Conditions: Merkel Cell Polyomavirus Infection Stage IV Merkel Cell Carcinoma AJCC v7
Allocation: NON_RANDOMIZED | Intervention model: PARALLEL
Enrollment: 8
Healthy volunteers: no

Brief summary:
This phase I/II trial studies the side effects and how well localized radiation therapy or recombinant interferon beta and avelumab with or without cellular adoptive immunotherapy works in treating patients with Merkel cell carcinoma that has spread to other parts of the body. Radiation therapy uses high energy x-rays to kill tumor cells and shrink tumors. Interferon beta is a substance that can improve the body's natural response and may interfere with the growth of tumor cells. Monoclonal antibodies, such as avelumab, may help T lymphocytes kill tumor cells. For cellular adoptive immunotherapy, specific white blood cells are collected from the patient's blood and treated in the laboratory to recognize Merkel cell carcinoma. Infusing these cells back into the patient may help the body build an effective immune response to kill Merkel cell carcinoma. Giving localized radiation therapy or recombinant interferon beta and avelumab with or without cellular adoptive immunotherapy may be a better treatment for Merkel cell carcinoma.

Detailed description:
PRIMARY OBJECTIVES:

I. Assess and compare the safety and potential toxicities associated with treating patients with metastatic Merkel cell carcinoma (MCC) with either major histocompatibility complex (MHC) up regulation and programmed cell death 1 (PD1)-axis blockade (Group 1), or MHC up-regulation, PD1-axis blockade and adoptive transfer of Merkel cell polyoma virus (MCPyV) T antigen (TAg)-specific polyclonal autologous cluster of differentiation (CD)8+ T cells (Group 2).

II. Assess and compare the antitumor efficacy associated with treating patients with metastatic MCC with either MHC up-regulation and PD1-axis blockade (Group 1), or MHC up-regulation, PD1-axis blockade and adoptive transfer of MCPyV TAg-specific polyclonal autologous CD8+ T cells (Group 2).

SECONDARY OBJECTIVES:

I. Examine the in vivo persistence and, where evaluable, migration to tumor sites of adoptively transferred polyclonal CD8+ T cells targeting the MCPyV TAg (Group 2).

II. Examine the in vivo functional capacity of adoptively transferred polyclonal CD8+ T cells targeting the MCPyV Tag (Group 2).

III. Examine and compare evidence of epitope spreading with either MHC up-regulation and adoptive transfer of MHC up-regulation and PD1-axis blockade (Group 1), or MHC up regulation, PD1-axis blockade and adoptive transfer of MCPyV TAg-specific polyclonal autologous CD8+ T cells (Group 2).

OUTLINE: Patients are assigned to 1 of 2 groups.

GROUP 1: Patients who do not have a human leukocyte antigen (HLA) type for which T cells can be generated or for whom T cells cannot be generated for technical issues receive avelumab intravenously (IV) over 1 hour every 2 weeks for 12 months. Within 7-10 days after completion of 1-3 doses of avelumab, patients receive MHC class I up-regulation intervention comprising either localized radiation therapy or recombinant interferon beta via intra-tumor injection.

GROUP 2: Patients who have an HLA type for which T cells can be generated receive avelumab IV over 1 hour every 2 weeks for 12 months. Patients also receive MHC class I up-regulation intervention as in Group 1 between 7-10 days after the first infusion of avelumab and 2-5 days before the first infusion of MCPyV TAg-specific polyclonal autologous CD8+ T cells. Patients receive two infusions of MCPyV TAg-specific polyclonal autologous CD8+ T cells IV over 60-120 minutes.

In both groups, MHC class I up-regulation treatment with or without T cell infusions may repeat if indicated.

After completion of study treatment, patients are followed up at 12 months and then periodically thereafter.

Arms:
arm 1: Patients who do not have a HLA type for which T cells can be generated or for whom T cells cannot be generated for technical issues receive avelumab intravenously (IV) over 1 hour every 2 weeks for 12 months. Within 7-10 days after completion of 1-3 doses of avelumab, patients receive MHC class I up-regulation intervention comprising either localized radiation therapy or recombinant interferon beta via intra-tumor injection. arm 2: Patients who have an HLA type for which T cells can be generated receive avelumab IV over 1 hour every 2 weeks for 12 months. Patients also receive MHC class I up-regulation intervention as in Group 1 between 7-10 days after the first infusion of avelumab and 2-5 days before the first infusion of MCPyV TAg-specific polyclonal autologous CD8+ T cells. Patients receive two infusions of MCPyV TAg-specific polyclonal autologous CD8+ T cells IV over 60-120 minutes.

Interventions: intervention 1: Avelumab intervention 2: Laboratory Biomarker Analysis intervention 3: MCPyV TAg-specific Polyclonal Autologous CD8-positive T Cells intervention 4: Radiation Therapy intervention 5: Recombinant Interferon Beta
Intervention descriptions: intervention 1: Given IV intervention 2: Correlative studies intervention 3: Given IV intervention 4: Undergo radiation therapy intervention 5: Given via intra-tumor injection

Protocol document:
Prot_SAP_000.pdf: 
    FRED HUTCHINSON CANCER RESEARCH CENTER (FHCRC) 
UNIVERSITY OF WASHINGTON SCHOOL OF MEDICINE (UW) 
 
 
PROTOCOL 9245 
 
Current Version: 06/04/2018 
Previous Version: 11/07/2017 
 
1. PROTOCOL: Study to Evaluate Cellular Adoptive Immunotherapy 
Using Polyclonal Autologous CD8+ Antigen-Specific T Cells for 
Metastatic Merkel Cell Carcinoma in Combination with MHC Class I Up-
regulation and the Anti-PD-L1 Antibody Avelumab. 
 
    
Investigator 
Professional Title 
Telephone 
Number 
Aude Chapuis, MD 
Principal Investigator (PI), 
Assistant Member, FHCRC 
Assistant Professor, 
UWMC 
(206) 667-4369 
Shailender Bhatia, MD 
 
Assistant Professor, 
Division of Oncology, 
UWMC 
(206) 606-6210 
Paul Nghiem, MD 
Professor, UWMC 
(206) 221-2632 
Upendra Parvathaneni, MD 
Assistant Professor, 
Radiation Oncology, 
UWMC 
(206) 598-4100 
 
Biostatistican:   
Ted Gooley  
 
 
     (206) 667-6533 
 
 
    Study Coordinator 
Cari Morin  
 
 
 
      (206) 667-5901 
 
 
 
 
       
 
 
 
Emergency 24-hour phone: (206) 598-6190 
 
IRO received 06/28/21
FHCRC IRB Approval  
07/23/2021 
Document Released Date


FHCRC 9245 
 
 
 
 
                  Page 2 
 
Table of Contents 
 
1. TITLE OF PROTOCOL: Study to Evaluate Cellular Adoptive 
Immunotherapy Using Polyclonal Autologous CD8+ Antigen-Specific T Cells 
for Metastatic Merkel Cell Carcinoma in Combination with MHC Class I Up-
regulation and the Anti-PD-L1 Antibody Avelumab ..................................................... 1 
2. INTRODUCTION ........................................................................................................... 4 
3. BACKGROUND ............................................................................................................ 5 
3.A.  MCC: An Often Lethal, Virus-driven Cancer ......................................................... 5 
3.B.  Spontaneous Immune Responses to MCPyV TAg ............................................... 6 
3.C.  Identification and Characterization of MCPyV-specific CD8+ T Cells from a  
MCC Patient’s Tumor and Peripheral Blood ................................................................. 7 
3.D.  Identification of Reactive MCPyV TAg-specific HLA-restricted Peptides .............. 9 
3.E.  Adoptive T-Cell Therapy ..................................................................................... 10 
3.F.  Induction of MHC Expression by Tumor Tissue .................................................. 12 
3.G.  Previous Human Experience: Adoptive Therapy of MCC Targeting  
MCPyV TAg ................................................................................................................ 14 
3.H.  Previous Clinical Experience: Results of Protocols FHCRC #2558 and #2586 .. 17 
3.I. Rationale for Combining MHC Class I Up-regulation, MCPyV-Specific T-Cell 
Infusions and PD1 - PD-L1 Axis Blockade .................................................................. 19 
3.J. Rationale for Assessing T-Cell Effectiveness by ‘Time to New Metastasis’ ......... 27 
4.  OBJECTIVES ............................................................................................................ 28 
4.A.  Primary Objectives ............................................................................................. 28 
4.B.  Secondary Objectives ........................................................................................ 28 
5.  STUDY ENDPOINTS ................................................................................................. 28 
5.A.  Primary Endpoints .............................................................................................. 28 
5.B.  Secondary Endpoints ......................................................................................... 28 
6.  STUDY DESIGN ........................................................................................................ 29 
7.  ELIGIBILITY FOR TREATMENT ............................................................................... 31 
7.A.  Inclusion Criteria ................................................................................................. 31 
7.B.  Exclusion Criteria ............................................................................................... 31 
8.  CONSENTING ........................................................................................................... 33 
9.  PROTOCOL REGISTRATION ................................................................................... 33 
10. STUDY AGENTS ...................................................................................................... 33 
10.A.  MCPyV-specific CD8+ Cells.............................................................................. 33 
10.B.  Anti PD-L1 Antibody Avelumab ........................................................................ 34 
10.C.  Radiation Therapy ............................................................................................ 36 
10.D.  Intralesional IFNβ-1B Injection ......................................................................... 37 
10.E. Guidelines for selecting radiation therapy or intralesional IFNβ-1B Injections…36 
11.  PLAN OF TREATMENT .......................................................................................... 38 
12.  EVALUATION .......................................................................................................... 39 
12.A.  Patient Evaluation at Each Planned Visit ......................................................... 39 
12.B.  Patient Evaluation During T Cell Infusions ....................................................... 40 
12.C.  Clinical and Laboratory Evaluation for Toxicity (General Toxicity Assessment) 40 
12.D.  Efficacy Assessment ........................................................................................ 40 
12.E.  Evaluation of Persistence and Function of Adoptively Transferred T Cells ...... 41 


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12.F.  Assessment of Epitope Spreading .................................................................... 43 
13.  MANAGEMENT OF TOXICITIES AND COMPLICATIONS ..................................... 43 
13.A.  Toxicity Grading ............................................................................................... 43 
13.B.  Regimen Related Toxicity................................................................................. 43 
13.C.  Management of Symptoms During Avelumab Infusions ................................... 45 
13.D.  Management of Symptoms During T-Cell Infusion ........................................... 49 
13.E.  Management of Severe Treatment-related Toxicities with Corticosteroids ....... 51 
13.F.  Concomitant Therapy ....................................................................................... 51 
13.G.  Off-study Criteria .............................................................................................. 52 
14.  TARGETED/PLANNED ENROLLMENT ................................................................. 53 
15.  GUIDELINES FOR ADVERSE EVENTS REPORTING ........................................... 54 
15.A.  Reporting of Adverse Events (AEs) .................................................................. 54 
15.B.  Definitions......................................................................................................... 54 
16.  DATA AND SAFETY MONITORING PLAN............................................................. 55 
16.A.  Primary Monitoring ........................................................................................... 55 
16.B.  Monitoring Plan ................................................................................................ 55 
16.C.  Monitoring the Progress of the Trial and the Safety of Participants .................. 55 
17.  RECORDS ............................................................................................................... 56 
18.  STATISTICAL CONSIDERATIONS ......................................................................... 57 
19.  ADMINISTRATIVE CONSIDERATIONS ................................................................. 60 
19.A.  Institutional Review Board ................................................................................ 60 
19.B.  Termination of Study ........................................................................................ 60 
20.  REFERENCES ......................................................................................................... 60 
APPENDIX A .................................................................................................................. 66 
APPENDIX B .................................................................................................................. 67 
APPENDIX C ………………………………………………………………………………….. 69 
 
 
 
 


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2. INTRODUCTION 
 
Merkel cell carcinoma (MCC) represents a highly aggressive neuroendocrine skin 
malignancy with a high disease-associated mortality and a high propensity for recurrence 
after initial treatment. The cause-specific mortality ranges from 23% to 80% at five years 
thus making it three times as lethal as melanoma, and the increase in annual incidence 
now makes MCC the second most common cause of non-melanoma skin cancer-
associated death.1 Strong epidemiologic data have linked MCC to patients with cellular 
immune deficiencies.2,3 In 2008, this association was further strengthened when a newly 
discovered viral agent related to SV40, the Merkel cell polyomavirus (MCPyV), was 
found to be clonally integrated into approximately 80% of MCC tumors.4 The virus 
produces T Antigen (TAg) oncoproteins. TAg oncoproteins are persistently expressed by 
MCC and are necessary for the survival and proliferation of tumor cells and thus 
represent an optimal target for immunotherapy.  
 
Original research in Dr. P. Nghiem’s lab has led to the initial identification of an 
HLA-A*2402-restricted MCPyV TAg epitope recognized by autologous CD8+ T cells in a 
patient with metastatic MCC followed by epitopes recognized by HLA-B*0702, 
HLA-B*3502, HLA-A*0201, HLA-A*0301, HLA-A*1101, and HLA-A*2301. As adoptively 
transferred antigen-specific T cells have demonstrated a significant clinical benefit in a 
subset of patients with metastatic melanoma targeting endogenous non-viral epitopes, 
we initially applied the use of antigen-specific T cells to target the TAg oncoproteins in 
patients with MCPyV-positive MCC tumors. Patients in Group 1 received targeted 
radiation therapy or intra-lesional injection of Interferon-beta 1B (IFNβ-1B) as a means to 
up-regulate previously down-regulated MHC expression by tumor tissue prior to adoptive 
transfer of antigen-specific CD8+ T cells, and followed by low-dose Interleukin-2 (IL-2) to 
improve T-cell survival. However, the transferred cells were shown to express the 
negative regulator antigen PD-1 in vivo; and although the treatment was well tolerated 
and safe, this may have accounted for the lack of tumor control in Group 1 / Protocol 
#2586.  
 
Therefore, we propose to assess whether the anti-tumor efficacy of adoptive transfer of 
MCPyV-specific T cells could be enhanced with concurrent PD-1/PD-L1 axis blockade 
with the anti-PD-L1 antibody Avelumab. This protocol proposes to treat 20 patients with 
metastatic MCC using a combined approach: Group 1 (10 pts) with MHC class I up-
regulation and Avelumab every 2 weeks for 6 months, and Group 2 (10 pts) with MHC 
class I up-regulation, Avelumab and adoptive transfer of MCPyV-specific T cells. The 
safety and efficacy of this approach will be examined along with the in vivo persistence 
and function of the transferred cells.  
 


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3. BACKGROUND 
3.A.  MCC: An Often Lethal, Virus-driven Cancer 
 
Despite its non-specific appearance as an asymptomatic red or purple rapidly growing 
nodule, MCC can often behave aggressively, spreading to local, nodal, and distant sites 
(Figure 1).  
 
 
Figure 1. Clinical appearance and progression of MCC 
Extensive nodal disease developed in this woman, months after a primary lesion on the 
eyelid. Within 1 year, she developed metastatic disease that became refractory to 
treatment. 
 
Even MCC tumors smaller than 1 cm at presentation carry a significant incidence of 
nodal and distant metastasis; likely accounting for the largely unsuccessful attempts at 
long-term control with surgery alone.5 Although radiation therapy can effectively reduce 
the size of MCC tumors, this treatment modality does not offer long-term disease 
control.6 Previous studies with concomitant or adjuvant chemotherapy with carboplatin or 
etoposide have also not shown increases in overall survival.7  
 
There is currently no Food and Drug Administration (FDA) approved treatment for non-
resectable, recurrent, advanced, or metastatic MCC. The National Comprehensive 
Cancer Network (NCCN) guidelines recommend that patients with metastatic disease 
consult with the multidisciplinary tumor board and consider radiation, surgery, 
chemotherapy, or a combination thereof.8 The specific treatment regimen for patients 
with distant metastasis must be individually tailored. Most NCCN institutions use only 
chemotherapy with or without surgery and/or radiation therapy for Stage IV, distant 
metastatic disease (M1). The most common regimen used for regional disease is 
cisplatin or carboplatin with or without etoposide.  
 
The NCCN panel recommends cisplatin or carboplatin with or without etoposide as the 
choice of treatment.9 Topotecan has also been used in some instances (for example, 
older patients). Cyclophosphamide in combination with doxorubicin and vincristine used 
to be a commonly administered regimen, but it is associated with significant toxicity.7,10,11 
Overall, although MCC is often initially chemosensitive, tumor responses are rarely 


FHCRC 9245 
 
 
 
 
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durable, with the 2-year survival rate for patients with stage IV disease at approximately 
26%.12 Furthermore, MCC reported incidence has quadrupled in the past 20 years to 
approximately 1,600 cases per year in the United States (US).13 MCC risk factors include 
UV exposure, age over 50, and long-term T-cell immune suppression.  
 
After recognition that MCC incidence was increased 11 fold among patients with AIDS,2 
newly developed molecular and bioinformatics methods led to the discovery of a novel 
polyomavirus, the MCPyV, expressed in approximately 80% of instances of MCC. 
MCPyV DNA shows random clonal integration into the cellular genome of MCC and is 
only expressed in tumor cells, as predicted for a direct viral carcinogen.4,14,15 MCPyV 
encodes a multiply spliced TAg protein complex that targets several suppressor proteins, 
such as RB1, similarly to the SV40 and murine polyomavirus family.15 Knockdown 
studies have shown that sustained TAg expression is required for the survival of 
virus-positive Merkel cell lines.4,16,17 
 
Skin carriage of the wild-type (WT) virus has been found to be common in adults.18 
However, the WT virus must undergo at least two mutations to promote tumorigenesis. 
The first mutation is non-homologous recombination in the host cell genome4, and all 
MCPyV genomes that have been obtained from MCC tumors have inactivating 
secondary mutations, such as inactivation or excision of its Helicase domain that is 
required to unwind the viral origin in order to allow viral DNA replication.19 If not for this 
second deletion, viral replication would proceed uncontrollably in the tumor cell from the 
integration site, deleting the developing tumor cells via genomic fidelity sensors and 
apoptosis. Thus integrated, MCPyV has no mechanism to excise its genome, is no 
longer transmissible (analogous to HPV in most cervical carcinomas), and has no 
DNA-replication capacity, thus rendering the virus resistant to putative antiviral 
agents.16,20 
3.B.  Spontaneous Immune Responses to MCPyV TAg 
 
As previously mentioned, epidemiologic data suggest a strong link of MCC to the 
immune system. Individuals with T-cell dysfunction (transplant recipients, HIV, or chronic 
leukemia patients) are at 5- to 50-fold increased risk of developing MCC.2,3 MCC tumors 
have been demonstrated to spontaneously regress following improvement in immune 
function suggesting an immune-mediated cancer control.21,22 Furthermore, intratumoral 
infiltration of CD8 lymphocytes has been documented as an independent predictor of 
improved survival for MCC patients, suggesting that improving T-cell function may be 
effective in treating MCC.23 As the TAg oncoproteins are persistently expressed in MCC 
tumors and are required for tumor growth, they may serve as ideal targets for 
immunotherapy. 
 
Similar to the relationship between EBV and Burkitt’s lymphoma, exposure to MCPyV is 
common in the general population (53%) as indicated by antibodies to virus-specific 
capsid protein VP1.24,25 In contrast, antibodies to the MCPyV TAg are rare in the general 
population ( less than 1%) but are commonly present (approximately 50%) at high titers 
in patients with active MCC.26,27 Moreover, antibodies to TAg oncoproteins dynamically 


FHCRC 9245 
 
 
 
 
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reflect disease burden and have already been used to identify MCC recurrences prior to 
clinical detection.27,28 Responses directed to MCPyV TAg, unlike antibodies directed to 
the capsid, are non-neutralizing and proportionate to tumor burden. Thus, the 
effectiveness of the treatment can be monitored by antibody titers in serial blood draws. 
3.C.  Identification and Characterization of MCPyV-specific CD8+ T Cells from a 
MCC Patient’s Tumor and Peripheral Blood 
 
To determine whether the integrated MCPyV TAg peptides presented by MHC could be 
recognized by virus-specific T cells in MCC, primary tumor from a patient (w447) with 
MCC was stained with anti-MCPyV TAg antibodies (CM2B4 antibody, Santa-Cruz 
Biotechnology17) by immunohistochemistry to confirm persistent T-Ag expression and 
assess CD8+ T-cell infiltration of the tumor tissue (Figure 2). 
 
 
 Note: Scale bar = 200 μm. 
 
Figure 2. Persistence of MCPyV TAg expression in MCC 
Histologic examination of serial sections from w447 MCC tumor. Left: Hematoxylin and 
eosin stain. Center: MCPyV TAg protein is expressed in the tumor. Right: CD8 
lymphocytes are detected in the tumor.  
 
A synthetic-peptide library spanning the MCPyV TAg was generated, and a total of 
93 peptides -- 13 amino acids in length and overlapping by 9 amino acids -- were then 
tested against tumor-infiltrating lymphocytes (TIL). Results demonstrated TIL secreted 
Interferon-gamma (INFγ) when exposed to the specific 10-mer peptide EWWRSGGFS 
(LT-92-101) (Figure 3). Sequencing of DNA amplified from this patient's tumor confirmed 
the predicted MCPyV large-TAg amino-acid sequence (amino acids 92-101, data not 
shown). 
 


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Figure 3. Reactivity to peptide 
IFNγ production by TIL exposed to MCPyV TAg peptide LT-92-101 (left panel) but not to 
DMSO (right panel) or any other of the 93 screened peptides. 
 
The patient’s HLA type was determined and the peptide was further identified as 
restricted to the HLA-A*2402 allele, because only HLA-A*2402 and TAg co-transfected 
Cos7 cells elicited IFNγ secretion in the TIL (Figure 4A) and target-cell lysis (Figure 4B). 
Specificity and HLA-restriction were confirmed with the synthesis of an MHC class I and 
peptide tetramer. T cells from both the tumor and peripheral blood successfully bound to 
the specific tetramer (Figure 5). These antigen-specific T cells were expanded to 
sufficient numbers to be adoptively transferred back into the patient (described in 
Section 3.H - Previous Human Experience).29 
 
 
 
 
Figure 4. Reactivity to HLA-A*2402 and T-Ag transfected Cos7 cell lines 
A) TIL isolated from this patient induce IFNγ after exposure to Cos7 cells that were transfected 
with HLA class-I A*2402 and T-Ag (right panel) but not with mock transfected Cos7 cells (left). 
B) w447 TIL clone was used as the effector cell against the specified target cells at the 
indicated Effector:Target cell ratios. Fractional cytotoxicity of Cos7 cells is presented as the 
percentage of all Cos7 cells, and does not take into account the decreased number of cells 
expected to be successfully double-transfected (50% or less). 
 
 


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Figure 5. Binding to tetramer 
TIL and T cells isolated from peripheral blood (PBMC) stain specifically with HLA-
A*2402/LT.92-101 tetramer, but not with negative control tetramer. 
 
3.D.  Identification of Reactive MCPyV TAg-specific HLA-restricted Peptides 
 
In collaboration with Sine Hadrup (Herlev Hospital, Denmark), a total of 
14 CD8+-restricted peptide-MHC pairs were identified within the persistently expressed 
LT/ST region of MCPyV (Table 1). All peptide-MHC pairs bind tetramers and were 
isolated from patients with a diagnosis of MCC, whereas no reactivity to these peptides 
was identified in 30 normal donors with identical HLA restrictions. Furthermore, the same 
epitopes were present in multiple MCC patients suggesting that these epitopes are 
uniquely recognized in patients with MCC (unpublished observations).  
 
To assess the fate of the transferred cells in vivo, all patients eligible for this study must 
have an available peptide-MHC pair that can be folded into a tetramer for which 
MCPyV TAg-specific cells can be generated. Additionally, peptide-MHC restricted CD8+ 
T cells must also show reactivity (lysis or INFγ secretion) in response to cell lines 
expressing MCPyV TAg with the corresponding HLA to verify cellular processing of the 
correct peptide. 
 


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Table 1. MCPyV peptide / HLA pairs for which functional tetramers exist 
 
HLA 
Restriction 
HLA 
Frequency 
Observed in 
MCC Patients 
(n=97) 
HLA  
Frequency 
Expected 
(US 
Caucasiansa) 
Viral T-Ag Domain 
LT: Large T 
ST: Small T 
CT: Common T 
Peptide(s) from 
MCPyV 
HLA-A*0201 
55% 
50% 
LT 
LT 
LT 
ST 
ST 
SMFDEVDEAPI 
KLLEIAPNC 
VIMMELNTL 
QLRDSKCACI 
KTLEETDYCLL 
HLA-B*0702 
16% 
25% 
LT 
LT 
LT 
APNCYGNIPL 
APNCYGNIPLM 
APIYGTTKF 
HLA-A*0301 
25% 
23% 
LT 
ASFTSTPPK 
HLA-A*2402 
14% 
15% 
LT 
EWWRSGGFSFb 
HLA-A*1101 
15% 
12% 
LT 
AAFKRSCLK 
HLA-B*3502 
3% 
2% 
CT 
ST 
DKGGNPVIM 
FPWEEYGTL 
HLA-A*2301 
3% 
1.3% 
LT 
EWWRSGGFSF 
aUS Caucasian HLA frequencies were obtained from allelefrequency.net 
bPreviously characterized and infused (see Section 3.H - Previous Human Experience) 
 
3.E.  Adoptive T-Cell Therapy 
 
3.E.1  Rationale as a treatment modality 
 
Adoptive therapy involves the transfer of ex vivo expanded effector cells as a means of 
augmenting the anti-tumor immune response. In contrast to tumor vaccination 
strategies, adoptive T-cell therapy can allow for far greater control over the magnitude 
and avidity of the targeted response by appropriate manipulation and selection in vitro of 
the T cells used for therapy.30,31 Tumor-reactive effector cells of a desired specificity can 
be selected based on expression of their T-cell receptor and expanded to very high 
numbers. Requisite numbers of effectors can be routinely achieved and produce 
frequencies of antigen-specific T cells in the peripheral blood that can be many times 
higher than those produced using current immunization regimens alone.  
 
To date, the majority of clinical trials in adoptive therapy have been performed in 
patients with metastatic melanoma for which T-cell-defined tumor antigens were first 
identified. Such studies designed to address the role of effector cells or conditioning 
regimens in optimizing the anti-tumor efficacy of adoptive cellular therapy in melanoma 
may be equally relevant for the use of adoptive therapy for the treatment of MCC. 
 


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3.E.2  Adoptive therapy of melanoma using antigen-specific CD8+ T-Cell Clones 
 
Historically, the median length of survival for patients with metastatic melanoma 
refractory to standard therapy has been four months. Among an initial cohort of 
10 patients receiving CD8+ T cells targeting the tumor-associated antigens, MART1 and 
MelanA, and gp100 followed by low-dose IL-2 for 14 days at doses up to 
3.3 x 109 cells/m2, no serious toxicity was observed. The adoptively transferred T cells 
persisted in vivo in response to low-dose IL-2, preferentially localized to tumor sites, and 
mediated an antigen-specific immune response characterized by the elimination of 
antigen-positive tumor cells and regression of individual metastases. Eight patients 
experienced minor, mixed, or stable responses for periods of 2 to 21 months (average of 
11 months).32  
 
An updated analysis of 18 patients with refractory progressive disease receiving 
adoptively transferred CD8+ T cells targeting, in addition to Mart1 and gp100, tyrosinase, 
and NYESO-1 at doses up to 1010 cells/m2, demonstrated no serious toxicities and mixed 
responses or stable disease in 10 of 18 patients, a partial response among four patients, 
two of these patients with near complete responses (CR) – (no activity by PET imaging) 
for periods of 3 to more than 33 months. Significant tumor regression and clinical 
responses were reported in 50% of patients receiving infusions of in vitro expanded TIL 
and high-dose IL-2 following non-myeloablative conditioning with cyclophosphamide 
(CY) (60 mg/kg for 2 days) and fludarabine (25 mg/m2 for 5 days).33  
 
In two of 13 patients receiving heterogeneous populations of up to 1010 cells/m2 of CD4+ 
and/or CD8+ T cells of various antigen-specificities (MART-1, gp100, tyrosinase, 
NYESO-1), over 90% of the reconstituted T-cell repertoire was comprised in the infused 
melanoma-specific T cells. T-cell receptor (TCR) analysis of circulating T cells in these 
patients suggested that a transferred clonal T-cell population had expanded, infiltrated 
tumor sites, and induced major clinical responses. However, this approach was 
accompanied by serious, potentially life-threatening toxicities as a consequence of 
profound immunosuppression and high-dose IL-2 therapy.34,35 Thus, the prolonged 
immune suppression resulting from the combination of high-dose cyclophosphamide and 
fludarabine may significantly contribute to serious toxicities.  
 
3.E.3  Adoptive therapy targeting viral antigens 
 
Since 1995, more than 250 patients with EBV-related diseases received donor or 
autologous-derived EBV-specific cells for both prophylaxis and treatment of 
post-transplant lymphoproliferative diseases (PTLD) arising in stem-cell or solid-organ 
transplant recipients.36 The safety of this approach was repetitively demonstrated, as no 
infusion-related serious adverse effects were recorded and the incidence of GVHD in 
allotransplant recipients was not increased after transfer.37 Furthermore, virological as 
well as immunological responses were noted in almost every patient after CTL 
infusion.38,39 Adoptive transfer of CTL specific for CMV after allogeneic HCT were also 
demonstrated to be safe in order to reconstitute deficient viral immunity, and a subset of 
the transferred T cells persisted long-term in vivo.40 Adoptive transfer of HIV-specific CTL 


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targeting various HIV proteins (gag, pol, and nef), as well as CMV-specific CTL targeting 
pp65, were infused into HIV+ patients and demonstrated transient efficacy in patients 
with high viral burdens and persistence in patients with undetectable viremias 
undergoing highly-active antiretroviral therapy.41,42  
 
Thus, virus-specific T cells that are derived and expanded from patients with established 
immunity to the pathogen can be safely transferred into autologous recipients without 
inducing significant side-effects and are capable of persisting, mediating effector 
functions, and controlling infection. The cell doses used in these settings ranged from 
2 x 108 to approximately 3.3 x 109 cells/m2.36 However, because CTL-mediated lysis of 
solid tumors likely requires increased frequencies of adoptively transferred cells to 
infiltrate tumor tissue, this protocol proposes to infuse cell doses of MCPyV-specific CTL 
that have shown safety and efficacy in solid tumors (1010 cells/m2) as well as in one 
patient with metastatic MCC (see Section 3.H – Previous Human Experience).  
 
3.F.  Induction of MHC Expression by Tumor Tissue 
 
Similar to other virus-associated cancers such as Kaposi sarcoma or cervical cancer 
that have been shown to directly down-regulate MHC I on tumor cell’s surface as a 
mechanism of tumor immune evasion,43,44 MHC class I expression on MCC is down-
regulated in more than 50% of cases,45 thus making tumor cells less susceptible to 
antigen-specific T-cell lysis. To reverse down-regulation,  radiation will be administered 
to the tumor or an intralesional stereotactic injection of IFNβ-1B will be administered 
within 24 to 72 hours prior to the CTL infusion to increase MHC expression. The method 
used will be dependent on the localization of metastasis at the time of treatment. Of 
note, MHC class I expression was decreased compared to epidermis on this patient’s 
primary tumor as shown in Figure 6. 
 
 
 
Figure 6. MHC class I downregulation in MCC 
MHC class I expression in epidermis (left), MCC with maintained MHC class I expression 
(middle), and primary tumor from patient W447 (right). MHC class I antibody: EMR8-5. 
 
3.F.1.  Localized radiation 
 
Intracellular peptide products are increased following irradiation due to increased 
degradation of proteins modified directly by radiation or indirectly by the oxidative stress 
exerted by free-radical formation after water radiolysis on amino acids.46 Under normal 
α-MHC class I 
Epidermis 
W447 primary MCC 
MCC - MHC I pos.  
10X 
10X 


FHCRC 9245 
 
 
 
 
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circumstances, TAP is not fully active in the cell and the peptide pool forms the limiting 
factor in the antigen presentation pathway.47 However, under stressful conditions such 
as irradiation, MHC class I expression is up-regulated presumably as a result of the 
increased degradation of intracellular peptides. The effects of radiation on MHC-I 
up-regulation likely play an important role in MCC. In MCC cell lines (such as the 
virus-positive, MHC-I-negative MKL-1 line), a single dose of gamma radiation (4-8 Gy) 
induces up-regulation of MHC-I that persists for at least 7 days (Figure 7).  
 
In mouse tumor models, irradiation of mouse colon adenocarcinoma MC38 cells with 
non-ablative doses of 8 or 10 Gy is not sufficient to inhibit in vivo tumor growth but 
induces MHC class I expression for up to 11 days. Furthermore, experiments in which 
both tumor and adoptively transferred tumor-specific T cells were infused in vivo in mice, 
only the mice which received the irradiated tumor were able to significantly reduce tumor 
growth.48 In a murine B16 melanoma tumor model, the administration of a single dose of 
radiation to established tumors affected tumor regression in a CD8+ T-cell-dependent 
manner; whereas sequential fractionated doses of radiation induced significantly fewer 
antigen-specific tumor responses, strongly suggesting that sequential doses likely 
contributed to ablate the induced T-cell responses.49,50  
 
Although MCC is an exquisitely radiosensitive tumor,51 the early micro-dissemination of 
the disease renders this modality short-lived. In a recent study of 43 MCC patients 
treated by radiation alone, the radiation in-field control rates were 75%, but the overall 
recurrences were 60%.52 We hypothesize that a combined approach involving low-dose 
radiation therapy followed 24 to 72 hours later by infusion of antigen-specific T cells has 
an increased potential to control disease at the irradiated tumor site as well as traffic to 
micro-metastatic sites. Similar to what was observed in mouse models and human MCC 
cell lines, radiation (such as a single dose of 8 Gy) is expected to up-regulate MHC 
sufficiently to allow tumor recognition and lysis by transferred CTL, Although based on 
our clinical experience the irradiation will also likely cause stabilization and/or regression 
of the treated lesion in the majority of cases,53 the primary goal of the radiation therapy is 
MHC upregulation/enhanced antigen presentation and ‘priming’ of a systemic immune 
response. 
  
 
 
Figure 7. MHC class I up-regulation 


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Up-regulation of MHC class I by gamma radiation and IFNγ administration in a MCPyV+ 
MCC cell line, MKL-1. MHC-I up-regulation persists up to 7 days with a single-dose 
radiation administration. 
 
3.F.2.  Intralesional IFNβ-1B injections  
 
This method will be used preferentially over radiation, if the tumor is accessible to 
stereotactic injections as, compared to radiation, a single injection of IFNβ-1B may less 
confound the effects of MCC T-cell-mediated lysis and tumor regression. Interferons 
mediate antiviral immune responses by up-regulation of MHC class I.54 Multiple 
interferons are approved for use in the US for varied indications such as chronic HCV, 
melanoma, and multiple sclerosis.55-57 Intralesional IFNβ-1B injections have previously 
been reported as primary therapy for MCC.58,59 Of nine patients treated at this institution 
with multiple IFNβ-1B injections at cutaneous MCC sites, in whom this was used as a 
unique treatment modality, tumor regression was observed in eight cases (see 
Appendix C).45 In patients in whom MCC tissue could be biopsied before and after 
treatment, MHC class I was significantly up-regulated (Figure 8). Expression of the 
MCPyV TAg by MCC was not modified, and intralesional CD8+ T-cell infiltrates were 
increased after treatment. In contrast, systemic interferon has not been reported to be 
effective in inducing MCC regression and may be due to inadequate delivery into the 
tumor micro-environment.16,60,61  
 
                      Patient #1 
 
 
 
                   Patient #2 
 
Figure 8. IFNβ-1B intralesional injections to induce MHC class I 
MHC class I expression in MCC tissue biopsies before (left) and after (right) IFNβ-1B 
intralesional injections for two individual patients. 
 
3.G.  Previous Human Experience: Adoptive Therapy of MCC Targeting MCPyV 
TAg 
MCPyV TAg represents an exceptionally attractive model of viral tumor antigen for 
evaluating the safety and efficacy of antigen-specific immunotherapy in MCC. MCPyV 
TAg is expressed in approximately 80% of MCC and, compared to mammalian tumor 
associated antigens that all have some degree of expression within normal tissue, 
MCPyV TAg expression is restricted to MCC. As described in Section 3.C, antigen-
specific CD8+ T cells targeting the HLA-A*2402-restricted MCPyV TAg epitope 
EWWRSGGFSF could be generated and infused in patient w447 (Protocol FHCRC 
#2558) and the results are now published.29 As soon as a first metastasis was detected 


FHCRC 9245 
 
 
 
 
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(Figure 9A), the patient underwent a leukapheresis to collect sufficient PBMC to derive 
and expand TAg-specific cells in the FHCRC cGMP facility.  
 
At the time of follow-up scans 140 days later, the patient’s initial metastasis had 
expanded and two additional metastases were detectable. To up-regulate MCH class I 
(see Section 3.F), the patient underwent intralesional IFNβ-IB injections to the largest 
metastasis before receiving a first infusion of 1010 MVPyV TAg-specific CD8+ T cells 
followed by low-dose subcutaneous (SC or s.c.) IL-2 (2.5 x 105 IU/m2 BID x 14 days) 
(Figure 9B). Restaging 28 days later showed a response in the IFNβ-IB infiltrated lesion, 
consistent with T-cell-mediated lysis following MHC class I up-regulation, but growth in 
the non-infiltrated lesions. The patient then went on to receive 8 Gy radiation to all three 
remaining lesions before a second infusion of 1010 MVPyV TAg-specific CD8+ T cells.  
 
Although two of three detected lesions disappeared 140 days after the first infusion, the 
pancreatic neck lesion first decreased in size and then re-increased, suggesting the 
possibility of tumor antigen escape. However, the patient did not develop additional 
metastasis elsewhere 274 days after starting treatment, which suggests a long-term 
control of the transferred T cells on MCC. Overall, the infusions were well tolerated: apart 
from the expected immediate toxicities associated with T-cell infusion (transient, <72 
hours Grade 2 Cytokine Release Syndrome [CRS] and Grade 2 lymphopenia), the 
patient did not develop changes in end organ function or inflammation-related 
complications such as an autoimmune process.  
 
 


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Figure 9. w447 response to MHC class I up-regulation and MCPyV-TAg-specific 
CD8+ T-cell infusions 
A) pre- (left) and post (right)-treatment PET scans. B) Evolution of the individual 
metastasis size in cm2 (y axis) over time (x axis) and corresponding RECIST 1.1 criteria. 
SD: stable disease, PR: partial remission.  
 
Assessment of the frequency of infused CD8+ T cells by their ability to bind tetramers in 
vivo demonstrated the cells peaked from 1 to 3 days after infusions (~8%) before 
persisting at least 140 days at frequencies of 1% to 1.5% or a 10-fold increase compared 
to pre-infusion levels (1.3%) (Figure 10).  
 
 
 
Figure 10. Persistence of MCPyV TAg-specific CD8+ T cells 
Top panels show flow plots of the infusion product and selected timepoints, and bottom 
panel shows the frequency of tetramer+ MCPyV-specific CD8+ T cells (x axis) over time 
(y axis). 
 
Prior to infusions, no MCPyV TAg-reactive cells could be detected within PBMC 
suggesting the detectable tetramer+ cells were non-functional. However, after infusions, 


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the frequency of MCPyV TAg-reactive cells measured by their ability to secrete IFNγ in 
response to cognate peptide increased with each infusion and were detectable at 
frequencies of 1% to 2% at least 140 days after the first infusion. These values 
correlated with the frequency of tetramer+ cells (Figure 11).29  
 
Overall, these results suggest the combination of MHC class I up-regulation with infusion 
of 1010 polyclonal ex vivo-expanded MCPyV TAg-specific cells was safe in this patient, 
established detectable cellular reactivity to MCPyV TAg for at least 140 days, and 
mediated tumor regression in 2 out of 3 detectable lesions without the occurrence of new 
distant metastasis for at least 274 days.  
 
 
 
Figure 11. Establishment of MCPyV TAg-reactive cells in vivo after CD8+ T-cell 
transfer 
Frequencies of IFNγ+CD8+ T cells (right y axis) correlate with frequencies of tetramer+ 
CD8+ T cells (left y axis) over time after transfer.  
 
3.H.  Previous Clinical Experience: Results of Protocols FHCRC #2558 and #2586  
 


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To date, 4 additional patients have been treated in Group 1 of protocol 2586 bringing the 
total number of patients who have received infusions of 1010 cells/m2 MCPyV-specific 
T cells to 5, for a total of 15 infusions, each followed by low-dose s.c. IL-2 for 14 days 
(Table 2).  
 
Table 2. Patient Characteristics 
Pt No 
Previous treatments 
Disease 
Burden (cm2) 
MHC class I 
up-regulation 
Targeted 
epitope 
Infusions 
received 
Pt-1 
#2558 
Surgery, 50Gy 
radiation 
Intermediate 
(20.03) 
IFNβ -1B and 
8Gy radiation 
HLA A*2402-
EWW 
3 
Pt-1 
#2586 
Surgery, IL-12 
electroporation, 
radiation, 
chemotherapy: 
cisplatin and etoposide 
x 5.  
High (51) 
IFNβ -1B 
HLA A*A0201 -
KLL and HLA 
A*2402-EWW 
2 
Pt-2 
#2586 
Surgery, radiation x 2. 
High (54) 
8Gy radiation 
HLA A*A0201 –
KLL 
4 
Pt-3 
#2586 
Surgery x 3, radiation x 
4. 
Low (1.25) 
IFNβ -1B 
HLA A*2402-
EWW 
4 
Pt-4 
#2586 
Surgery x 4, radiation x 
6, chemotherapy x 2, 
intratumoral IFNβ-1B 
High (56) 
8Gy radiation 
HLA B*3502 
2 
 
AEs were evaluated starting from the time of the first infusion to 30 days after the 
patients had taken the last dose of s.c. IL-2. AEs that were deemed possibly, probably or 
likely related were collected and graded according to the National Cancer Institute 
Common Terminology Criteria for Adverse Events version 4.0 (NCI CTCAE v4.0) (Table 
3). 
 
Table 3. Adverse events 
Categories 
NCI CTCAE v4.0a 
Grade 
1 
Grade 
2 
Grade  
3 
Grade 
4 
Cytokine Release 
Syndrome 
Fever 
6 
2 
  
  
Chills 
6 
  
  
  
 
LBBb 
 
1 
 
 
Hematological 
Abnormalities 
White blood cell count decreased 
4 
  
  
  
Lymphopenia 
  
3 
4 
1 
Thrombocytopenia 
1 
2 
  
  
IFNβ-1B-related 
Pain at injection site 
1 
  
  
  


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Wound infection 
  
1 
  
  
IL-2 related 
Injection site reaction 
2 
  
  
  
a National Cancer Institute Common Terminology Criteria for Adverse Events version 4.0. 
b  Left bundle branch block (new) in a patient with pre-existing coronary artery disease. 
 
Expected transient symptoms were observed due to a cytokine release syndrome (CRS) 
associated with activation of large numbers of antigen-specific CTL transferred into 
patients with targets expressing the antigen and/or low-dose s.c. IL-2. Specifically, all 
patients experienced fevers (≥ 38.3°C) with or without chills after at least 1 infusion. 
Blood cultures were all negative for bacterial or fungal growth. All symptoms were 
managed on the general hospital ward and resolved within 24 hours. One patient with 
pre-existing coronary artery disease experienced a transient new left bundle branch 
block while he was febrile. The symptoms resolved and he went on to receive a coronary 
artery stent 10 days later. The hematological abnormality most commonly encountered 
was lymphopenia, which is a predictable, transient side effect of T-cell infusions 
presumably reflecting redistribution of peripheral lymphocytes.62,63 The temporary drop in 
total lymphocyte counts returned to pre-infusion levels within 7 to 11 days in all patients. 
After the treatment, one patient experienced a temporary drop in platelet counts which 
was most likely associated with radiation administration to the pelvis. Of the patients who 
received IFNβ-1B injections, one experienced pain after the injection and another 
developed a localized wound infection which resolved with antibiotic treatment. One 
patient experienced injection-site reactions with low-dose s.c. IL-2 administration.  
 
3.I. Rationale for Combining MHC Class I Upregulation, MCPyV-Specific T-Cell 
Infusions and PD1 - PD-L1 Axis Blockade 
 
Of the patients that could be analyzed beyond 16 weeks (Table 4), one patient showed 
rapidly progressive disease (PD) at 8 weeks and was deceased by 16 weeks. Although 
the remaining 3 patients had either a partial response (PR) or stable disease (SD) at 8 
weeks, only 1 with SD continued on to have a PR at 16 weeks. The other 2 patients had 
progressed at 16 weeks from their maximal response at 8 weeks. Of the 3 patients who 
demonstrated progression after infusions, 2 had detectable transferred cells at the time 
of progression suggesting that the presence of transferred cells was not sufficient to 
suppress tumor growth.   
 
Table 4. Clinical outcomes 
Patient-
Number 
Protocol # 
Clinical Response by mWHO criteriaa 
(weeks after 1st infusion) 
T-cell 
persistence? 
8 weeks 
16 weeks 
Pt-1 #2558 
PR (-79%)b 
Progression of 1 
remaining lesion (-38%) 
Yes 
Pt-1 #2586 
PD (+100%) 
Deceased 
No 


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Pt-2 #2586 
SD (-10%) 
Mixed response (-34%): 
progression of non-
irradiated lesions 
Yes 
Pt-3 #2586 
SD (-13%) 
PR (-58%) 
Yes 
amWHO criteria=modified World Health Organization criteria based on the products of diameters of tumor 
lesions 64; b Note: numbers in parenthesis refer to an increase [positive number] or decrease [negative 
number] in tumor burden as a percentage of baseline values at the indicated timepoint. Definitions: 
Pt=patient, PR=partial response, PD=progressive disease, SD=stable disease. 
 
Analysis of the phenotype of the transferred cells revealed that most products expressed 
programmed death 1 (PD-1) before infusions (Figure 12, top row) and that products that 
expressed PD-1 continued to express PD-1 when they could be detected in vivo (Pt-1 
#2558, Pt-1 #2586) (Figure 12, bottom row). Pt-3 #2586 received a product with low 
PD-1 expression which remained detectable and continued to express low levels of PD-1 
in vivo. This observation coincided with the best clinical outcome as it occurred in the 
only patient whose MCC remained in a PR for 11 months. 
 
 
 
Figure 12. Phenotype of infused products before and after infusions 
Vertical bars indicate cumulative percentages of ecentss in the upper and lower left 
quadrants and represent percentages PD1-expressing cells. 
 
 
Because the ligand for PD-1 (PD-L1), is expressed on 50% of MCC, on 55% of MCC 
tumor infiltrating lymphocytes, and can be induced by local inflammation, this may 
provide a strong inhibitory signal by the tumor microenvironment, shielding the tumor 
from the transferred MCPyV-specific T cells upon PD-1/PD-L1 ligation.65,66 Thus, 
blocking this interaction could reverse the inhibitory signal and allow enhanced 
responses by the transferred T cells that could in turn more efficiently eliminate the MCC.  


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3.I.1.  Programmed death receptor and ligands 
 
The PD-1 receptor as well as PD-L1 and PD-1 ligand 2 (PD-L2) play integral roles in 
immune regulation. Expressed on activated T cells, PD-1 is activated by PD-L1 and 
PD-L2 expressed by stromal cells, tumor cells, or both, initiating T-cell death and 
localized immune suppression,67-70 potentially providing an immune-tolerant environment 
for tumor development and growth. Conversely, inhibition of this interaction can enhance 
local T-cell responses and mediate antitumor activity in nonclinical animal models.67,71 
 
In the clinical setting, treatment with antibodies that block the PD-1 – anti-PD-L1 
interaction have been reported to produce objective response rates of 7% to 38% in 
patients with advanced or metastatic solid tumors, with tolerable safety profiles.72-74 
Notably, responses appeared prolonged, with durations of 1 year or more for the majority 
of patients. 
 
Of importance to the current trial: previous studies have demonstrated PD-L1 tumor-cell 
expression in 49% of MCC patient samples (49 patients sampled) and expression in 
tumor-infiltrating lymphocytes in 55% of patient samples. Of note, of the PD-L1 positive 
samples, 28 of 29 samples showed co-localization with immune infiltrates.66 These 
findings, coupled with the sited response rates and long response durations in other solid 
tumors make PD-L1 an attractive target in patients with MCC. 
 
3.I.2.  The anti-PD-L1 therapeutic antibody Avelumab 
 
The Investigational Medicinal Product (IMP) for the present trial is Avelumab, a fully 
human monoclonal antibody of the immunoglobulin (Ig) G1 isotype. This anti-PD-L1 
therapeutic antibody concept is intended to be developed in oncological settings by 
Merck KGaA, Darmstadt, Germany, and by its subsidiary, EMD Serono Inc, Rockland, 
MA, USA. 
 
Avelumab 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.75 For complete details of the in vitro 
and nonclinical studies, please refer to the Investigator’s Brochure (See Appendix C). 
Avelumab is currently in clinical development with 2 ongoing Phase 1 studies in subjects 
with solid tumors: 
 
Trial EMR100070-001 is “a Phase 1, open-label, multiple-ascending dose trial to 
investigate the safety, tolerability, pharmacokinetics, biological, and clinical activity of 
Avelumab in subjects with metastatic or locally advanced solid tumors.” 
Trial EMR100070-002 is “a Phase 1 trial to investigate the tolerability, safety, 
pharmacokinetics, biological, and clinical activity of Avelumab in Japanese subjects with 


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metastatic or locally advanced solid tumors, with expansion part in Asian subjects with 
gastric cancer.” 
 
3.I.3.  Preliminary results of trial EMR10070-001 
 
This trial consists of two parts, a dose-escalation phase and an expansion phase, which 
is performed in selected tumor indications. 
A maximum of 42 subjects with advanced malignancies with no established therapy 
available were planned to be enrolled in the dose-escalation phase of this trial.  
Avelumab was administered intravenously at the assigned dose level as a 1-hour 
intravenous (IV) infusion once every 2 weeks. Dose escalation (3 + 3 design) was 
performed at the following dose levels:  
• Dose level 1: 1.0 mg/kg (Cohort 1)  
• Dose level 2: 3.0 mg/kg (Cohort 2) 
• Dose level 3: 10.0 mg/kg (Cohort 3) 
• Dose level 4: 20 mg/kg (Cohort 4) 
 
As of 29 August 2013, a total of 13 subjects were enrolled in the dose-escalation phase 
of the trial, with 4, 3, and 6 subjects being treated with Avelumab of 1, 3, and 10 mg/kg, 
respectively. All subjects completed the dose-limiting toxicity (DLT) evaluation period and 
received all planned treatment administrations except for 1 subject in the 1 mg/kg cohort 
(Subject 101-0002 who was replaced). Overall, the data reported under this section are 
based on a total of 51 administrations of Avelumab: 14 administrations of 1 mg/kg, 
20 administrations of 3 mg/kg, and 17 administrations of 10 mg/kg. 
 
Based on data of 13 subjects treated at 1, 3, or 10 mg/kg, the decision to use a dose of 
10 mg/kg for further evaluation in the expansion cohorts was made.  
 
No subject experienced a DLT. The maximum administered dose level of 10 mg/kg at 
this time was thus considered a safe and well-tolerated dose. 
 
Subsequently, the recruitment of Cohort 4 started (3 + 3). In addition, subjects are 
currently being enrolled at 3 mg/kg and 10 mg/kg for the purpose of generating additional 
PK and PharmDyn data. 
 
A total of 7 expansion cohorts are currently being enrolled in the US, EU, and Asia.  
Three cohorts (non-small-cell lung cancer, gastric cancer, metastatic breast cancer) are 
each planned to enroll 150 subjects to generate data that will support the development of 
a companion diagnosis based on immune-staining of PD-L1 that is specific to the 
membrane expression of the molecule. The other 4 cohorts (metastatic colorectal 
cancer, castrate-resistant prostate cancer, advanced melanoma, and ovarian cancer) are 
each planned to enroll 20 subjects. 
 


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3.I.3.A.  Safety results of trial EMR10070-001 
 
All adverse events (AE) 
All 13 subjects (100%) at all dose levels experienced at least 1 treatment-emergent AE 
(TEAE).  The TEAEs at the Preferred Term level reported for the greatest proportion of 
subjects (observed in ≥ 2 subjects) included lymphopenia (9 subjects; 69.2%), anemia 
(8 subjects; 61.5%), fatigue (7 subjects; 58.8%), pyrexia and white blood cell count 
decrease (each in 6 subjects; 46.1%), hyperglycemia (5 subjects; 38.5%), 
hypoalbuminemia and aspartate aminotransferase (AST) increase (each in 4 subjects; 
30.8%), chills and influenza-like illness (each in 3 subjects; 23.1%), and 
thrombocytopenia, blood alkaline phosphatase increase, blood creatinine increase, 
neutrophil count decrease, hypocalcemia, vomiting, and rash (each in 2 subjects; 
15.4%).  There were no relevant differences with regard to the frequency of the AEs 
across different dose levels. 
 
AEs of Grade ≥ 3 
Five subjects (38.5%), 4 receiving the 1 mg/kg dose and 1 receiving the 10 mg/kg dose, 
experienced at least 1 TEAE with a Grade ≥ 3. Of the Grade ≥ 3 TEAEs, anemia was 
reported in 2 subjects (15.4%) and all others, which included the Preferred Terms of 
peritonitis bacterial, staphylococcal infection, pneumonia, lymphopenia, hyperglycemia, 
hypoalbuminemia, hyponatremia, AST increase, blood alkaline phosphatase increase, 
dyspnea, ascites, pain, and gastric outlet (duodenal) obstruction, occurred in 1 subject 
each. No dose dependency for the occurrence of Grade ≥ 3 TEAEs was observed. In 
addition, Subject 101-0002 experienced a Grade 5 event of hypoxia, caused by a 
bacterial pneumonia, which led to the death of this subject.  A drug-induced pneumonitis 
caused by the drug was excluded by the Investigator. 
 
Treatment-related TEAEs and treatment-related Grade ≥ 3 AEs 
Eleven subjects (84.6%) experienced TEAEs assessed as related to Avelumab by the 
Investigator, including 3 of 4 subjects treated with the 1 mg/kg dose, 2 of 3 subjects 
treated with the 3 mg/kg dose, and all 6 subjects treated with the 10 mg/kg dose. 
  
The TEAEs reported for the greatest proportion of subjects and deemed to be related to 
Avelumab by the Investigator were fatigue and lymphopenia (each in 5 subjects; 38.5%), 
pyrexia (4 subjects; 30.8%), influenza-like illness and chills (each in 3 subjects; 23.1%), 
and AST increase (2 subjects; 15.4%).  All other treatment-related TEAEs were reported 
in 1 subject each. 
 
One subject (101-0004) treated with the 1 mg/kg dose experienced Grade ≥ 3 blood 
alkaline phosphatase increase and AST increase, both considered by the Investigator to 
be related to the trial treatment. 
 
Subsequent to the cutoff date of 29 August 2013, 2 Grade 4 infusion reactions have 
been reported (see Section 3.I.3.B) 
 
Serious adverse events (SAE) 


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As of 29 August 2013, 2 of 13 subjects (15.4%) experienced treatment-emergent SAEs 
(Subject 101-0003: gastric outlet obstruction, abdominal ascites, and bacterial peritonitis; 
and Subject 101-0002: hypoxia, shortness of breath, and lung infection bilateral 
pneumonia).  Both subjects were treated with 1 mg/kg of Avelumab.  None of the SAEs 
in either subject were considered related to trial treatment as assessed by the 
Investigator. 
 
As of 29 August 2013, 1 death as a result of a TEAE has been reported:  
• Subject 101-0002 (1 mg/kg of Avelumab):  The subject was reported to have died 
of bilateral pneumonia, with the SAE of hypoxia being considered as the fatal 
event.  The SAEs leading to death, and thus the event of the death, were not 
considered related to trial treatment by the Investigator.  In particular, the 
diagnosis of drug-induced pneumonitis was excluded by the Investigator. 
 
TEAEs leading to permanent discontinuation of Avelumab treatment 
As of 29 August 2013, 3 subjects (23.1%), all receiving 1 mg/kg of Avelumab, 
discontinued trial treatment permanently for TEAEs. These included the 2 subjects (101-
0003 and 101-0002) who experienced SAEs as described above and 1 additional subject 
(101-0004) who discontinued trial treatment for Grade 3 AST increased. 
 
Laboratory abnormalities 
Abnormal laboratory findings were reported as AEs if they were associated with clinical 
signs or symptoms, or led to treatment discontinuation, or were considered otherwise 
medically important by the Investigator.   
 
Hematology parameters were stable, except for transient decreases in white blood cell 
(WBC) count during trial treatment. No clinically significant hematology abnormalities 
were evident and no dose-dependent changes in hematology parameters were 
observed. 
 
Clinically significant increases in AST and alanine aminotransferase (ALT) leading to 
treatment discontinuations as well as increases in alkaline phosphatase were observed 
in 1 subject treated with 1 mg/kg of Avelumab, which were observed following 5 doses of 
trial drug infusion and were transient, with the ALT and AST abnormalities returning to 
normal within a week. An elevated gamma-glutamyl transferase (GGT) was observed in 
1, 2, and 1 subjects treated with 1, 3, and 10mg/kg of Avelumab respectively. In all these 
subjects, decreases in GGT were seen over time. Total bilirubin remained unchanged 
during the treatment. 
 
No clinically relevant changes in amylase, lipase, glucose, albumin, total protein, creatine 
kinase, lactate dehydrogenase (LDH), creatinine, urea, uric acid, electrolytes, 
cholesterol, and triglycerides were observed. Overall, no evidence of a dose dependency 
of Avelumab was observed for changes in blood chemistry variables during the trial. 
No clinically relevant changes in blood coagulation parameters, such as international 
normalization ratio, and activated partial thromboplastin time were identified in all 
subjects. 


FHCRC 9245 
 
 
 
 
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Vital signs, body weight, and electrocardiogram  
During trial treatment, most variations in vital sign parameters, including heart rate, 
systolic and diastolic blood pressures, respiratory rate, and body temperature as well as 
body weight were small and the values remained stable and within the normal range. No 
obvious differences were observed among different dose cohorts. Overall, no clinically 
significant changes in blood pressure or heart rate were observed during trial treatment.  
The electrocardiograms (ECGs) in 5 of 13 subjects were normal at baseline. During trial 
treatment, none of the 5 subjects had abnormal ECGs assessed by the Investigator as 
clinically significant. Eight subjects had abnormal ECG findings at baseline. All these 
abnormal ECG findings were discussed at the Safety Monitoring Committee meetings 
and had no impact on the continuation of trial treatment. 
 
Summary of safety 
Overall, Avelumab was well tolerated up to 10 mg/kg, no DLT has been reported, and no 
safety concerns have been identified. The majority of the observed AEs were consistent 
with the subjects’ underlying cancer disease. 
   
3.I.3.B.  Infusion-Related Reactions and Hypersensitivity 
 
As of 23 January 2014, 7 subjects with infusion-related hypersensitivity have been 
reported in 119 subjects treated with Avelumab in the 2 Phase 1 trials, including 3 SAEs 
(2 infusion-related reactions, 1 anaphylactic reaction). Of the 3 SAEs, 2 were assessed 
by the Investigator as Grade 4 infusion-related reactions; however, review of the cases 
revealed that the described reactions fulfilled the criteria for Grade 2 infusion-related 
reactions. Follow-up with the Investigator is ongoing. The remaining 4 subjects were 
reported as having non-serious infusion-related reactions, including 3 subjects with 
Grade 2 and 1 subject with Grade 3 infusion-related reactions. 
 
3.I.3.C. Pharmacokinetic and Pharmacodynamic Results 
 
At the analysis cut-off date of 29 August 2013, 13 evaluable subjects were included for 
PK analysis. The PK profile of Avelumab at the highest dose level of 10 mg/kg 
(compared with the 1 and 3 mg/kg doses) is characterized by the following parameters:  
• Average maximum concentration observed postdose (Cmax) was 230.0 µg/mL, with 
individual values ranging from 182.6 to 299.4 µg/mL compared with 18.41 µg/mL for 
the 1 mg/kg dose (14.49 to 22.64 µg/mL) and 86.54 µg/mL (76.89 to 101.1 µg/mL) 
for the 3 mg/kg dose.  
• Median time to reach Cmax (tmax) was approximately 1 hour after the start of infusion 
for all doses. 
• At Day 15 before the second dose administration, an average minimum postdose 
(trough) concentration (Cmin) of 17.14 µg/mL was observed for the subjects who 
received a dose of 10 mg/kg with individual values ranging from 11.72 to 
26.87 µg/mL compared with 0.23 µg/mL for the 3 subjects treated at a 1 mg/kg dose 


FHCRC 9245 
 
 
 
 
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(0 to 0.68 µg/mL) and 5.66 µg/mL (4.56 to 6.99 µg/mL) for the 3 subjects treated at a 
3 mg/kg dose.  
• The average area under the concentration-time curve (AUCtau) for the first infusion 
was 23485 µg/mL∙h (16439 to 29316 µg/mL∙h) compared with 1286 µg/mL∙h for the 
1 mg/kg dose (854 to 1669 µg/mL∙h) and 8067 µg/mL∙h (7516 to 8485 µg/mL∙h) for 
the 3 mg/kg dose. 
• Interindividual variability in exposure parameters (Cmax and AUCtau) was low. In the 
10 mg/kg cohort, the coefficient of variation for the geometric mean was 19.1% for 
Cmax and 21.9% for AUCtau.  
• No significant accumulation of drug concentrations was recorded in all dose levels.  
After the first infusion, trough levels did not fall beyond the lower limit of detection, 
but were very low (Cmin). Based on the limited number of PK data available after the 
second infusion, it was concluded that these residual trough values did not increase 
up to the fourth infusion. The peak values also remained at a similar level over time. 
• Half-life (t1/2) was 92.8 hours (77.2 to 117.4 hours) compared with 63.4 hours for the 
1 mg/kg dose (45.9 to 85.6 hours) and 91.3 hours (88.5 to 95.1 hours) for the 
3 mg/kg dose.  
 
From the PK perspective, no unexpected observations or findings that would raise safety 
concerns could be identified. The shift to a longer t1/2 from 1 to 3 mg/kg doses is 
consistent with a progressing target occupancy, which indicated that the fast target-
mediated clearance reached to a saturated stage, while a slow unspecific elimination 
became prominent. Similar half-lives for the 3 and 10 mg/kg doses indicated that the 
target-mediated clearance was nearly saturated by the 3 mg/kg dose. The PK 
parameters determined, support a DLT evaluation period of 21 days, which covered 
approximately 5 half-lives. 
 
These theoretical considerations, extrapolated from the PK behavior were supported by 
an initial set of target occupancy (TO) data. The TO, that is, the binding to the PD-L1 
molecules, was analyzed on circulating CD3+ lymphocytes. The investigation was 
performed by flow cytometry on blood samples collected on Day 1 before the start of the 
infusion, at 4 and 48 hours (Day 3) after the start of infusion, and before the start of each 
infusion on Days 15, 29, 43, and 85. 
 
At the Avelumab doses of 3 and 10 mg/kg, the TO prior to the second infusion on Day 15 
was greater than 90% for all subjects, while the Avelumab trough serum levels ranged 
from 4.56 to 6.99 µg/mL and from 19.42 to 26.87 µg/mL, respectively, at this time point.  
At the Avelumab dose of 1 mg/kg, 2 of 3 subjects displayed a TO of less than 90% at 
trough serum concentrations on Day 15, while the Avelumab serum levels were below 
the lower limit of quantification of 0.2 µg/mL in these 2 subjects.  
Before the initiation of the Phase 1 trial, a target trough concentration of 58.5 μg/mL was 
calculated (Avelumab Investigator Brochure Version 1.0) on the basis of TO data for 
Avelumab in mouse blood, fitted by a receptor binding model, and considering that 95% 
receptor occupancy on the peripheral blood mononuclear cell (PBMC) correlated with 


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tumor growth inhibition in a murine disease model. Subsequently, additional target 
occupancy data for primate blood cells showed that PD-L1 density was much lower on 
primate CD3+ lymphocytes than mouse PBMCs.  
Target occupancy was investigated on PBMC by flow cytometry-based assay, previously 
validated using PBMC from healthy volunteers. Validation data showed that near 
complete TO was achieved at anti-PD-L1 concentrations greater than 0.2 µg/mL. These 
new data explain why a near-complete TO on peripheral CD3+ was achieved at lower 
Avelumab serum levels than predicted. 
  
Though a TO of > 90% was observed for trough concentrations at the 3 mg/kg 
dose, the 10 mg/kg dose was selected as the clinical dose for the expansion 
cohorts because 
• it was considered conservative to compensate for possible differences in receptor 
density between CD3+ cells present in peripheral blood and tumor cells or 
immune-infiltrating cells that are present in the tumor micro-environment;  
• to compensate for differences in Avelumab serum concentration and tissue (tumor) 
concentrations; 
• there was no difference in terms of safety for any of the 3 doses, which was consistent 
with the findings reported for another anti-PD-L1 monoclonal antibody (0.3 to 
10 mg/kg), in which the spectrum, frequency, and severity of treatment-related TEAEs 
were similar among the dose levels, with the exception of infusion reactions (29); 
• the finding that the t1/2 for the 3 and 10 mg/kg doses were nearly identical is consistent 
with the view that the target-mediated clearance was saturated and that higher doses 
were not expected to result in a higher percentage of TO. 
 
3.J. Rationale for Assessing T-Cell Effectiveness by ‘Time to New Metastasis’ 
 
It cannot be excluded that up-regulation of MHC class I with either radiation therapy or 
intralesional IFNβ-1B injections may have an independent effect on the targeted 
metastasis, which could then be measurable by Response Evaluation Criteria In Solid 
Tumors (RECIST) criteria. Furthermore, as transferred T cells circulate systemically, 
their effect may be better assessed by the prevention of the establishment of 
micrometastasis and the occurrence of new detectable metastatic lesions. To capture 
the systemic effects of adoptively transferred T cells, the time for new metastasis to 
develop after treatment will be assessed, and this parameter will be compared to 
historical matched controls undergoing standard therapy by radiation or chemotherapy.  
 
 


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4.  OBJECTIVES 
 
4.A.  Primary Objectives 
 
1. Assess and compare the safety and potential toxicities associated with treating 
patients with metastatic MCC with either MHC up-regulation and PD1-axis blockade 
(Group 1), or MHC up-regulation, PD1-axis blockade and adoptive transfer of MCPyV 
TAg-specific polyclonal autologous CD8+ T cells (Group 2).  
 
2. Assess and compare the antitumor efficacy associated with treating patients with 
metastatic MCC with either MHC up-regulation and PD1-axis blockade (Group 1), or 
MHC up-regulation, PD1-axis blockade and adoptive transfer of MCPyV TAg-specific 
polyclonal autologous CD8+ T cells (Group 2). 
4.B.  Secondary Objectives 
 
1. Examine the in vivo persistence and, where evaluable, migration to tumor sites of 
adoptively transferred polyclonal CD8+ T cells targeting the MCPyV TAg (Group 2). 
 
2. Examine the in vivo functional capacity of adoptively transferred polyclonal 
CD8+ T cells targeting the MCPyV Tag (Group 2). 
 
3. Examine and compare evidence of epitope spreading with either MHC up-regulation 
and adoptive transfer of MHC up-regulation and PD1-axis blockade (Group 1), or 
MHC up-regulation, PD1-axis blockade and adoptive transfer of MCPyV TAg-specific 
polyclonal autologous CD8+ T cells (Group 2). 
 
 
5.  STUDY ENDPOINTS 
 
5.A.  Primary Endpoints 
 
1. Evidence and nature of toxicity related to the treatment.  
 
2. Evidence of response based on “median time to new metastasis” (See 
Section 12.D.1).  
 
5.B.  Secondary Endpoints 
 
1. Persistence of transferred T cells in blood and in tumor (Group 2). 
 
2. Functional capacity of transferred T cells (Group 2) (Please see Section 12.E.3). 
 
3. Evidence of epitope spreading (See Section 12.F). 
 


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4. Disease response by RECIST criteria, and MCC-specific survival (See 
Section 12.D.2). 
 
 
6.  STUDY DESIGN 
 
The proposed study is a Phase I/II trial aimed at treating 20 individuals with metastatic 
MCC.  
Group 1: Due to the availability of the anti-PD-L1 antibody, ten patients with metastatic 
MCC who do not have an HLA type for which T cells can be generated (for example, 
who DO NOT express HLA A*0201, HLA A*2402 or HLA B*3502) or for whom T cells 
cannot be generated for technical issues, will be treated on Group 1 ( 
Figure 13). They will be planned to receive the anti-PD-L1 antibody as an infusion every 
2 weeks for 12 months and an MHC class I up-regulation intervention (either localized 
radiation or an intra-tumor injection of 3.3 x 106 IU of IFNβ-1B) between 7 and 10 days 
after the preceding infusion of anti-PD-L1 (or 7 to 4 days before the next anti-PD-L1). 
Patients may receive preferably one, but up to 3 infusions of anti-PD-L1 before the first 
MHC class I up-regulation intervention. Depending on the continued presence of 
detectable metastatic disease and if they remain clinically eligible, patients could 
undergo MHC class I up-regulation during the time anti-PD-L1 infusions were ongoing 
starting within four weeks of the preceding restaging scans. 
 
Group 2: Ten patients with metastatic MCC who have an HLA type for which T cells can 
be generated (for example, who express HLA A*0201, HLA A*2402 and/or HLA B*3502) 
will be treated on Group 2. They will be planned to receive an infusion of anti-PD-L1 
antibody every 2 weeks for 12 months, an MHC class I up-regulation intervention (either 
localized radiation or an intra-tumor injection of 3.3 x 106 IU of IFNβ-1B) between 7 and 
10 days after the first infusion of anti-PD-L1 and 2 to 5 days prior to receiving a first of 2 
infusions (approximately 28 days apart) of no greater than 1010 cells/m2 of polyclonal 
autologous MCPyV TAg-specific CD8+ T cells. For example, a patient with a surface 
area of 2 m2 would receive no greater than 2 x 1010 total polyclonal autologous MCPyV 
TAg-specific CD8+ T cells. Restaging studies were performed 8 weeks after the first 
interventions to induce local MHC Class I upregulation. Similar for patients in Group 1, 
they may receive preferably one, but up to 3 infusions of anti-PD-L1 before the first MHC 
class I up-regulation intervention.  A second attempt will be made to generate MCPyV 
TAg-specific CD8+ T cells if the initial cell production was unsuccessful and the final cell 
product will be infused as soon as it becomes available.   
 
If patient has disease progression while on Group 1 alone, treatment may be 
stopped/interrupted to allow for disease control with other anti-cancer therapies. If T cells 
are generated and made available in the interim, the patient may resume therapy on 
Group 2 as long as eligibility criteria are met. 
For safety considerations, low-dose SC IL-2 will not be administered after the T-cell 
infusions due to concurrent anti-PD-L1 treatment. Restaging studies will be performed 8 
weeks after interventions to induce local MHC Class I upregulation by tumor tissue. 
Depending on the continued presence of detectable metastatic disease and if the patient 


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remains clinically eligible, patients could undergo the same treatment (MHC class I up-
regulation, two CTL infusions approximately 28 days apart) during the time anti-PD-L1 
infusions were ongoing starting within four weeks of the preceding restaging scans. 
 
 
 
 
Figure 13. Study decision tree (Groups 1 and 2) 


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7.  ELIGIBILITY FOR TREATMENT 
 
7.A.  Inclusion Criteria 
 
1. Signed written informed consent. 
 
2. Confirmation of MCC by internal pathology review of initial or subsequent biopsy 
or other pathologic material.  
 
3. If an accessible lesion is present, a biopsy will be performed within 6 weeks of the 
start of study intervention. The results of the biopsy must be obtained prior to 
initiation of study intervention.  
 
4. Evidence of MCPyV TAg tumor expression by immunohistochemistry on any prior 
or current tumor specimen or viral oncoprotein antibody confirmation within 6 
weeks of the start of study intervention. 
 
5. ECOG PS ≤ 2 at trial entry. 
 
6. 18 years of age or older.  
 
7. Patients must have at least one bi-dimensionally measurable lesion by palpation, 
clinical exam, or radiographic imaging within 6 weeks of the start of study 
intervention (X-ray, CT scan, PET scan, MRI, or Ultrasound). 
 
8. For patients designated to be treated on Group 2: cardiac ejection fraction ≥35%. 
For patients with significant risk factors for coronary artery disease (Framingham 
risk score > 15%), a cardiac stress test is recommended.  
 
9. At least 3 weeks must have passed since any of the following: systemic 
corticosteroids, immunotherapy (for example, T-cell infusions, immunomodulatory 
agents, interleukins, MCC vaccines, intravenous immunoglobulin, expanded 
polyclonal TIL or LAK therapy), pentoxifylline, other small molecule or 
chemotherapy cancer treatment, other investigational agents or other systemic 
agents that target Merkel cell carcinoma.  
 
 
7.B.  Exclusion Criteria 
 
1. Known active infections or oral temperature >38.2 C fewer than 72 hours prior to 
receiving study treatment or systemic infection requiring chronic maintenance or 
suppressive therapy.  
 
2. CBC profile prior to trial entry: 


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WBC < 2000/mcl 
Hb < 8 g/dL  
ANC < 1000/mcl 
Platelets < 50,000/mcl 
 
3. New York Heart Association functional class III-IV heart failure, symptomatic 
pericardial effusion, stable or unstable angina, symptoms of coronary artery 
disease, congestive heart failure, clinically significant hypotension, or history of an 
ejection fraction of ≤ 30 % (echocardiogram or MUGA). 
 
4. Clinically significant pulmonary dysfunction, as determined by medical history and 
physical exam. Patients so identified will undergo pulmonary functions testing and 
those with FEV1 < 2.0 L or DLco (corr for Hgb) < 50% will be excluded. 
 
5. Creatinine clearance < 30 ml/min which cannot be attributed to MCC metastasis.  
 
6. Total bilirubin > 1.5 x ULN, AST/ALT > 2.5 x ULN.  For patients with liver 
metastases, AST/ALT > 5x ULN 
 
7. Active autoimmune disease (e.g. systemic lupus erythematosus, vasculitis, 
infiltrating lung disease, inflammatory bowel disease) whose possible progression 
during treatment would be considered unacceptable by the investigators. 
 
8. Symptomatic and untreated central nervous system (CNS) metastasis. However, 
patients with 1 to 2 asymptomatic, less than 1 cm brain/CNS metastases without 
significant edema may be considered for treatment. If sub-centimeter CNS lesions 
are noted at study entry, then repeat imaging will be performed, if more than 4 
weeks have elapsed from the last scan. 
 
9. Any condition or organ toxicity that is deemed by the PI or the attending physician 
to place the patient at unacceptable risk for treatment on the protocol. 
 
10.  Pregnant women, nursing mothers, men or women of reproductive ability who are 
unwilling to use effective contraception or abstinence. Women of childbearing 
potential must have a negative pregnancy test within 2 - 6 weeks prior to 
treatment. 
 
11.  Clinically significant and ongoing immune suppression including, but not limited 
to, systemic immunosuppressive agents such as cyclosporine or corticosteroids, 
CLL, uncontrolled HIV infection, or solid organ transplantation. 
  
12.  Patients may not be on any other treatments for their cancer aside from those 
included in the protocol. Patients may not undergo another form of treatment 
concurrently with this study. 
 


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13.  Known severe hypersensitivity reactions to monoclonal antibodies (Grade ≥ 3 
NCI-CTCAE v 4.0), any history of anaphylaxis, or uncontrolled asthma. 
 
14.  Vaccination with live inactivated viral strains for the prevention of infectious 
diseases within 4 weeks of the start of the study treatment. Inactivated influenza 
vaccines are permitted while on trial.  
 
15.  Known alcohol or drug abuse. 
 
16.  Legal incapacity or limited legal capacity. 
8.  CONSENTING 
 
Prior to consenting to this study, patients will have been HLA typed. If they express an 
HLA type for which T cells can be generated (HLA A*0201, HLA A*2402 or HLA B*3502), 
they will have undergone a leukapheresis under protocols 2365 (see Section 10.A.1). 
Thus: at the time of the consent, they will already be designated for Group 1 or 2.  An 
enrollment conference will be held with the patient. The PI or a delegated representative 
will discuss this study and alternative treatments available for metastatic MCC. All known 
risks and potential hazards of treatment with IFNβ-IB, radiation and PD-L1 and MCPyV-
specific CD8+ T polyclonal cells (Group 2) will be discussed. Informed consent will be 
obtained from the patient using forms approved by the Institutional Review Board (IRB) 
of the FHCRC. The patient will also be informed that they may withdraw from the study 
at any time and for any reason without jeopardizing their future treatment. Patients who 
are designated for treatment on Group 2 will also be informed that they will receive 
alternative treatment on Group 1 in the unlikely case that MCPyV-specific T-cells cannot 
be generated for them. In accordance with federal regulations (21 CFR 312), the patient 
must sign the IRB-approved consent form in the presence of a witness.   
 
 
9.  PROTOCOL REGISTRATION 
 
Patients will be assigned to the protocol by the Clinical Coordinator who will register the 
patient with the Registration Office, (206) 667-4728, between 8:30 am and 4:00 pm, 
Monday through Friday. After hours, the Registration Office can be reached by paging 
(206) 995-7437.  
 
 
 
 
10. STUDY AGENTS 
 
10.A.  MCPyV-specific CD8+ Cells 
 
10.A.1.  Procedure to obtain PBMC for generation of MCPyV-specific CD8+ cells 
 
Patients will have undergone leukapheresis under FHCRC IRB-approved Protocol 
2365 to obtain PBMC for the generation of antigen-specific T cells in the lab. PBMC 


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may be cryopreserved under therapy conditions and used to generate T cells for this 
study. 
 
10.A.2.  Generation of MCPyV-specific CD8+ cells 
 
All T cells administered will be autologous antigen-specific T cells targeting MCPyV TAg 
and derived from the peripheral blood lymphocytes of a patient with an established 
diagnosis of MCC. Methods employed to generate and qualify product for infusions are 
outlined in IND 14279 (Sponsor Dr. Aude Chapuis). T cells demonstrating antigen 
specificity are further qualified using mycoplasma, fungal, and bacterial sterility testing. 
 
10.A.3.  Handling of T-cell products before infusion 
 
For each infusion, the cell product is formulated at the desired cell dose in a final volume 
of 500 mL. The final product will be prepared and labeled according to standard of 
operation procedure in the Cellular Processing Facility. The cell product will be 
transported to the infusion facility by a protocol 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 to the patient over one to two hours (as described in Section 
11.4). 
 
10.B.  Anti PD-L1 Antibody Avelumab 
 
10.B.1.  Packaging and labeling 
 
Avelumab is a sterile, clear, and colorless solution intended for IV administration 
supplied by the Sponsor in single-use glass vials at a concentration of 20 mg/mL closed 
with a rubber stopper and sealed with an aluminum yellow polypropylene flip off seal. 
Packaging and labeling will be in accordance with applicable local regulatory 
requirements and applicable Good Manufacturing Practice (GMP) guidelines. Avelumab 
will be packed in boxes containing a suitable number of vials. The information on the IMP 
will be in accordance with approved submission documents. Avelumab will be shipped in 
transport cool containers (2°C to 8°C) that are monitored with temperature control 
devices. 
 
10.B.2.  Storage, handling, and preparation 
 
The contents of the Avelumab vials are sterile and nonpyrogenic, and do not contain 
bacteriostatic preservatives. Any spills that occur should be cleaned up using the 
facility’s standard cleanup procedures for biologic products. 
 
Avelumab drug product must be stored at 2°C to 8°C until use, with a temperature log 
maintained daily.  All medication boxes supplied to each trial site must be stored 
carefully, safely, and separately from other drugs. 
 


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Avelumab drug product stored at room temperature (23°C to 27°C) or at elevated 
temperatures (38°C to 42°C) for extended periods is subject to degradation.   
 
Avelumab must not be frozen. Rough shaking of Avelumab must be avoided. 
For application in this trial, Avelumab drug product must be diluted with 0.9% saline 
solution (sodium chloride injection) supplied in an infusion bag. Detailed information on 
infusion bags and medical devices to be used for the preparation of the dilutions and 
subsequent administration will be provided in the MOP. 
 
Avelumab must not be used for any purpose other than the trial. The administration of 
IMPs to subjects who have not been enrolled into the trial is not covered by the trial 
insurance. 
 
Any unused portion of the solution should be discarded in biohazard waste disposal with 
final disposal by accepted local and national standards of incineration. 
 
10.B.3.  Avelumab product accountability 
 
The Investigator is responsible for ensuring accountability for IMP, including 
reconciliation of drugs and maintenance of drug records. 
 
• Upon receipt of IMP, the Investigator (or designee) will check for accurate delivery 
and acknowledge receipt by signing (or initialing) and dating the documentation 
provided by the Sponsor and returning it to the Sponsor. A copy will be retained for 
the Investigator File. 
• The dispensing of the IMP will be carefully recorded on the appropriate drug 
accountability forms provided by the Sponsor and an accurate accounting will be 
available for verification by the Sponsor’s Monitor at each monitoring visit. 
• IMP accountability records will include: 
 confirmation of IMP delivery to the trial site; 
 the inventory at the site of IMP provided by the Sponsor and prepared at the 
site; 
 the use of each dose by each subject; 
 the return to the Sponsor or alternative disposition of unused IMP; and 
 dates, quantities, batch numbers, expiry dates and (for IMP prepared at the site) 
formulation, as well as the subjects’ trial numbers. 
• The Investigator should maintain records that adequately document 
 that the subjects were provided the doses specified by the clinical trial 
protocol/amendment(s); and 
 That all IMP provided by the Sponsor was fully reconciled. 


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Unused IMP must not be discarded or used for any purpose other than the present trial.  
Any IMP that has been dispensed to a subject must not be redispensed to a different 
subject. 
 
The Sponsor’s Monitor will periodically collect the IMP accountability forms and will 
check all returns (both unused and used containers) before arranging for their return to 
the Sponsor or authorizing their destruction by the trial site. 
 
At the conclusion or termination of this trial, trial site personnel and the Clinical Trial 
Monitor (CTM) will conduct a final product supply inventory on the Investigational Drug 
Accountability Forms and all unused containers will be destroyed. Instructions for 
destruction of product will be provided to the site. The CTM will be supplied with a copy 
for filing of the Investigational Drug Accountability Forms. This documentation must 
contain a record of clinical supplies used, unused, and destroyed and shall include 
information on 
• all administered units, 
• all unused units, 
• all destroyed units (during the trial), 
• all destroyed units at the end of the trial, 
• date of destruction(s), 
• name and signature of the Investigator/pharmacist. 
 
It must be ensured at each trial site that the IMP is not used 
• after the expiry date, and 
• after the retest date unless the IMP is reanalyzed and its retest date extended. 
 
This is to be closely monitored by the trial monitor. 
 
10.C.  Radiation therapy 
 
The goal of the radiation is not to kill tumor cells, although some lesional 
stabilization/shrinkage is expected in many cases, but to induce the tumor cells to up-
regulate previously down-regulated MHC Class I to prime a systemic immune response 
mediated by the transferred CTL. A suggested regimen based on a large case series as 
well as preclinical data supporting MHC upregulation effect is 8 Gy dose given in single 
fraction,53 however we recognize that there are clinical scenarios in which alternate 
dosages and/or fractionated regimens may be more appropriate, and in this scenario the 
treating radiation oncologist may select an alternate regimen for the patient if felt that it is 
in the best interest of patient safety. Given that radiation is being given for MHC Class I 
upregulation/antigen presentation, the GCT (gross tumor volume) and the CTV (clinical 
tumor volume) are the same. For the planning tumor volume (PTV) which accounts for 


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day-to-day set-up variations, it is recommended a 5-10mm margin be added to the GTV 
(subject to individualization by the treating Radiation Oncologist). CT-based 3D-
conformal planning (e.g. Intensity Modulated Radiation Therapy [IMRT]) will be utilized to 
minimize dose to the surrounding critical structures. Image Guided Radiotherapy (IGRT) 
will be utilized to verify the target location at treatment with the planning CT scan. Choice 
of radiation modality will be at discretion of radiation oncologist. It is recommended that 
the field size be kept under 10 x 10 x 10 cm. Re-irradiation is allowed although it is 
recommended to limit re-irradiation to the spinal cord, neural structures and small bowel 
to no more than 2x single fraction 8Gy radiation unless no appropriate alternatives. 
Radiation will be completed within 2 to 5 days prior to a T-cell infusion and 7 to 4 days 
prior to the infusion of the next anti-PD-L1 infusion.  
If MHC class I up-regulation is not clinically indicated, (for example tumor site reduction 
without clearance and/or no discrete tumor metastasis that can be irradiated or infiltrated 
with intralesional IFNβ-1B) the second set of T cell infusions can proceed without MHC 
class I up-regulation.  
10.D.  Intralesional IFNβ-1B Injection 
 
IFNβ-1B (Betaseron, Bayer, Montville NJ; or Extavia, Novartis, East Hanover, NJ) will be 
used. One vial contains 0.3 mg or approximately 10 x 106 IU. The lyophilized contents 
will be suspended in the provided 1.2 ml sodium chloride 0.54%. 1/3 of the vial (0.1mg or 
approximately 3.3 x 106 UI or 0.1mg) is administered intralesional into an identified lesion 
after US-guided tumor core biopsy. IFNβ-1B administration will be completed 2 to 5 days 
prior to a T-cell infusion and 7 to 4 days prior to the infusion of the next anti-PD-L1 
infusion.  
 
10.E. Guidelines for selecting radiation therapy or intralesional IFNβ-1B Injections 
 
The decision to proceed to radiation of the lesions or administer intralesional IFNβ-1B 
will be based on clinical judgment with the following guidelines: 1. If more than one tumor 
lesions are present at different locations and no critical organ is involved, if feasible, 
approximately 50% of the lesions should receive radiation or intralesional IFNβ-1B 
initially; 2. At a given time, one patient should receive either radiation or intralesional 
IFNβ-1B but not both modalities; 3. Intralesional IFNβ-1B is the preferred modality for 
patients who have easily palpable superficial lesions (cutaneous, subcutaneous, or 
lymph nodal metastasis) that can be accurately localized, stabilized by palpation, and are 
superficial enough to enable intratumoral injections, or patients who have lesions that are 
attainable through endoscopy; 4. Radiation to a specific lesion or to multiple lesions 
within a specific field will be administered in the case that multiple radiographically 
detectable lesions are identified on the pre-treatment scans, lesions are inaccessible by 
interventional radiology, or the patient refuses intralesional injections. 5. If multiple 
lesions are present within a tight field, it is recommended not to irradiate some and not 
others, but rather deliver radiation to the whole field.  
 


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11.  PLAN OF TREATMENT 
 
 
 
Figure 14. Treatment schema 
 
 
1. Initial Evaluation/Consent (all Groups) 
Patients will be screened for their eligibility on the study and consented during this 
visit. Baseline anthropometry will be obtained.  
 
2. Administration of Avelumab (Groups 1 and 2) 
Subjects will receive an IV infusion of Avelumab within 4 weeks of the baseline 
staging scans at a dose of 10 mg/kg over the duration of 1 hour (-10/+20 minutes) 
following pretreatment with H1 blockers (diphenhydramine 12.5mg to 50 mg PO 
or IV, or equivalent), and acetaminophen po 500 to 650 mg, once every 2 weeks 
(See Appendix B). The dose of Avelumab will be calculated based on the weight 
of the subject determined at trial entry. Following Avelumab infusions, subjects will 
be observed for 1 hour post infusion for potential infusion-related reactions. 
Subjects will receive Avelumab once every 2 weeks for 12 months or until the 
criteria in Section 13.G. (Off Study Criteria) are met. Modifications of the infusion 
rate due to infusion-related reactions are described in Section 13.C.  
 


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In this trial, subjects will receive Avelumab IV infusions at the SCCA. Well trained 
medical staff will monitor and perform the trial drug administration.   
 
3. MHC class I up-regulation: Radiation or intralesional IFNβ-IB injection 
Radiation therapy or intralesional IFNβ-1B injection will be scheduled as an 
outpatient treatment at the SCCA or the University of Washington Medical Center 
(UWMC) between 7 and 10 days after the preceding dose of Avelumab (for 
Groups 1 and 2) and within 2 to 5 days prior to a subsequent T-cell infusion 
(Group 2).  
 
4. T-Cell Infusions 
T-cell infusions will be given at the Seattle Cancer Care Alliance at least 48 hours 
after MHC class I upregulation and preferably at least 48 hours before the next 
dose of Avelumab. All patients will be observed for 2 hours post infusion and will 
be released if no AE requiring prolonged hospitalization have occurred (see 
Section 13). 
T cells will be infused intravenously at a rate of not more than 500 cc/hour through 
an 18- or 20-gauge catheter inserted into a peripheral vein or through a central 
catheter. The infusion bag will be gently mixed periodically during the infusion. 
Subjects will have vital signs per Section 12.B.  
 
 
12.  EVALUATION  
 
See Protocol Appendix B for a summary of patient evaluation for Groups 1 and 2. The 
dates listed on the study calendar are approximate as many patients reside out of the 
area and may not always follow the time points as dictated by the protocol. Results of 
tests and, or procedures conducted as standard of care may be used for research 
purposes.  Potential patients will initially be identified and recruited through the SCCA 
MCC clinic. The informed consent form will be discussed and signed in the presence of a 
GCP-trained investigator. Patients who are found eligible to participate in this protocol 
will be followed closely by a multidisciplinary oncology team that includes medical 
oncologists, dermatologists and radiation oncologists (as appropriate). 
 
12.A.  Patient Evaluation at Each Planned Visit 
 
Patients will have a history taken, ECOG performance status (Appendix A), a physical 
exam with vital signs prior to the T-cell infusion and 1, 14, and 28 days after each 
infusion. Thyroxine 3 (T3), free thyroxine 4 (FT4) and thyroid stimulating hormone (TSH) 
at baseline, at cycle 6 and then every 6 cycles. Merkel virus oncoprotein serology at 
baseline and at time of radiographic evaluation. A comprehensive chemistry panel, and a 
complete blood count at each planned visit including the pre-treatment visit, prior to 
Avelumab infusion, prior to IFNβ-IB injections or radiation, prior to tissue biopsy or 
aspiration, prior to the T-cell infusion and 1, 3, 7, 14, and 28 days after each T cell 
infusion (Appendix B).  It is anticipated that some patients reside outside of the area and 


FHCRC 9245 
 
 
 
 
                  Page 40 
 
may have their +7, +14 and +28 day clinical evaluations completed locally (visits can 
occur+/- 1 day, weekly visits can occur +/- 3 days before or after the planned date).   
 
12.B.  Patient Evaluation During T-Cell Infusions 
 
Blood pressure, heart rate, temperature, respiratory rate, and O2 saturation (by pulse 
oximetry) will be recorded at time 0, every 15 minutes during the 1 to 2 hour T-cell 
infusion, end of infusion, hourly for 2 hours following the T-cell infusion. Adverse events 
will be managed by standard medical practice (see also Section 13). 
 
12.C.  Clinical and Laboratory Evaluation for Toxicity (General Toxicity 
Assessment) 
 
The period of monitoring for treatment-related toxicity will start with the first Avelumab 
infusion. All grade ≥ 3 CTCAE v.4.03 AEs will be recorded up to the last T cell or 
avelumab infusion. Beginning 4 weeks after the last study infusion, only study related 
toxicities will be collected per section 15. To evaluate for potential toxicities, patients will 
undergo a history and physical and, or laboratory evaluations (comprehensive chemistry 
panel and complete blood count) immediately prior to Avelumab infusion, prior to 
radiation therapy or intralesional IFNβ-1B injection, immediately prior to infusions, days 
+1, +3, +7, +14, and +28 after each T cell infusion, and then as clinically indicated after 
the last infusion (for example, every 3 months for the first year, every 6 months for a 
year and then yearly see Appendix B).  Clinical assessments at days +7, +14 and +28 
after T-cell infusions may be performed by the patient’s local provider and records 
assessed by the investigator within 14 days for any toxicities not previously assessed.   
 
12.D.  Efficacy Assessment 
 
12.D.1.  Clinical response by ‘median time to new metastasis’ (primary endpoint) 
 
All patients treated on this protocol will have developed metastatic disease. It is 
plausible that up-regulation of MHC class I with either radiation therapy or intralesional 
IFNβ-1B injection may have an independent effect on the targeted metastasis (as 
assessed by RECIST criteria). It is unlikely however, that localized radiation or 
intralesional interferon will have a significant effect on the risk of distant disease 
development. In contrast, T-cell infusions and/or anti-PD-L1 have the potential to 
generate a broader systemic effect by preventing the occurrence of new distant 
metastasis. To assess the potential systemic effects of adoptively transferred T cells, 
the time for a new metastasis to develop after treatment will be assessed, and this 
parameter will be compared to historical matched controls undergoing standard therapy. 
A preliminary assessment was carried out on 49 of the MCC patients in our cohort who 
originally presented with local or regional disease and later developed metastatic 
disease, at which time they underwent standard therapy (radiation or chemotherapy). 
Patients with systemic immune suppression were eliminated from the analysis as they 
would not be candidates for the combined MHC and T-cell therapy trial. Among these 
patients, the median time to developing a new metastasis was 200 days (see Section 


FHCRC 9245 
 
 
 
 
                  Page 41 
 
18 for further statistical details). Patients will be followed and restaged at least once 
every 3 months and as clinically indicated after 12 months since the last infusion has 
elapsed until new metastasis occur (see Appendix B).  
 
12.D.2  Clinical response by RECIST criteria (secondary endpoint)  
 
Tumor lesions and tumor responses will be assessed according to the Response 
Evaluation Criteria in Solid Tumors (RECIST) v 1.1. Radiographic imaging and clinical 
assessment of residual disease will be compared with pre-infusion assessment. 
Responses will be reported separately for lesions which did or did not received MHC 
Class I upregulation interventions, if the latter are present. Overall responses will be 
reported as well. 
Clinical response assessment of each individual lesions: Response assessment will 
be done through clinical (for example bi-directional measurements of palpable lesions) or 
radiographic (for example CT, PET, or MRI) measurement of each lesion at 8 weeks 
after the first intervention to induce MHC class I upregulation, 8 weeks after the second 
MHC class I upregulation, if the patient undergoes a second set of T cell infusions, and 
then at least once every 3 months (12 weeks) or as clinically indicated. Consistency of 
imaging modality over time is recommended for a given patient. Objective responses in 
lesions which received MHC Class I upregulation and lesions which did not receive MHC 
Class I upregulation, will be reported separately. For each individual lesion or group of 
lesions, a complete response (CR) will be defined as total regression of the lesions, a PR 
as 30% or greater decrease in the sum of the longest diameter of the lesions compared 
to baseline and PD as 20% increase in the sum of the longest diameter of the lesions 
compared to the smallest prior diameter (RECIST v1.1 criteria). 
Overall Response Assessment: 
A complete response (CR) will be defined as total regression of all tumors, a PR as 30% 
or greater decrease in the sum of the longest diameter of target lesions compared to 
baseline and PD as 20% increase in the sum of the longest diameter of target lesions 
compared to the smallest prior diameter (RECIST v1.1 criteria). This assessment will be 
performed 8 weeks after the first MHC class I upregulation, 8 weeks after the second 
MHC class I upregulation, if the patient undergoes a second set of T cell infusions, and 
then at least once every 3 months (12 weeks) or as clinically indicated.  
 
12.E.  Evaluation of Persistence and Function of Adoptively Transferred T Cells 
 
 
12.E.1.  Blood samples 
 
Patients will have 50-60 ml of heparinized blood (green top) and 6 ml of serum (red top) 
tube drawn during the pre-treatment visit, prior to Avelumab infusion, prior to receiving 
radiation or intralesional IFNβ-1B injections (pre-treatment baseline), prior to tissue 
biopsy or aspiration, prior to infusion, and at selected time points following infusion 
(Appendix B, research labs). Samples will be used to evaluate the duration of in vivo 
persistence and function of infused T cells, as well as evaluation of MCPyV TAg titers. 


F H C R C  9 2 4 5  
 
 
 
 
                  P a g e 4 2  
 
All bl o o d s a m pl e s will b e k e pt at r o o m t e m p er at ur e  a n d s e nt t o r o o m E 1 -3 0 5 , F H C R C 
R e s e ar c h C ell B a n k .  
 
1 2 . E. 2.  T u m or s a m pl e s  
 
O nl y p ati e nt s wit h hi st ol o gi c all y c o nfir m e d M C C  will b e e nr oll e d o n t h e st u d y. It i s 
a nti ci p at e d t h at m o st p ati e nt s will u n d er g o a c o nfir m at or y bi o p s y of m et a st ati c l e si o n s ; 
alt h o u g h t hi s m a y n ot b e p o s si bl e f or p ati e nt s wit h a hi g h cli ni c al li k eli h o o d of m et a st ati c 
M er k el c ar ci n o m a a n d l e si o n( s) t h at ar e diffi c ult t o a c c e s s. P ati e nt s u n d er g oi n g 
i ntr al e si o n al I F N β-1 B  i nj e cti o n s will h a v e a n a s pir at e or bi o p s y pri or t o t h e i nj e cti o n s. 
W h e n o bt ai n e d, t h e a s pir at e s  or bi o p si e s will s er v e a s a pr e -tr e at m e nt b a s eli n e f or t h e 
pr e s e n c e of M C P y V T A g -s p e cifi c c e ll s. L o c ali z ati o n of tr a n sf err e d M C P y V T A g - s p e cifi c 
C T L t o t u m or sit e s aft er tr e at m e nt will b e e v al u at e d  i n s el e ct e d p ati e nt s wit h s ur gi c all y 
a c c e s si bl e di s e a s e ( di s e a s e t h at i s a c c e s si bl e b y n e e dl e or c or e bi o p s y, or t h at c a n b e 
e x ci s e d fr o m s ki n or s u p er fi ci al l y m p h n o d e).  
P o st st u d y tr e at m e nt bi o p s y will b e d o n e at a p pr o xi m at el y 4 w e e k s aft er t h e fir st 
A v el u m a b i nf u si o n  ( gr o u p 1) a n d t w o w e e k s aft er t h e fir st T c ell i nf u si o n (gr o u p 2 ). If 
a c c e s si bl e a 2 n d  p o st tr e at m e nt bi o p s y m a y b e p erf or m e d if p ati e nt s u n d er g o a n ot h er 
c y cl e of M H C cl a s s I u p -r e g ul ati o n a n d C T L i nf u si o n s. If e n o u g h m at eri al i s o bt ai n e d, 
si n gl e c ell s u s p e n si o n s will b e pr e p ar e d a n d st ai n e d t o i d e ntif y a nti g e n -s p e cifi c C T L b y 
t etr a m er a n al y si s. Alt er n ati v el y, if li mit e d m at eri al i s  o bt ai n e d, D N A will b e i s ol at e d fr o m 
s a m pl e s a n d u s e d f or hi g h t hr o u g h p ut T C R a n al y si s. All r e s ult s will b e c o m p ar e d t o pr e -
tr e at m e nt p eri p h er al bl o o d a n d, w h er e p o s si bl e, t o pr e -i nf u si o n t u m or s a m pl e s.  
 
1 2 . E. 3.  A s s e s s m e nt of t h e F u n cti o n al C a p a cit y of T r a n sf err e d C ell s 
 
T o e v al u at e t h e dir e ct  e x vi v o  f u n cti o n of t h e tr a n sf err e d c ell s, w h er e  p o s si bl e, t etr a m er +  
c ell s  wit hi n c oll e ct e d P B M C will b e e v al u at e d f or pr o d u cti o n of i ntr a c ell ul ar c yt o ki n e s 
i n cl u di n g I F N, T N Fα  a n d I L -2 i n r e s p o n s e t o c o g n at e a nti g e n  u si n g a n i ntr a c ell ul ar 
c yt o ki n e a s s a y  (I C S).  
 
I ntr a n u cl e ar Ki-6 7 e x pr e s si o n will b e a s s e s s e d o n i nf u s e d t etr a m er+  T c ell s  to e v al u at e 
t h e i n vi v o pr olif er ati o n of  t h e tr a n sf err e d c ell s. T o e v al u at e t h e  e x vi v o pr olif er ati v e 
c a p a cit y of i nf u s e d c ell s c o m p ar e d t o t h e i nf u si o n pr o d u ct, c ar b o x y fl u or e s c ei n 
s u c ci ni mi d yl e st er  (C F S E ) d y e dil uti o n i n r e s p o n s e t o p e pti d e sti m ul ati o n  will b e u s e d .  
 
T h e p h e n ot y p e of t etr a m er +  a nti g e n -s p e cifi c c ell s i n t h e p eri p h er al bl o o d will al s o b e  
a s s e s s e d u si n g e st a bli s h e d i m m u n o p h e n ot y pi n g p a n el s i n cl u di n g a s s e s s m e nt of 
C D 4 5 R O ( m e m or y T c ell s), C D 2 8, C D 2 7, C D 1 2 7, C D 6 2 L a n d C C R 7 ( c e ntr al m e m or y 
T c ell s), C T L A -4 ( C D 1 5 2), P D -1, Ti m -3, a n d C D 5 7 ( e vi d e n c e of n e g ati v e r e g ul ati o n  or 
s e n e s c e n c e).  
 
T h e p er si st e n c e, f u n cti o n, pr olif er ati v e c a p a cit y a n d p h e n ot y p e of i nf u s e d T c ell s will b e 
c orr el at e d t o cli ni c al r e s p o n s e s.  


FHCRC 9245 
 
 
 
 
                  Page 43 
 
 
12.F.  Assessment of Epitope Spreading 
 
Quantification of the overall recognition of the MCPyV T-antigen for each patient will 
likely be performed by testing the reactivity of whole PBMC before and at indicated 
timepoints after treatment to peptides 15 amino acids (aa) in length offset by 5 aa bases 
spanning the whole T-antigen protein to include both CD8 and CD4 responses 
regardless of the HLA type of the patient. The reactivity will be detected using IFNγ 
secretion in a human IFNγ ELISpot assay.76 Results will be presented as the number of 
spot forming cells/105 PBMCs. 
 
 
13.  MANAGEMENT OF TOXICITIES AND COMPLICATIONS 
 
 
13.A.  Toxicity Grading 
 
 
Toxicity grading will be evaluated according to the current guidelines in NCI Common 
Toxicity Criteria version 4.0.77 The full text of the NCI CTCAE is available online at: 
http://evs.nci.nih.gov/ftp1/CTCAE/About.html.  
 
13.B.  Regimen Related Toxicity 
 
If the patient develops excessive Toxicity attributable to one or more component of the 
regimen, the patient will not receive additional study treatment and a treatment of 
corticosteroids will be given, if clinically indicated (please refer to Section 13.C). 
Excessive toxicity will be considered when a non-pre-existing Grade 3 or higher toxicity 
develops after the start of treatment with the following exceptions: 
 
13.B.1.  Expected toxicities attributable to anti-PD-L1 (Avelumab) infusions and 
considered exceptions to criteria for discontinuation include: 
1. Infusion-related reactions (see Section 13.C for management):  
i. Transient (≤ 6 hours) Grade 3 CRS, flu-like symptoms or fever, which is 
controlled with medical management. 
ii. Transient (≤ 24 hours) Grade 3 fatigue, local reactions, headaches, 
nausea, emesis that resolved to ≤ Grade 1.  
iii. Single laboratory values out of normal (excluding Grade ≥ 3 liver function 
tests increase) that are unlikely related to Avelumab according to the 
investigator, do not have a clinical correlate, and resolve to ≤ Grade 1 
within 7 days with adequate medical management. 
iv. Urticaria lesions responding to antihistaminics (Grade 2) 
v. Allergic reaction (flushing or rash) responding to antihistaminic treatment 
(Grade 2).  
 
2. Tumor flare phenomenon defined as local pain, irritation, or rash localized at 
sites of known or suspected tumor. 
 


FHCRC 9245 
 
 
 
 
                  Page 44 
 
3. Tumor-lysis syndrome: If symptoms of tumor lysis occur after the Avelumab 
infusions they will be managed according to best clinical practice (see Section 
13.C.4). Tumor lysis will not constitute a reason to discontinue the Avelumab 
infusions. 
 
4. Change in ECOG PS to ≥ 3 that resolved to ≤ 2 within 14 days (infusions 
should not be given on the following cycle if the ECOG PS is ≥3 on the day of the 
Avelumab administration.  
 
13.B.2.  Expected toxicities attributable to localized radiation of pre-existing 
symptomatic metastasis and considered exceptions to criteria for discontinuation 
include: 
 
i. Non-life-threatening - diarrhea (i.e. 4-6 stools over baseline stools a day) 
ii. Target organ inflammatory responses resulting in Grade 3 or 4 toxicities that 
return to baseline within 4 weeks.  For example: elevated lipase and/or 
amylase after radiation to a pancreatic lesion; or elevated transaminases after 
radiation to a liver lesion.  
iii. Hyperglycemia (Grade 3) lasting <14 days. 
iv. Dry skin, skin pain, and erythema lasting <14 days.  
v. Single laboratory values out of normal that according to the investigator do 
not have a clinical correlate and resolve to ≤ Grade 1 within 7 days with 
adequate medical management. 
 
13.B.3.  Expected toxicities attributable to intralesional IFNβ-1B injections and 
considered exceptions to criteria for discontinuation include: 
 
i. Non-life-threatening infections (i.e. absence of hemodynamic compromise 
requiring pressor support and non-opportunistic infection). 
ii. CRS Grade 3 or less, including but not limited to asthenia, flu-like symptoms, 
myalgia, lymphopenia, and rigors. 
iii. Target organ inflammatory responses resulting in Grade 3 or 4 toxicities that 
resolve within 4 weeks.  For example: elevated lipase and/or amylase after 
injection to a pancreatic lesion; or elevated transaminases after injection to a 
liver lesion. 
iv. Single laboratory values out of normal that according to the investigator do 
not have a clinical correlate and resolve to ≤ Grade 1 within 7 days with 
adequate medical management. 
 
 
13.B.4.  Expected toxicities attributable to T-cell infusions and considered exceptions 
to criteria for discontinuation include: 
 
i. CRS Grade 3 or less, including but not limited to asthenia, flu-like symptoms, 
myalgia, lymphopenia, and rigors. 
ii. Skin rash or erythroderma (Grade 3 toxicity) lasting for <7 days. 


FHCRC 9245 
 
 
 
 
                  Page 45 
 
iii. Hypoxemia requiring continuous oxygen <72 hours, but not requiring 
mechanical ventilation or intubation. 
iv. Fever that resolves within 36 hours of the T-cell infusion. 
v. Lymphopenia (lymphocytes <500) that resolves to baseline levels (pre-
therapy) within 4 weeks.  
vi. Single laboratory values out of normal that according to the investigator do 
not have a clinical correlate and resolve to ≤ Grade 1 within 7 days with 
adequate medical management. 
 
13.C.  Management of Symptoms During Avelumab Infusions 
 
13.C.1.  Special precautions  
 
As infusion reactions have been observed with Avelumab (see Section 3.J.3), 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 
each Avelumab infusion to mitigate infusion-related reactions including hypersensitivity 
reactions. As a routine precaution, subjects enrolled in this trial must also be observed 
for 1 hour post infusion, in an area with resuscitation equipment and emergency agents.  
 
Infusion of Avelumab 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 according 
to the NCI are as outlined in Sections 13.C.2, 13.C.3, and 13.C.4 below. 
 
13.C.2. Infusion-related reactions 
 
A. Symptoms 
• Fever 
• Chills 
• Rigors 
• Diaphoresis 
• Headache 
 


FHCRC 9245 
 
 
 
 
                  Page 46 
 
B. Management  
Table 5. Treatment modification for symptoms of infusion-related reactions 
caused by Avelumab 
NCI-CTCAE Grade 
Treatment Modification for Avelumab 
Grade 1 – mild 
Mild transient reaction; infusion 
interruption not indicated; intervention not 
indicated. 
 
Decrease the Avelumab infusion rate by 
50% and monitor closely for any 
worsening. 
The total infusion time for Avelumab 
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 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 infusion immediately 
and disconnect infusion tubing from the 
subject. 
Subjects must be withdrawn immediately 
from Avelumab treatment and must not 
receive any further Avelumab treatment. 
Definitions: IV=intravenous, NCI-CTCAE=National Cancer Institute-Common Terminology Criteria for 
Adverse Event, NSAIDs=nonsteroidal anti-inflammatory drugs,  
 
Once the Avelumab 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 
subject has a second infusion-related reaction of Grade ≥ 2 on the slower infusion rate, 
the infusion should be stopped and the subject should be removed from Avelumab 
treatment. If a subject experiences a Grade 3 or 4 infusion-related reaction at any time, 
the subject must discontinue Avelumab. 
 
13.C.3.  Management of severe hypersensitivity reactions and flu-like symptoms 
due to Avelumab infusions 
 
If hypersensitivity reaction occurs, the subject must be treated according to the best 
available medical practice. Subjects should be instructed to report any delayed reactions 
to the Investigator immediately. 


FHCRC 9245 
 
 
 
 
                  Page 47 
 
A. Symptoms 
• Impaired airway 
• Decreased oxygen saturation (< 92%) 
• Confusion 
• Lethargy 
• Hypotension 
• Pale / clammy skin 
• Cyanosis 
B. Management 
1. Epinephrine injection and dexamethasone infusion 
2. Patient should be placed on monitor immediately  
3. Alert ICU for possible transfer if required 
 
For prophylaxis of flu-like symptoms, 25 mg of indomethacin or comparable NSAID dose 
(for example, ibuprofen 600 mg, naproxen sodium 500 mg) may be administered 2 hours 
before and 8 hours after the start of each dose of Avelumab IV infusion. Alternative 
treatments for fever (for example, paracetamol) may be given to subjects at the 
discretion of the Investigator. 
 
13.C.4.  Tumor lysis syndrome 
 
In addition, since Avelumab can induce antibody-dependent cell-mediated cytotoxicity 
(ADCC), there is a potential risk of tumor lysis syndrome. Should this occur, subjects 
should be treated per the local guidelines and the management algorithm below (Figure 
135).78 


FHCRC 9245 
 
 
 
 
                  Page 48 
 
 
Definitions: TLS=tumor lysis syndrome 
Figure 135. Assessment and initial management of tumor lysis syndrome 
13.C.5.  Delayed toxicities and immune-related adverse events (irAE) secondary to 
Measure serum potassium, phosphorus, calcium, creatinine, uric acid, and urine output
No TLS at diagnosis
Assess cancer mass
Laboratory TLS
≥2 abnormal laboratory
test values
No symptoms
Small or resected
localized tumor
Medium-size 
cancer mass
Large cancer mass
Bulky tumor or organ 
infiltration
Bone marrow replaced
with cancer
Assess cell-lysis potential
Assess cell-lysis potential
Low
High
Medium or
unknown
Low
High
Medium or
unknown
Negligible Risk
of Clinical TLS
No prophylaxis
No monitoring
Low Risk or
of Clinical TLS
Intravenous fluids
Allopurinol
Daily laboratory tests
High Risk of
Clinical TLS
Intravenous fluids
Rasburicase
Cardiac monitoring
Laboratory tests every
6-8 hr 
Established
Clinical TLS
Intravenous fluids
Rasburicase
Cardiac monitoring
Intensive care unit
Laboratory tests every
4-6 hr 
High Risk of
Clinical TLS
Intravenous fluids
Allopurinol or ras-
buricase
Inpatient monitoring
Laboratory tests every
8-12 hr 
Assess patient
presentation
Preexisting nephropathy
Dehydration
Hypotension
Nephrotoxin exposure
No
Yes
Clinical TLS
Acute kidney injury
Symptomatic hypo-
calcemia
Dysrhythmia


FHCRC 9245 
 
 
 
 
                  Page 49 
 
Avelumab 
 
Patients will also be closely monitored for potential irAEs,79,80 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). IrAEs will be managed according to Section 13.E with 
steroids.  
 
Suggested management for Grade 2 adverse drug reactions (ADR): 
• If a Grade 2 ADR resolves to Grade ≤ 1 by the last day of the current cycle, treatment 
may continue. 
• If a Grade 2 ADR does not resolve to Grade ≤ 1 by the last day of the current cycle, 
infusions should not be given on the following cycle.  If at the end of the following 
cycle the event has not resolved to Grade 1, the subject should permanently 
discontinue treatment with a Avelumab ADR (except for hormone insufficiencies, that 
can be managed by replacement therapy; for these hormone insufficiencies, up to 2 
subsequent doses may be omitted). 
• Upon the second occurrence of the same Grade 2 ADR (except for hormone 
insufficiencies that can be managed by replacement therapy) in the same subject, 
treatment with Avelumab must be permanently discontinued. 
 
13.D.  Management of Symptoms During T-Cell Infusion 
 
Immediate reaction to infusion (i.e. defined as those occurring either during the first 
24 hours following T-cell infusion) may occur due to release of cytokines from T cells 
stimulated by the recognition of targets. Mild transient symptoms have been observed 
with lymphokine-activated killer cell (LAK) and TIL cell infusions, and with infusions of 
antigen-specific T-cell clones (FHCRC Protocol 2140).   
 
13.D.1.  Milder reactions (i.e. <Grade 3 CTCAE v.4 or less severe than specified below) 
would include symptoms such as:  
• Fever, chills, fatigue.  
• Dyspnea, chest tightness, or myalgia, 
• Alteration in vital signs such as:  
- 
Lowering of blood pressure, but with systolic BP ≥ 90 mm Hg, or ≤ 20 mm 
Hg below baseline.  
- 
Tachycardia, but with HR ≤ 130 or ≤ 30 above baseline. 
- 
Tachypnea, but with RR ≤ 32/min or ≤ 10 above baseline. 
- 
Hypoxemia, but O2 saturation ≥ 88% on room air, or ≤ 5% fall from 
baseline. 
- 
Skin changes such as erythema, urticaria, or other rash.  
 


FHCRC 9245 
 
 
 
 
                  Page 50 
 
Management will be by decreasing the rate of infusion and/or appropriate 
supportive care such as:  
• Acetaminophen or Demerol for fever and chills. (All subjects who develop 
fever or chills should have a blood culture drawn). 
• Acetaminophen for headache. 
• Diphenhydramine for nausea and vomiting. 
• Fluid administration for hypotension. 
• Supplemental oxygen for hypoxemia. 
 
13.D.2.  More severe reactions occurring during the 1-2 hour infusion would include 
symptoms such as: 
• Hypotension with systolic BP <90 mm Hg and >20 mm Hg below baseline 
• Tachycardia with HR >130 and >30 above baseline. 
• Tachypnea with RR >32 and >10 above baseline.  
• Hypoxemia with O2 saturation of <88% and >5% fall from baseline. 
 
Management will be by terminating the infusion and administering supportive 
medical care.  
• If patient responds to supportive care by normalization of vital signs or 
resolution of hypoxemia and PI deems it safe to continue, the infusion will be 
restarted at slower rate.   
• If the patient does not respond by normalization of vital signs or hypoxemia 
after supportive care alone, methylprednisolone to ablate the infused 
T cells will be administered as per Section 13.E.  
 
13.D.3.  Any unexpected severe toxicity (see Section 13.B) occurring in the first 
24 hours (due to the T-cell infusion and not attributable to a non-infusion-related cause). 
 
Management will be by supportive medical care and methylprednisolone will be 
administered as per Section 13.E. 
 
13.D.4.  Delayed toxicities secondary to T-cell infusions 
 
Although unlikely, it remains possible that the infusion of MCPyV TAg-specific T cells will 
result in delayed toxicities related to the recognition of tumor tissue. It is anticipated that 
the symptoms and signs will occur within hours to four weeks after completion of the T-
cell infusion. 
 
Any toxicities requiring treatment discontinuation (see Section 13.B) occurring 
within 4 weeks of study treatment (potentially due to the T-cell infusion and not 
attributable to a non-infusion-related cause) will be managed by supportive care and 
MP will be administered as per Section 13.E. 
 


FHCRC 9245 
 
 
 
 
                  Page 51 
 
13.E.  Management of Severe Treatment-related Toxicities with Corticosteroids 
 
i. 
The patient will receive corticosteroids if treatment-related toxicity 
warranting ablation of T cells is observed. 
 
 
ii. 
The patient will be hospitalized for the first 48 hours for monitoring. The 
following dose schedule is recommended: 
 
Day 1 
Intravenous Methylprednisolone at 2 mg/kg  
Day 2 
Intravenous Methylprednisolone at 2 mg/kg 
Day 3-4 
Prednisone at 30 mg po b.i.d. 
Day 5-6 
Prednisone at 15 mg po b.i.d. 
Day 7-8 
Prednisone at 10 mg po b.i.d. 
Day 9-10 
Prednisone at 10 mg po q.d 
Day11-12 
Prednisone at 5 mg po q.d. 
 
iii. 
The in vivo frequency of infused CD8+ T cells will be assayed immediately 
prior to and between 48 and 72 hours after the start of steroid therapy.  
 
13.F.  Concomitant Therapy 
 
13.F.1.  As with all monoclonal antibody therapies, Avelumab carries a risk of allergic 
reaction. Avelumab will be administered at the SCCA in a setting that allows for 
immediate access and administration of therapy for severe allergic or hypersensitivity 
reactions, such as the ability to implement immediate resuscitation measures. Steroids 
(dexamethasone 10 mg), epinephrine (1:1000 dilution), allergy medications 
(antihistamines), or equivalents are available for immediate access. If hypersensitivity 
reaction occurs, the subject must be treated according to the best available medical 
practice.  
 
13.F.2.  Subjects will be pretreated with H1 blockers (diphenhydramine 25-50 mg PO or 
IV, or equivalent), and acetaminophen 500 to 650 mg (orally), 30 to 60 minutes prior to 
each Avelumab and T-cell infusion. 
 
13.F.3.  Any medications (other than those excluded by the clinical trial protocol) that are 
considered necessary for the subjects’ welfare and will not interfere with the IMP may be 
given at the Investigator’s discretion. For example, active infections occurring after 
initiating the study should be treated according to the standard of care. Other agents 
might include analgesics, antinausea medications, antihistamines, diuretics, anti-anxiety 
medications, and medication for pain management, including narcotic agents. 
 


FHCRC 9245 
 
 
 
 
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13.F.4.  The following agents are not allowed while on study:   
• Systemic corticosteroids (except as outlined for management of toxicity of 
Avelumab and non-transduced CTL) 
• Immunotherapy (for example, interleukins, MCC vaccines, intravenous 
immunoglobulin, expanded polyclonal TIL or LAK therapy)  
• Pentoxifylline, other small-molecule or chemotherapy cancer treatment  
• Other investigational agents 
• Any vaccine therapies for the prevention of infectious disease (for example, 
seasonal flu vaccine, human papilloma virus vaccine) except administration of the 
inactive influenza vaccine 
• Growth factors (granulocyte colony stimulating factor or granulocyte macrophage 
colony stimulating factor). Exception: erythropoietin and darbepoietin alpha may 
be prescribed at the Investigator’s discretion 
• Bisphosphonate treatment is not allowed unless it has been initiated more than 
14 days prior to receiving the first administration of Avelumab 
 
13.G.  Off-study Criteria 
 
A patient’s participation on the protocol will be terminated for any of the reasons listed 
below: 
• Progressive disease or development of new metastasis requiring urgent additional 
drugs (for example chemotherapy) 
• The participant withdraws consent 
• Patient death 
• Occurrence of pregnancy  
• Participation in another 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 and / or avelumab infusions are not in 
the best interest of the patient 
• Occurrence of any non-pre-existing AEs / repetitive Grade 2 AEDRs that are 
deemed related to the treatment as defined in Section 13.C.2. 


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• Use of a non-permitted concomitant drug such as steroids, or as defined in Section 
13.F.4 for which the predefined consequence is withdrawal from the trial 
• 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 
 
 
14.  TARGETED/PLANNED ENROLLMENT 
 
TARGETED/PLANNED ENROLLMENT: 20 Subjects 
Ethnic Category 
Sex/Gender 
Females 
Males 
Total 
Hispanic or Latino  
0 
0 
0 
Not Hispanic or Latino 
9 
11 
20 
Ethnic Category: Total of All Subjects * 
9 
11 
20 
Racial Categories 
 
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 
9 
11 
20 
Racial Categories: Total of All Subjects * 
9 
11 
20 
* The “Ethnic Category: Total of All Subjects” must be equal to the “Racial Categories: Total of All” 
All racial groups and ethnicities will be included. 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.  
 
Sex/Gender 
Males  
 
 
 
 
 
 
60% 
Females  
 
 
 
 
 
 
40% 
 
Ethnicity 
Hispanic/Latino 
 
 
 
 
 
3.2% 
Not Hispanic/Latino 
 
 
 
 
96.8% 
 
Race 
American Indian/Alaska Native 
 
 
 
0.18% 
Asian  
 
 
 
 
 
 
1.4% 
Native Hawaiian/Pacific Islander  
 
 
0.98% 
Black/African American 
 
 
 
 
0.84% 
White  
 
 
 
 
 
 
96.6% 


FHCRC 9245 
 
 
 
 
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15.  GUIDELINES FOR ADVERSE EVENTS REPORTING 
15.A.  Reporting of Adverse Events 
 
All unexpected and serious AEs which may be due to study treatment or intervention 
must be reported to the FHCRC Institutional Review Office (IRO) per their current 
reporting requirements.  
 
All grade ≥ 3 CTCAE v.4  AEs will be collected through 4 weeks after the last T-cell 
infusion and per section 12.C. Beginning 4 weeks after the last study infusion, only study 
related toxicities will be collected. 
  
All SAEs that are unexpected and related to study treatment will be reported to the FDA 
on a MedWatch 3500 reporting form that will be submitted to the FDA within 15 calendar 
days. The reports will include the date and time of onset, severity and duration of the 
event, the relationship to study treatment, the treatment given, and the eventual 
outcome. 
 
15.B.  Definitions 
 
Definitions associated with reportable events can be found on the FHCRC IRO extranet 
website. (Relevant FHCRC policies include, but are not limited the documents listed 
[Table 6]. Please also refer to the FHCRC IRO website.) 
 
Table 6. FHCRC IRB policies for reportable events 
 
IRB Policy 2.6 
Adverse Events and Other 
Unanticipated Problems 
Involving Risks to Subjects or 
Others 
http://extranet.fhcrc.org/EN/sections/iro/ir
b/ae.html 
IRB Policy 1.9 
Noncompliance with the 
Office of the Director’s Human 
Research Protection Program 
Policy 
http://extranet.fhcrc.org/EN/sections/iro/ir
b/ae.html 
IRB Policy 1.1 
Reporting Obligations for 
Principal Investigators 
http://extranet.fhcrc.org/EN/sections/iro/ir
b/policy/index.html 
IRB Policy 2.2 
Continuing Review 
http://extranet.fhcrc.org/EN/sections/iro/ir
b/policy/index.html 
IRB Policy 1.13 
Investigational New Drugs 
(IND), Biologics and 
Investigational Device 
Exemptions (IDE) 
http://extranet.fhcrc.org/EN/sections/iro/ir
b/policy/index.html 


FHCRC 9245 
 
 
 
 
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16.  DATA AND SAFETY MONITORING PLAN 
 
16.A.  Primary Monitoring 
 
The PI of the study will have primary responsibility for ensuring that the protocol is 
conducted as approved by the Scientific Review Committee and IRB. The PI will ensure 
that the monitoring plan is followed, that all data required for oversight of monitoring are 
accurately reported to the Protocol Office and Data Safety Monitoring Board (DSMB), 
that all AEs are reported according to protocol guidelines, and that any AEs reflecting 
patient safety concerns are appropriately reported. 
 
16.B.  Monitoring Plan 
 
The PI, or a co-investigator on the study designated by the PI, will personally review with 
the Study Nurse the clinical course of all MCPyV TAg-specific T-cell infusions. The PI or 
his/her designee will review with the Study Nurse the progress of each patient 
undergoing therapy as well as the clinical course of all patients who have completed a 
course of Tcell therapy.  
 
16.C.  Monitoring the Progress of the Trial and the Safety of Participants 
 
The FHCRC PI is responsible for monitoring this clinical trial, with oversight by a DSMB, 
the Protocol and Data Monitoring Committee (PDMC) at the FHCRC, and the FHCRC 
IRB. This is a phase I/II study and the assessment of risk is considered above minimal.  
 
A DSMB will be in place and will meet after four patients on Group 2 have received 
treatment (or sooner if requested) to review the data as it relates to AEs and response to 
treatment. 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 III or greater NIH CTC v4.0 toxicities and SAEs and determine whether the study 
should be prematurely discontinued due to excessive toxicity consistent with Section 
13.B. Ad hoc meetings may be scheduled as needed.  
 
A report from the DSMB will be submitted to the Coordinating Center IRB (FHCRC) and 
the PI. The DSMB will discontinue the review of outcomes when all subjects on this trial 
have completed all protocol-specified follow-up. The DSMB for this study will be 
composed of Drs. Scott Tykodi, Barry Storer, and Michi Shinohara (Table 7). 
 
Table 7. DSMB Members 
 
Name 
Affiliation 
Position 
Dr. Scott Tykodi 
FHCRC  
DSMB Chair 
Dr. Barry Storer 
FHCRC 
DSMB Biostatistician 
Dr. Michi Shinohara 
University of Washington 
DSMB Clinical Investigator 
 


FHCRC 9245 
 
 
 
 
                  Page 56 
 
All members have experience in the management of patients with skin cancers and in 
the conduct and monitoring of clinical trials. Any DSMB member who has or develops a 
significant conflict of interest will be required to resign from the DSMB. DSMB 
membership is for the duration of the clinical trial. If any members leave the DSMB 
during the course of the trial, the PI will promptly appoint their replacement.  
 
A monitor will be retained to monitor study progress. The scope of monitoring will be 
based on the FHCRC/UW Data and Safety Monitoring Plan (DSMP): 
http://www.cancerconsortium.org/rto/prr/DSMPlan.pdf. Per the DSMP, an initial 
monitoring visit is expected after the first 4 patients have received treatment with 100% 
of records verification. Monitoring reports will be forwarded to the DSMB, the IND 
sponsor, and the PI at FHCRC.  
 
Content of Reports for the DSMB:  
• Study number and title. Brief summary of the study design  
• Protocol amendments  
• Status of accrual. If accrual is slower than expected, a plan for increasing 
enrollment 
• Compliance   
• Analyses of primary and secondary endpoints  
• Analyses of AEs and overall safety data. A listing of SAEs by subject and by body 
system  
• Analysis of lab values  
 
DSMB Meeting Minutes  
The CTSO will provide staff to assist with minutes for the DSMB meeting. The FHCRC 
IRB and Regulatory Affairs Manager will be included on the distribution list. At the time of 
IRB annual renewal, DSMB minutes will be required, if not already provided. 
 
Statistical Monitoring Guidelines  
The DSMB will review all Grade 3 or greater toxicities as defined by version 4.0 of NCI 
Common Toxicity Criteria and determine whether the study should be prematurely 
discontinued due to toxicity. Toxicity grading will be evaluated by the clinical 
investigators. Criteria for discontinuing of therapy in an individual patient are described in 
protocol section titled “Management of Toxicities and Complications”. Criteria for 
discontinuing the trial is described in section titled “Data and Safety Monitoring”.  
 
The type and grade of toxicities noted during therapy will be summarized for each dose 
level. All AEs noted by the investigator will be tabulated according to the affected body 
system. Descriptive statistics will be used to summarize changes from baseline in clinical 
laboratory parameters. Tumor responses will be determined as specified above. 
17.  RECORDS 
 
 
The Clinical Research Division at the FHCRC maintains a patient database that allows 
for the storage and retrieval of specific types of patient data including demographic 


FHCRC 9245 
 
 
 
 
                  Page 57 
 
information, protocol registration information, and data from the treatment course. These 
data are collected from a wide variety of sources and conform to institutionally 
established guidelines for coding, collection, key entry, and verification. Each patient will 
be assigned a unique patient number (UPN) to assure patient confidentiality. Any 
publication or presentation will refer to patients by this number and not by name. 
Information about patients enrolled on this protocol that is forwarded to agencies such as 
the FHCRC IRB, EMD Serono, Inc., NIH, and FDA will refer to the patients only by their 
UPN.  
 
Original inpatient and outpatient medical records will be maintained by the medical 
records departments at the institutions where the patients receive their care. The 
majority of their care related to this protocol will be received at the SCCA and UW 
Medical Center. The study nurse and/or data coordinator will maintain a Case Report 
Form (CRF) Notebook for each patient treated on this protocol. The CRF notebooks and 
their contents will be identified by the patient’s initials and UPN only. All supporting 
documents used to verify the accuracy of the data in the case report forms will be kept 
separately. Patient research files will be kept in a locked, controlled-access building. At 
least monthly, the PI or a designated co-investigator will review and cross check the data 
entered on the case report forms with the source documents. 
 
18.  STATISTICAL CONSIDERATIONS  
 
The primary objective of this trial is to assess and compare the safety and potential 
efficacy associated with treating patients who developed metastatic MCC by combined 
MHC up-regulation and anti-PD-L1 antibody treatment (Group 1); and combined MHC 
up-regulation, anti-PD-L1 antibody treatment, and adoptive transfer of MCPyV TAg-
specific polyclonal autologous CD8+ T cells (Group 2). The treatment for each group will 
be considered to have an acceptable safety profile, if the observed toxicity rate is 
consistent with a true rate that does not exceed 30%, whereas toxicity is defined in 
Section 13. The potential efficacy of treatment within each group will be assessed 
primarily by examining the time from first to second metastasis. Among 49 patients who 
would be considered eligible for this trial and who received standard treatment (localized 
radiation or chemotherapy) at UW, the median time to second metastasis was an 
estimated 200 days (Figure 146). This is the benchmark that we shall use in the current 
trial for assessment of potential efficacy.   
 


FHCRC 9245 
 
 
 
 
                  Page 58 
 
 
 
Figure 146. Observed and predicted times to second metastasis 
 
 
A total of 20 patients will be treated in the trial, including 10 in each of Groups 1 and 2 as 
long as the level of safety is deemed to be acceptable. This number was not derived 
based on any formal statistical considerations; rather, it is a number that is a 
compromise between the number of patients that can be enrolled in a reasonable time 
frame and the number of patients whose treatment will provide enough information to 
gain a preliminary assessment of efficacy. From our database of over 850 patients, we 
are in active contact with ~300 living MCC patients. Approximately 120 new patients are 
recruited each year to our cohort. One-third of MCC patients will have a first metastasis 
within 3 years of diagnosis. Each year, approximately 40 patients in our cohort 
experience a first metastasis. Because we now have tetramers that cover >80% of MCC 
patients based on their HLA alleles, we anticipate that generation of functional tetramer-
positive MCPyV-specific T cells from the peripheral blood will be feasible on a sufficient 
number of cases so that we can readily meet targeted enrollment of four cases per year. 
 
The observed time to second metastasis for the 49 patients is summarized in Figure 
146. Based on these data, and in collaboration with our study biostatistician (Ted 
Gooley, PhD), we fit a piecewise exponential model wherein we assumed a constant 
hazard of metastasis in each of nine different time windows as a means of specifying an 
assumed-true distribution for time to metastasis. At the end of each window, the 
assumed-true probability of metastasis is required to equal the observed value at this 
time. From this assumed-true distribution, the assumed-true distribution for the current 


F H C R C  9 2 4 5  
 
 
 
 
                  P a g e 5 9  
 
t h er a p y w a s g e n er at e d b y a s s u mi n g a h a z ar d of f ail ur e t h at i s pr o p orti o n al t o t h at fr o m 
t h e a s s u m e d-tr u e di stri b uti o n fr o m t h e hi st ori c al d at a. Fr o m t hi s di stri b uti o n, 5, 0 0 0 
si m ul at e d tri al s w er e r a n d o ml y g e n er at e d, a n d t h e l o w er li mit of a p oi nt -wi s e c o nfi d e n c e 
i nt er v al w a s e sti m at e d f or e a c h si m ul at e d tri al. T h e pr o p orti o n of tri al s w h er e t hi s 
c o nfi d e n c e li mit e x c e e d e d 2 0 0 d a y s w a s t a k e n a s t h e e sti m at e d p o w er t o o b s er v e a 
st ati sti c all y si g nifi c a ntl y i m pr o v e d m e di a n ti m e t o m et a st a si s c o m p ar e d t o t h e fi x e d ti m e 
of 2 0 0 d a y s. T a bl e 8  b el o w s u m m ari z e s t h e p o w er e sti m at e d i n t hi s m a n n er f or a v ari et y 
of a s s u m e d -tr u e di stri b uti o n s a n d o n e-si d e d si g nifi c a n c e () l e v el s wit h 10 p ati e nt s.  
 
T h er ef or e, wit h 1 0 p ati e nt s tr e at e d, w e’ll h a v e s uffi ci e nt p o w er t o o b s er v e a st ati sti c all y 
si g nifi c a nt i m pr o v e m e nt i n m e di a n ti m e  t o m et a st a si s r el ati v e t o t h e fi x e d hi st ori c al ti m e 
of 2 0 0 d a y s f or s c e n ari o s wit h a s s u m e d -tr u e p ar a m et er s a s s u m m ari z e d a b o v e. F or t h e 
a n al y si s, w e will  u s e ‘ Ti m e 0’ a s t h e fir st d a y of tr e at m e nt a s o p p o s e d t o t h e ti m e of 1 st 
m et a st a si s. P ot e nti al effi c a c y of Gr o u p s 1 a n d 2  will b e a s s e s s e d i n t hi s m a n n er.  If e a c h 
gr o u p i s d e e m e d t o b e p ot e nti all y effi c a ci o u s, t h e n w e will u s e a “ pl a y -t h e-wi n n er” 
a p pr o a c h t o c h o o s e t h e tr e at m e nt t h at will m o v e f or w ar d t o o ur n e xt tri al, w h er e i n t h e 
gr o u p wit h t h e l o n g e st m e di a n ti m e t o m et a st a si s will b e d e cl ar e d t h e wi n n er.  If o nl y o n e 
gr o u p i s d e e m e d t o b e p ot e nti all y effi c a ci o u s a s d e s cri b e d a b o v e, t h e n t hi s i s t h e 
tr e at m e nt t h at will b e t a k e n f or w ar d.  If n eit h er tr e at m e nt i s c o n si d er e d t o b e p ot e nti all y 
effi c a ci o u s, t h e n n eit h er tr e at m e nt will b e t a k e n f or w ar d.  
 
T a bl e 8 . P o w er e sti m at e s 
 
A s s u m e d -tr u e 
M e di a n  
A s s u m e d -tr u e 
H a z ar d R ati o  
Si g nifi c a n c e l e v el 
of 0. 1 0  
Si g nifi c a n c e l e v el 
of 0. 1 5  
3 5 2 d a y s  
3  
7 0 %  
8 8 %  
4 1 1 d a y s  
4  
8 5 %  
9 2 %  
  
 
S e c o n d ar y effi c a c y e n d p oi nt s  i n cl u d e r e s p o n s e t o tr e at m e nt b y R E CI S T crit eri a a n d 
M C C - s p e cifi c s ur vi v al. Alt h o u g h w e will r e p ort t h e o v er all r e s p o n s e r at e ( S e e S e cti o n 
1 2. D. 2 ) u si n g R E CI S T crit eri a t o d et er mi n e cli ni c al r e s p o n s e s i n all p ati e nt s, r a di ati o n 
t h er a p y or i ntr al e si o n al I F Nβ -1 B i nj e cti o n m a y h a v e a n i n d e p e n d e nt eff e ct o n t h e 
t ar g et e d m et a st a si s ( s e e S e cti o n 1 2 . D. 1) a n d m a y n ot a d e q u at el y c a pt ur e t h e eff e ct of 
tr a n sf err e d T c ell s or a nti -P D -L 1  a nti b o d y tr e at m e nt. F urt h er m or e, it i s e sti m at e d t h at 1 0 
p ati e nt s will n ot b e s uffi ci e nt t o e v al u at e t h e eff e ct of t hi s tr e at m e nt o n M C C -s p e cifi c  
s ur vi v al.  
 
H o w e v er, i n p ati e nt s w h o h a d b ot h l e si o n s w hi c h di d a n d di d n ot r e c ei v e M H C Cl a s s I 
u pr e g ul ati o n i nt er v e nti o n s, w e will r e p ort t h e r at e of r e s p o n s e of eit h er gr o u p  of l e si o n s . 
R e s p o n s e a s s e s s m e nt will b e c arri e d o ut p er R E CI S T v 1. 1 i n all l e si o n s w hi c h r e c ei v e d 
M H C Cl a s s I u pr e g ul ati o n a n d all l e si o n s w hi c h di d n ot r e c ei v e M H C Cl a s s I 
u pr e g ul ati o n.  R e s p o n s e s will b e cl a s sifi e d a s C R, P R, S D or P D p er R E CI S T d efi niti o n s. 
T h e O bj e cti v e r e s p o n s e r at e ( O R R) a n d 9 0 % c o nfi d e n c e i nt er v al will b e c al c ul at e d a s 
t h e pr o p orti o n of e v al u a bl e l e si o n s t h at a c hi e v e a C R or P R. T h e O R R f or l e si o n s w hi c h 


FHCRC 9245 
 
 
 
 
                  Page 60 
 
did and did not receive MHC Class I upregulation will be calculated separately, and 
confidence intervals will account for correlation of lesions within a patient.  
 
The study will be suspended pending a review by the DSMB if there is ever sufficient 
evidence to suggest that the true toxicity rate exceeds 30% in any group. In terms of 
toxicity of conventional therapy, fractionated ionizing radiation of 30-60Gy is typically 
effective in controlling the target MCC lesion, but it is commonly associated with transient 
grade ≤ 3 local toxicities such as mucositis and erythema, depending on the target site. 
However, radiation therapy cannot be used to repetitively control multiple recurrences.81 
The rate of grade ≥ 3 toxicities associated with platinum-based chemotherapy regimens 
for the systemic treatment of MCC is approximately 30% to 40%.82 Thus, in order not to 
exceed the toxicity rate induced by standard systemic treatment, enrollment to any group 
will be suspended if evidence suggests that the true toxicity rate exceeds 30%. Sufficient 
evidence will be taken to be an observed proportion of toxicities for which the associated 
lower 80% confidence limit exceeds 30%. Operationally, this limit will be met if any of the 
following proportions is observed: 2/2-3, 3/4-5, 4/6-8, 5/9-10. Based on this rule, if the 
true probability of toxicity is 10%, the probability of suspending one group after 
10 patients is approximately 0.03. If the true probability of toxicity is 50%, the probability 
of suspension is 0.76. 
 
 
19.  ADMINISTRATIVE CONSIDERATIONS 
 
19.A.  Institutional Review Board 
 
In accordance with federal regulations (21 CFR 312.66), an IRB that complies with 
regulations in 21 CFR 56 must review and approve this protocol and the informed 
consent form prior to initiation of the study. 
 
19.B.  Termination of Study 
 
The study will be stopped if any of the following events occur: 
 
• All 20 patients have completed treatment. 
 
• Stopping rules for toxicity have been met. Accrual will be put on hold for 
discussion with the DSMB regarding a possible change in study design. 
 
• The PI reserves the right to terminate the study at any time. The FDA may also 
terminate the study. 
 
20.  REFERENCES 
 
1. 
Henness S, Vereecken P. Management of Merkel tumours: an evidence-based review. 
Curr Opin Oncol. 2008;20(3):280-286. 


FHCRC 9245 
 
 
 
 
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2. 
Engels EA, Frisch M, Goedert JJ, Biggar RJ, Miller RW. Merkel cell carcinoma and HIV 
infection. Lancet. 2002;359(9305):497-498. 
3. 
Heath M, Jaimes N, Lemos B, et al. Clinical characteristics of Merkel cell carcinoma at 
diagnosis in 195 patients: the AEIOU features. J Am Acad Dermatol. 2008;58(3):375-381. 
4. 
Feng H, Shuda M, Chang Y, Moore PS. Clonal integration of a polyomavirus in human 
Merkel cell carcinoma. Science. 2008;319(5866):1096-1100. 
5. 
Tai P, Yu E, Assouline A, Lian JD, Kurian J, Krzisch C. Management of Merkel cell 
carcinoma with emphasis on small primary tumors--a case series and review of the current 
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6. 
Mojica P, Smith D, Ellenhorn JD. Adjuvant radiation therapy is associated with improved 
survival in Merkel cell carcinoma of the skin. J Clin Oncol. 2007;25(9):1043-1047. 
7. 
Poulsen M, Rischin D, Walpole E, et al. High-risk Merkel cell carcinoma of the skin 
treated with synchronous carboplatin/etoposide and radiation: a Trans-Tasman Radiation 
Oncology Group Study--TROG 96:07. J Clin Oncol. 2003;21(23):4371-4376. 
8. 
Bichakjian CK, Olencki T, Alam M, et al. Merkel cell carcinoma, version 1.2014. Journal 
of the National Comprehensive Cancer Network : JNCCN. 2014;12(3):410-424. 
9. 
Pectasides D, Pectasides M, Psyrri A, et al. Cisplatin-based chemotherapy for merkel cell 
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Tai PT, Yu E, Winquist E, et al. Chemotherapy in neuroendocrine/Merkel cell carcinoma 
of the skin: case series and review of 204 cases. J Clin Oncol. 2000;18(12):2493-2499. 
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Voog E, Biron P, Martin JP, Blay JY. Chemotherapy for patients with locally advanced or 
metastatic Merkel cell carcinoma. Cancer. 1999;85(12):2589-2595. 
12. 
Lemos BD, Storer BE, Iyer JG, et al. Pathologic nodal evaluation improves prognostic 
accuracy in Merkel cell carcinoma: analysis of 5823 cases as the basis of the first 
consensus staging system. J Am Acad Dermatol. 2010;63(5):751-761. 
13. 
Albores-Saavedra J, Batich K, Chable-Montero F, Sagy N, Schwartz AM, Henson DE. 
Merkel cell carcinoma demographics, morphology, and survival based on 3870 cases: a 
population based study. J Cutan Pathol.37(1):20-27. 
14. 
Sastre-Garau X, Peter M, Avril MF, et al. Merkel cell carcinoma of the skin: pathological 
and molecular evidence for a causative role of MCV in oncogenesis. J Pathol. 
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Shuda M, Feng H, Kwun HJ, et al. T antigen mutations are a human tumor-specific 
signature for Merkel cell polyomavirus. Proc Natl Acad Sci U S A. 2008;105(42):16272-
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Houben R, Shuda M, Weinkam R, et al. Merkel cell polyomavirus-infected Merkel cell 
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Shuda M, Arora R, Kwun HJ, et al. Human Merkel cell polyomavirus infection I. MCV T 
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18. 
Schowalter RM, Pastrana DV, Pumphrey KA, Moyer AL, Buck CB. Merkel cell 
polyomavirus and two previously unknown polyomaviruses are chronically shed from 
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Kwun HJ, Guastafierro A, Shuda M, et al. The minimum replication origin of merkel cell 
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FHCRC 9245 
 
 
 
 
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APPENDIX A 
 
ECOG / Zubrod Performance Status 
 
0 
Fully active, able to carry on 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 house work, office work 
 
2 
Ambulatory and capable of all self-care but unable to carry out any work activities. 
Up and about more than 50% of waking hours 
 
3 
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 
 


FHCRC 9245 
 
 
 
 
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APPENDIX B 
 
Group 1 Monitoring Schedulea 
GROUP 1 
Event 
History and 
PE 
Chemistry 
Panel 
CBC 
T3 FT4 
TSH 
Tumor 
Biopsy 
Blood for 
Research 
Radiographic 
Evaluation 
Oncoprotein 
Serology 
Pre-treatment 
Evaluation for 
treatment 
X 
X 
X 
Xf 
Xg 
X  
Xb 
X 
  
Day 0  
PD-L1 infusionc 
X 
X 
X 
 
 
X  
  
 
Week 
+1/+2 
Day 
+7 - 
+10 
Administer 
Radiation OR 
Intralesional 
IFNβ-1B 
X 
X 
X 
 
 
X 
  
 
Week 
+2 
Day 
+14 
PD-L1 infusion 
X 
X 
X 
 
 
X  
  
 
Week 
+4 
Day 
+28 
PD-L1 infusion 
X 
X 
X 
 
Xg 
X  
  
 
Week 
+6 
Day 
+42 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Week 
+8 
Day 
+56 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Week 
+9 
Day 
+63 
Staging 
  
  
  
 
 
  
X         
(Day 56-
70) 
X 
Week  
+10 
Day 
+70 
PD-L1 infusion 
X 
X 
X 
Xf 
 
X 
  
 
Week 
+11/+12 
Day 
+77- 
+80 
Administer 
Radiation OR 
Intralesional 
IFNβ-1B (if 
indicated)d 
(X) 
(X) 
(X) 
 
 
(X) 
  
 
Week  
+12 
Day 
+84 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Week 
+14 
Day 
+98 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Week 
+16 
Day 
+112 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Week 
+18 
Day 
126 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Week 
+19 
Day 
+133 
Staging 
  
  
 
 
  
X         
(Day 
126-140) 
X 
Week 
+20 
Day 
+140 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Week 
+22 
Day 
+154 
PD-L1 infusion 
X 
X 
X 
Xf 
 
X 
  
 
Week 
+24 
Day 
+168 
PD-L1 infusion 
X 
X 
X 
 
 
X 
  
 
Beyond +52 
weeks  
Study ends  
Xe 
Xe 
Xe 
 
 
Xe 
Xe 
X 
Beyond +24 
weeks infusion 
Q2 weeks for up 
to 1 year total 
on treatment 
PD-L1 infusion 
X 
X 
X 
 
 
X 
 
 


FHCRC 9245 
 
 
 
 
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Test results and procedures conducted as per standard of care may be used for eligibility determination and evaluation 
for toxicity. 
a The dates listed on the study calendar above are approximate as many patients are expected to reside out of state 
and his oncologists and laboratories may not always be able to follow the time points as dictated by the protocol. 
b Includes brain evaluation. 
c Patients may receive preferably one, but up to 3 infusions of anti-PD-L1 before the first MHC class I up-regulation 
intervention. 
d The MHC class I upregulation can be repeated once, at any time point, if there is radiological disease present. 
Treatment must start within 4 weeks of the preceding radiological evaluation.  
e At least every 3 months for one year, every 6 months for one year then yearly and as clinically indicated. 
f at baseline, at cycle 6 and then every 6 cycles. 
gIf accessible. 
 
Group 2 Monitoring Schedulea 
 
GROUP 2   
 Event   
History 
and PE   
Chemistry   
 CBC   
T3 
FT4 
TSH 
Tumor 
Biopsy 
 Blood for 
Research   
 
Radiographic 
Evaluation   
Oncoprotein 
Serology 
 Pre-
treatment   
 Evaluation for 
treatment   
 X   
 X   
 X   
 X   
Xf 
Xg 
 X  
 Xb 
X 
    
 Day 0   
 PD-L1 infusionc   
 X   
 X   
 X   
 
 
 X   
    
 
 Week 
+1/+2   
 Day+7 - +10   
 Administer Radiation OR 
Intralesional IFNβ-1B   
 X   
 X   
 X   
 
 
 X   
    
 
    
 Day +12  (T1+0)  
 Infusion #1 
(1010cells/m2)   
 X   
 X   
 X   
 
 
 X   
    
 
    
 Day +13  (T1+1)  
* Day after infusion 1   
 X   
 X   
 X   
 
 
 X   
    
 
 Week +2   
 Day +14   
 PD-L1 infusion   
 
 
 
 
 
 
    
 
Week +2 
Day +15 (T1+3) 
Day 3 after Infusion #1 
  
 X   
 X   
 
 
 X   
  
 
 Week +3   
 Day +19  (T1+7) 
* Day 7 after Infusion #1   
  
 X   
 X   
 
 
 X   
    
 
Week 
+3/+4 
Day +26 (T1+14) 
Day 14 after Infusion #1 
 X   
 X   
 X   
 
 
 X   
  
 
 Week +4   
 Day +28   
 PD-L1 infusion   
 X   
 X   
 X   
 
Xg 
 X   
    
 
 Week +5   
 Day 40  (T1+28 
/ T2+0) 
 Infusion #2 
(1010cells/m2)   
 X   
 X   
 X   
 
 
 X   
    
 
    
 Day +41  (T2+1) 
 * Day after infusion 2   
 X   
 X   
 X   
 
 
 X   
    
 
 Week +6   
 Day +42   
 PD-L1 infusion   
 
 
 
 
 
 
    
 
Week +6 
Day +43 (T2+3) 
Day 3 after Infusion #2 
  
 X   
 X   
 
 
  X  
  
 
  
 Day +47  (T2+7) 
* Day 7 after Infusion #2   
  
 X   
 X   
 
 
 X   
    
 
Week +7 
Day +54 (T2+14) 
Day 14 after Infusion #2 
 X   
 X   
 X   
 
 
X 
  
 
 Week +8   
 Day +56   
 PD-L1 infusion   
 X   
 X   
 X   
 
 
 X   
    
 
 Week +9   
 Day +63   
 Staging   
 X   
 X   
 X   
 
 
    
 X (Day 56-
70)   
X 
Week +9 
Day +68 (T2+28) 
Day 28 after Infusion #2 
 X   
 X   
 X   
 
 
 X   
  
 
 Week 
+10   
 Day +70   
 PD-L1 infusion   
 X   
 X   
 X   
Xf 
 
 X   
    
 
 Week 
+11/+12   
 Day +77- +80   
 Administer Radiation OR 
Intralesional IFNβ-1B (if 
indicated)d   
 (X)   
 (X)   
 (X)   
 
 
 (X)   
    
 
    
 Day +82  (T3+0) 
 * Infusion #3 (1010 
cells/m2) (if indicated)   
 (X)   
 (X)   
 (X)   
 
 
 (X)   
    
 
    
 Day +83  (T3+1) 
* Day after infusion 3  (if 
indicated)   
 (X)   
 (X)   
 (X)   
 
 
 (X)   
    
 
Week +11 
Day +85 (T3+3) 
Day 3 after Infusion #3 
 
X 
X 
 
 
 X   
  
 
 Week 
+12   
 Day +84   
 PD-L1 infusion   
 X   
 X   
 X   
 
 
 X   
    
 
 Week 
+13   
 Day +89  (T3+7) 
 * Day 7 after Infusion #3 
(if indicated)   
  
 X   
 (X)   
 
 
 (X)   
    
 
Week +13 
Day +96 (T3+14) 
Day 14 after Infusion #3 
 X   
 X   
 X   
 
 
 X   
  
 
 Week 
+14   
 Day +98   
 PD-L1 infusion   
 X   
 X   
 X   
 
 
 X   
    
 
 Week 
+15   
 Day +110  
(T3+28 / T4+0) 
 * Infusion #4 (1010 
cells/m2) (if indicated)   
 (X)   
 (X)   
 (X)   
 
 
 (X)   
    
 
    
 Day +111  
(T4+1) 
* Day after infusion 4  (if 
indicated)   
 (X)   
 (X)   
 (X)   
 
 
 (X)   
    
 
 Week 
+16   
 Day +112   
 PD-L1 infusion   
 X   
 X   
 X   
 
 
 X   
    
 
Week +16 
Day +113 (T4+3) 
Day 3 after Infusion #4 
 
 X   
 X   
 
 
X    
  
 


F H C R C  9 2 4 5  
 
 
 
 
                  P a g e 6 9  
 
 W e e k  
+ 1 7   
 D a y + 1 1 7  
( T 4 + 7) 
* D a y 7 aft er I nf usi o n # 4 
(if i n dic at e d)   
  
 X   
 ( X)   
 
 
 ( X)   
    
 
W e e k + 1 7  
D a y + 1 2 4 
( T 4 + 1 4) 
D a y 1 4 aft er I nf usi o n # 4  
 X   
 X   
 X   
 
 
X  
  
 
 W e e k  
+ 1 8   
 D a y 1 2 6   
 P D -L 1 i nf usi o n   
 X   
 X   
 X   
 
 
 X   
    
 
 W e e k 
+ 1 9   
 D a y + 1 3 3   
 St a gi n g   
    
    
    
 
 
    
 X ( D a y 1 2 6 -
1 4 0)  
X  
 W e e k  
+ 1 9   
 D a y + 1 3 8 
( T 4 + 2 8) 
D a y 2 8 aft er I nf usi o n # 4  
 X   
 X   
 X   
 
 
  
    
 
 W e e k  
+ 2 0   
 D a y + 1 4 0   
 P D -L 1 i nf usi o n   
 X   
 X   
 X   
 
 
 X   
    
 
 W e e k 
+ 2 2   
 D a y + 1 5 4   
 P D -L 1 i nf usi o n   
 X   
 X   
 X   
X f 
 
 X   
    
 
 W e e k 
+ 2 4   
 D a y + 1 6 8   
 P D -L 1 i nf usi o n   
 X   
 X   
 X   
 
 
 X   
    
 
B e y o n d 5 2 w e e ks   
 
St u d y e n ds  
 X e   
 X e   
 X e   
 
 
 X e   
X e 
X  
B e y o n d 
+ 2 4 
w e e ks 
i nf usi o n 
Q 2 w e e ks 
f or u p t o 
1 y e ar 
t ot al o n 
tr e at m e nt  
 
P D -L 1 i nf usi o n 
X  
X  
X  
 
 
X  
  
 
 
T e st r e s ult s a n d pr o c e d ur e s c o n d u ct e d a s p er st a n d ar d of c ar e m a y b e u s e d f or eli gi bilit y d et er mi n ati o n  a n d e v al u ati o n 
f or t o xi cit y. 
a  T h e d at e s li st e d o n t h e st u d y c al e n d ar a b o v e ar e a p pr o xi m at e a s m a n y p ati e nt s ar e e x p e ct e d t o r e si d e o ut of st at e 
a n d hi s o n c ol o gi st s a n d l a b or at ori e s m a y n ot al w a y s b e a bl e t o f oll o w t h e ti m e p oi nt s a s di ct at e d b y t h e pr ot o c ol.  
b  I n cl u d e s br ai n e v al u ati o n. 
c P ati e nt s m a y r e c ei v e pr ef er a bl y o n e, b ut u p t o 3 i nf u si o n s of a nti - P D-L 1 b ef or e t h e fir st M H C cl a s s I u p -r e g ul ati o n 
i nt er v e nti o n.  
d  T h e M H C cl a s s I u pr e g ul ati o n a n d i nf u si o n s c a n b e r e p e at e d o n c e, at a n y ti m e p oi nt , if t h er e i s r a di ol o gi c al di s e a s e 
pr e s e nt. Tr e at m e nt m u st st art wit hi n 4 w e e k s of t h e l a st r a di ol o gi c al e v al u ati o n. A c u m ul ati v e t ot al of 4 i nf u si o n s c a n 
b e a d mi ni st er e d . R a di ati o n t h er a p y or i ntr al e si o n al I F N-1 B i nj e cti o n  will t y pi c all y b e a d mi ni st er e d b ef or e t h e fir st 
i nf u si o n of s u b s e q u e nt c y cl e s. M o nit ori n g will e n d at l e a st 2 0 w e e k s aft er t h e l a st i nf u si o n or wit h t h e d e v el o p m e nt of 
n e w m et a st a si s.  
e At l e a st e v er y 3 m o nt h s  f or o n e y e ar, e v er y 6 m o nt h s f or o n e y e ar t h e n y e arl y a n d a s cli ni c all y i n di c at e d.  
f at b a s eli n e, at c y cl e 6 a n d t h e n e v er y 6 c y cl e s . 
g If a c c e s si bl e. 
 
 
 


FHCRC 9245 
 
 
 
 
                  Page 70 
 
APPENDIX C 
 
Outcomes of patients with MCC who received IFNβ-1B injections 
 
Patient 
Injected lesion(s) 
Number of injections 
Response  
Draining node 
swelling 
1 * 
3 in-transit 
metastases 
27 (to each of 3 lesions) 
over 37 days. 
Complete response 
Unknown 
2 (w419) 
Primary tumor 
14 over 14 days. 
Partial response 
Yes, no tumor by 
IHC 
3 (w380) 
2 recurrences 
15 injections per each of 
2 lesions over 15 days. 
Partial response 
No 
4 (w438) 
Primary tumor 
9 over 9 days. 
Complete response 
Yes, no tumor by 
IHC 
5 (w425) 
Recurrence 
15 over 15 days. 
 
Progressive disease 
No 
6 (w390) 
Recurrence 
12 over 24 days. 
4 week rest. 
3 over 5 days. 
Partial response 
No 
7 (w131) 
Recurrence 
12 over 15 days. 
 
Partial response 
No 
8 (w732) 
Multiple recurrences 
21 over 31 days. 
Complete response** 
No 
9 (w668) 
2 nodal metastases 
10 over 14 days  
Partial response 
N.A. 
*   Some details of this case were previously reported (Nakajima, et al 2009) 
** Ten un-injected lesions on the same leg also had complete responses to interferon.