Conditions: Carcinoma, Unspecified Site
Allocation: NA | Intervention model: SINGLE_GROUP
Enrollment: 14
Healthy volunteers: no

Brief summary:
Single-arm phase 2 study to examine pembrolizumab and concurrent radiation to induce an abscopal effect in patients with previously treated carcinoma of unknown primary (CUP16-268)

Detailed description:
This is a proof-of-principle single-arm phase 2 study in patients with previously treated CUP. All patients receive pembrolizumab combined with Radiation Therapy (RT) to a metastatic site, so as to induce an abscopal tumor response. The treatment combination will be repeated with RT delivery to a second metastatic site in a non-overlapping RT field.

The results will be compared with historical control. The primary endpoint is the confirmed response rate (RR) in a non-irradiated site based on best responding abscopal lesion. This study will also evaluate the following secondary endpoints: RR in a non-irradiated site based on RECIST 1.1, adverse events, progression-free survival (PFS), overall survival (OS), time-to-progression (TTP), and disease control rate (DCR).

Arms:
arm 1: Pembrolizumab + External Beam Radiation Therapy

Interventions: intervention 1: Pembrolizumab intervention 2: External Beam Radiation Therapy
Intervention descriptions: intervention 1: 200mg IV at day -21, then day 1 of each 21-day cycle for a maximum of 24 continuous months of Pembrolizumab (35 cycles) from Cycle 1 Day 1 (C1D1) can be administered. intervention 2: 20-30 Gy over five fractions for up to two cycles.

Protocol document:
Prot_SAP_000.pdf: 
Clinical Study Protocol 
HCRN CUP16-268 
 
 
Protocol Title 
Single-arm phase 2 study to examine pembrolizumab and concurrent radiation to induce an 
abscopal effect in patients with previously treated carcinoma of unknown primary (CUP16-268) 
 
 
Sponsor Investigator 
Harry H. Yoon MD MHS 
 
Co-Investigators 
Sean S. Park MD PhD, immunotherapy and radiation oncology 
Chris L. Hallemeier MD, radiation oncology 
Haidong Dong MD PhD, immunology 
Sarah E. Kerr, MD pathology 
 
Statistician 
Nathan Foster 
 
Trial Management Provided by  
Hoosier Cancer Research Network, Inc. 
500 N. Meridian, Suite 100 
Indianapolis, IN 46204 
 
 
Trial Supported by  
Merck Sharp & Dohme Corp. (“Merck”) 
 
 
Investigational New Drug (IND) #: IND exempt per FDA 7/19/17 
 
 
 
 
Initial Protocol Version Date: 19SEP2017 
Protocol Amendment Version Date:  
24JAN2018 
 
 
 


Clinical Study Protocol 
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PROTOCOL SIGNATURE PAGE 
 
Protocol title: Single-arm phase 2 study to examine pembrolizumab and concurrent radiation to 
induce an abscopal effect in patients with previously treated carcinoma of unknown primary 
(CUP16-268)  
 
 
VERSION DATE: 24JAN2018 
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  
HOOSIER CANCER RESEARCH NETWORK (HCRN) 
 
 
 


Clinical Study Protocol 
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SYNOPSIS 
TITLE 
Single-arm phase 2 study to examine pembrolizumab and concurrent 
radiation to induce an abscopal effect in patients with previously treated 
carcinoma of unknown primary (CUP16-268) 
SHORT TITLE 
Phase 2 study of pembrolizumab and concurrent radiation in patients with 
previously treated carcinoma of unknown primary 
PHASE  
II 
OBJECTIVES 
Primary Objective: 
Evaluate the abscopal response rate in CUP patients treated with the 
combination of pembrolizumab plus radiotherapy. 
 
Secondary Objectives: 
• Determine the response rate by RECIST 1.1 and informed by 
irRECIST, of non-irradiated metastatic sites when pembrolizumab 
is combined with radiotherapy. 
• Evaluate treatment-related toxicity. 
• Evaluate PFS, OS, TTP, and DCR 
• Explore the association between RR and other endpoints (e.g., OS, 
PFS) 
 
Exploratory Objectives: 
• Evaluate the association of MSI status and mutation load with 
clinical efficacy 
• Evaluate the association of tissue PDL1 expression with clinical 
efficacy 
• Evaluate the association of soluble PD-L1 with clinical efficacy 
• Evaluate changes in the CD11ahigh PD-1high CD8+ T effector 
population in association with pembrolizumab-based therapy 
• Evaluate changes and predictive value of BIM expression in T 
cells in circulation and the predictive value of BIM/PD-1 
expression in tumor infiltrating lymphocytes 
• Evaluate the prognostic and predictive value of tumor tissue of 
origin testing 
 
STUDY 
DESIGN 
This is a proof-of-principle single-arm phase 2 study in patients with 
previously treated CUP.  All patients receive pembrolizumab combined 
with RT to a metastatic site, so as to induce an abscopal tumor response.  
The treatment combination will be repeated with RT delivery to a second 
metastatic site in a non-overlapping RT field.   
 
The results will be compared with historical control.   The primary 
endpoint is the confirmed response rate (RR) in a non-irradiated site based 
on best responding abscopal lesion.  This study will also evaluate the 


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Version Date: 24JAN2018 
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following secondary endpoints: RR in a non-irradiated site based on 
RECIST 1.1, adverse events, progression-free survival (PFS), overall 
survival (OS), time-to-progression (TTP), and disease control rate (DCR). 
 
ELIGIBILITY 
CRITERIA  
Inclusion Criteria 
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 ≤ 2 within 28 days prior to registration. 
 
4. Archival tissue must be available and identified during screening and 
shipped prior to Day -21. If archival tissue is not available and the 
subject is not undergoing a standard of care biopsy, the subject must 
undergo a research biopsy to obtain fresh tissue prior to start of 
treatment. 
 
5. Carcinoma of unknown primary after the following diagnostic 
procedures have been performed if clinically indicated and are 
unrevealing of the primary site:  
• Complete history and clinically appropriate physical  
• CT scan of chest, abdomen, and pelvis 
• Directed evaluation of symptomatic areas  
• Mammogram in women  
• Colonoscopy in patients with liver metastasis or an elevated CEA 
• Direct pathologic comparison with prior tumor specimens, where 
possible, even if prior tumor is early-stage or clinically remote 
from current disease 
 
NOTE: Immunohistochemical stains will be performed according to 
institutional standards.  If a primary carcinoma is identified, the 
patient should undergo treatment as appropriate for that primary 
tumor and not be enrolled in the study.  The above diagnostic workup 
does not need to be performed if: (1) it was previously completed at 
the time of original diagnosis or (2) the investigator does not believe 
the workup has clinical utility at the current time, given that the 
patient has received interval therapy. 
 
6. Histologic confirmation of metastatic adenocarcinoma, poorly 
differentiated non-small cell carcinoma, or poorly differentiated 
squamous carcinoma. NOTE: Pathology consultation at Mayo Clinic 
is recommended if clinically indicated.  One scenario is where 
unknown primary is the most likely diagnosis but immunostains show 


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relatively site-specific marker staining (e.g., CD45, TTF1, 
chromogranin, GATA3, PAX8, PSA, melanocytic markers).   
Information provided for pathology consultation should include 
recent H&P and imaging reports.   
 
7. If and when available, submission of genomic sequencing and 
expression profiling results is mandatory. Results of testing are not 
required prior to treatment. 
 
8. At least one measurable lesion (per RECIST 1.1) outside the planned 
RT fields.   
 
9. Stable or progressive disease after, or was unable to tolerate, at least 
one line of prior anticancer therapy for this disease.  NOTE: For 
patients with stable disease, it is strongly encouraged to confirm the 
presence of active disease (eg, demonstrating FDG avidity via PET or 
repeat biopsy). 
 
10. Radiation oncology consultation at enrolling site ≤ 56 days prior to 
registration to confirm at least two metastatic lesions which are 
targetable by RT at doses and schedule prescribed in this study and 
which reside in non-overlapping RT fields. 
 
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 
Absolute Neutrophil Count (ANC) 
≥ 900 K/mm3 
Hemoglobin (Hgb) 
≥ 8.5 g/dL without transfusion or/ EPO 
dependency (≤ 7 days prior to 
assessment) 
Platelets 
≥ 90,000 / mcL 
Renal 
Creatinine OR 
Calculated creatinine clearance1 
≤ 1.5 X upper limit of normal (ULN) 
OR 
≥ 60 mL/min for subject with 
creatinine levels > 1.5 X institutional 
ULN 
Hepatic 
Bilirubin 
Total bilirubin ≤ 1.5 X ULN OR 
Direct bilirubin ≤ ULN for subjects 
with total bilirubin levels > 1.5 ULN 
OR total bilirubin ≤2 X ULN if liver 
metastases are present 
Aspartate aminotransferase (AST) 
and Alanine aminotransferase 
(ALT) 
≤ 2.5 X ULN OR 
≤ 5 X ULN for subjects with liver 
metastases 
Albumin 
> 2.5 g/dL 


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1 Cockcroft-Gault formula will be used to calculate creatinine clearance (See SPM) 
 
10. Females of childbearing potential must have a negative serum 
pregnancy test within 72 hours prior to registration. NOTE: If the 
urine test is positive or cannot be confirmed as negative, a serum 
pregnancy test will be required. NOTE: breast milk cannot be stored 
for future use while the mother is being treated on study 
 
11. Females of childbearing potential and males must be willing to 
abstain from heterosexual activity (abstinence) or use effective 
methods of contraception as described in Section 5.5 from the time of 
informed consent until 120 days after treatment discontinuation.  
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 
 
12. Willingness to return to the enrolling institution for follow up 
 
13. Willingness to provide tissue and blood samples for correlative 
research purposes 
 
Exclusion Criteria 
14. Prior radiation to an area of the body which, if included in the current 
radiation field, poses an unacceptably high risk of toxicity in the 
opinion of the investigator.  NOTE: A prior field that overlaps with 
the current field, by itself, does not exclude the patient. 
 
15. Any of the following 
• Melanoma.  NOTE: Positive tumor staining for S-100 or HMB45 
alone does not exclude patients. 
• If immunostains are performed, and any of the below tests are 
positive:  
o Hematologic CD45+ (others such as CD2, CD20, CD30, 
CD43 also suggest hematologic origin)  
o Lung or thyroid origin (Thyroid Transcription Factor [TTF-
1]). NOTE: Patients with biopsy proven TTF-1 positive 
tumor who do not have clinical evidence for either lung or 
thyroid cancer (e.g. a dominant lung mass) are still eligible. 
 
16. Progressed on 4 or more lines of prior chemotherapy for this cancer. 
NOTE: Bisphosphonates and neoadjuvant/adjuvant anticancer 
therapies (including locally directed therapies) do not count as a line 
of therapy with regard to this exclusion criteria. 
 
 


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17. Co-morbid systemic illnesses or other severe concurrent disease 
which, in the judgment of the investigator, would make the patient 
inappropriate for entry into this study or interfere significantly with 
the proper assessment of safety and toxicity of the prescribed 
regimens. 
 
18 Is currently participating and receiving study therapy or has 
participated in a study of an investigational agent and received study 
therapy or used an investigational device within 4 weeks of the first 
dose of treatment. 
 
19 Has a diagnosis of immunodeficiency or is receiving systemic steroid 
therapy or any other form of immunosuppressive therapy within 7 
days prior to the first dose of trial treatment. 
 
20 Has received prior therapy with an anti-PD-1, anti-PD-L1, or anti-
PD-L2 agent. 
 
21 Has a known history of Human Immunodeficiency Virus (HIV) (HIV 
1/2 antibodies). 
 
22 Has known active Hepatitis B (e.g., HBsAg reactive) or Hepatitis C 
(e.g., HCV RNA [qualitative] is detected). 
 
23 Has active autoimmune disease that has required systemic treatment 
in the past 2 years (i.e. with use of disease modifying agents, 
corticosteroids or immunosuppressive drugs). Replacement therapy 
(eg., thyroxine, insulin, or physiologic corticosteroid replacement 
therapy for adrenal or pituitary insufficiency, etc.) is not considered a 
form of systemic treatment.  NOTE:  Inhaled steroids or steroid 
injections for joint disease are allowed. 
 
24 Has known active Hepatitis B (e.g., HBsAg reactive) or Hepatitis C 
(e.g., HCV RNA [qualitative] is detected). 
 
25 Prior severe allergic reactions to a monoclonal antibody or 
hypersensitivity to pembrolizumab or any of its excipients. 
 
26 Has had a prior anti-cancer monoclonal antibody (mAb) within 4 
weeks prior to study Day 1 or who has not recovered (i.e., ≤ Grade 1 
or at baseline) from adverse events due to agents administered more 
than 4 weeks earlier. 
 
27 Other active malignancy which requires current treatment and which 
in the opinion of the site investigator is likely to interfere with 
evaluation of disease assessment.  NOTE:  Continuation of hormonal 
therapies is allowed.   


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28. Has known active central nervous system (CNS) metastases and/or 
carcinomatous meningitis.  Subjects with previously treated brain 
metastases may participate provided they are stable (without evidence 
of progression by imaging for at least four weeks prior to the first 
dose of trial treatment and any neurologic symptoms have returned to 
baseline), have no evidence of new or enlarging brain metastases, and 
are not using steroids for at least 7 days prior to trial treatment.  This 
exception does not include carcinomatous meningitis which is 
excluded regardless of clinical stability.  
 
29 Major surgery ≤ 4 weeks from registration.  NOTE: Diagnostic 
laparoscopy (without other intervention) and/or biopsies (needle 
aspirate, core biopsy, open biopsy, etc…) are not considered major 
surgery 
 
30 Uncontrolled intercurrent illness which in the opinion of the 
investigator poses unacceptably high risk when combined with study 
treatment, including but not limited to the following: 
• Symptomatic congestive heart failure 
• Unstable angina pectoris 
• Severely impaired lung function 
• Known history of active TB (Bacillus Tuberculosis) 
• Uncontrolled diabetes as defined by fasting serum glucose >1.5 x 
ULN (NOTE: Optimal glycemic control should be achieved 
before starting trial therapy.) 
• Significant underlying liver disease such as cirrhosis or severe 
hepatic impairment 
• Has known psychiatric or substance abuse disorders that would 
interfere with cooperation with the requirements of the trial. 
 
31 Has an active infection requiring systemic therapy. 
 
32 Has a history or current evidence of any condition, therapy, or 
laboratory abnormality that might confound the results of the trial, 
interfere with the subject’s participation for the full duration of the 
trial, or is not in the best interest of the subject to participate, in the 
opinion of the treating investigator.  
 
33 Is pregnant or breastfeeding, or expecting to conceive or father 
children within the projected duration of the trial, starting with the 
pre-screening or screening visit through 120 days after the last dose 
of trial treatment. 
 
 
 
 


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34 For patients in whom planned RT fields will include the heart, any of 
the following heart conditions, if in the opinion of the investigator 
they pose unacceptably high risk when combined with study 
treatment: 
• Prior symptomatic congestive heart failure 
• Documented myocardial infarction ≤6 months prior to registration 
(pretreatment ECG evidence of infarct only will not exclude 
patients) 
• Prior significant ventricular arrhythmia requiring medication 
• Prior 2nd or 3rd degree heart block or other types of clinically 
significant conduction delay ≤ 6 months prior to registration 
• Clinically significant pericardial disease (including pericardial 
effusion, pericarditis) or cardiac valvular disease ≤12 months 
prior to registration 
 
NOTE: As part of history and physical, all patients must be assessed 
for signs or symptoms of cardiac disease, or for prior history of 
cardiac disease. These conditions include but are not limited to 
diseases related to cardiac valves, pericardium, myocardium, 
atrioventricular delays or arrhythmias. It is strongly recommended 
that signs or symptoms of potentially clinically significant disease be 
evaluated with comprehensive cardiac echo. 
 
35 For patients in whom planned RT fields during the study will include 
the chest, any of the following, if in the opinion of the site 
investigator they pose unacceptably high risk when combined with 
study treatment:  
• Prior fistula within thorax, including bronchoalveolar or 
esophageal. 
• Respiratory condition that required oxygen supplementation ≤ 
3 months prior to registration 
• Clinically significant pulmonary hypertension ≤ 12 months prior 
to registration 
• Pneumonia requiring treatment ≤ 1 month prior to registration 
• Pulmonary embolism requiring treatment ≤ 6 months prior to 
registration 
• Pleural effusion requiring drainage ≤ 12 months prior to 
registration 
 
36 Has known history of or any evidence of active, non-infectious 
pneumonitis 
 
37 Currently uncontrolled hyper/hypothyroidism or hyper/hypocortism if 
in the opinion of the investigator they pose unacceptably high risk 
when combined with study treatment 
 


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38 Received live vaccine ≤ 30 days prior to registration. NOTE:  
Seasonal influenza vaccines for injection are generally inactivated flu 
vaccines and are allowed; however intranasal influenza vaccines 
(e.g., Flu-Mist®) are live attenuated vaccines, and are not allowed. 
 
TOTAL  
NUMBER OF 
SUBJECTS 
N = 34 
ESTIMATED 
ENROLLMEN
T PERIOD 
Estimated 23 months 
ESTIMATED 
STUDY 
DURATION  
Estimated 36 months 
 
 


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TABLE OF CONTENTS 
 
Schema ............................................................................................................................. 12 
1. 
Background and Rationale.................................................................................... 13 
2. 
Study Objectives and Endpoints........................................................................... 21 
3. 
Eligibility Criteria .................................................................................................. 21 
4. 
Subject Registration............................................................................................... 26 
5. 
Treatment Plan....................................................................................................... 27 
6. 
Toxicities and Dose Delays/Dose Modifications .................................................. 34 
7. 
Study Calendar & Evaluations ............................................................................. 43 
8. 
Biospecimen Studies and Procedures ................................................................... 45 
9. 
Criteria for Disease Evaluation ............................................................................ 50 
10. Drug Information ................................................................................................... 59 
11. Adverse Events ....................................................................................................... 61 
12. Statistical Methods ................................................................................................. 66 
13. Trial Management ................................................................................................. 69 
14. Data Handling and Record Keeping .................................................................... 70 
15 
Ethics ....................................................................................................................... 71 
16 
References ............................................................................................................... 73 
 
 
 
 
 


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SCHEMA 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Screening 
Registration 
Induction 
Pembrolizumab (Day -21) 
 
Cycle 1 (21 days) 
Pembrolizumab (Day 1) 
plus 
RT to 1st metastatic site (Day 1-5) 
 
Cycle 2 (21 days) 
Pembrolizumab  
plus 
RT to 2nd metastatic site (Day 1-5) 
 
Maintenance (21 days) 
Pembrolizumab (Day 1) 
for up to 22 months from C1D1 
Continue maintenance until: 
Progressive Disease 
Unacceptable Adverse Event 
Patient Refusal 


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1. 
BACKGROUND AND RATIONALE 
 
1.1 
Cancer of unknown primary (CUP) is an area of unmet need 
In the U.S., approximately 31,430 cases of CUP are diagnosed per year (ACS, 2015).  CUP is 
defined as biopsy-proven metastasis in the absence of an identifiable primary site.  While no 
standard 1st-line regimen exists, platinum-, taxane-, or gemcitabine-based therapies, such as 
carboplatin/paclitaxel, are most commonly used.  In clinical trials, including those led by Mayo 
(Holtan et al., 2012; Yoon et al., 2016), median overall survival (OS) from standard first-line 
therapies is 7-12 months.  OS in the 2nd-line setting is 3-5 months. 
 
1.1.1 Barriers in drug development in CUP   
• Trials in CUP examining various cytotoxic chemotherapy combinations, or biologic 
agents in molecularly unselected populations, have failed to improve outcomes.   
• An alternative approach is to test novel agents within subgroups possessing a specific 
targetable alteration.  However, trials which further shrink the study population in this 
way are not feasible, due to the low overall incidence of CUP, and the low frequency 
(<5% typically) of most potentially targetable genomic alterations in CUP (Ross et al., 
2015).   
• Alternatively, CUP patients may be “piggybacked” onto larger “basket” trials (e.g., M-
PACT, MATCH) in which the same agent is given to patients from diverse tumor types 
sharing a specific alteration (e.g., BRAF mutation).  However, it is unclear whether such 
data will be generalizable to CUP patients, who are likely to be poorly represented.  The 
complex cross talk and signaling pathways may differ among tumor types, as suggested 
by the lack of single-agent efficacy of vemurafenib in BRAF-mutated melanoma vs 
BRAF-mutated colorectal cancer (Chapman et al., 2011; Mao et al., 2013).   
• In addition, >15% of CUP patients do not have a genomic alteration that is potentially 
targetable with available therapies (Ross et al., 2015).   
 
The current phase 2 trial attempts to overcome these barriers through the use of an innovative 
small trial based on recent data suggesting that immunotherapy in combination with radiation 
may induce an abscopal effect that can improve patient outcomes (Golden et al., 2015).  The trial 
also attempts to take advantage of recent unpublished data that CUPs may have a relative high 
total mutation burden compared to other malignancies.  Total mutation burden is a putative 
biomarker that predicts enhanced efficacy to anti-PD-1 therapy (Hodi et al., 2016; Rizvi et al., 
2015).  
 
1.2 
Rationale for Therapeutic Approach 
 
1.2.1 Trial Design 
In this phase 2 study, patients with previously treated CUP will receive pembrolizumab in 
combination with concurrent RT, so as to induce an abscopal tumor response.  The treatment 
combination will be repeated with RT delivery to a second metastatic site in a non-overlapping 
RT field. 
 


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This trial design is modeled after the proof-of-principle trial examining GM-CSF plus RT in 
advanced solid tumors (see Clinical data for abscopal effect, below), in which RT to metastatic 
sites was delivered 1 week after administration of GM-CSF (Golden et al., 2015). 
 
1.2.2 Abscopal Effect 
 
1.2.2.1 
Pre-Clinical Data for Abscopal Effect 
Recent data, including from lab-based Mayo Clinic collaborators in the current trial (Park et al., 
2015), have generated the hypothesis that ionizing radiation (RT) can increase the production 
and presentation of tumor antigens in poorly immunogenic tumors.  This, in turn, could augment 
the antitumor immune responses elicited by anti–PD-1/PD-L1 agents (Demaria et al., 2005; Reits 
et al., 2006; Zhang et al., 2007).  The combination of anti-PD-L1 plus RT has been shown to 
induce an abscopal effect on non-irradiated distant tumors in a CD8-dependent, tumor antigen-
specific manner (Demaria et al., 2005; Deng et al., 2014).  RT can induce tissue PD-L1 
expression, a predictive biomarker of PD-1 blockade (Ansell et al., 2015; Herbst et al., 2014; 
Powles et al., 2014; Taube et al., 2014; Topalian et al., 2012; Topalian et al., 2014).  As shown 
by our collaborators, the combination of RT plus anti-PD-1 therapy elicited a 66% reduction in 
size of nonirradiated, secondary tumors outside the RT field (abscopal effect), compared to RT 
or PD-1 blockade alone (Park et al., 2015).   
 
1.2.2.2 
Clinical data for abscopal effect 
A recent proof-of-principle clinical trial from New York University (NYU) was performed in 
patients with stable or progressing metastatic solid tumors (n=41, mostly lung and breast cancer) 
(Golden et al., 2015).  Patients were treated with a combination of RT (35 Gy over 10 fractions 
to one metastatic site, repeated to a second metastatic site) plus GM-CSF.   RT to metastatic sites 
was delivered 1 week after administration of GM-CSF.  GM-CSF (granulocyte-macrophage 
colony-stimulating factor) is a potent immune adjuvant to dendritic cell maturation that has been 
shown to induce the differentiation of dendritic cells from bone marrow precursor cells (Blom et 
al., 2000; Inaba et al., 1992).  GM-CSF by itself is not expected to have a significant contributory 
role to the responses observed in the trial.  The primary endpoint was the abscopal response rate, 
defined as the proportion of patients with a response in the best responding abscopal lesion.  An 
abscopal response was defined as a decrease in the longest diameter of at least 30% in any 
measurable (≥1 cm) non-irradiated lesion from baseline.  In patients with more than three lesions, 
the non-irradiated lesions were measured individually for response to treatment.  The best 
abscopal responding lesion was reported.  The primary endpoint was achieved in 27% of 
patients.  As the investigators from this trial concluded, optimization of the strategy probably 
needs integration of many distinct and sequential signals to correct the preexisting immune 
suppression or tolerance of metastatic tumors. Whereas RT with GM-CSF actively recruits an 
inflammatory response within the irradiated field, T-cell exhaustion remains an obstacle for 
long-term anti-tumor immunity because acquired resistance to RT was shown to be mitigated by 
PD-1/PD-L1 axis blockade (Dovedi et al., 2014; Twyman-Saint Victor et al., 2015).  
 
These data represent a promising approach of utilizing RT to act as an “in-situ vaccine” to 
augment anti-tumor activity of immune blockade, including with anti-PD-1 antibodies.  This 
approach has not been examined in CUP and forms the framework for our study. 
 


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1.2.2.3 
Abscopal Response Rate 
Importantly, primary endpoint in the NYU trial was the abscopal response rate (defined above), 
not RECIST 1.1.  However, interestingly, a significantly better overall survival was noted in the 
patients with abscopal responses compared with those without an abscopal response: median 
overall survival was 21.0 (95% CI 11.1 - 31.0) months for abscopal responders vs 8.3 (95% CI 
5.0 – 13.3) months for non-responders (HR 2.06, 95% CI 1.04 – 4.11).  Median follow up was 
5.6 years.  A correlation between abscopal response rate and response per RECIST 1.1 was not 
reported.     
 
The abscopal response rate, as defined in the NYU study, does not distinguish between nodal and 
non-nodal lesions.  In our study, we will use RECIST criteria for defining a measurable or target 
lesion, which distinguishes between nodal and non-nodal lesions. 
 
The limitations of standard RECIST in measuring benefit from immunotherapies are increasingly 
recognized.  A significant number of patients have mixed responses (ie, some lesions shrink 
while others enlarge or remain stable).  It is possible that the abscopal response rate may be a 
better predictor of long-term outcome than RECIST 1.1 when studying the combination of PD-1 
inhibition and RT.  The risk of relying on RECIST 1.1 alone is that potentially benefitting 
patients may be overlooked.  Our purpose in this small phase 2 trial is to examine this issue in 
more detail.  In addition, our trial will use irRECIST for determination of progression. 
 
1.2.3 Radiation as Standard of Care 
The dose and fractionation of radiation planned within the current study is an accepted standard 
for the treatment of metastatic lesions in CUP patients, as supported by the NCI (NCI, 2016).  
RT in combination with pembrolizumab is hypothesized to enhance an abscopal effect (see 
above, abscopal effect). 
 
1.2.3.1 
Fractionated (vs single dose) Radiotherapy has Greater Synergy with Immune 
Therapy  
Dewan et al tested the hypothesis that the type of dose fractionation regimen of RT can 
determine the ability of RT to synergize with anti-CTLA-4 antibody(Dewan et al., 2009). In a 
mouse model where colon or breast cancer cells were injected into both flanks, single-dose or 
fractionated RT led to growth delay of the primary tumor, regardless of whether anti-CTLA-4 
antibody was administered, but had no effect on secondary tumors outside the RT field. 
However, an abscopal effect on the secondary tumor outside the RT field only occurred in mice 
treated with the combination of antibody plus fractionated RT, not single-dose RT. 
 
1.2.4 Induction Dose of Pembrolizumab on Day -21 
A major gap in knowledge in the field is the optimal timing of checkpoint inhibition in relation 
to RT and/or chemotherapy other therapies in the induction of an abscopal effect.  The available 
data suggests that the timing of the first dose of checkpoint inhibition in relation to other 
therapies may be critical (Dewan et al., 2009), including data from randomized controlled trials 
comparing the initiation of checkpoint inhibition concurrently vs non-concurrently with 
chemotherapy(Lynch et al., 2012; Reck et al., 2013).  Dewan et al. found that delaying the 
initiation of anti-CTLA-4 antibody until a few days after the completion of RT reduced the 
therapeutic effect on the primary and secondary (non-irradiated) tumor, as compared to initiating 


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antibody treatment on the first or final day of RT (RT was given in 3 fractions of 6 Gy each) 
(Dewan et al., 2009).  To date, no study has directly compared the initiation of checkpoint 
blockade on the same day as RT initiation vs prior to RT initiation. 
 
Some data support the initiation of checkpoint blockade on the same day as RT.  Deng et al 
demonstrated an abscopal effect from the combination of PD-L1 blockade and RT, and PD-L1 
blockade (administered every 3 days for a total of 4 times) was initiated on the day of IR or 1 day 
before IR (Deng et al., 2014).  However, the initiation of PD-1 axis blockade on the same day as 
(vs prior to) RT/chemotherapy can theoretical have a detrimental effect: cytotoxic T cells which 
become newly primed and recruited via anti-PD-1 therapy may be destroyed by RT and/or 
chemotherapy.  Deng et al did not directly compare different administration schedules of PD-
1/L1 with regard to RT.   
 
Other data or concepts support the initiation of checkpoint blockade prior to RT: 
• Collaborators in the current trial demonstrated an abscopal effect from the combination of 
PD-1 blockade and RT, and pharmacologic PD-1 blockade was initiated one day prior to 
RT (Park et al., 2015).  While Park et al did not directly compare different administration 
schedules of PD-1 Ab with regard to RT, the investigators found a similar abscopal effect 
in PD-1 knockout models, suggesting that baseline inhibition of PD-1 can induce an 
abscopal response. 
• Administration of carboplatin/paclitaxel in patients requires pre-medication with 
corticosteroids to reduce the likelihood and severity nausea/vomiting and hypersensitivity 
reactions.  However, the well-known immunosuppressive effect of corticosteroids, 
despite their short half-life, may blunt the effect of immune checkpoint inhibition.  The 
administration of pembrolizumab prior to the administration of cytotoxic agents may 
avoid this issue. 
 
1.3 
CUP Patient Population 
In addition to the poor prognosis of CUP patients despite treatment, CUP was selected because 
these tumors exhibit molecular characteristics that may increase their susceptibility to PD-1 
blockade.  Recent examination of 70 CUP cases revealed PDL1-expressing cancer cells in 21% 
of tumors (Gatalica et al., 2014), a frequency comparable to that found in other tumor types 
where single-agent PD-1 therapy has shown activity.  A high mutation load was detected in 13% 
of tumors of unknown primary; this frequency was the 16th highest out of >50 tumor types 
profiled using targeted next generation sequencing of >300 cancer genes (personal 
communication, unpublished data).  A high mutation load has been shown to correlate with 
pembrolizumab activity in NSCLC (Rizvi et al., 2015). 
 
At most cancer centers with a high volume of CUP patients, including at the Mayo Clinic, 
patients generally start standard chemotherapy with their local community oncologist prior to 
being seen at the larger center.  This referral pattern is primarily for logistic reasons – i.e., 
inability to secure an immediate appointment after the completion of the diagnostic workup, 
which itself can be time-consuming. 
 
 
 


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Accordingly, the vast majority of CUP patients seen at large centers have already initiated first-
line therapy.  While there is no standard front-line regimen, platinum-, taxane-, and/or 
gemcitabine combination regimens are common, and one regimen has not been shown to be 
superior compared to another.   
 
The rationale in the current study is to take advantage of this referral pattern by allowing the 
enrollment of patients who have started front-line therapy. 
 
1.4 
Potential Safety Concerns 
 
1.4.1 Pembrolizumab 
For pembrolizumab monotherapy, in clinical trials completed to date, most immune related 
adverse events associated with pembrolizumab were mild (grade 1–2), some were severe, and a 
few were lethal.   Refer to the Investigator’s Brochure (IB) for detailed safety information on 
MK-3475.  In brief, the most recent IB (updated 2/17/2017) presents safety data available from 
the Sponsor’s Reference Safety Dataset (n=2799), a locked and verified dataset with pooled data 
from clinical trials in melanoma and NSCLC, as part of the Sponsor’s product development of 
pembrolizumab. The Reference Safety Dataset comprises safety data from locked KN001, 
KN002, KN006, and KN010 [Sec. 1]. The Reference Safety Dataset is the dataset from which 
frequencies in this IB are derived.  The 5 most frequently reported AEs were: fatigue (37.3%), 
nausea (24.5%), decreased appetite (22.5%), diarrhea (22.3%), and cough (22%).  The 5 most 
frequently reported serious AEs (SAE) were pneumonia (3.0%), pleural effusion (1.7%), 
pneumonitis (1.6%), dyspnea (1.6%) and pulmonary embolism (1.5%).  The 5 most frequently 
reported AEs considered drug-related by the investigator were fatigue (24.2%), pruritus (16.7%), 
rash (13.8%), diarrhea (12.3%), and nausea (10.9%). The 5 most frequently reported SAEs 
considered drug-related by the investigator were pneumonitis (1.6%), colitis (0.9%), diarrhea 
(0.6%), pyrexia (0.4%), and autoimmune hepatitis (0.3%). 
 
1.4.2 Combination of Pembrolizumab and RT 
In the current trial, because RT may be given to lung or liver metastases, potential lung and liver 
toxicities of pembrolizumab are described further here: 
• Pneumonitis in association with pembrolizumab (according to IB 22-May-2015):   
o Frequency/grade:  Pneumonitis was quantitatively assessed using the following 
adverse events of special interest (AEOSI) preferred terms: acute interstitial 
pneumonitis, interstitial lung disease, pneumonitis, and idiopathic pneumonia 
syndrome. Out of a total of 1562 subjects with melanoma or lung cancer (P001 
and P002) treated with pembrolizumab (2 mg/kg q3w, 10 mg/k2 q3 w, or 10 
mg/k2 q2w), AEOSI of pneumonitis were identified in 45 subjects (2.9%), of 
which 42 (2.7%) were considered drug related. Serious AEOSI were identified in 
1.2% of subjects, and all serious AEOSI were considered by the Investigators to 
be drug related. One subject died due to pneumonitis (interstitial lung disease, 
dose level 10 mg/k2 q3w)). By worst toxicity grade, 14 subjects (0.9%) 
experienced Grade 1 pneumonitis, 15 subjects (1.0%) experienced Grade 2 
pneumonitis, 11 subjects (0.7%) experienced Grade 3 pneumonitis, and 2 
subjects (0.1%) experienced Grade 4 pneumonitis. For “interstitial lung disease,” 
1 subject (0.1%) experienced Grade 1 interstitial lung disease, 1 subject (0.1%) 


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experienced Grade 3 interstitial lung disease, and 1 subject (0.1%) experienced 
Grade 5 interstitial lung disease.   
o Time to development of pneumonitis:  Data are available from the 38 patients 
with previously-treated NSCLC treated with MK-3475 10mg/kg every 3 weeks 
in P001, of whom 2 patients have been reported with pneumonitis and one 
patient has been reported with steroid-responsive pulmonary edema. Two cases 
were grade 2; the case of pulmonary edema was grade 3. These cases began as 
early as four days after the first dose to several cycles (Week 18) after the first 
dose.  
• Hepatic abnormalities in association with pembrolizumab (according to IB 22-May-
2015):  Hepatic AEOSI was quantitatively assessed using the following AEOSI 
preferred terms: hepatitis, autoimmune hepatitis, hepatitis acute, hepatitis fulminant and 
drug-induced liver injury. Out of a total of 1562 subjects, hepatic AEOSI were identified 
in 8 subjects (0.5%). All the hepatic AEOSI were considered drug-related. Serious 
hepatic AEOSI were identified in 2 subjects (0.1%); both were considered by the 
Investigators to be drug related.  By worst toxicity grade, hepatitis was reported for 2 
subjects (0.1%). The 2 instances were Grade 1 and Grade 3. Autoimmune hepatitis was 
reported for 6 subjects (0.4%). Two subjects (0.1%) experienced Grade 2 autoimmune 
hepatitis, 3 subjects (0.2%) experienced Grade 3 autoimmune hepatitis, and 1 subject 
(0.1%) experienced Grade 4 autoimmune hepatitis. 
 
Delivery of RT to lung or liver lesion(s) in the current trial may theoretically cause an 
exacerbation of organ-specific toxicities and thus are considered in more detail as follows. 
 
Radiation pneumonitis has been reported in patients who have undergone mediastinal radiation 
therapy for lung cancer, Hodgkin’s lymphoma (HL), breast cancer, and other cancers that require 
radiation therapy to the thorax (Carver et al., 2007). A retrospective analysis of 1,911 patients 
who underwent combined-modality therapy for lung cancer demonstrated that the overall risk of 
radiation pneumonitis was 7.8% (Roach et al., 1995). Clinical factors that may increase the risk 
of radiation pneumonitis include concomitant chemotherapy, previous irradiation, and recent 
withdrawal of steroids (McDonald et al., 1995). Patient factors, such as lung function, age, and 
sex, do not adequately select patients at high risk for radiation pneumonitis or fibrosis (Dehing-
Oberije et al., 2009).   Radiation pneumonitis is typically a delayed acute reaction, usually 
occurring 1 to 3 months after completing mediastinal radiation therapy (McDonald et al., 1995). 
Patients who experience acute radiation pneumonitis will often have a self-limited course, with 
complete resolution of this process. However, a minority of patients may develop progressive 
pulmonary fibrosis, usually 6 to 24 months after treatment. Late complications of pulmonary 
fibrosis include cor pulmonale and respiratory failure (Carver et al., 2007).  Both the V20 (ie, the 
percentage volume of both lungs minus the planning target volume [PTV] receiving 20 Gy) and 
the mean lung dose (MLD) correlate with the risk for radiation pneumonitis (De Ruysscher et al., 
2010). In a landmark study (Graham et al., 1999), V20 was the single independent predictor of 
pneumonitis in a multivariate analysis. In this study, 42% of the patients received concurrent 
chemotherapy, and the incidence of grade >=2 pneumonitis was 7% when the V20 was 22 – 
31%. With the radiation doses we are proposing in this study, we expect V20 to be < 20% and 
therefore, the risk of radiation-related pneumonitis will be minimal. However, as a precaution to 


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reduce risk for thoracic toxicities following RT plus pembrolizumab, the following measures will 
be implemented: 
• Strict limits on normal tissue doses and other aspects of radiation delivery will be 
mandated per the most updated standard of care and as recommended(Tang et al., 
2014).  
• Patients with recent serious pulmonary illness will be excluded (see section 3). 
• Monitoring will include late lung toxicities, given that pneumonitis often does not 
appear until several months after radiation treatment is completed. 
 
Radiation-Induced Liver Disease (RILD) usually develops 1-2 months after RT and describes a 
clinical syndrome composed of fatigue, ascites, elevated LFTs and abdominal pain. Emami et al 
(Emami et al., 1991) estimated the acceptable liver tolerance dose (TD5/5; tolerance dose that 
may cause 5% complication rate within 5 years) of 3000, 3500, and 5000 cGy when whole, 2/3, 
and 1/3 of the liver was radiated, respectively. This task force estimation is limited by the 2D 
treatment technique employed at the time. The Quantitative Analysis of Normal Tissue Effects in 
the Clinic (QUANTEC) (Marks et al., 2010) provides the more recent guidelines using the 3D 
dose/volume/outcome data. In this analysis, preexisting liver dysfunction (Child-Pugh B or C), 
mean liver dose and V30 (volume receiving 30 Gy or higher) were associated with RILD (Pan et 
al., 2010).  Accordingly, patients with primary liver tumors have a lower radiation dose/volume 
threshold for developing RILD than patients with liver metastases. RILD data for partial liver 
radiation has been studied using the Lyman-Kutcher-Burman model by the investigators at the 
University of Michigan (Dawson et al., 2002).  In this study, the rate of RILD grade 3 or higher 
within 4 months in 203 patients treated with 3D conformal radiation and concurrent hepatic 
arterial chemotherapy were analyzed. Although some patients received 90 Gy, no RILD was 
noted when the mean liver dose was <31 Gy. Therefore, in the current study, we will use the 
following liver constraints as described in QUANTEC to minimize liver toxicity: mean liver <= 
30 Gy and >=700 ml of normal liver to receive <=15 Gy (which is a stricter constraint used for 5 
fraction SBRT regimen). 
 
The safety and tolerability of lung/liver RT in combination of pembrolizumab has not been 
extensively examined and, with the implementation of appropriate safeguards (see below: 
Eligibility, Study Design), is a focus of the current trial.  The combination of pembrolizumab + 
concurrent radio(chemo)therapy is being examined in multiple trials in other tumor types: 
• Pembrolizumab, Combination Chemotherapy, and Radiation Therapy Before 
Surgery in Treating Adult Patients with Locally Advanced Gastroesophageal 
Junction or Gastric Cardia Cancer That Can Be Removed by Surgery 
NCT02730546 – This study is led by the PI of the current study (Yoon HH) 
o Tumor: locally advanced gastroesophageal junction (GEJ) adenocarcinoma 
o Treatment: carboplatin/paclitaxel weekly for 5 weeks plus pembrolizumab every 
3 weeks, plus concurrent RT 
o Location/schedule of RT:  RT to GEJ at 41 Gy over 23 fractions 
o Estimated enrollment, N = 30 
o Dates study start, primary completion: Aug 2016, April 2018 
o Primary Outcome: Response 
 
 


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• Phase I Trial of MK-3475 and Concurrent Chemo/Radiation for the Elimination of Small 
Cell Lung Cancer (NCT02402920; for brevity, only “Part B” for extensive stage is 
shown):   
o Tumor: extensive SCLC (ECOG PS 0-2)  
o Treatment: Cis(carbo)platin/etoposide Days 1-3 (every 3 weeks) for up to 4 
cycles, plus pembrolizumab on Day 1, plus concurrent RT 
o Location/schedule of RT: RT to chest at 45 Gy once a day for 15 days.   
o Estimated enrollment, N = 80 
o Dates study start, primary completion: July 2015, July 2023 
o Primary Outcome: MTD of MK 3475. 
• A Randomized Multicenter Phase Ib/II Study to Assess the Safety and the 
Immunological Effect of Chemoradiation Therapy (CRT) in Combination with 
Pembrolizumab (MK-3475) Compared to CRT Alone in Patients with Resectable or 
Borderline Resectable Pancreatic Cancer (UVA-PC-PD101; NCT02305186):   
o Tumor: Resectable or borderline resectable pancreatic cancer (ECOG PS 0-1) 
o Treatment: 
▪ Experimental arm: Neoadjuvant capecitabine (825 mg/m2 orally twice 
daily, M-F on days of RT) and RT plus pembrolizumab 200 mg IV every 
3 weeks on days 1, 22, and 43 during concurrent chemoradiation.   
▪ Control arm: Chemoradiation alone. 
o Location/schedule of RT: RT to pancreas at 50.4 Gy in 28 daily fractions 
o Estimated enrollment, N = 56  
o Dates study start, primary completion: March 2015, June 2017).   
o Primary Outcome: Number of Tumor Infiltrating Lymphocytes (TILs) per high 
powered field (hpf) in resected pancreatic tissue and incidence of DLTs. 
• Phase I/II Trial of MK-3475 and Hypofractionated Stereotactic Radiation Therapy in 
Patients with Non-Small Cell Lung Cancer (NSCLC) (NCT02444741) 
o Tumor: metastatic NSCLC 
o Treatment – phase 1:  Pembrolizumab (starting at 100 mg) on Day 1 of each 3-
week cycle + SBRT to lung (50 Gy, 4 fractions)  
o Treatment – phase 2: Pembrolizumab (at MTD determined in phase 1) + SBRT, 
or pembrolizumab + wide-field radiation therapy (WFRT). 
o Location/schedule of RT: SBRT to lung at 50 Gy over 4 fractions, or WFRT to 
lung at 45 Gy in 15 daily fractions 
o Estimated enrollment, N = 104 
o Dates study start, primary completion: Sept 2015, Sept 2020 
o Primary Outcome: MTD (phase 1), PFS/response (phase 2)  
 
Reported results from the above studies will be followed to inform potential modifications, if 
appropriate, in the current trial. 
 
 
 


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1.6 
How Study Results Might Impact Future Trials or Practice 
This proof-of-principle study is intended to demonstrate that the combination of pembrolizumab 
with RT has the potential to induce an abscopal response.  If results are promising, the question 
of whether this therapeutic approach enhances anti-tumor activity through an abscopal response 
can be confirmed in future randomized trials.  A future trial in the 2nd-line setting may randomize 
patients to placebo vs placebo plus pembrolizumab vs pembrolizumab plus RT. 
 
2. 
STUDY OBJECTIVES AND ENDPOINTS 
 
2.1 
Objectives 
 
2.1.1 Primary Objective 
Evaluate the abscopal response rate in CUP patients treated with the combination of 
pembrolizumab plus radiotherapy. 
 
2.1.2 Secondary Objectives 
• Determine the response rate by RECIST 1.1 and informed by irRECIST, of non-
irradiated metastatic sites when pembrolizumab is combined with radiotherapy. 
• Evaluate treatment-related toxicity. 
• Evaluate PFS, OS, TTP, and DCR 
• Explore the association between RR and other endpoints (e.g., OS, PFS) 
 
2.1.3 Correlative/Exploratory Objectives 
• Evaluate the association of MSI status and mutation load with clinical efficacy 
• Evaluate the association of tissue PDL1 expression with clinical efficacy 
• Evaluate the association of soluble PD-L1 with clinical efficacy 
• Evaluate changes in the CD11ahigh PD-1high CD8+ T effector population in association 
with pembrolizumab-based therapy 
• Evaluate changes and predictive value of BIM expression in T cells in circulation and the 
predictive value of BIM/PD-1 expression in tumor infiltrating lymphocytes 
• Evaluate the prognostic and predictive value of tumor tissue of origin testing 
 
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 ≤ 2 within 28 days prior to registration. 
 


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4. Archival tissue must be available and identified during screening and shipped prior to 
Day -21. If archival tissue is not available and the subject is not undergoing a standard of 
care biopsy, the subject must undergo a research biopsy to obtain fresh tissue prior to 
start of treatment. 
 
5. Carcinoma of unknown primary after the following diagnostic procedures have been 
performed if clinically indicated and are unrevealing of the primary site:  
• Complete history and clinically appropriate physical  
• CT scan of chest, abdomen, and pelvis 
• Directed evaluation of symptomatic areas  
• Mammogram in women  
• Colonoscopy in patients with liver metastasis or an elevated CEA 
• Direct pathologic comparison with prior tumor specimens, where possible, even if 
prior tumor is early-stage or clinically remote from current disease 
 
NOTE: Immunohistochemical stains will be performed according to institutional 
standards.  If a primary carcinoma is identified, the patient should undergo treatment as 
appropriate for that primary tumor and not be enrolled in the study.  The above diagnostic 
workup does not need to be performed if: (1) it was previously completed at the time of 
original diagnosis or (2) the investigator does not believe the workup has clinical utility at 
the current time, given that the patient has received interval therapy. 
 
6. Histologic confirmation of metastatic adenocarcinoma, poorly differentiated non-small 
cell carcinoma, or poorly differentiated squamous carcinoma. NOTE: Pathology 
consultation at Mayo Clinic is recommended if clinically indicated.  One scenario is 
where unknown primary is the most likely diagnosis but immunostains show relatively 
site-specific marker staining (e.g., CD45, TTF1, chromogranin, GATA3, PAX8, PSA, 
melanocytic markers).  Information provided for pathology consultation should include 
recent H&P and imaging reports.   
 
7. If and when available, submission of genomic sequencing and expression profiling results 
is mandatory. Results of testing are not required prior to treatment. 
 
8. At least one measurable lesion (per RECIST 1.1) outside the planned RT fields.    
 
9. Stable or progressive disease after, or was unable to tolerate, at least one line of prior 
anticancer therapy for this disease.  NOTE: For patients with stable disease, it is strongly 
encouraged to confirm the presence of active disease (eg, demonstrating FDG avidity via 
PET or repeat biopsy). 
 
10. Radiation oncology consultation at enrolling site ≤ 56 days prior to registration to 
confirm at least two metastatic lesions which are targetable by RT at doses and schedule 
prescribed in this study and which reside in non-overlapping RT fields. 
 
 


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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 
Absolute Neutrophil Count (ANC) 
≥ 900 K/mm3 
Hemoglobin (Hgb) 
≥ 8.5 g/dL without transfusion or EPO 
dependency (≤ 7 days prior to assessment) 
Platelets 
≥ 90,000 / mcL 
Renal 
Creatinine OR 
Calculated creatinine clearance1 
≤ 1.5 X upper limit of normal (ULN) OR 
≥ 60 mL/min for subject with creatinine 
levels > 1.5 X institutional ULN 
Hepatic 
Bilirubin 
Total bilirubin ≤ 1.5 X ULN OR 
Direct bilirubin ≤ ULN for subjects with 
total bilirubin levels > 1.5 ULN OR total 
bilirubin ≤2 X ULN if liver metastases are 
present 
Aspartate aminotransferase (AST) and 
Alanine aminotransferase (ALT) 
≤ 2.5 X ULN OR 
≤ 5 X ULN for subjects with liver 
metastases 
Albumin 
> 2.5 g/dL 
1 
Cockcroft-Gault formula will be used to calculate creatinine clearance (See SPM) 
 
12. Females of childbearing potential must have a negative serum pregnancy test within 72 
hours prior to registration. NOTE: If the urine test is positive or cannot be confirmed as 
negative, a serum pregnancy test will be required. NOTE: breast milk cannot be stored 
for future use while the mother is being treated on study 
 
13. Females of childbearing potential and males must be willing to abstain from heterosexual 
activity (abstinence) or use effective methods of contraception as described in Section 5.5 
from the time of informed consent until 120 days after treatment discontinuation.  
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 
 
14. Willingness to return to the enrolling institution for follow up 
 
15. Willingness to provide tissue and blood samples for correlative research purposes 
 
3.2 
Exclusion Criteria 
Subjects meeting any of the criteria below may not participate in the study: 
 
16. Prior radiation to an area of the body which, if included in the current radiation field, 
poses an unacceptably high risk of toxicity in the opinion of the investigator.  NOTE: A 
prior field that overlaps with the current field, by itself, does not exclude the patient. 
 
 
 


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17. Any of the following 
• Melanoma.  NOTE: Positive tumor staining for S-100 or HMB45 alone does not 
exclude patients. 
• If immunostains are performed, and any of the below tests are positive:  
o Hematologic CD45+ (others such as CD2, CD20, CD30, CD43 also 
suggest hematologic origin)  
o Lung or thyroid origin (Thyroid Transcription Factor [TTF-1]). NOTE: 
Patients with biopsy proven TTF-1 positive tumor who do not have 
clinical evidence for either lung or thyroid cancer (e.g. a dominant lung 
mass) are still eligible. 
 
18. Progressed on 4 or more lines of prior chemotherapy for this cancer. NOTE: 
Bisphosphonates and neoadjuvant/adjuvant anticancer therapies (including locally 
directed therapies) do not count as a line of therapy with regard to this exclusion criteria. 
  
19. Co-morbid systemic illnesses or other severe concurrent disease which, in the judgment 
of the investigator, would make the patient inappropriate for entry into this study or 
interfere significantly with the proper assessment of safety and toxicity of the prescribed 
regimens. 
 
20. Is currently participating and receiving study therapy or has participated in a study of an 
investigational agent and received study therapy or used an investigational device within 
4 weeks of the first dose of treatment. 
 
21. Has a diagnosis of immunodeficiency or is receiving systemic steroid therapy or any 
other form of immunosuppressive therapy within 7 days prior to the first dose of trial 
treatment. 
 
22. Has received prior therapy with an anti-PD-1, anti-PD-L1, or anti-PD-L2 agent. 
 
23. Has a known history of Human Immunodeficiency Virus (HIV) (HIV 1/2 antibodies). 
 
24. Has active autoimmune disease that has required systemic treatment in the past 2 years 
(i.e. with use of disease modifying agents, corticosteroids or immunosuppressive drugs). 
Replacement therapy (eg., thyroxine, insulin, or physiologic corticosteroid replacement 
therapy for adrenal or pituitary insufficiency, etc.) is not considered a form of systemic 
treatment.  NOTE:  Inhaled steroids or steroid injections for joint disease are allowed. 
 
25. Has known active Hepatitis B (e.g., HBsAg reactive) or Hepatitis C (e.g., HCV RNA 
[qualitative] is detected). 
 
26. Prior severe allergic reactions to a monoclonal antibody or hypersensitivity to 
pembrolizumab or any of its excipients. 
27. Has had a prior anti-cancer monoclonal antibody (mAb) within 4 weeks prior to study 
Day 1 or who has not recovered (i.e., ≤ Grade 1 or at baseline) from adverse events due 
to agents administered more than 4 weeks earlier. 


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28. Other active malignancy which requires current treatment and which in the opinion of the 
site investigator is likely to interfere with evaluation of disease assessment.   
NOTE:  Continuation of hormonal therapies is allowed.   
 
29. Has known active central nervous system (CNS) metastases and/or carcinomatous 
meningitis.  Subjects with previously treated brain metastases may participate provided 
they are stable (without evidence of progression by imaging for at least four weeks prior 
to the first dose of trial treatment and any neurologic symptoms have returned to 
baseline), have no evidence of new or enlarging brain metastases, and are not using 
steroids for at least 7 days prior to trial treatment.  This exception does not include 
carcinomatous meningitis which is excluded regardless of clinical stability.  
 
30. Major surgery ≤ 4 weeks from registration.  NOTE: Diagnostic laparoscopy (without 
other intervention) and/or biopsies (needle aspirate, core biopsy, open biopsy, etc…) are 
not considered major surgery.  If subject received surgery, they must have recovered 
adequately from the toxicity and/or complications from the intervention prior to starting 
therapy.   
 
31. Uncontrolled intercurrent illness which in the opinion of the investigator poses 
unacceptably high risk when combined with study treatment, including but not limited to 
the following: 
• Symptomatic congestive heart failure 
• Unstable angina pectoris 
• Severely impaired lung function 
• Known history of active TB (Bacillus Tuberculosis) 
• Uncontrolled diabetes as defined by fasting serum glucose >1.5 x ULN  
(NOTE: Optimal glycemic control should be achieved before starting trial 
therapy.) 
• Significant underlying liver disease such as cirrhosis or severe hepatic impairment 
• Has known psychiatric or substance abuse disorders that would interfere with 
cooperation with the requirements of the trial. 
 
32. Has an active infection requiring systemic therapy. 
33. Has a history or current evidence of any condition, therapy, or laboratory abnormality 
that might confound the results of the trial, interfere with the subject’s participation for 
the full duration of the trial, or is not in the best interest of the subject to participate, in 
the opinion of the treating investigator.  
 
34. Is pregnant or breastfeeding, or expecting to conceive or father children within the 
projected duration of the trial, starting with the pre-screening or screening visit through 
120 days after the last dose of trial treatment. 
 
 


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35. For patients in whom planned RT fields will include the heart, any of the following heart 
conditions, if in the opinion of the investigator they pose unacceptably high risk when 
combined with study treatment: 
• Prior symptomatic congestive heart failure 
• Documented myocardial infarction ≤6 months prior to registration (pretreatment 
ECG evidence of infarct only will not exclude patients) 
• Prior significant ventricular arrhythmia requiring medication 
• Prior 2nd or 3rd degree heart block or other types of clinically significant 
conduction delay ≤ 6 months prior to registration 
• Clinically significant pericardial disease (including pericardial effusion, 
pericarditis) or cardiac valvular disease ≤12 months prior to registration 
 
NOTE: As part of history and physical, all patients must be assessed for signs or 
symptoms of cardiac disease, or for prior history of cardiac disease. These conditions 
include but are not limited to diseases related to cardiac valves, pericardium, 
myocardium, atrioventricular delays or arrhythmias. It is strongly recommended that 
signs or symptoms of potentially clinically significant disease be evaluated with 
comprehensive cardiac echo. 
 
36. For patients in whom planned RT fields during the study will include the chest, any of the 
following, if in the opinion of the site investigator they pose unacceptably high risk when 
combined with study treatment:  
• Prior fistula within thorax, including bronchoalveolar or esophageal. 
• Respiratory condition that required oxygen supplementation ≤ 3 months prior to 
registration 
• Clinically significant pulmonary hypertension ≤ 12 months prior to registration 
• Pneumonia requiring treatment ≤ 1 month prior to registration 
• Pulmonary embolism requiring treatment ≤ 6 months prior to registration 
• Pleural effusion requiring drainage ≤ 12 months prior to registration 
 
37. Has known history of, or any evidence of active, non-infectious pneumonitis 
 
38. Currently uncontrolled hyper/hypothyroidism or hyper/hypocortism if in the opinion of 
the investigator they pose unacceptably high risk when combined with study treatment 
 
39. Received live vaccine ≤ 30 days prior to registration.  NOTE:  Seasonal influenza 
vaccines for injection are generally inactivated flu vaccines and are allowed; however 
intranasal influenza vaccines (e.g., Flu-Mist®) are live attenuated vaccines, and are not 
allowed. 
 
40. History of organ or stem cell transplantation 
 
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 system. Subjects must 
be registered prior to starting protocol therapy. 


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5. 
TREATMENT PLAN 
 
5.1 
Pre-Medications  
There are no pre-medication or hydration requirements for pembrolizumab administration.  For 
radiation therapy please use institutional standards for pre-medications. 
 
5.2 
Study Treatment Administration 
 
Treatment 
Dose 
Route 
Schedule 
Cycle 
Length 
Pembrolizumab 
200 mg  
Intravenously (IV) 
over 30 minutes 
Day -21 (induction) then  
Day 1 of each cycle.  
A maximum of 24 
continuous months of 
pembrolizumab (35 cycles) 
will be administered.    
21 days 
RT 
20-30 Gy 
over  
5 fractions 
Extracorporeal 
Cycle 1- Day 1-5; 
1st metastatic lesion  
Cycle 2- Day 1-5; 
2nd metastatic lesion 
 
5.2.1 Pembrolizumab 
Pembrolizumab 200 mg will be administered as a 30 minute IV infusion. Sites should make 
every effort to target infusion timing to be as close to 30 minutes as possible. However, given the 
variability of infusion pumps from site to site, a window of -5 minutes and +10 minutes is 
permitted (i.e., infusion time is 30 minutes: -5 min/+10 min).  Additional information can be 
found in the pharmacy manual. 
 
A window of ± 3 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).  After the 
completion of the period of concurrent RT, pembrolizumab monotherapy may be given up to ± 7 
days of the scheduled dose on Day 1 of each cycle.  All trial treatments are administered on an 
outpatient basis. 
 
NOTE: Dosing of pembrolizumab may be withheld in the case of medical/surgical events or 
logistical reasons (i.e. elective surgery, unrelated medical events, subject vacation, holidays) not 
related to study therapy. Patients should be placed back on study therapy within 3 weeks of the 
scheduled interruption to remain aligned with the Q3W dosing interval. The reason for 
withholding dosing of either pembrolizumab should be documented in the subject's study record.   
 


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5.2.2 Radiation Therapy 
Palliative external beam radiation therapy is administered to 2 separate non-overlapping sites of 
metastatic disease.  Radiation therapy is administered to the 1st site of metastatic disease on Days 
1-5 of Cycle 1 and to the 2nd site of metastatic disease on Days 1-5 of Cycle 2.  Here are 
guidelines for treatment: 
• The treating radiation oncologist will select sites to be treated with radiotherapy with 
preference given to locations that are currently symptomatic or may become symptomatic 
if there is local tumor progression.  Lesions that may provide palliative benefit may also 
be selected. 
• The entire lesion does not need to be completely encompassed within the radiation field.   
• If there are > 2 candidate metastatic sites, the treating radiation oncologist should 
consider treating sites with lower risk of treatment related adverse effects.  No more than 
2 metastatic sites may be treated. 
• A third measurable lesion located outside the planned RT fields must NOT be radiated. 
 
5.2.2.1 
Dose specifications and technical factors 
The trial administers a total dose of 20, 25, or 30 Gy over 5 fractions per metastatic lesion. The 
treating radiation oncologist may select the prescription dose to meet the tumor coverage 
requirements and the organ at risk constraints (OAR) as specified below.  It is recommended that 
the highest dose possible be administered, so long as all mandatory organs at risk (OAR) criteria 
are met.  General recommendations for dose based on lesion site are described below.  
 
Total dose (Gy) 
Fractions 
Dose per fraction 
Example sites 
20-30 
5 
4-6 
Liver, lung, bone (non-spine) 
20-25 
5 
4-5 
Abdomen/pelvis 
20 
5 
4 
Spine 
 
Dose should be prescribed such that ≥ 95% of the PTV receives the prescription dose. All 
mandatory OAR criteria must be met at a given radiotherapy dose level, or else the dose must be 
lowered to the next lowest dose level. 
 
The energy of the treatment beam will be at least 6 MV and no greater than 18 MV. The use of 
intensity-modulated radiation therapy (IMRT) is allowed, however, proton beam radiation 
therapy and intraoperative radiotherapy (IORT) are NOT allowed. Radiation dose prescriptions 
will be calculated using heterogeneity corrections to take into account tissue in homogeneities in 
treatment planning. 
 
5.2.2.2 
Immobilization and Simulation 
After study enrollment, patients will undergo CT simulation.  All patients will undergo high-
resolution CT-based (≤ 2 mm slices) 3D treatment planning in custom made immobilization 
devices.  IV contrast and other contrast agents can be used at physician’s discretion and as per 
standard of care. Respiratory motion can be assessed and managed when clinically indicated (i.e. 
lung, liver, and adrenal gland metastases) with a 4D CT scan. Respiratory gating techniques 
(including breath hold techniques) may be employed for sites with tumor motion greater than 1 
cm.  


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5.2.2.3 
Treatment Planning and Target Volumes  
When available, the RT planning process can incorporate diagnostic image fusion (MRI, 
PET/CT) with planning CT as needed to delineate the target volume and OAR. Gross tumor 
volume (GTV) will cover the entire metastasis to be treated.  For metastases prone to respiratory 
motion, an internal target volume (ITV) may be generated based on the 4D CT images. Clinical 
target volume (CTV) expansion will be 5 mm on the GTV or ITV.  For spinal lesions, the entire 
involved vertebra(e) should be included in the CTV.  The CTV may be modified to exclude 
overlap with uninvolved adjacent OAR. The PTV will be a 5 mm uniform expansion of the CTV. 
 
5.2.2.4 
Treatment Delivery 
Daily verification of the treatment setup will be performed with kilovoltage x-ray (2D) images or 
volumetric (3D) imaging (kilovoltage or megavoltage CT). CT volumetric imaging (3D to 3D 
match) is recommended for lung, liver, soft tissue and lymph nodes metastases.  For bone 
metastases, a pair of kV orthogonal (2D) imaging can be used. RT treatments will be delivered 
on consecutive weekdays. RT treatments can be delivered on weekends and holidays but is not 
required. 
 
5.2.2.5 
Radiation Compliance Criteria 
 
Target coverage 
PTV: D95 ≥ 95% and max dose (0.03 cc) ≤120% (while hotspots in GTV is encouraged, 
hotspots in PTV should be avoided where it overlaps with organs at risk.) 
 
Organs at risk 
All mandatory organs at risk (OAR) constraints must be met for treatment on this protocol. If 
these constraints cannot be met at a given total dose, then the dose must be lowered (Lowest dose 
is 20 Gy in 5 fractions).  For example, if the mandatory constraints cannot be met at a 
prescription dose of 30 Gy in 5 fractions, then the dose must be lowered to 25 Gy in 5 fractions. 
Questions regarding radiation planning can be directed to Dr. Sean Park (Park.Sean@mayo.edu) 
or Dr. Christopher Hallemeier (hallemeier.christopher@mayo.edu). 
 
Mandatory OAR  
Optic chiasm: D0.03cc[Gy] < 23 Gy  
Optic nerves: D0.03cc[Gy] < 23 Gy  
Cochlea: D0.03cc[Gy] < 23 Gy 
Brainstem: D0.03[Gy] < 23 Gy 
Spinal cord: D0.03[Gy] < 23 Gy 
Cauda equina: D0.03[Gy] < 28 Gy 
Brachial plexus: D0.03[Gy] < 28 Gy 
Sacral plexus: D0.03[Gy] < 28 Gy 
Esophagus: D0.03[Gy] < 28 Gy 
Trachea: D0.03[Gy] < 33 Gy 
Proximal bronchial tree: D0.03[Gy] < 33 Gy 
Heart: D0.03[Gy] < 33 Gy 
Stomach: D0.03[Gy] < 28 Gy 
Duodenum: D0.03[Gy] < 28 Gy 


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Small bowel: D0.03[Gy] < 28 Gy 
Large bowel: D0.03[Gy] < 28 Gy 
Rectum: D0.03[Gy] < 33 Gy 
Bladder: D0.03[Gy] < 33 Gy 
Skin: D0.03[Gy] < 33 Gy 
Lung: V20 < 10% 
Liver: CV21Gy[cc] > 700 cc  
Kidney total: V12.5Gy[%] < 67 %, Mean[Gy] < 12.5 Gy  
 
Recommended (but not required) OAR 
Esophagus: V19.5Gy[cc] < 5 cc  
Trachea: V16.5Gy[cc] < 4 cc 
Proximal bronchial tree: V16.5Gy[cc] < 4 cc  
Stomach: V18Gy[cc] < 10 cc  
Duodenum: V18Gy[cc] < 5 cc, V12.5Gy[cc] < 10 cc  
Small bowel: V19.5Gy[cc] < 5 cc  
Large bowel: V25Gy[cc] < 20 cc  
Rectum: V25Gy[cc] < 20 cc  
Bladder: V18.3Gy[cc] < 15 cc 
Femoral heads: V30Gy[cc] < 10 cc  
 
5.2.2.6 
Treatment monitoring requirements 
During radiation therapy, all patients will be seen by a healthcare provider for a radiation therapy 
management visit at least once weekly, at which time pertinent history and physical exam will be 
noted, in addition to the patient’s weight and performance status. 
 
5.2.2.7 
Treatment interruptions 
Treatment interruptions should be minimized through the use of supportive care measures. 
Interruptions for life-threatening adverse events (CTCAE Grade 4) are allowed. 
 
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 any medication or vaccination 
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 investigator 
and/or the patient’s primary physician. 
 
5.3.1 Acceptable 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 medication will be recorded in the case report forms (CRF) 
including all prescription, over-the-counter (OTC), herbal supplements, and IV medications and 
fluids. If changes occur during the trial period, documentation of drug dosage, frequency, route, 
and date may also be included on the eCRF. 
 


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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 30 days after the last dose of trial treatment should be recorded for SAEs and ECIs. 
 
5.3.2 Prohibited Concomitant Medications 
The following medications are not permitted during the screening and treatment phase (including 
retreatment for post-complete response relapse) of this trial: 
• Anti-neoplastic systemic chemotherapy or biological therapy.  Continuation of hormonal 
therapies is allowed. 
• Immunotherapy not specified in this protocol 
• Chemotherapy not specified in this protocol 
• Investigational agents other than pembrolizumab 
• 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, varicella/zoster, yellow fever, intranasal influenza, 
rabies, BCG, and typhoid vaccine. 
• Systemic glucocorticoids for any purpose other than to modulate symptoms from an 
event of clinical interest of suspected immunologic etiology. However, exceptions may 
be possible if approved after consultation with the sponsor-investigator. 
 
Patients 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. Patients 
may receive other medications that the investigator deems to be medically necessary. 
 
5.4 
Supportive Care 
Subjects should receive appropriate supportive care measures as deemed necessary by the 
treating investigator.  Suggested supportive care measures for the management of adverse events 
with potential immunologic etiology 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.  
 
5.4.1 Blood products and growth factors  
Blood products and growth factors should be utilized as clinically warranted and following 
institutional policies and recommendations. The use of growth factors should follow published 
guidelines of the Journal of Clinical Oncology, Vol 24, No 18 (June 20), 2006: pp. 2932-2947. 
 
5.4.1.1 
Neutropenia 
Prophylactic use of colony-stimulating factors including Granulocyte Colony-Stimulating Factor 
(G-CSF), pegylated G-CSF or Granulocyte Macrophage Colony-Stimulating Factor GM-CSF is 
not allowed in this study. Therapeutic use of G-CSF is allowed in patients with Grade 3-4 febrile 
neutropenia. 


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5.4.1.2 
Anemia 
Transfusions and/or erythropoietin may be utilized as clinically indicated for the treatment of 
anemia, but should be clearly noted as concurrent medications. 
 
5.4.1.3 
Thrombocytopenia 
Transfusion of platelets may be used if clinically indicated. ITP should be ruled out before 
initiation of platelet transfusion. 
 
5.4.2 Anti-Infectives 
Patients with a documented infectious complication should receive oral or IV antibiotics or other 
anti-infective agents as considered appropriate by the treating Investigator for a given infectious 
condition, according to standard institutional practice. 
 
5.4.3 Corticosteroids 
Patients requiring chronic steroid administration (excluding inhaled steroids) are excluded from 
the trial. Patients may continue on inhalation therapy. Corticosteroids are known 
immunosuppressive agents that can mitigate the effects of pembrolizumab. Steroids should be 
generally reserved to treat side effects of pembrolizumab.  
 
5.4.4 Antiemetics 
Antiemetics may be used at the discretion of the attending physician. Nausea and vomiting 
should be treated aggressively, and consideration should be given to the administration of 
prophylactic antiemetic therapy according to standard institutional practice. Patients should be 
strongly encouraged to maintain liberal oral fluid intake. Volume depletion should be corrected 
before initiation of study drug. 
 
5.4.5 Anti-Diarrheals 
Patients should be 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). In symptomatic patients, infectious etiologies should be ruled out, 
and if symptoms are persistent and/or severe, endoscopic evaluation should be considered.  
 
All patients who experience diarrhea 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. 
 
NOTE: Loperamide/diphenoxylate/atropines should NOT be used for diarrhea symptoms unless: 
(1) it is believed that pembrolizumab-related enterocolitis is unlikely to be present after detailed 
evaluation by gastroenterology, including endoscopy; PLUS (2) approval is documented by a 
gastroenterology specialist. 
 
 
 


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5.5 
Contraception 
Pembrolizumab may have adverse effects on a fetus in utero. Furthermore, it is not known if 
pembrolizumab has transient adverse effects on the composition of sperm. 
 
For this trial, male patients will be considered to be of non-reproductive potential if they have 
azoospermia (whether due to having had a vasectomy or due to an underlying medical 
condition). 
 
Female patients will be considered of non-reproductive potential if they are either:  
(1) postmenopausal (defined as at least 12 months with no menses without an alternative medical 
cause; in women < 45 years of age a high follicle stimulating hormone (FSH) level in the 
postmenopausal range may be used to confirm a post-menopausal state in women not using 
hormonal contraception or hormonal replacement therapy. In the absence of 12 months of 
amenorrhea, a single FSH measurement is insufficient.); OR  
 
(2) have had a hysterectomy and/or bilateral oophorectomy, bilateral salpingectomy or bilateral 
tubal ligation/occlusion, at least 6 weeks prior to screening; OR 
 
(3) has a congenital or acquired condition that prevents childbearing. 
 
Female and male patients of reproductive potential must agree to avoid becoming pregnant or 
impregnating a partner, respectively, while receiving study drug and for 120 days after the last 
dose of study drug by complying with one of the following: 
 
(1) practice abstinence from heterosexual activity; Abstinence (relative to heterosexual activity) 
can be used as the sole method of contraception if it is consistently employed as the subject’s 
preferred and usual lifestyle and if considered acceptable by local regulatory agencies and 
ERCs/IRBs.  Periodic abstinence (e.g., calendar, ovulation, sympto-thermal, OR  
post-ovulation methods, etc.) and withdrawal are not acceptable methods of contraception. 
 
(2) use (or have their partner use) acceptable contraception during heterosexual activity. 
 
5.5.1 Acceptable Methods of Contraception 
 
Single method (one of the following is acceptable): 
• 
intrauterine device (IUD) 
• 
vasectomy of a female subject’s male partner  
• 
contraceptive rod implanted into the skin 
 
Combination method (requires use of two of the following): 
• 
diaphragm with spermicide (cannot be used in conjunction with cervical cap/spermicide) 
• 
cervical cap with spermicide (nulliparous women only)   
• 
contraceptive sponge (nulliparous women only)  
• 
male condom or female condom (cannot be used together) 


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• 
hormonal contraceptive: oral contraceptive pill (estrogen/progestin pill or progestin-only 
pill), contraceptive skin patch, vaginal contraceptive ring, or subcutaneous contraceptive 
injection 
 
If a contraceptive method listed above is restricted by local regulations/guidelines, then it does 
not qualify as an acceptable method of contraception for patients participating at sites in this 
country/region. 
 
Patients 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. In order to participate in the 
study patients of childbearing potential must adhere to the contraception requirement (described 
above) from the day of study medication initiation (or 14 days prior to the initiation of study 
medication for oral contraception) throughout the study period up to 120 days after the last dose 
of trial therapy. If there is any question that a patient of childbearing potential will not reliably 
comply with the requirements for contraception, that patient should not be entered into the study. 
 
5.5.2 Use in Pregnancy 
If a patient inadvertently becomes pregnant while on treatment with pembrolizumab, the patient 
will immediately be removed from the study. The site will contact the patient at least monthly 
and document the patient’s status until the pregnancy has been completed or terminated.  Please 
see Section 11 for reporting guidelines.  
 
The study investigator will make every effort to obtain permission to follow the outcome of the 
pregnancy and report the condition of the fetus or newborn. If a male subject impregnates his 
female partner the study personnel at the site must be informed immediately and the pregnancy 
reported to HCRN and to Merck as described below. 
 
5.5.3 Use in Nursing Women 
Since many drugs are excreted in human milk, and because of the potential for serious adverse 
reactions in the nursing infant, patients 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 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.   
 
 
 


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6.1 
Dose Delays for Pembrolizumab 
There are no dose reductions for pembrolizumab; the 200 mg flat dose is the only dose that will 
be administered. 
 
Strictly follow the modifications in this table for the first two cycles, until individual treatment 
tolerance can be ascertained.  Thereafter, these modifications should be regarded as guidelines to 
produce mild-to-moderate, but not debilitating, side effects.  If multiple adverse events are seen, 
administer dose based on greatest reduction required for any single adverse event observed.  
Reductions or increases apply to treatment given in the preceding cycle and are based on adverse 
events observed since the prior dose. 
 
Adverse events (both non-serious and serious) associated with pembrolizumab exposure may 
represent an immunologic etiology. These AEs 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 the Table below.  
 
Permanently discontinue for any severe or Grade 3 drug-related AE that recurs or any life-
threatening event.  If pembrolizumab-related toxicity does not resolve to Grade 0-1 within 12 
weeks after last administration of study drug, study therapy discontinuation is recommended.  
 
With sponsor-investigator agreement, patients with a laboratory adverse event still at Grade 2 
may continue in the study only if asymptomatic and controlled.  In patients who continue on 
study therapy after experiencing an Adverse Event warranting potential dose delay, if considered 
drug-related by the site investigator. 
 
For patients who experience a recurrence of the same severe AEs listed above with rechallenge 
of pembrolizumab, a consultation with the sponsor-investigator will occur to determine whether 
the patient should continue in the study. A patient who experiences the same SAE of the same 
NCI CTCAE grade or higher with rechallenge of pembrolizumab must discontinue 
pembrolizumab immediately. 


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Dose Modification and Toxicity Management Guidelines for Immune-related AEs Associated with Pembro
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
corticosteroid has been tapered. Pembrolizumab should be permanently discontinued if AE does not resolve within 12 weeks o
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 immunosuppre
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 s
symptoms of pneumoniti
• 
Evaluate participants with
pneumonitis with radiogr
initiate corticosteroid trea
• 
Add prophylactic antibio
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 s
of enterocolitis (ie, diarrh
pain, blood or mucus in s
without fever) and of bow
(ie, peritoneal signs and i
• 
Participants with ≥ Grade
suspecting colitis should 
consultation and perform
rule out colitis.  
• 
Participants with diarrhea
advised to drink liberal qu
fluids.  If sufficient oral f
feasible, fluid and electro
substituted via IV infusio
Grade 4 
Permanently 
discontinue 
 
 


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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  
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 functio
weekly or more frequentl
enzyme value returned to
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 h
other signs and symptom
Hypophysitis 
Grade 2 
Withhold 
• Administer corticosteroids and 
initiate hormonal replacements as 
clinically indicated.   
 
• 
Monitor for signs and sym
hypophysitis (including h
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 sym
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 sym
disorders.  
 
Nephritis and 
Renal 
dysfunction 
Grade 2 
Withhold  
• Administer corticosteroids 
(prednisone 1-2 mg/kg or 
equivalent) followed by taper. 
• 
Monitor changes of renal
 
Grade 3 or 4  
Permanently 
discontinue 
 
 


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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  
Myocarditis 
Grade 1 or 2 
Withhold 
• Based on severity of AE 
administer corticosteroids 
• 
Ensure adequate evaluati
etiology and/or exclude o
 
Grade 3 or 4 
Permanently 
discontinue 
All other 
immune-related 
AEs 
Intolerable/ 
persistent Grade 2 
Withhold 
• Based on type and severity of AE 
administer corticosteroids  
• 
Ensure adequate evaluati
etiology and/or exclude o
 
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 
when AE resolves to ≤ Grade 2 and is controlled with hormonal replacement therapy or achieved metabolic control (in case of T1D
 
 
 
 
 
 


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6.2 
Immunotherapy-related toxicities 
Suggested supportive care measures for the management of adverse events with potential 
immunologic etiology 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 site investigator determines the events to be 
related to pembrolizumab.  If after the evaluation, the event is determined not to be related, the 
investigator does not need to follow the treatment guidance (as outlined below).  
 
It may be necessary to perform conditional procedures such as bronchoscopy, endoscopy, or skin 
photography as part of evaluation of the event. 
 
Patients should be monitored for signs and symptoms of immunotherapy-related toxicities, which 
include but are not limited to the following: 
 
Pneumonitis 
o 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. 
o For Grade 3-4 events, immediately treat with intravenous steroids. Administer additional 
anti-inflammatory measures, as needed. 
o Add prophylactic antibiotics for opportunistic infections in the case of prolonged steroid 
administration. 
 
Diarrhea/colitis 
Patients 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). 
o All patients 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. 
o For Grade 2 diarrhea/colitis, administer oral corticosteroids. 
o For Grade 3 or 4 diarrhea/colitis, treat with intravenous steroids followed by high dose 
oral steroids. Recommended dose of 1 to 2 mg/kg/day of prednisone or equivalent. 
o When symptoms improve to Grade 1 or less, steroid taper should be started and 
continued over no less than 4 weeks. 
 
Type 1 Diabetes Mellitus (if new onset, including diabetic ketoacidosis [DKA]) or ≥Grade 3 
Hyperglycemia, if associated with ketosis (ketonuria) or metabolic acidosis (DKA) 
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.  


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o Evaluate patients with serum glucose and a metabolic panel, urine ketones, glycosylated 
hemoglobin, and C-peptide. 
 
Hypophysitis 
o 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. 
o 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. 
 
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. 
o Grade 2 hyperthyroidism events (and Grade 2-4 hypothyroidism): 
▪ In hyperthyroidism, non-selective beta-blockers (e.g. propranolol) are suggested 
as initial therapy. 
▪ In hypothyroidism, thyroid hormone replacement therapy, with levothyroxine or 
liothyroinine, is indicated per standard of care. 
o Grade 3-4 hyperthyroidism  
▪ 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. 
Hepatic 
o For Grade 2 events, monitor liver function tests more frequently until returned to 
baseline values (consider weekly). 
▪ Treat with IV or oral corticosteroids 
o 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. 
 
Renal failure or nephritis 
o For Grade 2 events, treat with corticosteroids. 
o 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.  
 
6.3 
Infusion Reaction 
Appropriate resuscitation equipment should be available in the room and a physician readily 
available during the period of study treatment administration.  Below are guidelines for 
management of Infusion Reactions 
 


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NCI CTCAE Grade 
Treatment 
Premedication at subsequent 
dosing 
Grade 1 
Mild reaction; infusion 
interruption not indicated; 
intervention not indicated 
Increase monitoring of vital signs as 
medically indicated until the subject is 
deemed medically stable in the opinion of 
the investigator. 
None 
Grade 2 
Requires infusion interruption 
but responds promptly to 
symptomatic treatment (e.g., 
antihistamines, NSAIDS, 
narcotics, IV fluids); 
prophylactic medications 
indicated for ≤24 hrs 
Stop Infusion and monitor symptoms. 
Additional appropriate medical therapy may 
include but is not limited to: 
IV fluids 
Antihistamines 
NSAIDS 
Acetaminophen 
Narcotics 
Increase monitoring of vital signs as 
medically indicated until the subject is 
deemed medically stable in the opinion of 
the investigator. 
If symptoms resolve within one hour of 
stopping treatment infusion, the infusion 
may be restarted at 50% of the original 
infusion rate (e.g., from 100 mL/hr to 50 
mL/hr). Otherwise dosing will be held until 
symptoms resolve and the subject should be 
premedicated for the next scheduled dose. 
Patients who develop Grade 2 toxicity 
despite adequate premedication should be 
permanently discontinued from further 
trial treatment administration. 
Subject may be premedicated 
1.5h (± 30 minutes) prior to 
infusion of pembrolizumab 
with: 
Diphenhydramine 50 mg po 
(or equivalent dose of 
antihistamine). 
Acetaminophen 500-1000 mg 
po (or equivalent dose of 
antipyretic). 
Grades 3 or 4 
Grade 3: 
Prolonged (i.e., not rapidly 
responsive to symptomatic 
medication and/or brief 
interruption of infusion); 
recurrence of symptoms 
following initial improvement; 
hospitalization indicated for 
other clinical sequelae (e.g., 
renal impairment, pulmonary 
infiltrates) 
Grade 4: 
Life-threatening; pressor or 
ventilatory support indicated 
Stop Infusion 
Additional appropriate medical therapy may 
include but is not limited to: 
IV fluids 
Antihistamines 
NSAIDS 
Acetaminophen 
Narcotics 
Oxygen 
Pressors 
Corticosteroids 
Epinephrine 
 
Increase monitoring of vital signs as 
medically indicated until the subject is 
deemed medically stable in the opinion of 
the investigator. 
Hospitalization may be indicated. 
Subject is permanently discontinued from 
further trial treatment administration. 
No subsequent dosing 
 


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6.4 
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 in the EDC system. 
• 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  
• Protocol therapy is interrupted for ≥ 12 weeks.  
• A maximum of 24 continuous months of pembrolizumab (35 cycles) was administered.    
 
6.5 
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 
Cycle = 21 days 
< 28 days 
prior to 
Registration 
Treatment 
End of 
treatment13 
~ 30 days 
after last 
dose of 
pembro 
Pre-RT 
(induction) 
During RT 
Cycle 3 and 
subsequent 
Day 1 
Cycle 1 
Cycle 2 
D -21 
D1 
D1 
REQUIRED ASSESSMENTS 
 
 
 
 
 
 
Informed Consent 
X 
 
 
 
 
 
Medical History1 
X 
 
 
 
 
 
Physical Exam 
X 
 
X 
X 
X 
X 
Vital signs and ECOG PS 2 
X 
 
X 
X 
X 
X 
ECG  
X 
 
 
 
 
 
AE and concomitant medications review 
X 
X 
X 
X 
X 
X 
LABORATORY ASSESSMENTS 
 
 
 
 
 
 
CBC with differential and platelet count 
X 
X 
X 
X 
X 
X 
Chemistry panel3 
X 
X 
X 
X 
X 
X 
T3, Free T4, TSH4 
X 
 
 
X 
X4 
X 
Pregnancy test5  
X 
 
 
 
 
 
Urinalysis for protein, glucose, blood 
X 
 
 
 
 
 
DISEASE ASSESSMENT  
 
 
 
 
 
 
CT (chest, abd/pelvis)6 
X6 
 
 
 
X6 
X6 
Mammogram (women only)7  
X7 
 
 
 
 
 
Colonoscopy (if liver metastasis or 
suspicion of colon primary)8  
X8 
 
 
 
 
 
SPECIMEN COLLECTION 
 
 
 
 
 
 
MANDATORY Blood for correlative 
research9 
 
X 
X 
X 
X9 
X 
MANDATORY Blood for somatic 
baseline10 
 
X 
 
 
 
 
MANDATORY: Archival Tissue, 
Research Biopsy or New Standard of 
Care Biopsy11 
 
X 
 
 
 
 
MANDATORY: Tumor Tissue from 
Standard of Care Biopsy12 
 
 
 
 
 
X12 
FOLLOW UP 
 
 
 
 
 
 
Survival, Subsequent Therapy14  
 
 
 
 
 
 
 
 


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Key Footnotes: 
1: Medical History to include a smoking history, how the subject learned about the study and prior anti-cancer treatment.  If available, s
sequencing or expression profiling results is mandatory. If and when available, submission of genomic sequencing and expression prof
Results of testing are not required prior to treatment. 
2: Vital signs to include blood pressure, weight, and height (screening only) and ECOG performance status.   
3: AST, ALT, alkaline phosphatase, total bilirubin (direct bilirubin if total is elevated), BUN, Cr, creatinine clearance, calcium, phosph
bicarbonate, uric acid, chloride. If there is a clinical need for more frequent testing, it may be performed as needed. 
4: Thyroid tests are collected every other treatment cycle, starting with the second cycle after the completion of RT 
5: For women of childbearing potential only. Must be done within 72 hours prior to registration 
6: Tumor measurements will be performed at the following time points: (1) ≤ 28 days prior to registration, (2) after the completion of R
initiation of post-RT therapy), and (3) every 9 weeks (± 7 days) afterward until the subject comes off study treatment.  Tumor measurem
once every 12 weeks (± 2 weeks) after 6 months of study treatment and once every 4 months (± 3 weeks) after 12 months of study treat
increased time intervals may be possible after discussion with and agreement by the sponsor-investigator. The type of tumor measurem
discretion of the site investigator although preferred is CT scan with contrast; if this cannot be done then a PET/CT; if this cannot be do
performed. The type of modality used should be consistent throughout the study treatment. All radiology imaging tests will be submitt
review once funding is available. Please see SPM regarding shipping of radiology imaging.  
7: Performed ≤ 60 days prior to original diagnosis or at any time thereafter. A mammography should be performed if initial histology o
possible diagnosis of breast cancer. Also, mammography should be performed if the clinical presentation suggests a possible diagnosis 
particularly important if there is no readily accessible site for biopsy. The cost of the mammogram should be covered if the treating phy
“diagnostic” as opposed to “screening.” A mammogram is not necessary for squamous carcinoma or neuroendocrine tumors where a br
clinically being considered.  
8 Performed within one year prior to original diagnosis or at any time thereafter. 
9: Whole blood samples for correlative studies will be sent to the Haidong Dong Lab at Mayo Clinic after collection at the following 5 
treatment Day -21 (within 21 days prior to initiating treatment), (2) prior to treatment Cycle 1 Day 1 (- 5 days), (3) prior to treatment C
(4) prior to treatment Cycle 3 Day 1 (first pembrolizumab after completion of RT (- 5 days) and (5) at the time of treatment discontinua
can be found in the Correlative Laboratory Manual (CLM) 
10: Whole blood samples for somatic baseline will be collected at any time prior to treatment Day -21 and after registration.   See CLM
11: Archival tissue must be identified during screening and shipped prior to Day -21. If archival tissue or standard of care biopsy is not
biopsy will be performed. Please see SPM for additional details.  
12: If a subject has a biopsy at progression as per standard of care, some of this tissue will be stored for research particularly directed at
resistance. 
13: The end of treatment visit should only occur when subjects permanently stop study treatment for whatever reasons (toxicity, progre
site investigator) and should be performed 30 days (±7 days) after the last dose of treatment. Subjects who have an ongoing Grade ≥ 2 o
this visit will continue to be followed until the AE resolves to ≤ Grade 1 or baseline, deemed clinically insignificant, and/or until a new
starts, whichever is earlier. 
14: Follow up will occur in all subjects until documented disease progression. Subjects will be followed for survival and initiation of an
3 months for 2 years from treatment discontinuation.  Follow up may be accomplished via clinic visit, phone call, or other avenues as a
 
 


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8. 
BIOSPECIMEN STUDIES AND PROCEDURES 
 
Summary Tables of Blood specimens 
*CLM = Correlative Laboratory Manual 
 
Summary Table of Tissue Specimens 
Type of tissue biospecimen to submit 
Mandatory  
When to submit 
Reason for submission 
(background/methodology 
section) 
Where to find specifi
details for biospecim
submission 
Formalin-fixed paraffin-embedded 
(FFPE) tissue blocks with corresponding 
H&E (OR unstained slides with 
corresponding H&E); archival or fresh 
biopsy 
Mandatory 
Prior to Day -21 
CLM 
CLM 
Formalin-fixed paraffin-embedded 
(FFPE) tissue block with corresponding 
H&E (OR unstained slides with 
corresponding H&E) 
Mandatory 
At time of tumor 
recurrence during 
standard of care biopsy. 
CLM 
CLM 
 
Type of 
biospecimen to 
submit 
Mandatory 
or optional 
When to 
submit 
Collection tube 
description 
and/or additive 
(color of tube 
top) 
Volume to 
collect per 
tube (number 
of tubes to be 
collected) 
Blood product 
being processed 
and submitted by 
participating site 
Additional 
processing 
required at 
site after 
blood draw? 
Storage/ 
shipping 
conditions 
Reason for 
submission 
(background/m
hodology secti
Blood for 
correlative 
research 
(Haidong Dong 
Lab)a 
Mandatory 
Pre-treatment 
Day -21, 
C1D1, C2D1, 
C3D1, EOT; 
See calendar 
above for 
additional 
details 
EDTA (purple 
top) 
10 ml (2) 
Whole blood 
No 
Ambient 
temperature 
(DO NOT 
FREEZE) 
Defined 
translational 
studies  
(Section 1.5) 
Blood for 
somatic baseline Mandatory 
Pre-treatment 
Day -21  
EDTA (purple 
top) 
10 ml (1) 
Whole blood 
No 
Frozen to 
HCRN 
Stored for futur
somatic baselin
testing 


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8.1 
Whole Blood Collection 
Please see Tables above and CLM for additional information regarding blood collection.  
Collection of these samples should occur Monday- Thursday. 
 
8.1.1 Return of Genetic Testing Research Results 
Since the results generated by the somatic baseline genetic testing included in this section are not 
currently anticipated to have clinical relevance to the patient or their family members, the genetic 
results will not be disclosed to the patients or their physicians. Sharing of research data with 
individual patients should only occur when data have been validated by multiple studies and 
testing has been done in CLIA-approved laboratories. 
 
8.2 
Tissue Collection Information  
 
8.2.1 Pretreatment Tissue 
Pretreatment tumor tissue must be submitted for this trial.  Archived tumor tissue is sufficient 
and tissue collected as close as possible to study treatment is strongly encouraged.  A 
documented attempt to locate and retrieve archival tissue should be made during screening. 
Please see the CLM for documentation form. If archival tissue is identified, it should be shipped 
prior to Day -21. If archival tissue is not available and a standard of care biopsy is not being 
performed, a research biopsy is required for study eligibility.   
 
Submitted tissue samples will be analyzed for the following biomarkers: 
• TOO assay 
• Immune status using such assays as mismatch repair, microsatellite instability, mutational 
load, genomic profile, PD-L1, BIM-PD-1 expression, tumor infiltrating lymphocytes, and 
others 
• Molecular pathways that are deemed to be relevant for prognosis or ability to predict 
outcomes from immune and/or radiotherapy 
 
The rationale for these tissue studies is explained below. 
 
8.2.2 Tissue from Standard of Care Biopsy at Progression 
If a subject has a biopsy at progression as per standard of care, some of this tissue will be stored 
for research particularly directed at mechanisms of resistance such as but not limited to BIM/PD-
1 expression, and CD3, CD8, granzyme B by IHC and other molecular markers in the PD-1 
pathway.  
 
8.3 
Sample Analyses 
 
8.3.1 MSI and Mutation Load 
Recent data implicate the potential value of MSI (mismatch repair) status (Le et al., 2015) and 
mutation load (Le et al., 2015; Rizvi et al., 2015) in predicting anti-PD-1/PD-L1 blockade.  As 
described above, CUPs appear to have a relatively high proportion of tumors with high mutation 
load.  MSI data are recently available on certain genomic profiles, which also provides 
information on alterations in hundreds of cancer-related genes and mutational load. Genomic 
testing is commercially available and has been obtained as part of clinical care in Mayo Clinic 


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patients to facilitate treatment options.   
Given that genomic testing is often routinely ordered in clinical settings, some subjects may 
already have had the necessary analyses performed.  Subjects who have had previous genetic 
results including MSI and mutation load must consent to permit use of those previous findings 
by the investigators and therefore not need additional genomic testing.  If previous results are not 
available, subjects must consent to providing tissue for this analysis. 
 
8.3.2 Tissue Expression of PD-L1 and Immune Markers 
PDL1 expression is an important marker that has potential predictive value for pembrolizumab 
therapy.  The density of markers of adaptive immunity such as CD3, CD8, FoxP3, and CD45RO 
infiltrating intratumoral/stromal tissue compartments have been associated with survival in 
patients with colorectal cancer, as we (Yoon et al., 2012) and others (Galon et al., 2006; Pages et 
al., 2005) have shown.  It is possible that adaptive immune markers in TILs, apart from PD-L1 
expression, can have predictive value for immunomodulating agents (Kluger et al., 2015). 
 
Pre-treatment tissue and tissue from standard of care biopsy at progression will be analyzed for 
multiple markers including PD-L1, CD3, CD8, FoxP3, CD45RO via IHC.   
 
8.3.3 Soluble PD-L1 
Release of inhibitory coregulatory proteins into the circulation may represent one mechanism by 
which tumors thwart immune responses. A collaborator in the current trial (Haidong Dong 
Laboratory) identified a soluble form of PD-L1 that retains immunosuppressive activity and is 
associated with aggressive renal cell carcinoma (Frigola et al., 2011).  This lab developed an 
ELISA for quantification of PD-L1 in biological fluids. Soluble PD-L1 was detected in the cell 
supernatants of some PD-L1-positive tumor cell lines. Protein sequencing established that the 
measured soluble PD-L1 retained its receptor-binding domain and could deliver proapoptotic 
signals to T cells. Higher preoperative soluble PD-L1 levels were associated with larger tumors 
(P < .001), tumors of advanced stage (P =.017) and grade (P =.044), and tumors with necrosis (P 
=.003). A doubling of soluble PD-L1 levels was associated with a 41% increased risk of death (P 
=.010). These observations suggest that soluble PD-L1 may be detected in the sera of cancer 
patients and that it may systemically impair host immunity, thereby fostering cancer progression 
and subsequent poor clinical outcome.  
 
Peripheral blood is collected for soluble PD-L1 analyses at the following 5 time points: (1) prior 
to treatment Day -21 (2) prior to treatment Cycle 1 Day 1 (- 5 days), (3) prior to treatment Cycle 
2 Day 1 (- 5 days), (4) prior to treatment Cycle 3 Day 1 (first pembrolizumab after completion of 
RT’ - 5 days) and (5) at the time of treatment discontinuation.   
 
8.3.4 Tumor-Reactive CD11ahighPD-1high CD8+ T Effector Population Following RT 
T cell receptor (TCR) tetramers can detect tumor-specific immunity but have limitations. Both 
the down regulation of the TCR induced by persistent tumor antigen encounter and the weak 
affinity of CD8+ T cells for tumor antigens contribute to the underestimation of anti-tumor 
responses measured by tetramers. In addition, TCR tetramers are not capable of detecting CD8 + 
T cell responses to subdominant epitopes. Therefore, a novel method of detecting tumor-related 
T cell response is necessary in situations where the identity of tumor antigens are unknown or 
suboptimal to be detected by tetramers. CD11a expression is required in the rejection of tumors 


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(Schmits et al., 1996). A collaborator laboratory recently established that the CD11ahigh PD-1+ 
CD8+ T cell population represent tumor-reactive CD8+ T cells(Liu et al., 2013). In addition, work 
from this lab determined that CD11ahighCD8+ T cells inside tumors are tumor-specific functional 
effector cells and that their function is enhanced by stereotactic ablative radiotherapy in PD-1 
knockout mice(Park et al., 2015). In the current study, we propose to investigate whether the 
CD11ahigh PD-1+ CD8+ T cell population in GEJ cancer patients following the initiation of 
chemoRT plus pembrolizumab can identify tumor-related T cell response.  
 
PBMCs and plasma will be collected at the following 5 time points: (1) prior to treatment Day -
21 (2) prior to treatment Cycle 1 Day 1 (- 5 days), (3) prior to treatment Cycle 2 Day 1 (- 5 days), 
(4) prior to treatment Cycle 3 Day 1 (first pembrolizumab after completion of RT’ - 5 days) and 
(5) at the time of treatment discontinuation.   
• T cell activation marker assay PD-1+ CD11ahigh CD8+ T cells will be stained with 
antibodies for CD69, CD25, CD45RO and CD62L which are upregulated upon T cell 
activation.  
• T cell proliferation assay The proliferation of PD-1+ CD11ahigh CD8+ T cells will be 
measured by their larger forward light scatter (FSC) and intranuclear expression of Ki67.  
• T cell differentiation assay The expression of transcription factors that controls T cell 
differentiation into effector or memory cells will be analyzed. We will measure the levels 
of T-bet (for effector T cells) and Eomes (for memory T cells) in PD-1 + CD11ahigh CD8+ 
T cells.  
• T cell function assay. Cytotoxic (CTL) T cell effector functions of CD11ahigh PD-1+ CD8+ 
T cells will be assessed by measuring degranulation (CD107a expression) and 
intracellular production of IFN-gamma following a 4-hour ex vivo stimulation with 
PMA/ionomycin or with any known prostate antigen peptides. The expression of effector 
molecule of CTL (granzyme B) and its inducer transcription factors (Bcl6 and Runx3), 
will be examined in PD-1+ CD11ahigh CD8+ T cells. 
 
8.3.5 Balance of Circulating CD8+ Effector Cell Subpopulation 
Given that the antigen spectrum for a given tumor is usually incomplete, it is possible that neo 
antigens could be generated and released from tumor cells following RT, which would not be 
identified by commercially available tetramers. Since the establishment of antigen specificity for 
the entire T-cell population is not possible, this study will ascertain whether the antigen burst 
and/or inflammatory factors released following RT would selectively expand or reduce T cell 
subpopulation. While RT-induced changes are initiated locally at tumor sites, RT can impact the 
systemic immune system as observed in the abscopal effect. We hypothesize that tumor antigens 
and/or inflammatory factors released from RT-induced tumor cell necrosis will trigger an efflux 
of effector or memory T cells in the peripheral blood since these subpopulation of T cells are 
antigen-primed and may have a rapid response to altered antigen stimulation in context with 
inflammatory cytokines. It has been reported that RT increases the serum levels of IL-1beta and 
IL-6 and reduces the serum levels of TGF-beta in prostate cancer patients after 1-2 weeks of RT. 
Among the cytokines, IL-6 is responsible for expansion of effector memory CD8 + T cells and IL-
1beta and TGF-beta regulate the transmigration or tissue retention of effector and memory CD8 + 
T cells. In this portion of the project, fluctuations in frequencies among naïve 
(CCR7+CD45RA+), central memory (CCR7+CD45RA-), effector/effector memory (CCR7-
CD45RA-), and terminally differentiated (CCR7-CD45RA+) CD8+ T cells will be determined and 


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evaluated together with the development of anti-gastric cancer immunity in relation to the timing 
of RT. PBMCs and plasma will be isolated at (baseline), after RT (day 8, 15, pre-surgery), after 
adjuvant therapy, and at tumor recurrence. Cells will be co-stained with antibodies for CD3, 
CD8, CD45RA, CCR7 to identify CD8+ T cells and their phenotype. The percentage of each 
subset will be determined based on the total CD8+ T cells when analyzed by flow cytometry. We 
will compare the frequency of each subset of CD8+T cells and their PD-1 expression in relation 
to the different time points following RT. If we identify any significant changes of T cell subset, 
we will measure the cytokine profiles of these patients at that time point using multiplex 
methods.  
 
8.3.6 BIM Expression in Circulating T Cells and Tumor-Infiltrating Lymphocytes 
The Haidong Dong lab recently reported that Bim is a downstream signaling molecule of the PD-
1 pathway, and its detection in T cells is significantly associated with expression of PD-1 and 
effector T cell markers (Dronca et al., 2016). High levels of Bim in circulating tumor-reactive 
(PD-1+CD11ahiCD8+) T cells were prognostic of poor survival in patients with metastatic 
melanoma who did not receive anti–PD-1 therapy and were also predictive of clinical benefit in 
patients with metastatic melanoma who were treated with anti–PD-1 therapy. Moreover, this 
circulating tumor-reactive T cell population significantly decreased after successful anti–PD-1 
therapy. This study supports a crucial role of Bim in both T cell activation and apoptosis as 
regulated by PD-1 and PD-L1 interactions in effector CD8+ T cells. Measurement of Bim levels 
in circulating T cells of patients with cancer may provide a less invasive strategy to predict and 
monitor responses to anti–PD-1 therapy.  This lab also has experience evaluating BIM/PD-1 
expression in tumor infiltrating T cells using double-stained IHC. 
 
8.3.7 Expression Profiling to Identify Tissue of Origin in CUP 
Accurate identiﬁcation of the tissue of origin (TOO) via molecular profiling and subsequent site-
speciﬁc therapy has been theorized to improve outcomes.  Gene expression profiling has 
accurately identified the tissue of origin in ∼85% of patients in patients with known primary, as 
a collaborator in the current study (Kerr et al., 2012) and others (Erlander et al., 2011; Monzon et 
al., 2009; Pillai et al., 2011) have shown.  In retrospective studies, molecular profiling rendered a 
prediction in the majority of CUP patients that was consistent with clinical and pathologic 
features (Horlings et al., 2008; Varadhachary et al., 2008). In CUP patients whose primary was 
detected later, molecular testing predicted the primary in 75% of cases (Greco et al., 2013).  
 
Our group recently found that a commercially available tumor gene expression profiling assay 
that predicts tissue of origin identified CUP patients who responded to carboplatin/paclitaxel-
based chemotherapy (Yoon et al., 2016).  In this multicenter NCI-sponsored cooperative group 
trial (N0871), patients with a tissue of origin where platinum/taxane is a standard had a higher 
response rate (53% vs 26%) and significantly longer progression free survival (6.4 vs 3.5 
months; HR 0.47; 95% CI 0.24–0.93; P =.026) and overall survival (17.8 vs 8.3 months, HR 
0.37; 95% CI 0.18–0.76; P =.005), compared with patients with a tissue of origin where 
platinum/taxane is not standard.  These data suggest the possible prognostic and predictive 
importance of evaluating tissue of origin in CUP patients.  Multiple commercially available TOO 
assays (CancerType ID, BioTheranostics; Cancer Genetics) can be ordered for clinical purposes 
that can aid in diagnosis and treatment decisions. Pre-treatment tissue (archival or biopsy) will be 
submitted for TOO assay analyses.  


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8.3 
Storage of Biospecimens 
At the completion of the study, any unused/remaining material will be stored for future research 
in the area of cancer.  Patients will be asked permission to store leftover samples of tissue or 
blood.   
 
8.4 
Confidentiality of Biospecimens 
Samples that are collected will be identified by a subject’s sequence ID 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. 
 
9. 
CRITERIA FOR DISEASE EVALUATION 
RECIST 1.1 will be adapted to account for the unique tumor response characteristics seen with 
treatment of pembrolizumab. Immunotherapeutic agents such as pembrolizumab may produce 
antitumor effects by potentiating endogenous cancer-specific immune responses. The response 
patterns seen with such an approach may extend beyond the typical time course of responses 
seen with cytotoxic agents, and can induce a clinical response after an apparent initial increase in 
tumor volume or even the appearance of new lesions. Standard RECIST may not provide an 
accurate response assessment of immunotherapeutic agents such as pembrolizumab. Immune-
related RECIST (irRECIST) is RECIST 1.1 adapted as described below to account for the unique 
tumor response seen with immuno-therapeutics.  In addition, an improved long-term survival has 
been observed when using the best abscopal response as the predictor variable. 
 
The version of RECIST used in this study will be the new international criteria proposed by the 
revised Response Evaluation Criteria in Solid Tumors (RECIST) guidelines (version 1.1) by 
Eisenhauer et al.  Changes in the largest diameter (unidimensional measurement) of the tumor 
lesions and the short axis measurements in the case of lymph nodes are used in the RECIST 
guideline. 
 
9.1 
Tumor Imaging and Assessment of Disease 
Each participating site will determine subject eligibility and response. All CT, PET, and MRI 
scans, scheduled and unscheduled, should be submitted to central imaging as soon as possible.  
Treatment decisions will be based on the site investigator’s radiographic assessment of disease, 
and not based on Research Base’s assessment.  There will be no real-time confirmation from 
central review.  Retrospectively, the Research Base will review radiologic images for a 
retrospective analysis of subject eligibility and response. 
 
Tumor imaging may be performed by computed tomography (CT) (preferred) or magnetic 
resonance imaging (MRI), but the same imaging technique, acquisition, and processing 
parameters should be used in a subject throughout the trial.  Scans performed as part of routine 
clinical management are acceptable for use as the baseline scan if they are of diagnostic quality.  
Scans performed as part of routine clinical management are acceptable for use as the baseline 
scan if they are of diagnostic quality.   
 
Imaging should follow calendar days and not be delayed for any dose interruptions that may 
occur. 
 


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Imaging should continue to be performed until whichever one of the following occurs first: 
• 
disease progression  
• 
start of new systemic anti-cancer treatment 
• 
withdrawal of consent 
• 
death 
• 
end of study 
 
9.1.1 Timing of Repeat Imaging to Confirm Response 
Best abscopal response should be confirmed by a repeat radiographic assessment not less than 4 
weeks from the date the response was first documented. 
 
9.2 
Assessment of Disease  
 
9.2.1 Determination of Response (tumor shrinkage) 
Best abscopal response will be applied as the primary measure for assessment of tumor 
shrinkage, following the NYU study by Golden et al (see Background 1.2.2).   
Best abscopal response is defined as either of the following: 
• Non-nodal lesions:  Decrease in the longest diameter by at least 30% in any measurable 
(≥1 cm) non-irradiated lesion from baseline 
• Nodal lesions:  Decrease in the short-axis diameter of any measurable (≥1.5 cm) non-
irradiated nodal lesion from baseline 
 
RECIST 1.1 will be applied as a secondary measure for assessment of tumor response. 
 
9.2.2 Determination of Progressive Disease (irRECIST) 
irRECIST is applied only in situations when first radiographic progression per RECIST 1.1 has 
occurred.  After initial radiographic progression per RECIST 1.1, irRECIST will be used by site 
investigators to assess tumor progression and to make treatment decisions.  
 
If radiologic imaging verifies initial disease progression per RECIST 1.1, treatment may 
continue at the discretion of the site investigator until repeat imaging ≥ 4 weeks later. The site 
investigator’s decision to continue treatment while awaiting repeat imaging should be based on 
the subject’s overall clinical condition guided by the following criteria: 
• Absence of signs and symptoms indicating disease progression 
• No decline in ECOG performance status 
• Absence of rapid progression of disease 
• Absence of progressive tumor at critical anatomical sites (eg, cord compression) 
requiring urgent alternative medical intervention 
 
Assessment of repeat imaging after initial RECIST 1.1-defined disease progression:  
Subject will be considered as not having disease progression as defined by irRECIST if repeat 
imaging shows: 
• < 20% increase in tumor burden compared to nadir, and  
• Stable or improved previous new lesion (if identified as cause for initial disease 
progression), and 
• Stable/improved non-target disease (if identified as cause for initial disease progression). 


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• Treatment plan:  Treatment may be continued/resumed. 
 
Subject will be considered as having disease progression as defined by irRECIST if repeat 
imaging confirms disease progression due to any of the scenarios listed below: 
• Tumor burden remains ≥20% and at least 5 mm absolute increase compared to nadir, or 
• Non-target disease resulting in initial disease progression is worse (qualitative), or 
• New lesion resulting in initial disease progression is worse (qualitative), or 
• Additional new lesion(s) since last evaluation 
• Treatment plan: Subjects will be discontinued from study therapy 
 
In determining whether or not the tumor burden has increased or decreased, site investigators 
should consider all target lesions as well as non-target lesions. 
 
In cases where irRECIST-defined progression is occurring at the same time as a best 
abscopal response, study treatment should generally be discontinued.  However, if the 
subject is achieving a clinically meaningful benefit, an exception to continue treatment may be 
considered. 
 
The guidance described above regarding treatment continuation/discontinuation and repeat 
imaging per irRECIST is described in the table below. 
 
When feasible, study treatment should continue until progression is confirmed. Subjects that are 
deemed clinically unstable are not required to have repeat imaging for confirmation of 
progressive disease.  Subsequent tumor imaging follows protocol schedule. 
 
Plan for Imaging and Treatment after 1st Radiologic Evidence of Progressive  
Plan for Imaging and Treatment after 1st Radiologic Evidence of Progressive Disease 
(application of irRECIST) 
Scenario 
Clinically stable  
Clinically unstable 
Imaging plan 
Treatment plan 
Imaging plan 
Treatment plan 
1st radiologic 
evidence of PD as 
defined by 
RECIST 1.1 
Perform repeat 
scan at ≥ 4 weeks 
to confirm PD. 
May continue study 
treatment at 
Investigator’s discretion 
while awaiting repeat 
scan 
Perform repeat scan 
at ≥ 4 weeks to 
confirm PD per 
Investigator 
discretion only 
Discontinue study 
treatment 
Repeat scan 
confirms PD as 
defined by 
irRECIST 
No additional 
scans required. 
Discontinue treatment 
(exception is possible 
upon consultation with 
Research Base)1 
No additional scans 
required 
N/A 
Repeat scan shows 
SD, PR, or CR as 
defined by 
irRECIST 
Continue regularly 
scheduled scans. 
Continue study treatment 
at Investigator’s 
discretion 
Continue regularly 
scheduled scans. 
May restart study 
treatment if condition 
has improved and/or 
clinically stable per 
Investigator’s discretion 
CR = complete response; PD = progressive disease; PR = partial response; SD = stable disease 
1 If a subject has confirmed radiographic progression per irRECIST (i.e., 2 scans at least 4 weeks apart demonstrating 
progressive disease), but the subject is achieving a clinically meaningful benefit (e.g., experiencing a response in a non-
irradiated lesion), an exception to continue treatment may be considered following consultation with the Research Base. 
 


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9.3 
Definitions of Measurable and Non-Measurable Disease according to RECIST 1.1 
 
9.3.1 Measurable Disease 
A non-nodal lesion is considered measurable if its longest diameter can be accurately measured 
as ≥ 2.0 cm with chest x-ray, or as ≥1.0 cm with CT scan, CT component of a PET/CT, or MRI.  
 
A superficial non-nodal lesion is measurable if its longest diameter is ≥ 1.0 cm in diameter as 
assessed using calipers (e.g. skin nodules) or imaging.  In the case of skin lesions, documentation 
by color photography, including a ruler to estimate the size of the lesion, is recommended. 
 
A malignant lymph node is considered measurable if its short axis is >1.5 cm when assessed by 
CT scan (CT scan slice thickness recommended to be no greater than 5 mm).   
NOTE: Tumor lesions in a previously irradiated area are not considered measurable disease.   
 
9.3.2 Non-Measurable Disease 
All other lesions (or sites of disease) are considered non-measurable disease, including 
pathological nodes (those with a short axis ≥1.0 to <1.5 cm).  Bone lesions, leptomeningeal 
disease, ascites, pleural/pericardial effusions, lymphangitis cutis/pulmonis, inflammatory breast 
disease, and abdominal masses (not followed by CT or MRI), are considered as non-measurable 
as well. 
 
NOTE: ‘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 patient, these are preferred for selection as target lesions.  In 
addition, lymph nodes that have a short axis < 1.0 cm are considered non- pathological (i.e., 
normal) and should not be recorded or followed. 
 
9.3.3 Guidelines for Evaluation of Measurable Disease according to RECIST 1.1 
 
Measurement Methods:   
All measurements should be recorded in metric notation (i.e., decimal fractions of centimeters) 
using a ruler or calipers. 
 
The same method of assessment and the same technique must be used to characterize each 
identified and reported lesion at baseline and during follow-up. For patients having only lesions 
measuring at least 1 cm to less than 2 cm must use CT imaging for both pre- and post-treatment 
tumor assessments.   
 
Imaging-based evaluation is preferred to evaluation by clinical examination when both methods 
have been used at the same evaluation to assess the antitumor effect of a treatment.  
 
9.3.4 Acceptable Modalities for Measurable Disease       
Conventional CT and MRI:  This guideline has defined measurability of lesions on CT scan 
based on the assumption that CT slice thickness is 5 mm or less. If CT scans have slice thickness 
greater than 5 mm, the minimum size for a measurable lesion should be twice the slice thickness.     
 


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As with CT, if an MRI is performed, the technical specifications of the scanning sequences used 
should be optimized for the evaluation of the type and site of disease.  The lesions should be 
measured on the same pulse sequence.  Ideally, the same type of scanner should be used and the 
image acquisition protocol should be followed as closely as possible to prior scans.  Body scans 
should be performed with breath-hold scanning techniques, if possible. 
 
PET-CT:  If the site can document that the CT performed as part of a PET-CT is of identical 
diagnostic quality to a diagnostic CT (with IV and oral contrast), then the CT portion of the PET-
CT can be used for RECIST measurements and can be used interchangeably with conventional 
CT in accurately measuring cancer lesions over time.   
 
Chest X-ray:  Lesions on chest x-ray are acceptable as measurable lesions when they are clearly 
defined and surrounded by aerated lung.  However, CT scans are preferable. 
Physical Examination:  For superficial non-nodal lesions, physical examination is acceptable, but 
imaging is preferable, if both can be done. In the case of skin lesions, documentation by color 
photography, including a ruler to estimate the size of the lesion, is recommended. 
 
FDG-PET:  FDG-PET scanning is allowed to complement CT scanning in assessment of 
progressive disease [PD] and particularly possible 'new' disease.  A ‘positive’ FDG-PET scanned 
lesion is defined as one which is FDG avid with an update greater than twice that of the 
surrounding tissue on the attenuation corrected image; otherwise, an FDG-PET scanned lesion is 
considered ‘negative.’  New lesions on the basis of FDG-PET imaging can be identified 
according to the following algorithm: 
 
• Negative FDG-PET at baseline with a positive FDG-PET at follow-up is a sign of PD 
based on a new lesion. 
 
• No FDG-PET at baseline and a positive FDG-PET at follow-up:   
o If the positive FDG-PET at follow-up corresponds to a new site of disease 
confirmed by CT, this is PD.   
o If the positive FDG-PET at follow-up is not confirmed as a new site of disease on 
CT at the same evaluation, additional follow-up CT scans (i.e., additional follow-
up scans at least 4 weeks later) are needed to determine if there is truly 
progression occurring at that site. In this situation, the date of PD will be the date 
of the initial abnormal PDG-PET scan.   
o If the positive FDG-PET at follow-up corresponds to a pre-existing site of disease 
on CT that is not progressing on the basis of the anatomic images, it is not 
classified as PD. 
 
9.3.5 Measurement at Follow-up Evaluation 
A subsequent scan must be obtained not less than 4 weeks following initial documentation of an 
objective status of either complete response (CR) or partial response (PR).  
 
In the case of stable disease (SD), follow-up measurements must have met the SD criteria at least 
once after study entry at a minimum interval of not less than 6 weeks. 
 


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The cytological confirmation of the neoplastic origin of any effusion that appears or worsens 
during treatment when the measurable tumor has met criteria for response or stable disease is 
mandatory to differentiate between response or stable disease (an effusion may be a side effect of 
the treatment) and progressive disease. 
 
Cytologic and histologic techniques can be used to differentiate between PR and CR in rare cases 
(e.g., residual lesions in tumor types such as germ cell tumors, where known residual benign 
tumors can remain.) 
 
9.4 
Measurement of Effect per RECIST 1.1 
 
9.4.1 Target Lesions & Target Lymph Nodes 
Measurable lesions up to a maximum of 5 lesions, representative of all involved organs, should 
be identified as “Target Lesions” and recorded and measured at baseline.  Lesions to be 
monitored for assessment of response must lie outside the RT field.  These lesions can be non-
nodal or nodal, where no more than 2 lesions are from the same organ and no more than 2 
malignant nodal lesions are selected. NOTE: If fewer than 5 target lesions and target lymph 
nodes are identified (as there often will be), there is no reason to perform additional studies 
beyond those specified in the protocol to discover new lesions.   
 
Target lesions and target lymph nodes should be selected on the basis of their size, be 
representative of all involved sites of disease, but in addition should be those that lend 
themselves to reproducible repeated measurements.  It may be the case that, on occasion, the 
largest lesion (or malignant lymph node) does not lend itself to reproducible measurements in 
which circumstance the next largest lesion (or malignant lymph node) which can be measured 
reproducibly should be selected.   
 
Baseline Sum of Dimensions (BSD):  A sum of the longest diameter for all target lesions plus the 
sum of the short axis of all the target lymph nodes will be calculated and reported as the baseline 
sum of dimensions (BSD).  The BSD will be used as reference to further characterize any 
objective tumor response in the measurable dimension of the disease.   
 
Post-Baseline Sum of the Dimensions (PBSD):  A sum of the longest diameter for all target 
lesions plus the sum of the short axis of all the target lymph nodes will be calculated and 
reported as the post-baseline sum of dimensions (PBSD).  If the radiologist is able to provide an 
actual measure for the target lesion (or target lymph node), that should be recorded, even if it is 
below 0.5 cm.  If the target lesion (or target lymph node) is believed to be present and is faintly 
seen but too small to measure, a default value of 0.5 cm should be assigned.  If it is the opinion 
of the radiologist that the target lesion or target lymph node has likely disappeared, the 
measurement should be recorded as 0 cm. 
 
The minimum sum of the dimensions (MSD) is the minimum of the BSD and the PBSD. 
 
 
 
 


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9.4.2 Non-Target Lesions & Non-Target Lymph Nodes 
Non-measurable sites of disease are classified as non- target lesions or non-target lymph nodes 
and should also be recorded at baseline.  Lesions to be monitored for assessment of response 
must lie outside the RT field. 
 
9.5 
Response criteria per best abscopal response 
A novel aspect of this study is the examination of best abscopal response. Response criteria per 
best abscopal response are identical to RECIST-defined response criteria, with the exception that 
tumor shrinkage by > 20% in only a single representative measurable lesion (not a sum of 
lesions) is sufficient to count as a best abscopal response.  In addition, up to 10 measurable 
lesions per organ may be recorded.  Best abscopal response is a binary variable: present or not 
present.  A subject can experience a best abscopal response at the same time as CR, PR, SD, or 
PD as defined by irRECIST/RECIST 1.1.   
 
See 9.2.2 for guidance on management of cases when a best abscopal response co-occurs with 
PD as defined by irRECIST/RECIST 1.1. 
 
9.6 
Response Criteria per RECIST 1.1 
All target lesions and target lymph nodes followed by 
• CT/MRI/PET-CT/Chest X-ray/physical examination must be measured on re-evaluation.  
Specifically, a change in objective status to either a PR or CR cannot be done without re-
measuring target lesions and target lymph nodes.  
 
NOTE: Non-target lesions and non-target lymph nodes should be evaluated at each assessment, 
especially in the case of first response or confirmation of response.  In selected circumstances, 
certain non-target organs may be evaluated less frequently.  For example, bone scans may need 
to be repeated only when complete response is identified in target disease or when progression in 
bone is suspected. 
 
Evaluation of Target Lesions 
• Complete Response (CR): All of the following must be true: 
o Disappearance of all target lesions. 
o Each target lymph node must have reduction in short axis to <1.0 cm. 
• Partial Response (PR): At least a 30% decrease in PBSD (sum of the longest diameter for 
all target lesions plus the sum of the short axis of all the target lymph nodes at current 
evaluation) taking as reference the BSD. 
• Progression (PD): At least one of the following must be true: 
o At least one new malignant lesion, which also includes any lymph node that was 
normal at baseline (< 1.0 cm short axis) and increased to ≥ 1.0 cm short axis 
during follow-up.  
o At least a 20% increase in PBSD (sum of the longest diameter for all target 
lesions plus the sum of the short axis of all the target lymph nodes at current 
evaluation) taking as reference the MSD (Section 11.41).  In addition, the PBSD 
must also demonstrate an absolute increase of at least 0.5 cm from the MSD.  
o See Section 11.1 on incorporation of irRECIST in determining whether PD on 
RECIST counts as true progression. 


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o See Section 11.32 for details regarding the requirements for PD via FDG-PET 
imaging. 
• Stable Disease (SD): Neither sufficient shrinkage to qualify for PR, nor sufficient 
increase to qualify for PD taking as reference the MSD. 
 
Evaluation of Non-Target Lesions & Non-target Lymph Nodes 
• Complete Response (CR): All of the following must be true: 
o Disappearance of all non-target lesions.  
o Each non-target lymph node must have a reduction in short axis to <1.0 cm. 
• Non-CR/Non-PD: Persistence of one or more non-target lesions or non-target lymph 
nodes. 
• Progression (PD): At least one of the following must be true: 
o At least one new malignant lesion, which also includes  
any lymph node that 
was normal at baseline (< 1.0 cm short axis) and increased to ≥  
1.0 cm short 
axis during follow-up.   
o Unequivocal progression of existing non-target lesions and non-target lymph 
nodes.  (NOTE:  Unequivocal progression should not normally trump target 
lesion and target lymph node status.  It must be representative of overall disease 
status change.)  
o See Section 11.1 on incorporation of irRECIST in determining whether PD on 
RECIST counts as true progression 
o See Section 11.32 for details regarding the requirements for PD via FDG-PET 
imaging. 
 
9.6.1 Overall Objective Status per RECIST 1.1 
The overall objective status for an evaluation is determined by combining the patient’s status on 
target lesions, target lymph nodes, non-target lesions, non-target lymph nodes, and new disease 
as defined in the following tables:  
 
Target Lesions & 
Target Lymph Nodes 
Non-Target Lesions & 
Non-Target Lymph 
Nodes 
New  
Sites of Disease 
Overall Objective 
Status 
CR 
CR 
No 
CR 
CR 
Non-CR/Non-PD 
No 
PR 
PR 
CR 
Non-CR/Non-PD 
No 
PR 
CR/PR 
Not All Evaluated* 
No 
PR** 
SD 
CR 
Non-CR/Non-PD 
Not All Evaluated* 
No 
SD 
Not all Evaluated 
CR 
Non-CR/Non-PD 
Not All Evaluated* 
No 
Not Evaluated 
(NE) 
PD 
Unequivocal PD 
CR 
Non-CR/Non-PD 
Not All Evaluated* 
Yes or No 
PD 


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Target Lesions & 
Target Lymph Nodes 
Non-Target Lesions & 
Non-Target Lymph 
Nodes 
New  
Sites of Disease 
Overall Objective 
Status 
CR/PR/SD/PD/Not all 
Evaluated 
Unequivocal PD 
Yes or No 
PD 
CR/PR/SD/PD/Not all 
Evaluated 
CR 
Non-CR/Non-PD 
Not All Evaluated* 
Yes 
PD 
 
Symptomatic Deterioration: Patients with global deterioration of health status requiring 
discontinuation of treatment without objective evidence of disease progression at that time, and 
not either related to study treatment or other medical conditions, should be reported as PD due to 
“symptomatic deterioration.”  Every effort should be made to document the objective 
progression even after discontinuation of treatment due to symptomatic deterioration.  A patient 
is classified as having PD due to “symptomatic deterioration” if any of the following occur that 
are not either related to study treatment or other medical conditions: 
• Worsening of tumor-related symptoms. 
• Decline in performance status of >1 level on ECOG scale. 
 
9.7 
Descriptive Factors   
• RT to bone on this study: ever vs. never 
• Number of metastases:  3 vs. 4-6 vs. 7-10 vs. 11 or more 
• Number of anatomic regions involved (1-3: thoracic, abdominal/pelvic, or bone) 
• Type of anatomic region involved (thoracic [yes/no], abdominal/pelvic [yes/no], bone 
[yes/no]) 
 
9.8 
Treatment/Follow–Up Decision at Evaluation of Patient   
 
9.8.1 CR, PR or SD 
Patients who are CR, PR, or SD will continue treatment per protocol.  Patients who achieve a CR 
should receive at least 2 further cycles of study treatment beyond the confirmation of CR.  While 
subsequent continuation of study treatment is at the Investigator’s discretion, it is strongly 
encouraged for pembrolizumab monotherapy to be continued.   
 
9.8.2 PD 
Patients who develop PD based on RECIST and, if appropriate, irRECIST while receiving 
therapy will continue to be monitored.  These patients should be treated, at the discretion of their 
treating physician, with alternative chemotherapy if their clinical status is good enough to allow 
further therapy. 
• Patients who develop PD in the CNS only should receive whole brain radiotherapy 
(WBRT) and continue treatment on study after completion of WBRT. 
• Patients who develop non-CNS PD by RECIST and irRECIST at any time should go to 
event monitoring.   
 
A maximum of 24 continuous months of pembrolizumab (35 cycles) can be administered.   If 
study treatment is discontinued while a patient is experiencing CR, PR, or SD, the patient should 


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undergo active monitoring.  If the cancer recurs, up to 12 months of further pembrolizumab 
monotherapy can be administered if deemed appropriate by the site investigator.   
 
A patient is deemed ineligible if after registration, it is determined that at the time of registration, 
the patient did not satisfy each and every eligibility criteria for study entry.  The patient may 
continue treatment off-protocol at the discretion of the treating physician as long as there are no 
safety concerns, and the patient was properly registered. The patient will go directly to the event-
monitoring phase of the study (or off study treatment, if applicable).  If the patient received 
treatment, all data up until the point of confirmation of ineligibility must be submitted.  If the 
patient never received treatment, on-study material must be submitted.  
 
A patient is deemed a major violation, if protocol requirements regarding treatment in Cycle 1 of 
the initial therapy are severely violated that evaluability for primary end point is questionable.  
All data up until the point of confirmation of a major violation must be submitted.  The patient 
will continue to be monitored per protocol.  The patient may continue treatment off-protocol at 
the discretion of the treating physician as long as there are no safety concerns, and the patient 
was properly registered.   
 
A patient is deemed a cancel if he/she is removed from the study for any reason before any study 
treatment is given.  No further data submission is necessary. 
 
10. 
DRUG INFORMATION 
 
10.1 
Pembrolizumab (Keytruda®) 
Please refer to the current version of the Investigator’s Brochure (IB) for additional information 
regarding this drug. 
 
Pembrolizumab is a potent and highly selective humanized mAb of the IgG4/kappa isotype 
designed to directly block the interaction between PD-1 and its ligands, PD-L1 and PD-L2.  
KeytrudaTM (pembrolizumab) has recently been approved in the United Stated for the treatment 
of subjects with unresectable or metastatic melanoma and disease progression following 
ipilumumab and, if BRAF V600 mutation positive, a BRAF inhibitor. It is also approved for the 
treatment of subjects with PD-L1 positive metastatic NSCLC who have disease progression on 
or after platinum chemotherapy. Please refer to the current version of the Keytruda® prescribing 
information and the pembrolizumab Investigator’s Brochure (IB) for additional information 
regarding this drug. 
 
10.1.1 Supplier/How Supplied 
Merck will supply pembrolizumab at no charge to subjects participating in this clinical trial, as 
summarized in the table below. 
 
Product Name & Potency 
Dosage Form 
Pembrolizumab 100 mg/ 4mL 
Solution for Injection 


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10.1.2 Preparation 
Please refer to the Pharmacy Manual for a comprehensive description of pembrolizumab 
preparation. 
 
10.1.3 Storage and Stability 
Clinical supplies must be stored in a secure, limited-access location under the storage conditions 
specified on the label.   
 
Clinical supplies may not be used for any purpose other than that stated in the protocol. 
 
10.1.4 Handling and Disposal 
Upon completion or termination of the study, all unused and/or partially used investigational 
product will be destroyed at the site per institutional policy. It is the Investigator’s responsibility 
to arrange for disposal of all empty containers, provided that procedures for proper disposal 
have been established according to applicable federal, state, local and institutional guidelines and 
procedures, and provided that appropriate records of disposal are kept. 
 
10.1.5 Dispensing, Returns and Reconciliation 
The investigator is responsible for keeping accurate records of the clinical supplies received from 
Merck or designee, the amount dispensed to and returned by the subjects and the amount 
remaining at the conclusion of the trial.  
 
10.1.6 Use in Pregnancy and Nursing Women 
Based on its mechanism of action, pembrolizumab can cause fetal harm when administered to a 
pregnant woman. Subjects who are pregnant or planning to become pregnant are not eligible for 
enrollment. It is unknown whether pembrolizumab 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. 
 
10.1.7 Adverse Events 
Please refer to the current prescribing information for pembrolizumab, and to the current version 
of the Investigator’s Brochure for a complete list of adverse events.  
 
The most common adverse reactions (reported in ≥20% of subjects in clinical trials of 
pembrolizumab in melanoma subjects) included fatigue, pruritus, rash, constipation, diarrhea, 
nausea and decreased appetite. The most common adverse reactions (reported in ≥20% of 
subjects in clinical trials of pembrolizumab in NSCLC subjects) included fatigue, decreased 
appetite, dyspnea and cough. 
 
Pembrolizumab is an immunomodulatory agent, and based on this mechanism of action, immune 
mediated adverse events (IMAEs) 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 mellitus, 
uveitis, and nephritis. After a recent review of data, events newly characterized as identified risks 
also include pancreatitis, myositis, and severe skin reaction. The majority of IMAEs was mild to 


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moderate in severity, manageable with appropriate care, and rarely required discontinuation of 
therapy. 
 
Further details around frequency, reporting, and management of IMAEs 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. 
 
11. 
ADVERSE EVENTS 
The descriptions and grading scales found in the NCI CTCAE v4 will be utilized for AE 
assessment. A copy of the CTCAE v4 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 meets any of the following: 
• 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. Hospitalization related to convenience (e.g. transportation issues etc.) will 
not be considered an SAE  


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• Results in persistent or significant disability/incapacity 
• Is a congenital anomaly or birth defect 
• Is a new cancer (that is not a condition of the study) 
• 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. 
 
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 study drug(s).  
• AEs will be recorded regardless of whether or not they are considered related to the study 
drug(s).   
• All AEs that are possibly, probably, or definitely related will be recorded in the subject’s 
medical record and on the appropriate study specific eCRF form within the EDC system. 
• AEs considered related to study drug(s) will be followed until resolution to Grade ≤ 1 or 
baseline, deemed clinically insignificant, and/or until a new anti-cancer treatment starts, 
whichever occurs first. 
• Asymptomatic laboratory abnormalities that do not require treatment will not be collected 
as adverse events. 
 


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11.2.2 Serious Adverse Events (SAEs) 
 
11.2.2.1 
Site Requirements for Reporting SAEs to HCRN 
• The time period for expedited reporting begins at registration and continues through 90 
days following cessation of treatment, or 30 days following cessation of treatment if the 
subject initiates new anticancer therapy, whichever is earlier.  
• SAEs will be reported on the SAE/ECI Submission Form. 
• For timelines of reporting, please see the table below. 
• SAEs include events related and unrelated to the study drug(s).  
• All SAEs will be recorded in the subject’s medical record and on the appropriate study 
specific eCRF form within the EDC system.   
• All SAEs regardless of relation to study drug will be followed until resolution to ≤ Grade 
1 or baseline and/or deemed clinically insignificant and/or until a new anti-cancer 
treatment starts, whichever occurs first. 
 
REPORTING REQUIREMENTS FOR SERIOUS ADVERSE EVENTS (21 CFR Part 312) 
NOTE: Investigators MUST immediately report to HCRN ANY Serious Adverse Events, 
whether or not they are considered related to the investigational agent(s)/intervention (21 CFR 
312.64).  HCRN will then report to Merck.  The definition of a serious adverse event can be 
found above in Section 11.1.2. 
ALL SERIOUS adverse events that meet the above criteria in Section 11.1.2 be immediately 
reported within the timeframes detailed in the table below. 
Hospitalization 
Grade 1 
Timeframe 
Grade 2 
Timeframes 
Grade 3 
Timeframe 
Grade 4 & 5 
Timeframe 
Resulting in 
Hospitalization 
≥ 24 hrs 
7 Calendar 
Days 
 
24-Hour; 3 
Calendar Days 
Not resulting in 
Hospitalization 
≥ 24 hrs 
Not required 
7 Calendar 
Days 
Expedited AE reporting timelines are defined as: 
o “24-Hour; 3 Calendar Days” - The AE must initially be reported within 24 hours of 
learning of the AE, followed by a complete expedited report within 3 calendar days of the 
initial 24-hour report. 
o “7 Calendar Days” - A complete expedited report on the AE must be submitted within 7 
calendar days of learning of the AE. 
1Serious adverse events that occur more than 30 days after the last administration of investigational 
agent/intervention and have an attribution of possible, probable, or definite require reporting as follows: 
 
Expedited 24-hour notification followed by complete report within 3 calendar days for: 
• 
All Grade 4, and Grade 5 AEs 
 
Expedited 7 calendar day reports for: 
• 
Grade 2 adverse events resulting in hospitalization or prolongation of hospitalization 
• 
Grade 3 adverse events  
 


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The site will submit the completed SAE/ECI Submission Form to HCRN within the timeframes 
described in the table above. 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.   
 
The original copy of the SAE/ECI Submission Form and the email correspondence must be kept 
within the study file at the study site. 
 
Until the SAE has resolved (see resolution guidelines listed in 11.2.2.1), sites must submit 
follow-up SAE/ECI Submission Forms when the AE has decreased by at least 1 Grade level to 
HCRN electronically to safety@hoosiercancer.org.   
 
11.2.3 Overdose 
For purposes of this trial, an overdose will be defined as any dose of 1,000 mg or greater (≥ 5 
times the indicated dose). No specific information is available on the treatment of overdose of 
pembrolizumab. In the event of overdose, pembrolizumab should be discontinued and the subject 
should be observed closely for signs of toxicity.  Appropriate supportive treatment should be 
provided if clinically indicated. 
 
If an adverse event is associated with the overdose of pembrolizumab, the adverse event is 
reported as a serious adverse event, even if no other seriousness criteria are met.  
 
If a dose of pembrolizumab meeting the protocol definition of overdose is taken without any 
associated clinical symptoms or abnormal laboratory results, the overdose is reported as a non-
serious ECI, using the terminology “accidental or intentional overdose without adverse effect.”  
 
All reports of overdose with and without an adverse event must be reported within 1 business 
day to HCRN and within 1 business day to Merck Global Safety. (Attn: Worldwide Product 
Safety; FAX 215 993-1220). 
 
11.2.4 Reporting of Pregnancy and Lactation  
Although pregnancy and lactation are not considered adverse events, it is the responsibility of 
investigators or their designees to report any pregnancy or lactation in a subject (spontaneously 
reported to them) that occurs during the trial. 
 
Pregnancies and lactations that occur after the consent form is signed but before treatment 
allocation must be reported by the investigator if they cause the subject to be excluded from the 
trial, or are the result of a protocol-specified intervention, including but not limited to washout or 
discontinuation of usual therapy, diet, placebo treatment or a procedure.   
 
Pregnancies and lactations that occur from the time of treatment allocation/randomization 
through 120 days following cessation of pembrolizumab, or 30 days following cessation of 
treatment if the subject initiates new anticancer therapy, whichever is earlier, must be reported by 
the investigator.  All reported pregnancies must be followed to the completion/termination of the 
pregnancy. Pregnancy outcomes of spontaneous abortion, missed abortion, benign hydatidiform 
mole, blighted ovum, fetal death, intrauterine death, miscarriage and stillbirth must be reported 


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as serious events (Important Medical Events). If the pregnancy continues to term, the outcome 
(health of infant) must also be reported. 
 
The outcome of the pregnancy will be reported to Hoosier Cancer Research Network (HCRN) 
within 1 business day and HCRN will report to Merck 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). 
 
11.2.5 Events of Clinical Interest (ECI) 
Selected non-serious and serious adverse events are also known as Events of Clinical Interest 
(ECI) and must be recorded as such on the SAE/ECI Submission Form.  ECIs (both non-serious 
and serious adverse events) identified from the date of first dose through 90 days following 
cessation of treatment, or 30 days after the initiation of a new anticancer therapy, whichever is 
earlier, will be reported from the site to HCRN within one business day.  HCRN will report to 
Merck within one business day. 
Events of clinical interest for this trial include: 
 
1. an overdose of Pembrolizumab, as defined in Section 10.3 - Definition of an Overdose and 
Reporting Guidelines, that is not associated with clinical symptoms or abnormal laboratory 
results.  
 
2. an elevated AST or ALT lab value that is greater than or equal to 3X the upper limit of 
normal and an elevated total bilirubin lab value that is greater than or equal to 2X the upper 
limit of normal and, at the same time, an alkaline phosphatase lab value that is less than 2X 
the upper limit of normal, as determined by way of protocol-specified laboratory testing or 
unscheduled laboratory testing.* 
 
*NOTE:  These criteria are based upon available regulatory guidance documents. The 
purpose of the criteria is to specify a threshold of abnormal hepatic tests that may require an 
additional evaluation for an underlying etiology.  The trial site guidance for assessment and 
follow up of these criteria can be found in the Investigator Trial Master File (or equivalent). 
 
11.3 
HCRN Requirements for Reporting SAEs to Merck  
HCRN will report all SAEs to Merck within 1 business day of receipt of the SAE/ECI 
Submission Form from a site. Follow-up information will be provided to Merck as it is received 
from site.  Merck Global Safety (Attn: Worldwide Product Safety; FAX 215 993-1220). 
 
11.4 
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.   
 
 
 


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11.5 
HCRN Responsibilities to FDA 
For protocols exempt from the requirements of an IND, the above stated requirements are not 
applicable. 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. 
 
12. 
STATISTICAL METHODS 
 
12.1 
Study Design 
This is a proof-of-principle single-arm phase 2 study in patients with previously treated CUP.  
All patients receive pembrolizumab combined with RT to a metastatic site, so as to induce an 
abscopal tumor response.  The treatment combination will be repeated with RT delivery to a 
second metastatic site in a non-overlapping RT field.   
 
The results will be compared with historical control.   The primary endpoint is the confirmed 
response rate (RR) in a non-irradiated site based on best responding abscopal lesion.  This study 
will also evaluate the following secondary endpoints: RR in a non-irradiated site based on 
RECIST 1.1, adverse events, progression-free survival (PFS), overall survival (OS), time-to-
progression (TTP), and disease control rate (DCR). 
 
12.2 
Endpoints 
 
12.2.1 Definition of Primary Endpoint 
The primary endpoint for this study will assess the confirmed response rate in the best 
responding abscopal lesion in a non-irradiated site.  Best abscopal response is defined as the 
frequency of patients whose best responding abscopal lesion demonstrates at least a 30% 
decrease in its longest diameter from baseline (Golden et al., 2015).  Responses need to be 
confirmed (2 consecutive responses at least 4 weeks apart) to count as a response (see Section 
11).  All eligible patients who are registered and start treatment will be evaluable for response.   
 
The study is powered based on the confirmed response rate.  In the largest second-line CUP trials 
where patients received cytotoxic therapy, patients received gemcitabine alone (N = 39) 
(Hainsworth et al., 2001) or gemcitabine plus irinotecan (N = 40) (Hainsworth et al., 2005). RRs 
were 8% and 10%, PFS was not reported, and OS was 3 months and 4.5 months— all 
respectively.   
 
A Simon’s optimal two-stage design will be utilized (see below). This design has 80% power to 
test the hypothesis of 15% improvement in RR (i.e. 24%) from a historical control of 9%, at one-
sided significance level of 0.10.  The 2-stage design details are below, where accrual will 
continue during the interim analysis phase unless we observe a much higher than expected 
accrual rate or toxicity rate. 
 
 
 


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Interim Analysis:  Enroll 14 eligible patients.  If at least 2 confirmed responses are observed in 
the first 14 eligible patients, we will continue the trial to the second stage and continue the study 
to a full accrual of 29 eligible patients.  Otherwise, the study will be permanently closed due to 
lack of efficacy.   
 
Final Analysis:  If the trial is a success during the interim analysis, we will enroll another 15 
eligible patients to the second and final stage (29 eligible total).  If at least 5 confirmed responses 
are observed in the first 29 eligible patients (17%), the treatment will be considered worthy of 
further investigation.  Otherwise, the study will be permanently closed due to lack of efficacy.   
 
12.2.2 Definition and Analysis of Secondary Endpoints 
• 
Response rate (RR) by RECIST 1.1. 
 
• 
Adverse events:  The maximum grade for each type of adverse event will be summarized 
using CTCAE version 4.0.  The frequency and percentage of grade 2+ adverse events will 
be assessed.  We will also explore the treatment related adverse events as well 
(toxicities).   
 
• 
Progression-free survival (PFS) is defined as the time from registration to the first of 
either disease progression or death from any cause, where disease progression will be 
determined based on RECIST 1.1 criteria.  PFS will be estimated using the Kaplan-Meier 
method, where patients that are alive and progression-free will be censored on their last 
tumor evaluation date. 
 
• 
Overall Survival (OS) is defined as the time from registration to death from any cause.  
OS will be estimated using the Kaplan-Meier method. 
 
• 
Time-to-Progression (TTP) is defined as the time from registration to disease 
progression, where patients that are progression-free will be censored on their last tumor 
evaluation date.  TTP will be estimated using the cumulative incidence method, where 
deaths will be treated as competing risks. 
 
• 
Disease Control Rate (DCR) is defined as the frequency (%) of patients who have a best 
response of PR, CR, or Stable disease.   
 
• 
The association between RR (using RECIST 1.1 (informed by irRECIST)) with other 
clinical endpoints (e.g., OS, PFS, etc.) will be performed.  For time-to-event endpoints, 
we will use Cox proportional hazards models and Kaplan-Meier methodology.  This 
analysis of RR as a predictor of OS, PFS, etc. will be based on a landmark analysis at 
various time points of interest post-baseline that correspond to expected tumor 
assessment time points.  This will eliminate the possible response bias of patients living 
longer or doing better because they lived long enough to respond to treatment. For any 
associations of RR with categorical data, we will use Chi-square or Fisher’s exact tests 
along with logistic regression models.  Descriptive statistics and graphical techniques will 
be used to summarize this data.  Given the small sample size, all these analyses will be 
considered hypothesis generating and exploratory.   


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12.3 
Sample Size and Accrual 
A maximum of 29 evaluable patients will be accrued onto this phase II study unless the study is 
closed early for excessive toxicity or lack of efficacy.  To account for potential ineligibles, major 
treatment violations during Cycle 1, or cancellations, we will enroll an additional 5 patients, for a 
total of 34 patients.  The expected accrual rate is about 1.5 patients per month across the Hoosier 
sites.  With this accrual rate, we expect to finish accrual within about 23 months, assuming we 
accrue 34 total patients.  We anticipate that the study will take approximately 36 months to 
complete.  This allows a 9-month follow-up for the final patient enrolled, along with data entry, 
data clean-up, and analysis. Given the expected accrual rate is around 1.5 patients per month, it is 
expected that the study will take around 23 months to fully accrue.  We plan to monitor the 
accrual continually and if we only end up accruing 6 patients or less in the first year (after study 
activation), we will consider stopping the trial for slow accrual. 
 
12.4 
Assessment of Safety 
Any subject who receives at least one dose of treatment on this protocol will be evaluable for 
toxicity. The National Cancer Institute (NCI) Common Terminology Criteria for Adverse Events 
(CTCAE) v4 will be used. The Study Calendar shows the schedule of toxicity assessments. 
 
12.5 
Assessment of Efficacy 
All eligible patients who are registered and receive at least one dose of trial drug will be 
evaluable for all efficacy endpoints.    
 
12.6 
Translational Studies 
We plan to correlate the tumor markers (MSI, tissue PDL1 expression, mutational load, genomic 
mutation analysis, BIM/PD-1 expression in tumor infiltrating T cells, and other immune markers, 
and tumor tissue of origin testing) and plasma-based markers (soluble PD-L1 and T cell 
subpopulations) with clinical endpoints like confirmed response, best responding abscopal 
lesion, time-to-event outcomes, and adverse events.  Statistical and graphical techniques will be 
used to evaluate the relationships between tumor markers and clinical data. For time-to-event 
endpoints, we will use Cox proportional hazards models, and for confirmed response we will use 
Logistic regression models. In addition, we will use the Chi-square or Fisher’s exact tests to test 
the association between categorical marker data and confirmed response/adverse events. 
Continuous marker data will be assessed using 2-sample t-tests (or the nonparametric equivalent 
for non-normal data).  Descriptive statistics and graphical techniques will be used to summarize 
this data.  Given the small sample size, all these analyses will be considered hypothesis 
generating and exploratory.   
 
12.7 
Interim Analysis/Criteria for Stopping Study 
Interim Analysis (based on primary endpoint):  Enroll 14 eligible patients.  If at least 2 
confirmed responses are observed in the first 14 eligible patients, we will continue the trial to the 
second stage and continue the study to a full accrual of 29 eligible patients.  Otherwise, the study 
will be permanently closed due to lack of efficacy.  Accrual will continue during the interim 
analysis phase unless we observe a much higher than expected accrual rate or toxicity rate. 
 
 
 


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Final Analysis (based on primary endpoint):  If the trial is a success during the interim analysis, 
we will enroll another 15 eligible patients to the second and final stage (29 eligible total).  If at 
least 5 confirmed responses are observed in the first 29 eligible patients (17%), the treatment will 
be considered worthy of further investigation.  Otherwise, the study will be permanently closed 
due to lack of efficacy.   
 
Adverse Event Stopping Rule:  The stopping rule specified below is based on the knowledge 
available at study development. We note that the Adverse Event Stopping Rule may be adjusted 
in the event of either (1) the study re-opening to accrual after any temporary suspension or (2) at 
any time during the conduct of the trial and in consideration of newly acquired information 
regarding the adverse event profile of the treatment(s) under investigation. The study team may 
also choose to suspend accrual because of unexpected adverse event profiles that have not 
crossed the specified rule below. 
 
Accrual will be temporarily suspended to this study if at any time we observe events considered 
at least possibly related to study treatment (i.e., an adverse event with attribute specified as 
"possible", "probable", or "definite") that satisfy any of the following criteria: 
• If at any time, 4 of the initial 10 treated patients or 40% or more of all patients (i.e. 
when accrual is greater than 10 patients) have experienced a Grade 4 non-
hematologic adverse event. 
• If at any time, 2 of the initial 10 treated patients or 20% or more of all patients (i.e., 
when accrual is greater than 10 patients) have experienced a Grade 5 adverse event 
(non -progressive disease). 
 
13. 
TRIAL MANAGEMENT 
 
13.1 
Data and Safety Monitoring Plan (DSMP) 
The study will be conducted with guidance from the Mayo Clinic Cancer Center’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). 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 
 


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During the active treatment phase of this trial, the DSMB will review the above information at 
least every 6 months.  The DSMB will provide a recommendation to the sponsor-investigator 
after all information is reviewed. 
 
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.  
 
The trial site may also be subject to quality assurance audit by Merck 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 Merck or its designee as well as 
inspection by appropriate regulatory agencies. 
 
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. All data will be collected and entered into the EDC 
system by study site personnel from participating institutions.   
 


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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. 
 
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, Merck, 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.  
 


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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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