumstances, unequivocal progression in non-target 
lesions may be accepted as disease progression. 
 
Note: Subjects 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.
 
Reference 
 
E.A. Eisenhauer, P. Therasse, J. Bogaerts, L.H. Schwartz, D. Sargent, R. Ford, J. Dancey, S. 
Arbuck, S. Gwyther, M. Mooney, L. Rubinstein, L. Shankar, L. Dodd, R. Kaplan, D. Lacombe, J. 
Verweij. New response evaluation criteria in solid tumours: Revised RECIST guideline (version 
1.1).  Eur J Cancer. 2009 Jan;45(2):228-47. 
 
 
 

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APPENDIX C: Description of the iRECIST Process For Assessment of Disease Progression  
 
Assessment at Screening and Prior to RECIST 1.1 Progression 
 
Until radiographic progression based on RECIST 1.1, there is no distinct iRECIST assessment. 
 
Assessment and Decision at RECIST 1.1 Progression 
 
In participants who show evidence of radiological PD by RECIST 1.1 the Investigator will decide 
whether to continue a participant on study treatment until repeat imaging is obtained using the 
criteria outlined in Section 4.10.1. 
 
If the Investigator decides to continue treatment, the participant may continue to receive study 
treatment and the tumor assessment should be repeated 4 to 8 weeks later to confirm PD by 
iRECIST, per Investigator assessment. 
 
Tumor flare may manifest as any factor causing radiographic progression per RECIST 1.1, 
including: 
 Increase in the sum of diameters of target lesion(s) identified at baseline to ≥ 20% and 
≥ 5 mm from nadir 
o Please note: the iRECIST publication uses the terminology “sum of measurements”, 
but “sum of diameters” will be used in this protocol, consistent with the original 
RECIST 1.1 terminology. 
 Unequivocal progression of non-target lesion(s) identified at baseline 
 Development of new lesion(s) 
iRECIST defines new response categories, including iUPD (unconfirmed progressive disease) and 
iCPD (confirmed progressive disease).  For purposes of iRECIST assessment, the first visit 
showing progression according to RECIST 1.1 will be assigned a visit (overall) response of iUPD, 
regardless of which factors caused the progression. 
At this visit, target and non-target lesions identified at baseline by RECIST 1.1 will be assessed as 
usual. 
New lesions will be classified as measurable or non-measurable, using the same size thresholds 
and rules as for baseline lesion assessment in RECIST 1.1.  From measurable new lesions, up to 5 
lesions total (up to 2 per organ), may be selected as New Lesions – Target.  The sum of diameters 
of these lesions will be calculated, and kept distinct from the sum of diameters for target lesions at 
baseline.  All other new lesions will be followed qualitatively as New Lesions – Non-target. 
Assessment at the Confirmatory Imaging 
On the confirmatory imaging, the participant will be classified as progression confirmed (with an 
overall response of iCPD), or as showing persistent unconfirmed progression (with an overall 
response of iUPD), or as showing disease stability or response (iSD/iPR/iCR). 
Confirmation of Progression 

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Progression is considered confirmed, and the overall response will be iCPD, if ANY of the 
following occurs: 
 Any of the factors that were the basis for the initial iUPD show worsening 
o For target lesions, worsening is a further increase in the sum of diameters of ≥ 5 mm, 
compared to any prior iUPD time point 
o For non-target lesions, worsening is any significant growth in lesions overall, compared 
to a prior iUPD time point; this does not have to meet the “unequivocal” standard of 
RECIST 1.1 
o For new lesions, worsening is any of these: 
 An increase in the new lesion sum of diameters by ≥ 5 mm from a prior iUPD time 
point  
 Visible growth of new non-target lesions  
 The appearance of additional new lesions 
 Any new factor appears that would have triggered PD by RECIST 1.1 
Persistent iUPD 
Progression is considered not confirmed, and the overall response remains iUPD, if: 
 None of the progression-confirming factors identified above occurs AND 
 The target lesion sum of diameters (initial target lesions) remains above the initial PD 
threshold (by RECIST 1.1) 
Additional imaging for confirmation should be scheduled 4 to 8 weeks from the scan on which 
iUPD is seen.  This may correspond to the next visit in the original visit schedule.  The assessment 
of the subsequent confirmation scan proceeds in an identical manner, with possible outcomes of 
iCPD, iUPD, and iSD/iPR/iCR. 
Resolution of iUPD 
Progression is considered not confirmed, and the overall response becomes iSD/iPR/iCR, if: 
 None of the progression-confirming factors identified above occurs, AND 
 The target lesion sum of diameters (initial target lesions) is not above the initial PD 
threshold.  
The response is classified as iSD or iPR (depending on the sum of diameters of the target lesions), 
or iCR if all lesions resolve. 
In this case, the initial iUPD is considered to be pseudo-progression, and the level of suspicion for 
progression is “reset”.  This means that the next visit that shows radiographic progression, 
whenever it occurs, is again classified as iUPD by iRECIST, and the confirmation process is 
repeated before a response of iCPD can be assigned.  

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Detection of Progression at Visits After Pseudo-progression Resolves 
After resolution of pseudo-progression (i.e., achievement of iSD/iPR/iCR), iUPD is indicated by 
any of the following events: 
 Target lesions 
o Sum of diameters reaches the PD threshold (≥ 20% and ≥ 5 mm increase from nadir) 
either for the first time, or after resolution of previous pseudo-progression.  The nadir 
is always the smallest sum of diameters seen during the entire trial, either before or 
after an instance of pseudo-progression. 
 Non-target lesions 
o If non-target lesions have never shown unequivocal progression, their doing so for the 
first time results in iUPD. 
o If non-target lesions had shown previous unequivocal progression, and this progression 
has not resolved, iUPD results from any significant further growth of non-target 
lesions, taken as a whole. 
 New lesions 
o New lesions appear for the first time 
o Additional new lesions appear 
o Previously identified new target lesions show an increase of ≥ 5 mm in the new lesion 
sum of diameters, from the nadir value of that sum 
o Previously identified non-target lesions show any significant growth 
If any of the events above occur, the overall response for that visit is iUPD, and the iUPD 
evaluation process (see Assessment at the Confirmatory Imaging above) is repeated.  Progression 
must be confirmed before iCPD can occur. 
The decision process is identical to the iUPD confirmation process for the initial PD, except in one 
respect. If new lesions occurred at a prior instance of iUPD, and at the confirmatory scan the 
burden of new lesions has increased from its smallest value (for new target lesions, their sum of 
diameters is ≥5 mm increased from its nadir), then iUPD cannot resolve to iSD or iPR. It will 
remain iUPD until either a decrease in the new lesion burden allows resolution to iSD or iPR, or 
until a confirmatory factor causes iCPD. 
Additional details about iRECIST are provided in the iRECIST publication 31. 
 
 
 
 
 
 
 

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Table 2: Comparison between RECIST 1.1 and iRECIST 
 
RECIST 1.1 
iRECIST 
Definitions of disease: 
numbers, sites and 
target or non 
Measurable are diameters greater 
than 10 mm (15 for nodes 
maximum of 5 (2 per organ)
No change from RECIST 1.1 
CR, PR or SD 
Cannot have met criteria for 
progression 
Can have had iUPD (more than 
once) but not iCPD before iCR, 
iPR or iSD
Confirmation of CR or 
PR 
Only in non-randomized studies 
As per RECIST 1.1 
Confirmation of SD 
Not required 
As per RECIST 1.1 
New lesions 
Progression: recorded but not 
measured 
iUPD but only becomes iCPD if 
on the next scan there are new 
lesions or the size increases by 
greater than 5 mm
Confirmation of 
progression 
Not required 
Required 
Consideration of 
clinical status 
Not required 
Clinical stability considered at 
iUPD to decide treatment 
continuation
 
Table 3:    Trajectory of progression in iRECIST 
 
Target Lesions: iCR, Non-
target: iCR, no new lesions 
iCR 
iCR 
Target lesions: iCR, Non-
target: non iCR/non iUPD, no 
new lesions 
iPR 
iPR 
Target Lesions: iPR, Non-
target: non iCR/non iUPD, no 
new lesions 
iPR 
iPR 
Target lesions: iSD, Non-
target: non iCR/non iUPD, no 
new lesions 
iSD 
iSD 
Target lesions: iUPD with no 
change or with a decrease 
from the last time point, Non-
target: iUPD with no change 
or decrease from last time 
point, new lesions 
NA 
 
 
New lesions confirm iCPD if 
new lesions previously 
identified and increased in 
size (≥ 5 mm in sum of 
measures for new lesions or 
any increase for new lesion 
non-target) or increase in 
number. If no change is seen 
in new lesions assignment 
remains iUPD 
Target lesions: iSD, iPR, 
iCR, non-target: iUPD, no 
iUPD 
Remains iUPD unless iCPD 
is confirmed by increase in 

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new lesions 
the size of non-targets (does 
not need to meet RECIST 1.1 
criteria) 
Target lesions: iUPD, non-
target: non iCR/non iUPD, no 
new lesions 
iUPD 
Remains iUPD unless iCPD 
confirmed on the basis of 
further increase ≥ 5 mm; 
otherwise stays as iUPD 
Target lesions: iUPD, non-
target: iUPD, no new lesions 
iUPD 
Remains iUPD unless iCPD 
confirmed on previously 
identified targets iUPD ≥ 5 
mm or non-target iUPD 
Target lesions: iUPD, non-
targets: iUPD, new lesions 
iUPD 
Remains iUPD unless iCPD 
confirmed by increase of ≥ 5 
mm previously identified 
target, or non-target or an 
increase in size or number of 
new lesions
Target lesions non iUPD or 
progression, non-targets: non 
iUPD or progression, new 
lesions
iUPD 
Remains iUPD unless iCPD 
confirmed by increase in size 
or number of new lesions 
previously identified.
Target lesions, non-target lesions and new lesions are defined according to RECIST 1.1 criteria: if 
no pseudoprogression occurs, RECIST 1.1 and iRECIST categories for CR, PR and SD are the 
same. * Previously identified in the assessment prior to this time point. ‘I’ indicates immune 
responses assigned using iRECIST