untitled h1
untitled h2

Abbreviated Title: Perioperative MVT-5873 CC Protocol \#: 19C0039 E
Version Date: 08/10/2020 NCT Number \#: NCT03801915 Title: Perioperative MVT-5873, a Fully Human Monoclonal Antibody Against a CA 19-9 Epitope, for Operable CA 19-9 Producing Pancreatic Cancers, Cholangiocarcinomas, and Metastatic Colorectal Cancers NCI Principal Investigator: Jonathan M. Hernandez, MD
Surgical Oncology Program (SOP), CCR, NCI Building 10, Room 4W-3740 10 Center Drive Bethesda, MD 20892 Phone: 240-760-6072 E-mail: jonathan.hernandez@nih.gov
untitled h1
investigational agents

| Drug Name:    | HuMab-5B1 (MVT-5873)           |
|---------------|--------------------------------|
| IND Number:   | 144112                         |
| Sponsor:      | NCI Center for Cancer Research |
| Manufacturer: | AAIPharma                      |  
Commercial Agents: None 1
untitled h1
precis

Background:
- Resections to remove tumors in the liver, bile ducts and pancreas are rarely curative, and patients frequently succumb to disease recurrence in the ensuing months to year(s) after the operation.  
- Standard adjuvant therapies, which typically begin 6-12 weeks after surgery, offer little demonstrable decreases in the rates of tumor recurrence.  
- The concept and implementation of immediate perioperative therapy has not been evaluated given the serious concerns related to healing and recovery with standard cytotoxic chemotherapy and newer targeted agents.  
- A significant percentage of metastatic colorectal cancers, and primary tumors of the pancreas and bile ducts express Sialyl Lewisa, an epitope on the well-established tumor marker, CA 19-9.  
- MVT-5873, a fully human antibody against Sialyl Lewisa, has displayed ADCC and CDC
in vitro, potentiated chemotherapeutic efficacy in mouse models and demonstrated efficacy in Phase 1 trials of patients with advanced inoperable HPB cancers.  
- MVT-5873 is well tolerated as a single agent; moderate elevations in AST/ALT appear to be dose-limiting.  
- Patients with resectable Sialyl Lewisa-expressing cancers represent an ideal population to explore the use of perioperative MVT-5873 given moderate level of CA 19-9 elevations, and the potential for extension of recurrence-free survival.  
Objectives:
- Document the safety of perioperative MVT-5873 in patients undergoing pancreas and liver resections.  
- Determine if perioperative MVT-5873 can decrease 1-year recurrence rates for patients with operable CA 19-9-producing cancers.
untitled h1
eligibility

- Histologically or cytologically confirmed adenocarcinoma of the o Colon (metastatic to liver) o Pancreas o Bile Ducts (Cholangiocarcinoma)
- Serum CA 19-9 levels greater than the upper limit of normal, but less than 2500.  
- Disease amenable to complete surgical extirpation.
untitled h1
design

- Pre-operative one-time treatment with MVT-5873, resection to remove all demonstrable disease in the liver, bile ducts and pancreas, and continuing MVT-5873 mono-therapy until off treatment criteria are met.  
| TABLE OF CONTENTS    |                                                                                                    |
|----------------------|----------------------------------------------------------------------------------------------------|
| PRÉCIS .             |                                                                                                    |
| TABLE OF CONTENTS.   |                                                                                                    |
| LIST OF TABLES .     |                                                                                                    |
| LIST OF FIGURES      |                                                                                                    |
| l                    | INTRODUCTION .                                                                                     |
| 1.1                  | Study Objectives .                                                                                 |
| Primary Objectives   |                                                                                                    |
| 1.1.1                |                                                                                                    |
| 1.1.2                | Secondary Objective                                                                                |
| 1.1.3                | Exploratory Objectives .                                                                           |
| 1.2                  | Background and Rationale .                                                                         |
| HPB Cancers          |                                                                                                    |
| 1.2.1                |                                                                                                    |
| 1.2.2                | Sialyl Lewis A Antigen - CA19-9                                                                    |
| 1.2.3                | MVT-5873 .                                                                                         |
| Preclinical Studies  |                                                                                                    |
| 1.2.4                | Effects in Humans                                                                                  |
| 1.2.5                |                                                                                                    |
| 1.2.6                | Pharmacokinetics .                                                                                 |
| 1.2.7                | Reductions in Serum CA19-9 Levels                                                                  |
| 1.2.8                | Responses to Therapy .                                                                             |
| 1.2.9                | RATIONALE                                                                                          |
| 1.2.10               | Metastasis Assays Using Ex Vivo Perfusion of Omentum........................................... 36 |
| 1.2.11               | Ex Vivo Hepatic Perfusion Model                                                                    |
| 1.2.12               | Simulated Metastasis Assay with Relevant Tissue (SMART) Chamber ............... 37                 |
| 1.2.13               | The SMART System                                                                                   |
| 2                    | ELIGIBILITY ASSESSMENT AND ENROLLMENT.                                                             |
| 2.1                  | Eligibility Criteria .                                                                             |
| 2.1.1                | Inclusion Criteria .                                                                               |
| 2.1.2                | Exclusion Criteria .                                                                               |
| 2.1.3                | Recruitment Strategies .                                                                           |
| Screening Evaluation |                                                                                                    |
| 2.2                  |                                                                                                    |
| 2.2.1                | Screening Activities Performed Prior to Obtaining Informed Consent .................. 43           |
| 2.2.2                | Screening Activities Performed After a Consent for Screening Has Been Signed. 43                   |  
| Version Date: 08/10/2020                             |                                                                                                                     |
|------------------------------------------------------|---------------------------------------------------------------------------------------------------------------------|
| 2.3                                                  | Participant Registration and Status Update Procedures ........................................................... 4 |
| 2.3.1                                                | Treatment Assignment and Randomization/Stratification Procedures .................. 44                              |
| 2.4                                                  | Baseline Evaluation                                                                                                 |
| 3                                                    | STUDY IMPLEMENTATION .                                                                                              |
| 3.1                                                  | Study Design .                                                                                                      |
| 3.1.1                                                | Dose Limiting Toxicity                                                                                              |
| 3.1.2                                                | Safety Lead-In Cohorts .                                                                                            |
| T Treatment Cohorts                                  |                                                                                                                     |
| 3.1.3                                                |                                                                                                                     |
| 3.2                                                  | Study Drug Administration                                                                                           |
| 3.2.1                                                | Preparation and Administration of MVT-5873                                                                          |
| 3.2.2                                                | Stability of MVT-5873 Following Preparation.                                                                        |
| Upper Limit to Administered Dose in Obese Subjects . |                                                                                                                     |
| 3.2.3                                                |                                                                                                                     |
| 3.2.4                                                | Infusion Rate of MVT-5873                                                                                           |
| 3.2.5                                                | Treatment Guidelines for MVT-5873-Related Infusion Reaction                                                         |
| Premedication                                        |                                                                                                                     |
| 3.2.6                                                |                                                                                                                     |
| Dose Modifications .                                 |                                                                                                                     |
| 3.3                                                  |                                                                                                                     |
| 3.4                                                  | Study Calendar .                                                                                                    |
| 3.5                                                  | Surgical Guidelines                                                                                                 |
| Preoperative Patient Management                      |                                                                                                                     |
| 3.5.1                                                |                                                                                                                     |
| 3.5.2                                                | Patient Management in the Operating Room                                                                            |
| 3.5.3                                                | Postoperative Care .                                                                                                |
| 3.5.4                                                | D Discharge                                                                                                         |
| Pathological Evaluation                              |                                                                                                                     |
| 3.6                                                  |                                                                                                                     |
| 3.7                                                  | Criteria for Removal from Protocol Therapy and Off Study Criteria ................................. 59              |
| 3.7.1                                                | Criteria for Removal from Protocol Therapy                                                                          |
| 3.7.2                                                | OFF-Study Criteria .                                                                                                |
| 4                                                    | CONCOMITANT MEDICATIONS/MEASURES .                                                                                  |
| S                                                    | BIOSPECIMEN COLLECTION .                                                                                            |
| 5.1                                                  | Specimen Collection Table                                                                                           |
| 5.2                                                  | Correlative Studies for Research/Pharmacokinetic Studies ..................................................... 62   |
| Tumor and Tissue Collection                          |                                                                                                                     |
| 5.2.1                                                |                                                                                                                     |
| 5.2.2                                                | RNA Evaluation                                                                                                      |
untitled h1
design

| Version Date: 08/10/2020 5.2.3 Metastasis Assays Using Ex Vivo Perfusion of Omentum 63 5.2.4 Ex Vivo Hepatic Perfusion Model 63 5.2.5 Circulating Tumor Cell Evaluation 63 5.2.6 Monocyte Phagocyte System (MPS) Profiling 64 Sample Storage, Tracking and Disposition 64 5.3.1 Samples Managed by the Blood Processing Core (BPC) 64 5.3.2 Procedures for Storage of Patient Samples in the Laboratory of Dr. Trepel 65 5.3.3 Procedures for Storage of Tissue Specimens in the Laboratory of Pathology 66 5.3.4 Procedures for Storage of Patient Samples in the Laboratory of Dr. Hernandez ... 66 5.3.5 Protocol Completion/Sample Destruction 66 5.3.6 A Certificate of Confidentiality 67 5.3.7 Management of Results 67 5.3.8 Genetic Counseling 67 6 DATA COLLECTION AND EVALUATION 67 Data Collection 67 6.1.1 Routine Data Collection 68 Data Sharing Plans 69 6.2.1 Human Data Sharing Plan 69 6.2.2 Genomic Data Sharing Plan 70 Response Criteria 70 6.3.1 Definitions 70 6.3.2 Disease Parameters 70 6.3.3 Methods for Evaluation of Measurable Disease 71 6.3.4 Response Criteria 73 6.3.5 Duration of Response 76 6.3.6 Progression-Free Survival 76 Toxicity Criteria 76 7 NIH REPORTING REQUIREMENTS / DATA AND SAFETY MONITORING PLAN .. 76 Definitions 76 OHSRP Office of Compliance and Training / IRB Reporting 76 7.2.1 Expedited Reporting 76 7.2.2 IRB Requirements for PI Reporting at Continuing Review 77   |
|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|  
Abbreviated Title: Perioperative MVT-5873  
| Version Date: 08/10/2020                     |                                                                                                                                                       |
|----------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------|
| NCI Clinical Director Reporting              |                                                                                                                                                       |
| 7.3                                          |                                                                                                                                                       |
| 7.4                                          | NIH Required Data and Safety Monitoring Plan .                                                                                                        |
| 7.4.1                                        | Principal Investigator/Research Team .                                                                                                                |
| 8                                            | SPONSOR SAFETY REPORTING .                                                                                                                            |
| 8.1                                          | DEFINITIONS                                                                                                                                           |
| 8.1.1                                        | A Adverse Event                                                                                                                                       |
| 8.1.2                                        | Serious Adverse Event (SAE)                                                                                                                           |
| Life-threatening                             |                                                                                                                                                       |
| 8.1.3                                        |                                                                                                                                                       |
| 8.1.4                                        | Severity .                                                                                                                                            |
| 8.1.5                                        | Relationship to Study Product                                                                                                                         |
| 8.2                                          | Assessment of Safety Events                                                                                                                           |
| Reporting of Serious Adverse Events          |                                                                                                                                                       |
| 8.3                                          |                                                                                                                                                       |
| 8.4                                          | Reporting Pregnancy                                                                                                                                   |
| 8.4.1                                        | Maternal Exposure                                                                                                                                     |
| Paternal Exposure                            |                                                                                                                                                       |
| 8.4.2                                        |                                                                                                                                                       |
| 8.5                                          | Regulatory Reporting for Studies Conducted Under CCR-Sponsored IND ................. 80                                                               |
| 9                                            | CLINICAL MONITORING                                                                                                                                   |
| 10                                           | STATISTICAL CONSIDERATIONS                                                                                                                            |
| Statistical Hypotheses                       |                                                                                                                                                       |
| 10.1                                         |                                                                                                                                                       |
| 10.1.1                                       | Primary Efficacy Endpoints ....................................................................................................................... 81 |
| 10.1.2                                       | Secondary Efficacy Endpoints                                                                                                                          |
| 10.2                                         | Sample Size Determination: .                                                                                                                          |
| Populations for Analysis                     |                                                                                                                                                       |
| 10.3                                         |                                                                                                                                                       |
| 10.3.1                                       | Evaluable for Toxicity .                                                                                                                              |
| 10.3.2                                       | Evaluable for Objective Response .                                                                                                                    |
| Evaluable Non-Target Disease Response        |                                                                                                                                                       |
| 10.3.3                                       |                                                                                                                                                       |
| 10.4                                         | Statistical Analyses                                                                                                                                  |
| 10.4.1                                       | General Approach .                                                                                                                                    |
| 10.4.2                                       | Analysis of the Primary Efficacy Endpoints                                                                                                            |
| Analysis of the Secondary Efficacy Endpoints |                                                                                                                                                       |
| 10.4.3                                       | Se                                                                                                                                                    |
| 10.4.4                                       |                                                                                                                                                       |
| 10.4.5                                       | Baseline Descriptive Statistics                                                                                                                       |  
Version Date: 08/10/2020  
| Version Date: 08/10/2020                                                                                                                           |                                                                                                                                                                |                                                                                                                                       |
|----------------------------------------------------------------------------------------------------------------------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------------------|---------------------------------------------------------------------------------------------------------------------------------------|
| Planned Interim Analyses ...................................................................................................................... 84 |                                                                                                                                                                |                                                                                                                                       |
| 10.4.6                                                                                                                                             |                                                                                                                                                                |                                                                                                                                       |
| 10.4.7                                                                                                                                             | Exploratory Analyses                                                                                                                                           |                                                                                                                                       |
| 11                                                                                                                                                 | COLLABORATIVE AGREEMENTS .                                                                                                                                     |                                                                                                                                       |
| Material Transfer Agreement (MTA)                                                                                                                  |                                                                                                                                                                |                                                                                                                                       |
| 1.1                                                                                                                                                |                                                                                                                                                                |                                                                                                                                       |
| 12  HUMAN SUBJECTS PROTECTIONS .                                                                                                                   |                                                                                                                                                                |                                                                                                                                       |
| 12.1                                                                                                                                               | Rationale For Subject Selection                                                                                                                                |                                                                                                                                       |
| Participation of Children                                                                                                                          |                                                                                                                                                                |                                                                                                                                       |
| 12.2                                                                                                                                               |                                                                                                                                                                |                                                                                                                                       |
| 12.3                                                                                                                                               | Participation of Subjects Unable to Give Consent .                                                                                                             |                                                                                                                                       |
| 12.4                                                                                                                                               | Evaluation of Benefits and Risks/Discomforts                                                                                                                   |                                                                                                                                       |
| 12.4.1                                                                                                                                             | Risks                                                                                                                                                          |                                                                                                                                       |
| 12.4.2                                                                                                                                             | Risks of Exposure to Ionizing Radiation                                                                                                                        |                                                                                                                                       |
| 12.4.3                                                                                                                                             | Non-Physical Risks of Genetic Research ................................................................................ 88                                     |                                                                                                                                       |
| 12.4.4                                                                                                                                             | Benefits .                                                                                                                                                     |                                                                                                                                       |
| 12.4.5                                                                                                                                             | Certificate of Confidentiality .                                                                                                                               |                                                                                                                                       |
| Consent Process and Documentation                                                                                                                  |                                                                                                                                                                |                                                                                                                                       |
| 12.5                                                                                                                                               | Request for Waiver of Consent for Screening Activities................................................. 89                                                     |                                                                                                                                       |
| 12.5.1                                                                                                                                             |                                                                                                                                                                |                                                                                                                                       |
| 13                                                                                                                                                 | PHARMACEUTICAL AND INVESTIGATIONAL DEVICE INFORMATION ............. 89                                                                                         |                                                                                                                                       |
| Drug MVT-5873 (IND # 144112)                                                                                                                       |                                                                                                                                                                |                                                                                                                                       |
| 13.1                                                                                                                                               | SOURCE .                                                                                                                                                       |                                                                                                                                       |
| 13.1.1                                                                                                                                             |                                                                                                                                                                |                                                                                                                                       |
| 13.1.2                                                                                                                                             | T Toxicity .                                                                                                                                                   |                                                                                                                                       |
| 13.1.3                                                                                                                                             | Formulation and Preparation................................................................................................................... 90              |                                                                                                                                       |
| 13.1.4                                                                                                                                             | Stability and Storage                                                                                                                                          |                                                                                                                                       |
| Administration Procedures                                                                                                                          |                                                                                                                                                                |                                                                                                                                       |
| 13.1.5                                                                                                                                             |                                                                                                                                                                |                                                                                                                                       |
| 13.1.6                                                                                                                                             | Incompatibilities                                                                                                                                              |                                                                                                                                       |
| 14                                                                                                                                                 | REFERENCES .                                                                                                                                                   |                                                                                                                                       |
| APPENDICES .                                                                                                                                       |                                                                                                                                                                |                                                                                                                                       |
| 15                                                                                                                                                 | 15.1                                                                                                                                                           | Appendix A: Performance Status Criteria .......................................................................................... 94 |
| 15.2                                                                                                                                               | Appendix B: Toxicity Tables ............................................................................................................................... 95 |                                                                                                                                       |
| 15.3                                                                                                                                               | Appendix C: Investigator Agreement                                                                                                                             |                                                                                                                                       |
untitled h1
list of tables

Table 1: Pharmacokinetic Parameters for MVT-5873 Following Single Intravenous Doses of 10, 30, and 100 mg/kg of MVT-5873 to Cynomolgus Monkeys................................ 17 Table 2: Pharmacokinetic Parameters for MVT-5873 Following Intravenous Administration of 30 mg/kg and 100 mg/kg of MVT-5873 to Cynomolgus Monkeys on Day 1 and Day 8 .................................................................................................................... 17 Table 3: Dose Cohorts, Cycles Administered and Patient Disposition ........................................ 29 Table 4: Summary of MVT-5873 Dose Limiting Toxicities and Treatment Disposition ..... 30 Table 5: All Adverse Events, All Grades, in ≥ 30% of Subjects .................................................. 31 Table 6: All Treatment-Related Adverse Events ≥ Grade 3 ......................................................... 32 Table 7: MVT-5873 Human Pharmacokinetics ............................................................................ 32 Table 8: Normalization (< 37U/mL) of CA 19-9 Levels Following Adminstration of MVT5873............................................................................................................................. 33 Table 9: Safety Lead-In Study Design .......................................................................................... 46 Table 10: Safety Lead-In Cohorts ................................................................................................. 47 Table 11: Treatment Study Design ............................................................................................... 49 Table 12: Treatment Guidelines (If Two Different Pre-Operative MTDs by LFT Elevation are Determined from Safety Lead-In) ......................................................................... 49 Table 13:Treatment Study Design (If Two Different Pre-Operative MTDs by LFT Elevation are Determined from Safety Lead-In) ......................................................................... 50
untitled h1
list of figures

Figure 1: Top left: pancreas, ductal adenocarcinoma, stage III .................................................... 13 Figure 2: Tumor Growth Over Time - BxPC3 Xenograft Model with MVT-5873 Doses of 5, 15, and 30 mg/kg as a Single Agent and in Combination with Gemcitabine/nabpaclitaxel ..................................................................................................................... 14 Figure 3: Immunohistochemical Staining of BxPC3 Tumor Tissue Samples at 20X
Magnification; MVT-5873 (top panel) and Human IgG (bottom panel) .................... 15 Figure 4: Mean MVT-5873 Serum Concentrations Over Time in BxPC3 Tumor-Bearing Mice and Nontumor-Bearing Mice ............................................................................. 16 Figure 5: MVT-5873 Serum Concentration and BxPC3 Tumor Volume Versus Time ............... 16 Figure 6: Serum Levels (AUC0-168 and Cmax) of MVT-5873 after Repeated Intravenous Dosing in Cynomolgus Monkeys (Males and Females Combined) on Day 1 and Day 8 ........................................................................................................................... 18 Figure 7: Liver Function Test with 3mg/kg Weekly Dosing of MVT-5873 from Six Subjects ... 28 Figure 8: Efficacy for MVT-5873................................................................................................. 34 Figure 9: Percent Change in Target Lesions in Cohort B3 (0.125 mg/kg MVT-5873) ................ 35  
| Figure 10: The SMART System Consisting of SMART Platform 38 Figure 11: Ex Vivo Normal and Tumor Preservation in the SMART System 39 Figure 12: SMART Platforms are Valuable Tools for Drug Evaluations Such as  Pembrolizumab 40 Figure 13: Operating Room Workflow 58   |
|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|  
Figure 14: Pathologic Evaluation of Tumor and Liver Tissue Removed at the Time of  
Surgery ........................................................................................................................ 59  
Figure 15: Scientific Endpoints. Evaluation of Circulating Tumor Cells (top) and Ex Vivo Tissue Modeling (bottom)........................................................................................... 63
untitled h1
1 introduction study objectives 1.1.1

- Document the safety of perioperative MVT-5873 in patients undergoing pancreas and liver resections.  
- Determine if perioperative MVT-5873 can decrease 1-year recurrence rates for subjects with operable CA 19-9 producing cancers.
untitled h1
1.1.2 secondary objective

- Define disease free survival (DFS) for subjects treated with preoperative MVT-5873.
untitled h1
1.1.3 exploratory objectives

- Define overall survival (OS) for subjects treated with preoperative MVT-5873.  
- Determine the mechanism(s) of liver function test abnormalities associated with MVT5873 administration.  
- Determine the tumor penetration of MVT-5873 in primary tumors of the pancreas and bile ducts, and in metastatic colorectal cancers to the liver.  
- Evaluate the impact of MVT-5873 therapy on circulating tumor cells. - Develop ex *vivo* models for pancreatic cancer, cholangiocarcinoma, and metastatic colorectal cancer.  
- Evaluate chemotherapeutic and immunotherapeutic agent effects in tumors from patients undergoing pancreas and liver resections after pre-operative MVT-5873 therapy using a novel ex *vivo* platform, the SMART System.
untitled h1
background and rationale

1.2.1 **HPB Cancers**
Cancers of the pancreas, liver (primary and metastatic) and bile ducts represent some of the most recalcitrant diseases encountered by the practicing oncologist. When identified early, these cancers can be removed with technically challenging operations. Fortunately, experience has decreased the perioperative complications associated with these operations. However, the ability to prevent disease recurrence following the procedure has not yet come to fruition. Although resection remains the only hope for cure, this end-point is achieved by very few. Analysis of randomized prospective trial data and single institution series strongly suggests that nearly half of all patients undergoing resections for pancreatic cancer, cholangiocarcinoma and metastatic colorectal cancer to the liver will experience a recurrence of disease in the first 12-15 months following removal of tumor(s).[1-3] In other words, these patients underwent an operation for likely abbreviated benefit in terms of survival. The recently reported, not yet published, data on the BILCAP (cholangiocarcinoma) is very similar to these outcomes below. It has been postulated for some time that the operation itself may hasten demise for some patients by inciting an inflammatory milieu and triggering an angiogenic switch in remote dormant tumor cells. Substantial data exist in mice[4] supporting this concept but data in humans is admittedly lacking. We do however know that surgery is associated with increases in circulating tumor cell numbers, likely secondary to manipulation during the procedure.[5] It would be ideal to give a perioperative agent with efficacy against the tumor cells at the time of resection. Unfortunately, chemotherapy has deleterious effects on wound healing that would make resection an extremely-risky endeavor. However, a well-tolerated monoclonal antibody targeting a tumor cell epitope may hold promise in mitigating those early recurrences and thereby increasing the efficacy of resection for cancers of the pancreas, liver and bile ducts. This concept of immediate perioperative therapy is unique and additive to what is generally undertaken with high-risk cancers. For example, patients could receive standard neo-adjuvant and/or adjuvant therapies in addition to immediate perioperative therapy without altering the standard regimens. Moreover, antibody penetration and binding to tumor cells would be directly assessed on resected specimens, as would the effects on circulating tumor cell populations (which have been demonstrated to be prognostic[6-8]), which could be used to stratify patients for combination antibody/chemotherapy should disease recur. In short, the employment of immediate perioperative therapy, if demonstrated to be safe, could add an additional tool in the oncologist's armamentarium for resectable high-risk cancers.
untitled h1
1.2.2 sialyl lewis a antigen

The Sialyl Lewis A (sLea) antigen is an epitope present on Carbohydrate Antigen 19-9 (CA 19-9),
defined by the mouse monoclonal antibody 116-NS-19-9, shown to be overexpressed on epithelial cell tumors.[9, 10] SLea is an oligosaccharide expressed primarily as a proteoglycan that is secreted and circulates as a mucin form, and also as a less well studied glycolipid form.[10, 11] The sLea antigen is predominantly expressed on cancer cells.[12] As a ligand for E selectin, sLea facilitates tumor adhesion and extravasation, key events for tumor metastasis, and is thus a marker of an aggressive tumor phenotype.[13] Glycolipids, such as sLea, are established targets for cancer immunotherapies.[14] The monoclonal antibody MVT-5873 (also referred to as HuMab-5B1 or 5B1), the investigational agent in this study, specifically targets the sLea antigen.  
CA 19-9 is widely expressed on tumors of the gastrointestinal tract, with up to 94% of pancreatic cancers positive for CA 19-9 expression and high expression rates also seen in bile duct carcinomas and transitional cell carcinomas.[15, 16] Additionally, expression of CA 19-9 is frequently seen in ovarian, colon, stomach, and distal esophagus/stomach cancers. Circulating serum levels of CA 19-9 have been validated as a biomarker for assessing the metastatic potential of pancreatic ductal adenocarcinomas (PDAC)[17, 18] and have been used to evaluate the aggressiveness of other epithelial cell cancers.[19, 20] As a known ligand for endothelial leukocyte adhesion molecules, CA 19-9 expression is associated with increased metastatic potential in colon cancer[13, 21, 22] and pancreatic adenocarcinoma.[23] Serum CA 19-9 levels have also been found to be informative with respect to prognosis and treatment effect in patients with pancreatic cancer, with several studies correlating increasingly higher serum levels with poorer survival outcomes.[17, 18, 24] In a Phase I/II clinical trial of nabpaclitaxel and gemcitabine in patients with advanced pancreatic cancer, decreases in CA 19-9 levels correlated with tumor response, PFS, and OS.[25] In a Phase II study of 5-fluorouracil-based chemoradiotherapy in patients with locally advanced pancreatic cancer, a greater than 90% reduction in CA 19-9 levels from baseline was associated with significantly improved median survival time, with a multivariate analysis finding a-post therapy CA 19-9 level of less than 85.5 U/mL to be an independent prognostic factor for survival.[26] On normal cells, expression of CA 19-9 is restricted to the apical brush border, a relatively inaccessible location due to the basement membrane and the tight junctions of mature and polarized ductal epithelium.[27] In patients with CA 19-9 positive tumors, about 5% are negative for circulating levels of CA 19-9, and this appears to be related to poor secretion or release of CA 19-9 into the blood stream, not necessarily to expression of the CA 19-9 antigen by the tumor cell.[28-30] Because CA 19-9 has a high expression rate in patients with pancreatic cancer, this antigen was identified as a target for antibody-based biopharmaceuticals, and specifically this investigation of MVT-5873 in patients with PDAC.
untitled h1
1.2.3 mvt-5873

MVT-5873 (HuMab-5B1) is a fully human IgG1 lambda antibody discovered by MabVax in collaboration with investigators at Memorial Sloan Kettering Cancer Center (MSKCC). MVT5873 was identified from blood lymphocytes from a breast cancer patient following immunization with a sLea-KLH vaccine.[31] This vaccine has been shown to induce high titers of both IgG and IgM antibodies against sLea in mice and humans without cross-reactivity to other similar blood group carbohydrate antigens.[31] Two human monoclonal antibodies (5B1 and 7E3) with highaffinity for sLea were identified, subsequently expressed as a recombinant antibodies, and further characterized.[32]
Evaluation of the specificity of MVT-5873 demonstrated cell surface binding in sLea positive colon cancer (HT29 and Colo205), ovarian cancer (SW626), small cell lung cancer (DMS79), and pancreatic cancer (BxPC3) cell lines, but not to a sLea negative melanoma (SK-MEL28) cell line.[32] Importantly, MVT-5873 did not bind to Lea, sLex, Ley, or other related carbohydrates when evaluated by enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance.[32] In addition, an independent binding analysis conducted by the Consortium on Functional Glycomics Core H group using a glycan array with 465 distinct carbohydrates revealed that MVT-5873 had exquisite specificity for the carbohydrate epitope of the sLea antigen.  
MVT-5873 has been shown to be potent in complement-dependent cytotoxicity (CDC) assays and antibody-dependent cell mediated cytotoxicity (ADCC) assays (see below). Both of these mechanisms are consistent with antitumor activity. Based on these findings, MVT-5873 was selected for further development as a potential therapy for patients with CA 19-9 expressing tumor types.
untitled h1
1.2.4 preclinical studies 1.2.4.1 tissue

MVT-5873 binding to tumor TMA appeared to be specific, differentially expressed, and restricted, with most normal tissues lacking reactivity. Expression of sLea was present on several adenocarcinomas, including pancreatic cancers, a finding consistent with literature reports of sLea expression identified by CA 19-9 antibody binding (**Figure 1**). Most of the human specimens evaluated in the TMA analysis were negative for staining. However, expression of sLea antigen was found on mucous glands of the small intestine, colon (goblet cells), and rectum. There was also expression in exocrine pancreas, germinal center of lymph node, gallbladder, breast (ductal cells), and uterine cervix of normal human tissue. The observed binding pattern is consistent with expression in tissues producing exocrine secretions, i.e. mucins that carry the sLea antigen.  
Strong positive staining was seen in 23/34 colon adenocarcinomas (68%), 34/58 adenocarcinoma  
 metastases to the ovary (59%), and 7/9 pancreatic ductal cancers (62%) at various stages. As shown in the figure below, typical reactivity shows as diffuse cytoplasmic staining, with some tumor cells clearly showing distinct staining at the cell membrane level. In addition, signet ring-metastatic to ovaries, some lung (i.e. bronchioalveolar, mucoepidermoid) and metastatic breast cancer lymph node biopsy cores were found to be strongly positive, albeit at lower frequencies. In contrast, only 2/43 prostate cancer samples (5%) and 0/51 gastrointestinal stromal tumors (GIST) specimens were positive (data not shown).  
Figure 1: Top left: pancreas, ductal adenocarcinoma, stage III
Top center: sigmoid colon, carcinoma stage IIIB Top right: lung adenocarcinoma, stage IB
Bottom left: urinary bladder, mucinous adenocarcinoma, stage IV
Bottom center: ovary, metastatic carcinoma from colon Bottom right: lymph node, breast carcinoma, stage IIIA
untitled h1
1.2.4.2 evaluation of mvt-5873 in

In an early exploratory study, the antitumor potential of MVT-5873 was assessed using a Colo205luc xenograft model of metastatic colon cancer in SCID mice.[32] Across two experiments, groups of 5 mice were given either 4 doses of MVT-5873 at 5 mg/kg (20 mg/kg total dose) or 6 doses of MVT-5873 at 5 mg/kg (30 mg/kg total) intraperitoneally, each over the course of 21 days, while animals in the control groups received injections of phosphate buffered saline (PBS). Across all the control animals, the average median survival was 102 days and all died within 155 days. Treatment improved survival significantly; median survival in animals that received 4 doses of MVT-5875 was 207 days (p < 0.05; HR = 3.46), and in the animals that received 6 doses of MVT5873, median survival had not yet been reached at day 301, the termination of the experiment (p
< 0.01; HR =6.375).  
To further evaluate the antitumor activity of MVT-5873, a series of *in vivo* studies were conducted utilizing human pancreatic cancer (BxPC3) xenograft models.  
The combination of MVT-5873, both a single agent and in combination with gemcitabine and nabpaclitaxel in mice bearing BxPC3 tumor xenografts was evaluated. In this study MVT-5873 was administered as doses of 5, 15, and 30 mg/kg, blood samples for PK analysis were collected from all study groups, and tumor tissues were harvested at the end of study for IHC staining. As had been observed in the previous BxPC3 xenograft experiments, single-agent MVT-5873  
 demonstrated tumor growth inhibition and growth delay at all dose levels compared to controls
(**Figure 2**).  
Tumor tissue samples taken at the end of treatment were sectioned and stained for the presence of the sLea target using MVT-5873 as a probe **Figure 3**. Human IgG using goat anti-human IgG
antibody was used as a probe for residual MVT-5873 bound to tumor post administration. Representative tissue staining samples are depicted. IHC staining signal intensities, recorded as optical densities, across the two panels are shown in the **Figure 3** below.  
In the **Figure 3**, top panel, staining the presence of the sLea was high and similar in all samples, indicating a relative abundance of the sLea target in these tissues. These data provide evidence that upon treatment of gemcitabine plus nab-paclitaxel there is no change in expression of the target epitope. In the bottom panel, when stained with human IgG that lacks binding to the target epitope, the presence of residual MVT-5873 in tumor tissue was observed across all MVT-5873 treatment groups, with staining density correlating with response. The complete lack of staining in the chemotherapy alone group serves as a negative control.  
As shown in the **Figure 3**, signal intensities of IgG staining across all MVT-5873 treatment groups  
  
were higher than that of the human IgG control, indicating binding selectivity of MVT-5873 in tumor tissue. There was a significant difference in signal intensity between the MVT-5873 5 mg/kg + chemotherapy group and the MVT-5873 15 mg/kg + chemotherapy group (P < 0.05), suggesting dose-dependent binding of MVT-5873 to tumor tissues.  
These xenograft studies indicate that MVT-5873 has activity in the known sLea-bearing tumor models. Analysis of MVT-5873 binding to tumor tissue by IHC revealed an apparent dose-dependent uptake of MVT-5873 in tumor tissue.  
Serum concentrations were measured. Further analysis of these data shown in **Figure 4** and Figure 5 indicate an inverse relationship between MVT-5873 serum concentrations and tumor volume.  
This suggests that the presence or expansion of the sLea target, either on tumor tissue or in circulation, decreases the amount of free MVT-5873 in the serum and contributes to the systemic clearance of MVT-5873.  
  
  
These studies support the clinical evaluation of MVT-5873, as a single agent and in combination with chemotherapy, in subjects with pancreatic cancer and other sLea positive tumor types.
untitled h1
1.2.4.3 pharmacokinetics 1.2.4.4 single-dose pharmacokinetics

In a single-dose IV PK study in cynomolgus monkeys (non-GLP Study 20068188), MVT-5873 serum levels were determined over a one-month period using an ELISA procedure. MVT-5873 exhibited a low clearance from serum (0.384 to 0.503 mL/min/kg) and a low volume of distribution (40.9 to 83.0 mL/kg), with a resultant long half-life (65.2 to 217 hours) at dose levels of 10, 30 or 100 mg/kg MVT-5873 (Table 1).  
Table 1: Pharmacokinetic Parameters for MVT-5873 Following Single Intravenous Doses of 10, 30, and 100 mg/kg of MVT-5873 to Cynomolgus Monkeys  
Dose  
(mg/kg) Sex **Tmax**  
(h)  
Cmax  
(µg/mL)  
AUC0-t (µg∙h/mL)  
t1/2 (h)  
CL  
(mL/min/kg)  
Vd (mL/kg)  
10  
Female 0.250 141 12,800 NC NC NC  
Male 0.250 254 22,900 65.2 0.424 40.9  
30  
Female 0.280 499 41,900 NC NC NC  
Male 0.250 736 64,700 217 0.384 120  
100 Female 0.750 1780 169,000 103 0.503 74.9 Male 0.750 2190 213,000 126 0.456 83.0 Abbreviation: NC=Not Calculated (coefficient of determination less than 0.800 or the extrapolation of the AUC to infinity represented more than 20% of the total area)
The low serum clearance seen with MVT-5873 is typical of monoclonal antibodies, where the main route of elimination is generally thought to be by receptor binding and internalization and/or protein catabolism. The low volume of distribution is typical for a monoclonal antibody, since it is similar to the volume of plasma for the cynomolgus monkey and suggests limited distribution into tissue. 1.2.4.5 Repeat-Dose Pharmacokinetics A multiple-dose toxicity study in cynomolgus monkeys (non-GLP Study 20068122) evaluated the effects of MVT-5873 (30 or 100 mg/kg) administered IV once a week for 2 weeks (2 doses). The toxicokinetic evaluations were conducted over each dosing interval on Days 1-7 and Days 8-15, revealing that exposure, measured by AUC0-168, was approximately dose-proportional and similar across the dosing interval for both sexes, ranging from 26,600 to 146,00 µg•hr/mL for dose levels of 30 and 100 mg/kg, respectively (Table 2, **Figure 6**). There was minor MVT-5873 plasma accumulation when administered once weekly. Similarly, in a GLP 4-week once weekly (4 doses) IV toxicity study, trough plasma levels measured weekly were either similar to or greater than Week 1 indicating good exposure to MVT-5873 over the 4-week dosing period.  
Table 2: Pharmacokinetic Parameters for MVT-5873 Following Intravenous Administration of 30 mg/kg and 100 mg/kg of MVT-5873 to Cynomolgus Monkeys on Day 1 and Day 8  
| Day   | Dose (mg/kg)   | Sex   | Cmax (μg/mL)   | AUC0-168 (μg•h/mL)   |
|-------|----------------|-------|----------------|----------------------|
| 30    | Female         | 843   | 33,300         |                      |
| 30    | Male           | 977   | 34,800         |                      |
| 30    | Mean (M&F)     | 910   | 34,050         |                      |
| 100   | Female         | 2260  | 151,000        |                      |
| 100   | Male           | 2380  | 118,000        |                      |
| 100   | Mean (M&F)     | 2320  | 134,500        |                      |
| 30    | Female         | 760   | 38,400         |                      |
| 30    | Male           | 881   | 48,900         |                      |
| 30    | Mean (M&F)     | 821   | 43,650         |                      |
| 100   | Female         | 2540  | 212,000        |                      |
| 100   | Male           | 2450  | 148,000        |                      |
| 100   | Mean (M&F)     | 2495  | 180,000        |                      |  
| 1  8   |
|--------|  
  
In addition to assessing PK in cynomolgus monkeys, serum levels of MVT-5873 were measured at various times during a multiple dose PK study in both nontumor- and tumor-bearing SCID mice. After intraperitoneal administration, MVT-5873 serum levels were maintained at an average of 46.1 µg/mL in nontumor-bearing mice. Mice with established CA 19-9 expressing tumor xenografts demonstrated lower serum levels, indicating a relatively higher clearance compared to nontumor-bearing mice. Further information on the pharmacokinetics of MVT-5873 in mice the presence and absence of tumor was collected as part of the second BxPC3 xenograft study described above, which evaluated MVT-5873 as a single agent at doses of 5, 15, and 30 mg/kg as a single agent and in combination with 80 mg/kg of gemcitabine plus 20 mg of nab-paclitaxel. This study included an additional treatment group of nontumor-bearing mice that were treated with MVT-5873 15 mg/kg. Serial pre-dose (Cmin) concentrations of MVT-5873 were collected on Days 13, 16, 23, 30, and 37, with an additional sample collected 1 week later, on Day 44. As shown in the figure below, in the absence of tumor, serial Cmin MVT-5873 concentrations (15 mg/kg dose level) appeared to slightly accumulate throughout treatment. However, in all tumor-bearing animals, serial Cmin values declined over time, as had been observed previously. 1.2.4.6 Preclinical Safety The preclinical studies conducted to determine the safety of MVT-5873 are summarized in the table below. Several toxicology studies have been conducted with MVT-5873 in cynomolgus monkeys and included a 4-week (4 dose) GLP safety study, a 2-week (2 dose) exploratory nonGLP toxicology study, and a single dose tolerability study. All studies were conducted at Charles River Laboratories (Reno, NV). To support the cynomolgus monkey as an appropriate model for MVT-5873 toxicology studies, pivotal (GLP) tissue cross-reactivity studies were conducted to assess the binding of MVT-5873 to cryosections (or to fixed blood) of normal human tissues (GLP Study 20067038) and normal cynomolgus monkey tissues (GLP Study 20074271); these studies were conducted at Charles River Laboratories (Pathology Associates, Frederick, MD).  
| Study                                                                                                                                      | Type                                                                             | and              | Route           | /        | Mode           | of       | Species   | Doses   | Study   | No.   |
|--------------------------------------------------------------------------------------------------------------------------------------------|----------------------------------------------------------------------------------|------------------|-----------------|----------|----------------|----------|-----------|---------|---------|-------|
| Duration                                                                                                                                   | Administration                                                                   | Compliance       |                 |          |                |          |           |         |         |       |
| Single dose Single-dose tolerability  and pharmacokinetics  with 4 week postdose  observation/blood  sampling period                       | IV slow bolus (1-3 min) or  30-min infusion via a  saphenous vein                | Monkey/          | 0, 10, 30 mg/kg |          |                |          |           |         |         |       |
| Cynomolgus                                                                                                                                 | (slow bolus) 100 mg/kg (30- min infusion)                                        | 20068188 Non-GLP |                 |          |                |          |           |         |         |       |
| Repeat-dose studies Once weekly (2 doses)  toxicity and  toxicokinetic study  with terminal sacrifice  2 weeks after the last  dose        | IV slow bolus (1-3 min) or  30-min infusion via a  saphenous vein                | Monkey/          | 0,              | 30 mg/kg |                |          |           |         |         |       |
| Cynomolgus                                                                                                                                 | (slow bolus) 100 mg/kg (30- min infusion)                                        | 20068122 Non-GLP |                 |          |                |          |           |         |         |       |
| Once weekly (4 doses)  toxicity and  toxicokinetic study  with terminal sacrifice  1 week after the last  dose and with a 4-week  recovery | IV slow bolus (1-3 min) or  30-min infusion via a  saphenous vein                | Monkey/          | 10,             | 30 mg/kg |                |          |           |         |         |       |
| Cynomolgus                                                                                                                                 | (slow bolus) 0, 100 mg/kg  (30-min  infusion) Recovery (0,  30, 100 mg/kg)       | 20068123 GLP     |                 |          |                |          |           |         |         |       |
| Other toxicity studies Tissue cross-reactivity (40 tissues from  3 donors)                                                                 | In                                                                               | vitro            | (cyrosectioned  | Monkey/  | 2 and 10 µg/mL | 20067038 |           |         |         |       |
| tissue)                                                                                                                                    | Cynomolgus                                                                       | GLP              |                 |          |                |          |           |         |         |       |
| Tissue cross-reactivity (40 tissues from  3 donors)                                                                                        | In                                                                               | vitro            | (cyrosectioned  | Human    | 2 and 10 µg/mL | 20074271 |           |         |         |       |
| tissue)                                                                                                                                    | GLP                                                                              |                  |                 |          |                |          |           |         |         |       |
| Blood                                                                                                                                      | compatibility                                                                    |                  |                 |          |                |          |           |         |         |       |
| (hemolysis                                                                                                                                 | and                                                                              |                  |                 |          |                |          |           |         |         |       |
| erythrocyte clumping)                                                                                                                      | In vitro heparinized blood  from Lewis A positive  phenotype on red blood  cells | Human            | Final           | blood    |                |          |           |         |         |       |
| concentrations:  57.5 - 384  µg/mL                                                                                                         | 323966                                                                           |                  |                 |          |                |          |           |         |         |       |
| 1.2.4.7                                                                                                                                    | Species Selection                                                                |                  |                 |          |                |          |           |         |         |       |  
The cynomolgus monkey is considered a relevant biological species for evaluating the safety of MVT-5873, since this species, like humans, expresses the gene (*FUT3*) responsible for expression of the pharmacologic target sLea. In contrast, both mice and rats lack a functional *FUT3* and do not express sLea. To establish the tissue expression of the sLea target in standard toxicology animal models, binding of MVT-5873 to tissue microarrays obtained from cynomolgus monkey, mouse, and rat was studied. As expected from the literature and from previous studies with MVT-5873 using normal human tissue, binding of MVT-5873 to cynomolgus monkey tissue was seen in mucus secreting epithelium in the gastrointestinal tract, as well as in other related tissues (e.g., salivary glands). Unlike cynomolgus monkey tissue microarrays, the tissue microarrays from mouse and rat showed no specific binding for MVT-5873. These results confirm literature reports that rodents do not express sLea and thus are not appropriate for nonclinical safety testing, and that cynomolgus monkeys do express the pharmacologic target and are appropriate for toxicity testing with MVT-5873. 1.2.4.8 Repeat Dose Toxicity
untitled h1
1.2.4.8.1 mvt-5873 2-week once weekly

In a pilot toxicity study in cynomolgus monkeys (nonGLP Study 20068122), MVT-5873 was administered IV as a slow bolus dose at 0 and 30 mg/kg or as a 30-min infusion at 100 mg/kg on study Days 1 and 8. The vehicle was 25 mM histidine, 150 mM sucrose, 55 mM sodium chloride, 0.02% Tween 80 at pH 6.0. Observations were recorded during the study. Samples for hematology, coagulation, urinalysis and clinical chemistry were collected for all animals once prestudy and on Day 23 or, for urinalysis, on Day 25. Animals were necropsied on Day 25, organs weighed and tissue collected for histopathological examination. Toxicokinetic samples were drawn for all animals on Days 1 and 8 at 0.25, 6, 12, 24, 72, and 168 hours post dose. Samples were also collected on Day 23 prior to sacrifice (see Section **1.2.4.8.2**
for discussion of repeat-dose toxicokinetics). There was no mortality, no changes in clinical observations, no changes in food consumption, or adverse reactions to MVT-5873. There were no MVT-5873-related changes in hematology, serum chemistry, urinalysis or coagulation parameters, and no MVT-5873-related macroscopic findings noted at terminal necropsy. Non-adverse microscopic findings considered related to MVT-5873 were noted at 100 mg/kg in the gallbladder (minimal eosinophilic infiltration) in both the male and female monkeys and in the liver (mild Kupffer cell hypertrophy) in the male. Mild mononuclear cell infiltrates noted in the kidney of the female at 100 mg/kg was of uncertain relationship to the test article. These histological changes were not seen in the pivotal GLP 4-week study.
untitled h1
1.2.4.8.2 mvt-5873 4-week once weekly

In the pivotal GLP 4-week toxicity study in cynomolgus monkeys (GLP Study 20068123), MVT-5873 was administered IV as a 30-minute infusion at 0 and 100 mg/kg and as a slow bolus dose at 10 and 30 mg/kg on study Days 1, 8, 15 and 22. The vehicle was the same as the clinical formulation and consisted of 25 mM histidine, 150 mM sucrose, 55 mM sodium chloride, 0.02%
Tween 80, pH 6.0. Observations were recorded during the study. Ophthalmic examinations were conducted predose and during Weeks 4 and 8. Electrocardiograms, heart rate, and waveform intervals (PR, QRS, RR, QT, and QTc Bazett's) were recorded predose and during Weeks 4 and 8. Samples for hematology, coagulation, and clinical chemistry were collected for all animals once predose, on Day 30, and at the end of the 4-week reversal period (Day 58). Samples for assessment of anti-MVT-5873 antibodies were drawn predose, on Day 29, and near the end of the 4-week reversal period (Day 57). Animals were necropsied, organs weighed and tissue collected for histopathological examination on Day 30 for the main sacrifice groups (3/sex/group) and on Day 58 for the reversal groups (2/sex at 0, 30 and 100 mg/kg). Toxicokinetic samples were drawn at various times over a one-week period following the first dose administration, at weekly intervals until the main sacrifice, and then again at the end of the 4-week reversal period. There was no mortality and no MVT-5873 related changes in clinical observations, body weight, food consumption, ophthalmic exams, or cardiac parameters. There were no MVT-5873 related changes in hematology, serum chemistry, or coagulation parameters and no MVT-5873 related macroscopic findings noted at the main necropsy or at the reversal necropsy. In contrast to the previous cynomolgus monkey toxicity study, no MVT-5873 related microscopic findings were observed in this GLP study. The toxicokinetics of MVT-5873 in female and male cynomolgus monkey serum demonstrated similar systemic exposure as in the previous toxicity study, with mean weekly trough values over the 4-week study period being similar to, or greater than, those observed during Week 1. Although MVT-5873, a fully human monoclonal antibody, generated anti-MVT-5873 antibodies in most animals at all dose groups, the presence of these antibodies had no apparent effect on serum levels of MVT-5873 and was not associated with any microscopic changes. Two monkeys tested positive for anti-MVT-5873 antibodies prior to treatment with MVT-5873 and also demonstrated higher serum clearance of MVT-5873 on Day 1 versus monkeys with no predose antibodies; these results indicate that the positive results prestudy were likely cross-reactive antibodies. Development of antibodies during the study was expected following administration of a human monoclonal antibody to cynomolgus monkeys. In summary, intravenous administration of MVT-5873 up to 100 mg/kg once weekly for 4 weeks generated no adverse toxicological effects. The dose of 100 mg/kg is near the maximum feasible dose based on maximum formulation concentration and maximum acceptable dose volume in monkeys. An expected immune reaction was induced in cynomolgus monkeys after treatment with MVT-5873, but had no apparent effect on serum levels of MVT-5873, which generally remained constant throughout the 4-week study period.
untitled h1
1.2.4.9 tissue cross-reactivity studies

The objective of this study was to determine the potential cross-reactivity of MVT-5873, a monoclonal human IgG antibody directed against Sialyl Lewis (a), with cryosections of human tissues. Sialyl Lewis (a) is a carbohydrate antigen present on O-glycans on the surfaces of cells and is often associated with high grade tumors from various tissues.[12, 33] Sialyl Lewis (a) has been reported to be expressed in a number of normal tissues as well.[34-37]
Pivotal (GLP) tissue cross-reactivity studies of MVT-5873 with cryosections (or with fixed blood)
of normal human tissues (GLP Study 20067038) and normal cynomolgus monkey tissues (Study 20074271) were conducted using 41 different tissue types from typically 3 donors/tissue. In addition, positive control (cryosections of human DMS 79 cells), negative control (cryosections of human Jurkat cells), and ancillary control (cryosections of human colon which was previously shown to cross react with MVT-5873) tissues were used to ensure integrity of the staining. As a further control to ensure adequacy of the tissue sample, sections were immunostained for β2-microglobulin, a relatively ubiquitous epitope. To ensure specificity of the MVT-5873 antibody, all tissue sections were concurrently stained with a negative control human IgG1 kappa antibody. The optimal concentration of MVT-5873 was determined by finding the lowest concentration of test article that produced the maximum (plateau) binding to the target antigen (2 µg/mL). This concentration and a 5-fold increase (10 µg/mL), which did not incur excessive nonspecific staining of control samples and/or test tissues, were used for the definitive tissue cross-reactivity studies. MVT-5873 produced moderate to intense membrane, cytoplasm, and cytoplasmic granule staining of the positive control material (cryosections of human DMS 79 cells) at both staining concentrations. MVT-5873 did not specifically react with the negative control material (cryosections of human Jurkat cells) at either staining concentration. The control article, HuIgG1, did not specifically react with either the positive or negative control materials or with the any of the tissue slides. There also was no staining of the assay control slides. MVT-5873 produced moderate to intense staining of membrane, membrane granules, cytoplasm, and cytoplasmic granules in frequent mucosal epithelial cells and weak to strong staining of occasional extracellular material with morphology and location expected for mucus in the ancillary control material (cryosections of human colon) at both staining concentrations. There was no staining of any human colon tissue elements from the ancillary control material with either concentration of HuIgG1 or in the respective assay control slide. The excellent specific reactions of MVT-5873with the positive control material and the lack of specific reactivity with the negative control material, as well as the lack of reactivity of the control article, indicated that the assay was sensitive, specific, and reproducible. The results of human tissue staining with MVT-5873 at 10 and 2 µg/mL and with the control article, HuIgG1, at 10 and 2 µg/mL are presented in GLP Study 20067038 and summarized below.
untitled h1
epithelium

MVT-5873 variably stained membrane, membrane granules, cytoplasm, and/or cytoplasmic granules in epithelium in a variety of human tissues including bladder (transitional), breast, colon (mucosa), eye (conjunctiva), Fallopian tube (mucosa), gallbladder (mucosa), esophagus (squamous, submucosal glands), small intestine (mucosa), stomach (mucosa), kidney (tubular), liver (bile duct), lung (bronchial, bronchiolar), ovary (surface, follicle), pancreas (acinar, ductal), pituitary (adenohypophysis), prostate (glandular, ductal), rectum (mucosa), salivary gland (acinar, ductal), skin (sweat gland, hair follicle, epidermis), testis (seminiferous tubules), thymus, tonsil (squamous, surface, crypt), ureter (transitional), cervix (external ostium [squamous], internal ostium, endocervical glands), and uterus (endometrium, surface, glandular). This staining was equivalent, reduced in intensity and/or frequency, or absent at the lower staining concentration. Sialyl Lewis (a) background expression has been reported in a variety of tissues including pancreas, gallbladder, stomach, colon, bronchial tree, endometrium, salivary glands, kidney and prostate. Based on the wide range of Sialyl Lewis (a) background expression, the observed staining of epithelium in the human tissues was judged expected.[34-37]
untitled h1
mononuclear cells

MVT-5873 weakly to moderately stained cytoplasm and cytoplasmic granules in rare to occasional resident, migrating, and/or infiltrating mononuclear cells in lymph node at the higher staining concentration. This staining was reduced in frequency to rare at the lower staining concentration. No literature was available describing Sialyl Lewis (a) expression by mononuclear cells; thus, the observed staining of this cell type was judged unexpected.
untitled h1
extracellular material

MVT-5873 variably stained extracellular material in a number of tissues including bladder, colon, eye, Fallopian tube, gall bladder, esophagus, small intestine, stomach, kidney, lung, ovary, pancreas, pituitary, placenta, prostate, rectum, salivary gland, testis, thymus, thyroid, tonsil, ureter, cervix, and uterus. This staining was generally equivalent at both staining concentrations. MVT- 5873 stained extracellular material generally had morphology and location consistent with that expected for mucus, serum components, colloid, lens protein, and/or serum components. Sialyl Lewisa is reported to be secreted into the serum in the form of high-molecular mucin-like glycoproteins; thus, the observed staining of extracellular material was judged expected.[12, 33] Mesothelium In one testis sample, MVT-5873 weakly to moderately stained the cytoplasm of occasional mesothelial cells at both staining concentrations. No literature was available describing Sialyl Lewis (a) expression by mesothelium; thus, the observed staining of this cell type was judged unexpected.
untitled h1
summary

Consistent with the literature for expression of sLea, MVT-5873 stained a variety of epithelial cell types and related extracellular mucus-like material in both human and cynomolgus monkey tissues, all consistent with epithelial cell secretions. The structures stained included mucosal/secretory epithelium of the bladder, breast, eye (conjunctiva), Fallopian tube, gastrointestinal tract, kidney tubules, liver (bile duct), lung (bronchial, bronchiolar) ovary, pancreas (acinar ducts), pituitary (adenohypophysis), prostate (ducts), salivary gland (ducts), skin (sweat gland, hair follicle, epidermis), testis (seminiferous tubules), thymus, tonsils, ureter, cervix, and uterus. These epithelial cells were similarly stained in both human and cynomolgus monkey tissues, except for the ovary and testes, which were only stained in human tissue; however, extracellular mucus material in cynomolgus monkey testes did stain positive. Additional staining was seen intracellularly in lymph node of one human and one cynomolgus monkey donor tissue and in the mesothelium of one human teste. Staining of these tissues is considered to be of little-to-no toxicologic relevance as the cytoplasmic compartment is generally thought to be inaccessible to monoclonal antibodies when administered intravenously to animals. The lack of toxicological significance of the epithelial secretory cell staining is supported by the lack of toxicology findings at doses up to 100 mg/kg/week in the GLP 4-week MVT-5873 toxicology study in cynomolgus monkeys.
untitled h1
1.2.5 effects in humans

MVT-5873 is currently being evaluated in a Phase I/II study both as a single agent and in combination with gemcitabine/nab-paclitaxel. Trial MV-0715-CP-001.01 is a Phase 1 Safety and Tolerability Study of Human Monoclonal Antibody 5B1 (MVT-5873) as Monotherapy and with Chemotherapy in Subjects with Pancreatic Cancer or Other CA 19-9 Positive Malignancies. The primary objective for this trail is to determine the safety and maximum tolerated dose of MVT5873 administered as monotherapy, determine safety and maximum tolerated dose of MVT-5873 administered in combination with nab-paclitaxel plus gemcitabine and to determine the pharmacokinetics of MVT-5873. Secondary objectives are to evaluate tumor response rate (based on RECIST 1.1) to MVT-5873 and evaluate the duration of response This study is currently ongoing in the dose escalation phase for both the monotherapy arm and the combination chemotherapy arm is being evaluated in an expansion cohort at an MVT-5873 dose of 0.125 mg/kg. Trial MV-0715-CP-001.01 enrolled the first subject on 17-Feb 2016 and as of 15-Dec 2017 had accrued 32 subjects with pancreatic adenocarcinoma or other CA 19-9 positive malignancies in the single agent study as summarized below. The population of this study consists of adults (males and females), 18 years of age and older who have locally advanced or metastatic pancreatic ductal adenocarcinoma or other CA 19-9 positive malignancies.  
| adenocarcinoma or other CA 19-9 positive malignancies.  Demographics/Baseline Characteristics Single Agent Subjects Male : Female 16 : 16 Median age, (range), years 65 (51-87) ECOG PS 0 : 1, n (%) 7(22) : 25(78) Prior Therapy: No. of systemic therapies  1   2   3   4+  Radiation Surgery   Primary tumor  Metastatic disease   8  5  11  8  9  17  16  2 Primary Disease Site (n)  Pancreas Colon/rectum  Other (unknown primary) 29  1  2 Stage at entry (n)  III 2   IV 30   |
|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|  
MVT-5873 was administered on an every-other-week schedule (Q2W) (Cohorts A1 and A2) and  
 then modified to a weekly schedule (QW) based dose limiting toxicity at 3 mg/kg and on initial PK data. The QW dose escalation schema is shown below and further summarized in the subsequent table.  
| Parameter          | Cohort Identifier A1 A2 A4    | A5      | A6    | A7   |    |     |
|--------------------|-------------------------------|---------|-------|------|----|-----|
| Number of subjects | 6                             | 3       | 4     | 3    | 6  | 4   |
| Dose (mg/kg)       | 1                             | 3       | 1     | 2    | 3  | 2.5 |
| Schedule           | Q2W                           | Q2W     | QW    | QW   | QW | QW  |
| Dose               | intensity                     | (weekly | mg/kg |      |    |     |
| equivalent)        | 0.5                           | 1.5     | 1     | 2    | 3  | 2.5 |
| 1.2.5.1            | Summary of Safety Information |         |       |      |    |     |  
As of December 15, 2017, the single agent MTD for every other week (Q2W) and weekly (QW) schedules were defined as 1 mg/kg with dose limiting liver toxicity observed at 3 mg/kg for Q2W and 2 mg/kg for QW. The single agent weekly schedule has explored MVT-5873 doses ranging from 1 to 3.0 mg/kg (1.0, 2.0, 2.5 and 3.0 mg/kg). Based on the occurrence of dose limiting liver toxicity characterized by increased transaminases (AST/ALT) and blood bilirubin (Grade 3 and single Grade 4), MVT-5873 doses ≥ 2.0 mg/kg were considered to have exceeded the maximum tolerated dose (MTD). The MTD for single agent MVT-5873 was determined to be 1 mg/kg. However, based on clinical data single-agent MVT-5873 at a dose of 3 mg/kg appears to demonstrate dose limiting toxicity with increases in liver transaminases and blood bilirubin. When seen, these abnormalities occurred within a few days of drug administration, were self-limited and resolved spontaneously within eight days and resulted in no permanent liver abnormalities. Moreover, dosing >1.0 mg/kg appeared to be a more effective dose in terms of RECIST responses than 1.0 mg/kg, and subjects on doses >1.0 mg/kg received more doses before PD was documented. A secondary aim of Trial MV-0715-CP-001.01 was to determine the mechanism(s) of this DLT, which may be related to Kupffer cell activation and not necessarily any direct injury related to the antibody. Non-human primate data support this hypothesis.  
  
  
Patient \#1
untitled h1
patient #3

Patient \#6  
 As a single agent, severe (Grade 3+) toxicities considered related to MVT-5873 have predominately included reversible liver function abnormalities (manifest as increased transaminases and blood bilirubin), and infusion reactions. Infusion reactions decreased in frequency and severity following initiation of a prophylactic regimen (slower infusion rate and IV steroids). Low-grade (Grades 1 and 2) treatment-related toxicities consist predominately of toxicities related to immune activation (chills, fever, malaise, pruritus, rash, and skin induration). No significant related hematologic toxicity has been observed. To date, no significant EKG changes (e.g., increases in QTcF) have been observed. In summary, the predominate toxicity of MVT-5873 as a single agent and/or in combination with gemcitabine/nab-paclitaxel is dose limiting liver function test abnormalities. Increases in liver function tests are generally seen early (Day 3) and are reversible. Infusion reactions, when seen, generally appear during the infusion or shortly thereafter and appear less likely with slower infusion rates or with the use of steroid premedication. A summary of the safety profile and adverse events are summarized below.  
| Table 3: Dose Cohorts, Cycles Administered and Patient Disposition  Baseline CA19-9 (U/mL) Patient Disposition  Cohort  < 2500 ≥ 2500 Off Study On  Study n Median Cycles  (range) n Median Cycles  (range) Toxicity PD   | Other   |                    |    |               |    |    |    |    |
|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|---------|--------------------|----|---------------|----|----|----|----|
| A1                                                                                                                                                                                                                        | 1       | 4                  | 5  | 1.25 (1-3)    | 0  | 0  | 5  | 1  |
| A2                                                                                                                                                                                                                        | 2       | 4.5 (4-5)          | 1  | 7.5           | 0  | 0  | 2  | 1  |
| A4                                                                                                                                                                                                                        | 4       | 5.625 (1.5-13.5)   | 2  | 2 (1.25-2.75) | 0  | 0  | 5  | 1  |
| A5                                                                                                                                                                                                                        | 5       | 2 (1.5-6.25)       | 1  | 2             | 0  | 0  | 6  | 0  |
| A6                                                                                                                                                                                                                        | 0       | -                  | 6  | 1.375 (1-2)   | 0  | 0  | 6  | 0  |
| A7                                                                                                                                                                                                                        | 4       | 4.375 (0.25-10.75) | 1  | 1             | 1  | 1  | 3  | 0  |
| Total                                                                                                                                                                                                                     | 16      | 4 (0.25-10.75)     | 16 | 1.375 (1-7.5) | 1  | 1  | 27 | 3  |
| B3                                                                                                                                                                                                                        | 2       | 0.875 (0.75 - 1.0) | 1  | 1             | 0  | 0  | 1  | 2  |
| B0                                                                                                                                                                                                                        | 3       | 2.0 (0.5-6.75)     | 1  | 3.5           | 2  | 0  | 1  | 1  |
| Total                                                                                                                                                                                                                     | 5       | 1 (0.5-6.75)       | 2  | 2.25 (1-3.5)  | 2  | 0  | 2  | 3  |  
| Table 4: Summary of MVT-5873 Dose Limiting Toxicities and Treatment Disposition Cohort (n) Individual Patient Events (Cycle 1) Treatment Disposition CTCAE Day Toxicity Duration Total (days) Adjustment Grade Cycles A1 (6) None  3 ↑ Bilirubin 3 12 ↓ dose 4  A2 (3)  3 ↑ ALT 3 2 ↓ dose 5  A4 (6) None  8 ↑ Bilirubin 3 7 ↓ dose 1.5  A5 (6)  3 ↑ ALT,↑ AST 3, 3 3,3 ↓ dose 1.25*  5 ↑ ALT,↑ AST 3, 3 3, 3 ↓ dose (x2) 1.25  A6 (6)  3 ↑ AST 3 6 ↓ dose 1.5  3 ↑ ALT,↑ AST 3, 3 5,5 d/c 0.25# A7 (5)  29 ↑ ALT,↑ AST 3, 3 2,2 ↓ dose 3.25*  3 ↑ ALT,↑ AST 3, 4 3,5 ↓ dose 3*  8 ↑ ALT 3 7 delay 0.75  B3 (3)  3 ↑ Bilirubin 3 5 delay 1  *Continued on treatment; # discontinued (d/c) after 1st dose, resolution to Grade 2 by Day 5   |
|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|  
The adverse event experience for MVT-5873 as a single agent is listed in the **Appendix B** and summarized below in the accompanying **Table 5** and **Table 6**.  
| Table 5: All Adverse Events, All Grades, in ≥ 30% of Subjects Cohort At MTD Cohort Exceeds MTD Preferred Term n (%) A1 A4 A2 A5 A6   | A7      |         |         |         |         |    |    |
|--------------------------------------------------------------------------------------------------------------------------------------|---------|---------|---------|---------|---------|----|----|
| (n = 6)                                                                                                                              | (n = 6) | (n = 3) | (n = 6) | (n = 6) | (n = 5) |    |    |
| ↑ AST                                                                                                                                | 23 (72) | -       | 2       | -       | 5       | 5  | 11 |
| ↑ ALT                                                                                                                                | 18 (56) | -       | 1       | 1       | 4       | 2  | 10 |
| ↑ Alkaline phosphatase                                                                                                               | 22 (69) | 4       | 4       | -       | 2       | 4  | 8  |
| ↑ Bilirubin                                                                                                                          | 20 (63) | 3       | 1       | -       | 9       | 1  | 6  |
| Fatigue                                                                                                                              | 19 (59) | 4       | 4       | 1       | 1       | 5  | 4  |
| Nausea                                                                                                                               | 13 (41) | 3       | 1       | 2       | 5       | -  | 2  |
| Hypoalbuminemia                                                                                                                      | 12 (38) | 1       | 2       | -       | -       | 1  | 8  |
| Hyperglycemia                                                                                                                        | 10 (31) | 4       | 2       | -       | 2       | -  | 2  |
| Vomiting                                                                                                                             | 10 (31) | 4       | 1       | 3       | 1       | 1  | -  |
| Back Pain                                                                                                                            | 10 (31) | 1       | -       | 1       | 2       | 6  | -  |
| Anemia                                                                                                                               | 10 (31) | 1       | 2       | -       | -       | -  | 7  |
| Pneumonitis                                                                                                                          | -       | -       | -       | -       | -       | -  | -  |
| Total* (n = 32)                                                                                                                      |         |         |         |         |         |    |    |  
| Table 6: All Treatment-Related Adverse Events ≥ Grade 3  Cohort At MTD Cohort Exceeds MTD Preferred Term n (%) A1 A4 A2 A5 A6   | A7      |         |         |         |         |    |    |
|---------------------------------------------------------------------------------------------------------------------------------|---------|---------|---------|---------|---------|----|----|
| (n = 6)                                                                                                                         | (n = 6) | (n = 3) | (n = 6) | (n = 6) | (n = 5) |    |    |
| ↑ ALT                                                                                                                           | 9 (30)  | -       | -       | 1       | 2       | 1  | 5  |
| ↑ AST                                                                                                                           | 8 (27)  | -       | -       | -       | 2       | 2  | 4  |
| ↑ Alkaline phosphatase                                                                                                          | 5 (17)  | -       | 1       | -       | 1       | -  | 3  |
| ↑ Bilirubin                                                                                                                     | 4 (13)  | 1       | -       | -       | 3       | -  | -  |
| Pneumonitis                                                                                                                     | -       | -       | -       | -       | -       | -  | -  |
| ↓ Lymphocyte count                                                                                                              | 2 (7)   | -       | -       | -       | 1       | -  | 1  |
| ↑ Lipase                                                                                                                        | 1 (3)   | -       | -       | -       | 1       | -  | -  |
| Hyperbilirubinemia                                                                                                              | 1 (3)   | -       | -       | 1       | -       | -  | -  |
| Infusion reaction                                                                                                               | 1 (3)   | 1       | -       | -       | -       | -  | -  |
| *Total adverse events recorded in all subjects across all cycles; # data as of 08/30/2017                                       |         |         |         |         |         |    |    |
untitled h1
1.2.6 pharmacokinetics

MVT-5873 displays a half-life of 117.6 ± 33.4 h, clearance of 279.6 ± 313.0 mL/h, and volume of distribution at steady-state of 17.7 ± 21.0 L. These data demonstrate that MVT-5873 displays properties that are in the typical range for monoclonal antibodies. These initial PK values for the two different schedules of administration using a two-compartment model are summarized in the Table 7.  
Regimen Subject **Initial t1/2**  
(h)  
Terminal t1/2 (h)  
AUC0-∞  
(ng•h/mL) Vss (mL) **Cl (ml/h)**  
Q2W Mean 20.8 193.0 2339353 17147 226.3  
SD 9.5 93.4 3256439 27582 290.1  
QW Mean 37.3 833.1 2812486 14922 65.1  
SD 14.1 1164.4 3247729 12999 68.9
untitled h1
1.2.7 reductions in serum ca19-9

In study MV-0715-CP-001.01, serum CA 19-9 levels are assessed both pre- and post-MVT-5873 administration. Data on serum CA 19-9 levels indicate that serum CA 19-9 levels are uniformly reduced in all subjects regardless of dose. However, the probability of a CA 19-9 level within normal limits (i.e., < 37 U/mL) post MVT-5873 appears to be related to dose. As shown in the table below, at a dose of 1 mg/kg, MVT-5873 appears to normalize an elevated CA 19-9 level in about 50% of subjects on Cycle 1 Day 1 and in about 20% of subjects across all administration days. In contrast, doses of 2 mg/kg or higher normalize CA 19-9 levels in essentially all subjects.  
Dose MVT-5873  
(mg/kg)
normal
1.2.7 reductions in serum ca19-9

Post C1D1 * Percent **# Normal**  
Post All Days** **Percent**  
1 3/6 50% 4/20 20% 2 3/3 100% 10/10 100% 3 1/1 100% 12/13 92%  
TOTAL 7/10 70% 26/43 60%  
*Number of subjects with CA 19-9 level > 37U/mL pre MVT-5873 that was reduced to < 37U/mL post MVT-5873 on Cycle 1 Day 1 only.  
Table 8: Normalization (< 37U/mL) of CA 19-9 Levels Following Adminstration of MVT-5873
**Number of subjects with CA 19-9 level > 37U/mL pre MVT-5873 that was reduced to < 37U/mL post MVT-5873 for all administration days.
normal
1.2.8 responses to therapy

Upon review of the single agent dosing cohorts, dosing > 1 mg/kg appears to be a more effective dose based on RECIST responses than 1 mg/kg. Moreover, subjects on doses > 1 mg/kg remained on therapy longer before PD was documented indicating longer duration of response. Most importantly however the data clearly indicate efficacy for MVT-5873 when given as a single agent, and the subjects that experienced the most efficacy were those with CA 19-9 values less than 2,500 U/ml (**Figure 8**).  
inge from pretreatment (Day 1) CA19-9 level  
  
Cycles of therapy administered
normal
figure 8 efficacy for mvt-5873

The preliminary response data for MVT-5873 in combination with a standard of care chemotherapy are promising with 5 PR and 1 SD as best response. Figure 9 illustrates the percent change in target lesion for the first six subjects at a dose of 0.125 mg/kg (Cohort B3).
normal
baseline ca19-9 # < 2500

Figure 9: Percent Change in Target Lesions in Cohort B3 (0.125 mg/kg MVT-5873)
normal
1.2.9 rationale

In this study we are planning to give subjects a pre-operative, one-time infusion of 1 mg/kg of MVT-5873 at Day -3 before surgery, perform the resection and continue treatment with MVT5873 at the dose of 1 mg/kg. We believe that peri-operative use of MVT-5873 adds "systemic" treatment during a time when none is given, despite the fact that the resection is associated with large increases in circulating tumors cells, and an inflammatory milieu believed to be, at least in part, responsible for the early recurrences observed in a substantial number of subjects. We believe peri-operative MVT-5873 will decrease early recurrence rates and increase DFS for subjects with CA 19-9 producing cancers included in this study. To assess safety of the combination of treatment with MVT-5873 and surgery, we are planning to start with enrollment into safety lead-in cohorts and carefully monitor status of the subjects during 10 days after initial treatment and surgery. We will continue enrollment into the study only if treatment in the safety lead-in cohorts are proven to be safe. If safety is documented with the safety-lead in cohorts at 1 mg/kg, we will dose escalate the single pre-operative dose of MVT-5873 to 3 mg/kg. We want to explore this higher dose for pre-operative treatment because we consider the drug to be more effective at this dose, and we believe the days following the operation are the most critical time to for the reasons outlined above. We do not believe there will be undo risk as subjects will receive the 3 mg/kg dose once.
normal
1.2.10 metastasis assays using ex

A major goal of Dr. Hernandez's lab is to understand how tumor cells adapt to and co-opt the liver microenvironment for eventual metastatic outgrowth. With tumor cells labelled and observable throughout week-long time courses, many downstream applications will be possible, including (but not limited to) interrogation of the tumor cells at various time-points and at various stages of the metastatic cascade, with and without forced over-expression of various genes of interest. In order to accomplish this, cholangiocarcinoma and pancreatic cancer tissue removed during pancreatic resections will be submitted to Dr. Hernandez's laboratory. Tumor cells will be immediately dissociated into single cells using the Miltenyi Tumor Dissociator and labelled using fluorescent protein tags (Cell tracking Dyes). Labelled single tumor cells will then be injected back into the perfusate to simulate the metastatic process. The cells will be tracked using specialized camera/detector system in combination with our multi-photon microscope.
normal
1.2.11 ex vivo hepatic perfusion

This study is an important step toward our goal of implementing an ideal tumor model system. The ability to recapitulate the complexities of solid human tumors for the purposes of drug development and testing has been, and remains, a major obstacle in the progress of cancer care. Despite great efforts expended on pre-clinical optimization using existing validation models, most drugs simply fail to demonstrate efficacy when subjected to Phase III clinical trial scrutiny. Our interpretation is that the currently available model systems lack the appropriate clinical predictive power. The reasons for the inadequacies of these model systems, which are largely based upon cell lines, mouse models or patient-derived xenografts, are myriad but are almost certainly related to the absence of the human stromal component and its intricate relationship with tumor cells. Indeed, modeling the multi-faceted interactions between human cancer cells and the multitude of various stromal components, including activated fibroblasts, immune cell infiltrate and the abnormal vasculature is, at present, an impossibility. The advent of a model system capable of accurately reproducing human tumors *ex vivo* carries broad implications across many fields of investigational medicine and brings with it the potential to fundamentally alter the translational research landscape. Moreover, if it were possible to represent a given patient's tumor in that model system, the benefits of personalized medicine could leap from generating lists of potentially useful drugs to designation of efficacious agents. We intend to satisfy all the requirements of an ideal tumor model system by utilizing resected segments of tumor-containing liver and repurposing the Liver-Assist perfusion device for prolonged ex *vivo* animation via native blood vessel perfusion. This is entirely novel and has not been attempted elsewhere in the world. Moreover, the expertise required to accomplish this make it unlikely to be successful anywhere other than the clinical center. This study expands our CRC patient population and importantly expands it to include subjects with intrahepatic cholangiocarcinoma. We will start the model with optimization of perfusion conditions for maintenance of both tumor and normal liver with serial evaluation to include the following:
1) Ultrasonographic evaluation to document preservation of tumor echogenic properties. 2) Core needle biopsy to document histopathologic preservation of the microscopic architecture and cellular viability. Sufficient tissue will be obtained for any necessary staining. 3) Transcriptomic profiling will be undertaken using snap-frozen biopsies. Samples from a given tumor will be run together to minimize variability. Standard HiSeq RNA platform with a read depth of 30-40 million will be employed. 4) Metabolomic profiling will be undertaken using snap-frozen biopsies. Samples from a given tumor will be run together to minimize variability.  
We've chosen strategically to use Genomic Expression's OneRNA platform in the present protocol
(refer to Section **5.2.2**). This technology is designated to quickly sequence 20K genes and provide lists of potentially usable drugs in addition to the sequencing data from tumor and normal liver.  
The platform will be of use to us as we move forward with our ex *vivo* tumor model, as we anticipate patterns will emerge among tumors such that likely drug combinations can be estimated for use in our ex *vivo* model. We will facilitate transfer of material from any pre-operative biopsies/tumor removal to genomic expressions for evaluation. This will allow us to utilize various agents predicted by OneRNA as efficacious. This model system will enable us to evaluate drug delivery, cell based therapies, etc. given the unlimited use of repeat biopsies. The perfusate is also the ideal medium for biomarker discovery, which can be related to actual changes demonstrable in the tumor. Although we will not be able to use efficacious drugs in the model to treat subjects, we believe the information will be of immediate value in guiding subjects toward subsequent trials should they recur. Finally, we will be able to conduct metastases assays using the ex vivo tumor containing liver as the target organ and the tumor dissociator as above (see Section **1.2.10**).
normal
1.2.12 simulated metastasis assay with

Dr. Hernandez's laboratory in the Surgical Oncology Program is also developing a novel tumor modeling system, referred to as the Simulated Metastasis Assay with Relevant Tissue (SMART) Chamber. Correlative studies in proteogenomic characterization of primary and metastatic cancers and *ex vivo* tumor tissue modeling of peritoneal metastasis will be added to this study, utilizing the SMART Chamber. In conjunction with ongoing laboratory work, segments of the peritoneum are removed from patients with peritoneal carcinomatosis at the time of operation. This peritoneal tissue is utilized in the SMART Chamber system to imitate conditions of metastasis within the abdominal cavity. This *ex vivo* tissue model system allows for observation of peritoneal metastasis, real-time cellular imaging, and manipulation of tumor-environment interactions.
normal
1.2.13 the smart system

The SMART System is an important step toward our goal of realizing personalized translational cancer care. The ability to recapitulate the complexities of solid human tumors for the purposes of drug development and testing has been, and remains, a major obstacle in the progress of cancer therapy. Despite great efforts expended on pre-clinical optimization using existing validation models, most drugs simply fail to demonstrate efficacy when subjected to Phase III clinical trial scrutiny. Our interpretation is that the currently available model systems lack the appropriate clinical predictive power. The reasons for the inadequacies of these model systems, which are largely based upon cell lines, mouse models or patient-derived xenografts, are myriad but are almost certainly related to the absence of the human stromal component and its intricate relationship with tumor cells. There has been a sizable expansion of oncologic pharmaceuticals that have targeted and/or  
 immunomodulatory effects. However, the predictive accuracy of drug testing is predicated on models that accurately represent the architectural/cellular organization of the tumor and faithfully reflect the *in vivo* biology. We have satisfied many of the requirements of an ideal tumor model utilizing resected mesothelial metastases to create the SMART (Sample Microenvironment of Resected Metastatic Tumor) System. Multiple solid tumors commonly manifest with metastatic disease to the mesothelial surface (peritoneum, pleura, liver capsule) and some are candidates for metastasectomy. During these operations, resected tumor-bearing mesothelial tissue is affixed to a specialized platform and perfused in a sterile, oxygenated circuit at 370C to maintain normal physiologic parameters (**Figure 10**). Circulating oxygenated perfusate within the system is comprised of human plasma (from the patient), DMEM media, antibiotics, insulin (slow infusion).  
The SMART platform (Top Left) is designed to mount the human mesothelial tissue containing tumor and be suspended in the reservoir of perfusate (Bottom Left), the SMART chamber. The chambers suspend maximum of 4 SMART platforms and placed on stir plate in the incubator (Middle). The SMART system boasts a physiologic oxygenated perfusion circuit that is suitable to keep tissue viable for up to four days. The Right Panel demonstrates the customized holder for each SMART platform that is imaged using complex imaging techniques.  
We have evaluated gastric adenocarcinoma, pancreatic adenocarcinoma, gastrointestinal stromal tumor, and breast adenocarcinoma in the SMART system and documented ex *vivo* viability at 4 days with expected immune cell populations, intact architecture, and expected mitotic activity
(**Figure 11A**). RNA sequencing was performed as well to compare transcriptomics between Day 0 and Day 4. Genes associated with hypoxia, oncogenes/tumor suppressor, cancer stem cells,  
 metabolism, and epithelial-mesenchymal transition were isolated from the sequencing data and demonstrated conservation of transcriptomics (**Figure 11B**). These results ultimately demonstrate that tumor nodules in their native, functional microenvironment can be preserved *ex vivo* for 4 days with fidelity.  
Figure 11: Ex Vivo Normal and Tumor Preservation in the SMART System
A (Left): Demonstration of various histologies on H&E, CD3, CD68 and Ki67 stains on Day 0 and Day 4. B (Right): Conservation of transcriptomics in hypoxic panel, oncogene/tumor suppressor, cancer stem cell, metabolism and EMT panels over breast, gastric, pancreatic and mesothelioma histology.  
Due to the translucent nature of the mesothelial tissue, the system is readily amenable to dynamic interrogation with confocal, two photon and light sheet microscopy. This avenue of investigation is supported by a research collaborative agreement (RCA) with BioLegend and our affiliation with CAT-I to develop fluorescently labelled Fab fragment for live cell labelling. For example, with the increasing use of checkpoint inhibitors like Pembrolizumab in solid tumors, we decided to evaluate the drug in the SMART system using tumor from a patient with gastric adenocarcinoma and a patient with colorectal adenocarcinoma. We demonstrated immune cell engagement with tumors cells for both gastric and colorectal adenocarcinoma, and captured tumor cell lysis with the latter
(**Figure 12**). Such responses at the cellular level are key to understanding why some cells respond to the immune system while others are ostensibly resistant. As the only currently known platform that has an intact, immunocompotent tumor microenvironment, the SMART System allows for the analysis of agents utilizing real controls to increase current understandings of the tumor microenvironment. Analyses can be readily imaged (taking video up to an hour) to obtain tangible visualizations of the tested agent effects (see Figure 12).  
  
Figure 12: SMART Platforms are Valuable Tools for Drug Evaluations Such as Pembrolizumab Left: SMART platforms with peritoneum from a patient with gastric cancer incubated with immune checkpoint inhibitor Pembrolizumab for 2 days followed by Alexa488CD44/
Alexa594CD45 antibodies. Maximum projections of 2-photon images illustrate the intimate contact between CD45-positive cells and the cancer cells. Right: Timeseries montage taken from liver capsule tissue mounted on SMART platforms from a colorectal cancer patient with liver metastases incubated with 5 g/mL pembrolizumab for 4 hours. CD45-positive cells bound to CD44-positive cancer cell undergoing apoptosis.
normal
2 eligibility assessment and enrollment

2.1.1 **Inclusion Criteria** 2.1.1.1 Subjects must have histologically or cytologically confirmed diagnoses of adenocarcinoma in one of the following scenarios:
o Primary tumors of the pancreas o Primary tumors of the bile duct and ampulla o Metastatic colorectal cancers to the liver 2.1.1.2 Subjects must have disease resectable with a standard pancreatectomy
(pancreaticoduodenectomy or distal pancreatectomy) or liver resection.  
2.1.1.3 Subjects may have received prior therapy, including neoadjuvant regimens. 2.1.1.4 Subjects must have serum CA 19-9 elevations greater than the upper limit of normal but less than 2500 U/mL.  
2.1.1.5 Age ≥ 18 years.  
2.1.1.6 ECOG performance status ≤ 1 (**Appendix A**).  
2.1.1.7 Subjects must have adequate organ and marrow function as defined below:
o leukocytes > 3,000/mcL
o absolute neutrophil count > 1,500/mcL o platelets > 90,000/mcL
2.1.1.8 For subjects with Periampullary cancers that require a pancreaticoduodenectomy for complete tumor extirpation:  
o total bilirubin < 10 ULN*
o AST(SGOT)/ALT(SGPT) < 5 X institutional upper limit of normal o creatinine < 1.5 X institutional upper limit of normal
*Subjects with periampullary cancers typically present with biliary obstruction resulting in significant abnormalities in liver function tests that do not reflect liver dysfunction. These values normalize after tumor removal. They can be normalized preoperatively with biliary stenting butseveral large studies have demonstrated an increase in infectious complications with drainage. As such, a practice standard has been to avoid stenting until bilirubin level rises above 10 X ULN.  
2.1.1.9 For subjects with liver tumors (cholangiocarcinoma or metastatic colorectal cancer)
requiring a hepatectomy for complete tumor extirpation:  
o total bilirubin < 2.5 X institutional upper limit of normal* o AST(SGOT)/ALT(SGPT) < 5 X institutional upper limit of normal*
o creatinine < 1.5 X institutional upper limit of normal
*Liver abnormalities in this range are consistent with parenchymal destruction from the tumor.  
2.1.1.10 For subjects with pancreas tumors that require a distal pancreatectomy for extirpation:  
o total bilirubin < 1.5 X institutional upper limit of normal* o AST(SGOT)/ALT(SGPT) < 2 X institutional upper limit of normal*
o creatinine < 1.5 X institutional upper limit of normal
*Liver abnormalities in this range are consistent with pancreas cancer destruction from the tumor.  
2.1.1.11 The effects of MVT-5873 on the developing human fetus are unknown. For this reason, women of child-bearing potential and men must agree to use adequate contraception (hormonal or barrier method of birth control; abstinence) prior to study entry and for the duration of study participation and for 3 months after completion of study treatment. Should a woman become pregnant or suspect she is pregnant while she or her partner is participating in this study, she should inform her treating physician immediately.  
2.1.1.12 Ability of subject to understand and the willingness to sign a written informed consent document.  
2.1.1.13 Subjects must agree to co-enrollment on the tissue collection protocol 13C0176,
"Tumor, Normal Tissue and Specimens from Patients Undergoing Evaluation or Surgical Resection of Solid Tumors".  
2.1.2 **Exclusion Criteria**
2.1.2.1 Presence of disease outside the confines of a standard operation for subjects with periampullary cancers (pancreatic and cholangiocarcinoma).  
2.1.2.2 Presence of disease outside the liver for subjects with intrahepatic/hilar cholangiocarcinoma or metastatic colorectal cancer, other than a primary tumor for subjects with metastatic colorectal cancer.  
2.1.2.3 Subjects who are receiving any other investigational agents. 2.1.2.4 Fewer than 28 days (or 5 half-lives for systemic agents, whichever is shorter) from the last day of prior anticancer therapy, including chemotherapy, hormonal, investigational, and or biological therapies and irradiation.  
2.1.2.5 Uncontrolled inter-current illness including, but not limited to, ongoing or active infection, symptomatic congestive heart failure, unstable angina pectoris, cardiac arrhythmia, or psychiatric illness/social situations that would limit compliance with study requirements.  
2.1.2.6 Active concurrent malignancies within the last five years other than the primary tumor in subjects with metastatic colorectal cancer, basal or squamous cell skin carcinoma or non- medullary thyroid carcinoma.  
2.1.2.7 Pregnant women are excluded from this study because of the potential for teratogenic or abortifacient effects of the MVT-5873. Because there is an unknown but potential risk for adverse events in nursing infants secondary to treatment of the mother, breastfeeding should be discontinued if the mother is treated. These potential risks may also apply to other agents used in this study.  
2.1.2.8 Subjects with active, Hepatitis B or C infection because of the potential for increased liver toxicity given the damaging effects of the virus.  
2.1.2.9 Allergic to chimeric, humanized or human antibodies. 2.1.2.10 Received live vaccine within 4 weeks prior to the first date of study intervention. 2.1.2.11 Infection requiring hospitalization or herpes zoster treatment within 2 weeks prior to the first date of study intervention.  
2.1.2.12 Long-term infectious diseases (tuberculosis, fungal infections) active within 2 years prior to the first date of study intervention.
normal
2.1.3 recruitment strategies

This protocol may be abstracted into a plain language announcement posted on NIH websites and on NIH social media platforms. This study will also be listed on available websites (e.g., www.clinicaltrials.gov) and participants will be recruited from the current patient population at NIH. Participants may also be recruited through self-referrals and physician referrals. Dr. Hernandez is a member of the clinical trials committee of America's HepatoPancreatoBiliary Association and will also solicit eligible subjects through the organization. Advertisements approved under this protocol may be distributed in local media (electronic and print) with recruitment efforts focused on, but not limited to, NIH and the greater Washington DC area; some recruitment efforts may take place nationwide.
normal
screening evaluation 2.2.1 screening activities

Minimal risk activities that may be performed before the subject has signed a consent include the following:  
- Email, written, in person or telephone communications with prospective subjects.  
- Review of existing medical records to include H&P, laboratory studies, etc. - Review of existing MRI, x-ray, or CT images. - Review of existing photographs or videos.  
- Review of existing pathology specimens/reports from a specimen obtained for diagnostic purposes.
A waiver of consent for these activities has been requested in Section **12.5.1**.
normal
2.2.2 screening activities performed after

The following activities will be performed only after the subject has signed the study consent OR the consent for study 01C0129 (provided the procedure is permitted on that study) on which screening activities may also be performed. Assessments performed at outside facilities or on another NIH protocol within the timeframes below may also be used to determine eligibility once a patient has signed the consent. Studies should be done within 6 weeks prior to being registered for treatment, unless otherwise noted below:
- History and Physical Examination:  
o Complete medical history and physical examination (including height, weight, vital signs, EKG and ECOG performance status).
- Laboratory Evaluation:
o CA 19-9 levels o Hematological profile: CBC with differential and platelet count.  
o Biochemical profile: electrolytes, BUN, creatinine, AST, ALT, total bilirubin, alkaline phosphatase, calcium, phosphorus, albumin, magnesium, amylase.  
o Hep B and C  
o Serum or urine pregnancy test for female participants of childbearing age (in the absence of prior hysterectomy), performed within 7 days prior to treatment.  
- CT of chest, abdomen and pelvis. (Note: This scan does not need to be repeated at NIH if it has already been done at an external institute within 6 weeks prior to treatment and the investigator determines that the scan is of sufficient detail for study diagnostics.)
- Histologic confirmation by the Laboratory of Pathology, NIH (at any time point prior to enrollment). If there is no available tumor sample, biopsy will be performed to confirm the diagnosis.  
- MRI (for subjects with liver disease). (Note: This scan does not need to be repeated at NIH
if it has already been done at an external institute within 6 weeks prior to treatment and the investigator determines that the scan is of sufficient detail for study diagnostics.)
normal
participant registration and status update

Registration and status updates (e.g., when a participant is taken off protocol therapy and when a participant is taken off-study) will take place per CCR SOP ADCR-2: CCR Participant Registration & Status Updates found here.  
2.3.1 **Treatment Assignment and Randomization/Stratification Procedures** 2.3.1.1 Cohorts  
| 2.3.1.1   | Cohorts        |                                                                                                                                                                                                                                                                                               |
|-----------|----------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Number    | Name           | Description                                                                                                                                                                                                                                                                                   |
| 1         | Cohort 1       | Subjects with pancreatic cancer, cholangiocarcinoma or metastatic to the liver colorectal cancer, having low elevated LFTs (AST/ALT  ≤ 2 X ULN and/or bilirubin ≤ 1.5 X ULN), enrolled into preoperative MVT-5873 escalation dose levels.                                                                                                                                                                                                                                                                                               |
| 2         | Cohort 2       | Subjects with pancreatic cancer, cholangiocarcinoma or metastatic  to the liver colorectal cancer, having moderate elevated LFTs (LFTs  above what is required for Cohort 1 AND AST/ALT < 5 X ULN  and/or bilirubin ≤ 3 X ULN), enrolled into pre-operative MVT-5873  escalation dose levels. |
| 3         | Cohort 3       | Subjects with pancreatic cancer enrolled at the pre-operative  recommended dose (RD) of MVT-5873.                                                                                                                                                                                             |
| 4         | Cohort 4       | Subjects with cholangiocarcinoma enrolled at the pre-operative  recommended dose (RD) of MVT-5873.                                                                                                                                                                                            |
| 5         | Cohort 5       | Subjects with metastatic to the liver colorectal cancer enrolled at the  pre-operative recommended dose (RD) of MVT-5873.                                                                                                                                                                     |
| 2.3.1.2   | Arms           |                                                                                                                                                                                                                                                                                               |
| Number    | Name           | Description                                                                                                                                                                                                                                                                                   |
| 1         | Arm 1          | Pre-operative escalation doses of MVT-5873, pancreatectomy or  hepatectomy and post-operative MVT-5873 treatment.                                                                                                                                                                             |
| 2         | Arm 2          | Pre-operative RD of MVT-5873, pancreatectomy or hepatectomy and post-operative MVT-5873 treatment.                                                                                                                                                                                            |
| 2.3.1.3   | Arm Assignment |                                                                                                                                                                                                                                                                                               |  
2.3.1.3 Arm Assignment Subjects in Cohorts 1 and 2 will be directly assigned to Arm 1.  
Subjects in Cohorts 3-5 will be directly assigned to Arm 2.
normal
baseline evaluation

Tests listed below that were performed within the appropriate timeframe at screening need not be repeated. Within 3 weeks prior to study intervention unless otherwise indicated:  
- Complete physical examination (including weight, vital signs, EKG and ECOG
performance status).  
- Laboratory Evaluation:
o CA 19-9 levels o Hematological profile: CBC with differential and platelet count. o Biochemical profile: electrolytes, BUN, creatinine, AST, ALT, total bilirubin, alkaline phosphatase, calcium, phosphorus, albumin, magnesium, amylase.  
o PT/PTT/INR o Serum or urine pregnancy test for female participants of potential childbearing age (in the absence of prior hysterectomy), performed within 3 days prior to resection.  
- CT of chest, abdomen and pelvis
- MRI (for subjects with liver disease) - Concomitant medications
- Baseline adverse events
normal
3 study implementation study design

This is a trial designed to determine if perioperative MVT-5873 is safe and effective for subjects with operable cancers of the pancreas and liver. Eligible candidates will receive one dose of MVT-5873 (1 mg/kg (DL1) or 3 mg/kg (DL2) 3 days prior to resection (Day -3) as part of the safety lead-in cohorts (Cohorts 1 and 2). See **Table 9**
below for expected course. Once the pre-operative safe dose of MVT-5873 is identified, additional subjects will be evaluated within the treatment cohorts (Cohorts 3-5) at that MTD. Resection will be undertaken to remove all demonstrable disease in the liver, bile ducts and pancreas on Day 0. The operation may be delayed for up to 4 days until any AST/ALT elevations are ≤ 10 X ULN, and Bilirubin is ≤ 2 X the baseline value or < 3 X ULN. All subjects will start post-operative infusions of MVT-5873 (1 mg/kg) in cycles consisting of 21 days. During Cycle 1, subjects will receive 2 weekly infusions of MVT-5873 (on Day 1 and Day 8). During Cycle 2, subjects will receive an infusion of MVT-5873 every 2 weeks (Day 1 and Day 15). Evaluations will be performed on C1D1, C1D8, C2D1 and C2D15 (+/- 48 hours) as indicated in Section 3.4.  
Cycle 1 Day 1 will be on any day between Day 4 and Day 10 after surgery, when AST/ALT
elevations ≤ 10 X ULN and decreasing for 2 consecutive days, and Bilirubin ≤ 2 X ULN or decreasing for 2 consecutive days. If LFTs do not decrease as described within 14 days after surgery, patient will be taken off treatment. Treatment with MVT-5873 will continue until off treatment criteria are met.  
| Pre-operative Dose                                                                                                                                     | Operation         | Weekly dosing          | Q2 Weekly dosing        |       |    |
|--------------------------------------------------------------------------------------------------------------------------------------------------------|-------------------|------------------------|-------------------------|-------|----|
| Day -3                                                                                                                                                 | Day 0             | Cycle 1:  Days 1 and 8 | Cycle 2:  Days 1 and 15 |       |    |
| MVT-5873                                                                                                                                               |                   |                        |                         |       |    |
| DL1                                                                                                                                                    | 1 mg/kg           | Pancreatectomy         | 1                       | mg/kg | MVT    |
| 5873*                                                                                                                                                  | 1 mg/kg MVT-5873* |                        |                         |       |    |
| or Hepatectomy                                                                                                                                         |                   |                        |                         |       |    |
| MVT-5873                                                                                                                                               |                   |                        |                         |       |    |
| DL2                                                                                                                                                    | 3 mg/kg           |                        |                         |       |    |
| * Post-operative infusions of MVT-5873 will consist of 1 mg/kg in all cycles. MVT-5873 dose  variability is only within the single pre-operative dose. |                   |                        |                         |       |    |
normal
3.1.1 dose limiting toxicity

The DLT period is 10 days after pre-operative MVT-5873 treatment. Any Grade ≥ 3 adverse event (AE), occurring during the DLT evaluation period, possibly or definitely attributed to MVT-5873 with the exception of:  
- Bilirubin < 6X ULN if resolved to < 3 or ≤ 2X the baseline in 7 days after first MVT-5873 treatment.  
- AST/ALT < 20X ULN if resolved to < 10X ULN in 7 days after first MVT-5873 treatment.
Resection will be undertaken to remove all demonstrable disease in the liver, pancreas or bile ducts on Day 0.  
Note: The operation may be delayed until AST/ALT elevations are ≤ 10X ULN, and Bilirubin is
≤ 2X the baseline value or < 3X ULN. If the operation is delayed for more than 4 days due to failure of LFT to fall within the values outlined above, this will be considered DLT.
normal
3.1.2 safety lead-in cohorts

In order to minimize patient risk, we will initially accrue six (6) subjects sequentially in two safety lead-in cohorts based upon AST/ALT/Bili abnormalities and irrespective of tumor histology. For each safety lead-in cohort we will evaluate two dose levels, 1 mg/kg and 3 mg/kg as summarized in **Table 10** below.  
The safety lead-in continues until at least two subjects among the cohort of six subjects experience dose-limiting toxicities (ie, ≥33% of subjects with a dose-limiting toxicity at that dose level). If none (0) or one (1) of the six subjects at 1 mg/kg experiences a dose-limiting toxicity, then escalation to 3 mg/kg may proceed within each safety lead-in cohort and 1 mg/kg is deemed safe.  
If none (0) or one (1) of the six subjects experiences a dose-limiting toxicity at 3 mg/kg within each safety lead-in cohort, then 3 mg/kg is deemed safe at the respective safety lead in cohort. Dose esclation maximum will be 3 mg/kg.  
Table 10: Safety Lead-In Cohorts  
| Safety Lead-In                                                                                                                                       | Safety Lead-In   |                  |           |          |
|------------------------------------------------------------------------------------------------------------------------------------------------------|------------------|------------------|-----------|----------|
| Cohort 1 (Low elevated LFTs)                                                                                                                         | Cohort           | 2*               | (Moderate | elevated |
| LFTs)                                                                                                                                                |                  |                  |           |          |
| LFT                                                                                                                                                  | N=6              | N=6              |           |          |
| AST/ALT                                                                                                                                              | ≤ 2X ULN, and/or | < 5X ULN, and/or |           |          |
| Bilirubin                                                                                                                                            | ≤ 1.5X ULN       | ≤ 3X ULN         |           |          |
| *LFTs above what is required for Cohort 1 AND AST/ALT < 5X ULN and/or bilirubin ≤ 3X ULN.  Note: LFTs determined at screening for cohort assignment. |                  |                  |           |          |  
Every subject of each safety lead-in cohort will be observed for at least 10 days after first dose of MVT-5873 before the subsequent subject can be treated. Subjects who do not complete the DLT observation period for reasons other than a DLT will be replaced and will not be included in the response evaluation. Intra-patient dose de-escalation is not permitted for subjects enrolled in the safety lead-in cohorts. Subjects will receive assigned dose and in case of DLT will be taken off protocol. Every subject enrolled onto safety lead-in cohorts will be part of the DLT evaluation of its respective dose level. Subjects who experience DLT after pre-operative dose of MVT-5873 may have surgery when DLT
is resolved and will be taken off MVT-5873 treatment after operation (see Section 3.5 for surgical guidelines and post-operative care).
normal
safety lead-in cohort 1 (low

Six (6) subjects will be enrolled at Dose Level 1, treated with MVT-5873 (**1 mg/kg on Day -3**)
and have surgery on Day 0. If two (2) subjects experience a DLT during DLT period at dose level DL1, no further attempts at completing the safety run-in of MVT-5873 in combination with surgery will be conducted, and the trial will be closed to accrual. If < 2 of 6 subjects experience DLT during DLT period, DL1 will be considered to be safe to use in subjects with low elevated LFTs and enrollment into Cohort 2 will begin at DL1. Concurrently, enrollment into Cohort 1 Dose Level 2 will begin.
normal
cohort 1 dose level 2

Six (6) subjects will be enrolled at Dose Level 2, treated with MVT-5873 (**3 mg/kg on Day -3**)
and have surgery on Day 0. If two (2) subjects experience a DLT during DLT period at dose level DL2, no further attempts at completing the safety run-in at DL2 will be conducted at trial will continue with DL1 only. If < 2 of 6 subjects experience DLT during DLT period, DL2 will be considered to be safe to use in subjects with low elevated LFTs.
normal
safety lead-in cohort 2 (moderate

If DL1 is proved to be safe in Cohort 1 Dose Level 1, 6 subjects will be enrolled in Cohort 2 Dose Level 1. Subjects will be treated with MVT-5873 (**1 mg/kg on Day -3**) and have surgery on Day 0. If two (2) subjects experience a DLT during DLT period at dose level DL1, no further attempts at completing the safety run-in of MVT-5873 in combination with surgery in subjects with moderate elevated LFTs will be conducted. If < 2 of 6 subjects experience DLT during DLT period, DL1 will be considered to be safe to use in subjects with moderate elevated LFTs. If DL1 is safe to use in subjects with moderate elevated LFTs, 6 subjects will be enrolled at Dose Level 2 in Cohort 2.
normal
cohort 2 dose level 2

Six (6) subjects will be treated with MVT-5873 (**3 mg/kg on Day -3**) and have surgery on Day 0.  
If two (2) subjects experience a DLT during DLT period at dose level DL2, no further attempts at completing the safety run-in at DL2 will be conducted at trial will continue with DL1 only. If < 2 out of 6 experience DLT during DLT period, DL2 will be considered to be safe to use in subjects with moderate elevated LFTs.
normal
3.1.3 treatment cohorts

3.1.3.1 Treatment Cohorts at Pre-Operative MTD Once the pre-operative safe dose of MVT-5873 and allowed elevated level of LFTs is estimated, additional subjects will be evaluated at that MTD (see **Table 11**) within Cohorts 3-5.  
Treatment subjects will be accrued with the following histologies:  
o Metastatic Colorectal Cancer (n=24)
o Cholangiocarcinoma (n= 24)
o Pancreatic Cancer (n=23)
Note: Subjects treated during the safety lead-in (Cohorts 1 and 2) will complete the study as indicated in Sections **3.1.2** and 3.5.  
| Pre-operative Dose   | Operation         | Weekly dosing          | Q2 Weekly dosing        |    |
|----------------------|-------------------|------------------------|-------------------------|----|
| Day -3               | Day 0             | Cycle 1:  Days 1 and 8 | Cycle 2:  Days 1 and 15 |    |
| MVT-5873 at MTD      | Pancreatectomy    | 1                      | mg/kg                   | MVT    |
| 5873*                | 1 mg/kg MVT-5873* |                        |                         |    |
| or Hepatectomy       |                   |                        |                         |    |  
3.1.3.2 Treatment Cohorts at Two Different Pre-Operative MTDs by LFT Elevation are Determined from Safety Lead-In If two different pre-operative safe doses of MVT-5873 are determined from the safety lead-in due to the difference of LFT elevation, subjects within the treatment cohorts will each be treated by their elevated level of LFTs as identified at screening (refer to **Table 12** and Table 13).  
Table 12: Treatment Guidelines (If Two Different Pre-Operative MTDs by LFT Elevation are Determined from Safety Lead-In)  
| Low elevated LFTs                   | Moderate elevated LFTs                          |                                                                                             |
|-------------------------------------|-------------------------------------------------|---------------------------------------------------------------------------------------------|
| Treatment                           | (AST/ALT ≤ 2X ULN and/or  bilirubin ≤ 1.5X ULN) | (LFTs above what is required for  Cohort 1 AND AST/ALT < 5X ULN  and/or bilirubin ≤ 3X ULN) |
| Cohorts Cohorts 3-5                 | MVT-5873 at                                     | MVT-5873 at                                                                                 |
| Low elevated LFTs MTD               | Moderate elevated LFTs MTD                      |                                                                                             |
| Note: LFTs determined at screening. |                                                 |                                                                                             |  
* Baseline LFTs for a subject may be used if the subject's screening and baseline LFTs are different and the change is determined by the PI to be clinically indicated.  
Table 13:Treatment Study Design (If Two Different Pre-Operative MTDs by LFT Elevation are Determined from Safety Lead-In)  
| Determined from Safety Lead-In)  Pre-operative Dose Operation                               | Weekly dosing                             | Q2 Weekly dosing       |                         |       |    |
|---------------------------------------------------------------------------------------------|-------------------------------------------|------------------------|-------------------------|-------|----|
| Day -3                                                                                      | Day 0                                     | Cycle 1:  Days 1 and 8 | Cycle 2:  Days 1 and 15 |       |    |
| MVT-5873 at                                                                                 |                                           |                        |                         |       |    |
| Low  elevated  LFTs                                                                         | Low elevated  LFTs MTD                    | Pancreatectomy         | 1                       | mg/kg | MVT    |
| 5873*                                                                                       | 1 mg/kg MVT-5873*                         |                        |                         |       |    |
| or Hepatectomy                                                                              |                                           |                        |                         |       |    |
| Moderate  elevated  LFTs                                                                    | MVT-5873 at  Moderate  elevated LFTs  MTD |                        |                         |       |    |
| * Post-operative infusions of MVT-5873 will consist of 1 mg/kg in all cycles. MVT-5873 dose |                                           |                        |                         |       |    |  
* Post-operative infusions of MVT-5873 will consist of 1 mg/kg in all cycles. MVT-5873 dose variability is only within the single pre-operative dose.
normal
study drug administration 3.2.1 preparation

MVT-5873 is provided as a single-use vial containing 3.0 mL of sterile solution at 10 mg/mL in a buffer at pH 6.0, containing 25 mM histidine, 150 mM sucrose, 55 mM sodium chloride, and 0.02% polysorbate 80. Prior to infusion, MVT-5873 is diluted in normal saline to obtain a final concentration of less than 5 mg/mL. The administration apparatus must include a 0.2-micron inline filter.
normal
3.2.3 upper limit to administered

In subjects who weigh more than 100 kg (220 pounds), the administered dose should be calculated based on a theoretical weight of 100 kg.
normal
3.2.4 infusion rate of mvt-5873

The first dose of MVT-5873 for each subject will be administered as an intravenous infusion over at least 120 minutes. In subjects who tolerate an initial infusion without difficulty, subsequent infusions may be reduced by 30 minutes to a minimum infusion duration of 60 minutes (i.e., a 120minute infusion during the first infusion, a 90-minute infusion during the second infusion, a 60minute infusion during the third and subsequent infusions). The duration of infusion for MVT5873 should never be less than 60 minutes.  
Vital signs will be taken pre-and post-MVT-5873 administration.
normal
3.2.5 treatment guidelines for mvt-5873-related

Please refer to table, **Treatment Guidelines for MVT-5873-Related Infusion Reaction,** for guidance on managing infusion related reactions during MVT-5873 administration.  
| Treatment Guidelines for MVT-5873-Related Infusion Reaction Grade Definition Treatment Guideline (CTCAEv4) 1 Mild transient reaction; infusion  interruption not indicated;  intervention not indicated  Monitor for at least 2 hours (120 minutes)  after end of infusion; provide patient  instructions and prophylactic medications  at discharge as appropriate 2 Therapy or infusion interruption  indicated but responds promptly to  symptomatic treatment Infusion may be restarted once following  a minimal 60-minute delay. A second  infusion interruption requires  discontinuation of MVT-5873  administration. Administration of medications (e.g., antihistamines, steroids, NSAIDS and IV  fluids etc.) as appropriate; monitor in  clinic for at least 2 hours (120 minutes)  after completion of infusion; provide  prophylactic medications as appropriate  for subsequent MVT-5873 doses  3 Prolonged (e.g., not rapidly  responsive to symptomatic  medication and/or brief  interruption of infusion);  recurrence of symptoms following  initial improvement,  hospitalization indicated for  clinical sequelae Administration of medications (e.g., antihistamines, steroids, NSAIDS and IV  fluids etc.) as appropriate; hospitalization  for medical care and observation;  Discontinue MVT-5873 permanently  4 Life-threatening consequences;  Hospitalization for urgent medical  urgent intervention indicated  treatment; Discontinue MVT-5873  permanently   |
|---|
normal
3.2.6 premedication

Subjects should receive their initial dose of MVT-5873 in the absence of premedication. In subjects who experience infusion reactions that are manageable and allow for additional administration of MVT-5873, premedication may be administered for subsequent doses consistent with institutional guidelines. Starting in February 2017 (as documented in the MabVax 0715 Clinical Study Note to File dated 13-Feb 2017), the Safety Committee allowed for the use of prophylactic medications for the control of infusion reactions. The recommended dose was 12 - 20 mg of decadron IV at least 30 minutes prior and 4 mg BID PO of decadron for two days following each MVT-5873 administration. The investigator was free to manage the taper of decadron in those subjects who did not demonstrate infusion reactions.
normal
dose modifications

Adverse Events: All adverse events in this trial will be graded using the NCI Common Terminology Criteria for Adverse Events (CTCAE) version 5.0 - a complete listing is available at the CTEP website: http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm.  
Dose adjustments for pre-operative MVT-5873 are explained in Section **3.1.2**.  
Summary of dose holding/interruptions for MVT-5873 in case of MVT-5873-related adverse events (graded according to NCI-CTCAE v5.0).  
| General Adverse Events                                                                                                                    | Action                                                                                                                                                                                                                                                                                                                                                                                                                               |
|-------------------------------------------------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Non-hematological, Grade 1 or 2                                                                                                           | Continue MVT-5873 therapy at full dose prescribed.  Apply maximum supportive care recommendations.  If prolonged duration of Grade 2 adverse event (≥ 7  days) is affecting quality of life, consider  discontinuation of the drug.                                                                                                                                                                                                  |
| Non-hematological, Grade 3 or 4                                                                                                           | Apply maximum supportive care recommendations.  Hold MVT-5873 therapy until recovery to Grade ≤ 1  (up to 14 days).                                                                                                                                                                                                                                                                                                                  |
| (excluding cardiac and hepatobiliary  events)                                                                                             | For Grade 3 or 4 interstitial pneumonitis or Grade 3 or  4 rash manifested as toxic epidermal necrolysis (e.g.  Stevens-Johnson Syndrome etc.) MVT-5873 must be  discontinued.  If recurrence of adverse event after drug hold/  interruptions is observed, and maximum supportive  care measures applied, hold drug once again until  recovery to Grade ≤ 1 (up to 14 days). If no recovery,  consider discontinuation of the drug. |
| Non-hematological, Grade 3 or 4  adverse events NOT resolved to  Grade ≤ 1 within a maximum of 2  weeks from last planned  administration | Action (discontinue or resume MVT-5873 therapy) in  individual cases after discussions with the Sponsor.                                                                                                                                                                                                                                                                                                                             |  
| General Adverse Events                                                                                                                                                                                                                                                                           | Action                                                                                                                                                    |           |     |       |    |                                                |
|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------|-----------|-----|-------|----|------------------------------------------------|
| Cardiac Adverse Events Cardiac (Severity corresponding to                                                                                                                                                                                                                                        | MVT-5873 therapy to be discontinued permanently in                                                                                                        |           |     |       |    |                                                |
| NYHA criteria)                                                                                                                                                                                                                                                                                   | case of symptomatic NYHA class III and IV CHF.  MVT-5873 therapy to be held, continued, or resumed  accordingly for subjects with NYHA class I or II CHF. |           |     |       |    |                                                |
| Hepatobiliary Adverse Events  Grade 1-2 bilirubin or Grade 1-2                                                                                                                                                                                                                                   | Continue MVT-5873 therapy at full dose prescribed.                                                                                                        |           |     |       |    |                                                |
| AST/ALT                                                                                                                                                                                                                                                                                          | Apply maximum supportive care recommendations.                                                                                                            |           |     |       |    |                                                |
| Grade                                                                                                                                                                                                                                                                                            | 3                                                                                                                                                         | bilirubin | or  | Grade | 3  | Apply maximum supportive care recommendations. |
| AST/ALT                                                                                                                                                                                                                                                                                          | Hold MVT-5873 therapy until recovery to Grade < 1  If not resolved for 14 days, discontinue treatment.                                                    |           |     |       |    |                                                |
| Grade                                                                                                                                                                                                                                                                                            | 2                                                                                                                                                         | bilirubin | and | Grade | 3  |                                                |
| AST/ALT                                                                                                                                                                                                                                                                                          | *Discontinue MVT-5873                                                                                                                                     |           |     |       |    |                                                |
| Grade 4 events *Unless Surgery-related cause Note: In case of multiple short interruptions of dose due to either adverse events or drug  supply or other reasons the sum of days without MVT-5873 treatment should not exceed 28  days between two treatments or in any 90-day treatment period. |                                                                                                                                                           |           |     |       |    |                                                |  
| Cycle 1                       | Cycle 2   | Long  Term FU4   |     |      |             |    |    |    |    |    |    |
|-------------------------------|-----------|------------------|-----|------|-------------|----|----|----|----|----|----|
| D15                           | D8        | D1               | D15 | EOT2 | Safety  FU3 |    |    |    |    |    |    |
| Screening1                    | Baseline1 | D-3              | D01 |      |             |    |    |    |    |    |    |
| MVT-5873                      | X         | X                | X   | X    | X           |    |    |    |    |    |    |
| Surgery                       | X         |                  |     |      |             |    |    |    |    |    |    |
| Informed Consent              | X         |                  |     |      |             |    |    |    |    |    |    |
| Medical History               | X         |                  |     |      |             |    |    |    |    |    |    |
| Concomitant Medications       | X         | X                | X   | X    | X           | X  | X  | X  |    |    |    |
| Adverse Event Evaluation      | X         | X                | X   | X    | X           | X  | X  | X  |    |    |    |
| Physical Exam                 | X         | X                | X   | X    | X           | X  | X  | X  | X  | X  |    |
| Vital Signs                   | X         | X                | X   | X    | X           | X  | X  | X  | X  | X  |    |
| Height                        | X         |                  |     |      |             |    |    |    |    |    |    |
| EKG                           | X         | X                |     |      |             |    |    |    |    |    |    |
| Weight                        | X         | X                | X   | X    | X           | X  | X  | X  | X  | X  |    |
| Performance Status            | X         | X                | X   | X    | X           | X  | X  | X  | X  | X  |    |
| CBC w/Differential, Platelets | X         | X                | X   | X    | X           | X  | X  | X  | X  | X  |    |
| PT, INR, PTT                  | X         | X                | X   | X    | X           | X  | X  | X  | X  |    |    |
| Serum Chemistry6              | X         | X                | X   | X    | X           | X  | X  | X  | X  | X  |    |
| CA 19-97                      | X         | X                | X7  | X    | X7          | X7 | X7 | X7 | X  | X  | X  |  
| Cycle 1                                                                        | Cycle 2   | Long  Term FU4   |     |      |             |    |    |
|--------------------------------------------------------------------------------|-----------|------------------|-----|------|-------------|----|----|
| D15                                                                            | D8        | D1               | D15 | EOT2 | Safety  FU3 |    |    |
| Screening1                                                                     | Baseline1 | D-3              | D01 |      |             |    |    |
| Hepatitis B and C Serology and/or                                              | X         |                  |     |      |             |    |    |
| Viral Load  Radiologic Evaluation8                                             | X         | X                | X   | X    | X           | X  | X  |
| Serum or Urine Pregnancy Test                                                  | X         | X                |     |      |             |    |    |
| Confirmation of dx by NCI LP                                                   | X         |                  |     |      |             |    |    |
| Tissue Collection for Research                                                 | X         |                  |     |      |             |    |    |
| Research Blood Collection (see                                                 | X         | X                | X   | X    |             |    |    |
| Section 5.1) Annual Telephone or Other NIH  Approved Remote Platforms  Contact | X         |                  |     |      |             |    |    |  
1 Day 0 operation may be delayed for 4 days until AST/ALT elevations are ≤ 10X ULN, and Bilirubin is ≤ 2X the baseline value or < 3X ULN. 2 Day 7 (+/-7 days) after treatment discontinuation. If patient is taken off treatment on the day of regular visit to Clinical Center, this day will be EOT visit.  
3 Safety Follow Up visits/labs will occur if patient is agreeable to coming back and physically able to return for the visit. This visit should occur around Day 28 post the last dose of study drug. If toxicities cause discontinuation of therapy, subjects will be followed until resolution of toxicity to at least Grade 1. If the patient cannot return to the Clinical Center for this visit, a request will be made to collect required clinical labs from a local physician or laboratory (as these are routine, standardized laboratory tests, interlaboratory variability is not a concern). If this is not possible, subjects may be assessed by telephone or other NIH approved remote platforms for symptoms.  
4 Follow-up will be telephone or other NIH approved remote platforms contact of patient or medical oncologist every 4 months (+/- 2 weeks) to assess disease and survival status. Medical records from visits done with the home physician may be securely sent to the study team per CC policy and procedures.  
5 Cycle 1 Day 1 will be on any day between Day 4 and Day 10 after surgery, when AST/ALT elevations ≤ 10 X ULN and decreasing for 2 consecutive days, and Bilirubin ≤ 2 X ULN OR decreasing for 2 consecutive days.  
6 Electrolytes, BUN, creatinine, AST, ALT, total bilirubin, alkaline phosphatase, calcium, phosphorus, albumin, magnesium, amylase.  
7 Serum CA19-9 collection pre-dose and 2 hours post dose (+30 minutes) as noted in treatment cycles.  
8 Evaluations will be performed on C1D1, C1D8, C2D1 and C2D15 (+/- 48 hours) and EOT. CT scan of chest, abdomen and pelvis in all subjects. MRI with contrast for subjects with liver disease. Note: Exceptions for the use of contrast during this MRI may be made using the best clinical judgement of the investigator.
normal
surgical guidelines

Pancreas tumors will be extirpated according to standard practice. Resections involving the pancreas may require either a pancreaticoduodenectomy or distal pancreatectomy, with or without splenectomy. Liver operations will be undertaken according to practice norms. 3.5.1 **Preoperative Patient Management** Subjects will receive standard preoperative care as appropriate to the planned surgical intervention and the patient's underlying health status. This will include:
- Clear liquid-diet the evening prior to operation, without additional bowel preparation.  
- Hibiclens shower the night before operation.  
- CA 19-9 levels will be drawn the morning of the procedure. - 20 mL of peripheral blood will be drawn prior to induction of anesthesia for circulating tumor cell acquisition and analysis (see Section **5.2.6**).  
- Preoperative IV antibiotics administered in pre-op holding. - Subcutaneous heparin administration for venous thromboembolism prophylaxis just prior to operation start.  
- Sequential compression devices will be placed on the lower extremities prior to induction of general anesthesia.  
3.5.2 **Patient Management in the Operating Room** 3.5.2.1 At Operation:
- Subjects will undergo a midline line or sub-costal incision. A thorough examination of the abdomen will ensue to rule-out occult metastatic disease on the peritoneal surfaces, or unrecognized suspicious lymphadenopathy. The operation will then be carried out according to standard practice.  
- Prior to tumor extirpation, a 24-gage needle will be used to draw 20 mL of blood from the portal vein OR suprahepatic vena cava.  
- Normal liver core biopsies and excisional biopsies will take place for exploratory endpoints but will not exceed 100 grams.  

3.5.3 **Postoperative Care**
3.5.3.1 Patient Monitoring
- The subjects will be monitored in the ICU or on the surgical ward per routine.  
- On the ward, vital signs (blood pressure, temperature, pulse, respirations) will be taken per routine (every 2-6 hours and as clinically indicated).  
- Subjects will receive routine post-operative care; early ambulation will be encouraged.  
Note: Other standard of care may be performed as deemed clinically necessary by the PI.  
- Laboratory evaluations will include:
o CBC, platelets, acute care, mineral and hepatic panel on post-operative Days 1 through 7, and then as clinically indicated until discharge.  
o Subjects will be transfused as appropriate to maintain a hemoglobin greater than or equal to 8 g/dl.
normal
3.5.4 discharge

- Total hospitalization may be approximately 7-14 days.  
Subjects who are discharged within this time frame should be able to tolerate an oral diet with or without dietary supplements.
normal
pathological evaluation

The histopathologic evaluation, in addition to standard practice of lymph nodes, margin status, etc, will involve an evaluation by a third party laboratory for antibody/tumor cell binding (**Figure 14**
top image) and a detailed evaluation of the liver parenchyma under the supervision of Dr. David Kleiner (**Figure 14** bottom image). The liver parenchyma evaluation will include standard immunohistochemical agents as well as any special evaluative measures as deemed necessary by Dr. Kleiner.
normal
criteria for removal from protocol

Prior to removal from study, effort must be made to have all subjects complete a safety visit approximately 28 days following the last dose of study therapy.
normal
3.7.1 criteria for removal from

- Excessive toxicity (see Sections **3.1.1** and 3.3).  
- Completion of 2 cycles of post-operative MVT-5873 treatment.  
- Recurrent disease. - CA 19-9 level > 2 X the lowest recorded post-operative dose AND > ULN.  
- Participant requests to be withdrawn from active therapy.  
- Investigator discretion. - Positive pregnancy test.  
- Start of standard adjuvant chemotherapy (per medical oncologist decision). **Note:**
Patients need to be removed from therapy so that MVT-5873 will not be given within 2 weeks of the planned start date for adjuvant chemotherapy.  
- Elapse of > 84 days from the start of Day 1, Cycle 1.
normal
3.7.2 off-study criteria

- Participant requests to be withdrawn from study.  
- Death.  
- Investigator discretion. - Loss of follow up. - PI decision to end the study.  
- Screen failure.
normal
4 concomitant medications/measures

During the post-operative period, subjects will receive all standard of care supportive measures, including possible nasogastric tube drainage and bowel rest for ileus, pulmonary toilet teaching and incentive spirometry to prevent atelectasis, transfusions, and antibiotics as indicated. During receipt of study drug, standard adjunctive medications inclusive of anti-emetic, anti-nausea and bowel function therapy will be employed.
normal
5 biospecimen collection

| SPECIMEN COLLECTION TABLE                                         |                          |                                    |                                 |                                            |         |     |    |
|-------------------------------------------------------------------|--------------------------|------------------------------------|---------------------------------|--------------------------------------------|---------|-----|----|
| Test/Assay                                                        | Volume  Blood  (approx.) | Type of                            | Collection                      |                                            |         |     |    |
| Tube                                                              | Points                   | Location of  Specimen  Processing/ | Location of  Specimen  Analysis |                                            |         |     |    |
| (+/-48hrs)                                                        | Storage                  |                                    |                                 |                                            |         |     |    |
| Circulating  Tumor Cell  Analysis                                 | 20 mL                    | CellSave  Preservati on Tubes      | D-3, D0 &                       | (Delivered                                 | by      |     |    |
| C1D1                                                              | study team to)           | Trepel Lab                         |                                 |                                            |         |     |    |
| Trepel Lab                                                        |                          |                                    |                                 |                                            |         |     |    |
| Monocyte  Phagocyte  System  (MPS)  Profiling                     | 17 mL                    | SST                                | D-3 & D0                        | Hernandez Lab  for storage  until shipment | Zamboni | Lab | at |
| UNC Chapel Hill                                                   |                          |                                    |                                 |                                            |         |     |    |
| Biomarker                                                         | 10 mL                    | EDTA                               | D-3, D0,                        |                                            |         |     |    |
| Studies                                                           | (Lavender)               | C1D1 &  EOT                        | Hernandez Lab                   | Hernandez Lab                              |         |     |    |
| (Blood  Processing  Core (BPC) as  necessary, see  Section 5.2.1) |                          |                                    |                                 |                                            |         |     |    |  
| Metastasis  Assays Using  Ex Vivo  Perfusion of  Omentum          | Omentum                                       | N/A                                           | D0 (at time         | (Immediately   |
|-------------------------------------------------------------------|-----------------------------------------------|-----------------------------------------------|---------------------|----------------|
| of surgery)                                                       | to)                                           | Hernandez Lab                                 |                     |                |
| Hernandez Lab                                                     |                                               |                                               |                     |                |
| Ex Vivo  Hepatic  Perfusion  Models                               | Tumor                                         | N/A                                           | D0 (at time         | (Immediately,  |
| tissue                                                            | of surgery)                                   | when necessary)                               | Hernandez Lab       |                |
| Hernandez Lab                                                     |                                               |                                               |                     |                |
| Tumor  tissue with  mesothelial  metastases                       |                                               |                                               |                     |                |
| Proteogenomi c Analysis                                           | N/A                                           | D0 (at time                                   | Hernandez Lab       |                |
| of surgery)                                                       | Hernandez Lab                                 |                                               |                     |                |
| (Blood  Processing  Core (BPC) as  necessary, see  Section 5.2.1) |                                               |                                               |                     |                |
| Drug  Penetration  Analysis                                       | Tumor                                         | N/A                                           | D0 (at time         | Hernandez Lab  |
| tissue                                                            | of surgery)                                   | Hernandez Lab                                 |                     |                |
| (Blood  Processing  Core (BPC) as  necessary, see  Section 5.2.1) |                                               |                                               |                     |                |
| RNA                                                               | 10 mL                                         | EDTA                                          | D0                  | Hernandez Lab  |
| Analysis                                                          | (Lavender)                                    | for storage  until shipment  (by study  team) | Genomic  Expression |                |
| RNA-Seq                                                           | Tumor  tissue                                 | N/A                                           | D0 (at time         | Hernandez Lab  |
| of surgery)                                                       | for storage  until shipment  (by study  team) | Genomic  Expression                           |                     |                |
| (and  normal  tissue, as  available)                              |                                               |                                               |                     |                |
normal
correlative studies for research/pharmacokinetic studies

Tumor and normal tissue samples will be collected at the time of surgery. Tissue will only be taken for research purposes after it is removed as part of the standard operation, as viable tissue that will otherwise be removed as part of the operation, or as core needle biopsies of normal liver for subjects undergoing pancreatectomies. The total amount of normal liver tissue removed for research purposes will not exceed 100 grams.  
Some of the samples will be immediately used in the Dr. Hernandez's lab (see Section **5.3.4**
below), and the rest will be sent for barcoding and initial storage to Blood Processing Core (BPC) as necessary per PI instruction. RNA-Seq data will be analyzed for correlation with recurrence patterns. The analysis will utilize logistic regression and corrections for false discovery (5%). The analysis will be undertaken in conjunction with the Branch's bioinformatics specialist. 5.2.2 **RNA Evaluation** Tumor and normal tissue samples (coded-linked) will be sent to Genomic Expression for RNA
analysis under MTA (Section **11.1**):
Gitte Pedersen, CEO Genomic Expression, Inc. 100 Cummings Center, Suite 451C Beverly, MA 01915 Phone: 203-812-0690 Email: glp@genomicexpression.com Samples sent to Genomic Expressions for RNA analysis will be coded before shipment. Genomic Expression has reengineered quantitative RNA sequencing for clinical applications, with the goal of enabling oncologists to routinely use tumor gene expression profiles to help select the best treatment options for their subjects. OneRNA, the proprietary platform from Genomic Expression, uses a novel approach to prepare sequencing libraries from tumor messenger RNA. The OneRNA library preparation technology, generates a library of nucleic acid tags from the 3' ends of mRNA molecules. These tag libraries can be "read out," meaning sequenced and counted, on next generation sequencers. This method reduces the number of reads to detect weakly expressed genes by 5- to 10-fold and simplifies the generation of a global expression profile from the sequencing data. OneRNA proprietary sample prep is integrated with proprietary analytics. After correcting for PCR duplicates, the sequenced tags are mapped to the human genome and/or transcriptome and compiled into gene expression profiles. Tumor expression profiles are normalized alongside normal tissue expression profiles to put them on a common scale. Using outlier detection methods, over and under expressed genes are identified by comparing the expression level of each gene in the tumor to the distribution of expression levels for that same gene in normal tissue(s).
normal
5.2.3 metastasis assays using ex

For instances in which a standard Whipple procedure is done, omentum obtained from the operating room (as part of the standard Whipple procedure) will be immediately transported to Dr. Hernandez's lab. The gastroepiploic arterial arcade will be cannulated and the omentum kept viable using Dr. Hernandez's Liver Assist Device (**Figure 4** bottom).
normal
5.2.4 ex vivo hepatic perfusion

Resected tumor-bearing liver and pancreas will be perfused through native blood vessels in order to develop ex *vivo* models for pancreatic cancer, cholangiocarcinoma, and metastatic colorectal cancer as indicated in Section **1.2.11**.
normal
5.2.5 circulating tumor cell evaluation

Blood will be collected for circulating tumor cell (CTC) assays to be performed in the Dr. Trepel Laboratory (Developmental Therapeutics Branch, CCR).  
Peripheral blood will be drawn from a peripheral vein and the portal or supra-hepatic IVC (intraop) for enumeration and characterization of CTCs. CTCs will be assessed using ferrofluidic enrichment and multiparameter flow cytometric detection. CTCs will be identified as viable, nucleated cells, that positively express one or more epithelial or tumor markers and are negative for expression of hematopoietic markers. The number of CTCs in peripheral blood will be counted. In addition to quantification, we will evaluate CTCs for CA 19-9 expression using flow cytometry.
normal
5.2.6 monocyte phagocyte system (mps)

Blood will be collected for monocyte phagocyte system (MPS) profiling, to be performed in the Zamboni Laboratory - University of North Carolina at Chapel Hill. MPS profiling will be used to evaluate the impact of MVT-5873 therapy on circulating tumor cells. Coded linked samples will be sent to Dr. Zambroni at the Zamboni Laboratory under MTA (see Section **11.1**):
Zamboni Laboratory - UNC at Chapel Hill Attn: Andrew Lucas 120 Mason Farm Road 1022A Genetic Medicine Building, CB\#7361 Chapel Hill, NC 27599 Lab Phone: (919) 966-1622
normal
sample storage tracking and disposition

Samples will be ordered in CRIS and tracked through a Clinical Trial Data Management system. Should a CRIS screen not be available, the CRIS downtime procedures will be followed. Samples will not be sent outside NIH without appropriate approvals and/or agreements, if required. Tissue samples will be sent to Blood Processing Core (BPC) for barcoding and initial storage until they are distributed to Dr. Hernandez for sample analysis as described in the protocol.
normal
5.3.1 samples managed by the

Please e-mail NCIBloodcore@mail.nih.gov at least 24 hours before transporting samples (the Friday before is preferred). For sample pickup, page 102-11964. For immediate help, call 240-760-6180 (main blood processing core number) or, if no answer, 240-760-6190 (main clinical pharmacology lab number). For questions regarding sample processing, contact NCIBloodcore@mail.nih.gov. 5.3.1.2 Sample Data Collection All samples sent to the Blood Processing Core (BPC) will be barcoded, with data entered and stored in the Labmatrix utilized by the BPC. This is a secure program, with access to Labmatrix limited to defined Figg lab personnel, who are issued individual user accounts. Installation of Labmatrix is limited to computers specified by Dr. Figg. These computers all have a password restricted login screen. Labmatrix creates a unique barcode ID for every sample and sample box, which cannot be traced back to subjects without Labmatrix access. The data recorded for each sample includes the patient ID, name, trial name/protocol number, time drawn, cycle time point, dose, material type, as well as box and freezer location. Patient demographics associated with the clinical center patient number are provided in the system. For each sample, there are notes associated with the processing method (delay in sample processing, storage conditions on the ward, etc.).
normal
5.3.1.3 sample storage and destruction

Barcoded samples are stored in barcoded boxes in a locked freezer at either -20 or -800C according to stability requirements. These freezers are located onsite in the BPC and offsite at NCI Frederick Central Repository Services in Frederick, MD. Visitors to the laboratory are required to be accompanied by laboratory staff at all times. Access to stored clinical samples is restricted. Samples will be stored until requested by a researcher named on the protocol. All requests are monitored and tracked in Labmatrix. All researchers are required to sign a form stating that the samples are only to be used for research purposes associated with this trial (as per the IRB approved protocol) and that any unused samples must be returned to the BPC. It is the responsibility of the NCI Principal Investigator to ensure that the samples requested are being used in a manner consistent with IRB approval. Following completion of this study, samples will remain in storage as detailed above. Access to these samples will only be granted following IRB approval of an additional protocol, granting the rights to use the material. If, at any time, a patient withdraws from the study and does not wish for their existing samples to be utilized, the individual must provide a written request. Following receipt of this request, the samples will be destroyed or returned to the patient, if so requested. The PI will record any loss or unanticipated destruction of samples as a deviation. Reporting will be per the requirements of Section 7.2.  
Sample barcodes are linked to patient demographics and limited clinical information. This information will only be provided to investigators listed on this protocol, via registered use of the Labmatrix. It is critical that the sample remains linked to patient information such as race, age, dates of diagnosis and death, and histological information about the tumor, in order to correlate genotype with these variables.
normal
5.3.2 procedures for storage of

Contact the Trepel Lab by email (Jane Trepel: trepel@helix.nih.gov; Min-Jung Lee: leemin@mail.nih.gov; Akira Yuno: akira.yuno@nih.gov and Sunmin Lee: lees@pop.nci.nih.gov) when the patient is scheduled and by phone as soon as the blood is drawn at 240-760-6330. A lab member will come to pick up the blood. Please keep blood at ambient temperature. Members of the lab will enter the samples into a secure password protected patient's sample tracking database (Translational Pharmacodynamics Research Group Patient Sample Management System) and process the samples. Sample barcodes are linked to patient demographics and limited clinical information. This information will only be provided to investigators listed on this protocol. It is critical that the sample remains coded and linked to patient information such as race, age, dates of diagnosis and death, and histological information about the tumor, in order to correlate with these variables. Blood samples will be stored initially in the Trepel Lab in the Magnuson Clinical Center. If, at any time, a subject withdraws from the study and does not wish for their existing samples to be utilized, the individual must provide a written request. Following receipt of this request, the samples will be destroyed (or returned to the patient, if so requested). When a patient withdraws consent the participant's data will be excluded from future distributions, but data that have already been distributed for approved research use will not be able to be retrieved. The PI will record any loss or unanticipated destruction of samples as a deviation. Reporting will be per the requirements of Section 7.2.
normal
5.3.3 procedures for storage of

Tissues designated for clinical diagnostics are transported to the Laboratory of Pathology (LP) where they are examined grossly and relevant portions are fixed, embedded in paraffin and sectioned and stained for diagnostic interpretation. Unutilized excess tissue that is not embedded in paraffin is stored in formalin for up to three months, in accordance with College of American Pathologists/Joint Commission on Accreditation of Healthcare Organizations (CAP/JCAHO) guidelines, and then discarded. Following completion of the diagnostic workup, the slides and tissue blocks are stored indefinitely in the LP's clinical archives. All specimens are catalogued and retrieved utilizing the clinical laboratory information systems, in accordance with CAP/JCAHO regulations. The use of any stored specimens for research purposes is only allowed when the appropriate IRB approval has been obtained. In some cases, this approval has been obtained via the original protocol on which the patient was enrolled.
normal
5.3.4 procedures for storage of

Patient samples, collected for the purpose of research under IRB approved protocols where Dr. Hernandez is Principal Investigator, may be archived in the Laboratory of Dr. Hernandez. All data associated with archived clinical research samples is entered into the NCI Labmatrix database. All staff in Dr. Hernandez's laboratory have received updated NIH/CIT training and maintain standards of computer security. The data recorded for each sample includes the patient ID, trial name/protocol number, date drawn/collected, treatment cycle/time point, cell source (e.g. peripheral blood, marrow, biopsy) as well as box and freezer location. Patient demographics that correlate treatment outcomes and therapies with the samples can be obtained only through the NCI clinical records. All received samples will receive a unique bar code number, which will be added to the sample NCI Labmatrix database. Only this bar code will be recorded on the sample vial and the vials will not be traceable back to patients without authorized access to the NCI Labmatrix database. Samples are stored in freezers at -80°C (sera, plasma, tissue samples) or under liquid nitrogen (cells), according to stability requirements. These freezers are located onsite at Dr. Hernandez's laboratory. Access to samples from a protocol for research purposes will be by permission of the Principal Investigator.
normal
5.3.5 protocol completion/sample destruction

All specimens obtained in the protocol are used as defined in the protocol. Any specimens that are remaining at the completion of the protocol will be transferred to protocol 13C0176. Subjects will be co-enrolled on protocol 13C0176. Thus, all tissue will be stored, tracked and disposed after transfer as specified in protocol 13C0176.  
If the patient withdraws consent the participant's data will be excluded from future distributions, but data that have already been distributed for approved research use will not be able to be retrieved. The PI will record any loss or unanticipated destruction of samples as a deviation. Reporting will be per the requirements of Section 7.2.
normal
5.3.7 management of results

Subjects will be contacted if a clinically actionable gene variant is discovered. Clinically actionable findings for the purpose of this study are defined as disorders appearing in the American College of Medical Genetics and Genomics recommendations for the return of incidental findings that is current at the time of primary analysis. (A list of current guidelines is maintained on the CCR intranet: https://ccrod.cancer.gov/confluence/display/CCRCRO/Incidental+Findings+Lists). Subjects will be contacted at this time with a request to provide a blood sample to be sent to a CLIA certified laboratory.
normal
5.3.8 genetic counseling

If the research findings are verified in the CLIA certified lab, the subject will be offered the opportunity to come to NIH (at our expense) to have genetic education and counseling with the NCI Genetics Branch to explain this result. If the subject does not want to come to NIH, a referral to a local genetic healthcare provider will be provided (at their expense). This is the only time during the course of the study that incidental findings will be returned. No interrogations regarding clinically actionable findings will be made after the primary analysis.