ATTAC-MCC: Phase I/II Study of Autologous CD8+ and CD4+ Transgenic T Cells Expressing High Affinity MCPyV-specific TCRs Combined With Avelumab and Class I MHC-upregulation in Patients With Metastatic MCC Refractory to PD-1 Axis Blockade
Phase: PHASE1
Conditions: Other Skin
Sponsor: Fred Hutchinson Cancer Center

 
 
Official Title 
ATTAC-MCC: Phase I/II study of Autologous CD8+ and 
CD4+ Transgenic T cells expressing high affinity MCPyV-
specific TCRs combined with Avelumab and Class 
I MHC-upregulation in patients with metastatic MCC 
refractory to PD-1 axis blockade 
NCT Number 
NCT03747484 
Document Type 
Protocol and Statistical Analysis Plan 
Document Date 
12/6/2023 
 
 

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FRED HUTCHINSON CANCER CENTER 
Title:  
ATTAC-MCC: Phase I/II study of Autologous CD8+ and CD4+ Transgenic T cells 
expressing high affinity MCPyV-specific TCRs combined with Avelumab and Class 
I MHC-upregulation in patients with metastatic MCC refractory to PD-1 axis 
blockade 
Protocol Number:  
IRB 9845; Consortium RG1003611 
Current Version: 
Version  15 - 02/06/2023 
Prior Version(s):  
Version 14 – 11/01/2022 
IND Number: 
FDA IND 18931 
Investigational Agent: 
FH-MCVA2TCR: Autologous CD8+ and CD4+ T cells transduced with transgenic 
TCR A2-MCC1, interferon gamma 
Study Regimen: 
Interferon gamma, FH-MCVA2TCR, and Avelumab or pembrolizumab 
Principal Investigator: 
Joshua Veatch, MD, PhD 
 
Associate in Clinical Research, Fred Hutchinson Cancer Center 
 
Acting Assistant Professor, University of Washington 
(O): 206-667-5108 
Emergency (24-hour): (206) 598-8902 (ask for the Oncology Triage Provider) 
 
Sponsor: 
Fred Hutchinson Cancer Center 
 
1100 Fairview Ave N, M/S J3-329 
 
Seattle, WA 98109-1024 
 
(O): 206-667-4247 
 
(F):  206-667-1370 
 
ISIOC@fredhutch.org 
 
Medical Monitor:  
Mary-Elizabeth Percival, MD 
 
Associate Professor, Fred Hutch Clinical Research Division 
 
(O): 206-606-1320 
 
mperciva@uw.edu 
 
Authorized Sponsor Representative: 
Name: Mary-Elizabeth Percival, MD 
Signature:  ______________________________________________ 
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Signature Page: 
Investigator Statement: 
I have carefully read Protocol 9845 entitled ATTAC-MCC: Phase I/II study of Autologous CD8+ and 
CD4+ Transgenic T cells expressing high affinity MCPyV-specific TCRs combined with Avelumab and 
Class I MHC -upregulation in patients with metastatic MCC refractory to PD-1 axis blockade 
(“Protocol”) version date: 02/06/2023.   
I agree to carry out my responsibilities in accordance with the Protocol, applicable laws and regulations 
(including 21 CFR Part 312), Good Clinical Practice: Consolidated Guidance (ICH-E6), and applicable 
policies of Fred Hutch.   
 
Principal Investigator: 
Name: 
Joshua Veatch, MD 
Institution:  Fred Hutchinson Cancer Center 
1100 Fairview Avenue N., M/S D3-100 
Seattle, WA 98019-1024 
Phone: 
206-667-5108 
 
 
Signature:_____________________________________________ Date: ______________ 
 
 
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PROTOCOL SYNOPSIS 
Protocol:  9845  
RG: 1003611 
Product Name:    FH-MCVA2TCR 
Title of Study: ATTAC-MCC: Phase I/II study of Autologous CD8+ and CD4+ Transgenic T cells 
expressing high affinity MCPyV-specific TCRs combined with Avelumab and Class I MHC -
upregulation in patients with metastatic MCC refractory to PD-1 axis blockade 
Sponsor:  Fred Hutch 
Phase of Development:  I/II 
Study Objectives: 
Primary: Safety, Efficacy   
Secondary/Exploratory: T cell persistence, T cell phenotype, T cell function, T cell localization into 
tumor, epitope spreading of T cell responses, recurrence-free survival, overall survival 
Study Purpose and Rationale:   
Metastatic Merkel cell carcinoma is an aggressive skin cancer caused by a virus. There are currently no 
effective treatments for individuals whose cancer progresses on or after treatment with PD-1 axis 
immunotherapies. We have developed an engineered autologous T cell therapy targeting Merkel cell 
polyomavirus.  In a phase I/II trial, we seek to determine the safety and potential efficacy of this T cell 
therapy combined with interferon gamma, in addition to standard treatments (anti-PDL1/avelumab) for 
patients whose virus-associated metastatic/unresectable MCC tumors are resistant to PD-1 axis 
treatments. 
Study Population:   
Individuals that are HLA-A02+ with metastatic Merkel cell polyomavirus associated Merkel cell 
carcinoma whose disease has progressed on or after treatment with a PD-1 axis checkpoint inhibitor
Test Product:   
- 
FH-MCVA2TCR: Autologous CD8+ and CD4+ T cells transduced with transgenic TCR A2-
MCC1. 
- 
Interferon gamma  
Additional Interventions (not test product, given as standard of care): 
- 
Avelumab 
- 
Pembrolizumab (as an alternative to avelumab) 
Safety Assessments: Ongoing; with evaluation of part I/II transition after three patients have been 
treated 
DLT Definition:  
DLTs will be assessed for 28 days following the infusion of the TCR transgenic T cell product. The 
following events will be considered DLTs if they are attributed as at least possibly related to TCR T 
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cell administration. Grading will be done in accordance with NCI CTCAE version 5.0 unless otherwise 
specified.  
1. Grade 3 allergic reaction or other event that leads to discontinuation of TCR T cell infusion  
2. Grade 3 autoimmune reactions  
3. Grade 3 neurotoxicity that does not resolve to grade 1 or less within 7 days  
4. Grade 3 CRS that does not resolve to < grade 3 within 7 days.  
5. Grade 4 organ toxicity not pre-existing or due to the underlying malignancy and occurring within 28 
days of study product infusion.  
6. Any other toxicity not meeting above criteria that is deemed by the PI to represent a DLT.  
Efficacy Assessments:  At completion of study 
Statistical Methods: Phase I/II interventional trial, not masked, dose-escalation. Goal grade 3+ 
treatment related toxicity rate of <40%; likelihood of stoppage if true rate 25% is 0.05.  Response 
determination by RECIST 1.1 with goal response rate statistically >5%; power to detect this is true 
response rate is 20% is 0.84.  All secondary and exploratory endpoints exploratory. 
Sample size:  up to 16 participants 
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TABLE OF CONTENTS 
 
Protocol Synopsis ............................................................................................................................................... 3 
1. 
BACKGROUND ....................................................................................................................................................9 
2. 
STUDY PURPOSE AND RATIONALE .................................................................................................................. 10 
2.1 
Rationale for Investigation of FH-MCVA2TCR ............................................................................................ 10 
2.1.1 
Scientific basis for T cell therapy targeting MCPyV in MCC .................................................................10 
2.1.2 
Target epitope: “KLLEIAPNC” MCPyV Epitope ..................................................................................11 
2.1.3 
Clinical experience with T cells targeting MCPyV in MCC: ..................................................................11 
2.1.4 
 Rationale for Transgenic T cell approach .............................................................................................13 
2.1.5 
Transgenic TCR vs. Transgenic CAR......................................................................................................14 
2.1.6 
Safety of TCR transgenic T cells for other cancers .................................................................................14 
2.1.7 
TCR A2-MCC1 is a highly potent natural TCR that effectively lyses MCC cell lines ............................14 
2.1.8 
Evaluation of potential for off-target activity and cross or allo-reactivity of TCR A2-MCC1 ...............15 
2.1.9 
Rationale for using a third-generation lentiviral vector .........................................................................15 
2.1.10 
Rationale for substrate T cell of CD62L+ (“young” but well differentiated) CD8+ T Cells .................17 
2.1.11 
Rationale for inclusion of transgenic CD4+ T cells in addition to transgenic CD8+ T cells ................17 
2.1.12 
FH-MCVA2TCR Dose Rationale ............................................................................................................18 
2.1.13 
Rationale for omission of Lymphodepleting Chemotherapy prior to T cell infusion ..............................18 
2.2       Rationale for inclusion of subcutaneous interferon gamma ........................................................................19 
2.2.1 
Scientific basis.........................................................................................................................................19 
2.2.2 
Interferon Gamma- Preclinical results ...................................................................................................19 
2.3 
Rationale for administration of TCR-transgenic T cells in the context of treatment with avelumab or 
Pembrolizumab as a standard of care ..................................................................................................................... 21 
2.3.1 
Avelumab and Pembrolizumab - Scientific basis ....................................................................................21 
2.3.2 
Avelumab - Clinical experience & safety ................................................................................................21 
2.4 
Rationale for inclusion of biopsies ............................................................................................................. 22 
3. 
STUDY OBJECTIVES ......................................................................................................................................... 22 
3.1 
Primary Objectives ..................................................................................................................................... 22 
3.1.1 
Safety .......................................................................................................................................................22 
3.1.2 
Efficacy....................................................................................................................................................23 
3.2 
Secondary Objectives ................................................................................................................................. 23 
3.2.1 
Persistence ..............................................................................................................................................23 
3.2.2 
Migration ................................................................................................................................................23 
3.2.3 
Progression free and overall survival .....................................................................................................23 
3.2.4 
Determination of preferred dose of FH-MCVA2TCR .............................................................................23 
3.2.5 
Response rate by immune RECIST ..........................................................................................................23 
3.3 
Exploratory Objectives ............................................................................................................................... 23 
3.4 
Projected Target Accrual ............................................................................................................................ 24 
4. 
STUDY DESIGN AND INVESTIGATIONAL PLAN ................................................................................................ 24 
4.1 
Design: Phase I/II single-arm dose-escalation non-masked interventional study ..................................... 24 
4.2 
Schematic ................................................................................................................................................... 26 
4.3 
Protocol Enrollment ................................................................................................................................... 26 
5. 
STUDY POPULATION AND ELIGIBILITY ............................................................................................................ 27 
5.1 
Inclusion Criteria – Evaluation .................................................................................................................... 28 
5.2 
Inclusion Criteria ......................................................................................................................................... 28 
5.2.1 
Metastatic or unresectable VP-MCC ......................................................................................................28 
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5.2.2 
Previous treatment with a PD-1 axis inhibitor or contraindication to treatment with a PD-1 axis 
inhibitor 28 
5.2.3 
HLA type HLA-A*02:01 ..........................................................................................................................29 
5.2.4 
Life expectancy must be anticipated to be >3 months at trial entry .......................................................29 
5.2.5 
18 years of age or older ..........................................................................................................................29 
5.2.6 
Capable of understanding and providing a written informed consent ....................................................29 
5.2.7 
If fertile, willingness to comply with reproductive requirements. ...........................................................29 
5.2.8 
Karnofsky performance status of >= 60% ..............................................................................................29 
5.2.9 
Willing to undergo serial tumor biopsies ................................................................................................29 
5.2.10 
Participants must be at least three weeks from last systemic MCC treatment ........................................29 
5.2.11 
Acceptable organ function ......................................................................................................................29 
5.3 
Exclusion Criteria ........................................................................................................................................ 30 
5.3.1 
Active autoimmune disease .....................................................................................................................30 
5.3.2 
Prior solid organ transplant or allogeneic hematopoietic stem cell transplant. ....................................30 
5.3.3 
Corticosteroid therapy at a dose equivalent of >10 mg prednisone per day. .........................................30 
5.3.4 
Concurrent use of other investigational agents or MCC directed therapies. .........................................30 
5.3.5 
Any Medical or psychological condition other than Merkel cell carcinoma that would significantly 
increase the risk of harm to a subject or interfere with the interpretation of trial endpoints. .................................30 
5.3.6 
Active uncontrolled infection. .................................................................................................................30 
5.3.7 
Uncontrolled concurrent illness. .............................................................................................................30 
5.3.8 
Untreated brain metastases.....................................................................................................................30 
5.3.9 
Active treatment for prior immune related adverse event to any immunotherapy. .................................30 
5.3.10 
Significant underlying neurologic disease. .............................................................................................31 
5.3.11 
Other medical, social, or psychiatric factor that interferes with medical appropriateness and/or ability 
to comply with study, as determined by PI. ..............................................................................................................31 
5.4 
Reproductive Potential and Contraception Requirements ........................................................................ 31 
6. 
TREATMENT PLAN ........................................................................................................................................... 31 
6.1 
Overall schema ........................................................................................................................................... 31 
6.2 
Leukapheresis ............................................................................................................................................. 32 
6.3 
Interferon Gamma treatment .................................................................................................................... 32 
6.4 
FH-MCVA2TCR Infusions ............................................................................................................................ 32 
6.4.1 
Preparation of Cell Product....................................................................................................................33 
6.4.2 
T Cell Infusions .......................................................................................................................................33 
6.4.3 
Acute infusion reactions ..........................................................................................................................34 
6.4.4 
Guidelines for the determination of 1 or 2 cycles of T cell therapy ........................................................34 
6.5 
Avelumab or Pembrolizumab – standard of care ....................................................................................... 35 
6.5.1 
Provision of avelumab or pembrolizumab ..............................................................................................35 
6.5.2 
Avelumab or pembrolizumab infusion ....................................................................................................35 
6.5.3 
Recommended Supportive Care, Additional Treatment, and Monitoring ...............................................36 
7. 
POTENTIAL RISKS AND TOXICITY MONITORING ............................................................................................. 37 
7.1 
Toxicity associated with interferon gamma administration ...................................................................... 37 
7.2 
Management of toxicity associated with interferon gamma ..................................................................... 37 
7.3 
Acute Toxicity Associated with T-Cell Infusion (Infusion reaction) ............................................................ 37 
7.4 
Management of Acute Infusion Reaction(s) ............................................................................................... 37 
7.5 
Acute Toxicity Associated with Avelumab or Pembrolizumab Infusion ..................................................... 38 
7.6 
Cytokine Release Syndrome (CRS).............................................................................................................. 40 
7.7 
Neurologic Toxicity ..................................................................................................................................... 41 
7.8 
Tumor Lysis Syndrome ............................................................................................................................... 41 
7.9 
Macrophage Activation Syndrome ............................................................................................................. 41 
7.10 
Adrenal Toxicity .......................................................................................................................................... 41 
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7.11 
Immune mediated adverse events ............................................................................................................. 42 
7.12 
Persistent Uncontrolled T-cell Proliferation ............................................................................................... 42 
7.13 
Replication-Competent Lentivirus .............................................................................................................. 42 
7.14 
Management of Other Toxicities ................................................................................................................ 42 
8. 
INFORMED CONSENT OF PATIENT .................................................................................................................. 43 
9. 
PATIENT REGISTRATION .................................................................................................................................. 43 
10. 
TRAVEL REIMBURSEMENT .......................................................................................................................... 43 
11. 
CLINICAL AND LABORATORY EVALUATIONS .............................................................................................. 43 
11.1 
Visits required to be at Fred Hutch/UWMC ............................................................................................... 43 
11.2 
Screening Evaluations ................................................................................................................................. 44 
11.3 
Pre-Treatment Evaluations ......................................................................................................................... 44 
11.4 
Evaluations prior to T cell Infusion ............................................................................................................. 46 
11.5 
Evaluations following T cell infusion .......................................................................................................... 46 
11.6 
Response Assessment ................................................................................................................................ 48 
11.7 
Participant discontinuation of active treatment ........................................................................................ 48 
11.8 
Long-term follow-up (LTFU) ....................................................................................................................... 49 
12. 
ADVERSE EVENT REPORTING...................................................................................................................... 49 
12.1 
Adverse Event Definitions .......................................................................................................................... 49 
12.2 
Monitoring and Recording Adverse Events ................................................................................................ 50 
12.3 
Grading of the Severity of an Adverse Event ............................................................................................. 50 
12.4 
Attribution of Adverse Event ...................................................................................................................... 50 
12.5 
Adverse Event Reporting Period - .............................................................................................................. 51 
12.6 
Expected Grade 3-4 Adverse Events .......................................................................................................... 52 
12.6.1 
Expected Adverse Events: T cell infusion ...............................................................................................52 
12.6.2 
Expected Adverse Events: Interferon gamma .........................................................................................53 
12.6.3 
Expected Adverse Events: Avelumab or pembrolizumab ........................................................................53 
12.7 
Adverse Event Reporting Requirements .................................................................................................... 53 
12.7.1 
Reporting to IRB .....................................................................................................................................53 
12.7.2 
Reporting to Sponsor ..............................................................................................................................53 
13. 
STATISTICAL CONSIDERATIONS .................................................................................................................. 54 
13.1 
Sample size and Power ............................................................................................................................... 54 
13.2 
Primary Endpoint: Safety ............................................................................................................................ 54 
13.2.1 
Definition of treatment related adverse event .........................................................................................54 
13.2.2 
Definition of success and probability of suspension ...............................................................................54 
13.3 
Primary Endpoint: Efficacy ......................................................................................................................... 55 
13.3.1 
Definition of response .............................................................................................................................55 
13.3.2 
Definition of success and power of success ............................................................................................55 
13.4 
Secondary and Exploratory analyses .......................................................................................................... 56 
14. 
DATA AND SAFETY MONITORING PLAN ..................................................................................................... 56 
14.1 
Overall Scope of Monitoring Activities ....................................................................................................... 56 
14.2 
Monitoring the Progress of Trial and Safety of Participants ...................................................................... 56 
15. 
DATA MANAGEMENT/CONFIDENTIALITY ................................................................................................... 57 
16. 
TERMINATION OF STUDY ........................................................................................................................... 57 
17. 
REFERENCES ............................................................................................................................................... 58 
APPENDIX A:  Schedule of Evaluations .................................................................................................................... 64 
APPENDIX B: Research Sample Checklist ................................................................................................................. 69 
APPENDIX C: Karnofsky Performance Status Scale ................................................................................................. 73 
APPENDIX D: ECOG Performance Status Scale ........................................................................................................ 74 
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APPENDIX F: Recommended Management of CRS ................................................................................................. 76 
APPENDIX G - Long Term Follow-Up ....................................................................................................................... 77 
 
 
 
 
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1. 
BACKGROUND 
This document is a clinical research protocol and the described study will be conducted in compliance 
with the IRB approved protocol  
 
Merkel cell carcinoma (MCC) is a highly aggressive skin cancer, with an incidence that has doubled 
since the year 2000 to over 2500 cases/year in the US.1 Over one-third of patients will develop 
widespread disease, and survival in these patients has been historically poor with a 5-year survival rate 
of <10%.2 MCC is very immune sensitive, with better immune responses being associated with better 
disease outcomes.3 This is due to the strong antigenicity (visibility to the immune system) of the cancer-
causing viral (Merkel cell polyomavirus; MCPyV) oncoproteins expressed in most MCC tumors.4,5 
Hence, immune therapies, particularly those targeting the viral oncoproteins may be effective for MCC 
treatment. Indeed, recent international studies led out of Fred Hutchinson Cancer  Center (Fred Hutch) 
have demonstrated that immune checkpoint inhibitors (ICIs) targeting the PD-1 axis (including 
pembrolizumab/Keytruda and avelumab/Bavencio) have efficacy, with one third to one half of the 
treated patients experiencing disease shrinkage and a small minority of patients (<10%) experiencing 
complete response.6,7  These demonstrate the potential for immunotherapy in MCC. However, for the 
majority of patients these treatments alone are ineffective and additional treatments are needed. We 
hypothesize that the patients who do not respond to checkpoint therapy alone have insufficient quantity 
of and/or poor-quality antiviral (and thus antitumor) T cells. Thus, we believe adding T cell therapies to 
the standard of care ICI avelumab 7-12 and administration of interferon gamma to facilitate T cell 
expansion (and increase the visibility of the tumor to the immune system) may be able to rescue patients 
who do not respond to an ICI alone. 
 
In our previous Fred Hutch protocol 9245, we have combined avelumab, class I HLA upregulation, and 
autologous bulk expanded antiviral T cells in patients with advanced MCC.13 Seven patients have 
received T cells as of October, 2018, with no unexpected toxicities. Encouragingly, three patients had 
durable complete responses (CRs) and a fourth patient had a non-durable partial response (PR). ICI have 
only recently become available for treatment of MCC, and treated patients on 9245 represent a mix of 
those with and without prior immune checkpoint inhibitor (ICI exposure). However, due to availability, 
accessibility, safety and cost considerations for the current protocol 9845/ATTAC we will focus on 
patients in second-line (failure of ICI monotherapy) where the clinical need is greatest and where there 
are no available FDA-approved treatments. 
 
Although previous protocols had success with bulk expanded/endogenous T cells, generation of these 
endogenous T cells is an expensive and slow process. In addition, it is difficult to achieve the required 
dose as patient-derived MCPyV-specific T cells are poorly avid and are rare or undetectable.5 Therefore, 
in the current proposed ATTAC regimen we are switching to a transgenic approach, which will allow us 
to deliver T cells to more patients, more cheaply, in greater numbers, and of higher quality (higher 
avidity).  Similar transgenic T cells with a different target protein have been delivered to individuals 
with other malignancies including non-small cell lung cancer, mesothelioma, and acute myeloid 
leukemia.14 
 
The present protocol, Protocol 9845: The ATTAC-MCC trial, is a phase I/II trial of the ATTAC regimen 
in patients with advanced MCC that has progressed on or after treatment with immune checkpoint (PD-1 
or PD-L1 inhibitor) therapy.  The ATTAC regimen is a combination of the FH-MCVA2TCR autologous 
transgenic CD8+ and CD4+ T cells carrying the transgenic TCR A2-MCC1 targeting the MCPyV, 
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avelumab or pembrolizumab (PD-L1 and PD-1 inhibitors approved by the FDA for treatment of Merkel 
Cell Carcinoma), and class I MHC upregulation in the tumor tissue with interferon gamma. In part I 
(until safety demonstrated in 3 consecutive participants), participants will receive a lower dose of T cells 
for first infusion (5-fold dose reduction from target dose).  If adequate safety profile is observed, we will 
proceed to part II; if adverse safety signal is seen, we will re-design trial in consultation with our DSMB 
and restart part I. In part II, participants will receive full dose of T cells for both infusions. We anticipate 
enrolling up to 16 HLA-A02+ participants with Merkel cell polyomavirus positive advanced MCC over 
2 years.  
 
We hope that study participation will provide direct benefits to the treated patients. This study will also 
benefit the research field for immunotherapy development for MCC and cancer more generally. Merkel 
cell carcinoma is an ideal cancer in which to develop immunotherapies due to the target of the viral 
oncoprotein which is immunodominant. This allows us to give T cells that are more on-target (with less 
risk of immune related adverse events; iRAEs) and also to 
more closely monitor immune responses to the cancer. If 
treatment is successful, as we are hopeful based on our 
early success with similar approaches as above, similar 
regimens could be expanded to other malignancies. If 
treatment is unsuccessful, our sophisticated 
immunological monitoring of the infused cells, native 
immune cells, and tumor tissues will allow for a cutting 
edge, comprehensive understanding of the mechanisms 
for tumor immune evasion that will provide essential data 
for developing new and better immunotherapies for MCC. 
 
2. 
STUDY PURPOSE AND RATIONALE 
2.1 
Rationale for Investigation of FH-
MCVA2TCR  
2.1.1 Scientific basis for T cell therapy targeting 
MCPyV in MCC 
MCC is a highly immune sensitive tumor. Patients 
with immune suppression have very poor outcomes3 
whereas those with brisk T cell responses have 
excellent outcomes15(Figure 1). Furthermore, 
immunotherapy is effective in MCC, with up to 50% 
of cases responding to PD-1 axis blockade (Figure 
2).6,7,10 
 
The majority of MCC tumors (>80%) are caused by the Merkel cell polyomavirus (MCPyV). 16 The 
wild-type virus is a frequent skin commensal on healthy adults,17-19 but it has been well demonstrated to 
play a causative role in virus-positive MCCs (VP-MCC). To become tumorigenic, MCPyV must 
undergo non-homologous recombination with the host cell genome and acquire an inactivating 
Figure 1: Improved MCC-Specific Survival in Patients with 
Brisk CD8+ T cell Infiltrate 
Figure 2: Response to first line avelumab (anti-PD-L1) in 
metastatic MCC10 
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secondary mutation to block 
viral DNA replication, which 
would otherwise lead to host 
cell apoptosis (Figure 3).16 
The integrated virus is thus 
not transmissible (analogous 
to human papilloma virus in 
most cervical carcinomas). 
With no DNA replication 
capacity, integrated MCPyV 
is resistant to standard 
antiviral agents. However, 
viral early proteins (T- 
antigens) remain highly 
expressed and promote cell 
cycle progression through multiple mechanisms including inhibition of the tumor suppressor 
Retinoblastoma protein.20 
 
In MCPyV-positive MCC tumors, the tumor cells are addicted to high expression of the viral 
oncoproteins (“T-Antigens”)(Figure 3) and these antigens thus cannot be readily lost.20 Importantly, the 
viral gene expression is only limited to the tumor tissue and is not present in healthy tissues, making 
these proteins a highly specific target for immunotherapies. In addition, these antigens are recognized by 
the immune system21 and are immunodominant.22-24 Patients with improved CD8+ T cell immune 
responses to the T antigen (as defined by quantity of T cells, number of reactive clones, and increased 
avidity of reactive T cells)  have improved disease specific outcomes including survival.5 Altogether, 
these antigens are compelling targets for tumor-specific immunotherapy. 
 
2.1.2 Target epitope: “KLLEIAPNC” MCPyV Epitope 
Effective T cell responses depend on the binding of the T cell 
receptor on the T cell surface to the class I HLA bound to viral 
peptide on the tumor cell surface. T cell receptor-based therapy is 
thus necessarily restricted to a single HLA type. For purposes of 
having the best data for safety and efficacy in the setting of a small 
trial, we will focus on a single epitope (T cell target) within the 
MCPyV-- the “KLL epitope.” This epitope (KLLEIAPNC) is in the 
shared/common portions of the MCPyV oncoproteins (effectively 
doubling its expression within any given cell), is well conserved 
across MCC tumors,25 and is not present in the human proteome by 
BLASTp search.  It is restricted by HLA*A0201 which is the most 
common HLA allele detectable in MCC patients and is found in 
>59% of virus positive cases (Table 1).  
 
2.1.3 
Clinical experience with T cells targeting MCPyV in MCC:  
A total of 12 patients have been treated on three separate trials with 
autologous ex vivo expanded T cells, class I HLA upregulation with 
intralesional interferon beta and/or single fraction radiation ).13,26  
Figure 3: Merkel Cell Polyomavirus and MCC. 
The majority of MCC tumors are caused by 
persistent expression of MCPyV oncoproteins. 
These are ideal targets for immune therapy as 
they are not expressed in non-tumor tissues. A 
representative tumor is shown at left, with 
MCPyV stain in brown color. 
Table 1: Epitopes in MCPyV T antigens. 
Class I HLA restricted epitopes in MCPyV 
oncoproteins are listed, with AA1-3 (first 
three amino acids). HLA restrictions and 
percent of VP-MCC patients (n=127) 
carrying allele listed. KLL (top row) is the 
target of TCR A2-MCC1 and was selected 
as it is the highest frequency target among 
VP-MCC patients and has been effectively 
targeted with endogenous cell therapy 
(detailed in 2.1.3) 
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Seven of these patients were additionally treated with avelumab, closely mimicking the proposed 
regimen (which differs in its use of transgenic T cells). In these trials (Table 1), Grade 3-4 T cell-related 
adverse events were as anticipated in both groups, and included transient lymphopenia and modest 
cytokine release syndrome lasting<48 hours, manageable on the general ward with supportive care 
alone. The safety profile was similar between patients who received avelumab and those that did not. 
There have been no treatment related deaths nor sustained grade 3 or IV toxicities. 
 
T cell persistence: Infused MCPyV specific autologous endogenous T cells persisted for years after 
initial infusion as confirmed by tetramer staining and demonstrated an effector memory (TEM) phenotype 
during sustained remission (Figure 4).  
 
Efficacy: Responses to T cell therapy to date for MCC are outlined in Table 2. In brief, in patients 
receiving endogenous T cells and HLA-upregulation, there was one objective response of 5 patients 
treated (ORR 20%). In patients receiving endogenous T cells, HLA upregulation and avelumab, there 
were 4 objective responses of 6 evaluable patients (ORR 67%). Patients had received between 0-3+ lines 
of prior systemic therapy prior to T cell therapy trial enrollment. Example responding patient shown in 
Figure 5. 
 
Table 2: Results of endogenous cell therapy to date in MCC. Results of double therapy protocols (2558/2586) 
and triple therapy protocols (9245) are shown.  Patient 2586-4 developed a subsequent late partial response 
lasting 22 months when immune checkpoint inhibitors were restarted after completing protocol treatment, disease 
had previously been immunotherapy refractory. 
Patient
Targeted 
Epitope(s)
HLA 
restrict.
HLA 
upreg. 
Method
Grade III-IV 
AE or II-IV 
CRS
Best 
respons
e 
Response 
duration 
(months)
2558-1
EWW
A*2402
IFN/XRT
Lymphopen
ia
PD
2586-1
KLL
A*0201
IFN
PD
2586-2
EWW & 
KLL
A*2402, 
A*0201
XRT
Lymphopen
ia
PD
2586-3
EWW
A*2402
IFN/XRT
CRS (gr. 2)
CR
13
2586-4
FPW
B*3502
XRT
CRS (gr. 2)
PD*
9245-1
EWW & APN
A*2402, 
B*0702
XRT
Lymphopen
ia
CR
24, ongoing
9245-2
KLL
A*0201
IFN
Lymphopen
ia
CR
24, ongoing
9245-3
KLL & FPW
A*0201, 
B*35
IFN/XRT
Lymphopen
ia, CRS (gr. 
CR
18
9245-5
KLL
A*0201
XRT
Lymphopen
ia
PR
3
9245-6
KLL
A*0201
IFN
PD
9245-7
EWW
A*2401
XRT
PD
9245-8
KLL
A*0201
XRT
Double therapy 
(HLA ⬆ + T cells)
Triple Tx (HLA ⬆+ T cells + 
anti-PDL1)
Treatment Ongoing
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2.1.4  Rationale for Transgenic T cell approach 
A transgenic approach offers following advantages over the prior 
endogenous T cell therapy approaches: time, potency, dose, 
number of patients, and cost. These are further detailed:  
a) Time to T cell product: On our previous ex vivo 
expanded T cell trial for MCC patients, the time to T cell 
product production ranged from 49-80 days. Given 
MCC’s rapid growth pattern, this delay is significant. A 
transgenic approach allows for T cells to be produced in 
less than one-quarter the time, approximately 10 days.  
b) Ability to deliver a more potent T cell product: With 
an ex vivo expanded T cell product, the avidity of T cells 
(ability of T cells to bind virus-expressing tumor cells) varies greatly between patients. Indeed, 
for MCC, there is a 3-order of magnitude range of antiviral T cell avidity. Furthermore, patients 
with the best outcomes (thus least likely to need our treatments) have the best/most avid T cells, 
and the patients with advanced disease generally have worse T cells.5 A highly avid T cell offers 
the best opportunity to “stick to” and kill a cell that has weak expression of the class I HLA 
genes, such as MCC tumor cells that are infected with MCPyV.27 By transgenically transferring a 
highly avid T-cell receptor specifically targeting MCPyV oncoproteins, we can improve the 
ability of transferred T cells to respond to tumor.  
c) Ability to deliver a larger number of transgenic T cells: Most endogenous cell products for 
MCC have not hit targeted dose and have instead ranged from 0.25-14% of targeted dose. As 
dose delivered is directly related to T cell persistence (which is thought to be essential for long 
term tumor control for T cell therapy,28 it is important to reach as close to target dose as possible. 
By starting with a larger number of cells and transducing/gene-modifying them, target doses are 
more achievable.  
d) Ability to reach patients that are not otherwise targetable: In a study of 69 virus positive, 
HLA-A02 positive patients with MCC, only 14% had detectable T cells targeting the MCPyV in 
their peripheral blood.5 The other 86% of patients would be not be treatable by ex vivo expansion 
but would be treatable by transgenic approaches.  
e) Reduced treatment costs: The ability to generate more potent T cell products in a relatively 
short-time and in a large scale will also ultimately reduce the cost associated with treatment.  
Figure. 5: MCC regression after TT. CT 
scans of irradiated and non-irradiated 
lesions are shown before and 8 mos after 
therapy began for Pt 9245-1, who achieved 
a biopsy-proven CR, ongoing at 24 mos  
Figure 4: (A) persistence of MCPyV-
specific T cells. %Tet+ CD8+ in peripheral 
blood (y-axis) after the first infusion (x-axis). 
The limit of detection (0.01%) is indicated. 
(B) Phenotype and function of transferred 
cells. Analysis of Pt 9245-3, on day 203 after 
transfer, showing % expression of TCM (RA–
,R7+), TEM (RA–,R7–) and TTD (RA+, R7–) 
(yellow), PD-1 (red) and %Tet+ cells 
producing IFN, TNF and IL-2 (green) in 
response to cognate antigen. 
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2.1.5 Transgenic TCR vs. Transgenic CAR 
Given the unqualified success of gene-modified CAR T cells in the treatment of lymphoid leukemias,29 we 
carefully considered gene modified T cell therapy for MCC. T cell receptors (TCRs) and chimeric antigen 
receptors (CARs) have their distinct advantages and disadvantages for engineering therapeutic T cells.30,31 We 
opted to design our MCC therapy with TCRs as the MCPyV T antigens have nuclear expression. We opted to 
design our MCC therapy with TCRs as the MCPyV T-Ag antigens have nuclear expression. Such intracellular 
targets are inaccessible to a CAR, whereas MCPyV peptide epitopes can be processed and presented to a TCR 
via HLA Class I. Additionally, naturally occurring TCRs, even high affinity TCRs, having survived negative 
selection during normal development in a human thymus that deletes self-reactive TCRs, further reducing the 
likelihood for an autologous T cell expressing such a TCR to mediate off-target toxicity.32,33 Other groups are 
pursuing high-affinity TCRs for MCPyV, which have been shown to be effective in mouse models,34 
supporting the general approach. However, these TCRs were identified in mice, and such artificially derived 
TCRs have been associated with toxicity in other disease contexts.35 Thus, we believe by exploiting the in vivo 
safety screen provided by the human thymus, we will have a safer naturally-isolated TCR product for our trials. 
 
2.1.6 Safety of TCR transgenic T cells for other cancers 
All prior MCPyV targeting T cell trials have been with non-transgenic T cell product. However, we have 
experience in preparing and delivering transgenic T cells expressing natural human transgenic TCRs 
isolated using similar approaches and in the same clinical vector in other disease processes. In clinical 
trial # NCT01640301,36 we have treated 24 acute myeloid leukemia patients post allogeneic stem cell 
transplant with transgenic CD8+ T cells targeting the WT1 antigen. Apart from expected grade 1-2 
cytokine release syndromes (<24 hour duration) and transient lymphopenia (<10 days), no immediate or 
delayed off-target or on-target/off-tissue toxicity was observed, and clinical benefit observed.37  
 
2.1.7 TCR A2-MCC1 is a highly potent natural TCR that effectively lyses MCC cell lines 
FH-MCVA2TCR cells are autologous CD8+ and CD4+ T cells gene modified to carry the TCR A2-
MCC1, a high affinity T cell receptor identified from the natural repertoire from a healthy individual.  
For an effective TCR for clinical translation, we sought several features, most notably high-affinity (to 
generate highly avid T cells better able to stick to and recognize MCC tumors that have weak antigen 
presentation), safety (as further detailed below), and ability to recognize MCPyV variants.  We screened 
20 TCRs and identified the best candidates that were isolated from three approaches: the highest affinity 
TCR from traditional cloning from MCC TIL (TCR A2-Trad), the highest affinity TCR from an 
endogenous cell therapy product that eradicated an MCC tumor (TCR A2 9245-3), and a screen of the 
peripheral repertoire of 10 healthy HLA-A2 blood donors (TCR A2-MCC1) (Figure 6). Although both 
TCR A2 9245-3 and TCR A2-MCC1 demonstrated much higher affinity than those identified by 
standard cloning (Figure 6E), TCR A2-MCC1 was chosen for clinical translation. This is because TCR 
A2-MCC1 exhibited superior killing of MCC cell lines in vitro (Figure 6F) and recognized a frequent 
variant MCPyV sequence KLLEISPNC25 in addition to the standard sequence KLLEIAPNC obviating 
the need for tumor DNA sequencing to confirm MCPyV epitope presence. 
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2.1.8 Evaluation of potential for off-target activity and cross or 
allo-reactivity of TCR A2-MCC1  
An alanine scan38 of target peptide recognition performed for TA2-MCC1 
identified four residues not critical for binding to the TCR (Figure 7). A 
BLAST search of the human genome for peptides with allowed 
substitutions at each non-critical residue identified 10 potentially 
recognizable self-peptides,39 none of which elicited responses.  In 
addition, TA2-MCC1 elicited no response to six HLA-A2+ normal tissue cells 
and a panel of HLA-mismatched lymphoblastoid cells, indicating lack of 
cross- and allo-reactivity, respectively (Table 3).  
 
2.1.9 Rationale for using a third-generation lentiviral vector  
The theoretic risk of insertional mutagenesis with retroviral vectors 
became realized during two X-linked severe combined 
immunodeficiency gene therapy trials in which five patients developed acute lymphoblastic leukemia, 
shown to reflect transactivation of either the LMO2 or CCND2 gene by the retroviral insert in 
transduced CD34+ hematopoietic stem cells (HSCs). This highlighted the propensity of gamma-
retroviral vectors (γ-RV) to preferentially insert near genes that are actively transcribed, which may be 
particularly problematic in transduced HSC that express genes that confer self-renewal capacity and a 
proliferative/survival advantage and are then transplanted into a setting in which they are driven to 
extensively expand.40 
 
 
Figure 7: Alanine scan of TCRA2-MCC1 reveals absence of cross reactivity. Left panel: results of alanine scan reveals non-
critical residues.  Center panel: Results of BLAST search identifies 10 potentially cross-reactive peptides in human proteome requiring further 
evaluation.  Right panel: absence of detectable cross-reactivity to any of the 10 identified peptides. 
Figure 6. High-throughput TCR isolation. (A) The top 2% of cells are selected, using serial sorts at decreasing [tet.]. (B) Ratios 
(sort:pooled line frequency) identify clonotypes enriched with decreasing tetramer concentrations (red line –TCRA2-MCC1). (C) TCR enrichment 
ratio (y-axis) inversely correlates with clonotype frequency in total pool. (D) EC50 of IFN secretion to decreasing [peptide] for T cell clones 
isolated from MCC pts with different clinical outcomes (adapted from 17). Clinical outcomes: Rapid- rapid progression, Slow - slow 
progression, NED - no evidence of disease. Circle identifies TA2-trad. (E) Tet binding (affinity) and (F) Specific lysis of MCC Waga cell line by 
TCRA2-trad.(black), TCRA2-9245-3 (blue) and TCRA2-MCC1 (red). 
Table 3: TA2-MCC1 cross- and allo-
reactivity. 
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By contrast, the risks of insertional oncogenesis or other vector-related cellular toxicities are extremely 
low when targeting peripheral blood T cells that lack the same self-renewal capacity of HSC, with no 
significant toxicities reported to date in any clinical trial. As described below, this risk can be further 
decreased with the use of a third-generation self-inactivating (SIN) lentivirus (LV), in which the 
promoter regions of the long terminal repeats (LTRs) have been truncated and the insert, such as a TCR 
gene, is expressed under control of an internal murine stem cell virus (MSCV)-based promoter.41 
 
Insertional oncogenesis was not observed in the long-term results from three clinical trials evaluating γ-
RV-engineered T cells for patients with HIV.42 The transduced cells persisted long term, in some cases 
>10 years, and clinical monitoring of the patients at yearly intervals for cumulatively >540 patient-years 
of observation has not detected evidence of retroviral genotoxicity. In more recent trials with less 
extensive retrospective data, no toxicities attributable to the administration of T cells transduced to 
express HLA*0201-restricted MART1 or gp100-specific TCRs using the MSGV1-based RV were 
observed in 33 patients with metastatic melanoma. The MSGV1-based RV uses the same promoter from 
the LTR of MSCV that will be used in the lentiviral vector in our trial.43 The MSGV1-based γ-RV was 
also used to transduce autologous T cells to express an NY-ESO-1-specific TCR that was infused in 17 
patients with metastatic synovial sarcoma or melanoma43 and to transduce cells to express a 
carcinoembryonic antigen (CEA)-specific TCR in T cells that were infused into three patients. Again, no 
insertional mutagenesis events were identified.  
 
Transcriptionally active enhancer/promoter elements may influence, expression of cellular genes at a 
distance from the insertion site, independently of the vector type used to introduce the gene (RV vs LV; 
LTR-based or SIN).44 However, γ-RVs have a predilection toward integrating in the immediate 
proximity of transcription start sites and deoxyribonuclease I hypersensitivity sites, increasing the 
probability that the viral LTR transcriptional enhancer will interfere with gene regulation and potentially 
activate cancer-causing genes.45 In contrast, LVs commonly insert into introns, and thus are more likely 
to integrate further away from the transcription start sites of active transcription units, making them less 
likely to induce transcriptional activation.46 In vivo genotoxicity assays based on the transplantation of 
transduced tumor-prone Cdkn2a-/- murine hematopoietic progenitor cells have also directly compared 
the effect of promoter location within γ-RVs and SIN lentiviral constructs on the oncogenic potential. 
Placing the strong LTR-based spleen focus-forming virus promoter as an internal promoter in a 
construct containing a SIN LTR rather than as a component of the LTR further reduced the propensity 
for insertional mutagenesis and lymphoid tumors.47   
 
Importantly, >1000 patients have been treated with TCR transgenic or CAR-T cells on protocols 
nationwide that include T cell modifications with similar vectors.  A recent review in the New England 
Journal of Medicine determined that among these >1000 individuals no occurrence of an oncogenic 
transformation has yet been reported.29 
 
Despite the risk of insertional mutagenesis being extremely low, it remains justified to initially examine 
safety/toxicity and potential efficacy of therapy with transduced T cells in patients with MCC. The 
potential toxicities are listed in the protocol consent form and will be discussed with patients as a part of 
the consent process. Eligible patients who elect to enroll on the protocol will be monitored and managed 
for potential toxicities as per current guidelines, which are outlined in Section 12. Stopping rules will 
also be applied, as described in Section 13. 
 
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2.1.10 Rationale for substrate T cell of CD62L+ (“young” but well differentiated) CD8+ T Cells 
Establishing a persistent functional population of antigen-specific T cells after adoptive transfer will 
likely be necessary to eliminate MCC cells and prevent recurrence. Previous trials of T cell therapies in 
multiple malignancies have often been limited by the inability of transferred T cells to expand and 
persist post- transfer. The in vivo fate of transferred T cells is dependent in part on the intrinsic 
properties of the T cells from which the infused cells are derived,48,49 and we can choose the T cell type 
used for adoptive cell therapy.  
 
Conventional CD8+ T cells can be divided into naïve and antigen-experienced, memory T cell subtypes 
(TN, TM). TM cells can be further divided into TCM and effector memory T cell (TEM) subsets, which 
have distinct transcriptional programs associated with defined characteristics, including homing, 
phenotype, and function. When the TEM cells are stimulated in vitro, they differentiate largely into short-
lived effector cells that kill targets, but generally have limited proliferative capacity and fail to persist 
for long periods of time after transfer.50 
 
Studies of transferred purified CD8+ T cell subsets in a murine virus infection model revealed that 
transferred TCM provide enhanced protective immunity from in vivo challenge compared to TEM. 
Although TCM cells expand and also differentiate into effector cells in response to in vitro stimulation, 
adoptive transfer studies in non-human primates revealed that TCM–derived effectors had been 
imprinted and retained some of the beneficial properties of their parent TCM cell, in particular the 
capacity for self-renewal, which translates into improved in vivo persistence and response to antigen 
challenge.49 
 
For this proposed trial, we will employ CD62L expressing CD8+ T cells. This will include lymphocytes 
subsets that are TCM, TN and TSCM (“young” but well differentiated, with increased persistence potential). 
In exploratory analyses, we will determine which subset of cells was associated with best persistence 
and tumor localization and use this information to help to better refine T cell therapies.  
 
2.1.11 Rationale for inclusion of transgenic CD4+ T cells in addition to transgenic CD8+ T cells 
TCR-based immunotherapies have primarily exploited CD8+ T cells, which recognize tumor antigens in 
the context of Class I presentation/restriction.51 However, co-transferred CD4+ T cells can provide major 
direct and indirect anti-tumor effects, including promoting expansion and survival of tumoricidal CD8+ 
T cells, as we showed long ago in an FBL murine leukemia model.52 More recently this has been shown 
with therapeutic CD19-CAR T cells,53,54 and supports the concept of engaging both subsets to target the 
same tumor. Tumor-specific, HLA Class II-restricted CD4+ T cells promote Class I-restricted CD8+ T 
cell proliferation, survival, and effector functions in part by producing IL-2, and facilitate activation of 
dendritic cells (DC) for de novo 
immune responses.38,55 CD4+ T cells 
expressing Class II-restricted TCRs 
exhibit direct cytolytic activity in 
metastatic melanoma and have had 
anti-tumor activity against human 
cholangiocarcinoma.56,57 However, 
Class II expression is rare in solid 
tumors58 and absent in MCC. An 
alternative might be to confer 
Figure. 8. TCRA2-MCC1 engages CD4+ T cell effector mechanisms. (A) Cytokine 
production after cognate antigen encounter of TA2-MCC1 CD4+ and CD8+ T cells. (B) CFSE 
dilution of proliferating tCD4+ day 5 post stimulation (red=TCRA2-MCC1, blue= TCRA2-9245-3). 
(C) Specific lysis by TA2-MCC1 CD4+ (dashed line) and CD8+ T cells (solid line) of MCPyV 
expressing fibroblasts.
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function to CD4+ T cells through expression of a high-affinity Class I TCR (TCRClass-I), that would co-
localize the engineered CD8+ and CD4+ T cells on the same tumor target without requiring Class II 
expression. These T cells can both perform traditional CD4 functions (production of T cell supportive 
cytokines, DC activation) and have direct tumoricidal effect.  Similar approaches have been safely 
employed for melanoma by other groups, in T cell therapy trials targeting the MART-1 antigen.59 
 
TCR A2-MCC1 can engage CD4+ T cells even when transduced in absence of CD8+ coreceptor. CD4+ 
T cells carrying TCR A2-MCC1 can produce beneficial cytokines (Figure 8A), proliferate (Figure 8B), 
and kill MCPyV oncoprotein expressing cells (Figure 8C).  Therefore, FH-MCVA2TCR will include 
both CD8+ and CD4+ T cells transduced to carry TCR A2-MCC1. 
 
2.1.12 FH-MCVA2TCR Dose Rationale 
Our prior endogenous cell therapy trials targeted an infused dose of 10 billion (1010) MCPyV-specific 
endogenous CD8+ T cells per square meter, or approximately 20 billion (2*1010) CD8+ T cells per 
infusion. No dose limiting toxicities were observed and the maximum actual infused dose was 26 billion 
for protocol 2586 (HLA upregulation + T cells) and 17.7 billion for protocol 9245 (HLA upregulation + 
T cells + avelumab).   
 
For the present study, our goal dose is 1 billion (109) MCPyV-specific TCR transgenic CD8+ T cells for 
the initial low dose part I infusion, and 5 billion (5x109) MCPyV-specific TCR transgenic CD8+ T cells 
for the second dose infusion or part 2 infusions. CD8+ and CD4+ cells are co-cultured and transgenic 
CD4+ cells will also be included. The dosing will be based on CD8+ T cells for consistency in 
formulation. Please see Section 6.4 for additional dosing information. This dose was selected for several 
reasons: it is achievable in an approximately 2 week production, it is similar to dosing for TCR 
transgenic T cells for other disease processes, and it is similar to doses given to 3 previous patients on 
previous versions of this trial with no toxicity attributed to the transferred T cells. All patients included 
in this study will be adult and are thus expected to have similar body surface areas, and thus we have 
selected a flat rather than weight-based dosing; this approach is used with dosing with that of other 
transgenic T cell products for adults including FDA-approved CAR-T therapies (for example, Kymriah).  
 
 
2.1.13 Rationale for omission of Lymphodepleting Chemotherapy prior to T cell infusion 
Chemotherapy lymphodepletion, typically with fludarabine and cyclophosphamide, is frequently used 
for transgenic T cell immunotherapy trials, in order to induce a favorable cytokine environment for T 
cell engraftment and promote expansion and persistence.60,61 We have elected to omit this chemotherapy 
conditioning for several reasons. 
a. Safety: Cytokine-mediated toxicity of transgenic CAR-T therapies is increased in the context of 
lymphodepletion,61 and it is plausible that such toxicities could extend to transgenic TCR 
therapy. It is also possible that depletion of endogenous T cell responses to the tumor could lead 
to faster progression of disease.  One subject received lymphodepleting chemotherapy in a 
previous version of this trial, and concerns for increased rapidity of disease progression 
following lymphodepletion compared to before were noted. 
b. Non-necessity: T cell persistence has been demonstrated for patients with MCC out to years 
without preceding lymphodepletion (Figure 4), and a patient treated on an earlier version of this 
trial achieved a clinical objective response without lymphodepleting chemotherapy. Two patients 
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who received lymphodepleting chemotherapy had inferior expansion and persistence of 
transgenic T cells in an earlier version of this trial.  
c. Potential to abrogate epitope spreading: Epitope spreading has been observed in multiple patients 
(Figure 9) and we believe will be essential to combat immunotherapy escape by antigen loss, a 
common mechanism of escape for TCR and CAR-T cell therapies.29 
Chemotherapy has potential to reduce epitope spreading by depleting 
endogenous T cells.  
 
2.2 
Rationale for inclusion of subcutaneous interferon gamma 
2.2.1 
Scientific basis 
As our therapy utilizes a transgenic TCR, we are dependent on antigen 
presentation by the primary tumor through class I MHC. We and others have 
shown that approximately 80% of MCC tumors downregulate MHC-I presenting 
an obstacle to T cell efficacy, and this is further true of tumors that have escaped 
immunotherapy.27,62 In 5 patients previously treated in an earlier version of this 
clinical trial, the only patient with a meaningful clinical response was the single 
patient with robust baseline class I MHC expression.  Furthermore, that patient 
had loss of class I MHC expression in a single resistant lesion.  Despite the small 
number of patients, this suggests that downregulation of class I MHC may be a 
key mechanism of resistance to transgenic TCR T cell therapy. However, this 
class I downregulation is typically reversible with one of several interventions, which include radiation 
treatment, epigenetic modifiers, chemotherapy, and cytokine treatments.27 Earlier versions of this trial 
included either single fraction radiation to a subset of lesions (n=3 patients) or chemotherapy treatment 
(n=2 patients) without clinical efficacy in patients without baseline class I MHC expression.  This 
indicated a different strategy was necessary to efficiently upregulate class I MHC in tumors. 
 
2.2.2 Interferon Gamma- Preclinical results 
In order to identify molecules that might be effective at clinically meaningful upregulation of class I 
MHC, a merkel cell line WAGA was treated with multiple either FDA-approved or investigational small 
Figure 9: Epitope spreading. % IFN-
gamma secreting (A) CD4+, Patient 
9245-1 peripheral blood mononuclear 
cells (PBMCs) and (B) CD8+ Patient 
9245- 3 TIL, exposed to cT, sT and LT 
peptide pools. Black arrows indicate 
infused CD8+ specificities; red arrows 
indicate expanded/new reactivities.
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molecule agents thought to have class I upregulating effects, and interferon gamma showed the most  
robust upregulation of class I MHC on these cells.  We found that the most dramatic effects on class I 
upregulation were seen with interferon gamma (Figure 10A), and these effects were seen at 
concentrations that could realistically be achieved in vivo with standard dosing (Figure 10B).  
Importantly, interferon gamma not only upregulated class I MHC but also allowed for recognition of 
endogenous antigen in Merkel cells in vitro (Figure 10C). Importantly, mimicking conditions that 
would be seen in vivo with standard dosing of interferon gamma (Peak concentration 0.6ng/ml, half life 
of 6 hours) but using 0.6ng/ml for 6 hours and 0.3ng/ml for 6 hours followed by washout showed robust 
upregulation of class I MHC across 4 different Merkel cell carcinoma cell lines (Figure 10D). 
 
2.2.3 Interferon gamma- safety and clinical results 
Interferon gamma is FDA-approved for the treatment of chronic granulomatous disease at a dosage of 
50mcg/m2 given three times weekly, and is generally tolerated with the known common (>10%) side 
effects of fever, headache, chills, fatigue, rash, diarrhea, vomiting and injection site reactions.  The peak 
concentration of interferon gamma occurs in 7 hours and is 0.6ng/ml in plasma, with a half-life of 6 
hours.  Importantly this concentration is associated with significant class I MHC upregulation in Merkel 
cell carcinoma cells in vitro (Figure 10B).  Use of subcutaneous interferon gamma in cancer patients 
has been explored in a previously reported phase 0 trial conducted in sarcoma patients.63  In this trial, 
administration of interferon gamma was well tolerated and resulted in upregulation of class I MHC in 
patient tumors and increases in measures of tumor inflammation.  Interferon gamma at standard dosing 
has been given in combination with the PD-1 inhibitor nivolumab in a phase I clinical trial of over 20 
patients with no evidence of increased immune related adverse events related to nivolumab (no cases of 
 
 
Figure 10: HLA upregulation by interferon gamma in Merkel cell carcinoma cell lines 
WAGA merkel cell line was incubated with the indicated agents for 48 hours and Class I MHC expression was measured by flow cytometry 
(A-B) and recognition of cells by the TCR-T cells was measured with and without addition of exogenous peptide (C).  D. Each of 4 indicated 
Merkel cell lines were incubated with either a stable, high concentration of IFNg (10ng/ml) or mimicking of in vivo pharmacokinetics with 
0.6ng/ml for 6 hours, followed by 0.3ng/ml for 6 hours, followed by washout and class I MHC expression was measured after 48 hours by 
flow cytometry 
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grade III or greater immune toxicity during combination treatment) or potentiation of side effects from 
interferon gamma (Matthew Zibelman, personal communication, manuscript in preparation). 
 
2.3 
Rationale for administration of TCR-transgenic T cells in the context of treatment with 
avelumab or Pembrolizumab as a standard of care  
2.3.1 Avelumab and Pembrolizumab - Scientific basis 
Our data from current T cell therapy trials, protocols 2586 and 9245, strongly support inclusion of 
avelumab. In protocol 2586, combining autologous ex vivo expanded anti-MCPyV T cells with HLA 
upregulation but without immune checkpoint blockade for MCC, we observed only one patient out of 
four responding. We further noted that the patient who responded had much lower PD-1 expression on 
their infused T cells, as compared to patients who did not respond. We thus hypothesized efficacy may 
be augmented by combination treatment 
(Figure 5), combining three immunotherapies 
(“triple therapy”): T cells, class I MHC 
upregulation, and PD-L1 blockade with 
avelumab. Indeed, in protocol 9245 combining 
T-cells with HLA-upregulation and avelumab, 
we noted improved response rate with 4 of 6 
patients responding, including several durable 
complete responses. We have noted no 
worsened toxicity with addition of avelumab. 
 
Multiple ICIs have been studied in MCC, 
including avelumab (anti-PD-L1), 
pembrolizumab, and nivolumab (anti-PD-1). 
All are showing clinical effect, and all would 
be expected to synergize with T cell therapy. 
Both Avelumab and Pembrolizumab have been 
FDA approved as standard of care treatments in 
MCC, and for this reason, they have been 
chosen as two alternatives for use in this trial. 
 
2.3.2 Avelumab - Clinical experience & safety 
Avelumab (trade name Bavencio) is a recently FDA-approved fully humanized monoclonal antibody 
targeting PD-L1 with indications for Merkel cell carcinoma and advanced bladder cancer.7,64 As 
described above, we believe the addition PD-1 axis blockade to T cell therapy essential due to high PD-1 
expression on infused T cells and improved efficacy signal combination therapy. Major side effects 
include: infusion reactions, immune related adverse events (iRAEs), and other.  Infusion reactions are a 
well-characterized side effect of infusional antibody therapy and occur with 17-22% of patients 
receiving Bavencio7. Consistent with package insert/manufacturer’s guidelines, patients will be 
premedicated with acetaminophen and an H1 blocking antihistamine prior to each infusion, infusions 
will be given slowly, and patients will be monitored closely for the development of infusion reaction. 
iRAEs are a well-described complication of any PD-1 axis blocking agent; for avelumab the most 
commonly reported include pneumonitis, colitis, hepatitis, endocrinopathies, and type 1 diabetes.  
Thyroid endocrinopathies are the most common iRAE (occurring in up to 6% of patients) and will be 
closely monitored for with regular blood work according to manufacturer’s recommendations. 
Table 4:  Toxicities of avelumab observed among 88 patients with 
advanced MCC treated as part of the JAVELIN-200 trial 
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Pneumonitis is the most severe reported complication. Among all patients receiving avelumab, 
pneumonitis is reported per FDA labelling insert as occurring in 1.2% (21/1738) of patients receiving 
avelumab. There has been one reported case (0.1%) of a patient with Grade 5 pneumonitis resulting in 
death, one (0.1%) with Grade 4, and five (0.3%) with Grade 3 pneumonitis. Immune-mediated 
pneumonitis led to permanent discontinuation of avelumab in 0.3% (6/1738) of patients.  Finally, other 
side effects include fatigue, arthralgias, and headache (see table 4).  Avelumab has been successfully 
given with ex vivo expanded T cells in 6 MCC patients, with no increased toxicity seen.13 As detailed 
below, avelumab dosing will be given at standard dose level and interval. 
 
Pembrolizumab is a fully humanized anti PD-1 antibody also FDA approved in MCC.  It has a similar 
incidence of immune related adverse events to avelumab in trials of MCC patients and has a lower 
incidence of infusion reactions than avelumab.  It has also been given safely in combination with 
adoptive transfer of Chimeric antigen receptor modified T cells in several trials65-67, suggesting it is 
likely to be safe in combination with adoptive T cell transfer in this trial. 
 
2.4 
Rationale for inclusion of biopsies 
Serial biopsies are very informative for determining whether T 
cells are preferentially localizing into the tumor environment and 
interactions between tumor and microenvironment. These can 
help to guide future studies. If treatment is effective, features of 
tumor-enriched T cells can be studied to optimize future T cell 
therapy products, and TIL can be investigated for epitope 
spreading. If the treatment is ineffective, study of the tumor 
microenvironment will allow determination of the reason for 
treatment failure and whether it is a tumor intrinsic or T cell 
intrinsic defect. We have developed a suite of tools to optimize 
information from small biopsies (multiplex 
immunohistochemistry, single cell RNA sequencing) in order 
to maximize benefit from tiny clinical specimens and reduce 
risk to research participants (Figure 11 and 12). 
 
3. 
STUDY OBJECTIVES  
3.1 
Primary Objectives 
3.1.1 
Safety 
To evaluate the safety of adoptive T cell therapy using FH-
MCVA2TCR (defined as an observed treatment-related 
grade 3 or higher toxicity rate consistent with a true rate 
<= 40%.) 
Patients who have metastatic MCC that has progressed beyond 
treatment with first line PD-1 inhibitors have few therapeutic 
options and no FDA-approved options. Commonly used 
treatments in this scenario include platinum-based doublet 
chemotherapy which has guarded efficacy and a 30 to 40% 
grade ≥ 3 toxicity rate.68 In terms of non-chemotherapeutic 
options, combination ICI immunotherapy (such as CTLA-
4/PD-1, ipilimumab + nivolumab) is occasionally used. This 
combination has not been well studied in MCC but carries a 
Figure 11: Multispectral IHC. A MCC tumor 
is shown. Multicolor IHC permits multiple 
studies to be done on a FFPE slide, maximizing 
information from precious biopsy specimens. 
Figure 12: Single Cell RNA sequencing identifies 
mechanism of T cell escape in an MCC tumor. 
Expression of the relevant gene is shown by orange 
color, with darker color indicating increased 
expression. Each tumor cell is indicated by a dot. 
This patient was treated with MCPyV targeting T 
cells restricted to ah HLA-B3502 epitope. After 
initial 22 month response to treatment the tumor later 
progressed despite persistence of infused T cells, and 
the tumor tissue was found to have lost the HLA-B 
expression necessary for T cell recognition 
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55% grade 3-4 toxicity rate in metastatic melanoma.69 Therefore, a toxicity rate of 40% is acceptably 
safe in this high-risk population with advanced disease that without treatment is uniformly fatal.  
 
Please see biostatistical section for further details on toxicity assessments.  
 
3.1.2 Efficacy. 
To demonstrate clinical activity of FH-MCVA2TCR (defined as an observed response rate that 
statistically exceeds 5%) 
Without treatment, metastatic MCC is near uniformly fatal. Spontaneous regressions have been reported 
in up to 1% of patients, therefore we will conservatively define a comparison response rate as 5% to 
allow for this and also for the possibility of very delayed checkpoint inhibitor response (although these 
have not been reported in MCC).6,7 There are no FDA-approved or known durably effective treatments 
for patients with advanced MCC tumors refractory to checkpoint blockade (hence the rationale for 
pursuing this population). Therefore, success in even a minority of patients would significantly advance 
the field. 
 
Please see biostatistical section for further details on efficacy assessment and power calculation. In brief, 
in order to meet this endpoint, at least 3 of 16 patients will need an objective response (best response of 
partial response or complete response) by RECIST 1.1 criteria.70 
 
3.2 
Secondary Objectives 
3.2.1 Persistence  
Persistence of infused transgenic T cells in peripheral blood 
3.2.2 Migration  
Migration of infused transgenic T cells into tumor tissue 
3.2.3 Progression free and overall survival 
Determine progression free and overall survival of treated individuals 
3.2.4 Determination of preferred dose of FH-MCVA2TCR 
Please see section 4.4 for details on dose escalation 
3.2.5 Response rate by immune RECIST 
Determine objective response rates by immune related RECIST criteria. Please see Hodi et al 
2018 for definition of iRRC criteria. 71 
 
3.3 
Exploratory Objectives 
Our study includes a number of exploratory objectives. These include phenotype of infused T cells that 
persist and localize to tumor, functional capacity of infused transgenic T cells, determination of the 
presence or absence of epitope spreading (broadening of immune responses) to other MCPyV and/or 
neoantigen epitopes in the peripheral blood and/or tumor environment, and evaluation of changes in the 
tumor tissue and microenvironment that may correlate with success and/or failure of the ATTAC 
regimen. These exploratory objectives are not anticipated to result in direct conclusions but instead 
develop hypotheses for future trials. 
 
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3.4 
Projected Target Accrual 
 
Table 5: Targeted / Planned Enrollment 
Ethnic Category 
            Patient Numbers                  
Females
    Males 
Total 
Hispanic or Latino
0
1
1
Not Hispanic or Latino
6
9
15
Ethnic Category Total of All Patients
6
10
16
Racial Category 
American Indian / Alaska Native
0
0
0
Asian
0
0
0
Native Hawaiian or Other Pacific 
Islander
0 
0 
0 
Black or African American
0
0
0
White
6
10
16
More than One Race
0
0
0
Racial Categories:  Total of All 
Patients 
6 
10 
16 
 
All racial groups and ethnicities will be included and participation of members of under-represented 
groups welcomed. These targeted/planned enrollment numbers are based on relative percentages 
of race/ethnicity reported for MCC cases in the Surveillance, Epidemiology, and End Results 
(SEER) cancer registry database.1 There is a white predominance due to the well described MCC risk 
factor of UV exposure.72 
 
Sex/Gender 
Males  
 
 
 
 
 
 
63% 
Females  
 
 
 
 
 
 
37% 
 
Ethnicity 
Hispanic/Latino 
 
 
 
 
 
5% 
Not Hispanic/Latino  
 
 
 
 
95% 
 
Race 
American Indian/Alaska Native 
 
 
 
<1% 
Asian  
 
 
 
 
 
 
<1% 
Native Hawaiian/Pacific Islander 
 
 
 
1% 
Black/African American 
 
 
 
 
1% 
White  
 
 
 
 
 
 
96% 
 
4. 
STUDY DESIGN AND INVESTIGATIONAL PLAN 
4.1 
Design: Phase I/II single-arm dose-escalation non-masked interventional study 
The study will be a phase I/II, non-masked (not blinded), interventional study aimed at treating up to 16 
HLA*A0201 patients with metastatic MCC that has progressed despite immune checkpoint inhibitor 
therapy (PD-1 or PD-L1 inhibitor). All patients will receive study interventions.  
 
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Patients may receive the other, standard-of-care portions of the ATTAC regimen in addition to T cells. 
This includes the PD-L1 inhibitor avelumab. Please see Section 6/treatment plan for further details). 
 
Patients will receive up to two T cell infusions. Please see 6.4.4 for details regarding selection of one 
versus two cycles of T cell therapy. 
 
Any patient experiencing treatment related grade 3 or 4 toxicity (except for the expected toxicity 
specifically excepted/outlined in Section 12.6) will immediately stop receiving T cell infusions, 
avelumab, and class I HLA upregulation. They will receive treatment as appropriate for their condition 
(see Section 7 and Section 12). They will continue to be followed for response/outcome.  Patients with 
infusion reactions to avelumab may receive pembrolizumab for first or subsequent infusions. 
 
As a criterion for enrollment into the trial, participants must agree to biopsies of tumor tissue if 
medically feasible. A minimum of 4 biopsies will be performed as clinical status allows: prior to 
interferon gamma administration, 1-2 days prior to T cell infusion, 2 weeks after, and 12 weeks after the 
first full dose T cell infusions. However, should there be no tumor tissue that is accessible for biopsy, 
patients will still be considered for participation at discretion of the investigator. Similarly, should an 
investigator or treating physician determine that a biopsy cannot be performed safely for clinical reasons 
(e.g., no safe approach), biopsies may be cancelled or retimed at the investigator or treating physician’s 
discretion. 
 
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4.2 
Schematic 
 
 
 
 
4.3 
Protocol Enrollment 
Enrollment to study commences after eligibility confirmation and just prior to leukapheresis. Enrollment 
is expected to take approximately 2 years. Study treatment with initial follow-up is expected to be 
approximately 1 year. All patients who receive treatment with FH-MCVA2TCR should be followed for 
at least 15 years after the final infusion for safety evaluations in the long-term follow-up (LTFU) portion 
of the study. The anticipated duration of the study, excluding LTFU assessments, is approximately 3.5 
years. 
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4.4 
Staggering and Dose Escalation 
At least three patients will be treated on part I.  In part I, patients will be treated with a lower-dose first 
infusion.  Each part I patient must complete their dose-limiting-toxicity (DLT) evaluation period (28 
days) after their first T cell infusion before the next part I patient may receive their first infusion. This 
allows for an ample buffer/window to evaluate for potential cytokine release syndrome and/or 
neurotoxicity, before proceeding to next patient. After three patients have received their first T cell 
infusion and completed the 28 day DLT evaluation after the lower first dose infusion, with one or fewer 
patients experiencing grade 3-4 d related adverse events as defined in Section 12, the study may move to 
part II. If two or more of the first three patients have a treatment-related grade 3 or 4 toxicity, or if any 
of the first three patients experiences a treatment-related death, at that point the trial will be immediately 
suspended. We will consult with the data and safety monitoring board (DSMB), and a detailed review of 
the potential causes of toxicity undertaken. We will reconvene with the DSMB and consideration will be 
given to changing the cell dose or construct if the T cells were implicated. In that scenario, the dose 
escalation will be restarted and an additional three patients treated before proceeding to the part II 
portion of the study. Part I will continue until 3 patients are treated at the lower dose level.  If the 
manufacturing process is short of the intended dose targeted for the patient, whatever smaller dose is 
available from a single manufacturing run will be given. 
 
For the first three patients treated on part II, the staggering interval will continue as for part I, with each 
treated patient completing their 28-day DLT evaluation period after first T cell infusion before the next 
part II patient may be treated at that dose. If two or more of the first three part II patients have a 
treatment-related grade 3 or 4 toxicity, unless a toxicity specifically accepted in Section 12.5 and 12.6, 
or if any of the first three patients experiences a treatment-related death, at that point the trial will be 
immediately suspended. We will consult with the data and safety monitoring board (DSMB), and a 
detailed review of the potential causes of toxicity undertaken. We will reconvene with the DSMB and 
consideration will be given to changing the cell dose to the part I dose level and/or other protocol 
modifications on guidance of the DSMB. If instead three part II patients have received their first T cell 
infusion and completed the 28 day DLT evaluation after the lower first dose infusion, with one or fewer 
patients experiencing grade 3-4 related adverse events as defined in Section 12, staggering restrictions 
will be lifted and there will be no specific staggering requirement. 
 
As patients eligible for second infusion will have tolerated a previous exposure to the same T cell 
product, between-patient staggering intervals will not apply for second infusion. However, patients will 
be monitored for a second 28-day DLT period and safety events recorded. Patients previously treated 
with lower doses of T cells on earlier versions of this trial will be eligible to receive up to an additional 2 
doses of TCR transgenic T cells, starting at the dose level for newly entering patients provided that they 
meet eligibility criteria in Section 5 before each additional dose. 
 
 
5. 
STUDY POPULATION AND ELIGIBILITY  
The target study population consists of adult patients with metastatic or unresectable MCPyV-associated 
Merkel cell carcinoma (VP-MCC) that has progressed on or after prior treatment with a PD-1 axis 
immune checkpoint inhibitor. Patients must meet all of the inclusion and exclusion criteria to be enrolled 
in this study.  
 
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Results of tests and/or procedures conducted as per standard of care purposes may be used for research 
purposes if conducted within the protocol-defined window prior to screening/leukapheresis and/or T-
Cell Therapy.  
 
5.1 
Inclusion Criteria – Evaluation  
Individuals that may be consented to undergo evaluation to determine potential eligibility must have a 
history of metastatic or unresectable MCC (as documented by medical record), be 18 years of age or 
older and be capable of understanding and providing informed consent. 
 
5.2 
Inclusion Criteria   
5.2.1 Metastatic or unresectable VP-MCC 
Participants must have metastatic or unresectable, histologically confirmed virus-positive MCC. 
Confirmation of diagnosis must be or have been performed by internal pathology review of initial or 
subsequent biopsy or other pathologic material at Fred Hutch. 
 
Approximately 80% of MCCs are caused by MCPyV T Antigens,16 and the treatment is expected to only 
be effective in this population. Merkel cell polyomavirus positivity may be established through one of 
two means: positive Merkel cell polyomavirus T antigen serology (preferred) or immunohistochemistry 
of a of primary or metastatic MCC tumor lesion (if T antigen seronegative).  
 
MCPyV T Antigen serology will be performed with the AMERK assay run through the University of 
Washington Medical Center Laboratory Medicine Clinical Immunology Laboratory 
(https://testguide.labmed.uw.edu/public/view/AMERK).73,74 Positivity will be defined as a Merkel 
oncoprotein antibody titer (MSCTT) of >= 75 standard titer units (STU) at any time point from initial 
diagnosis onward.   Patients with negative T antigen serology but MCPyV positive tumor by other 
methodologies will be considered for additional MCPyV testing as clinically appropriate, as the T 
antigen serology assays are highly specific 73,75 but incompletely sensitive for MCPyV status.73  In this 
case, immunohistochemistry of any tumor lesion will be performed with the CM2B4 Merkel cell 
polyomavirus T antigen antibody76.  Expression of MCPyV in at least 10% of tumor cells will be 
considered positive. CM2B4 staining performed at any point clinically and at any clinical laboratory 
may be accepted. However, if CM2B4 staining has not been previously performed by a clinical 
pathology laboratory as part of MCC diagnostic workup, it will be performed in a clinical diagnostic 
pathology laboratory at UW/Fred Hutch as per standard staining protocols. Persons are not required to 
have both CM2B4 positivity and seropositivity; either will be acceptable confirmation of viral status and 
if one negative and the other positive the patient will remain eligible provided other criteria are met. 
 
5.2.2 Previous treatment with a PD-1 axis inhibitor or contraindication to treatment with a PD-1 
axis inhibitor 
Patients must have been previously treated with at least one dose of a PD-1 axis inhibitor (e.g., PD-1 or 
PD-L1 inhibiting monoclonal antibody such as pembrolizumab, nivolumab, avelumab, atezolizumab, 
durvalumab), developed progression of their MCC tumor on or after treatment, or have biopsy 
confirmed residual disease following treatment. At least four weeks must have passed between the 
administration of the first dose of PD-1 axis inhibitor and determination of progression or residual 
disease. If there is significant clinical concern for pseudoprogression (i.e., progression developed rapidly 
after checkpoint inhibitor therapy), biopsy must be performed to demonstrate true progression. Patients 
may have received 1 or more prior systemic regimens for MCC. There is no upper limit on prior 
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regimens. Patients may have received prior anti-PD-1/anti-PD-L1 in the neoadjuvant or adjuvant setting.  
Patients are also eligible if they have a contraindication to PD-1/PD-L1 axis blockade such as a history 
of an autoimmune disease. 
 
5.2.3 HLA type HLA-A*02:01 
Participants must be HLA-A*02:01 in order for infused transgenic T cells to recognize antigen-MHC 
complexes.  HLA typing for HLA-A2 should be determined through molecular approaches at a clinical 
laboratory licensed for HLA testing. 
 
5.2.4 Life expectancy must be anticipated to be >3 months at trial entry 
 
5.2.5 18 years of age or older 
Fewer than 0.5% of MCC occur in individuals aged 30 years or younger, thus the protocol includes only 
adult patients. Participants may be of any gender, race, or ethnicity. 
 
5.2.6 Capable of understanding and providing a written informed consent 
 
5.2.7 If fertile, willingness to comply with reproductive requirements. 
These are outlined in detail in Section 5.4.  
 
5.2.8 Karnofsky performance status of >= 60%  
Please see Appendix C for Karnofsky performance status 
 
5.2.9 Willing to undergo serial tumor biopsies 
Should there be no tumor tissue that is accessible for biopsy, patients will still be considered for 
participation, at discretion of the investigator. Similarly, should an investigator determine that a biopsy 
cannot be performed safely for clinical reasons biopsies may be cancelled or retimed. 
 
5.2.10 Participants must be at least three weeks from last systemic MCC treatment 
At least 3 weeks must have passed since any: immunotherapy (for example, T-cell infusions, 
immunomodulatory agents, interleukins, MCC vaccines, intravenous immunoglobulin, expanded 
polyclonal TIL or LAK therapy), NK therapy, small molecule or chemotherapy cancer treatment, other 
investigational agents or other systemic agents that target MCC. There is no washout period for 
radiation. 
 
5.2.11 Acceptable organ function 
Acceptable organ function is defined as:  
 Renal: serum creatinine < 2 or eGFR > 45 
 Hepatic: tBili < 3.0, AST and ALT <5x upper limit of normal (ULN). Patients with suspected 
Gilbert syndrome may be included if Tbili >3 but no other evidence of hepatic dysfunction. 
 Pulmonary:  ≤ grade 1 dyspnea and SaO2 ≥ 92% on ambient air. If PFTs are performed based on 
the clinical judgment of the treating physician, patients with FEV1 ≥ 50% of predicted and 
DLCO (corrected) of ≥ 40% of predicted will be eligible. 
 Cardiac: Patients 50 years of age or older are required to have left ventricular ejection fraction 
(LVEF) evaluation performed within 2 months prior to study treatment. LVEF may be 
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established with echocardiogram or MUGA scan and must be ≥ 35%. Cardiac evaluation for 
other patients is at the discretion of the treating physician. 
 Hematologic: ANC > 1000 cells/mm3, ALC > 200 cells/mm3, HCT > 30%, platelet count > 
50K.  
 
5.3 
Exclusion Criteria  
5.3.1 Active autoimmune disease 
Active autoimmune disease requiring immunosuppressive therapy is excluded unless discussed with the 
PI. Patients with a history of autoimmune disease that was a contra-indication to PD-1 axis 
blockade treatment will be eligible, but will not receive PD-1 axis blockade treatment on trial.  
 
5.3.2 Prior solid organ transplant or allogeneic hematopoietic stem cell transplant.  
Kidney transplant will be considered on a case-by-case basis requiring discussion with PI. If 
kidney transplant, patient must have dialysis access, dialysis plan, supportive nephrologist, willingness 
to stop transplant immunosuppression, and express understanding that rejection is likely. Dialysis 
or costs related to transplant kidney will not be supported by the study. Participants having had any 
other solid organ transplants will be excluded, as will those with a history of allogeneic stem cell 
transplant.   
 
5.3.3 Corticosteroid therapy at a dose equivalent of >10 mg prednisone per day.  
 
5.3.4 Concurrent use of other investigational agents or MCC directed therapies. 
 
5.3.5  Any Medical or psychological condition other than Merkel cell carcinoma that would 
significantly increase the risk of harm to a subject or interfere with the interpretation of 
trial endpoints. 
 
5.3.6 Active uncontrolled infection.  
HIV positive participants on HAART with a CD4 count > 500 cells/mm3 are considered controlled, as 
are individuals with a history of hepatitis C who have successfully completed antiviral therapy with an 
undetectable viral load, and those with hepatitis B who have hepatitis well controlled on medication.  
 
5.3.7 Uncontrolled concurrent illness. 
Participants may not have uncontrolled or concurrent illness including, but not limited to, symptomatic 
congestive heart failure, unstable angina pectoris, cardiac arrhythmia, or psychiatric illness/social 
situations that would limit compliance with study requirements.    
 
5.3.8 Untreated brain metastases.  
Participants with small asymptomatic brain metastases (<1 cm) or those with brain metastases 
previously treated with surgery or radiotherapy will be considered for inclusion at discretion of principal 
investigator, so long as other eligibility criteria are met. 
 
5.3.9 Active treatment for prior immune related adverse event to any immunotherapy.  
Participants receiving treatment for prior Immune Related Adverse Events (iRAE) are excluded, with 
exception of hormone supplementation or corticosteroid therapy at equivalent of up to 10 mg prednisone 
per day, or immune related adverse events that have resolved to grade 1, unless otherwise approved by 
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PI. Patients with Grade 3 or higher Immune related adverse event (iRAE) to any prior PD-1 axis 
blocking agent will be eligible provided these have resolved to grade 1 or better, and are not requiring 
treatment greater than prednisone 10mg daily.  These patients with prior high grade iRAE will not 
receive a PD1/PD-L1 axis inhibitor on the trial. 
 
iRAEs are persistent T cell mediated inflammatory syndromes caused by PD-1 or PD-L1 inhibitors 
including colitis, nephritis, pneumonitis, myositis, hepatitis, encephalitis.  
 
5.3.10 Significant underlying neurologic disease.  
Study participants must not have significant active underlying neurologic disease, unless approved by 
PI. Mild neuropathy related to diabetes or prior chemotherapy is acceptable.  
 
5.3.11 Other medical, social, or psychiatric factor that interferes with medical appropriateness 
and/or ability to comply with study, as determined by PI. 
 
5.4 
Reproductive Potential and Contraception Requirements 
Any female patient who does not meet at least one of the following criteria will be considered to have 
reproductive potential: 
 Post-menopausal for at least 12 consecutive months (i.e., no menses), or 
 Undergone a sterilization procedure (hysterectomy, salpingectomy, or bilateral oophorectomy; 
tubal ligation is not considered a sterilization procedure) 
Pregnancy test for females of reproductive potential must be negative within 14 days before 
leukapheresis. 
Female patients with reproductive potential who are not sexually abstinent and male patients who are 
sexually active with females of reproductive potential must agree to use a suitable method of 
contraception for the duration of the study (from date of consent through treatment period). 
6. 
TREATMENT PLAN 
Please see Section 4.4 for information on patient staggering and escalation from part I to part II. 
6.1 
Overall schema  
 
In part I, the first T cell infusion will be up to 1 x 109 tetramer positive CD8+ T cells. Although targeted 
at 1:1, the actual number of transgenic CD4+ T cells will vary between individuals due to interpersonal 
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variability (cells are co-cultured). Please see Section 6.4 for maximum infused transgenic T cells, and 
Section 2.1.12 for dose rationale.  If patients have progressive disease and tetramer positive T cell 
persistence below 5% of peripheral CD8+ T cells, a second infusion may be administered at a higher 
dose of  up to 5 x 109 tetramer positive CD8+ T cells.  In part II, the initial and potential second T cell 
dose will be up to 5 x 109 tetramer positive CD8+ T cells.   
 
Transgenic T cells and interferon gamma are investigational, and avelumab or pembrolizumab are 
provided as standard of care. Therefore, duration of avelumab or pembrolizumab infusions is provided 
as a suggestion only and may be shortened or extended on the medical judgement of the treating 
oncologist. 
 
6.2 
Leukapheresis 
Patients must be eligible and consented for treatment portion of study prior to leukapheresis because cell 
product preparation will begin immediately upon leukapheresis in most cases. Following enrollment on 
the study, a leukapheresis collection will be performed on each patient to obtain peripheral blood 
mononuclear cells (PBMCs) for the production of the FH-MCVA2TCR investigational product. 
Leukapheresis will be performed by the Fred Hutch Apheresis Unit using standard operating procedures 
(SOPs) for obtaining PBMCs. If a technical issue arises during the procedure or in the processing of the 
product, or if insufficient FH-MCVA2TCR cells are manufactured for the prescribed dose, the patient 
may undergo a second collection procedure.   
Patients ineligible for a vein-to-vein apheresis may elect to have a percutaneous central venous access 
catheter inserted to support this collection.   
The leukapheresis product will be delivered to the Cell Processing Facility (CPF) at Fred Hutch or the 
Cell Therapy Laboratory (CTL) at Fred Hutch for cell selections then transported to the Fred Hutch CPF 
for study product manufacturing according to methods outlined in the Chemistry, Manufacturing and 
Controls (CMC) section of the Investigational New Drug (IND) application to the FDA. PBMCs from 
the leukapheresis and cell selection products may be archived for research. 
6.3 
Interferon Gamma treatment 
Interferon gamma is being administered with the goal of upregulation of class I MHC on Merkel cell 
carcinoma cells to allow recognition of tumor cells by TCR transgenic T cells. Interferon gamma 
(Actimmune) will be administered at the FDA-approved dosing (for the indication of chronic 
granulomatous disease) of 50mcg/m2 given subcutaneously in the abdomen, anterior thigh, or deltoid 
region three times weekly, starting 5-7 days prior to the T cell infusion and continuing for a total of 4 
weeks.  Patients will receive premedication with acetaminophen prior to each dose to minimize flu-like 
symptoms. Doses will be held if creatinine rises above 2.0; creatinine will be monitored by regular 
comprehensive metabolic panels per Appendix A. 
Dose reduction or skipping, as clinically required in the event of adverse events is permissible at the 
discretion of the PI or designee.  
6.4 
FH-MCVA2TCR Infusions 
FH-MCVA2TCR will be administered by IV infusion at Fred Hutch (or at University of Washington 
Medical Center (UWMC) if the patient is hospitalized). T-cell infusions will be given preferably at least 
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48 hours before the next dose of avelumab or pembrolizumab. However, the infusion may be delayed if 
required for clinical reasons. Patients will receive either 1 or 2 infusions of FH-MCVA2TCR. Infusions 
will be ideally scheduled for 12 weeks apart; however, they may be given earlier in the case of early 
disease progression or later in the event of a need for bridging chemotherapy treatment. 
Patients in the part I portion will receive a lower dose for the first infusion of up to 109 (one billion) 
TCR A2-MCC1 transgenic CD8+s, and then up to 5 x 109 (five billion) TCR A2-MCC1 transgenic 
CD8+ T cells at the second infusion. Patients in the part II portion will receive a full dose of up to 5 x 
109 (five billion) TCR A2-MCC1 transgenic CD8+ T cells at both infusions. Infusions will also include 
CD4+ T cells that will vary naturally between patients as the cells are co-cultured; the total allowable 
transgenic cell dose (including transgenic CD8+s and CD4+s) will be 10 times higher than the targeted 
CD8+ dose (e.g., 1010 for low dose infusion and 5 x 1010 for standard dose infusion respectively). 
Patients previously treated on the protocol at lower cell doses than currently available will be eligible to 
re-enter the protocol to receive up to 2 additional infusions. 
T cells will be infused intravenously through a blood component filter set on an electronic infusion 
device over approximately 60 minutes. The infusion bag will be gently mixed periodically during the 
infusion. All patients will be observed for 2 hours post infusion and will be released if no AE requiring 
prolonged hospitalization have occurred. 
 
6.4.1 Preparation of Cell Product 
All manufacturing activities are conducted using approved written procedures performed by trained, 
qualified personnel in accordance with current Good Manufacturing Practices (cGMP) and current Good 
Tissue Practices (cGTP) guidelines. Quality assurance oversight and review is provided to assure 
regulatory compliance and safety of therapeutic products. It is anticipated to take approximately 9-14 
days to manufacture a product in most cases. 
 
If a product cannot be formulated to meet a target cell dose, the cell product should be infused at or as 
close as possible to the total transgenic CD8+ T-cell dose. A product will be considered evaluable for 
response if it contains ≥10% of the scheduled CD8+ transgenic T cell dose; however, lower dosages of 
cells may be administered at the discretion of the investigator. 
 
6.4.2 T Cell Infusions 
1. TCR-T cells should be administered approximately 5-7 days after the initiation of interferon 
gamma A window of 4-8 days is acceptable if a 5-7 day window is not possible for logistical 
reasons.   
2. On the day of scheduled T cell infusion, the patients should undergo a clinical evaluation and a 
clinical determination for appropriateness to proceed with T cell administration. 
3. Patients deemed not appropriate for T cell administration may receive a T cell infusion outside the 
specified window if they subsequently resolve the clinical and/or laboratory concerns which 
deemed them inappropriate within 60 days of the planned initial infusion. In these cases, the 
product will be frozen and thawed on the new day of administration, as per facility policy and 
procedures.  
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4.  Patients who are unable to undergo TCR-T cell infusion within the intended window due to 
manufacturing or release issues may receive the cell infusion outside of this window if the issues 
can be resolved. Discussion with the protocol PI and notification of IND holder is required. 
FH-MCVA2TCR will be provided as a single cell product at the assigned cell dose.   
The final product will be prepared and labeled according to standard operating procedure in the Cellular 
Processing Facility. The cell product will be transported to the infusion facility by a delegated staff 
member. During the time of transportation, the cell product will be kept in a cooler with a cool pack. A 
nurse will then administer the cells using standard/universal institutional precautions for blood borne 
pathogens. 
 
All patients will be monitored during each T-cell infusion with vital signs and O2 saturation being 
recorded (+/- 5 minutes) within 15 minutes prior to the start of infusion, at thirty-minute intervals 
during, at completion of the infusion, and hourly after the infusion for at least 2 hours after the infusion. 
 
6.4.3 Acute infusion reactions 
Acute infusion reactions may occur with administration of FH-MCVA2TCR. Guidelines for the 
monitoring and treatment of acute infusion reactions are provided in Sections 7.3 and 7.4. 
6.4.4 Guidelines for the determination of 1 or 2 cycles of T cell therapy 
Each “cycle” of T cell therapy consists of one cycle of interferon gamma plus T cell infusion.  Interferon 
gamma will be given with second infusions unless patient has had a grade 3 or higher toxicity attributed 
to interferon gamma. 
 
Twelve weeks after first T cell infusion, restaging and evaluation of peripheral blood persistence of 
infused transgenic T cells will be performed, and patients will be considered for a second cycle (second 
class I MHC upregulation procedure plus second infusion of transgenic T cells). A second cycle may be 
given between 6 and 12 weeks after the first cycle in patients who have progressive disease and <5% 
TCR T cells among CD8 T cells in the blood. Avelumab or pembrolizumab infusions will continue up to 
the second infusion. Decision-making surrounding a second cycle will be at the discretion of the 
investigator and will be generally made following the below guidelines:  
 
Individuals with progressive disease and non-persisting T cells (<5% of CD8) may be considered for a 
second infusion if they meet eligibility criteria for treatment as described in Section 5 above.  Patients 
with symptomatic progression may receive debulking radiotherapy or chemotherapy for one or more 
cycles per standard of care between the first and second T cell infusions, but the interval between the 
end of chemotherapy and the second infusion must be at least 3 weeks.  Treatment with avelumab or 
pembrolizumab will be discontinued during chemotherapy and will be resumed 1 week before the 
second infusion of T cells.  
 
Individuals with progressive disease and persistent T cells (>5% of CD8) will no longer receive 
additional T cells or avelumab or pembrolizumab on study. They will leave the treatment phase of the 
study and will transition to long-term follow-up. 
 
Individuals with complete response will remain on the treatment phase of the study and will continue to 
receive avelumab or pembrolizumab to complete a year of treatment. However, they will not receive 
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additional T cells, given that they have no lesions with which to target with class I MHC upregulation 
and also that the risk-benefit profile favors fewer interventions in the setting of complete response. For 
these patients, T cell product will be cryopreserved, and they may be considered for a second cycle of T 
cell therapy in the future, at the discretion of the investigator, should their MCC recur. 
 
Individuals with partial response or stable disease (per RECIST 1.1) will remain on study and will 
continue to receive avelumab or pembrolizumab to complete a year of treatment. They will have 
evaluation of the persistence of their transgenic T cell product as well as the suitability of residual 
disease for class I MHC upregulation. Should they have poor to moderate persistence (for instance <5% 
TCR A2-MCC1 CD8+s) in the peripheral blood of the transgenic T cells by tetramer stain, a second T 
cell infusion may be given. If there is a lesion suitable for class I MHC upregulation, they will also 
receive a second class I MHC upregulation. 
 
6.5 
Avelumab or Pembrolizumab – standard of care 
Avelumab is a fully human monoclonal antibody of the immunoglobulin (Ig) G1 isotype that selectively 
binds to PD-L1 and competitively blocks its interaction with PD-1. Compared with anti-PD-1 antibodies 
that target T cells, Avelumab targets tumor cells and therefore is expected to have fewer side effects, 
including a lower risk of autoimmune-related safety issues, as blockade of PD-L1 leaves the PD-L2 / 
PD-1 pathway intact to promote peripheral self-tolerance.77 In the event of an infusion reaction to 
avelumab prior to or during the study,  the standard of care for the patient may change from avelumab to 
pembrolizumab. Pembrolizumab is a fully human monoclonal antibody that binds to and inhibits PD-1.  
Pembrolizumab may have a slightly higher incidence of immune related adverse events in patients with 
MCC78. If the patient has a history of grade 3 or higher immune related adverse event with prior PD-1 
axis blockade, or if the patient has a history of a significant autoimmune disease that precluded PD-1 
axis blockade treatment, the patient will not be treated with avelumab or pembrolizumab on this study.  
Patients who experience grade 3 or greater toxicity with ipilimumab containing regimens will be eligible 
to receive avelumab or pembrolizumab on study if they subsequently tolerated PD-1 axis inhibition 
without immune adverse event prior to enrollment. 
 
Avelumab (Bavencio) and pembrolizumab (Keytruda) are both FDA-approved for the treatment of 
MCC. 
 
6.5.1 Provision of avelumab or pembrolizumab 
Avelumab will be given as standard of care and will be billed to the study participant’s insurance.   
 
Since the initial writing of this protocol, the PD-1 inhibitor pembrolizumab has been FDA approved for 
the treatment of metastatic Merkel cell carcinoma. For patients with a history of infusion reactions to 
avelumab, pembrolizumab can be administered with this study in place of avelumab at 200 mg every 3 
weeks or at any subsequent FDA approved dosing regimen for this indication, starting 14 days following 
investigational T cell infusion. 
 
6.5.2 Avelumab or pembrolizumab infusion 
Avelumab will be administered as standard of care. Currently, the FDA recommended dosing is 
10 mg/kg over the duration of 1 hour (-10/+70 minutes) at an interval of once every 2 weeks.  
Premedications suggested by package label include pretreatment with H1 blockers (diphenhydramine 
12.5mg to 50 mg PO or IV, or equivalent) and acetaminophen PO 500 to 650 mg for first several doses.  
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First avelumab dose is intended 14 +/- 3 days after first T cell infusion up to 50 weeks (total of 25 
infusions) or longer, as clinically indicated under the standard of care or until Off Study Criteria are met 
as described in Section 10. The first dose of avelumab will be calculated based on the actual weight of 
the participant determined at study entry. As avelumab is being administered as standard of care, if 
alternate doses or schedules are approved by the FDA for avelumab for treatment of advanced MCC (for 
instance q4 week dosing or flat instead of weight-based dosing), these doses and schedules may be 
substituted at the discretion of the investigator and treating oncologist. Following first avelumab 
infusion, participants will be observed for 1 hour post infusion for potential infusion-related reactions. 
Participants will receive avelumab once every 2 weeks for 50 weeks or until the criteria in Section 11.7 
(Off Study Criteria) are met. Modifications of the infusion rate due to infusion-related reactions and 
management of acute infusion reactions are below as recommended in product label.   
 
The first avelumab infusion after each T cell infusion should be administered at Fred Hutch for 
monitoring in case of acute adverse event. Subsequent infusions may be given locally by a licensed 
oncology professional experienced in prescribing immunotherapy. A “Dear Physician” letter detailing 
the study treatment and providing contact information to the study team including emergency contact 
information will be provided to local/non-Fred Hutch oncologists.   
 
Monitoring exams and blood work associated with avelumab infusion will be performed clinically at the 
discretion of the treating oncologist and in keeping with institutional standards for avelumab infusion.   
 
If pembrolizumab is given instead of avelumab, it will be administered over 30 minutes intravenously 
with similar monitoring as above and without premedications unless at the direction of the clinical team. 
 
Typical practice at Fred Hutch: A physical exam is usually performed by an APP or MD within 48 
hours of infusion and blood work is obtained. Typically, this blood work typically includes a CBC with 
differential, basic metabolic panel, liver function panel, and LDH with each infusion, and a serum 
cortisol, TSH, free T4, amylase, and lipase with every other infusion.  Cortisol, TSH, free T4, amylase 
and lipase do not often result prior to that day’s infusion, however these are followed longitudinally to 
determine whether there is a need for hormone replacement. However, different institutions have 
different standards and policies for avelumab or pembrolizumab monitoring, and labs may need to be 
more or less often for safety reasons or include additional labs depending on patient’s course, therefore 
the protocol will allow flexibility in the avelumab or pembrolizumab.  
 
Avelumab or pembrolizumab doses may be held, rescheduled or dosages reduced at the discretion of 
treating oncologist in keeping with standard clinical practice.  If immune related adverse event 
secondary to avelumab or pembrolizumab suspected or confirmed, treating oncologist to discuss 
management with study team and joint management decision established. 
 
6.5.3 Recommended Supportive Care, Additional Treatment, and Monitoring 
Prophylactic treatment/measures are recommended for patients at risk for tumor lysis syndrome (TLS) 
according to institutional or clinical standards. Supportive care for the management of TLS is detailed in 
Section 7. 
 
The use of red blood cells and platelet transfusions, and/or colony-stimulating factors is permitted 
according to institutional or clinical standards.  
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The use of prophylactic or empiric anti-infective agents (e.g., trimethoprim/sulfamethoxazole for 
pneumocystis pneumonia [PCP] prophylaxis, broad spectrum antibiotics, antifungals, or antiviral agents 
for febrile neutropenia) is permitted according to institutional standards. 
Hospitalization may be required after study treatment to manage treatment-associated toxicities.   
 
7. 
POTENTIAL RISKS AND TOXICITY MONITORING 
7.1 
Toxicity associated with interferon gamma administration 
Administration of interferon gamma can commonly cause fever, headache, chills, fatigue, rash, nausea 
and vomiting, diarrhea, myalgia, abdominal pain, depression, arthralgias, and local injection site 
reactions.  More rare but serious adverse events include colitis, hypersensitivity reactions, elevations in 
liver function tests, myocardial infarction, cytopenias, hypokalemia, stevens johnson syndrome and 
neurologic effects. 
 
7.2 
Management of toxicity associated with interferon gamma  
Patients will be premedicated with acetaminophen 650 mg prior to each dose. 
Mild acute administration reactions should be managed with antipyretics, antihistamines, and anti-
emetics as clinically appropriate, and these may be added to prophylaxis for future doses. 
Corticosteroids should be avoided. Appropriate emergency medications (e.g., epinephrine, 
diphenhydramine, etc.) should be readily available during administration and institutional guidelines 
should be followed for the treatment of anaphylaxis. 
Fever and flu like symptoms should be treated with antipyretics as needed. 
Local reactions can be managed by choosing alternate sites. 
Severe symptoms or those involving hypoxemia or hypotension, particularly following T cell infusion, 
should be treated as presumptive cytokine release syndrome and treated as below. 
Doses may be held or treatment discontinued due to intolerable side effects or grade III or higher 
adverse events attributable to interferon gamma. Adverse events attributable to T cell infusion (e.g. 
cytokine release syndrome) will be assumed to be synergistic with interferon gamma, and interferon 
gamma will be held in these cases.  
 
7.3 
Acute Toxicity Associated with T-Cell Infusion (Infusion reaction) 
Administration of MCPyV-specific TCRs may cause infusion reactions, including fever, rigors, rash, 
urticaria, dyspnea, hypotension, and/or nausea. Mild infusion reactions should be managed with 
antipyretics, antihistamines, and anti-emetics as clinically appropriate. Corticosteroids should be 
avoided. Appropriate emergency medications (e.g., epinephrine, diphenhydramine, etc.) should be 
readily available during administration and institutional guidelines should be followed for the treatment 
of anaphylaxis. 
7.4 
Management of Acute Infusion Reaction(s) 
For infusion reactions or any clinically significant changes from baseline, stop administration of 
MCPyV-specific TCRs, notify the provider to have the patient assessed and administer appropriate 
symptomatic treatment. 
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 Fever, chills, and temperature elevations > 38.0°C may be managed with acetaminophen 650 mg 
PO every 4 to 6 hrs. All patients who develop fever > 38.0°C or chills should have a blood 
culture drawn and the PI or designee should be notified. 
 Nausea and/or vomiting may be managed with ondansetron 8 mg po or other antiemetics 
(excluding corticosteroids). 
 Hypotension should be managed initially by fluid administration. 
 Hypoxemia should be managed initially with supplemental oxygen. 
After the resolution of symptoms, an order may be obtained to resume administration at a reduced rate. 
If symptoms re-occur, stop the administration, administer symptomatic treatment as ordered and contact 
the IMTX attending for the final determination to discontinue the infusion. 
If an adverse reaction occurred during the IEC infusion or post-monitoring period, the provider will 
determine in collaboration with the research team if any workup is needed.  
Patients requiring discontinuation of the infusion may be eligible for re-treatment if the cause is deemed 
not related to the infusion. 
7.5 
Acute Toxicity Associated with Avelumab or Pembrolizumab Infusion 
As with all monoclonal antibody therapies, avelumab or pembrolizumab carries a risk of allergic 
reaction, although infusion reactions are more common with avelumab10 (20%) than pembrolizumab78 
(2%) in MCC. If hypersensitivity reaction occurs, the participant must be treated according to the best 
available medical practice. Hypersensitivity reactions associated temporally with infusion of 
pembrolizumab or avelumab will be attributed to pembrolizumab or avelumab. Reactions not temporally 
linked with pembrolizumab or avelumab will be attributed to interferon gamma.  
Avelumab: special precautions  
As infusion reactions have been commonly observed with avelumab, pre-treatment with H1 blockers 
(diphenhydramine 12.5-50 mg PO, IV, or equivalent), and acetaminophen (500 to 650 mg orally) will be 
administered 30 to 60 minutes prior to the first avelumab infusion to mitigate infusion-related reactions 
including hypersensitivity reactions. Pretreatment for subsequent infusions or pembrolizumab infusions 
will be per treating physician/institutional standard of care. 
 
Infusion of avelumab or pembrolizumab will be stopped in case of Grade ≥ 3 hypersensitivity, 
inflammatory response, or infusion-related reaction. The treatment recommendations for infusion-
related reactions, severe hypersensitivity reactions, and tumor lysis syndrome are as outlined below. 
These may be substituted with an institution’s specific guideline/standard practice for avelumab infusion 
related reactions. 
 
Avelumab or pembrolizumab: Infusion-related reactions 
 
Symptoms 
Fever 
Chills 
Rigors 
Diaphoresis 
Headache 
 
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Management  
Table 6. Treatment modification for symptoms of infusion-related reactions caused by Avelumab 
or Pembrolizumab 
 
NCI-CTCAE Grade
Treatment Modification for Avelumab
Grade 1 – mild 
Mild transient reaction; infusion 
interruption not indicated; intervention not 
indicated. 
Decrease the Avelumab or 
Pembrolizumab infusion rate by 50% and 
monitor closely for any worsening. 
The total infusion time for Avelumab or 
Pembrolizumab should not exceed 
120 minutes.
Grade 2 – moderate 
Therapy or infusion interruption indicated 
but responds promptly to symptomatic 
treatment (for example, antihistamines, 
NSAIDs, narcotics, IV fluids); prophylactic 
medications indicated for  24 hours.
Stop Avelumab or Pembrolizumab 
infusion. 
Resume infusion at 50% of previous rate 
once infusion-related reaction has resolved 
or decreased in severity to < Grade 1 and 
monitor closely for any worsening. 
Grade 3 or Grade 4 – severe or 
life-threatening 
Grade 3: Prolonged (for example, not 
rapidly responsive to symptomatic 
medication and/or brief interruption of 
infusion); recurrence of symptoms 
following initial improvement; 
hospitalization indicated for clinical 
sequelae. 
Grade 4: Life-threatening consequences; 
urgent intervention indicated. 
Stop the avelumab or pembrolizumab 
infusion immediately and disconnect 
infusion tubing from the participant. 
Participants who have a reaction associated 
with avelumab may not receive any further 
administration of avelumab as part of this 
study, but they may receive administration 
of pembrolizumab as part of this study. 
Participants who have a reaction associated 
with pembrolizumab may not receive any 
further administration of pembrolizumab as 
part of this study.  In addition, they may not 
receive any administration of avelumab as 
part of this study.” 
 
Definitions: IV=intravenous, NCI-CTCAE=National Cancer Institute-Common 
Terminology Criteria for Adverse Event, NSAIDs=nonsteroidal anti-inflammatory 
drugs, 
 
Once the avelumab or pembrolizumab infusion rate has been decreased by 50% or interrupted due to an 
infusion-related reaction, it must remain decreased for all subsequent infusions. If the participant has a 
second infusion-related reaction of Grade ≥ 2 on the slower infusion rate, the infusion should be 
stopped, and the participant should be removed from avelumab or pembrolizumab treatment. If a 
participant experiences a Grade 3 or 4 infusion-related reaction at any time, the participant must 
discontinue avelumab or pembrolizumab.  Patients who experience infusions reactions with avelumab 
may receive pembrolizumab for future infusions.  Patients who experience infusion reactions with 
pembrolizumab may not receive avelumab for future infusions.  
 
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Management of severe hypersensitivity reactions and flu-like symptoms due to avelumab or 
pembrolizumab infusions 
 
If hypersensitivity reaction occurs, the participant must be treated according to the best available 
medical practice. Participants should be instructed to report any delayed reactions to the Investigator 
immediately. 
 
Symptoms of severe hypersensitivity reaction 
Impaired airway 
Decreased oxygen saturation (< 92%) 
Confusion 
Lethargy 
Hypotension 
Pale / clammy skin 
Cyanosis 
 
Management 
Epinephrine injection and dexamethasone infusion 
Patient should be placed on monitor immediately  
Alert ICU for possible transfer if required 
 
7.6 
Cytokine Release Syndrome (CRS) 
Administration of transgenic T cells such as FH-MCVA2TCR may be associated with CRS. CRS may 
be characterized by high fever, fatigue, nausea, headache, dyspnea, tachycardia, rigors, hypotension, 
hypoxia, myalgia/arthralgia, anorexia, coagulation abnormalities, organ dysfunction, and neurologic 
abnormalities.   
If a patient becomes febrile or develops symptoms of CRS, cytokine levels, serum ferritin, C-reactive 
protein (CRP), coagulation studies, and/or markers of tumor lysis syndrome (e.g., chemistry, uric acid, 
lactate dehydrogenase [LDH]) may be measured, and persistence and/or phenotype of the transgene-
expressing cells may be evaluated, as clinically indicated. 
Any patient who develops clinical evidence of symptoms related to CRS will have a work-up to exclude 
infection or other causes, as clinically appropriate. Initial treatment should consist of supportive 
measures as dictated by the clinical and laboratory findings, and may include fluid replacement, 
medications to support blood pressure, antipyretics, oxygen supplementation, anti-seizure medications, 
and broad-spectrum antibiotics if infection cannot be excluded as a potential etiology for the signs and 
symptoms.   
The 2019 Lee et al consensus guidelines for immune effector cell will be utilized to quantify T cell 
associated CRS,79,80 as detailed in Appendix E.    
General guidelines for management of CRS are provided in Appendix F based on our current institution 
standard practice. Other cytokine-directed therapies may be considered after discussion with the PI 
and/or immunotherapy attending. 
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7.7 
Neurologic Toxicity 
Neurotoxicity, manifested as delirium, seizures, focal neurologic deficits, and/or coma has been reported 
after CAR T-cell therapy. Although neurotoxicity has not been reported after transgenic TCR therapy, if 
neurotoxicity develops management will be given analogous to CAR therapy. Neurotoxicity to CAR-T 
is usually reversible but can be irreversible or fatal. Neurotoxicity/ICANS will be graded according to 
2019 Lee et al consensus guidelines, 79 as detailed in Appendix E.    
For patients who develop mild neurologic manifestations (Grade 1), symptomatic care and levetiracetam 
are recommended.  Discussion with the PI or designee is recommended. For patients with worsening 
neurologic changes, the addition of corticosteroids should be considered (e.g., dexamethasone 10 mg IV 
every 6 to 24 hours). Tocilizumab (4-8 mg/kg IV) or other cytokine-directed therapies may be used 
based on clinical judgment; at this time, it is unclear if these approaches are of benefit to patients with 
neurotoxicity. Cerebrospinal fluid (CSF) assessments and CNS imaging should be considered if 
clinically indicated. 
7.8 
Tumor Lysis Syndrome 
Although rarely reported for MCC, FH-MCVA2TCR may cause tumor lysis syndrome (TLS) in patients 
with high disease burden. Patients with large bulky tumors will be considered for allopurinol 
prophylaxis to start prior to interferon gamma and continued for as long as the medical team determines 
appropriate after the infusion, unless there are contraindications. Patients may receive additional 
hydration for the first 2 weeks after infusion. If TLS develops, as defined by the Cairo Bishop criteria,81 
the Attending Physician will direct patient management with guidance from the study staff. 82  
Conservative therapy, including allopurinol and IV fluid hydration may be instituted immediately for 
suspected TLS.  Hyperkalemia may be treated with potassium-binding resins, diuresis, or 
insulin/dextrose therapy. Hyperphosphatemia may be treated with phosphate-binding resins. In severe 
cases, rasburicase (in non-G6PD-deficient individuals) or renal dialysis may be necessary. 
7.9 
Macrophage Activation Syndrome 
Macrophage activation syndrome (MAS) is a serious disorder potentially associated with uncontrolled 
activation and proliferation of T cells and subsequent activation of macrophages. MAS is typically 
characterized by high-grade, non-remitting fever, cytopenias, and hepatosplenomegaly, and may be 
difficult to distinguish from CRS. Laboratory abnormalities found in MAS include elevated 
inflammatory cytokine levels, serum ferritin, soluble IL-2 receptor (sCD25), triglycerides, and decreased 
circulating NK cells. Other findings include variable levels of transaminases, signs of acute liver failure, 
coagulopathy, and disseminated intravascular coagulopathy. While there are no definitive diagnostic 
criteria for MAS, it is typically diagnosed using published criteria for hemophagocytic 
lymphohistiocytosis.83  
Patients treated with FH-MCVA2TCR should be monitored for MAS, and cytokine-directed therapy or 
corticosteroids should be considered as clinically indicated. 
7.10 
Adrenal Toxicity 
Immune-mediated damage to the adrenal gland is a potential toxicity and could be manifested as 
primary mineralocorticoid or glucocorticoid deficiency.  
If mineralocorticoid and/or glucocorticoid insufficiency should occur, diagnosis may be difficult in the 
acute setting of CRS, and attention should be given to ensuring there is sufficient replacement therapy.  
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For patients with hypotension in the absence of fever that is unresponsive to fluid boluses, 
fludrocortisone or other mineralocorticoid administration should be considered.  If CRS requiring 
intervention with tocilizumab and dexamethasone should occur, the activity of dexamethasone 10 
mg/bid will likely be sufficient; however, if hypotension recurs following cessation of dexamethasone or 
persists despite dexamethasone, stress-dose hydrocortisone is recommended. Samples will be assayed 
for renin, aldosterone and cortisol levels. To detect subclinical hypoadrenalism, an adrenocorticotropic 
hormone (ACTH) stimulation test will be performed at baseline and approximately 4 weeks after 
infusion.   
7.11 
Immune mediated adverse events 
Avelumab and pembrolizumab are associated with a known risk of irAEs. Patients will be closely 
monitored for potential irAEs,84,85 which may manifest earliest after weeks of treatment. Such events 
may consist of persistent rash, diarrhea and colitis, autoimmune hepatitis, arthritis, glomerulonephritis, 
cardiomyopathy, or uveitis and other inflammatory eye conditions. The spectrum of hypothetical irAEs 
also includes formation of auto-antibodies like anti-nuclear antibodies (ANAs) or antineutrophil 
cytoplasmic antibodies (ANCAs). Severe irAEs will be managed with steroids. Where possible, irAEs 
definitely, probably, or possibly secondary to avelumab or pembrolizumab will be managed by the 
Attending Physician in consultation with the study team in accordance with ASCO society guidelines.86 
Both interferon gamma and avelumab/pembrolizumab can cause GI toxicity. More commonly GI 
toxicity due to pembrolizumab/avelumab will manifest as diarrhea that is persistent. Persistent diarrhea 
will be attributed to pembrolizumab/avelumab and managed by the attending physician in consultation 
with the study team in accordance with ASCO society guidelines. Episodic diarrhea or vomiting 
associated temporally with interferon gamma administration will be attributed to interferon gamma and 
managed with dose interruption or cessation in cases of grade 3 or higher toxicity. 
 
7.12 
Persistent Uncontrolled T-cell Proliferation 
Uncontrolled proliferation of TCR modified T cells has not been observed in clinical trials to date.29 
However, in the unlikely event that clinically significant uncontrolled and persistent proliferation of T 
cells occurs in a study patient, initial therapy may involve treatment with corticosteroids (e.g., 
methylprednisolone 1 g IV). If there is a progressive increase in TCR T cells to greater than 30% of T 
cells at more than 3 months after the last infusion is observed, an analysis for clonal expansion by deep 
sequencing of the T-cell receptor (TCR) beta gene (Adaptive Biotechnology) may be conducted.   
7.13 
Replication-Competent Lentivirus 
All patients will be followed in this study for presence of replication-competent lentivirus (RCL) for up 
to 15 years following the last dose of FH-MCVA2TCR.  Research studies to detect presence and 
phenotype T cells may be conducted on samples of tissue biopsies or blood performed for clinical 
conditions occurring during follow-up considered possibly related to FH-MCVA2TCR. 
7.14 
Management of Other Toxicities 
If a new onset CTCAE v5 Grade ≥ 3 toxicity is observed following infusion, the patient will receive 
investigation and medical treatment appropriate for the physiological abnormalities. Grade ≥ 3 toxicity 
that is attributed to the infusion may be treated with corticosteroids (e.g., dexamethasone 10 mg IV 
every 4 to 12 hours), tocilizumab, or other cytokine-directed therapy after discussion with the PI or 
designee. 
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8. 
INFORMED CONSENT OF PATIENT 
8.1 
Patient screening  
An evaluation consent, separate from the treatment consent, will be used for the patient to provide 
consent for records review, pathology review, MCPyV tumor testing (if previously unknown), and HLA 
typing. Where possible, consent for evaluation will be performed in person. However, for many patients 
this will not be possible for travel/logistical reasons. In this case, evaluation consent discussion may be 
performed by telephone by trained study personnel, and consent documents exchanged by email, mail or 
fax. 
8.2 
Treatment  
Patients will be seen at Fred Hutch/UWMC for consideration of treatment options for their disease. The 
protocol will be discussed thoroughly with the patient and other family members if appropriate and all 
known and potential risks to the patient will be described. The procedure and alternative forms of 
therapy will be presented as objectively as possible and the risks and hazards of the procedure will be 
explained to the patient. Consent from the patient will be obtained using forms approved by the Fred 
Hutch IRB. A summary of the clinic visits detailing what was covered will be dictated for the medical 
record. 
 
9. 
PATIENT REGISTRATION 
Eligible patients will be identified and registered into the system by the Clinical Coordinator’s Office 
(CCO; Intake Office) and assigned a Unique Patient Number (UPN).  The CCO will register the patient 
for the protocol through the Data Management Office. 
Patients will initially be screened.  Enrollment to the therapy portion of the study will occur at the 
conclusion of the pre-T-cell work-up when data are reviewed for all inclusion and exclusion criteria for 
therapy by the Immunotherapy attending physician and the patient signs consent for therapy.  
A study number will be allocated to each patient and a log of enrolled patients will be maintained.   
 
10. 
TRAVEL REIMBURSEMENT 
Patients who consent to the treatment arm of the study will be eligible for limited travel reimbursement 
of actual expenses incurred, up to $3,500. Please see consent form for additional details. 
 
11. 
CLINICAL AND LABORATORY EVALUATIONS 
A tabular schedule of events is provided in Appendix A. The proposed days of all treatments and 
assessments are approximate and may vary due to scheduling, clinical or other factors. 
11.1 
Visits required to be at Fred Hutch/UWMC 
Many of the patients with MCC travel >500 miles to receive care at Fred Hutch/UWMC. Although up to 
two T cell infusions are given, the avelumab is given every 2 weeks for a year (or pembrolizumab every 
3 weeks for a year), as a standard of care treatment, using standard of care FDA approved dosing and 
frequency. Therefore, for interventions surrounding T cell infusions and the first avelumab or 
pembrolizumab infusion after each T cell infusion, we request care to be at Fred Hutch/UWMC, in order 
to ensure safety. However, for “maintenance” avelumab or pembrolizumab visits, we will allow patients 
to receive these infusions with their local/primary oncologist if that is patient preference, except as 
detailed below.  
 
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Leukapheresis, pre-treatment biopsy, and pre-treatment physical exam, and treatment consenting need 
be performed through Fred Hutch/UWMC.  
 
For cycle 1 of T cell therapy, the patient is required to receive care at Fred Hutch/UWMC and stay 
within 30 minutes of Fred Hutch/UWMC from the first dose of interferon gamma (day -5) through T 
cell infusion and the 28-day dose-limiting toxicity (DLT) period thereafter. This period includes the first 
(d+14) avelumab or pembrolizumab infusion, if eligible/stable.  If the patient shows no signs of T cell 
mediated toxicity within 17 days of T cell infusion (and 3 days for PD-1/PD-L1 inhibitor infusion), they 
may return to the care of their treating oncologist with permission of the PI. 
 
Day 28, 42, 56, 70, and 84 avelumab (or day 35, 56 and 77 pembrolizumab) may be given locally or at 
Fred Hutch and research blood may be shipped/mailed. Day +84/cycle 1 evaluations including research 
and clinical blood work, exams, imaging, and biopsy should be performed if possible at Fred Hutch or 
UWMC. Patient must remain within 30 minutes of Fred Hutch/UWMC for second T cell infusion and at 
least 28 days thereafter, during which avelumab or pembrolizumab is likely to be administered. 
Subsequent avelumab or pembrolizumab may be given locally. If the patient shows no signs of T cell 
mediated toxicity within 17 days of T cell infusion (and 3 days for PD-1/PD-L1 inhibitor infusion), they 
may return to the care of their treating oncologist with permission of the PI. 
 
6 month, 9 month and 12 month imaging and laboratory evaluations are requested to be performed at 
Fred Hutch; however, they may be performed locally if circumstances preclude travel to Fred Hutch.  
 
11.2 
Screening Evaluations 
1. Determination of viral status if not previously determined (MCPyV AMERK serology assay 
and/or testing of archival tumor tissue). 
2. HLA Typing (HLA typing will be organized by and billed to study) 
11.3 
Pre-Treatment Evaluations 
Results of tests and procedures conducted as per standard of care purposes prior to screening may be 
used for research purposes if conducted within the protocol-defined window.  
 
The screening consent and HIPAA authorization must be signed before any non-standard of care 
evaluations are performed. 
 
Baseline evaluations include: 
1. Medical history and physical examination including prior therapies and response to therapy if 
known, and Karnofsky performance status. 
2. Laboratory tests, including: 
a. CBC, differential, platelet count 
b. Comprehensive metabolic panel 
c. LDH 
d. Fred Hutch (PSBC) Donor Battery Panel (should be performed within 30 days of 
leukapheresis) 
e. Serum ferritin 
f. Serum CRP 
g. Uric acid 
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h. Serum IL-6 
i. Prothrombin time (PT), partial thromboplastin time (PTT) (should be performed                    
within 30 days of leukapheresis) 
j. Serum pregnancy test for females of childbearing potential within 14 days of planned 
leukapheresis. 
3. Confirmation of diagnosis by internal pathology review of initial or subsequent biopsy or other 
pathologic material at Fred Hutch. 
4. Baseline pulse oximetry and documentation of O2 saturation on room air 
5. Baseline 12-lead EKG 
6. Patients age 60 years or older: Echocardiogram or MUGA scan 
7. Baseline imaging (generally CT chest/abdomen/pelvis (and if present affected extremity), MRI 
brain if history of brain metastasis or neurologic symptoms, CT neck if prior cervical 
adenopathy, MRI or PET may be substituted as clinical judgement dictates). Imaging must be 
obtained within 45 days of prior to start of first planned FH-MCVA2TCR infusion 
8. For skin/cutaneous tumors, representative/target tumors should be photographed pre treatment 
for baseline measurements (optional). 
9. Clinical documentation of relapsed disease (imaging and/or biopsy). 
 
Research samples: 
1. PBMC Baseline Samples: A 60 ml blood sample (in EDTA [lavender top] tube) for research 
should be sent to the Specimen Processing/Research Cell Bank lab at the Fred Hutch for PBMC 
isolation and cryopreservation. These cells will be used as PBMC baseline samples for 
comparison to post-treatment samples for all cellular correlative tests, including T cell 
persistence and immunophenotype by flow cytometry or Q-PCR or TCR-β deep sequencing 
(Adaptive Biotechnologies) assays, assessment of anti-transgene, and anti-tumor immune 
responses. These tests are all optional and may be performed depending on tumor response and T 
cell persistence. 
2. Baseline serum: A 20 ml blood sample (serum separator tube) should be sent to the Specimen 
Processing/Research Cell Bank lab at the Fred Hutch to be used as serum baseline to assess anti-
transgene and anti-tumor immune responses. These tests may be performed depending on tumor 
response and T cell persistence. 
3. Other biopsy or sample: If biopsy or sampling of tissues other than bone marrow (i.e., CSF, 
pleural fluid, etc.) is performed for clinical indications, then additional tissue may be obtained 
during the same procedure and sent to the Specimen Processing/Research Cell Bank Lab at the 
Fred Hutch for research studies. Please discuss the planned procedure with the study PI. 
4. RCL Testing: A 10 ml (EDTA/lavender top) blood sample should be collected as baseline before 
both cycles 1 and 2 for RCL testing and sent to the Specimen Processing Lab/ Research Cell 
Bank Lab at the Fred Hutch. 
5. Research RECIST 1.1 read of baseline scans  
6. Biopsy samples: If clinical biopsy performed within 42 days of study enrollment documenting 
persistent/progressive MCC and research samples not collected, a core needle (6 needle passes) 
or punch biopsy (1 x 6 mm or 2 x 4 mm) or equivalent sample will be collected for research if 
clinically feasible. Where possible, research samples will be coordinated with clinical biopsies. 
 
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11.4 
Evaluations prior to T cell Infusion 
On the day of scheduled T cell infusion, the patient should undergo a clinical evaluation and a clinical 
determination for appropriateness to proceed with T cell administration.  
      1.   Interval history and physical exam 
      2.   Research biopsy to evaluate class I MHC upregulation 
      3.   Blood draw for laboratory studies: 
 CBC, differential, and platelet count 
 Comprehensive metabolic panel 
 LDH 
 Uric acid 
 
Research Labs: Cytokine levels (research); T cell baseline persistence, function  
11.5 
Evaluations following T cell infusion 
The following evaluations will be performed after the T cell infusion.  
1. Record new findings on history and physical exam 1 day after the T cell infusion and at least 
weekly for 3 weeks. 
2. The following laboratory studies at least twice weekly for 2 weeks, then weekly until 4 weeks after 
the T cell infusion: 
a. CBC, differential, and platelet count 
b. Comprehensive metabolic panel 
c. LDH 
d. Uric acid  
3. If patients become febrile or develop symptoms of cytokine release or tumor lysis between the 
indicated time points, we may measure serum ferritin, IL-6, CRP, DIC panel, and tumor lysis 
markers at additional times, as clinically indicated. 
 
4. Tumor biopsy: between approximately 10 to 16 days following the T cell infusion, all patients with 
pathologic, palpable or radiologically accessible lymphadenopathy will be asked to undergo a 
punch, excisional or a core biopsy of an accessible tumor. The biopsy at 2-week time point will be 
for research. The requirement for a biopsy will be waived for patients who do not have tumor 
amenable to biopsy. Patients will also be asked to undergo a biopsy at time of 12-week re-
evaluation and may also be asked to undergo a tumor biopsy at the time of relapse. The goal of the 
12 week biopsy will be to confirm that there is progressive/residual tumor if tumors are stable or 
growing (pseudoprogression has been reported at up to 3 months in the JAVELIN trials of 
avelumab for MCC), and to confirm that there is residual tumor if tumors are shrinking (would not 
give second T cell infusion if no residual viable tumor). The purpose of the repeat biopsy at time of 
late relapse would be to confirm that the relapsed lesion represents MCC. These 12-week and 
relapse biopsies will thus be clinical with additional portion/core for research. 
5. Research labs: 
a. Leftover materials will be archived for future studies of T cell function.  All research assays are 
optional and may be performed or omitted or replaced with similar studies based on patient’s 
clinical course.  
b. Serum storage for measurement of serum cytokine levels 
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i. 
Serum cytokine levels. Serum should be collected in serum separator tube at volumes 
indicated. Samples should be sent to the Specimen Processing/Research Cell Bank Lab at 
Fred Hutch.  Serum cytokine levels will be run at selected time points (optional). 
ii. 
If patients become febrile, develop signs of CRS, or cytokine assessment is clinically 
appropriate at times other than those indicated, we may measure cytokine levels at 
additional times (optional). 
c. Evaluation for persistence and phenotype of TCR transgenic T cells 
i. 
PBMC for TCR T cell persistence:  Blood samples (EDTA [lavender top] tube) should be 
obtained on approximately days 1, 3, 7, 14, 21, and 28 during the first month after the T 
cell infusion, and also at 2, 3, 6, 9, and 12 months after the T cell infusion for analysis of 
the persistence and phenotype of transferred T cells. Samples should be sent to the 
Specimen Processing/Research Cell Bank Lab at Fred Hutch. Additional samples may be 
collected at other times than those indicated, including beyond 12 months, if required for 
evaluation of persistence of TCR T cells. Conversely, persistence monitoring may be 
discontinued beyond day 28 after each infusion in patients who do not have detectable 
transgene-expressing T cells on two consecutive occasions. The cells from these blood 
samples may also be analyzed by multiparameter flow cytometry for the phenotype of 
persisting TCR T cells (optional).  
ii. 
If patients become febrile, develop possible signs of toxicity, or assessment of TCR T cell 
persistence is clinically appropriate at times other than those indicated, we may measure 
the persistence of transferred T cells at additional times. 
d. Evaluation of migration of adoptively transferred TCR T cells 
i. 
Single cell tumor digests will be generated from research biopsy tissue and for selected 
patients single cell RNA sequencing will be performed (optional). 
ii. 
Multiplex immunohistochemistry will be performed on clinical and/or research biopsies 
to evaluate T cell localization (optional). 
e. Evaluation for development of endogenous anti-tumor immune responses and epitope 
spreading 
We may evaluate whether any cellular or humoral anti-tumor immune responses resulting from 
activation of endogenous immune cells have occurred. For this purpose, we will collect the 
following. Additional and/or fewer time points may be performed.  Research samples will be 
collected; however, performance of research studies is optional and assays may be substituted 
with alternate assays or omitted as clinical course dictates. 
i. 
Serum anti-tumor immune responses: Serum for humoral immune responses around day 
14, and at approximately 3, 6, and 12 months after T cell infusion to the Specimen 
Processing/Research Cell Bank lab at Fred Hutch. This is for full serologic panel testing 
for research purposes and is separate from the clinical AMERK MCPyV serology. 
ii. 
PBMC anti-tumor immune responses: (lavender top tubes) for cellular immune responses 
around day 14, and at approximately 3, 6, and 12 months after T cell infusion to the 
Specimen Processing/Research Cell Bank lab at Fred Hutch 
iii. 
T-cell repertoire by deep sequencing: Blood (10 ml EDTA [lavender top] tubes) around 
day 14, and at approximately 3 and 6 months after the T cell infusion to the Specimen 
Processing/Research Cell Bank lab at Fred Hutch to evaluate the T-cell repertoire by 
deep sequencing of TCR-β sequence  
Whole exome sequencing and RNA Seq: For patients in whom endogenous immune 
responses are detected, more detailed evaluation may be performed to characterize T cell 
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responses by predicting which tumor mutations are likely to be immunogenic and then 
evaluating peripheral blood T cells for reactivity to these epitopes. To identify mutations 
and determine the patient’s HLA alleles, whole exome sequencing and RNA Seq of 
baseline tumor as well as whole exome sequencing of normal tissue may be performed 
(optional).  
f. RCL testing 
Blood samples should be collected at approximately 3, 6, 9, and 12 months after the TCR T 
cell infusion and sent to the Specimen Processing Lab for RCL testing, as per FDA guidelines.  
 
11.6 
Response Assessment 
Participants will have evaluation of tumor burden at baseline (physical exam with tumor measurements, 
cross-sectional imaging as detailed above, RECIST and imRECIST reads of cross-sectional imaging), at 
4 and 12 weeks after first T cell infusion, at 4 and 12 weeks after second infusion (if given), and then at 
discretion of treating oncologist (suggested frequency every 3-6 months).  Patients with progressive 
disease on 4 week imaging but clinical stability may remain on study given the possibility of delayed 
clinical responses to immunotherapy. Imaging should also be performed if suspicion of progression. 
Imaging will be performed on a clinical basis with RECIST/imRECIST reads on a research basis. For 
skin/cutaneous tumors, representative/target tumors should (optionally) be photographed at key response 
assessment/measurement time points (day 14, 12 weeks, months 3, 6, 9, and 12 post infusion).  Patients 
without measurable lesions will not be evaluable for response, but will be evaluable for safety endpoints. 
 
Participants will have clinical pathology evaluation of tumor at baseline, at approximately 12 weeks 
after first T cell infusion, and if clinically suspected to have progression. Additional research biopsies 
are planned prior to T cell infusion, at 2 weeks post T cell infusion and at 12 weeks after second T cell 
infusion, and additional research tissue may be requested from clinical biopsies. 
 
11.7 
Participant discontinuation of active treatment 
A patient will no longer be able to receive active treatment on study (T cells, avelumab or 
pembrolizumab, or class I MHC upregulation) for any of the reasons listed below. 
 Progressive disease or development of new metastasis requiring urgent change in treatment after 
the second T cell infusion. Patients with progressive disease on 4 week imaging but clinical 
stability may remain on study given the possibility of pseudoprogression and delayed clinical 
responses to immunotherapy. 
 
The participant withdraws consent 
 
Patient death 
 
Occurrence of pregnancy  
 
Participation in another therapeutic trial during the treatment duration of this trial 
 
Occurrence of an exclusion criterion, which is clinically relevant and affects the subject’s safety, 
if discontinuation is considered necessary by the Investigator or Sponsor  
 
A patient will no longer be eligible to receive additional therapy if the PI or his designee 
determines that additional T cell infusions are not in the best interest of the patient 
 Occurrence of any non-pre-existing grade 3 or higher AEs / repetitive Grade 2 Adverse events 
deemed related to the treatment, except for expected, transient toxicities associated with 
avelumab, pembrolizumab, interferon gamma and T cell infusion. 
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 
Cytokine release syndrome requiring pressors for >24 hours or tocilizumab 
 
The reasons for premature discontinuation must be recorded on the case report form. The patient 
may re-enter the study after premature discontinuation only by approval of the PI 
Unless there is withdrawal of consent or death, the patient will transition to the LTFU phase of study. 
 
11.8 
Long-term follow-up (LTFU) 
Enrolled patients who receive FH-MCVA2TCR will be asked to participate in LTFU according to 
guidelines set forth by the FDA’s Biologic Response Modifiers Advisory Committee that apply to gene 
transfer studies. Current recommendations from the FDA suggest a minimum of 15 years of follow-up.  
Annual blood work (20 cc lavender top/10 cc serum separator) will be recommended to be used for RCL 
(replication competent lentivirus; required), persistence, and immune monitoring (optional). Medical 
records will be requested at least annually to determine long term outcomes. Should the participant 
develop a late relapse, a 50 cc blood draw and/or leftover/excess biopsy tissue (fresh and/or archival) 
will be requested for evaluation as to mechanisms of late relapse. Recommendations will be made for an 
autopsy to be conducted if the research participant dies.   
 
12. 
ADVERSE EVENT REPORTING 
12.1 
Adverse Event Definitions 
 Adverse Event 
An Adverse Event (AE) is any untoward medical occurrence in a clinical investigation patient 
administered a medicinal product; the event does not necessarily have a causal relationship with 
study drug administration or usage. An adverse event can therefore be any unfavorable and 
unintended sign (including an abnormal laboratory finding), symptom, or disease temporally 
associated with the use of a medicinal product, whether or not considered related to the 
medicinal product. 
 
 Serious Adverse Event  
A serious adverse event (SAE) is defined as an untoward medical occurrence that results in any 
of the following outcomes: 
1. Death. 
2. Life-threatening situation (i.e., with an immediate risk of death from the event as it occurred 
but not including an event that, had it occurred in a more serious form, might have caused 
death). 
3. In-patient hospitalization or prolongation of existing hospitalization. Inpatient hospitalization 
comprises formal admission to a hospital for medical reasons, for any length of time, whether 
or not hospitalization extends overnight. However, hospital admissions for administration of 
the study drug, procedures required by the study protocol, or tumor-related diagnostic 
procedures are not considered serious. 
4. Persistent or significant disability/incapacity or substantial disruption of the ability to 
conduct normal life functions. 
5. Congenital anomaly/birth defect. 
6. An important medical event that requires intervention to prevent one of the above outcomes. 
 
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 Unexpected Adverse Event 
An unexpected adverse event is defined as an event that has a nature or severity, or frequency 
that is not consistent with the investigator brochure, protocol, or consent form. Please see 
Section 12.6 for expected toxicities. 
 
12.2 
Monitoring and Recording Adverse Events 
Adverse events will be assessed by the investigator or qualified designee and recorded in the CRFs. The 
investigator should attempt to establish a diagnosis of the event on the basis of signs, symptoms and/or 
other clinical information. In such cases, the diagnosis should be documented as the adverse event 
and/or serious adverse event and not described as the individual signs or symptoms. The following 
information should be recorded: 
 Description of the adverse event using concise medical terminology 
 Description as to whether or not the adverse event is serious  
 The start date (date of adverse event onset) 
 The stop date (date of adverse event resolution or return to baseline) 
 The severity (grade) of the adverse event  
 A description of the potential relatedness of the adverse event to study drug or a study procedure, 
as determined by the PI  
 Expectedness of the adverse event based on prior observed and documented adverse events, as 
determined by the PI 
 The outcome of the adverse event 
 
12.3 
Grading of the Severity of an Adverse Event 
Immune effector cells are recognized to have a unique set of toxicities associated with T cell activation 
including cytokine release syndrome (CRS) and neurotoxicity/ICANS. In general, TCR-T therapies have 
not been described to be associated with CRS and neurotoxicity to the same severity and extent that 
certain CAR-T therapies have.  However, given that the underlying mechanism is T cell activation, CRS 
and neurotoxicity management scales designed for CAR-T are reasonably applied to TCR-T.  Therefore, 
for grading cytokine release syndrome and neurotoxicity associated with TCR-T cells we will utilize the 
recently published international consensus guidelines: ASTCT Consensus Grading for Cytokine Release 
Syndrome and Neurologic Toxicity Associated with Immune Effector Cells (Lee et al; 2019).79  
Individual components of CRS (e.g. fever, hypotension, hypoxia) will be graded as part of the CRS 
score, and will not count as separate adverse events, so long as the timing and clinical picture is 
consistent with cytokine release syndrome. 
 
All other AEs will be graded in severity according to the NCI Common Terminology Criteria for 
Adverse Events (CTCAE) Version 5.0. If a CTCAE criterion does not exist, the investigator should use 
the grade or adjectives: Grade 1 (mild), Grade 2 (moderate), Grade 3 (severe), Grade 4 (life-
threatening), or Grade 5 (fatal) to describe the maximum intensity of the adverse event.  
12.4 
Attribution of Adverse Event 
Association or relatedness to the study agent will be assessed by the PI as follows: 
 
Definite 
(must have all 4) 
 
Has a reasonable temporal relationship to the intervention 
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 
Could not have readily been produced by the patient's clinical state 
or have been due to environmental or other interventions 
 
Follows a known pattern of response to intervention 
 
Disappears or decreases with reduction in dose or cessation of 
intervention and recurs with re-exposure
Probable 
(must have 3) 
 
Has a reasonable temporal relationship to the intervention 
 
Could not have readily been produced by the patient's clinical state 
or have been due to environmental or other interventions 
 
Follows a known pattern of response to intervention 
 
Disappears or decreases with reduction in dose or cessation of 
intervention
Possible 
(must have 2) 
 
Has a reasonable temporal relationship to the intervention 
 
Could not have readily been produced by the patient's clinical state
 
Could not readily have been due to environmental or other 
interventions 
 
Follows a known pattern of response to intervention
Unlikely 
(must have 2) 
 
Does not have a temporal relationship to the intervention 
 
Could readily have been produced by the patient's clinical state 
 
Could have been due to environmental or other interventions 
 
Does not follow a known pattern of response to intervention 
 
Does not reappear or worsen with reintroduction of intervention
 
 
For general AE assessment, an AE is considered related if it is assessed as definitely, probably, or 
possibly related; unrelated if it is assessed as unlikely related or unrelated.  
 
For determination of IND safety reporting, AE attribution will be assessed according to the suspected 
adverse reaction definition described in 21 CFR 312.32 as an AE for which there is a reasonable 
possibility that the drug caused the adverse event where “reasonable possibility” means there is evidence 
to suggest a causal relationship between the drug and the AE. The IND Sponsor will report suspected 
adverse reactions that are both serious and unexpected to the FDA as an IND safety report, in 
accordance with regulations under 21 CFR 312.32. 
12.5 
Adverse Event Reporting Period -  
Adverse events of grades 3-5 will be monitored and recorded in study-specific case report forms (CRFs) 
throughout the investigative phases of the study: during and for 48 hours after leukapheresis and then 
from the start of interferon gamma through day 28 after each T cell infusion. Additionally, grade 1-2 
adverse events will be monitored and recorded from the start of T cell infusion through day 28 after each 
T cell infusion. Adverse events will continue to be monitored and recorded in conjunction with standard 
follow-up time points during the first year following T cell infusion. The detailed collection of AEs will 
stop at the time of commencement of new anti-tumor therapy.  
 
AEs with an onset date prior to apheresis will not be recorded, except in the case of clinically significant 
worsening of the AE during the specified monitoring time frame. A patient withdrawn from the study 
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because of an adverse event must be followed until the clinical outcome from the adverse event is 
determined.  
 
The following events are not identified as AEs in this study:  
 Disease progression or relapse.   
 Hospitalization for the purpose of facilitating T cell infusion is not considered an AE.  Any AE 
requiring prolongation of this hospitalization will be recorded and patient to applicable SAE 
reporting. 
 Medical or surgical procedures in and of themselves, including those that require hospitalization 
(e.g., surgery, endoscopy, biopsy procedures) are not considered AEs.  However, an event or 
condition requiring such procedures may be an AE. 
12.6 
Expected Grade 3-4 Adverse Events 
Avelumab, pembrolizumab, interferon gamma, and T cell infusions each have associated, expected, 
transient toxicities that will not be considered grounds for therapy discontinuation nor counted as 
adverse events for determination of treatment safety/dose limiting toxicity. These will however be 
appropriately logged and reported as with any other grade 3-4 toxicity below. 
 
12.6.1 Expected Adverse Events: T cell infusion 
 
The following toxicities are expected adverse events and will not be counted as dose limiting toxicities 
- 
Grade 3-4 lymphopenia. 
Rationale: Lymphopenia is an expected toxicity of T cell therapy, even without lymphodepleting 
chemotherapy.87,88 This was also observed in prior trials of endogenous T cells in MCC (Table 2), 
where 6/12 patients had grade 3 or 4 lymphopenia. In most cases, lymphopenia had onset of 24-72 hours 
after T cell infusion and in all cases lymphopenia had recovered to grade 2 or less by day +28 after T 
cell infusion.  
 
T cell activation has a known constellation of inflammatory responses that is well defined. This is an 
expected toxicity of cellular therapy as the immune system activates and seeks to immunologically reject 
tumor.  There are defined guidelines for cytokine release syndrome (CRS) as well as immune-effector 
cell-associated neurotoxicity syndrome (ICANS). 
 
Therefore, the following toxicities will be evaluated under consensus evaluations for CRS and/or 
ICANS if suspected due to T cell activation (Appendix E)79 and thus individual toxicities will not be 
considered DLT unless CRS and/or ICANS is grade III or higher 
- 
Fevers, hypoxia, hypotension: Fevers, hypoxia, hypotension suspected to be related to cytokine 
release syndrome and occurring within 28 days after T cell infusion will be graded in aggregate 
under the Lee et al cytokine release syndrome criteria consensus guidelines (Appendix E).  The 
individual components will NOT count as DLTs based on CTCAE criteria. 
- 
Tachycardia, flushing, tachypnea: Tachycardia and flushing often accompany CRS as a result of 
the fevers and inflammation and are included in some grading criteria.  Therefore, tachycardia 
(so long as sinus), flushing, and tachypnea will not individually count as DLTs if suspected to be 
due to CRS and CRS will instead be graded with Lee criteria, as above.   
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- 
Headache, confusion, seizure, delirium, and encephalopathy will be evaluated under 
neurotoxicity/ICANS criteria if occurring within 28 days of T cell infusion (Appendix E), and 
ICANS criteria will be used for grading/DLT determination. 
 
12.6.2 Expected Adverse Events: Interferon gamma 
Expected toxicities attributable to interferon gamma and considered exceptions to criteria for 
discontinuation include transient inflammatory responses (Fever, rash, flu like symptoms, outlined in 
7.1) resulting in Grade 3 or 4 toxicities that return to baseline within 2 weeks.   
 
12.6.3 Expected Adverse Events: Avelumab or pembrolizumab 
Expected toxicities attributable to anti-PD-L1 (avelumab) or anti-PD-1 (pembrolizumab) infusions and 
considered exceptions to criteria for discontinuation include infusion-related reactions with Transient (≤ 
24 hours) Grade 3-4 toxicity (outlined in 7.3) that resolved to ≤ Grade 1. Patients with infusion reactions 
to avelumab will be able to receive pembrolizumab for future infusions.  Patients with infusion reactions 
to pembrolizumab will not be able to receive avelumab for future infusions.  
 
12.7 
Adverse Event Reporting Requirements 
12.7.1 Reporting to IRB 
The investigator or designee must report events to the Fred Hutch IRB in accordance with the policies of 
the IRB. 
 
12.7.2 Reporting to Sponsor 
Classification of an event as serious or non-serious determines the reporting procedures to be followed 
by the site for reporting the event to the IND Sponsor.  
PI to IND Sponsor Reporting Requirements for Adverse Events 
 
Classification
Reporting Time
Reporting Action
Contact Information
Serious 
Adverse Event 
(SAE) 
Fatal or 
life-
threatening
Within 24 hours of 
research team* 
awareness 
Email notification 
to Sponsor’s 
Medical Monitor 
& ISIOC 
Administrator
Medical Monitor email: 
mpwerciva@uw.edu 
ISIOC email: 
ISIOC@fredhutch.org 
All SAEs 
Within 2 business 
days of research 
team* awareness 
Submit completed 
Institution-
Sponsored IND 
SAE Reporting 
Form signed by 
PI or designated 
sub-Investigator
ISIOC email: 
ISIOC@fredhutch.org 
Non-serious AE 
Per CRF 
completion 
guidelines 
Record 
information on 
appropriate CRFs 
N/A 
 
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*Research team is defined as the individuals listed on the delegation of authority log. Physicians listed 
on the study’s delegation of authority log as attending physicians with delegated authority to administer 
informed consent will not be considered part of the research team unless additional responsibilities 
related to the conduct of the study have been delegated to them by the Principal Investigator.  
The sponsor assumes responsibility for IND safety reporting to the FDA and participating investigators, 
in accordance with regulations under 21 CFR 312.32. 
The information in the Institution-Sponsored IND SAE Reporting Form must match or be reconciled 
with the information recorded in the adverse events section on the CRF and study database. For 
example, the same adverse event term should be used on both forms. 
 
Each serious adverse event report received from the investigator will be evaluated by the Medical 
Monitor who will assess the seriousness of the event, the expectedness of the event, and the relationship 
to participation in the study, after reviewing the assessments and supporting documentation provided by 
the PI. For regulatory reporting purposes, the IND Sponsor will independently determine expectedness 
relating to the investigational product using safety information specified in the Investigator Brochure, 
the consent document and the protocol document. An event will be classified as related if either the 
investigator or the IND Sponsor determines that the event may be related to the study drug. 
 
The IND Sponsor or designee will provide all investigators with a safety letter notifying them of an 
event that meets FDA IND Safety Reporting criteria. Investigators will be requested to provide written 
notification of safety report to the Fred Hutch IRB as soon as is practical, consistent with IRB 
requirements. 
 
13. 
STATISTICAL CONSIDERATIONS 
13.1 
Sample size and Power 
Our study is a phase I/II, non-blinded interventional trial. Our target sample size is up to 16 patients (8 
per year/2 years). This target is chosen for three reasons: it is achievable from an enrollment standpoint 
(our clinic sees between 15-20 potentially eligible patients per year), it is logistically possible from a 
funding and cell processing facility standpoint, and it allows >80% power to observe a statistically 
significant (at one-sided 0.05 level) efficacy signal that is clinically meaningful. (see below for details).  
With the eligibility expansion, we expect roughly 12 patients will receive a PD1 checkpoint inhibitor.  
For some analyses, all 16 patients will be analyzed, yet some analyses will be restricted to patients who 
receive a PD1 checkpoint inhibitor.  See below for the impact of this on statistical power. 
 
13.2 
Primary Endpoint: Safety 
13.2.1 Definition of treatment related adverse event 
For the purposes of statistical analysis and estimation of toxicity rate, a safety event will be any grade 3 
or higher adverse event determined to be possibly, probably, or definitely secondary to any of the study 
treatments, except as expected in Sections 12.5 and 12.6. Please see Section 12.1 for adverse event 
definition. 
 
13.2.2 Definition of success and probability of suspension 
Platinum-based combination chemotherapy is associated with 30 to 40% grade ≥ 3 toxicities46 and 
although data are not yet available for MCC, combination checkpoint inhibitors see 45-55% treatment 
associated grade ≥ 3 toxicity in patients with metastatic melanoma.69 To avoid exceeding these toxicity 
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rates, the treatment will be considered to have an acceptable safety profile if the observed toxicity rate is 
consistent with a true rate that does not exceed 40%. 
 
Evidence of excessive toxicity will be an observed proportion of toxicities for which the associated 
lower 80% confidence limit exceeds 40%. Operationally, this limit will be met if any of the following 
proportions is observed: 2/2, 3/3-4, 4/5-6, 5/7-8, 6/9-10, 7/11-12, 8/13-14, 9/15-16. Under these rules, if 
the true probability of toxicity is 20%, the probability of suspension is approximately 0.06. If the true 
probability of toxicity is 60%, the probability of suspension is approximately 0.84 (based on 5,000 
simulations). 
 
The initial three part1 patients will be treated and followed for toxicity. All three of these patients will 
receive their first cycle of treatment (T cell infusions) before additional patients will be enrolled. If zero 
or one of the first three part I patients experiences treatment-related and grade ≥3 toxicity, the remaining 
(up to) 13 pts will be treated on the part II portion of the trial. If instead two or more of the first three 
patients experience grade 3 toxicity, at the point that the two toxicities occur, the trial will be suspended 
and, after a detailed review of the potential causes of toxicity, consideration given to changing the cell 
dose or construct pending DSMB review. If any of the first three patients experience treatment related 
death, the trial will be suspended and, after a detailed review of the potential causes of toxicity, 
consideration given to changing the cell dose or construct pending DSMB review. As detailed in Section 
11.7, if at any point any patient experiences grade 3 or higher treatment-related toxicity, that particular 
patient will no longer receive study related active therapies, except as specified in Section 12.6.1. They 
will continue to be followed for response and outcome. 
 
The above toxicity considerations above will be examined among all patients, regardless of their PD1-
contraindication status and whether they get PD-1/PD-L1 inhibition therapy on the trial.  The delay of 
PD-1/PD-L1 inhibition until 14 days after T cell infusion will help differentiate toxicity related 
specifically to PD-1 axis inhibition.  However, if any patient who does not receive a checkpoint inhibitor 
develops a toxicity, enrollment to such patients will be suspended pending review by the DSMB and the 
PI, at which point a decision will be made to continue the trial as is, modify the trial, and halt enrollment 
of patients for whom a checkpoint inhibitor is contraindicated.  
 
13.3 
Primary Endpoint: Efficacy 
13.3.1 Definition of response 
Irradiated and non-irradiated lesions will be separately tracked but response determined in totality. As 
indicated, patients who will be evaluated for this endpoint will have at least one trackable lesion by 
response evaluation criteria in solid tumors (RECIST) 1.1. Response will be a defined as best overall 
response by RECIST 1.1 of complete or partial response for patients with at least one measurable lesion. 
 
13.3.2 Definition of success and power of success 
Responses to alternative therapies in this population of patients with advanced MCC resistant to PD1-
axis blockade are rare, and we therefore will use a 5% response rate as the fixed benchmark upon which 
we hope to improve with the proposed treatment. If the true response rate with TCR-transduced cells is 
25%, 16 patients yield 84% power to detect a statistically significantly improved response rate (one-
sided .05) over the fixed rate of 5%. Three or more responses among 16 pts would occur with 
probability .04 if the true response rate is 5%, so if three or more patients respond (an observed response 
rate of at least 19%), we will conclude that this regimen is potentially efficacious and worthy of further 
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study. If power is instead assessed among the anticipated 12 patients who will receive a checkpoint 
inhibitor, the statistical power is reduced to 78%. 
 
13.4 
Secondary and Exploratory analyses 
Progression-free and overall survival will be estimated using the method of Kaplan and Meier, with time 
zero the time of first T cell infusion. Given the small size of the trial, many of the secondary and 
exploratory endpoints are descriptive or hypothesis generating in nature and are thus underpowered for 
formal statistical analysis. Standard methods will be used to estimate each of the secondary endpoints. 
 
14. 
DATA AND SAFETY MONITORING PLAN 
14.1 
Overall Scope of Monitoring Activities 
Institutional support of trial monitoring will be in accordance with the Fred Hutch /University of 
Washington Cancer Consortium Institutional Data and Safety Monitoring Plan (DSMP). Under the 
provisions of this plan, Fred Hutch Clinical Research Support coordinates data and compliance 
monitoring conducted by consultants, contract research organizations, or Fred Hutch employees 
unaffiliated with the conduct of the study. Independent monitoring visits occur at specified intervals 
determined by the assessed risk level of the study and the findings of previous visits per the institutional 
DSMP.  
 
In addition, protocols are reviewed at least annually and as needed by the Consortium Data and Safety 
Monitoring Committee (DSMC), Fred Hutch Scientific Review Committee (SRC) and the Fred Hutch 
/University of Washington Cancer Consortium Institutional Review Board (IRB). The review 
committees evaluate accrual, adverse events, stopping rules, and adherence to the applicable data and 
safety monitoring plan for studies actively enrolling or treating patients.  The IRB reviews the study 
progress and safety information to assess continued acceptability of the risk-benefit ratio for human 
patients.  Approval of committees as applicable is necessary to continue the study. 
 
The trial will comply with the standard guidelines set forth by these regulatory committees and other 
institutional, state and federal guidelines. The conduct of this trial will be further monitored by an 
independent Data and Safety Monitoring Board (DSMB) in accordance with an approved DSMB 
Charter.  
 
14.2 
Monitoring the Progress of Trial and Safety of Participants 
The first level of trial oversight for this protocol will be provided by the Principal Investigator, the 
Research Nurse, and the Research Coordinator(s), who will provide continuous oversight of the trial.  
These individuals will meet at least monthly to review recently acquired data, stopping rules, and 
adverse events. Serious adverse events will be reviewed upon occurrence to ensure prompt and accurate 
reporting to the IND Sponsor, appropriate committees, and regulatory agencies as described above. The 
data recorded in the research charts and protocol database will be compared with the actual data 
available from the medical record and/or clinical histories. Data detailed in the research case report 
forms (CRFs) will include the nature and severity of all grade 3-5 adverse events. The Principal 
Investigator and all other investigators on the protocol have received formal training in the ethical 
conduct of human research. 
 
The IND Sponsor will ensure routine trial monitoring as described above and will review Serious 
Adverse Events and other reports of safety issues promptly upon receipt from the PI. 
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A DSMB will be in place and will meet after three patients have received treatment (or sooner if 
requested) to review the data as it relates to AEs and determine whether trial may proceed to part II dose 
level.  The purpose of the DSMB meetings is to review the conduct of the trial to date and assess safety 
and toxicity of the study intervention. The DSMB will review all grade 3 or greater CTCAE v5.0 
toxicities and SAEs and determine whether the study should be prematurely discontinued due to 
excessive toxicity consistent with Section 13. Ad hoc meetings may be scheduled as needed.  
 
 
15. 
DATA MANAGEMENT/CONFIDENTIALITY 
The medical record containing information regarding treatment of the patient will be maintained as a 
confidential document, within the guidelines of the Fred Hutch and the University of Washington 
Medical Center.  
The investigator will ensure that data collected conform to all established guidelines for coding 
collection, key entry, and verification. Each patient is assigned a unique patient number to assure patient 
confidentiality. Information forwarded to the FDA, NIH, NCI or other agencies about patients on this 
protocol refers to patients by a coded identifier and not by name.  Patients will not be referred to by this 
number, by name, or by any other individual identifier in any publication or external presentation. The 
licensed medical records department, affiliated with the institution where the patient receives medical 
care, maintains all original inpatient and outpatient chart documents. Additional clinical data may be 
made available from the Fred Hutch core database which is managed and verified independent of the 
research group.   
The research team will maintain Case Report Forms (CRFs) and associated research documentation for 
each patient treated under the protocol. This documentation includes both clinical data and study-
specific documents for each patient. The Principal Investigator or a designee will verify completed 
CRFs against source documentation on an ongoing basis as they are completed for individual patients. 
Data required for analysis of patients treated on this protocol will be maintained in a password-protected 
study-specific database. Data from the CRFs are keyed directly into the database by authorized research 
staff and verified on an ongoing basis.  
 
16. 
TERMINATION OF STUDY 
The study will terminate after the last treated patient has completed 15 years of follow-up as described 
in this protocol.  
 
The PI or IND Sponsor may terminate the study at any time. The IRB and FDA also have the authority 
to terminate the study should it be deemed necessary. 
 
 
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Hodi, F. S. et al. Immune-Modified Response Evaluation Criteria In Solid Tumors (imRECIST): 
Refining Guidelines to Assess the Clinical Benefit of Cancer Immunotherapy. J Clin Oncol 36, 
850-858, doi:10.1200/JCO.2017.75.1644 (2018). 
72 
Heath, M. et al. Clinical characteristics of Merkel cell carcinoma at diagnosis in 195 patients: the 
AEIOU features. J Am Acad Dermatol 58, 375-381, doi:10.1016/j.jaad.2007.11.020 (2008). 
73 
Paulson, K. G. et al. Antibodies to merkel cell polyomavirus T antigen oncoproteins reflect 
tumor burden in merkel cell carcinoma patients. Cancer Res 70, 8388-8397, doi:10.1158/0008-
5472.CAN-10-2128 (2010). 
74 
Paulson, K. G. et al. Viral oncoprotein antibodies as a marker for recurrence of Merkel cell 
carcinoma: A prospective validation study. Cancer 123, 1464-1474, doi:10.1002/cncr.30475 
(2017). 
75 
Moshiri, A. S. et al. Polyomavirus-Negative Merkel Cell Carcinoma: A More Aggressive 
Subtype Based on Analysis of 282 Cases Using Multimodal Tumor Virus Detection. J Invest 
Dermatol 137, 819-827, doi:10.1016/j.jid.2016.10.028 (2017). 
76 
Busam, K. J. et al. Merkel cell polyomavirus expression in merkel cell carcinomas and its 
absence in combined tumors and pulmonary neuroendocrine carcinomas. Am J Surg Pathol 33, 
1378-1385, doi:10.1097/PAS.0b013e3181aa30a5 (2009). 
77 
Latchman, Y. et al. PD-L2 is a second ligand for PD-1 and inhibits T cell activation. Nature 
immunology 2, 261-268, doi:10.1038/85330 (2001). 
78 
Nghiem, P. et al. Durable tumor regression and overall survival in patients with advanced merkel 
cell carcinoma receiving pembrolizumab as first-line therapy. Journal of Clinical Oncology 37, 
693-702 (2019). 
79 
Lee, D. W. et al. ASTCT Consensus Grading for Cytokine Release Syndrome and Neurologic 
Toxicity Associated with Immune Effector Cells. Biol Blood Marrow Transplant 25, 625-638, 
doi:10.1016/j.bbmt.2018.12.758 (2019). 
80 
.  (!!! INVALID CITATION !!! ). 
81 
Coiffier, B., Altman, A., Pui, C. H., Younes, A. & Cairo, M. S. Guidelines for the management 
of pediatric and adult tumor lysis syndrome: an evidence-based review. J Clin Oncol 26, 2767-
2778, doi:10.1200/JCO.2007.15.0177 (2008). 
82 
Howard, S. C., Jones, D. P. & Pui, C. H. The tumor lysis syndrome. N Engl J Med 364, 1844-
1854, doi:10.1056/NEJMra0904569 (2011). 
83 
Schulert, G. S. & Grom, A. A. Pathogenesis of Macrophage Activation Syndrome and Potential 
for Cytokine-Directed Therapies. Annual Review of Medicine 66, 145-159 (2015). 
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84 
Weber, J. S., Kahler, K. C. & Hauschild, A. Management of immune-related adverse events and 
kinetics of response with ipilimumab. J Clin Oncol 30, 2691-2697, 
doi:10.1200/JCO.2012.41.6750 (2012). 
85 
Wolchok, J. D. et al. Guidelines for the evaluation of immune therapy activity in solid tumors: 
immune-related response criteria. Clin Cancer Res 15, 7412-7420, doi:1078-0432.CCR-09-1624 
[pii] 
10.1158/1078-0432.CCR-09-1624 (2009). 
86 
Brahmer, J. R., Lacchetti, C. & Thompson, J. A. Management of Immune-Related Adverse 
Events in Patients Treated With Immune Checkpoint Inhibitor Therapy: American Society of 
Clinical Oncology Clinical Practice Guideline Summary. J Oncol Pract 14, 247-249, 
doi:10.1200/JOP.18.00005 (2018). 
87 
Chapuis, A. G. et al. Tracking the Fate and Origin of Clinically Relevant Adoptively Transferred 
CD8(+) T Cells In Vivo. Sci Immunol 2, doi:10.1126/sciimmunol.aal2568 (2017). 
88 
Chapuis, A. G. et al. T-Cell Therapy Using Interleukin-21-Primed Cytotoxic T-Cell 
Lymphocytes Combined With Cytotoxic T-Cell Lymphocyte Antigen-4 Blockade Results in 
Long-Term Cell Persistence and Durable Tumor Regression. J Clin Oncol 34, 3787-3795, 
doi:10.1200/JCO.2015.65.5142 (2016). 
 
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APPENDIX A:  Schedule of Evaluations 
Assessments
Screening Evaluation (Fred Hutch/local) 
Pre-treatment Evaluation a  
Leukapheresis 
Interferon Gamma 
Pre-Treatment Biopsy 
 T-cell Infusion #1 (Day 0) 
Fred Hutch Post T-
cell Infusion #1 
Evaluations 
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
12-Week Restaging c 
Pre-treatment Evaluationa 
Leukapheresis (if indicated) 
Interferon Gamma 
Pre-treatment Biopsy 
T-Cell Infusion #2 (Day 0)d 
Fred Hutch Post T-cell 
Infusion #2 Evaluations 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
12-Week Restagingc 
Short-term 
Follow-Upc 
Day +1 
Day +3 
Day +7 
Day +10 
Day +14  
(Avelumab  - dose 1) 
Day +21 
 
D+28, D+42, D+56, 
D+70, D+84 
Day +28  
Day +56 
Day +1 
Day +3 
Day +7 (+ Avelumab) 
Day +10 
Day +14 
Day  +21 
 (+ Avelumab) 
Day +28 
Day +56 
Approximately every 
2 weeks 
Month 6, 9, and 12 
post T-cell Infusion 
1#1 
I/E criteria
X
X
Informed 
consent/HIPAA
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Medical history 
– Completeb 
 
X 
 
 
 
 
 
 
 
 
 
 
 
Per 
institutional 
or provider 
routine for 
Avelumabi 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Per 
institutional 
or provider 
routine for 
Avelumabi 
 
 
Medical history 
- Focused
 
 
X
 
 
 
 
X
 
X
 
 
X
 
 
X
 
X
 
X
 
X
 
X
 
 
 
 
 
X
 
X
 
 
X
 
 
X
 
X
 
X
 
X
X 
 
X
Physical exam 
with Vital signs 
(including O2 
sat)
 
X 
 
 
 
X 
 
X
 
 
X
 
 
X 
 
X
 
X 
 
X 
 
X 
 
 
 
 
 
X 
 
X
 
 
X 
 
 
X
 
X 
 
X 
 
X 
 
X 
 
X 
Height/weight 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Karnofsky 
performance 
satus
 
X 
 
 
 
 
X 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
X 
 
 
 
 
 
 
 
 
 
X 
 
X 
Pregnancy test 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
12-lead ECG
X
MUGA/ECHO 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Evaluation of Therapeutic Potential (may be performed at any time prior to treatment) 
Determination 
of tumor viral 
statuse 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Fred Hutch 
pathology 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Assessments
Screening Evaluation (Fred Hutch/local) 
Pre-treatment Evaluation a  
Leukapheresis 
Interferon Gamma 
Pre-Treatment Biopsy 
 T-cell Infusion #1 (Day 0) 
Fred Hutch Post T-
cell Infusion #1 
Evaluations 
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
12-Week Restaging c 
Pre-treatment Evaluationa 
Leukapheresis (if indicated) 
Interferon Gamma 
Pre-treatment Biopsy 
T-Cell Infusion #2 (Day 0)d 
Fred Hutch Post T-cell 
Infusion #2 Evaluations 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
12-Week Restagingc 
Short-term 
Follow-Upc 
Day +1 
Day +3 
Day +7 
Day +10 
Day +14  
(Avelumab  - dose 1) 
Day +21 
 
D+28, D+42, D+56, 
D+70, D+84 
Day +28  
Day +56 
Day +1 
Day +3 
Day +7 (+ Avelumab) 
Day +10 
Day +14 
Day  +21 
 (+ Avelumab) 
Day +28 
Day +56 
Approximately every 
2 weeks 
Month 6, 9, and 12 
post T-cell Infusion 
1#1 
review of any 
tumor lesion 
HLA typing 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Imaging and Tumor Measurementsf 
CT C/A/P or  
PET-CT 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
Xj 
 
 
X 
 
X 
 
 
 
 
 
 
 
 
 
 
 
Xj 
 
 
 
X 
 
X 
CT affected 
area (if not 
covered w/ 
above)
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
MRI Brain 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
Measurement 
and photo of 
palpable tumors
 
X 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
X 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
Interventions 
Clinical Biopsy 
(to pathology)g 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
Research 
Biopsy (may be 
combined w/ 
clinical; to 
Chapuis Lab D3-
235)g 
 
X 
 
 
 
 
X
k 
 
 
 
 
 
 
 
X 
 
 
 
 
 
 
X 
 
 
X 
 
 
 
 
Xk
 
 
 
 
 
 
 
 
 
 
 
 
X 
 

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Assessments
Screening Evaluation (Fred Hutch/local) 
Pre-treatment Evaluation a  
Leukapheresis 
Interferon Gamma 
Pre-Treatment Biopsy 
 T-cell Infusion #1 (Day 0) 
Fred Hutch Post T-
cell Infusion #1 
Evaluations 
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
12-Week Restaging c 
Pre-treatment Evaluationa 
Leukapheresis (if indicated) 
Interferon Gamma 
Pre-treatment Biopsy 
T-Cell Infusion #2 (Day 0)d 
Fred Hutch Post T-cell 
Infusion #2 Evaluations 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
12-Week Restagingc 
Short-term 
Follow-Upc 
Day +1 
Day +3 
Day +7 
Day +10 
Day +14  
(Avelumab  - dose 1) 
Day +21 
 
D+28, D+42, D+56, 
D+70, D+84 
Day +28  
Day +56 
Day +1 
Day +3 
Day +7 (+ Avelumab) 
Day +10 
Day +14 
Day  +21 
 (+ Avelumab) 
Day +28 
Day +56 
Approximately every 
2 weeks 
Month 6, 9, and 12 
post T-cell Infusion 
1#1 
Leukapheresis 
 
 
X
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Interferon 
Gammah 
 
 
 
 
X
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
T-cell Infusion
X
X
Avelumabi 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
 
 
 
 
 
 
 
 
 
X 
 
 
X 
 
 
X 
 
 
Clinical Blood Work / Safety Laboratory Evaluations 
CBC w/ diff, 
platelet count 
 
 
X 
 
 
 
 
 
X 
 
X
 
X
 
X
 
X
 
X 
 
X
 
 
 
 
 
 
 
 
X 
 
X 
 
X 
 
X 
 
 
 
 
X 
 
X
 
X
 
X 
 
X
 
X
 
X 
 
X 
 
X 
 
 
 
 
 
 
 
 
X 
 
X 
Comprehensive 
Metabolic 
Panel
 
X 
 
 
 
X X X X X
X 
X
X 
X 
X 
X 
 
 
 
X X X
X X X
X 
X 
X 
X 
X 
LDH 
 
X 
 
 
 
X X X X X
X 
X
X 
 
X 
X 
 
 
 
X X X
X X X
X 
X 
 
X 
X 

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Assessments
Screening Evaluation (Fred Hutch/local) 
Pre-treatment Evaluation a  
Leukapheresis 
Interferon Gamma 
Pre-Treatment Biopsy 
 T-cell Infusion #1 (Day 0) 
Fred Hutch Post T-
cell Infusion #1 
Evaluations 
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
12-Week Restaging c 
Pre-treatment Evaluationa 
Leukapheresis (if indicated) 
Interferon Gamma 
Pre-treatment Biopsy 
T-Cell Infusion #2 (Day 0)d 
Fred Hutch Post T-cell 
Infusion #2 Evaluations 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
12-Week Restagingc 
Short-term 
Follow-Upc 
Day +1 
Day +3 
Day +7 
Day +10 
Day +14  
(Avelumab  - dose 1) 
Day +21 
 
D+28, D+42, D+56, 
D+70, D+84 
Day +28  
Day +56 
Day +1 
Day +3 
Day +7 (+ Avelumab) 
Day +10 
Day +14 
Day  +21 
 (+ Avelumab) 
Day +28 
Day +56 
Approximately every 
2 weeks 
Month 6, 9, and 12 
post T-cell Infusion 
1#1 
Amylase 
 
 
 
 
 
 
X 
 
Per 
institutional 
or provider 
routine for 
Avelumabi 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
X 
 
 
Per 
institutional 
or provider 
routine for 
Avelumabi 
 
 
 
Lipase
X
X
X
Uric acid 
X
 
 
X
X X X X
X 
X
X 
 
X 
X 
 
 
 
X X X
X X X
X 
X 
 
X 
X 
Serum ferritin 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
CRP 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
IL-6 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
Cortisol 
 
X 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
TSH 
 
X 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
Free T4 
 
X 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
DIC Panel w/o 
platelets
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
X
 
X
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
Fred Hutch 
(PSBC) donor 
battery panel
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
MCPyV 
Serology 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
Research Blood Work (all to Research Cell Bank) 
60 mL lavender 
top
 
X  
 
 
 
 
 
X  
X 
 
 
 
 
X  
 
 
 
 
 
 
X 
 
X  
 
 
 
 
X 
20 mL serum 
sep
 
X  
 
 
 
 
 
 
 
X 
 
 
 
 
X  
 
 
 
 
 
 
 
 
X  
 
 
 
 
X 
40 mL lavender 
top
 
 
 
 
 
X 
X X  
 
 
 
 
X 
X 
 
 
 
 
 
X 
X X  
 
 
 
X 
X 
 
 
 

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Assessments
Screening Evaluation (Fred Hutch/local) 
Pre-treatment Evaluation a  
Leukapheresis 
Interferon Gamma 
Pre-Treatment Biopsy 
 T-cell Infusion #1 (Day 0) 
Fred Hutch Post T-
cell Infusion #1 
Evaluations 
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
12-Week Restaging c 
Pre-treatment Evaluationa 
Leukapheresis (if indicated) 
Interferon Gamma 
Pre-treatment Biopsy 
T-Cell Infusion #2 (Day 0)d 
Fred Hutch Post T-cell 
Infusion #2 Evaluations 
(Fred 
Hutch/local-
may combine 
w/ Avelumab 
eval 
bloodwork)
Fred 
Hutch/Local 
Avelumab 
infusion 
daysi 
12-Week Restagingc 
Short-term 
Follow-Upc 
Day +1 
Day +3 
Day +7 
Day +10 
Day +14  
(Avelumab  - dose 1) 
Day +21 
 
D+28, D+42, D+56, 
D+70, D+84 
Day +28  
Day +56 
Day +1 
Day +3 
Day +7 (+ Avelumab) 
Day +10 
Day +14 
Day  +21 
 (+ Avelumab) 
Day +28 
Day +56 
Approximately every 
2 weeks 
Month 6, 9, and 12 
post T-cell Infusion 
1#1 
10 mL serum 
sep
 
 
 
 
 
X 
X X X  
 
X  
X 
X 
 
 
 
 
 
X 
X X X 
 
 
X 
X 
X 
 
 
 
20 mL lavender 
top
 
 
 
 
 
 
 
 
 
 
 
X  
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
 
NOTE: The proposed days of all treatments and assessments are approximate and may vary due to scheduling, clinical, or other factors 
a 
Tests obtained as part of routine clinical care may be substituted, and may be omitted for Cycle 2 evaluation as long as within protocol-specified window 
b 
Medical history should initially include a full history of all findings at diagnosis, time of enrollment, prior history, and response to therapy, as well as 
relevant other medical issues, family and social history, medications, allergies.  Subsequent medical histories may be focused as appropriate 
c 
12-week evaluation strongly preferred to be at Fred Hutch and 6, 9, and 12-month evaluations preferred at Fred Hutch. However, if patient cannot travel, 
these evaluations may be performed locally 
d 
Please see section 6.4 for details on determination of one versus two cycles if T cell therapy, and guidance for timing of second T-cell administration 
e 
Please see inclusion criteria for further details on determination of tumor virus status 
f 
Diagnostic imaging evaluations preferred to be at Fred Hutch but may be performed elsewhere if travel or insurance issues preclude Fred Hutch scans. 
Baseline imaging must be obtained within 45 days prior to start of first planned FH-MCVA2TCR infusion.  
g 
Biopsies may be omitted or retimed at discretion of PI based on clinical appropriateness 
h 
Administered 3 times weekly through d28 
i 
Please see section 6.5 for additional details and suggested guidance on standard of care avelumab or pembrolizumab. If patients receive pembrolizumab, 
certain evaluations may be rescheduled to match the pembrolizumab schedule (e.g., the D28 evaluations may be performed at D35, and the D84 evaluations 
may be performed at D77). 
j 
CT chest/abdomen/pelvis (CAP) + involved areas w/ IV contrast mandatory, PET CT optional per clinical status 
k 
Biopsy after 2-4 doses of IFN-G, day of or day before cell infusion 
 

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APPENDIX B: Research Sample Checklist  
 
Please note that many of the listed research studies are optional pending clinical course, and may 
be replaced by newer or alternate assays that answer the same general question. Note that the 
selected studies will be batched, and results are not anticipated to be available in “real-time”.  
Leftover materials from all time points may be archived. 
 
RECIPIENT RESEARCH EVALUATIONS BEFORE T CELL INFUSION 
SAMPLE
 
TIME 
TEST 
TUBE 
VOL. 
(Approx
)
LAB 
Blood 
At 
Enrollment 
T cell persistence and 
functional studies, epitope 
spreading, scRNAseq, 
clonality, RCL baseline
Lavender 
top 
60 mL 
RCB 
Blood 
At 
Enrollment
Cytokine levels and MCPyV 
antibody levels (research)
Serum 
separator
20 ml 
RCB 
Tumor 
At 
Enrollment
Single cell RNA sequencing 
RPMI 
4 cores 
or equiv
ICTPL 
Tumor 
At 
Enrollment 
T cell localization and tumor 
microenvironment 
characteristics 
Formalin 
2 core or 
equiv; in 
addition 
to 
clinical 
core
Chapuis 
D3-235 
Tumor 
At 
Enrollment 
Archive 
RPMI 
Any 
addtl 
tissue
Chapuis 
D3-235 
Tumor 
Pre-
Treatment
Single cell RNA sequencing 
RPMI 
4 cores 
or equiv
ICTPL 
Tumor 
Pre-
Treatment 
T cell localization and tumor 
microenvironment 
characteristics 
Formalin 
2 core or 
equiv; in 
addition 
to 
clinical 
core
Chapuis 
D3-235 
Tumor 
Pre-
Treatment 
Archive 
RPMI 
Any 
addtl 
tissue
Chapuis 
D3-235 
Blood 
Day 0 
Cytokine levels (research) 
Serum 
separator
10 ml 
RCB 
Blood 
Day 0 
T cell baseline persistance, 
function
Lavender 
top
40 ml 
RCB 
 
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RECIPIENT RESEARCH EVALUATIONS AFTER T CELL INFUSION 
SAMPLE
TIME 
TEST 
TUBE 
VOL. 
(Approx) 
LAB 
Blood 
Day 1 
Cytokine levels (research)  
Serum 
separator 
10 ml 
RCB 
Blood 
Day 1 
T cell persistence, phenotype 
Lavender 
top
40 ml 
RCB 
Blood 
Day 3 
Cytokine levels (research)  
Serum 
separator
10 ml 
RCB 
Blood 
Day 3 
T cell persistence and 
functional studies
Lavender 
top
   40 ml 
RCB 
Blood 
Day 7 
Cytokine levels (research) 
Serum 
separator
10 ml 
RCB 
Blood 
Day 7 
T cell persistence and 
functional studies
Lavender 
top
    60 ml 
RCB 
Blood 
Day 14 
Cytokine levels (research) 
Serum 
separator
20 ml 
RCB 
Blood 
Day 14 
T cell persistence and 
functional studies, epitope 
spreading, scRNAseq, clonality
Lavender 
top 
60 ml 
RCB 
Blood 
Day 21 
Cytokine levels (research) 
Serum 
separator
10 ml 
RCB 
Blood 
Day 21 
T cell persistence 
Lavender 
top
20 ml 
RCB 
Blood 
Day 28 
Cytokine levels (research) 
Serum 
separator 
10 ml 
RCB 
Blood 
Day 28 
T cell persistence, 
immunophenotype
Lavender 
top
40 ml 
RCB 
Blood 
Day 56 
Cytokine levels (research) 
Serum 
separator
10 ml 
RCB 
Blood 
Day 56 
T cell persistence, 
immunophenotype
Lavender 
top
40 ml 
RCB 
Blood 
Day 84 
T cell persistence, epitope 
spreading, scRNAseq, 
immunophenotype, clonality, 
RCL
Lavender 
top 
60 ml 
RCB 
Blood 
Day 84 
Cytokines, research serologies 
Serum 
separator
20 ml 
RCB 
If second infusion, please refer to study schedule 
Blood 
Day 180 
T cell persistence, epitope 
spreading, immunophenotype, 
clonality, RCL
Lavender 
top 
60 ml 
RCB 
Blood 
Day 180 
Cytokines, research serologies 
Serum 
separator
20 ml 
RCB 
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Blood 
Day 270 
T cell persistence, epitope 
spreading, immunophenotype, 
clonality, RCL
Lavender 
top 
60 ml 
RCB 
Blood 
Day 270 
Cytokines, research serologies 
Serum 
separator
20 ml 
RCB 
Blood 
Day 365 
T cell persistence, epitope 
spreading, immunophenotype, 
clonality, RCL
Lavender 
top 
60 ml 
RCB 
Blood 
Day 365 
Cytokines, research serologies 
Serum 
separator
20 ml 
RCB 
Blood 
Annual years 
2-15
T cell persistence, RCL 
Lavender 
top
20 ml 
RCB 
Blood 
Annual years 
2-15
serologies 
Serum 
separator
10 ml 
RCB 
Blood 
Clinical 
events 
T cell persistence, epitope 
spreading, immunophenotype, 
clonality, RCL, scRNAseq (all 
optional/TBD)
Lavender 
top 
Up to 60 
ml 
Discuss 
with 
study 
staff
Blood 
Clinical 
events 
Cytokine levels 
Serum 
separator 
20 mL 
Discuss 
with 
study 
staff
Tumor 
Day + 14 
Single cell RNA sequencing 
RPMI 
4 cores 
or equiv
Chapuis 
D3-235
Tumor 
Day + 14 
T cell localization and tumor 
microenvironment 
characteristics
Formalin 
2 cores 
or equiv 
Chapuis 
D3-235 
Tumor 
Day +14 
Archive 
RPMI 
Any 
addtl 
tissue
Chapuis 
D3-235 
Tumor 
Day + 84 
Single cell RNA sequencing 
RPMI 
4 cores 
or equiv
Chapuis 
D3-235
Tumor 
Day + 84 
T cell localization and tumor 
microenvironment 
characteristics 
Formalin 
2 cores 
or equiv; 
in 
addition 
to 
clinical 
core(s)
Chapuis 
D3-235 
Tumor 
Day + 84 
Archive 
RPMI 
Any 
addtl 
tissue
Chapuis 
D3-235 
Tumor 
Clinical 
event
Single cell RNA sequencing 
RPMI 
4 cores 
or equiv
Chapuis 
D3-235
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Tumor 
Clinical 
event 
T cell localization and tumor 
microenvironment 
characteristics 
Formalin 
2 cores 
or equiv; 
in 
addition 
to 
clinical 
core
Chapuis 
D3-235 
 
 
 
 
 
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APPENDIX C: Karnofsky Performance Status Scale 
 
KARNOFSKY PERFORMANCE STATUS SCALE 
 
General
Index
Specific Criteria
Able to carry on normal activity; no 
special care needed 
100 
Normal, no complaints, no evidence of 
disease 
 
90 
Able to carry on normal activity, 
minor signs or symptoms of disease 
 
80 
Normal activity with effort, some 
signs or symptoms of disease 
Unable to work, able to live at home and 
care for most personal needs, varying 
amount of assistance needed 
70 
Care for self, unable to carry on 
normal activity or to do work 
 
60 
Requires occasional assistance from 
others but able to care for most needs 
 
50 
Requires considerable assistance from 
others and frequent medical care 
Unable to care for self, requires 
institutional or hospital care or 
equivalent; disease may be rapidly 
progressing
40 
Disabled; requires special care and 
assistance 
 
30 
Severely disabled, hospitalization 
indicated, death not imminent 
 
20 
Very sick, hospitalization necessary, 
active supportive treatment necessary 
 
10 
Moribund 
 
0 
Dead 
 
 
 
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APPENDIX D: ECOG Performance Status Scale 
 
ECOG Performance Status Scale 
 
GRADE
SCALE
0 
Fully active, able to carry out all pre-disease performance without restriction 
1 
Restricted in physically strenuous activity but ambulatory and able to carry 
out work of a light or sedentary nature, e.g., light housework, office work 
2 
Ambulatory and capable of all self-care but unable to carry out work 
activities. Up and about more than 50% of waking hours.  
3 
Capable of only limited self-care, confined to bed or chair more than 50% of 
waking hours.  
4 
Completely disabled. Cannot carry on any self-care. Totally confined to bed 
or chair. 
5 
Dead 
 
 
 
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APPENDIX E: CRS and Neurotoxicity Grading Criteria 
 
 
 
CRS grading will be as per ASTCT CRS Consensus Grading. Table reprinted for reference from 
the guideline of Lee et al, 2019 ASTCT Consensus Grading for Cytokine Release Syndrome and 
Neurologic Toxicity Associated with Immune Effector Cells, Biol Blood Marrow Transplant 
25(2019) 625-638 
 
 
 
 
 
Neurotoxicity/ICANS grading will be as per ASTCT CRS Consensus Grading. Table reprinted 
for reference from the guideline of Lee et al, 2019 ASTCT Consensus Grading for Cytokine 
Release Syndrome and Neurologic Toxicity Associated with Immune Effector Cells, Biol Blood 
Marrow Transplant 25(2019) 625-638 
 
 
 
 
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APPENDIX F: Recommended Management of CRS 
 
Grade (Lee et al 2019) 
Suggested Treatment 
1 
- 
Manage symptomatically with acetaminophen and 
antiemetics prn  
- 
Fluids 
- 
Administer antibiotics per institutional guidelines, 
evaluate fever 
- 
If not on Keppra, initiate Keppra 500mg PO BID 
2 
- 
Manage symptomatically with fluids and oxygen 
- 
If not responsive to oxygen and fluids, Tocilizumab 
8mg/kg IV, max 800 mg in a single dose, may repeat 
q8h if needed, to max of 3 doses in 24 hours 
- 
Consider dexamethasone 10 mg IV q12H x 2 doses 
- 
If not on Keppra, initiate Keppra 500mg PO BID 
- 
Administer antibiotics per institutional guidelines, 
evaluate fever 
- 
Continue symptomatic support
3 
- 
Transfer to ICU if vasopressors are required. Other 
clinical settings may also require ICU transfer 
- 
Tocilizumab 8 mg/kg IV, may repeat q8h if needed, to 
max of 3 doses in 24 hours 
- 
Dexamethasone 10 mg IV q6-12H 
- 
Administer antibiotics per institutional guidelines, 
evaluate fever 
- 
If not on Keppra, initiate Keppra 500mg PO BID
4 
- 
Transfer to ICU 
- 
Tocilizumab 8 mg/kg IV, may repeat q8h if needed, to 
max of 3 doses in 24 hours 
- 
Methylprednisolone 1000 IV qd x 3 days  
- 
If not on Keppra, initiate Keppra 500mg PO BID 
- 
Administer antibiotics per institutional guidelines, 
evaluate fever 
- 
Contact study PI and immunotherapy team for 
additional anti-cytokine therapy recommendations
Suggested guidelines for CRS management based on Lee et al 2019 CRS grade.  Clinical flexibility is 
allowed by treating Immunotherapy attending physician based on his/her clinical judgement and 
experience. Recommendations for CRS management are evolving rapidly; guidelines may be substituted 
with Fred Hutch/UW institutional best practices and/or national companion guidelines to Lee et al 
guidelines if newer information available. 
 
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APPENDIX G - Long Term Follow-Up 
Study participants should be asked to participate in long term follow-up, as directed by the FDA 
Guidance for Industry – Gene Therapy Clinical Trials: Observing Subjects for Delayed Adverse 
Events. 
(http://www.fda.gov/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/Guid
ances/CellularandGeneTherapy/ucm072957.htm#5). 
 
Long term follow-up should commence one year after the final T cell infusion. The planned 
recommendations for follow-up are as follows:  
 
Years 1 - 15: 
 
1. Recommendation that patients undergo at least annual history and physical examination with 
their primary physician: 
 Adverse event screening guidance for the primary physician in the form of a gene 
therapy LTFU-directed screening survey may be available. 
 A request for the study team to be notified of all new malignancies and unexpected 
illnesses. 
 The primary physician may be provided with a blood draw courier kit to enable 
samples to be returned to the Chapuis Lab for archival purposes, and for analysis for 
transgene and vector persistence, and RCL, as dictated by studies of transferred T cell 
persistence. 
 If the patient develops a late recurrence, request for up to 50 cc of research blood as 
well as leftover or archival biopsy/surgical tissue, in order to evaluate for mechanisms 
of late recurrence (may be collected remotely with a courier kit) 
 
2. Annual phone call survey or questionnaire to the participant to screen for adverse events. 
 
3. Offer the opportunity to return to Fred Hutch for an annual LTFU clinic visit. 
 
4. Compliance with 21 CFR 312.32 in adverse event reporting. 
 
5. Research bloodwork: 30 cc of blood annually for persistence and RCL monitoring 
 
 
 
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