Clinical trial NCT03476174
Conditions: Melanoma
Sponsor: Ralph Hauke
Start date: 2018-05-07; Completion date: 2019-10-16
Enrollment: 1 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
The primary objective of this single arm phase 2 trial is to assess the response rate \[complete response (CR) + partial response (PR)\] of sequential therapy of pembrolizumab followed by HD IL-2 in subjects with stage IV malignant melanoma. Response assessment will be performed after pembrolizumab therapy, and response reassessment will be performed after HD IL-2 therapy using revised RECIST 1.1.

Detailed description:

Arms: arm 1: Pembrolizumab 200 mg IV over 30 minutes; Day 1 of each cycle 3 weeks (21 days) for 2 cycles. IL-2 600,000 IU/kg2 IV over 15 minutes every 8 hours for up to 14 doses over 5 days; Days 1-5 = Cycle 1; 9 days of rest in between; Days 15-19 = Cycle 2
Interventions: intervention 1: Pembrolizumab intervention 2: Interleukin-2
Intervention descriptions: intervention 1: Subjects will receive 200 mg pembrolizumab every 3 weeks for two cycles. Cycle length is 21 days (i.e., 3 weeks). On the last day of Cycle 2 (+/- 3 days), disease status will be assessed via imaging. intervention 2: Following the pembrolizumab treatment, HD IL-2 treatment will commence. After completing the 2 cycles of pembrolizumab, High Dose Interleukin-2 (HD IL-2) will be administered. This will require a hospital stay of at least 5 days for each treatment cycle. Established guidelines will be followed for safely administering HD-IL-2 and managing toxicities from this treatment. Two treatment cycles will be administered, Cycle 2 being separated from Cycle 1 by approximately 9 days after completion (assuming subject has recovered from Cycle 1 adequately to proceed with Cycle 2). Four weeks after completion of the 2 cycles (called a course of HD IL-2 therapy), disease status will be monitored via CT of the chest and abdomen/pelvis using revised RECIST guidelines

Protocol documents:
Prot_SAP_000.pdf: 
Clinical Study Protocol 
HCRN MMP-MEL16-261 
 
MMP-01: Pilot Study Evaluating Activity of the Combination of Anti-PD-1 Antibody with 
High Dose IL-2 in Metastatic Melanoma 
 
 
Sponsor Investigator 
Ralph Hauke, MD 
Nebraska Cancer Specialists 
Midwest Cancer Center-Legacy 
17201 Wright Street, Suite 200 
Omaha, NE 68130 
Tel: 402-334-4773 
Fax 402-330-7463 
 
Co-Investigator 
Arkadiusz Z. Dudek, MD, PhD  
HealthPartners Regions Cancer Care and Frauenshuh Cancer Care Centers 
Suite 1161 Mail Stop 11101G 
640 Jackson Street, St. Paul, MN 55101 
Tel: 651 254-3321 
Fax: 651 254-3102 
 
Statistician 
Menggang Yu PhD 
 
Trial Management Provided by  
Hoosier Cancer Research Network, Inc. 
500 N. Meridian, Suite 100 
Indianapolis, IN 46204 
 
 
Trial Supported by  
Prometheus Laboratories 
 
IND exempt per FDA: 12/12/17 
 
 
Initial Protocol Version Date: 28FEB2018 
Protocol Amendment Version Date:  
 
 
 
 


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PROTOCOL SIGNATURE PAGE 
 
MMP-01: Pilot Study Evaluating Activity of the Combination of Anti-PD-1 Antibody with High 
Dose IL-2 in Metastatic Melanoma 
 
 
VERSION DATE: 28FEB2018 
I confirm I have read this protocol, I understand it, and I will work according to this protocol and 
to the ethical principles stated in the latest version of the Declaration of Helsinki, the applicable 
guidelines for good clinical practices, whichever provides the greater protection of the 
individual. I will accept the monitor’s overseeing of the study. I will promptly submit the 
protocol to applicable institutional review board(s). 
 
 
____________________________________ 
________________________ 
Signature of Site Investigator  
 
 
  
Date 
 
 
____________________________________ 
Site Investigator Name (printed) 
 
 
 
Site Investigator Title 
 
 
 
Name of Facility 
 
 
 
Location of Facility (City and State) 
 
 
 
 PLEASE COMPLETE AND EMAIL A COPY TO HCRN 
 
 
 
____________________________________ 
____________________________________ 
____________________________________ 


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SYNOPSIS 
TITLE 
MMP-01: Pilot Study Evaluating Activity of the Combination of anti-
PD-1 Antibody with High Dose IL-2 in Metastatic Melanoma 
SHORT TITLE 
Anti-PD-1 Antibody with High Dose IL-2 in Metastatic Melanoma 
PHASE  
Phase II Pilot 
OBJECTIVES 
Primary Objective 
Assess the response rate [complete response (CR) + partial response 
(PR)] of sequential therapy of pembrolizumab followed by high dose 
IL-2 (HD IL-2) in subjects with stage IV malignant melanoma. 
Response assessment will be performed using revised RECIST 1.1. 
 
Secondary Objectives 
• Characterize safety, tolerability and adverse effects (AE) profile 
of pembrolizumab followed by HD IL-2 in subjects with stage IV 
malignant melanoma. 
• Measure Progression-Free Survival (PFS) using revised RECIST 
1.1 after completion of 2 cycles of pembrolizumab and 2 cycles of 
HD IL-2 in all the subjects enrolled in the study. 
• Measure overall survival (OS) at 12 months in subjects with stage 
IV malignant melanoma who showed a response [stable disease 
(SD), complete response (CR) or partial response (PR)] following 
completion of 2 cycles of pembrolizumab and 2 cycles of HD IL-
2. 
 
Exploratory Objectives 
• Correlate PD-L1 expression in archived diagnostic tumor tissue 
with best clinical response (revised RECIST criteria) and 
treatment outcome in subjects with stage IV malignant melanoma. 
• Correlate Th1/Th2 ratio and myeloid-derived suppressor cells in 
peripheral blood during therapy with best clinical response 
(revised RECIST criteria) and treatment outcome in subjects with 
stage IV malignant melanoma. Data will be collected prior to 
treatment, during treatment after the 2nd dose of pembrolizumab 
and after each cycle of HD IL-2. 
• Correlate the change in soluble PD-L1 and arginine levels in 
serum/plasma during therapy with best clinical response (revised 
RECIST criteria) and treatment outcome in subjects with stage IV 
malignant melanoma. Data will be collected prior to treatment, 
during treatment after the 2nd dose of pembrolizumab and after 
each cycle of HD IL-2. 
• Correlate intestinal microbiome at baseline with best clinical 
response (revised RECIST criteria) and treatment outcome in 
subjects with stage IV malignant melanoma.  


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STUDY DESIGN 
Single arm 2-stage Phase II Simon design 
KEY 
ELIGIBILITY 
CRITERIA (See 
Section 3 for full 
eligibility criteria) 
1. Age ≥ 18 years at the time of consent. 
2. Histologically-confirmed diagnosis of unresectable stage IV or 
metastatic melanoma not amenable to local therapy. 
3. Measurable disease, defined as at least 1 tumor that fulfills the 
criteria for a target lesion according to RECIST 1.1 (Section 9), and 
obtained by imaging within 28 days prior to registration for 
protocol therapy. 
4. Has an Eastern Cooperative Oncology Group (ECOG) performance 
status of 0 or 1 within 28 days prior to registration for protocol 
therapy. 
5. Life expectancy of 6 months or greater as determined by the 
treating physician. 
6. Adequate hepatic function within 28 days prior to registration for 
protocol therapy defined as meeting all of the following criteria: 
i. 
total bilirubin ≤ 1.5 mg/dl × upper limit of normal (ULN) OR 
direct bilirubin ≤ ULN for subjects with total bilirubin levels 
> 1.5 mg/dl x ULN (except in patients with Gilbert’s 
syndrome who must have a total bilirubin less than 3.0 
mg/dl.) 
ii. 
and aspartate aminotransferase (AST) ≤ 2.5 × ULN or ≤ 5 × 
ULN for subjects with known hepatic metastases 
iii. 
and alanine aminotransferase (ALT) ≤ 2.5 × ULN or ≤ 5 × 
ULN for subjects with known hepatic metastases 
7. Adequate renal function within 28 days prior to registration for 
protocol therapy defined by either of the following criteria:  
i. 
Serum creatinine ≤ 1.5 mg/dL 
ii. 
OR if serum creatinine > 1.5 mg/dL, estimated glomerular 
filtration rate (GFR) ≥ 40 mL/min 
8. Adequate hematologic function within 28 days prior to registration 
for protocol therapy defined as meeting all of the following criteria: 
i. 
hemoglobin ≥ 9.0 g/dL 
ii. 
and absolute neutrophil count (ANC) ≥ 1000/L without the 
support of filgrastim 
iii. 
white blood cells (WBC) ≥ 3000/L 
iv. 
and platelet count ≥ 100 × 109/L 
9. Adequate coagulation functioning within 28 days prior to 
registration for protocol therapy defined by either of the following 
criteria:  
i. 
INR < 1.5 × ULN 
ii. 
OR for subjects receiving warfarin or LMWH, the subjects 
must, in the investigator’s opinion, be clinically stable with 
no evidence of active bleeding while receiving anticoagulant 
therapy. The INR for these subjects may exceed 1.5 × ULN if 
that is the goal of anticoagulant therapy. 


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10. Adequate baseline pulmonary function test (FEV1 > 2 L or ≥ 75% 
of predicted for height and age). 
11. Because the teratogenicity of pembrolizumab and IL-2 is not 
known,  
• Female subject of childbearing potential (WOCP) must not be 
pregnant confirmed by a negative urine or serum pregnancy 
test within 72 hours of the study registration. If the urine test 
cannot be confirmed as negative, a serum pregnancy test will 
be required. 
12. < 2 lines of prior therapy for metastatic melanoma. Cannot have 
received prior therapy with HD IL-2. May have had one prior line 
of therapy that included a check point inhibitor.  
13. Not received radiation therapy within 21 days of initiation of study 
treatment, and the measurable disease must have been outside of 
the radiation port. 
14. More than 3 weeks must have elapsed since any prior systemic 
therapy at the time the subject receives the first dose of 
pembrolizumab. Subject’s toxicities must have recovered to a grade 
1 or less (except for toxicities such as alopecia or vitiligo) 
15. No second active malignancy.  
16. No autoimmune diseases (see full eligibility criteria) 
17. Willingness and ability to comply with scheduled visits, treatment 
plans, laboratory tests, and other study procedures. 
18. Provide written informed consent and HIPAA authorization for 
release of personal health information, approved by an Institutional 
Review Board/Independent Ethics Committee (IRB/IEC). HIPAA 
authorization may be included in the informed consent or obtained 
separately. 
STATISTICAL 
CONSIDERATIONS 
 
The optimal two-stage design to test the null hypothesis that P 
≤ 0.260 versus the alternative that P ≥ 0.500 with 80% power 
and 0.05 level of significance proceeds as follows. After 
testing the drug on 11 patients in the first stage, the trial will 
be terminated if 3 or fewer respond. If the trial goes on to the 
second stage, a total of 29 patients will be studied. If the total 
number responding is less than or equal to 11, the drug is 
rejected. The probability of early termination at stage 1 with a 
total sample size of 11 is 0.685; the probability of going on to 
stage 2 with a total sample size of 29 is 0.315. The sample 
size calculation is conducted using the software PASS 13 
(NCSS, Kaysville, Utah 2014) 
TOTAL  
NUMBER OF 
SUBJECTS 
N = Up to 29 subjects 


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ESTIMATED 
ENROLLMENT 
PERIOD 
Estimated 36 months 
ESTIMATED 
STUDY 
DURATION  
Estimated 40 months 
 
 


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TABLE OF CONTENTS 
 
1. 
Background and Rationale ......................................................................................... 9 
2. 
Study Objectives and Endpoints .............................................................................. 13 
3. 
Eligibility Criteria ..................................................... Error! Bookmark not defined. 
4. 
Subject Registration ................................................................................................. 18 
5. 
Treatment Plan ......................................................................................................... 18 
6. 
Toxicities and Dose Delays/Dose Modifications ..................................................... 24 
7. 
Study Calendar & Evaluations ................................................................................. 30 
8. 
Biospecimen Studies and Procedures ...................................................................... 33 
9. 
Criteria for Disease Evaluation ................................................................................ 34 
10 
Drug Information ..................................................................................................... 38 
11 
Adverse Events ........................................................................................................ 41 
12 
Statistical Methods ................................................................................................... 44 
13 
Trial Management .................................................................................................... 46 
14. Data Handling and Record Keeping ........................................................................ 47 
15 
Ethics........................................................................................................................ 48 
16 
References ................................................................................................................ 49 
Appendix 1: ECOG Performance Status........................................................................... 51 
 
 
 
 
 


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SCHEMA 
 
 
 
 
 
 
 
 
 
 


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1. 
BACKGROUND AND RATIONALE 
 
1.1 
Disease Background 
Metastatic melanoma continues to be a challenging disease to treat, with an estimated 9,940 
expected deaths in the United States in 2015 [1].  The 5 year survival rate ranges between 5-19% 
[2] and the 10 year survival is less than 10% [3]. Patients with Stage IV melanoma continue to 
have a poor prognosis, with a mean survival of 8–10 months in large cohort analysis studies [4]. 
The National Comprehensive Cancer Network (NCCN) currently recommends the following 
therapies for treatment of advanced melanoma: 1) immunotherapy with anti-programmed cell 
death protein 1 (PD-1) monotherapy (Pembrolizumab or Nivolumab (NCCN category 1, which 
means it is based on high-level evidence, leading to NCCN consensus that the intervention is 
appropriate), 2) combination immunotherapy with Nivolumab/Ipilimumab, 3) targeted therapy if 
B-Raf proto-oncogene, serine/threonine kinase (BRAF)- mutated, 4) Dabrafenib/Trametinib 
(category 1 and preferred by NCCN), 5) Vemurafenib (category 1), 6) Dabrafenib (category 1) or 
7) enrollment in a clinical trial. Although no longer recommended as a single agent for metastatic 
melanoma, Ipilimumab has been associated with long term survival in approximately 20% of 
patients [5]. The current recommended regimen of ipilimumab/nivolumab, has not provided 
mature data yet on overall survival (OS). Upon disease progression, a selected number of 
patients with Eastern Cooperative Oncology Group ECOG performance status (PS) 0-2 are 
offered the same agents (provided they did not have them before). In this setting (progression 
after initial treatment) High Dose interleukin-2 (HD IL-2) is one of the therapies recommended 
by NCCN. Despite these recent advances in increased options for treatment of advanced 
melanoma the prognosis remains poor for the majority of patients, and improved strategies to 
increase the number of patients benefited with longer and sustained responses are needed. 
 
We propose to test two drugs in combination in order to determine if response rates can be 
improved over either agent as single therapy. The drugs which will be testing in combination are 
pembrolizumab and HD-IL-2. The next two sections will briefly describe the two drugs (their 
mechanism of action and some seminal clinical trials studies), and then we will conclude with 
providing the rationale for 1) combining the two drugs to test in this pilot study, and 2) the trial 
design and treatment sequencing we plan to use. 
 
1.2 
Pembrolizumab: Immune Checkpoint Inhibitor 
Interrupting the PD-1/programmed death-ligand 1 (PD-L1) interaction has emerged as an 
important weapon in immunotherapy [6]. Anti-PD-1 antibodies have been shown to induce 
positive radiological responses and improved survival rates in patients with melanoma. Sustained 
remissions remain low with average PFS still less than 6 months [7]. 
 
Pembrolizumab (formerly known as MK-3475) is a highly selective, humanized monoclonal 
IgG4–kappa isotype antibody against PD-1 that can disrupt the engagement of PD-1 with its 
ligands and impede inhibitory signals in T Cells, with resultant tumor recognition by cytotoxic T 
cells. The anti-tumor activity, side effects and safety of pembrolizumab in patients with 
melanoma was initially studied as part of the large, international, phase 1 KEYNOTE-001 trial. 
 
 
 


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In a pooled analysis of 411 patients with advanced melanoma enrolled in KEYNOTE-001, the 
response rate (RR) was 34% and was maintained in 81% of those patients, the median overall 
survival (OS) was 25.9 months [8] after a median follow-up duration of 18 months.  The 
KEYNOTE-002 study of pembrolizumab versus chemotherapy confirmed the benefit of 
pembrolizumab in patients who had disease progression during or after ipilimumab therapy [9]. 
 
In this open-label, international expansion cohort of a phase 1 trial, patients (aged >= 18 years) 
with advanced melanoma whose disease had progressed after at least two ipilimumab doses were 
randomly assigned to intravenous pembrolizumab at 2 mg/kg every 3 weeks or 10 mg/kg every 3 
weeks until disease progression, intolerable toxicity, or consent withdrawal. A total of 173 
patients received pembrolizumab 2 mg/kg (n=89) or 10 mg/kg (n=84). Median follow-up 
duration was 8 months. ORR was 26% at both doses (21 of 81 patients in the 2 mg/kg group and 
20 of 76 in the 10 mg/kg group) (difference 0%, 95% CI -14 to 13; p=0.96). Treatment was well 
tolerated, with similar safety profiles in both groups and no drug-related deaths. The most 
common drug-related adverse events of any grade in the 2 mg/kg and 10 mg/kg groups were 
fatigue (29 [33%] vs 31 [37%]), pruritus (23 [26%] vs 16 [19%]), and rash (16 [18%] vs 15 
[18%]). Grade 3 fatigue, reported in five (3%) patients in the 2 mg/kg pembrolizumab group, 
was the only drug-related grade 3 to 4 adverse event reported in more than one patient [8]. 
 
Based on these preliminary trials, the landmark KEYNOTE 006 trial [10] was launched and 
ultimately lead to the FDA approval of pembrolizumab for ipilimumab refractory melanoma. In 
this randomized controlled, phase 3 study, 834 patients with advanced melanoma in a 1:1:1 ratio 
were assigned to receive pembrolizumab (at a dose of 10 mg per kilogram of body weight) every 
2 weeks or every 3 weeks or four doses of ipilimumab (at 3 mg per kilogram) every 3 weeks. 
Primary end points were progression-free and overall survival. The estimated 6-month 
progression-free-survival (PFS) rates were 47.3% for pembrolizumab every 2 weeks, 46.4% for 
pembrolizumab every 3 weeks, and 26.5% for ipilimumab (hazard ratio [HR] for disease 
progression, 0.58; P<0.001 for both pembrolizumab regimens versus ipilimumab; 95% 
confidence intervals [CIs], 0.46 to 0.72 and 0.47 to 0.72, respectively). Estimated 12-month 
survival rates were 74.1%, 68.4%, and 58.2%, respectively (hazard ratio [HR] for death for 
pembrolizumab every 2 weeks, 0.63; 95% CI, 0.47 to 0.83; P=0.0005; HR for pembrolizumab 
every 3 weeks, 0.69; 95% CI, 0.52 to 0.90; P=0.0036). The RR was improved with 
pembrolizumab administered every 2 weeks (33.7%) and every 3 weeks (32.9%), as compared 
with ipilimumab (11.9%) (P<0.001 for both comparisons). Responses were ongoing in 89.4%, 
96.7%, and 87.9% of patients, respectively, after a median follow-up of 7.9 months. Efficacy 
was similar in the two pembrolizumab groups. Rates of treatment-related adverse events of grade 
3 to 5 severity were lower in the pembrolizumab groups (13.3% and 10.1%) than in the 
ipilimumab group (19.9%), and the specific adverse events were similar to that observed in the 
KEYNOTE-002 trial. 
 
1.3 
High Dose Interleukin-2 (HD IL-2) 
HD IL-2 has been approved since 1998 for the treatment of stage IV malignant melanoma. 
Interleukin-2 is mostly produced by antigen-stimulated CD4 + T helper cells, and to a lesser 
extent by CD8 + T cells, natural killer (NK) cells, natural killer T (NKT) cells, and activated 
Dendritic Cells [11]. IL-2 induces the proliferation of NK cells augmenting their cytolytic 
capacity, drives proliferation and activation of CD8 + T cells and promotes the proliferation of B 


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cells and antibody secretion [12].  IL-2 was originally seen as the key factor of augmenting an 
effector lymphocyte immune response. However, it also serves as a potent immune regulator by 
expanding immunosuppressive CD4+ FOXP3+ T-regulatory cells (Treg) [13] as well as 
promoting activation-induced cell death (AICD) of overactivated T cells [14]. Hence, rather than 
simply enhancing cytotoxic effects it acts as an immune modulator with different activities 
depending on the target cell and tumor microenvironment. 
 
Studies with high-dose bolus (HDB) IL-2 used doses of 600,000–720,000 U/kg every 8 hours 
from days 1–5 (cycle 1) and 15–19 (cycle 2) with a maximum of 14 doses per cycle or 28 doses 
per course (one course = two cycles). IL-2 was also administered either as a single agent or in 
combination with immunologically active cells: lymphokine-activated killer (LAK) cells and 
tumor-infiltrating lymphocytes (TILs). A meta-analysis [15,16] of all seven National Cancer 
Institute-sponsored trials and the one Chiron Corporation sponsored trial conducted between 
1985 and 1993 (270 patients) on metastatic melanoma patients revealed an overall objective 
response of 16 % (95% confidence interval, 12% to 21%); there were 17 complete responses 
(CRs) (6%) and 26 partial responses (PRs) (10%). Responses occurred with all sites of disease 
and in patient with large tumor burdens. Nineteen of the 43 responding patients (44%) and 11% 
of all patients were alive with survivals as long as 150 months. Twelve of these responding 
patients were continuously disease free. The median response duration was 8.9 months (range, 
4–106+months). Twenty-eight percent of responding patients, including 59% of those patients 
who achieved a complete response remained progression-free at a median follow-up of 62 
months. No patient with an ongoing response at 30 months has relapsed, with follow-up 
extending beyond 20 years in some cases, suggesting that such patients are likely cured [15]. 
However, the major toxicities associated with this regimen, including hypotension, renal 
insufficiency and hypoxia, have precluded its widespread application. The use of HD IL-2 is 
limited to specialized programs with experienced personnel, and it is generally offered only to 
patients with good performance and excellent organ function [17]. However, in contrast with low 
dose IL-2, HD IL-2 has shown the ability to provide durable responses in a subset of patients. 
For this reason, HD IL-2 is selected as the dose to use in this study. 
 
1.4 
Study Rationale 
One of the several identified difficulties with not obtaining better outcomes to checkpoint 
blockade is T-cell exhaustion. T cell exhaustion describes a state of T cell dysfunction that was 
initially observed during chronic lymphocytic choriomeningitis virus (LCMV) infection in mice 
[18]. Exhausted T cells fail to proliferate and exert effector functions such as cytotoxicity and 
cytokine secretion in response to antigen stimulation. Further studies identified that exhausted T 
cells are characterized by sustained expression of the inhibitory molecule PD-1 (programmed 
cell death 1) and that blockade of PD-1 and PD-L1 interactions can reverse T cell exhaustion and 
restore antigen-specific T cell responses in LCMV-infected mice [19]. Several lines of evidence 
also implicate the PD-1–PD-L pathway in T cell exhaustion in cancer [20]. However, targeting 
the PD-1/PD-L1 pathway does not always result in reversal of T cell exhaustion [21]. IL-2 has 
the potential to augment memory T-cells [22] in addition to other possible beneficial effects such 
as increasing NK cells [23]. Administration of IL-2 in combination with anti-PD-L1 blockade in 
mouse models has been shown to be synergistic in greatly increasing antigen-specific CD8 T cell 
numbers and increasing their function [24]. For these reasons we are using both pembrolizumab 


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and IL-2 in the present trial to determine if RR (CR+PR) can be enhanced over either agent 
alone. 
 
1.5 
Rationale for Trial Design and Treatment Sequencing 
Despite advances in our understanding of immunology, we lack data on specific responses of 
cellular subtypes to these different agents and perhaps more pertinent, the interactions of the 
various immunocellular components. The sequencing of the drugs emerges as an important 
potential determinant. An ongoing study of ipilimumab with HD IL-2 is exploring sequencing of 
the agents as well [25] and there is an additional ongoing clinical trial using pembrolizumab, 
lymphodepletion and TIL effectors in combination with high or low dose IL-2 [26]. 
 
Given the limited number of centers and patients that can undertake HD IL-2 therapy, the most 
practical design would be a single arm study of anti-PD1 with HD IL-2 to determine if response 
rates can be improved over either agent as single therapy. Indeed, a recent retrospective study 
[27] using the PROCLAIM registry [28] the largest collection of IL-2 treated patients in the US, 
revealed that the median overall survival (mOS) for the 236 patients with metastatic melanoma 
was 18.4 months with a median follow-up of 21.7 months. Patients were stratified into three 
groups; HD IL-2 only (n=123), HD IL-2 followed by ipilimumab (IL-2→ipi, n=78), and HD IL-
2 followed by PD-1 inhibitors (IL-2→aPD-1, n=35). The majority of patients (22 of 35) in the 
IL-2→aPD-1 group had progressive disease before receiving subsequent treatment with anti-PD-
1/PD-L1-containing regimens. Patients in the HD IL-2 only, IL-2→ipi, and IL-2→aPD-1 groups 
achieved a median OS of 14, 15.8, and 28.7 months, respectively. The estimated 12-month 
survival rates were 57%, 64%, and 97%, respectively. There were 10/78 (13%) and 2/35 (5.7%) 
post therapy treatment-related incidences of autoimmune events in the IL-2→ipi and IL-2→aPD-
1 groups, respectively. This study was the first report of clinical data relating to HD IL-2 use 
followed by checkpoint blockade of the PD-1 pathway. Treatment with anti-PD-1 after initial 
therapy with HD IL-2 had significantly prolonged survival compared to patients treated with 
ipilimumab (28.7 vs 15.8 months). Moreover, improved survival was not observed in patients 
treated with follow-on ipilimumab compared to patients treated only with HD IL-2. Anti-PD-1 
therapy after HD IL-2 appears to be safe and therapeutically active in this retrospective study 
[27]. Given that anti PD-1 treatment can result in activation of T-cell cytotoxic effectors by 
partially reversing T-cell exhaustion, it seems likely that the amplitude of this cytotoxic response 
can be greatly augmented by adding HD IL-2 resulting in increased tumor destruction and in the 
generation of a durable T-Cell Memory effector population which may result in a sustained and 
prolonged remission. 
 
 
 


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2. 
STUDY OBJECTIVES AND ENDPOINTS 
 
2.1 
Objectives 
 
2.1.1 Primary Objective 
Assess the response rate [complete response (CR) + partial response (PR)] of sequential therapy 
of pembrolizumab followed by HD IL-2 in subjects with stage IV malignant melanoma. 
Response assessment will be performed using revised RECIST 1.1 [29]. 
 
2.1.2 Secondary Objectives 
• Characterize safety, tolerability and adverse effect (AE) profile of pembrolizumab 
followed by HD IL-2 in subjects with stage IV malignant melanoma. 
• Measure Progression-Free Survival (PFS) using revised RECIST guideline (version 1.1) 
after completion of 2 cycles of pembrolizumab and 2 cycles of HD IL-2 in all the subjects 
enrolled in the study. 
• Measure overall survival (OS) at 12 months in subjects with stage IV malignant 
melanoma who showed a response [stable disease (SD), complete response (CR) or 
partial response (PR)] following completion of 2 cycles of pembrolizumab and 2 cycles 
of HD IL-2. 
 
2.1.3 Correlative/Exploratory Objectives 
• Correlate PD-L1 expression in archived diagnostic tumor tissue with best clinical 
response (revised RECIST 1.1 criteria) and treatment outcome in subjects with stage IV 
malignant melanoma. 
• Correlate Th1/Th2 ratio and myeloid-derived suppressor cells (MDSCs) in peripheral 
blood during therapy with best clinical response (revised RECIST criteria) and treatment 
outcome in subjects with stage IV malignant melanoma. Data will be collected prior to 
treatment, during treatment after the 2nd dose of pembrolizumab and after each cycle of 
HD IL-2. 
• Correlate the change in soluble PD-L1 and arginine level in plasma during therapy with 
best clinical response (revised RECIST1.1 criteria) and treatment outcome in subjects 
with stage IV malignant melanoma. Data will be collected prior to treatment, during 
treatment after the 2nd dose of pembrolizumab and after each cycle of HD IL-2. 
• Correlate intestinal microbiome at baseline with best clinical response (revised RECIST 
1.1 criteria) and treatment outcome in subjects with stage IV malignant melanoma.  
 
2.2 
Endpoints 
 
2.2.1 Primary Endpoint 
Assess the response rate [complete response (CR) + partial response (PR)] of sequential therapy 
of pembrolizumab followed by HD IL-2 in subjects with stage IV malignant melanoma, using 
revised RECIST 1.1. Response rate will be computed with associated 95% confidence intervals. 
 


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2.2.2 Secondary Endpoints 
• Proportion of subjects with each grade of adverse events as defined by CTCAE v4.03 
will be computed along with 95% confidence intervals, and reported in a tabular and 
descriptive manner. 
• Median PFS times will be computed, and PFS rate at 1 year +/- 3 months will be 
calculated with associated 95% confidence intervals based on the Kaplan-Meier method.  
Median OS times will be computed, and OS rate at 1 year +/- 3 months will be calculated 
with associated 95% confidence intervals based on the Kaplan-Meier method.   
 
3. 
ELIGIBILITY CRITERIA 
 
3.1 
Inclusion Criteria 
Subject must meet all of the following applicable inclusion criteria to participate in this study: 
 
1. Written informed consent and HIPAA authorization for release of personal health 
information prior to registration. NOTE:  HIPAA authorization may be included in the 
informed consent or obtained separately. 
 
2. Age ≥ 18 years at the time of consent. 
 
3. ECOG Performance Status of 0-1 within 28 days prior to registration (Appendix 1). 
 
4. Life expectancy of 6 months or greater as determined by the site investigator. 
 
5. Histologically-confirmed diagnosis of unresectable stage IV or metastatic melanoma not 
amenable to local therapy. 
 
6. Measurable disease, defined as at least 1 tumor that fulfills the criteria for a target lesion 
according to RECIST 1.1 (Section 9), and obtained by imaging within 28 days prior to 
registration for protocol therapy. 
 
7. Archival tissue (from the primary tumor or metastases) is mandatory if available for 
correlative studies. If archived tumor tissue is not available, the patient does not need to 
undergo a biopsy to obtain tissue and is still eligible for study. 
 
8. < 2 lines of prior therapy for metastatic melanoma. Cannot have received prior therapy 
with HD IL-2. May have had one prior line of therapy that included a check point 
inhibitor.  
 
9. Prior systemic cancer treatment must be completed at least 21 days prior to first dose of 
study drug and the subject must have recovered from all reversible acute toxic effects of 
the regimen (other than alopecia or vitiligo) to Grade ≤ 1 or baseline. 
 
10. Not received radiation therapy within 21 days of initiation of study treatment, and the 
measurable disease must have been outside of the radiation port. 
 


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11. Demonstrate adequate organ function as defined in the table below. All screening labs to 
be obtained within 28 days prior to registration. 
 
System 
Laboratory Value 
Hematological 
White blood cell (WBC) 
≥ 3,000/L 
Absolute Neutrophil Count (ANC) 
≥ 1,000/L without the support of filgrastim 
Hemoglobin (Hgb) 
≥ 9 g/dL 
Platelet Count 
≥ 100 × 109/L 
Renal 
Serum creatinine 
Calculated creatinine clearance1 
≤ 1.5 mg/dL  
≥ 40 mL/min; for subjects with serum 
creatinine > 1.5 mg/dL 
Hepatic 
Total Bilirubin2 OR 
Direct Bilirubin 
≤ 1.5 × upper limit of normal (ULN) 
≤ ULN for subjects with total bilirubin levels 
> 1.5 x ULN 
Aspartate aminotransferase (AST) 
≤ 2.5 × ULN or ≤ 5 x ULN for subjects with 
known hepatic metastasis 
Alanine aminotransferase (ALT) 
≤ 2.5 × ULN or ≤ 5 x ULN for subjects with 
known hepatic metastasis 
Coagulation 
International Normalized Ratio (INR) or  
 
≤ 1.5 × ULN; For subjects receiving 
warfarin or LMWH, the subjects must, in the 
site investigator’s opinion, be clinically 
stable with no evidence of active bleeding 
while receiving anticoagulant therapy. The 
INR for these subjects may exceed 1.5 × 
ULN if that is the goal of anticoagulant 
therapy. 
1: Cockcroft-Gault formula will be used to calculate creatinine clearance (See SPM) 
 
2: Except in patients with Gilbert’s syndrome who must have a total bilirubin less than 3.0 mg/dl. 
 
12. Adequate baseline pulmonary function test (PFT) (FEV1 > 2 L or ≥ 75% of predicted for 
height and age). 
 
13. Documented left ventricular ejection fraction (LVEF) of > 45%, testing is required in 
patients with: 
• Age ≥ 60 years old 
• Clinically significant atrial and or ventricular arrhythmias including but not limited 
to: atrial fibrillation, ventricular tachycardia, second or third degree heart block 
• History of coronary revascularization or ischemic symptoms 
 
14. Females of childbearing potential must have a negative serum pregnancy test within 3 
days prior to registration. NOTE: Females are considered of child bearing potential 
unless they are surgically sterile (have undergone a hysterectomy, bilateral tubal ligation, 
or bilateral oophorectomy) or they are naturally postmenopausal for at least 12 
consecutive months. 


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15. Females of childbearing potential and males must be willing to abstain from heterosexual 
activity or to use 2 forms of effective methods of contraception from the time of informed 
consent until 120 days after treatment discontinuation. The two contraception methods 
can be comprised of two barrier methods, or a barrier method plus a hormonal method. 
 
16. Male subjects who are not surgically sterile (vasectomy) must agree to use an adequate 
method of contraception. Male subjects with female sexual partners who are pregnant, 
possibly pregnant or who could become pregnant during the study must agree to use 
condoms from the first dose of study drug through 120 days after the last dose of study 
therapy. Total abstinence for the same study period is an acceptable alternative. 
 
17. As determined by the enrolling physician or protocol designee, ability of the subject to 
understand and comply with study procedures for the entire length of the study 
 
18. Willingness and ability to comply with scheduled visits, treatment plans, laboratory tests, 
and other study procedures. 
 
3.2 
Exclusion Criteria 
Subjects meeting any of the criteria below may not participate in the study: 
 
1. Active infection requiring systemic therapy 
 
2. Pregnant or breastfeeding. NOTE: breast milk cannot be stored for future use while the 
mother is being treated on study. 
 
3. Known additional malignancy that is active and/or progressive requiring treatment; 
exceptions include basal cell or squamous cell skin cancer, in situ cervical or bladder 
cancer, or other cancer for which the subject has been disease-free for at least five years. 
 
4. Active central nervous system (CNS) metastases. NOTE: if prior metastasis but treated 
and clinically stable for 1 month after treatment are eligible. Subjects with 3 or fewer 
brain metastases that are less than 1 cm in diameter and asymptomatic are eligible. 
 
5. Surgery within 4 weeks prior to study treatment except for minor procedures. NOTE: 
Hepatic biliary stent placement is allowed.  
 
6. Uncontrolled or poorly-controlled hypertension (> 160 mmHg systolic or > 100 mmHg 
diastolic for > 4 weeks) despite standard medical management. 
 
7. Serious or non-healing wound, ulcer, or bone fracture within 28 days prior to initiation of 
study treatment. 
 
8. Any Grade 3-4 GI bleeding within 3 months prior to initiation of study treatment. 
 
 
 


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9. History of deep vein thrombosis, pulmonary embolism, or any other significant 
thromboembolism (venous port or catheter thrombosis or superficial venous thrombosis 
are not considered “significant”) during the 3 months prior to initiation of study 
treatment. 
 
10. Any arterial thromboembolic events, including but not limited to myocardial infarction, 
transient ischemic attack, cerebrovascular accident, or unstable angina, within 6 months 
prior to initiation of study treatment. 
 
11. Gross hemoptysis within 2 months of initiation of study treatment. 
 
12. Has any condition that, in the opinion of the investigator, might jeopardize the safety of 
the patient or interfere with protocol compliance. 
 
13. Has any mental or medical condition that prevents the patient from giving informed 
consent or participating in the trial. 
 
14. Known hypersensitivity to pembrolizumab or IL-2 or any of their components. 
 
15. Known history of active tuberculosis.  
 
16. Concurrent systemic steroid therapy with doses above physiologic level. 
 
17. History of autoimmune disease, including but not limited to myasthenia gravis, myositis, 
autoimmune hepatitis, systemic lupus erythematosus, rheumatoid arthritis, inflammatory 
bowel disease, vascular thrombosis associated with anti-phospholipid syndrome, 
granulomatosis with polyangiitis, Sjögren’s syndrome, Guillain-Barré syndrome, multiple 
sclerosis, vasculitis, or glomerulonephritis.  See Study Procedures Manual (SPM) for a 
detailed list. 
• Subjects with a history of autoimmune-related hypothyroidism on a stable dose of 
thyroid replacement hormone may be eligible for this study. 
• Subjects with controlled Type I diabetes mellitus on a stable dose of insulin 
regimen may be eligible for this study. 
• Subjects with a history of celiac disease may be eligible if controlled with diet. 
 
18. Treatment with any investigational agent within 21 days prior to initiation of study 
treatment and the subject must have recovered from the acute toxic effects of the 
regimen. 
 
19. Prior organ transplant including allogeneic transplantation. 
 
3.3 
Selection of Subjects 
Study entry is open to adults regardless of gender or ethnic background.  While there will be 
every effort to seek out and include women and minorities, the patient population is expected to 
be no different than that of other advanced metastatic melanoma (cutaneous, unresectable) 
cancer studies at institutions that have experience in administration of HD IL-2. 


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4. 
SUBJECT REGISTRATION 
All subjects must be registered through HCRN’s electronic data capture (EDC) system. A subject 
is considered registered when an “On Study” date is entered into the EDC. 
 
Subjects must be registered prior to starting protocol therapy. Subjects must begin therapy 
within 7 business days of registration.  
 
5. 
TREATMENT PLAN 
The primary objective of this single arm phase 2 trial is to assess the response rate [complete 
response (CR) + partial response (PR)] of sequential therapy of pembrolizumab followed by HD 
IL-2 in subjects with stage IV malignant melanoma. Response assessment will be performed 
after pembrolizumab therapy, and response reassessment will be performed after HD IL-2 
therapy using revised RECIST 1.1.  
 
5.1 
Pembrolizumab Administration 
Subjects will receive 200 mg pembrolizumab every 3 weeks for two cycles. Cycle length is 21 
days (i.e., 3 weeks). On the last day of Cycle 2 (+ 3 days), disease status will be assessed via 
imaging. Following the pembrolizumab treatment, HD IL-2 treatment will commence. 
 
Correlative research analyses during pembrolizumab administration include examining the 
relationship between clinical response at the end of Cycle 2 with PD-L1 expression of the 
archived diagnostic tumor tissue. Research analyses also include examining the relationship 
between changes in soluble PD-L1 level and Type 1 T helper cell/Type 2 T helper cell (Th1/Th2) 
ratios during therapy (assessed after the 2nd dose) relative to baseline and characterization of 
intestinal microbiome to the best clinical response (RECIST 1.1 criteria) and to treatment 
outcome.  
 
Drug 
Dose 
Route 
Schedule 
Cycle 
Length 
Pembrolizumab 200 mg 
Intravenously (IV) over  
30 minutes 
Day 1 of each cycle 
3 weeks 
(21 days) 
1 A window of - 1 or + 2 days may be applied to all study visits to accommodate observed holidays, inclement 
weather, scheduling conflicts etc. Date and time of each drug administration should be clearly documented in 
subject’s chart and electronic case report forms (eCRFs). 
 
5.1.1 Pre-Medication for Pembrolizumab Administration 
Premedication will be given at the discretion of the site investigator. For patients who experience 
a Grade 1 or Grade 2 infusion reaction, acetaminophen 650 mg PO and/or 50 mg IV ranitidine 
(or equivalent H2 antagonist) may be considered prior to subsequent infusions. 
 
5.1.2 Supportive Care for Pembrolizumab 
Subjects should receive full supportive care during the study, including transfusion of blood and 
blood products, treatment with antibiotics, analgesics, erythropoietin, or bisphosphonates, when 
appropriate.  
 
Although acetaminophen at doses of ≤ 2 grams/day is permitted, it should be used with caution 
in subjects with impaired liver function.  


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Subjects should receive appropriate supportive care measures as deemed necessary by the 
treating investigator. Suggested supportive care measures for the management of adverse events 
that are related to pembrolizumab are outlined below. Where appropriate, these guidelines 
include the use of oral or intravenous treatment with corticosteroids as well as additional anti-
inflammatory agents if symptoms do not improve with administration of corticosteroids. Note 
that several courses of steroid tapering may be necessary as symptoms may worsen when the 
steroid dose is decreased. For each disorder, attempts should be made to rule out other causes 
such as metastatic disease or bacterial or viral infection, which might require additional 
supportive care. The treatment guidelines are intended to be applied when the investigator 
determines the events to be related to pembrolizumab. 
 
Pneumonitis 
• For Grade 2 events, treat with systemic corticosteroids. When symptoms improve to 
Grade 1 or less, steroid taper should be started and continued over no less than 4 weeks.  
• For Grade 3-4 events, immediately treat with intravenous steroids. Administer additional 
anti-inflammatory measures, as needed.  
• Add prophylactic antibiotics for opportunistic infections in the case of prolonged steroid 
administration.  
• For guidelines on treatment holds, timing for restarting treatment or discontinuing 
treatment permanently, see Section 6.1.3, Table 2. 
 
Diarrhea/Colitis 
• Subjects should be carefully monitored for signs and symptoms of enterocolitis (such as 
diarrhea, abdominal pain, blood or mucus in stool, with or without fever) and of bowel 
perforation (such as peritoneal signs and ileus).  
• All subjects who experience diarrhea/colitis should be advised to drink liberal quantities 
of clear fluids. If sufficient oral fluid intake is not feasible, fluid and electrolytes should 
be substituted via IV infusion. For Grade 2 or higher diarrhea, consider GI consultation 
and endoscopy to confirm or rule out colitis.  
• For Grade 2 diarrhea/colitis that persists greater than 3 days, administer oral 
corticosteroids.  
• For Grade 3 or 4 diarrhea/colitis that persists > 1 week, treat with intravenous steroids 
followed by high dose oral steroids.  
• When symptoms improve to Grade 1 or less, steroid taper should be started and 
continued over no less than 4 weeks.  
• For guidelines on treatment holds, timing for restarting treatment or discontinuing 
treatment permanently, see Section 6.1.3, Table 2. 
 
Type 1 diabetes mellitus (if new onset, including diabetic ketoacidosis [DKA]) or ≥ Grade 3 
Hyperglycemia, if associated with ketosis (ketonuria) or metabolic acidosis  
• For T1DM or Grade 3-4 Hyperglycemia  
o Insulin replacement therapy is recommended for Type I diabetes mellitus and for 
Grade 3-4 hyperglycemia associated with metabolic acidosis or ketonuria.  
o Evaluate patients with serum glucose and a metabolic panel, urine ketones, 
glycosylated hemoglobin, and C-peptide.  
 


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Hypophysitis  
• For Grade 2 events, treat with corticosteroids. When symptoms improve to Grade 1 or 
less, steroid taper should be started and continued over no less than 4 weeks. 
Replacement of appropriate hormones may be required as the steroid dose is tapered.  
• For Grade 3-4 events, treat with an initial dose of IV corticosteroids followed by oral 
corticosteroids. When symptoms improve to Grade 1 or less, steroid taper should be 
started and continued over no less than 4 weeks. Replacement of appropriate hormones 
may be required as the steroid dose is tapered. 
• For guidelines on treatment holds, timing for restarting treatment or discontinuing 
treatment permanently, see Section 6.1.3, Table 2. 
 
Hyperthyroidism or Hypothyroidism  
Thyroid disorders can occur at any time during treatment. Monitor patients for changes in 
thyroid function (at the start of treatment, periodically during treatment, and as indicated 
based on clinical evaluation) and for clinical signs and symptoms of thyroid disorders.  
 
• Grade 2 hyperthyroidism events (and Grade 2-4 hypothyroidism):  
o In hyperthyroidism, non-selective beta-blockers (e.g. propranolol) are suggested as 
initial therapy.  
o In hypothyroidism, thyroid hormone replacement therapy, with levothyroxine or 
liothyroinine, is indicated per standard of care.  
• Grade 3-4 hyperthyroidism  
o Treat with an initial dose of IV corticosteroid followed by oral corticosteroids. When 
symptoms improve to Grade 1 or less, steroid taper should be started and continued 
over no less than 4 weeks. Replacement of appropriate hormones may be required as 
the steroid dose is tapered.  
o For guidelines on treatment holds, timing for restarting treatment or discontinuing 
treatment permanently, see Section 6.1.3, Table 2. 
 
Hepatic 
• For Grade 2 events, monitor liver function tests more frequently until returned to 
baseline values (consider weekly).  
o Treat with IV or oral corticosteroids  
• For Grade 3-4 events, treat with intravenous corticosteroids for 24 to 48 hours.  
o When symptoms improve to Grade 1 or less, a steroid taper should be started and 
continued over no less than 4 weeks.  
o For guidelines on treatment holds, timing for restarting treatment or discontinuing 
treatment permanently, see Section 6.1.3, Table 2. 
Renal Failure or Nephritis  
• For Grade 2 events, treat with corticosteroids.  
• For Grade 3-4 events, treat with systemic corticosteroids.  
o When symptoms improve to Grade 1 or less, steroid taper should be started and 
continued over no less than 4 weeks.  
o For guidelines on treatment holds, timing for restarting treatment or discontinuing 
treatment permanently, see Section 6.1.3, Table 2. 
 


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Management of Infusion Reactions  
• Signs and symptoms usually develop during or shortly after drug infusion and generally 
resolve completely within 24 hours of completion of infusion.  
• See Table 1 in Section 6.1.2, Management of Allergic Reaction/Hypersensitivity, for 
detailed information on how Grades 1-4 reactions are treated.  
 
5.2 
HD IL-2 Administration 
After completing the 2 cycles of pembrolizumab, High Dose Interleukin-2 (HD IL-2) will be 
administered. This will require a hospital stay of at least 5 days for each treatment cycle. 
Established guidelines will be followed for safely administering HD-IL-2 and managing 
toxicities from this treatment [31,32]. Two treatment cycles will be administered, Cycle 2 being 
separated from Cycle 1 by approximately 9 days after completion (assuming subject has 
recovered from Cycle 1 adequately to proceed with Cycle 2). Four weeks after completion of the 
2 cycles (called a course of HD IL-2 therapy), disease status will be monitored via CT of the 
chest and abdomen/pelvis using revised RECIST guidelines (version v1.1, Section 9 of protocol).  
 
Correlative research analyses in this phase include examining the relationship between changes 
in soluble PD-L1 level and Type 1 T helper cell/Type 2 T helper cell (Th1/Th2) ratios assessed 
after each of the two HD IL-2 cycles relative to baseline (i.e., pre-dose on Day 1 of Cycle 1 of 
pembrolizumab treatment), to the best clinical response (modified RECIST criteria) and to 
treatment outcome. 
 
Drug 
Dose 
Route 
Schedule 
Cycle Length 
IL-2 
600,000 
IU/kg2 
Intravenously (IV) 
over 15 minutes 
Every 8 hours for 
up to 14 doses over 
5 days   
Days 1-5 = Cycle 1 
9 days of rest in between 
Days 15-19 = Cycle 2 
1 A window of - 1 or + 2 days may be applied to all study visits to accommodate observed holidays, inclement 
weather, scheduling conflicts etc. Date and time of each drug administration should be clearly documented in 
subject’s chart and electronic case report forms (eCRFs). 
2Actual weight will be used unless the patient is 30% over ideal body weight in which case adjusted body 
weight will be used (Males: IBW = 50 kg + 2.3 kg for each inch over 5 feet. 
Females: IBW = 45.5 kg + 2.3 kg for each inch over 5 feet); ABW = IBW + 0.4(actual weight - IBW) 
 
5.2.1 Supportive Care specific to HD IL-2 
Subjects should receive appropriate supportive care measures as deemed necessary by the site 
investigator. Several treatment-related toxicities have been uniquely associated with the 
administration of HD IL-2. Suggested management are listed in the Study Procedures Manual 
(SPM). 
 
5.3 
Concomitant Medications 
Medications or vaccinations specifically prohibited in the exclusion criteria are not allowed 
during the ongoing trial. If there is a clinical indication for one of these or other medications or 
vaccinations specifically prohibited during the trial, discontinuation from trial therapy or 
vaccination may be required. The final decision on any supportive therapy or vaccination rests 
with the site investigator and/or the subject's primary physician. 
 


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5.3.1 Allowed Concomitant Medications 
All treatments that the investigator considers necessary for a subject’s welfare may be 
administered at the discretion of the investigator in keeping with the community standards of 
medical care. All concomitant medications will be recorded on the electronic case report form 
(eCRF). 
  
All concomitant medications received within 28 days before the first dose of trial treatment and 
30 days after the last dose of trial treatment should be recorded. Concomitant medications 
administered beyond 30 days after the last dose of trial treatment should be recorded for SAEs 
and ECIs. 
 
Myeloid growth factors to treat subjects with neutropenia according to the American Society of 
Clinical Oncology (ASCO) Guidelines are permitted. Myeloid growth factors should be avoided 
(if medically appropriate) in Cycle 1 until subjects have developed a dose-limiting Grade 4 
neutropenia.  
 
Antiemetic agents may be administered at the discretion of the site investigator but are not 
commonly required as a prophylactic agent. All other manifestations of the subject’s malignancy 
should be treated at the discretion of the investigator.  
 
Medications with potential central nervous system (CNS) effects are not prohibited in this study, 
but it is recommended that their use be minimized to avoid confusion in the interpretation of 
CNS effects should they occur during the course of treatment with pembrolizumab. 
 
In appropriate settings, such as combinations with agents known to produce frequent 
thrombocytopenia, restricted uses of anticoagulants should be considered.  
 
All other medical conditions should be treated at the discretion of the site investigator in 
accordance with local community standards of medical care. 
 
5.3.2 Prohibited Concomitant Medications 
Subjects are prohibited from receiving the following therapies during the Screening and 
Pembrolizumab Treatment Phase of this trial: 
• Anti-cancer systemic chemotherapy or biological therapy not specified in this protocol 
• Immunotherapy not specified in this protocol 
• Concomitant use of chemotherapy  
• Investigational agents  
• Radiation therapy  
▪ Note: Radiation therapy to a symptomatic solitary lesion or to the brain may be allowed 
after consultation with study chair.  
• Live vaccines within 30 days prior to the first dose of trial treatment and while 
participating in the trial. Examples of live vaccines include, but are not limited to, the 
following: measles, mumps, rubella, chicken pox, yellow fever, rabies, BCG, and typhoid 
(oral) vaccine. Seasonal influenza vaccines for injection are generally killed virus 
vaccines and are allowed; however, intranasal influenza vaccines (e.g. Flu-Mist®) are live 
attenuated vaccines, and are not allowed.  


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• Systemic glucocorticoids for any purpose other than to modulate symptoms from an 
event of clinical interest of suspected immunologic etiology. The use of physiologic 
doses of corticosteroids may be approved after consultation with the sponsor-investigator.  
 
Subjects who, in the assessment by the site investigator, require the use of any of the 
aforementioned treatments for clinical management should be removed from the trial. 
 
5.4 
Contraception 
It is not known if pembrolizumab has adverse effects on a fetus in utero nor if there are transient 
adverse effects on the composition of sperm.  Non-pregnant, non-breast-feeding women may be 
enrolled if they are willing to use 2 methods of birth control or are considered highly unlikely to 
conceive. Highly unlikely to conceive is defined as 1) surgically sterilized, or 2) postmenopausal 
(a woman who is ≥45 years of age and has not had menses for greater than 1 year will be 
considered postmenopausal), or 3) not heterosexually active for the duration of the study. The 
two birth control methods can be either two barrier methods or a barrier method plus a hormonal 
method to prevent pregnancy. Subjects should start using birth control from time of informed 
consent throughout the study period and up to 120 days after the last dose of study drug.  
 
The following are considered adequate barrier methods of contraception: diaphragm, condom (by 
the partner), copper intrauterine device, sponge, or spermicide. Appropriate hormonal 
contraceptives will include any registered and marketed contraceptive agent that contains an 
estrogen and/or a progestational agent (including oral, subcutaneous, intrauterine, or 
intramuscular agents). 
 
Subjects should be informed that taking the study medication may involve unknown risks to the 
fetus (unborn baby) if pregnancy were to occur during the study. To participate in the study, they 
must adhere to the contraception requirements described above for the duration of the study and 
120 days following the last dose of study drug. If there is any question that a subject will not 
reliably comply with the requirements for contraception, that subject should not be entered into 
the study. 
 
Male subjects who are not surgically sterile (vasectomy) must agree to use an adequate method 
of contraception. Male subjects with female sexual partners who are pregnant, possibly pregnant 
or who could become pregnant during the study must agree to use condoms from the time of 
informed consent through 120 days after the last dose of study therapy. Total abstinence for the 
same study period is an acceptable alternative. 
 
5.5 
Use in Pregnancy 
If a subject inadvertently becomes pregnant while on study treatment, the subject will 
immediately be removed from the study. The site will contact the subject at least monthly and 
document the subject’s status until the pregnancy has been completed or terminated. The 
outcome of the pregnancy will be reported to Hoosier Cancer Research Network (HCRN) within 
1 business day if the outcome is a serious adverse experience (e.g., death, abortion, congenital 
anomaly, or other disabling or life-threatening complication to the mother or newborn). HCRN 
will report this information within 1 business day to Prometheus. The study investigator will 
make every effort to obtain permission to follow the outcome of the pregnancy and report the 


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condition of the fetus or newborn to HCRN. If a male subject impregnates his female partner, he 
must immediately inform the site study personnel and the pregnancy reported and followed as 
described above. 
 
5.6 
Breastfeeding 
It is unknown whether pembrolizumab or IL-2 is excreted in human milk. Since many drugs are 
excreted in human milk, and because of the potential for serious adverse reactions in the nursing 
infant, subjects who are breast-feeding are not eligible for enrollment. 
 
6. 
TOXICITIES AND DOSE DELAYS/DOSE MODIFICATIONS 
The NCI Common Terminology Criteria for Adverse Events (CTCAE) v4.03 will be used to 
grade adverse events.  
 
Subjects enrolled in this study will be evaluated clinically and with standard laboratory tests 
before and at regular intervals during their participation in this study as specified in Study 
Calendar & Evaluations. 
 
Subjects will be evaluated for adverse events (all grades), serious adverse events, and adverse 
events requiring study drug interruption or discontinuation as specified in Study Calendar & 
Evaluations.   
 
6.1 
Dose Delays for Pembrolizumab Administration  
 
6.1.1 Start of a New Cycle 
A new treatment cycle will only be initiated when all of the following conditions are met: 
• ANC ≥ 1,500× 109/L  
• platelets ≥ 100 × 109/L  
• Non-hematologic treatment related toxicities have improved to ≤ Grade 1 or to the 
subject’s baseline values (except alopecia) 
 
If blood counts (first two bullet points above) are below this threshold, blood work is to be 
repeated weekly until counts are at an acceptable level. If treatment is unable to restart within 12 
weeks of the planned treatment date, the subject will be permanently discontinued from study 
therapy.   
 
 
 


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6.1.2 Management of Hypersensitivity Reaction 
 
Table 1: Management of allergic reaction/hypersensitivity based on CTCAE Grade 
 
Description 
Action 
Grade I Allergic 
Reaction/Hypersensitivity: (Transient 
flushing or rash, drug fever < 38°C):  
Supervise without further treatment for 
allergic reaction/hypersensitivity. 
Grade 2 Hypersensitivity Reaction: 
(Rash, flushing, dyspnea, urticaria, drug 
fever greater than or equal to 38°C) and/or 
asymptomatic bronchospasm: 
Interrupt the infusion. If symptoms 
abate, attempt re-infusion at a slower 
rate. If the symptoms recur, discontinue 
infusion and follow for recurrent 
allergic reaction/hypersensitivity in the 
next paragraph. 
Recurrent Grade 2 or Grade 3 or 4 
Hypersensitivity Reaction: 
Stop the infusion. Administer 
additional doses of H1 and H2 blockers 
intravenously. Administer IV steroids 
and consider epinephrine and 
bronchodilators as clinically indicated. 
Grade 3 or 4 Hypersensitivity Reaction: 
Will be permanently discontinued from 
study treatment. 
 
Prior to re-challenge of Grade 2 allergic reaction/HSR and if there is a subsequent cycle, the 
following prophylactic premedications are recommended prior to the pembrolizumab infusion: 
give both an H1 and H2 blocker intravenously within 30 to 60 minutes before pembrolizumab 
infusion.  If treatment delay necessitates a period longer than 12 weeks, treatment is stopped and 
the subject is discontinued from the study. 
 
6.1.3 Dose Modification for Toxicity 
Adverse events (both non-serious and serious) associated with pembrolizumab exposure may 
represent an immunologic etiology. These adverse events may occur shortly after the first dose 
or several months after the last dose of treatment. Pembrolizumab must be withheld for drug-
related toxicities and severe or life-threatening AEs as per Table 2 below.  


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Table 2: Dose Modification and Toxicity Management Guidelines for Immune-related AEs Associated with Pembrolizumab 
General instructions: 
1. Corticosteroid taper should be initiated upon AE improving to Grade 1 or less and continue to taper over at least 4 weeks. 
2. For situations where pembrolizumab has been withheld, pembrolizumab can be resumed after AE has been reduced to Grade 1 or 0 and 
corticosteroid has been tapered. Pembrolizumab should be permanently discontinued if AE does not resolve within 12 weeks of last dose or 
corticosteroids cannot be reduced to ≤10 mg prednisone or equivalent per day within 12 weeks.  
3. For severe and life-threatening irAEs, IV corticosteroid should be initiated first followed by oral steroid. Other immunosuppressive treatment 
should be initiated if irAEs cannot be controlled by corticosteroids. 
 
Immune-related 
AEs 
Toxicity grade or 
conditions 
(CTCAEv4.0) 
Action taken to 
pembrolizumab 
irAE management with 
corticosteroid and/or other 
therapies  
Monitor and follow-up  
Pneumonitis  
Grade 2 
Withhold  
• Administer corticosteroids (initial 
dose of 1-2 mg/kg prednisone or 
equivalent) followed by taper 
 
• 
Monitor participants for signs and 
symptoms of pneumonitis 
• 
Evaluate participants with suspected 
pneumonitis with radiographic imaging and 
initiate corticosteroid treatment 
• 
Add prophylactic antibiotics for 
opportunistic infections 
Grade 3 or 4, or 
recurrent Grade 2 
Permanently 
discontinue 
Diarrhea / Colitis  Grade 2 or 3 
Withhold 
• Administer corticosteroids (initial 
dose of 1-2 mg/kg prednisone or 
equivalent) followed by taper 
 
• 
Monitor participants for signs and symptoms 
of enterocolitis (ie, diarrhea, abdominal 
pain, blood or mucus in stool with or 
without fever) and of bowel perforation 
(ie, peritoneal signs and ileus). 
• 
Participants with ≥ Grade 2 diarrhea 
suspecting colitis should consider GI 
consultation and performing endoscopy to 
rule out colitis.  
• 
Participants with diarrhea/colitis should be 
advised to drink liberal quantities of clear 
fluids.  If sufficient oral fluid intake is not 
feasible, fluid and electrolytes should be 
substituted via IV infusion. 
Grade 4 
Permanently 
discontinue 


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AST / ALT 
elevation or 
Increased 
bilirubin 
Grade 2 
Withhold 
• Administer corticosteroids (initial 
dose of 0.5- 1 mg/kg prednisone or 
equivalent) followed by taper 
• 
Monitor with liver function tests (consider 
weekly or more frequently until liver 
enzyme value returned to baseline or is 
stable 
Grade 3 or 4  
Permanently 
discontinue 
• Administer corticosteroids (initial 
dose of 1-2 mg/kg prednisone or 
equivalent) followed by taper 
Type 1 diabetes 
mellitus (T1DM) 
or 
Hyperglycemia 
 
Newly onset 
T1DM or  
Grade 3 or 4 
hyperglycemia 
associated with 
evidence of -cell 
failure 
Withhold 
• Initiate insulin replacement 
therapy for participants with 
T1DM  
• Administer anti-hyperglycemic in 
participants with hyperglycemia  
 
• 
Monitor participants for hyperglycemia or 
other signs and symptoms of diabetes. 
Hypophysitis 
Grade 2 
Withhold 
• Administer corticosteroids and 
initiate hormonal replacements as 
clinically indicated.   
 
• 
Monitor for signs and symptoms of 
hypophysitis (including hypopituitarism and 
adrenal insufficiency)  
Grade 3 or 4  
Withhold or 
permanently 
discontinue1  
Hyperthyroidism  
Grade 2 
Continue  
• Treat with non-selective beta-
blockers (eg, propranolol) or 
thionamides as appropriate  
• 
Monitor for signs and symptoms of thyroid 
disorders.  
 
Grade 3 or 4  
Withhold or  
permanently 
discontinue1 
Hypothyroidism 
Grade 2-4 
Continue  
• Initiate thyroid replacement 
hormones (eg, levothyroxine or 
liothyroinine) per standard of care 
• 
Monitor for signs and symptoms of thyroid 
disorders.  
 
Nephritis and 
Renal 
dysfunction 
Grade 2 
Withhold  
• Administer corticosteroids 
(prednisone 1-2 mg/kg or 
equivalent) followed by taper. 
• 
Monitor changes of renal function 
 
Grade 3 or 4  
Permanently 
discontinue 
Myocarditis 
Grade 1 or 2 
Withhold 
• Based on severity of AE 
administer corticosteroids 
• 
Ensure adequate evaluation to confirm 
etiology and/or exclude other causes 
 
Grade 3 or 4 
Permanently 
discontinue 


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All other 
immune-related 
AEs 
Intolerable/ 
persistent Grade 2 
Withhold 
• Based on type and severity of AE 
administer corticosteroids  
• 
Ensure adequate evaluation to confirm 
etiology and/or exclude other causes 
 
Grade 3 
Withhold or 
discontinue based 
on the type of 
event.  Events that 
require 
discontinuation 
include and not 
limited to:  
Gullain-Barre 
Syndrome, 
encephalitis 
Grade 4 or 
recurrent Grade 3  
Permanently 
discontinue 
1. Withhold or permanently discontinue pembrolizumab is at the discretion of the investigator or treating physician.   
NOTE:  
For participants with Grade 3 or 4 immune-related endocrinopathy where withhold of pembrolizumab is required, pembrolizumab may be resumed 
when AE resolves to ≤ Grade 2 and is controlled with hormonal replacement therapy or achieved metabolic control (in case of T1DM).   
 
 


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6.2 
Dose Delays for HD IL-2 
 
6.2.1 Start of New Cycle 
A new treatment cycle will only be initiated when all of the following conditions are met: 
• 
ANC ≥ 1,500× 109/L  
• 
Platelets ≥ 100 × 109/L  
• 
Non-hematologic treatment related toxicities have improved to ≤ Grade 1 or to the 
subject’s baseline values (except alopecia) 
 
If blood counts are below this threshold (first two bullet points), blood work is to be repeated 
weekly until counts are at an acceptable level. If treatment is unable to restart within 12 weeks of 
the planned treatment date, the subject will be permanently discontinued from study therapy.   
 
6.2 
Protocol Therapy Discontinuation 
In addition to discontinuation from therapy related to toxicities as outlined above, a subject will 
also be discontinued from protocol therapy and followed per protocol under the following 
circumstances outlined below.  The reason for discontinuation of protocol therapy will be 
documented on the electronic case report form (eCRF)   
• Documented disease progression  
• 
Site investigator determines a change of therapy would be in the best interest of the 
subject 
• 
Subject requests to discontinue protocol therapy, whether due to unacceptable toxicity or 
for other reasons 
o In a subject decides to prematurely discontinue protocol therapy (“refuses 
treatment”), the subject should be asked if he or she may still be contacted for further 
scheduled study assessments. The outcome of that discussion should be documented 
in both the medical records and in the eCRF. 
• 
Female subject becomes pregnant  
• HD IL-2 will be discontinued if there is a ≥ 12-week delay between cycles 
• Pembrolizumab will be discontinued if there is a ≥ 12-week delay between cycles 
 
6.3 
Protocol Discontinuation  
If a subject decides to discontinue from the protocol (and not just from protocol therapy) all 
efforts should be made to complete and report study assessments as thoroughly as possible. A 
complete final evaluation at the time of the subject’s protocol withdrawal should be made with 
an explanation of why the subject is withdrawing from the protocol. If the reason for removal of 
a subject from the study is an adverse event, it will be recorded on the eCRF. 


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7. 
STUDY CALENDAR & EVALUATIONS 
 
Study Evaluations  
Cycle = 21 days for Pembrolizumab 
Cycle = 5 days for HD IL-2 
Screening 
Cycles 1-2 
Pembrolizumab (P) 
End of 
Cycle 2 
Pembro 
Cycle 1 
HD IL2 
Cycle 2 
HD IL2 
Safety follow 
up visit11 
Long-term 
Follow up12 
-28 days 
Day 1 
(± 3 days) 
Day 21 
(± 3 days) 
Days  
1-5 
Days  
15-19 
28 days post 
last dose 
(± 7 days) 
Every 3 
months  
(±14 days) 
REQUIRED ASSESSMENTS 
 
 
 
 
 
 
 
Informed Consent/HIPPA authorization 
X 
 
 
 
 
 
 
Medical History1 
X 
 
 
 
 
 
 
Diagnosis and Staging2 
 
 
 
 
 
 
 
Physical Exam 
X 
X 
X 
X 
X 
X 
 
Vital signs and ECOG Performance Status3 
X 
X 
X 
X 
X 
X 
 
AEs & concomitant medications 
X 
X 
X 
X 
X 
X 
 
LABORATORY ASSESSMENTS 
 
 
 
 
 
 
 
Complete Blood Cell Count with diff (CBC) 
X 
X10 
X 
X 
X 
X 
 
Comprehensive Metabolic Profile (CMP) 
X 
X10 
X 
X 
X 
X 
 
PT/INR and aPTT 
X 
X10 
 
 
 
 
 
Thyroid Function (TSH, T4, free T3) 
X 
 
X 
 
 
 
 
Vitamin D level 
X 
 
 
 
 
 
 
Pregnancy test (serum or urine) (WOCBP)4 
X 
X4 
 
 
 
 
 
Urinalysis 
X 
X10 
X 
 
X 
 
 
DISEASE ASSESSMENT 
 
 
 
 
 
 
 
Chest X-ray 
 
 
 
X 
 
 
 
CT of chest5 
X 
 
X 
 
 
X 
 
CT or MRI of abdomen and pelvis5 
X 
 
X 
 
 
X 
 
CT or MRI Brain5 
X 
 
X 
 
 
X 
 
TREATMENT EXPOSURE 
 
 
 
 
 
 
 
Pembrolizumab 
 
X 
 
 
 
 
 
HD IL-2 
 
 
 
X 
X 
 
 
SPECIMEN COLLECTION  
 
 
 
 
 
 
 
Archival Tumor Tissue6 
X 
 
 
 
 
 
 
Blood Samples7 
 
X7 
X 
X 
X7 
 
 
Stool sample8 
 
X 
 
 
 
 
 


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Study Evaluations  
Cycle = 21 days for Pembrolizumab 
Cycle = 5 days for HD IL-2 
Screening 
Cycles 1-2 
Pembrolizumab (P) 
End of 
Cycle 2 
Pembro 
Cycle 1 
HD IL2 
Cycle 2 
HD IL2 
Safety follow 
up visit11 
Long-term 
Follow up12 
-28 days 
Day 1 
(± 3 days) 
Day 21 
(± 3 days) 
Days  
1-5 
Days  
15-19 
28 days post 
last dose 
(± 7 days) 
Every 3 
months  
(±14 days) 
BANKING SAMPLES 
 
 
 
 
 
 
 
Whole blood, Plasma and PBMCs9 
 
X9 
X 
 
 
X 
 
FOLLOW-UP 
 
 
 
 
 
 
 
Survival Status, Subsequent Therapy 
 
 
 
 
 
 
X 
 
Key to Footnotes  
CBC with differential and platelet to include: Hgb, Hct, WBC, ANC, lymphocyte count, platelets. CMP to include sodium, potassium, chloride, 
creatinine, blood urea nitrogen; liver function tests (LFTs) to include AST, ALT, total bilirubin, alkaline phosphatase  
 
1: Medical History to include documentation of prior anti-cancer treatment including medications (chemotherapy, checkpoint inhibitors, etc.), 
radiation, or surgery.  A question of how the subject learned about the study will be asked. 
2: Diagnosis and staging to include pathology report and Tumor Node Metastasis (TNM) staging.   
3: Vital signs to include blood pressure, weight, and height (screening only) and ECOG performance status.   
4: For women of childbearing potential (WOCBP): urine or serum βhCG if clinically appropriate will be done during screening and within 72 
hours of initiation of study treatment. If a urine test is done and it is positive or cannot be confirmed as negative, a serum pregnancy test will be 
required.   
5: Appropriate scans including CT chest, abdomen/pelvis to assess disease status will be obtained within 4 weeks before initiation of study 
treatment (screening). Post treatment scans will be performed on Cycle 2 Day 21 (end of Cycle 2 pembrolizumab; +3 days) of pembrolizumab and 
Cycle 2 of HD IL-2; +9 days) of HD IL-2 including CT chest, abdomen/pelvis. CT or MRI of the brain should be done if clinically indicated at 
site investigator’s discretion.  
6: Required if available: submission of unstained slides from archived tumor tissue for PD-L1 expression are to be identified during screening and 
submitted after the patient is registered to the trial prior to Cycle 2 of pembrolizumab. If archived tumor tissue is not available, the patient does not 
need to undergo a biopsy to obtain tissue and is still eligible for study. 
7: Required: submission of blood for serum (soluble PD-L1, Th1/Th2 ratio and arginine levels) will be collected pre-dose Cycle 1 Day 1 of 
pembrolizumab, and on Cycle 2 Day 21 (+3 days) of pembrolizumab. Blood samples for these levels will also be collected at the end of each of 
the HD IL-2 cycles; End of Cycle 1 sample will be drawn (+3 days) prior to dosing Cycle 1 Day 5 (after completion of HD IL-2). The end of 
Cycle 2 sample will be drawn (+ 9 days) at the end of Cycle 2 Day 19 (after completion of HD IL-2.  


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8: Required: submission of a stool specimen (for microbiome characterization) will be collected at baseline (pre-dose Cycle 1 Day1 of 
pembrolizumab). Stool for microbiome analysis will be performed prior to treatment. Subjects will be provided a kit with detailed instructions 
regarding collection of the sample prior to the timepoint it is due. Please see the CLM for additional details. 
9: Optional: submission of whole blood, plasma and PBMCs for banking to be collected at baseline pre-dose Cycle 1 Day 1 of pembrolizumab and 
on Cycle 2 Day 21 (+3 days) of pembrolizumab. This sample will also be collected about 4 weeks after the last dose of HD IL-2 (safety visit) 
10: For Cycle 1 Day of pembrolizumab only: labs do not need to be repeated if done within 7 days of Day 1. 
11: The safety follow-up visit should occur when subjects permanently stop study treatment for whatever reasons (toxicity, progression, or at 
discretion of site investigator) and should be performed 30 days (±7 days) after the last dose of treatment. Subjects who have an ongoing Grade ≥ 
2 or serious AE (SAE) at this visit will continue to be followed until the AE resolves to ≤ Grade 1 or baseline, deemed clinically insignificant, 
and/or until a new anti-cancer treatment starts, whichever is earlier. 
12: Long-term follow up will occur in all subjects after the safety follow up visit. Subjects who discontinue treatment for any reason will be 
followed every 3 months for 12 months. Follow up may be accomplished via clinic visit, phone call, or other avenues as appropriate.   
 
 
 
 
 
 
 
 
 
 
 


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8. 
BIOSPECIMEN STUDIES AND PROCEDURES 
 
8.1 
Correlate PD-L1 expression in archived diagnostic tumor tissue with best clinical 
response (revised RECIST criteria) and treatment outcome in subjects with stage IV 
malignant melanoma 
Expression of PD-L1 in archived diagnostic tumor tissue will be determined by IHC by referring 
laboratory using anti-PD-L1 antibody clone 22C3, and will be correlated with clinical response 
assessed by imaging. Unstained slides are to be submitted; a tumor block is not acceptable. 
 
8.2 
Correlate the change in soluble PD-L1 and arginine levels in plasma during therapy 
with best clinical response (revised RECIST criteria) and treatment outcome in 
subjects with stage IV malignant melanoma 
Whole blood for serum submission will be collected pre-dose Cycle 1 Day 1 of pembrolizumab, 
and on Cycle 2 Day 21 (+3 days) of pembrolizumab. Blood samples for these levels will also be 
collected at the end of each of the HD IL-2 cycles; End of Cycle 1 sample will be drawn prior to 
dosing Cycle 1 Day 5 (+3 days). The end of Cycle 2 sample will be drawn (+ 9 days) at the end 
of Cycle 2 Day 19 for analysis of soluble PD-L1 and arginine levels. Change in soluble PD-L1 
and arginine levels as a result of treatment will be examined in relation to the best clinical 
response (RECIST, v1.1) and treatment outcome.  
 
8.3 
Correlate Th1/Th2 ratio and MDSCs in peripheral blood during therapy with best 
clinical response (revised RECIST criteria) and treatment outcome in subjects with 
stage IV malignant melanoma 
Whole blood for serum submission will be collected pre-dose Cycle 1 Day 1 of pembrolizumab, 
and on Cycle 2 Day 21 (+3 days) of pembrolizumab. Blood samples for these levels will also be 
collected at the end of each of the HD IL-2 cycles; End of Cycle 1 sample will be drawn prior to 
dosing Cycle 1 Day 5 (+3 days). The end of Cycle 2 sample will be drawn (+ 9 days) at the end 
of Cycle 2 Day 19 for analysis of Th1/Th2 ratio levels and MDSC quantification. Change in 
Th1/Th2 ratio and MDSCs levels as a result of treatment will be examined in relation to the best 
clinical response (RECIST, v1.1) and treatment outcome.  
 
8.4 
Correlate intestinal microbiome at baseline with best clinical response (revised 
RECIST criteria) and treatment outcome in subjects with stage IV malignant 
melanoma 
Samples of stool will be collected prior to pembrolizumab treatment in a stool collection tube per 
manufacturer's guidelines. Microbial DNA will be isolated using the Invitek PSP® Spin Stool 
DNA Plus Kit (with lysis enhancer) as described by the manufacturer [29]. Subsequent 
microbiome characterization will be performed after study complete accrual using Illumina 16S 
Metagenomic sequencing protocol. 
 
8.5 
Correlative samples for future unspecified cancer related studies 
Subject consent will be obtained for additional samples (whole blood, plasma and PBMCs) to be 
collected and banked for future unspecified cancer related research. Samples will be collected at 
baseline (pre-dose Cycle 1 Day 1 of pembrolizumab) and on Cycle 2 Day 21 (+3 days) of 
pembrolizumab. This sample will also be collected about 4 weeks after the last dose of HD IL-2 
(safety visit). These samples will be banked at HCRN. 


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8.5.1 Storage of Biospecimens 
Remaining specimens once protocol described biospecimen-based studies are complete will be 
stored for future unspecified cancer related research. Permission for this will be obtained from 
subjects in the informed consent. 
 
8.5.2 Confidentiality of Biospecimens 
Samples that are collected will be identified by a subject’s sequence ID number assigned at the 
time of registration to the trial. Any material issued to collaborating researchers will be 
anonymized and only identified by the subject’s sequence ID number. 
 
9. 
CRITERIA FOR DISEASE EVALUATION 
Response assessments will be made using RECIST v1.1 [30], allowing additional comparisons 
among these criteria for disease response assessment. The same measurable and non-measurable 
lesions will be followed by RECIST v1.1. Disease progression will be determined using RECIST 
v1.1 criteria. 
 
9.1 
Measurable Disease 
The presence of at least one measurable lesion. If the measurable disease is restricted to a 
solitary lesion, its neoplastic nature should be confirmed by cytology/histology. 
 
9.2 
Measurable Lesions 
Measurable lesions are defined as the presence of at least one measurable lesion. Measurable 
lesions are defined as those that can be accurately measured in at least one dimension (longest 
diameter to be recorded) as >20 mm by chest x-ray, as >10 mm with CT scan, or >10 mm with 
calipers by clinical exam. All tumor measurements must be recorded in millimeters (or decimal 
fractions of centimeters). 
 
9.2.1 Malignant Lymph Nodes 
To be considered pathologically enlarged and measurable, a lymph node must be >15 mm in 
short axis when assessed by CT scan (CT scan slice thickness recommended to be no greater 
than 5 mm). At baseline and in follow-up, only the short axis will be measured and followed. 
 
9.3 
Non-measurable Lesions 
All other lesions (or sites of disease), including small lesions (longest diameter <10 mm or 
pathological lymph nodes with ≥ 10 to <15 mm short axis), are considered non-measurable 
disease. Bone lesions, leptomeningeal disease, ascites, pleural/pericardial effusions, lymphangitis 
cutis/pulmonitis, inflammatory breast disease, and abdominal masses (not followed by CT or 
MRI), are considered as non-measurable. 
 
NOTE:  Cystic lesions that meet the criteria for radiographically defined simple cysts should not 
be considered as malignant lesions (neither measurable nor non-measurable) since they are, by 
definition, simple cysts. ‘Cystic lesions’ thought to represent cystic metastases can be considered 
as measurable lesions, if they meet the definition of measurability described above. However, if 
non-cystic lesions are present in the same subject, these are preferred for selection as target 
lesions. 
 


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9.4 
Target Lesions 
All measurable lesions up to a maximum of 2 lesions per organ and 5 lesions in total, 
representative of all involved organs, should be identified as target lesions and recorded and 
measured at baseline. Target lesions should be selected on the basis of their size (lesions with the 
longest diameter), be representative of all involved organs, but in addition should be those that 
lend themselves to reproducible repeated measurements. It may be the case that, on occasion, the 
largest lesion does not lend itself to reproducible measurement in which circumstance the next 
largest lesion which can be measured reproducibly should be selected. A sum of the diameters 
(longest for non-nodal lesions, short axis for nodal lesions) for all target lesions will be 
calculated and reported as the baseline sum diameters. If lymph nodes are to be included in the 
sum, then only the short axis is added into the sum. The baseline sum diameters will be used as 
reference to further characterize any objective tumor regression in the measurable dimension of 
the disease. 
 
9.5 
Non-target Lesions 
All other lesions (or sites of disease) including any measurable lesions over and above the 5 
target lesions should be identified as non-target lesions and should also be recorded at baseline. 
Measurements of these lesions are not required, but the presence, absence, or in rare cases 
unequivocal progression of each should be noted throughout follow-up. 
 
9.6 
Evaluation of Target Lesions 
NOTE: In addition to the information below, also see section 4.3.2 in the international criteria 
proposed by the Response Evaluation Criteria in Solid Tumors (RECIST) Committee, version 
1.1 (Eur J Cancer 45;2009:228-247) for special notes on the assessment of target lesions.  
 
 
Complete Response (CR) 
Disappearance of all target lesions. Any pathological lymph 
nodes (whether target or non-target) must have reduction in 
short axis to <10 mm. 
Partial Response (PR) 
At least a 30% decrease in the sum of the diameters of target 
lesions, taking as reference the baseline sum diameters 
Progressive Disease (PD) 
At least a 20% increase in the sum of the diameters of target 
lesions, taking as reference the smallest sum on study (this 
includes the baseline sum if that is the smallest on study). In 
addition to the relative increase of 20%, the sum must also 
demonstrate an absolute increase of at least 5 mm. (Note:  
the appearance of one or more new lesions is also considered 
progressions). 
Stable Disease (SD) 
Neither sufficient shrinkage to qualify for PR nor sufficient 
increase to qualify for PD, taking as reference the smallest 
sum diameters while on study 
 
 
 


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9.7 
Evaluation of Non-Target Lesions 
Complete Response (CR) 
Disappearance of all non-target lesions and normalization 
of tumor marker level.  
Incomplete Response/Stable 
Disease 
Persistence of one or more non-target lesion(s) and/or 
maintenance of tumor marker level above the normal 
limits 
Progressive Disease (PD) 
Appearance of one or more new lesions and/or 
unequivocal progression of existing non-target lesions.  
 
Although a clear progression of “non-target” lesions only is exceptional, the opinion of the site 
investigator should prevail in such circumstances, and the progression status should be 
confirmed at a later time by the sponsor investigator. 
 
9.8 
Evaluation of Best Overall Response 
Target Lesions 
Non-Target Lesions 
New Lesions 
Overall 
Response 
CR 
CR 
No 
CR 
CR 
Non-CR/ Non-PD 
No 
PR 
CR 
Not evaluated 
No 
PR 
PR 
Non-PD/ or not all 
evaluated 
No 
PR 
SD 
Non-PD or not all evaluated 
No 
SD 
Not all evaluated 
Non-PD 
No 
Non-evaluable 
PD 
Any 
Yes or No 
PD 
Any 
PD* 
Yes or No 
PD 
Any 
Any 
Yes 
PD 
*In exceptional circumstances, unequivocal progression in non-target lesions may be 
accepted as disease progression. 
 
Subjects with a global deterioration of health status requiring discontinuation of treatment 
without objective evidence of disease progression at that time should be classified as having 
“symptomatic deterioration.” Every effort should be made to document the objective progression 
even after discontinuation of treatment. 
 
In some circumstances, it may be difficult to distinguish residual disease from normal tissue. 
When the evaluation of complete response depends on this determination, it is recommended that 
the residual lesion be investigated (fine needle aspirate/biopsy) to confirm the complete response 
status. 
 
 
 


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9.9 
Definitions for Response Evaluation – RECIST 1.1 
 
9.9.1 First Documentation of Response 
The time between initiation of therapy and first documentation of PR or CR. 
 
9.9.2 Confirmation of Response 
To be assigned a status of complete or partial response, changes in tumor measurements must be 
confirmed by repeat assessments performed no less than four weeks after the criteria for response 
are first met. 
 
9.9.3 Duration of Response 
Duration of overall response—the period measured from the time that measurement criteria are 
met for complete or partial response (whichever status is recorded first) until the date that 
recurrent or progressive disease is objectively documented (taking as reference for progressive 
disease the smallest measurements recorded since treatment started). 
 
9.9.4 Duration of Overall Complete Response 
The period measured from the time that measurement criteria are met for complete response until 
the first date that recurrent disease is objectively documented. 
 
9.9.5 Objective Response Rate  
The objective response rate is the proportion of all subjects with confirmed PR or CR according 
to RECIST 1.1, from the start of treatment until disease progression/recurrence (taking as 
reference for progressive disease the smallest measurements recorded since the start of 
treatment). 
 
9.9.6 Clinical Benefit Rate 
The proportion of patients with a confirmed complete response, a partial response or stable 
disease based on RECIST v1.1. 
 
9.9.7 Progression-Free Survival 
A measurement from the date of treatment initiation until the criteria for disease progression is 
met as defined by RECIST 1.1 or death occurs. Subjects who have not progressed will be right-
censored at the date of the last disease evaluation. 
 
9.9.8 Overall Survival 
Overall survival is defined by the date of treatment initiation to date of death from any cause. 
 
 
 


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10 DRUG INFORMATION 
 
10.1 
Keytruda® (Pembrolizumab; MK-3475 [Anti-PD-1 Antibody MK-3475] 
 
Please refer to the current version of the Investigator’s Brochure (IB) for additional information 
regarding this drug. 
 
10.1.1 Chemical name and properties 
Humanized X PD-1_mAb (H409A11) IgG4 
 
10.1.2 Availability 
Pembrolizumab is commercially available and will be ordered through outpatient pharmacy 
where the subject is being treatment. 
 
10.1.3 Storage 
Clinical supplies must be stored in a secure, limited-access location under the storage conditions 
specified on the label. Receipt and dispensing of trial medication must be recorded by an 
authorized person at the trial site. Clinical supplies may not be used for any purpose other than 
that stated in the protocol. 
 
10.1.4 Dosage and Administration 
Pembrolizumab is a white to off-white lyophilized powder (50 mg/vial) or as a liquid solution 
(100 mg/vial) in Type I glass vials intended for single use only. Pembrolizumab Powder for 
Solution for Infusion, 50 mg/vial, is reconstituted with sterile water for injection prior to use. 
Pembrolizumab is formulated with L-histidine as buffering agent, polysorbate 80 as surfactant, 
sucrose as stabilizer/tonicity modifier, and hydrochloric acid (HCl) and/or sodium hydroxide 
(NaOH) for pH adjustment (if necessary). The drug product is stored as a stable lyophilized 
powder or liquid solution under refrigerated conditions (2°C - 8°C). 
 
The product after reconstitution with sterile water for injection and the liquid drug product is a 
clear to opalescent solution, which may contain extraneous and proteinaceous particulates. The 
reconstituted product and liquid product is intended for IV administration. The reconstituted drug 
product solution and liquid drug product can be further diluted with normal saline in IV 
containers made of polyvinyl chloride (PVC) or non-PVC material. Reconstituted vials should be 
immediately used to prepare the infusion solution in the IV bag and the infusion solution should 
be immediately administered. If not used immediately, vials and/or IV bags may be stored at 2-8 
°C for up to a cumulative time of 20 hours. If refrigerated, the vials and/or IV bags should be 
allowed to equilibrate to room temperature prior to subsequent use. Pembrolizumab solutions 
may be stored at room temperature for a cumulative time of up to 4 hours. This includes room 
temperature storage of reconstituted drug product solution and liquid drug product in vials, room 
temperature storage of infusion solution in the IV bag and the duration of infusion. 
 
10.1.5 Side Effects 
Please refer to the current version of the Investigator’s Brochure for a complete list of adverse 
events. 
 


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Pembrolizumab is generally well tolerated and demonstrates a favorable safety profile in 
comparison to chemotherapy. Pembrolizumab is an immunomodulatory agent, and based on this 
mechanism of action, immune mediated adverse events are of primary concern. Important 
identified risks for pembrolizumab are of an immune mediate nature, including: pneumonitis, 
colitis, thyroid disorders (hypothryoidism/hyperthyroidism), hepatitis, hypophysitis, Type I 
diabetes mellitis, uveitis, nephritis, pancreatitis, myositis, and severe skin reaction. After a recent 
review of data, events newly characterized as identified risks also include vitiligo, Guillan-Barre 
Syndrome, and adrenal insufficiency. The majority of immune-mediated adverse events were 
mild to moderate in severity, were manageable with appropriate care, and rarely required 
discontinuation of therapy. 
 
Further details around frequency, reporting, and management of immune-related adverse events 
(irAEs) can be found in the current version of the Investigator’s Brochure. In addition to the 
previously noted identified risks, infusion-related reactions are a risk but are not considered 
immune mediated; these are also further described in the current IB. 
 
10.2 
Proleukin (Aldesleukin; Interleukin-2) 
 
10.2.1 Chemical name and properties 
Desalanyl-1, serine-125 human recombinant interleukin-2 
MW: 15.3 KD 
This IL-2 differs from native interleukin-2 in the following ways:  
▪ Not glycosylated 
▪ No N-terminal alanine 
▪ C125S mutation 
▪ Aggregation state 
Biological potency: 1.1 mg = 18 x 106 IU 
 
10.2.2 Availability 
Proleukin is commercially available and will be ordered through outpatient pharmacy where the 
subject is being treatment. 
 
10.2.3 Package 
Sterile single-use vials intended for intravenous administration. Each vial contains 22 x 106  IU 
of lyophilized Proleukin.  
 
10.2.4 Storage 
Clinical supplies must be stored in a secure, limited-access location under the storage conditions 
specified on the label. Store vials of lyophilized Proleukin in a refrigerator at 2° to 8°C. Protect 
from light. Receipt and dispensing of trial medication must be recorded by an authorized person 
at the trial site. Clinical supplies may not be used for any purpose other than that stated in the 
protocol. 
 
10.2.5 Dosage and Administration 
Please refer to the Pharmacy Manual for a comprehensive description of Proleukin preparation. 
Reconstituted or diluted Proleukin is stable for up to 48 hours at 2° to 8° C. 


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Prometheus Laboratories will supply directly to sites at no cost to patients in this clinical trial. 
Proleukin is supplied as a sterile, white to off-white, lyophilized cake in single-use vials intended 
for intravenous administration. When reconstituted with 1.2 mL sterile water for injection, USP, 
each mL contains 18 x 106 IU (1.1 mg) Proleukin, 50 mg mannitol, and 0.18 mg sodium dodecyl 
sulfate, buffered with approximately 0.17 mg monobasic and 0.89 mg dibasic sodium phosphate 
to a pH of 7.5 (range 7.2 to 7.8). The manufacturing process for Proleukin involves fermentation 
in a defined medium containing tetercycline hydrochloride. The presence of the antibiotic is not 
detectable in the final product. Proleukin contains no preservatives in the final product. 
 
The recommended Proleukin treatment regimen is administered in the inpatient setting on a 
monitored unit. Each course of treatment consists of two 5-day treatment cycles, the cycles 
separated by rest period for 9 days. In each cycle, 600,000 IU/kg (0.037 mg/kg) will be 
administered as a 15-minute intravenous infusion every 8 hours for up to 14 consecutive doses 
over 5 days.  
 
10.2.6 Side Effects 
Please refer to the current version of the Investigator’s Brochure for a complete list of adverse 
events. 
 
The following adverse events (Grades 1-4) were seen in ≥30% of 525 patients (255 with 
metastatic renal cell cancer and 270 with metastatic melanoma) treated with Proleukin®: 
hypotension (71%), diarrhea (67%), oliguria (63%), chills (52%), vomiting (50%), dyspnea 
(43%), rash (42%), bilirubinemia (40%), thrombocytopenia (37%), nausea (35%), confusion 
(34%), and creatinine increase (33%). Due to the potential severe adverse events that might 
accompany Proleukin therapy at the recommended dosages, thorough clinical evaluation will be 
performed to exclude subjects that are contraindicated to Proleukin therapy and patients that 
have significant cardiac, pulmonary, renal, hepatic, or CNS impairment. Adverse events are 
frequent, often serious, and sometimes fatal. Further details around frequency, reporting, and 
management of immune-related adverse events (irAEs) can be found in the current version of the 
Investigator’s Brochure. In addition to the previously noted identified risks, infusion-related 
reactions are a risk but are not considered immune mediated; these are also further described in 
the current IB. 
 
 
 


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11 
ADVERSE EVENTS 
The descriptions and grading scales found in the NCI CTCAE v4.03 will be utilized for AE 
assessment. A copy of the CTCAE v4.03 can be downloaded from the CTEP website at 
http://ctep.cancer.gov. All forms for AE/SAE recording and reporting can be found in the Study 
Procedure Manual or in the EDC system (Documents and Information Tab). 
 
11.1 
Definitions 
 
11.1.1 Adverse Event (AE) 
An AE is any untoward medical occurrence whether or not considered related to the study drug 
that appears to change in intensity during the course of the study. The following are examples of 
AEs:  
• Unintended or unfavorable sign or symptom 
• A disease temporally associated with participation in the protocol 
• An intercurrent illness or injury that impairs the well-being of the subject 
 
Abnormal laboratory values or diagnostic test results constitute AEs only if they induce clinical 
signs or symptoms or require treatment or further diagnostic tests   
 
Hospitalization for elective surgery or routine clinical procedures that are not the result of an AE 
(e.g., surgical insertion of central line) should not be recorded as an AE.  
 
Disease progression should not be recorded as an AE, unless it is attributable to the study 
regimen by the site investigator. 
 
11.1.2 Serious Adverse Event (SAE) 
A SAE is an adverse event that: 
• Results in death. NOTE: Death due to disease progression should not be reported as a 
SAE, unless it is attributable by the site investigator to the study drug(s) 
• Is life-threatening (defined as an event in which the subject was at risk of death at the 
time of the event; it does not refer to an event which hypothetically might have caused 
death if it were more severe) 
• Requires inpatient hospitalization for >24 hours or prolongation of existing 
hospitalization. NOTE: Hospitalization for anticipated or protocol specified procedures 
such as administration of chemotherapy, central line insertion, metastasis interventional 
therapy, resection of primary tumor, or elective surgery, will not be considered serious 
adverse events. 
• Results in persistent or significant disability/incapacity 
• Is a congenital anomaly or birth defect 
• Is an important medical event (defined as a medical event(s) that may not be immediately 
life-threatening or result in death or hospitalization but, based upon appropriate medical 
and scientific judgment, may jeopardize the subject or may require intervention (e.g., 
medical, surgical) to prevent one of the other serious outcomes listed in the definition 
above). Examples of such events include, but are not limited to, intensive treatment in an 
emergency room or at home for allergic bronchospasm; blood dyscrasias or convulsions 
not resulting in hospitalization; or the development of drug dependency or drug abuse. 


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11.1.3 Unexpected Adverse Event 
For this study, an AE is considered unexpected when it varies in nature, intensity or frequency 
from information provided in the current IB, prescribing information or when it is not included in 
the informed consent document as a potential risk. Unexpected also refers to AEs that are 
mentioned in the IB as occurring with a class of drugs or are anticipated from the 
pharmacological properties of the drug, but are not specifically mentioned as occurring with the 
particular drug under investigation.   
 
11.1.4 Relatedness 
AEs will be categorized according to the likelihood that they are related to the study drug(s). 
Specifically, they will be categorized using the following terms:  
 
Unrelated 
Adverse Event is not related to the study drug(s)  
Unlikely 
Adverse Event is doubtfully related to the study drug(s)  
Possible 
Adverse Event may be related to the study drug(s)  
Probable 
Adverse Event is likely related to the study drug(s)  
Definite 
Adverse Event is clearly related to the study drug(s)  
 
11.2 
Reporting 
 
11.2.1 Adverse Events  
• AEs will be recorded from time of signed informed consent until 30 days after 
discontinuation of HD IL-2.  
• AEs will be recorded regardless of whether or not they are considered related to the 
pembrolizumab or HD IL-2.   
• All AEs will be recorded in the subject’s medical record and on the appropriate study 
specific eCRF form within the EDC system. 
• Asymptomatic laboratory abnormalities that do not require treatment will not be collected 
as adverse events. 
 
11.2.2 Serious Adverse Events (SAEs) 
 
11.2.2.1 
Site Requirements for Reporting SAEs to HCRN 
• SAEs will be reported from time of signed informed consent until 30 days after 
discontinuation of HD IL-2.  
• SAEs will be reported on the SAE Submission Form within 1 business day of discovery 
of the event.   
• SAEs include events related and unrelated to pembrolizumab or HD IL-2. 
• All SAEs will be recorded in the subject’s medical record and on the appropriate study 
specific eCRF form within the EDC system.   
 
The site will submit the completed SAE Submission Form to HCRN within 1 business day of 
discovery of the event. The form may be submitted to HCRN electronically to 
safety@hoosiercancer.org. The site investigator is responsible for informing the IRB and/or other 
local regulatory bodies as per local requirements.   


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The original copy of the SAE Submission Form and the email correspondence must be kept 
within the study file at the study site. 
 
Once the SAE has resolved (see resolution guidelines listed above), sites must submit a follow-
up SAE Submission Form within a reasonable timeframe to HCRN electronically to 
safety@hoosiercancer.org.   
 
11.2.2.2 
HCRN Requirements for Reporting SAEs to Prometheus 
HCRN will report all SAEs to Prometheus as stipulated by each company (i.e., all SAEs, only 
those related etc.) within 1 business day of receipt of the SAE Submission Form from a site. 
Follow-up information will be provided to Prometheus as it is received from site.    
 
Contact information for Prometheus 
Drug Safety Email: drugsafety@prometheuslabs.com 
Drug Safety Fax: (858) 754-3046 
 
 
11.3 
Sponsor-Investigator Responsibilities  
HCRN will send a SAE summary to the sponsor-investigator within 1 business day of receipt of 
SAE Submission Form from a site.  The sponsor-investigator will promptly review the SAE 
summary and assess for expectedness and relatedness.   
 
11.4 
HCRN Responsibilities to FDA 
HCRN will continue to facilitate compliance of applicable requirements for the sponsor-
investigator in relation to this study. This includes but is not limited to 21 CFR 50.20 informed 
consent, 21 CFR Part 56 IRB, and pertinent sections of the Public Health Service Act and 
FDAAA. 
 
11.5 
IND Safety Reports Unrelated to this Trial 
Prometheus will provide to HCRN IND safety reports from external studies that involve the 
study drug(s) per their guidelines.  HCRN will forward safety reports to the sponsor-investigator 
who will review these reports and determine if revisions are needed to the protocol or consent. 
HCRN will forward these reports to participating sites within 1 business day of receiving the 
sponsor-investigator’s review.  Based on the sponsor-investigator’s review, applicable changes 
will be made to the protocol and informed consent document (if required).  All IND safety 
reports will also be made available to sites via the EDC system. Upon receipt from HCRN, site 
investigators (or designees) are responsible for submitting these safety reports to their respective 
IRBs, as per their IRB policies. 
 
 
 


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12 
STATISTICAL METHODS 
 
12.1 
Study Design 
The primary objective of this single arm Phase II trial is to assess the response rate [complete 
response (CR) + partial response (PR)] of sequential therapy of pembrolizumab followed by HD 
IL-2 in subjects with stage IV malignant melanoma. Response assessment will be performed 
using revised RECIST guideline (version 1.1). The trial will consist of two phases. In the first 
phase of this trial, therapy will be initiated with pembrolizumab followed by HD IL-2 therapy. 
Pembrolizumab 200 mg will be given every 3 weeks for 2 doses (cycles). Staging will occur 
approximately 20 days after the last treatment dose (i.e., Day 21 [+3 days] of Cycle 2). Then 
subjects will receive HD IL-2 for 2 cycles, Cycle 2 separated from Cycle 1 by 9 days (one 
course). Subjects are hospitalized during each cycle which consists of up to 14 infusions over a 5 
day period (3 infusions/day spaced 8 hours apart). Four weeks after the last HD IL-2 
administration of Cycle 2, the subjects will be restaged (i.e., their disease status reassessed). For 
subjects with progressive disease as revealed by the restaging, they will be followed for OS up to 
one year from initiation of the first treatment (pembrolizumab) of the study. For subjects with 
CR, PR, and SD, scans will be performed at 3-month intervals until disease progression or up to 
a year after initiation of the first treatment (with pembrolizumab) on study. For the subjects who 
progress, although they will be taken off the study they will be followed for OS up to one year 
from initiation of the first treatment (pembrolizumab) of the study. 
 
12.2 
Endpoints and Analyses 
 
12.2.1 Primary Endpoint 
The primary endpoint of this single arm Phase II trial is to assess the response rate [complete 
response (CR) + partial response (PR)] of sequential therapy of pembrolizumab followed by HD 
IL-2 in subjects with stage IV malignant melanoma, using revised RECIST guideline (version 
1.1). Response rate will be computed with associated 95% confidence intervals constructed from 
the exact Binomial method. 
 
12.2.2 Secondary Endpoints 
To characterize safety, tolerability and adverse effects (AE) profile of pembrolizumab followed 
by HD IL-2 in subjects with stage IV malignant melanoma. Proportion of subjects with each 
grade of adverse events as defined by CTCAE v4.03 will be computed along with 95% 
confidence intervals, and reported in a tabular and descriptive manner. 
 
To measure Progression-Free Survival (PFS) using revised RECIST 1.1 after completion of 2 
cycles of pembrolizumab and 2 cycles of HD IL-2 in all the subjects enrolled in the study. 
Median PFS times will be computed if reached, and PFS rates at 1 year +/- 3 months will be 
calculated with associated 95% confidence intervals based on the Kaplan-Meier method.  
 
To measure overall survival (OS) at 12 months in subjects with stage IV malignant melanoma 
who showed a response [stable disease (SD), complete response (CR) or partial response (PR)] 
following completion of 2 cycles of pembrolizumab and 2 cycles of HD IL-2. Median OS times 
will be computed, and OS rate at 1 year +/- 3 months will be calculated with associated 95% 
confidence intervals based on the Kaplan-Meier method.  


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12.2.3 Analysis for Correlative Objectives 
Correlative research analyses include examining the relationship between best clinical 
response/treatment outcome and PD-L1 expression in archived diagnostic tumor tissue, as well 
as examining the relationship between best clinical response/treatment outcome and change in 
soluble PD-L1 level and Th1/Th2 ratio level in serum during treatment after the 2nd dose of 
pembrolizumab, after each cycle of HD IL-2, relative to baseline (i.e., pre-pembrolizumab dose 
Day 1 of Cycle 1) and baseline microbiome diversity. PD-L1 expression can be discretized into 
different levels and tests such as Fisher’s exact test and log-rank test will be used for its related 
analysis depending on the outcome types. Similar type of analyses will also be conducted to 
examine the change in soluble PD-L1 level, change in Th1/Th2 ratio level, arginine levels and 
type of intestinal microbiome at baseline. 
 
12.3 
Sample Size and Accrual 
Response rates for anti-PD-1 treatment ranges from 20%-40% [9,33-35]. Response for IL-2 in 
the pivotal study was 16% for melanoma [15,16]. We based our sample size calculation on the 
premise that response rate must be 38% or higher – otherwise the drug treatment regimen 
described in this protocol will be rejected (i.e., not considered further for clinical trials). 
 
The optimal two-stage design to test the null hypothesis that P ≤ 0.260 versus the alternative that 
P ≥ 0.500 with 80% power and 0.05 level of significance proceeds as follows. After testing the 
drug on 11 patients, the trial will be terminated if 3 or fewer respond. If the trial goes on to the 
second stage, a total of 29 patients will be studied. If the total number responding is less than or 
equal to 11, the drug is rejected. The probability of early termination at stage 1 with a total 
sample size of 11 is 0.685; the probability of going on to stage 2 with a total sample size of 29 is 
0.315. The sample size calculation is conducted using the software PASS 13 (NCSS, Kaysville, 
Utah 2014). 
 
12.4 
Analysis Datasets 
 
Population 
Definition 
Enrolled 
This will comprise all subjects who meet the 
eligibility criteria and are registered onto the study. 
  
Evaluable 
This will comprise all subjects who receive at least 
one dose of trial drug and either undergo at least 
one post-baseline assessment or die before any 
evaluation. Unevaluable patients will be replaced. 
   
Safety 
This will comprise all subjects that receive at least 
one dose of study drug. 
 
 
12.5 
Interim Analysis/Criteria for Stopping Study 
A Two-Stage Phase II Simon design will be used in this study. After testing the study drug on 11 
patients in the first stage, the trial will be terminated if 3 or fewer respond. During this analysis, 
accrual to the trial will be halted. 


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13 
TRIAL MANAGEMENT 
 
13.1 
Data and Safety Monitoring Plan (DSMP) 
The study will be conducted with guidance from the Health Partner’s DSMP.  
 
HCRN oversight activities include: 
• Review all adverse events requiring expedited reporting as defined in the protocol 
• Provide trial accrual progress, safety information and data summary reports to the 
sponsor-investigator  
• Submit data summary reports to the DSMB for review according to DSMB Charter  
 
13.2 
Data Safety Monitoring Board 
This study will have a Data and Safety Monitoring Board (DSMB). The DSMB is chaired by an 
independent medical oncologist external to this trial. The DSMB will provide a recommendation 
to the sponsor-investigator after all information is reviewed. This information will also be 
provided to HCRN who will distribute to the site investigator/participating sites for submission 
to their respective IRB according to the local IRB’s policies and procedures. 
 
The DSMB review will include but is not limited to: 
• Adverse event summary report 
• Audit results if applicable 
• Data related to stopping/decision rules described in study design 
• Study accrual patterns  
• Protocol deviations 
 
13.3 
Data Quality Oversight Activities 
Remote validation of the EDC system data will be completed on a continual basis throughout the 
life cycle of the study. Automated edit check listings will be used to generate queries in the EDC 
system and transmitted to the site to address in a timely fashion. Corrections will be made by the 
study site personnel.  
 
Participating sites may also be subject to quality assurance audits by Prometheus or its designee 
as well as inspection by appropriate regulatory agencies. 
 
13.3.1 Onsite Monitoring 
There will be at least one routine visit per site per year for sites that have accrued.  Additional for 
cause visits may occur as necessary. Selected source documents will be reviewed for verification 
of agreement with data entered into the EDC system.  It is important for the site investigator and 
their relevant personnel to be available for a sufficient amount of time during the monitoring 
visits or audit, if applicable. The site investigator and institution guarantee access to source 
documents by HCRN or its designee. 
 
The trial site may also be subject to quality assurance audit by Prometheus or its designee as well 
as inspection by appropriate regulatory agencies. 
 


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13.4 
Compliance with Trial Registration and Results Posting Requirements 
Under the terms of the Food and Drug Administration Modernization Act (FDAMA) and the 
Food and Drug Administration Amendments Act (FDAAA), the sponsor-investigator of the trial 
is solely responsible for determining whether the trial and its results are subject to the 
requirements for submission to the Clinical Trials Data Bank, http://www.clinicaltrials.gov. All 
results of primary and secondary objectives must be posted to CT.gov within a year of 
completion.  The sponsor-investigator has delegated responsibility to HCRN for registering the 
trial and posting the results on clinicaltrials.gov. Information posted will allow subjects to 
identify potentially appropriate trials for their disease conditions and pursue participation by 
calling a central contact number for further information on appropriate trial locations and study 
site contact information.   
 
14. 
DATA HANDLING AND RECORD KEEPING 
 
14.1 
Data Management 
HCRN will serve as the Clinical Research Organization for this trial. Data will be collected 
through a web based clinical research platform, a system compliant with Good Clinical Practices 
and Federal Rules and Regulations.  HCRN personnel will coordinate and manage data for 
quality control assurance and integrity. Select data will be collected and entered into the EDC 
system  by study site personnel from participating institutions.   
 
14.2 
Case Report Forms and Submission 
Generally, clinical data will be electronically captured in the EDC system and correlative results 
will be captured in the EDC system or other secure database(s). If procedures on the study 
calendar are performed for standard of care, at minimum, that data will be captured in the source 
document. Select standard of care data will also be captured in the EDC system, according to 
study-specific objectives.  
 
The completed dataset is the sole property of the sponsor-investigator’s institution and should 
not be exported to third parties, except for authorized representatives of appropriate 
Health/Regulatory Authorities, without permission from the sponsor-investigator and HCRN. 
 
14.3 
Record Retention 
To enable evaluations and/or audits from Health Authorities/HCRN, the site investigator agrees 
to keep records, including the identity of all subjects (sufficient information to link records; e.g., 
hospital records), all original signed informed consent forms, copies of all source documents, and 
detailed records of drug disposition. All source documents are to remain in the subject’s file and 
retained by the site investigator in compliance with the site contract with HCRN.  No records 
will be destroyed until HCRN confirms destruction is permitted. 
 
 
 


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14.4 
Confidentiality 
There is a slight risk of loss of confidentiality of subject information. All records identifying the 
subjects will be kept confidential and, to the extent permitted by the applicable laws and/or 
regulations, will not be made publicly available. Information collected will be maintained on 
secure, password protected electronic systems. Paper files that contain personal information will 
be kept in locked and secure locations only accessible to the study site personnel.  
Subjects will be informed in writing that some organizations including the sponsor-investigator 
and his/her research associates, HCRN, Prometheus, IRB, or government agencies, like the FDA, 
may inspect their medical records to verify the information collected, and that all personal 
information made available for inspection will be handled in strictest confidence and in 
accordance with local data protection laws. 
 
If the results of the study are published, the subject’s identity will remain confidential. 
 
15 
ETHICS 
 
15.1 
Institutional Review Board (IRB) Approval 
The final study protocol and the final version of the informed consent form must be approved in 
writing by an IRB. The site investigator must submit written approval by the IRB to HCRN 
before he or she can enroll subjects into the study.   
 
The site investigator is responsible for informing the IRB of any amendment to the protocol in 
accordance with local requirements. In addition, the IRB must approve all advertising used to 
recruit subjects for the study. The protocol must be re-approved by the IRB, as local regulations 
require.  
 
Progress reports and notifications of serious and unexpected adverse events will be provided to 
the IRB according to local regulations and guidelines. 
 
15.2 
Ethical Conduct of the Study 
The study will be performed in accordance with ethical principles originating from the 
Declaration of Helsinki.  Conduct of the study will be in compliance with ICH Good Clinical 
Practice, and with all applicable federal (including 21 CFR parts 56 & 50), state, or local laws. 
 
15.3 
Informed Consent Process 
The site investigator will ensure the subject is given full and adequate oral and written 
information about the nature, purpose, possible risks and benefits of the study. Subjects must 
also be notified they are free to discontinue from the study at any time. The subject should be 
given the opportunity to ask questions and allowed time to consider the information provided. 
 
The subject’s signed and dated informed consent must be obtained before conducting any 
procedure specifically for the study. The site investigator must store the original, signed 
informed consent form. A copy of the signed informed consent form must be given to the 
subject.  
 
 
 


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Clinical Study Protocol 
HCRN MMP-MEL16-261 
Version Date: 28FEB2018 
Confidential 
Page 51 of 51 
APPENDIX 1: ECOG Performance Status 
 
ECOG Score 
Performance Status 
0 
Asymptomatic 
1 
Symptomatic, fully ambulatory 
2 
Symptomatic, in bed < 50% of the day 
3 
Symptomatic, in bed > 50% of the day but not bedridden 
4 
Bedridden 
5 
Dead