TREATMENT RESISTANCE FOLLOWING ANTI-CANCER THERAPIES (TRANSLATE)
Phase: NA
Conditions: Disease Progression
Sponsor: Pfizer

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TREATMENT RESISTANCE FOLLOWING ANTI-CANCER THERAPIES
(TRANSLATE)
Therapeutic Area (TA):
Oncology
Protocol Number:
A9001502
This document and accompanying materials contain confidential information belonging to Pfizer.  Except as 
otherwise agreed to in writing, by accepting or reviewing these documents, you agree to hold this information in 
confidence and not copy or disclose it to others (except where required by applicable law) or use it for 
unauthorized purposes.  In the event of any actual or suspected breach of this obligation, Pfizer must be promptly 
notified.

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Document History
Document
Version Date
Summary of Changes and Rationale
Amendment 2
12 May 2020
Schedule of Activities updated to reflect 
changes to Informed Consent and 
Screening process.
Section 3. Study design description
updated to reflect changes to Informed 
Consent and Screening process.
Figure 1 Study schema updated to reflect 
changes to Informed Consent and 
Screening process.
Table 1 updated to reflect changes to 
permitted anti-cancer therapies and 
estimated cohort size.
Inclusion Criterion 1c revised to allow 
specific axitinib combinations in order to 
reflect current standard of care therapy.
Patient enrollment cohorts 1 and 2 
combined and cohort sample size 
decreased by a total of 50 in order to 
reflect current standard of care therapy.
Prior Protocol Administrative 
Clarification Letters (PACLs) 
incorporated (Section 4.1).
Pre-screening and main Informed 
Consent processes merged in order to 
simplify the consent process (Section 5).
Window defined for de novo tumor 
tissue biopsy laboratory shipment in 
order to allow operational flexibility 
while maintaining timely return of Next 
Generation Sequencing analyses 
(Section 5.2.1.2).
Window defined for research blood 
draws in order to allow operational 
flexibility while maintaining a collection 
date contemporaneous with the de novo
tumor tissue biopsy (Section 5.2.1.3).

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Clarification added regarding screening 
(Section 5.1), re-screening (Section 
5.1.5), enrollment (Section 5.2), and 
follow-up (Section 5.3) activities.
Clarification added regarding allowing 
patients to participate in other concurrent 
investigational treatment clinical trials, 
with prior Sponsor agreement (Section 
5.1.2).
Clarification added regarding adverse 
event reporting.
Sample size determination re-worded for 
clarity.
 
 
 
Country-Specific (France) Contrat 
Unique added to Appendix.
Administrative updates made throughout 
to ensure consistent terminology.
Amendment 1
10 Jun 2019
Added country specific request for 
Austria to include pregnancy testing in 
the Schedule of Activities.
Added country specific request for 
Austria to include exclusion criterion for 
pregnant/breast-feeding patients.
Added Section 6.1.3 Pregnancy Testing 
(Specific to Austria).
Original protocol
03 Aug 2018
Not applicable (N/A).
This amendment incorporates all revisions to date, including amendments made at the 
request of country health authorities and IRBs/ECs.
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TABLE OF CONTENTS
LIST OF TABLES.....................................................................................................................6
LIST OF FIGURES ...................................................................................................................6
APPENDICES ...........................................................................................................................6
SCHEDULE OF ACTIVITIES..................................................................................................7
1. INTRODUCTION AND RATIONALE..............................................................................10
1.1. Clinical Mechanisms of Resistance.........................................................................10
1.1.1. Immune Checkpoint (PD-1/PD-L1) Inhibition...........................................11
1.1.1.1. The Antigenicity of the Tumor..................................................11
1.1.1.2. Tumor Cell Intrinsic Immunogenicity.......................................11
1.1.1.3. Tumor Cell Extrinsic Immunogenicity .....................................12
1.1.2. Cell Cycle and CDK4/6 Inhibition .............................................................13
1.1.2.1. Preclinical Mechanisms of Resistance ......................................13
1.1.2.2. Clinical Mechanisms of Resistance...........................................15
1.1.3. Androgen Receptor.....................................................................................15
1.1.3.1. AR Point Mutations...................................................................16
1.1.3.2. AR Amplification......................................................................16
1.1.3.3. AR Splice Variants....................................................................16
1.1.3.4. Trans-differentiation as a Mechanism of AR Resistance..........17
1.1.3.5. Perturbation of Cell Cycle.........................................................17
1.1.4. DNA Damage Response and PARP Inhibition ..........................................18
1.2. Potential Risks and Possible Benefits .....................................................................20
2. STUDY OBJECTIVES AND ENDPOINTS.......................................................................20
3. STUDY DESIGN.................................................................................................................22
4. PATIENT ELIGIBILITY CRITERIA.................................................................................24
4.1. Inclusion Criteria.....................................................................................................24
4.2. Exclusion Criteria....................................................................................................26
5. STUDY PROCEDURES .....................................................................................................27
5.1. Screening (Study Day -45 to Day 1) .......................................................................27
5.1.1. Informed Consent .......................................................................................27
5.1.2. Preliminary Eligibility Assessment ............................................................27
5.1.3. Archival Tumor Tissue Assessment...........................................................27
5.1.4. Confirmation of Eligibility .........................................................................28

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5.1.5. Re-Screening...............................................................................................28
5.2. Enrollment (Study Day 1) .......................................................................................28
5.2.1. Biospecimen Collection..............................................................................29
5.2.1.1. Archival Tumor Tissue Submission..........................................29
5.2.1.2. De Novo Tumor Biopsy.............................................................29
5.2.1.3. Research Blood Specimens .......................................................30
5.2.2. Additional Medical and Oncological History.............................................30
5.2.3. Pregnancy Testing (Specific to Austria) .....................................................30
5.3. Post-Biospecimen Collection Follow-Up (≤30 days after receipt of NGS 
results at the provider’s facility)................................................................................31
5.3.1. NGS Results Consultation..........................................................................31
5.3.3. Next Anti-Cancer Therapy .........................................................................31
5.4. Patient Withdrawal..................................................................................................31
6. ASSESSMENTS..................................................................................................................32
6.3. Pregnancy Testing (Specific to Austria)..................................................................33
7. SAFETY ..............................................................................................................................33
7.1. Adverse Events........................................................................................................33
7.1.1. Research Related Injury..............................................................................33
8. DATA ANALYSIS/STATISTICAL METHODS...............................................................34
8.1. Analysis Population.................................................................................................34
8.2. Sample Size Determination.....................................................................................34
8.3. Efficacy Analysis ....................................................................................................35
8.4. Safety Analysis........................................................................................................35
8.5. Analysis of Other Endpoints ...................................................................................35
8.5.1. Analysis of Primary Endpoint ....................................................................35
8.5.2. Analysis of Secondary Endpoints...............................................................35
8.5.3. Analysis of Asset Specific Secondary Endpoints.......................................36
8.6. Interim Analysis ......................................................................................................37
9. QUALITY CONTROL AND QUALITY ASSURANCE...................................................37
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10. DATA HANDLING AND RECORD KEEPING .............................................................38
10.1. Case Report Forms/Data Collection Tools/Electronic Data Record.....................38
10.2. Record Retention...................................................................................................38
11. ETHICS..............................................................................................................................39
11.1. Institutional Review Board/Ethics Committee......................................................39
11.2. Ethical Conduct of the Study ................................................................................39
11.3. Patient Information and Consent...........................................................................39
12. PUBLICATIONS BY INVESTIGATORS........................................................................40
13. REFERENCES ..................................................................................................................42
LIST OF TABLES
Table 1.
Targeted Biology and Disease Cohorts ................................................................23
Table 2.
Sample Size And 95% Score Based Confidence Intervals For Change In 
The Frequency Of Gene Alterations After Disease Progression..........................35
Table 3.
Analysis of Asset Specific Secondary Endpoints.................................................36
LIST OF FIGURES
Figure 1.
Study Schema .......................................................................................................23
APPENDICES
Appendix 1. Abbreviations......................................................................................................49
Appendix 4. Country-Specific Requirements..........................................................................56
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SCHEDULE OF ACTIVITIES
The Schedule of Activities table provides an overview of the protocol visits and procedures.  Refer to STUDY PROCEDURES and 
ASSESSMENTS sections of the protocol for detailed information on each procedure and assessment required for compliance with the 
protocol. 
The investigator may schedule visits (unplanned visits) in addition to those listed in the Schedule of Activities table, in order to 
conduct evaluations or assessments required to protect the well-being of the patient.
Visit Identifiers
Screening
Day -45 to Day 1
Enrollment
Biospecimen Collection 
Day 1
Post-Biospecimen Follow-up
≤30 days after receipt of NGS 
results at the provider’s facility
Informed Consent1
X
Preliminary eligibility assessment2
X
Archival tumor tissue assessment3
X
Confirmation of eligibility4
X
Additional medical and oncological history5
X
Archival tumor tissue submission6
X
Procedures 
De novo tumor biopsy7
X
Research blood collection8
a.
circulating free (cf) DNA sequence analysis;
b.
germline DNA sequence analysis;
c.
translational analyses.
X
Pregnancy test9
X
X
X
Clinical Assessments
Adverse events monitoring10
X
X
NGS results consultation11
X
Other Assessments
Record next anti-cancer treatment following de novo
biopsy14
X
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cf = circulating free; DNA=deoxyribonucleic acid; NGS = Next generation sequencing.
1.
Informed Consent: Study consent is obtained before undergoing study-related activities and should be provided as close as possible to 
the day of biospecimen collection, optimally the day of, but no more than 45 days before the de novo tumor biopsy or research blood 
collection
2.
Preliminary Eligibility Assessment: Confirmation of demographics, histological diagnosis, prior systemic anti-cancer therapy 
regimens (and discontinuation reasons, if appropriate) or radiation therapy. Patients who have started new anti-cancer therapy prior to 
performing the de novo tumor tissue biopsy or research blood collection are not eligible and should be discontinued from the study. 
Screen failures are defined as participants who consent to participate in the clinical study but do not subsequently undergo biospecimen 
collection (ie, de novo tumor biopsy and research blood collection are not performed).
3.
Archival tumor tissue assessment: The archival tumor tissue from the most recent primary or metastatic tumor biopsy or resection is 
checked to confirm availability and adequacy of sample. See protocol Section 5.1.3.  If the archival tumor tissue does not meet protocol 
requirements, the patient must be discontinued and should not proceed with any further enrolment activities; the participant is recorded 
as a screen failure. 
4.
Confirmation of Eligibility: Confirmation of any changes to systemic anti-cancer therapy regimens, radiation therapy or reasons for 
treatment discontinuation; confirmation of radiographic evidence of disease progression for the lesion being targeted for the de novo
tumor tissue biopsy.
5.
Additional medical and oncological history: Following Enrollment, smoking history, prior cancer surgery, duration and best overall 
response for each anti-cancer therapy is recorded.
6.
Archival tumor tissue submission: If the archival tumor tissue is confirmed as meeting protocol requirements (Section 5.1.3), the 
patient may be considered for enrollment. The archival tumor tissue must be processed and sent to the Central Laboratory as detailed in 
the laboratory manual.
7.
De novo tumor biopsy: Occurs only if archival tumor tissue meets protocol requirements (see Section 5.1.3).  De novo biopsy is 
performed according to institutional guidelines. Pre-biopsy clinical and laboratory assessments per institutional guidelines must be 
completed and reviewed prior to biopsy procedure.  Anti-coagulation therapy needs to be held prior to the biopsy according to 
institutional guidelines.  The biopsy must be performed 45 days after the last dose of standard of care (SOC) treatment.  Post 
progression de novo biopsy specimens must be shipped no later than 2 weeks after collection.  Please refer to Section 5.2.1.2 and the 
laboratory manual for detailed instructions. 
8.
Research blood collection: Research blood collection may be performed at a separate visit (which may be a home visit, if appropriate) 
to the de novo tumor tissue biopsy but must occur ≤5 days before or ≤1 day (24 hours) after the de novo biopsy. Please refer to the 
laboratory manual for detailed instructions.  See Section 8 for details.  Research blood specimens must be shipped no later than one 
week after collection.  Please refer to the laboratory manual for detailed collection, processing, storage, and shipment instructions. 
9.
Pregnancy Test: Specific only to Austria. Monthly pregnancy tests per institutional guidelines. 
10. Adverse event monitoring: Adverse event collection will begin at the time of the de novo tumor biopsy and research blood collection, 
whichever occurs first, through and including 24 hours post-biospecimen collection. 
11. NGS results consultation: NGS consultation will occur within 30 days of receipt of the de novo tumor biopsy or cfDNA NGS results at 
the provider’s facility.  These visits should preferably be conducted in person, although phone visits are permitted if the patient is unable 
or unwilling to travel to the facility. 

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14. Next anti-cancer therapy decision: Next anti-cancer medication following de novo tumor biopsy or cfDNA sample collection and 
whether impacted by the NGS results will be recorded in the appropriate CRF page.
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1. INTRODUCTION AND RATIONALE
The overall objective of this study is to better understand why tumors become resistant to 
treatment with standard of care (SOC) therapies that target critical aspects of tumor biology.  
Limited availability of post-progression tumor tissue has compromised efforts to rigorously 
characterize the tumor genomic and microenvironmental factors that drive resistance.  
Understanding mechanisms of resistance is critical to developing better anti-cancer 
treatments and treatment combinations.  This study specifically addresses the key challenges 
associated with studying clinical mechanisms of resistance.
Intrinsic or baseline tumor genetic heterogeneity, reflecting varying degrees of both intra-and 
inter-patient biologic diversity, represents a defining characteristic of cancer biology.1  In 
addition, the host response to cancer further defines the tumor microenvironment within 
which each patient’s cancer is subject to further selection, establishing distinct immune 
landscapes that reflect the complex tumor-immune cell interactions.1,2  The introduction of 
targeted therapeutic agents that potently modulate tumor and/or immune cell biology impose 
additional, very specific, selection pressures that further define the evolving biology of each 
patient’s cancer, frequently resulting in resistance following an initially beneficial treatment.  
Given the underlying genetic heterogeneity and the diversity of potential immune landscapes 
in cancer, informative analysis of mechanisms of resistance requires the application of robust 
platform technologies supporting immuno-genomic profiling of post-progression tumor 
tissue applied uniformly across a large series of patients.
1.1. Clinical Mechanisms of Resistance
Understanding clinical mechanisms of resistance has proven challenging because obtaining 
the required clinical tumor samples once resistance is established is difficult and the rationale 
for patients to undergo a de novo tumor biopsy after progression has been limited.  As a 
result, studies of clinical mechanisms of resistance often derive at best from very limited 
number of studies or case reports, each describing results from analysis of small numbers of 
patients (see Section 1.1.1 - Section 1.1.4).  Furthermore, these independent studies do not 
employ common or broad molecular profiling technologies, further limiting the utility of the 
information.  This information gap compromises efforts to develop therapeutic agents or 
combinations of therapeutic agents that might overcome these mechanisms of resistance to 
initially effective anti-cancer therapies.  Despite these limitations initial progress has been 
made in identification of clinical mechanisms of resistance to different therapeutic agents, 
and this has been complemented by studies of resistance in preclinical models, as 
summarized below.  The development of high quality clinical Next Generation Sequencing 
(NGS) tests, together with the availability of a broad spectrum of targeted therapeutic agents 
that are approved or in clinical development, provides additional rationale to pursue a 
de novo tumor biopsy for patients in whom a biopsy procedure can be safely performed 
(see Section 1.2).  Broad and deep genomic characterization by NGS in larger cohorts 
increases the probability of identifying clinically relevant resistance mechanisms.

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1.1.1. Immune Checkpoint (PD-1/PD-L1) Inhibition
Agents targeting programmed cell death-receptor 1(PD)-1, or programmed cell death-ligand 
1 (PD-L1) [PD-1/-L1], have demonstrated significant clinical benefit across a wide range of 
different tumor types.  However, in the majority of settings, responses are limited to a sub-set 
of patients, and, while responses are highly durable, progression following prolonged 
response does occur.  These phenomena speak to mechanisms of both intrinsic (also referred 
to as primary or inherent) and acquired (secondary) resistance, which if better understood 
may allow for more effective treatments. 
Studies conducted over the last several years have revealed a complex series of tumor cell 
intrinsic and extrinsic factors that contribute to the response and resistance to anti-PD-1/-L1 
targeting therapies. 
1.1.1.1. The Antigenicity of the Tumor 
The process of mutation, which is fundamental to the emergence and progression of cancer, 
also has the potential to generate unique protein sequences that are recognizable by T cells.  
The frequency of these sequences, called neoepitopes, is proportional to the overall 
frequency of mutations within a tumor, termed the tumor mutational burden (TMB).  Both 
the number of neoepitopes, and the overall TMB of tumors are emerging as critical 
determinants of response to anti-PD-1/-L1 therapy in several settings.51,52,53  Further, a recent 
study of non-small cell lung cancer (NSCLC) patients indicated that, while changes in the 
absolute TMB or number of neoepitopes did not correlate with acquired resistance to 
anti-PD-1, loss of neoepitopes shown to be actively engaging T-cell responses at the time of 
response was correlated with acquired resistance.54  This suggests that an evolution of the 
tumor neoepitope landscape and the overall antigenicity of a tumor, in response to treatment 
with anti-PD-1/-L1, may also represent a mechanism of resistance. 
1.1.1.2. Tumor Cell Intrinsic Immunogenicity 
A number of cell intrinsic factors influence how effectively a tumor cell is detected and 
eliminated by the immune system.  These factors include the surface expression of immune 
stimulatory or inhibitory signaling proteins, such as PD-L1, the ability to process and present 
antigens via the major histocompatibility complex (MHC), also termed human leukocyte 
antigen (HLA), and the ability to respond to inflammatory signals.  Through their impact on 
one or more of these factors, mutations in specific genes and pathways can impact tumor cell 
intrinsic immunogenicity, and response to anti-PD-1/-L1. 
Recent work has suggested that loss of heterozygosity (LOH) in the HLA genes can be a 
mechanism of immune escape during the development and evolution of a tumor.55  In 
keeping with this finding, loss of HLA, or other components of the antigen processing and 
presentation machinery, and presence of specific HLA haplotypes with reduced affinity for 
the T-cell receptor (TCR) have also been associated with resistance to anti-PD-1/-L1 
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The up-regulation of many surface proteins critical to the effective recognition of tumor cells 
by the immune system, including MHC, is facilitated by the interferon-(IFN-) response, 
and mutations in genes that are critical to this response, such as janus kinase (JAK) 1 and 
2 and the IFN-receptor, have also been associated with resistance to treatment with 
anti-PD-1/-L1 and anti-cytotoxic T-lymphocyte-associated protein (CTLA)-4.58,59,60  
Preclinically, studies have indicated that epigenetic, as well as mutational, changes can 
impact the production of key cytokines and chemokines, negatively impacting tumor cell 
immunogenicity61 and as such could represent an alternative route to the development of 
resistance. 
Mutations in key oncogenes, as well as mutations in genes associated with immune function, 
have also been shown to impact tumor immunogenicity and response to anti-PD-1/-L1 
treatment.  For example, mutations in the epithelial growth factor receptor (EGFR) gene in 
NSCLC have been associated with reduced immunogenicity and impaired response to 
anti-PD-1/-L1 treatment.62  Increased -catenin signaling has been associated with reduced 
immunogenicity in melanoma patients, and shown to reduce response to anti-PD-L1 in 
preclinical models.63  Similarly loss of phosphatase and tensin homolog (PTEN) in 
melanoma patients has been associated with reduced T-cell infiltration and reduced tumor 
shrinkage in response to anti-PD-1.  In contrast, mutations in BReast CAncer susceptibility 
gene (BRCA) 2 have been shown to be enriched among melanoma patients responding to 
anti-PD-1.64
1.1.1.3. Tumor Cell Extrinsic Immunogenicity 
The signals and cells present in the microenvironment surrounding the tumor cells represent 
a third, and likely major, driver of response, and resistance to anti-PD-1/-L1 therapy. 
The quantity and quality of the T cells responding to a tumor is likely one important factor 
related to both response and resistance to anti-PD-1/-L1 treatment.  In keeping with this 
hypothesis, the overall shape of the T cell response, as measured by TCR repertoire 
sequencing, has been shown to relate to response to anti-PD-1/-L1 therapy in some 
settings65,66 and changes in the presence of T cells specific for certain neoepitopes has been 
associated with acquired resistance.54  In both NSCLC67 and squamous cell carcinoma of the 
head and neck (SCCHN)68 phenotyping of tumor infiltrating T cells has indicated that the 
expression of an increasing number of inhibitory cell surface receptors, such as PD-1, T-cell 
immunoglobulin and mucin-domain containing-3 (TIM-3) and lymphocyte activation gene-3 
(LAG-3), is associated with a reduced responsiveness to ex vivo antigen stimulation, and in 
keeping with these findings, preclinical studies in mice have indicated that increased levels of 
PD-1 high T cells are associated with reduced response to anti-PD-1 treatment.69
Immune cells other than T cells have also been shown to impact the response to 
anti-PD-1/-L1.  Most notably an increased level of immunosuppressive myeloid derived 
suppressor cells (MDSCs) has been shown to correlate to reduced likelihood of response to 
treatment.70,71  In addition, broader assessment of transcriptional signatures associated with 
resistance to anti-PD-1 treatment in melanoma patients has identified an innate anti-PD-1 
resistance (IPRES) signature, characterized by genes associated with mesenchymal 
transition, angiogenesis, hypoxia and wound healing.64  A similar transcriptomic study of 

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multiple, differentially responsive lesions within a single patient also identified a signature 
enriched for genes associated with mesenchymal transition.72
Finally, it is worth noting that in addition to the factors described above, the treatment 
regimen received prior to anti-PD-1 treatment can significantly alter the immunogenicity of a 
tumor.  This is highlighted in recent studies of melanoma patients, in which tumors 
demonstrating acquired resistance to mitogen-activated protein kinase (MAPK)/BRAF 
directed therapies also demonstrated a decrease in T cell infiltration, an increase in T-cell 
exhaustion and a potential shift toward a suppressive, M2 macrophage driven inflammatory 
microenvironment.73  In keeping with these findings a separate study indicated that patient 
outcomes following treatment with the anti-CTLA-4 ipilimumab were poorer in patients who 
had initially progressed on previous BRAF targeted therapies prior to ipilimumab, as 
compared to those who had not received these therapies.74
1.1.2. Cell Cycle and CDK4/6 Inhibition 
The treatment of women with hormone receptor (HR)-positive, human epidermal growth 
factor receptor 2 (HER2)-negative advanced or metastatic breast cancer has been 
significantly improved by combining a cell cycle inhibitor (eg, palbociclib, abemaciclib, 
ribociclib) that blocks the function of both cyclin-dependent kinase (CDK) 4 and the 
redundant CDK6 with an aromatase inhibitor (AI) or fulvestrant.3  CDK4 or CDK6 
complexed with cyclin D1 (CCND1) phosphorylates the retinoblastoma gene product (Rb), 
releasing the E2F and DP transcription factors that regulate the expression of genes required 
for entry into the S phase of the cell cycle.  CDK4/6 kinase activity and progression through 
the G1 phase are negatively regulated by CDK4/6 interacting protein-kinase inhibitory 
protein (Cip-Kip) and by the inhibitor of the cyclin-dependent kinase (INK) 4 families, 
typified by cyclin-dependent kinase inhibitor 2A (CDKN2A) (gene product p16).  The 
frequent deletion or mutation of CDKN2A in tumor cells suggests that p16 acts as a tumor 
suppressor.  CCND1 is a direct transcriptional target of the estrogen receptor (ER)4-7 and 
antiestrogen-induced growth arrest of ER-positive breast cancer cells is accompanied by 
decreased CCND1 expression,8 while endocrine resistance is associated with persistent 
CCND1 expression and Rb phosphorylation.9  It has been shown that inhibition of CDK4/6 
prevents deoxyribonucleic acid (DNA) replication by arresting progression from G1 to 
S phase during cell division.  Thus, interruption of this mechanism should prevent tumor cell 
proliferation through control of the cell cycle.
1.1.2.1. Preclinical Mechanisms of Resistance
Despite a detailed understanding of the mechanisms underlying cell cycle regulations, based 
in large part on data from preclinical models, our understanding of the clinical mechanisms 
of resistance to CDK4/6 inhibitors remains limited.  Cancer cell line models have often 
proven to be limited in their ability to elucidate mechanisms of cell cycle regulation in 
patients.
Preclinical studies have indicated that increased expression of CCND1, Rb phosphorylation 
and decreased expression of p16 are associated with response to palbociclib.  However, 
results from biomarker analyses in clinical trials of palbociclib (PALOMA-1, -2, and -3), 
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CDKN2A and immunohistochemistry (IHC) for CCND1, p16, and Rb, showed that patients 
benefitted from palbociclib plus endocrine therapy regardless of differences in the expression 
of these biomarkers.10,11
Additional preclinical studies indicated that in a panel of breast cancer cell lines intact Rb 
signaling is needed for palbociclib to induce its cytostatic effect, and in ex vivo cultures of 
human breast tumors Rb loss was associated with resistance to palbociclib.12  However, 
although retinoblastoma gene (RB1) mutations are detected in tumors from patients treated 
with CDK4/6 inhibitors, they are rare and therefore unlikely to represent a dominant response 
determinant for CDK4/6 inhibition.13
Further, preclinical studies showed that sensitivity to CDK4/6 blockade could be mediated by 
abundant expression of both cyclin D3 and CDK6.14  However, analysis of Cyclin D3 
(CCND3) and CDK6 gene expression in clinical studies of palbociclib (PALOMA-2, -3) 
demonstrated the efficacy of palbociclib in combination with endocrine therapy did not 
correlate with expression of these genes.11,15
In addition to intrinsic and acquired resistance, rapid adaptation to resist drug pressure, 
typically by mechanisms other than genetic alterations, has been observed in various 
preclinical systems treated with different therapeutic agents.  Preclinical studies assessing 
mechanisms of early adaptation and secondary resistance of breast cancer cells to palbociclib 
treatment highlighted a potential role for either phosphoinositide 3-kinase (PI3K) signaling 
or CDK2 signaling, which fueled cell-cycle progression despite successful CDK4/6 
inhibition.16  Also, potential secondary resistance was found to be mediated by acquired 
cyclin E1 gene (CCNE1) amplification.  Cyclin E1 functions as an obligate partner 
complexed with CDK2 to activate its kinase activity which appears to be able to compensate 
for CDK4/6 inhibition in certain contexts. Acquired CDK6 amplification promotes breast 
cancer resistance to CDK4/6 inhibitors and loss of ER signaling and dependence.  Long-term 
exposure of ER-positive breast cancer cells to CDK4/6 blocking agents resulted in the 
emergence of acquired resistance after an initial period of cell-cycle arrest; this was mediated 
by the acquired CDK6 gene amplification and the subsequent overexpression, resulting in 
therapeutic resistance to abemaciclib treatment.  Interestingly, the acquired CDK6 
amplification resulted also in reduced ER/progesterone receptor (PgR) expression, and thus 
reduced sensitivity to endocrine treatment.17  These results indicated that CDK6 
amplification/overexpression could mediate secondary resistance in patients with hormone 
receptor-positive, HER2-negative breast cancer receiving combined ER-CDK4/6 blockade.18
The recent review by Klein et al79 suggested that tumor cells upon CDK4/6 inhibition could 
lead to either quiescence or senescence.  Such changing in state of cells may depend on the 
down regulation of MDM2, redistribution of ATRX, and repression of HRAS.  In addition, 
although changes in metabolism after CDK4/6 inhibition are likely to be context specific, 
cellular metabolism could also lead to non-Rb targets of CDK4/6 in the metabolic machinery 
including AMPKa2 and GCN5 and/or Rb-dependent manner in the altering glycolytic and 
oxidative metabolism.  Moreover, in preclinical models, both abemaciclib and palbociclib 
could induce growth arrest and upregulate antigen processing and presentation in tumor cells, 
changing their tumor microenvironment. 

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1.1.2.2. Clinical Mechanisms of Resistance
An increasing body of preclinical and early clinical evidence suggests that cyclin E1 
expression is a marker of resistance to CDK4/6 inhibition.  High cyclin E1 expression 
correlates with resistance to CDK4/6 inhibitor treatment in cell line models of breast cancer 
and ovarian cancer.  In triple negative breast cancer cell lines resistant to CDK4/6 inhibition, 
cyclin E1 expression remains high directly after mitosis, bypassing the restriction point at 
which CDK4/6 has traditionally been viewed as being required for G1 transition.  
Upregulation of cyclin E1 expression promoted sufficient CDK2 activity to initiate 
retinoblastoma phosphorylation thereby bypassing CDK4/6 inhibition.  Limited biomarker 
work from pre-operative palbociclib clinical trials supports cyclin E1 as a biomarker 
identifying ER-positive, HER2-negative breast cancers that are resistant to CDK4/6 
inhibition.  In the NeoPalAna neo-adjuvant study, breast cancers that were intrinsically 
resistant to CDK4/6 had high levels of CCNE1.19  The baseline gene expression patterns of 
CDK4/6 inhibitor treatment in naïve or refractory HR+, HER2- advanced breast cancer in the 
phase Ib study of ribociclib plus everolimus plus exemestane showed that higher levels of 
CCNE1 expression were observed in patients with progressive disease as compared to those 
with stable disease.20  Further, gene expression analysis of baseline breast cancer tissues in 
the palbociclib PALOMA-3 trial showed that high expression of CCNE1, along with high 
expression of pathways of E2F activation, predicted for lower benefit from palbociclib.15
Cancers with high CCNE1 expression were more likely to progress early on treatment.  In 
contrast, there is no evidence that either ER expression or luminal subtype predicts for 
benefit from palbociclib.  These data suggest that the phenotype observed in PALOMA-3 
may be present in primary breast cancer, although progression on prior endocrine therapy 
may conceivably effect CCNE1 expression.  Future research will be required to assess to 
what extent the acquisition of resistance to endocrine therapy could amplify cyclin E1 
expression, and also identify the cellular processes that allow cyclin E1 expression to become 
de-coupled from the requirement for prior CDK4/6 activation.21  
Using driver mutation targeted sequencing, a longitudinal circulating tumor DNA analysis in 
193 pairs of baseline and end of treatment (EOT) plasma samples of PALOMA-3 was 
conducted.  The results show that breast cancer driver mutation landscapes after treatment 
with either palbociclib + fulvestrant or with fulvestrant alone are largely similar, with 
acquired PIK3CA and estrogen receptor 1 (ESR1) Y537S mutations likely contributing to 
fulvestrant resistance.  Acquired RB1 mutations are selected, albeit very infrequently, by 
palbociclib + fulvestrant.22  These findings demonstrate the critical need to understand the 
genomic profile of each patient’s tumor both before and after treatment in order to decipher 
potential resistance mechanisms.
1.1.3. Androgen Receptor 
Second generation androgen receptor (AR) antagonists (eg, enzalutamide, abiraterone) have 
demonstrated significant clinical activity in metastatic castration resistant prostate cancer 
(CRPC), both in the first and second line settings.  While responses are observed in the 
majority of patients treated not all patients respond and many patients eventually relapse.  
Thus, there is a clear need to identify biomarkers that distinguish responders from 
non-responders and to identify mechanisms driving acquired resistance to therapeutic 
inhibition of the androgen receptor. 

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Resistance to androgen receptor antagonists, such as enzalutamide (Xtandi), remains poorly 
understood.  However, a number of studies conducted in recent years have begun to elucidate 
several potential mechanisms of clinical resistance. 
1.1.3.1. AR Point Mutations
There is strong evidence that point mutations within the AR increase in frequency in late 
stage CRPC patients from <1% to 10-20%.23,24  Moreover, these mutations are also 
associated with resistance to second generation AR antagonists.  For example, F876L results 
in altered enzalutamide binding such that enzalutamide now functions as a partial AR agonist 
and increases AR activity.25  This mutation was found in a patient that no longer responded 
to apalutamide25 or to enzalutamide.26  Additional mutations in AR have been detected in 
patients progressing on enzalutamide, including L702H, H875Y, and T878A.27  Importantly, 
~8% of patients progressing on enzalutamide harbor multiple AR mutations,27 emphasizing 
the difficulty of targeting a specific mutation as a way to combat resistance.  While AR 
mutations driving resistance to enzalutamide may also be associated with resistance to 
abiraterone (Zytiga), there are some notable differences which may be attributed to their 
different modes of action.  For example, T878A and L702H are found in abiraterone 
progressing patients.26,28,81  These mutations are particularly revealing, as they confer 
progesterone and, to a lesser extent, estradiol responsiveness to AR suggesting that 
alternative hormones may be able to bind and activate AR in place of androgens.29,30  
Importantly, CYP17A1 inhibition not only reduces AR activity by decreasing testosterone 
levels but it also increase progesterone levels, suggesting that these mutations may become 
less sensitive to CYP17A1 inhibitors, possibly suggesting that enzalutamide may benefit 
patients harboring a T878A or L702H mutation.82
1.1.3.2. AR Amplification 
A second emerging mechanism also centers on AR and involves amplification or copy 
number gain of the gene encoding AR.26,31  The prevalence of AR amplification has been 
observed to increase in a number of studies comparing pre-treated samples with 
tumors/plasma of patients that have progressed on enzalutamide.  Interestingly, amplification 
of AR has been noted to occur to a greater extent in response to enzalutamide compared to 
abiraterone in two independent studies.  In one study amplification of AR was more 
prevalent in patients progressing on enzalutamide (53%) compared to patients progressing on 
abiraterone (17%).26  In a second study, the amplification frequency of AR was 15% at 
baseline but increased to 30% in patients progressing on enzalutamide; in contrast, no 
increase was found in patients progressing on abiraterone.27  Due to the limited number of 
patients and the number of independent studies examined, it is difficult to conclude if AR 
amplification status represents a true mechanistic difference in resistance to these agents. 
1.1.3.3. AR Splice Variants 
Another well described mechanism driving resistance is increased expression of splice 
variants of the AR, (AR-V’s).  The most well described of these is AR-V7, which lacks the 
ligand binding domain of the AR and is constitutively active in the absence of androgens.  It 
should be noted that there are as many as 18 different splice variants of AR32 with AR-V7 
representing the most extensively studied as there are significantly better quality reagents to 

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detect this variant; thus, the true role of the other variants in driving resistance to AR 
antagonists remains undetermined.  The presence of AR-V7 transcripts in circulating tumor 
cells (CTCs) has been associated with poor survival33 and with reduced response to both 
enzalutamide and abiraterone.34  However, there is no consensus as to whether these variants 
are drivers of resistance, or are merely passengers that either mask the true mechanism or act 
in concert with a second driver. 
1.1.3.4. Trans-differentiation as a Mechanism of AR Resistance 
Emerging data now suggests that tumors can evade AR modulators by trans-differentiation 
into a neuroendocrine like phenotype that is characterized by mutations that include loss of 
RB, p53, or PTEN, and an increase in copy number of N-MYC genes.35,36  These 
neuroendocrine prostate cancer (NEPC) tumors are thought to transdifferentiate from a 
CRPC luminal adenocarcinoma cell by divergent clonal evolution.35,36  Additionally, NEPCs 
are often characterized by loss of AR expression and are hormone refractory.  This is an 
especially aggressive form of prostate cancer with a survival of approximately 1 year.  In a 
recent report of 114 metastatic biopsies, tumors progressing on enzalutamide or abiraterone 
were found to have neuroendocrine like features, including lower AR expression levels and 
loss of RB and p53.35  Epigenetic reprogramming involving the histone methyltransferase 
enhancer of zeste homolog 2 (EZH2) has also been implicated in driving neuroendocrine 
prostate cancer,37 suggesting that novel agents such as EZH2 inhibitors may have utility in 
this setting. 
1.1.3.5. Perturbation of Cell Cycle 
Emerging data suggests that mutations that increase cell cycle progression from the G1- to 
the S-phase of the cell cycle are implicated in resistance to enzalutamide.  Patients 
progressing on enzalutamide have mutations in a number of genes involved in regulating this 
transition, including amplification of CCND1 and CCNE1,26,27 loss of RB,26 amplification of 
CDK6 and an inactivating mutation in the tumor suppressor CDKN2A.38  All of these 
chromosomal aberrations converge on proteins involved in regulating the ability of cells to 
initiate DNA replication, suggesting that drugs such as CDK4/6 inhibitors could have utility 
in the resistance setting. 
In addition to these clinically identified resistance mechanisms, a number of other potential 
resistance pathways have been proposed based on preclinical models, including loss of AR, 
ERG gene rearrangements, PTEN deficiency and stabilization of the AR.39  Further, it is 
plausible that resistance to androgen receptor antagonists is driven by multiple genomic 
aberrations.  For example, in a recent study 8% of patients progressing on enzalutamide were 
found to harbor multiple mutations within AR.  It’s not known if these mutations coexist in 
the same lesion or if different metastatic sites harbor different AR mutations.  Additionally, 
patients can display either inherent or acquired resistance to enzalutamide that does not have 
chromosomal aberrations in any of the genes mentioned above or others known to be 
involved in AR signaling in prostate cancer.  Thus, there are other mechanisms that remain 
unidentified. 

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1.1.4. DNA Damage Response and PARP Inhibition
Poly (ADP-ribose) polymerase (PARP) inhibitors exert cytotoxic effects by two mechanisms: 
inhibition of PARP catalytic activity and PARP trapping.  Inhibition of PARP catalytic 
activity results in persistent single-strand DNA breaks that culminate in double-strand DNA 
breaks, creating a dependency on homologous recombination-mediated DNA damage repair 
(HR DDR) for cell survival.  PARP trapping prevents a PARP inhibitor-bound PARP protein 
complex from readily dissociating from DNA, thereby inhibiting DNA repair, replication, 
and transcription, resulting in double-strand DNA breaks and cytotoxicity.40,41  PARP
inhibitors induce synthetic lethality in tumor cells bearing mutations and/or deletions in 
genes involved in homologous recombination or other DNA repair pathways, most notably 
BRCA1 and BRCA2.42
PARP inhibitors are highly active in patients bearing tumors with HR DDR deficiency.  
However there is evidence for both intrinsic and acquired resistance, with some patients not 
responding and others exhibiting transient responses with subsequent disease progression.42  
There has been substantial progress in recent years in elucidating the mechanisms underlying 
clinical resistance to PARP inhibition.  However, as described above and shared by 
essentially all analyses of clinical mechanisms of resistance, results derive from very small 
series of patients, typically do not employ uniform or broad molecular profiling technologies, 
and may be of relevance to only a fraction of patients. 
The primary clinically identified mechanism of resistance to PARP inhibition is acquired 
reversion mutations in BRCA1 and BRCA2.  Moreover, in non-BRCA mutated tumors there
is anecdotal but compelling evidence that acquired resistance to PARP inhibitors in the clinic 
is consistently associated with reversion mutations in other HR DDR genes, including 
Partner and localizer of BRCA2 (PALB2), RAD51C and RAD51D.  For example, in
ARIEL2 Part 1, a Phase 2 study of rucaparib in platinum-sensitive, relapsed ovarian cancer, 
core HR pathway genes in 12 pairs of pre-treatment and post-progression tumor biopsy 
samples were sequenced.43  In 6 of 12 pre-treatment biopsies, a truncation mutation in 
BRCA1 (n=4), RAD51C (n=1) or RAD51D (n=1) was identified.  In five of six paired 
post-progression biopsies from these 6 patients, one or more secondary mutations restored 
the open reading frame. 
In another analysis, BRCA1 or BRCA2 reversion mutations were identified in circulating 
free DNA (cfDNA) from 4 patients with ovarian cancer (21%) and from two patients with 
breast cancer (40%) who were resistant or refractory to platinum-based chemotherapy or 
PARP inhibitors (PARPi).44  Serial analysis of plasma from one breast cancer patient 
demonstrated the presence of such BRCA2 reversion mutations post carboplatin and prior to 
talazoparib, consistent with BRCA reversion mutations also being an established mechanism 
of resistance to platinum-based therapy.77  This patient did not respond to talazoparib and 
displayed a greater diversity in BRCA2 reversion mutations post talazoparib.44
In an analysis of patients with metastatic prostate cancer,45 10 of 16 patients with initial 
response to the PARPi olaparib had cfDNA samples acquired at the time of resistance and 
disease progression.  In 2 of 2 patients with a germline BRCA2 frameshift mutation, at the 
time of tumor progression additional somatic BRCA2 mutations restoring the open reading 
frame (ORF) were detected in cfDNA.  In a patient with a somatic BRCA2 mutation, 

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reversion mutations restoring the ORF were detected at progression. Moreover, in a patient 
with a somatic PALB2 mutation, reversion mutations restoring the ORF were also detected. 
These and similar analyses confirm that such reversion mutations as a mechanism of 
acquired resistance are not confined to BRCA1 or BRCA2, but also evident in genes 
encoding other proteins involved in HR DDR.  Moreover, such reversion mutations can 
occur regardless of whether the initial mutations were germline or somatic in origin. 
Multiple additional potential mechanisms of intrinsic and/or acquired resistance to PARPi 
have been identified based on work in preclinical models.  These include: 
Loss of DNA repair proteins 53BP1 or REV7 which results in the restoration of 
HR-mediated DNA repair.46,47
Loss of a number of proteins, including PARP1 itself, which are involved in 
maintaining replication fork stability.48
Loss of Schlafen 11 (SLFN11) in small cell lung cancer cell lines grown in culture 
and in vivo.  SLFN11 expression was demonstrated to predict responsiveness to 
talazoparib, while loss of SLFN11 via Clustered Regularly Interspaced Short 
Palindromic Repeats (CRISPR)/Cas9 or short hairpin ribonucleic acid (RNA) 
conferred resistance to PARPi.49
Expression of BRCA1 variants.  Some BRCA1 truncation mutations result in 
expression of a RING-less BRCA1 isoform that is associated with resistance to 
PARPi in cells and in vivo models.50
Loss of AT-rich interactive domain-containing protein 1A (ARID1A) or GC-Rich 
Promoter Binding Protein 1 (GPBP1), based on a quantitative chemotherapy genetic 
interaction map and limited clinical correlative analyses.78
Mutations in PARP1.  A combination of genome-wide and high density 
CRISPR-Cas9 screens identified point mutations in PARP1 which were associated 
with resistance to PARPi in nonclinical models.  These mutations impacted DNA 
binding and/or PARP trapping on DNA.  Importantly, a similar mutation was 
observed in a patient with de novo resistance to olaparib. 
It should be noted that these additional potential mechanisms of resistance have generally not 
been identified clinically and if identified, prevalence would need to be defined to better 
understand clinical relevance.
In summary, various potential mechanisms of resistance to therapeutics that are the subject of 
the current study have emerged from preclinical and clinical assessment in previous efforts, 
with limitations noted on patient cohort sizes, technology platforms utilized, and preclinical 
observations only.  The current study seeks to confirm and expand upon this early body of 
work.  All previously identified resistance mechanisms summarized above will be captured 
by the methodologies applied in this study, and with a larger number of patients pairing 

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pretreatment and post-progression biopsies for analysis.  This will afford clinical and 
independent confirmation and improved frequency assessment of these previously identified 
resistance mechanisms.  In addition, broader reaching approaches (eg, whole exome 
sequencing, whole transcriptome) will be employed to identify new/unknown potential 
resistance mechanisms. 
1.2. Potential Risks and Possible Benefits 
The primary potential risk to patients who participate in the study is associated with the 
de novo tumor biopsy performed as a study-related procedure.  To be eligible for the study 
patients must have a tumor lesion that is considered to be safely accessible to a de novo
biopsy performed in accord with local institutional practice standards.  See Section 4 for 
details. 
Possible benefits for patients who participate in the study derive from the NGS results from 
the analysis of their tumor tissue and/or blood performed by a third-party laboratory.  These 
results are returned to the patient’s physician/ health care provider who will then discuss the 
results with the patient.  To enable sufficient time to thoroughly discuss the results and 
possible treatment implications, the study also supports the direct interaction between the 
patient and the physician by incorporating dedicated consultation time (See Schedule of 
Activities).  Further, patients will be able to obtain a personal copy of the NGS results 
through their health care provider.  It is recognized that results from NGS analysis of the 
patient’s tumor tissue and/or blood may not directly impact the subsequent treatment plan.  
However, only by performing such analyses can low probability treatment opportunities be 
identified.  
2. STUDY OBJECTIVES AND ENDPOINTS
Primary Objective:
Primary Endpoint:

To obtain and analyze archival pre-treatment 
tumor samples and post-progression tumor 
biopsies to identify molecular markers of 
resistance to selected anti-cancer therapies.

Change in the frequency of gene alterations 
between pre-treatment tumor samples and 
post-progression tumor biopsies.
Secondary Objectives:
Secondary Endpoints:

To evaluate the success rate in obtaining 
paired archival and post-progression tumor 
biopsies that are adequate to meet the 
objectives of the study; 

To determine concordance in gene alterations 
identified by NGS analysis of post-progression 
tumor tissue and blood.

Proportion of patients with fully evaluable 
archival and post-progression tumor biopsy 
(eg, sample sufficient for all intended analyses 
at all measured time points); 

Overall agreement rates of gene alterations 
between post-progression biopsy tissue and 
blood NGS results.

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Secondary Objectives by Target Biology:
Secondary Endpoints by Target Biology:
PD-1/-L1 inhibition:
•
To evaluate alterations in genes encoding 
HLA, β2-Microglobulin, Signal transducer and 
activator of transcription 1(STAT1), JAK1, 
JAK2, IFN-γ, and interferon-gamma receptor
(IFNGR) as mechanisms of resistance to 
PD-1/PD-L1 checkpoint blockade.
• Change in the frequency of alterations in genes 
encoding HLA, β2-Microglobulin, STAT1, JAK1, 
JAK2, IFN- γ and IFN-γR between pre-treatment 
archival and post-progression samples;
• The frequency of alterations in genes encoding 
HLA, β2-Microglobulin, STAT1, JAK1, JAK2, 
IFN-γ and IFNGR in cfDNA.
CDK4/6 inhibition:
•
To evaluate alterations in the Rb gene as a 
mechanism of resistance to palbociclib.
• Change in the frequency of RB1 gene alterations 
between pre-treatment archival and 
post-progression samples.
• The frequency of RB1 gene alterations in cfDNA.
AR inhibition:
•
To evaluate AR gene alterations as 
mechanisms of resistance to enzalutamide or 
abiraterone;
•
To evaluate changes in expression of nuclear 
hormone receptor genes as a mechanism of 
resistance to enzalutamide or abiraterone. 
• Change in the frequency of AR gene alterations 
between pre-treatment archival and 
post-progression samples;
• The frequency of AR gene alterations in cfDNA;
• Changes in the expression of nuclear hormone 
receptors or related RNA signatures reflecting 
nuclear receptor pathway activity between 
pre-treatment archival and post-progression 
samples.
PARP inhibition:
•
To evaluate somatic reversion of germline 
BRCA (gBRCA) gene alterations as a 
mechanism of resistance to monotherapy 
PARP inhibition. 
• Change in the frequency of somatic reversion 
alterations in gBRCA mutant allele between 
pre-treatment archival and post-progression 
samples.
•
 
 
 
 
 
 
 
 
 
 
 
  
 
 
 
 
 
 
 
CCI
CCI
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3. STUDY DESIGN
This is a multi-national, multi-center study to collect and analyze archival pre-treatment 
tumor and post-progression biospecimens (de novo tumor biopsy and blood) from patients 
with advanced or metastatic cancer whose disease has progressed on selected SOC therapy.  
Approximately 500 evaluable patients with different tumor types will participate in the study.  
Evaluable patients are defined as the ones with both archival tumor tissue and de novo tumor 
biopsy that is sufficient to enable the intended sample analyses. 
Following Informed Consent, Screening assessments are performed to confirm availability 
and adequacy of archival tumor tissue and to confirm preliminary eligibility criteria.  
Once disease progression on SOC therapy has occurred, patients who remain eligible are 
enrolled in the study and undergo post-progression biospecimen collection (de novo tumor 
tissue biopsy and research blood draws) and other study-related activities. Molecular 
profiling, including NGS analysis, will be performed on the archival and post-progression 
biospecimens. 
As part of study participation, results from the NGS analysis of the de novo post-progression 
biopsy or research blood specimens will be returned to the patient’s physician/health care 
provider.  Results will come from tests performed in a College of American Pathologists 
(CAP) CAP-accredited, Clinical Laboratory Improvements Amendments (CLIA)-certified, 
and good clinical practice (GCP)-compliant clinical reference third-party laboratory using a 
commercially available NGS test.  
A study-specific follow-up consultation between the treating physician and patient will be 
conducted to discuss the NGS results and potential treatment options (see Schedule of 
Activities).  All treatment decisions, including potential referral to a clinical trial, will be per 
the treating physician as part of the patients’ clinical care and are not dictated or defined by 
the study.  See Figure 1. 
 
CCI

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Figure 1.
Study Schema
Table 1 describes the target areas of biology, disease cohorts, prior SOC therapy, indications 
and cohort sizes.  All cohorts will enroll concurrently.  Individual cohorts and cohort sizes 
may be adjusted based upon emerging data.  Cohort size refers to evaluable patients as 
defined above.
Table 1.
Targeted Biology and Disease Cohorts
CDK = Cyclin-dependent kinase; CTLA-4 = cytotoxic T-lymphocyte-associated protein 4; 
gBRCAm = germline mutated BRCA; HER2 = human epidermal growth factor receptor 2; NSCLC = non-small 
cell lung cancer; PARP = Poly (ADP-ribose) polymerase; PD-1/L1 = programmed cell death receptor 1 or 
programmed cell death ligand 1; RCC = renal cell carcinoma.
CCI

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4. PATIENT ELIGIBILITY CRITERIA
This study can fulfill its objectives only if appropriate patients are enrolled.  The following 
eligibility criteria are designed to select patients for whom participation in the study is 
considered appropriate.  All relevant medical and nonmedical conditions should be taken into 
consideration when deciding whether a particular patient is suitable for this protocol.
4.1. Inclusion Criteria
Patients must meet all of the following inclusion criteria to be eligible for enrollment in the 
study:
Histological diagnosis of locally advanced (primary or recurrent) or metastatic solid 
tumors treated as follows (See Table 1 for cohorts):
a.
NSCLC monotherapy (Cohort 1):

Disease progression (PD) on 1st-line monotherapy anti-PD-1/-L1.
b. NSCLC combination (Cohort 2):

PD on 1st-line anti-PD-1/-L1 plus standard doublet platinum-containing 
regimen; or

PD on 1st-line anti-PD-1/-L1 plus standard doublet platinum-containing 
regimen followed by continuation of single agent anti-PD-1/-L1).
c.
Renal cell carcinoma (RCC) with clear cell component (Cohort 3):

PD on 2nd-line monotherapy anti-PD-1/-L1; or 

PD on 1st-line combination of doublet anti-PD-1/-L1 with anti-CTLA-4; or

PD on 1st-line combination of avelumab with axitinib or pembrolizumab with 
axitinib.
d. HR+ HER2- adenocarcinoma of the breast (Cohort 4):

PD on 1st-line combination of doublet palbociclib with hormonal therapy. 
e.
Castrate-resistant adenocarcinoma of the prostate (Cohort 5):

PD on enzalutamide monotherapy.
f.
Castrate-resistant adenocarcinoma of the prostate (Cohort 6):

PD on abiraterone in combination with prednisone.

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g. germline mutated BRCA (gBRCAm), HER2- adenocarcinoma of the breast 
(Cohort 7):

PD on a PARP inhibitor monotherapy in patients previously treated with 
chemotherapy in the neoadjuvant, adjuvant, or metastatic setting. 
Radiographic evidence of PD, including the target lesion being subjected to biopsy 
for the study, on the most recent regimen that requires a change in anti-cancer 
treatment. 
Medically stable for a biopsy procedure as defined by the local institutional 
guidelines.
A tumor lesion for the de novo biopsy that meets the following criteria:
a.
Is safely accessible to a biopsy procedure (ie, core needle, excisional) performed 
in accordance with local institutional practice standards;
b. Is anticipated to yield an amount of tumor tissue sufficient to meet the objectives 
of the study (see Section 2); 
c.
Not previously irradiated; 
d. Does not require decalcification for subsequent processing. (ie, no bone lesions).
Availability and adequacy of an archival, formalin-fixed, paraffin embedded 
(FFPE) tumor tissue block confirmed during Screening as containing sufficient 
tumor tissue to allow for sectioning of up to 25 slides each containing tissue 
sections that are 5 microns thick with a minimum tissue cross-sectional area of 
approximately 5 mm2 and containing a minimum of approximately 20-40% tumor 
by ratio of tumor nuclei to benign nuclei.  Fewer slides may be required for FFPE 
tissue blocks containing tissue that is greater than 5 mm2 in cross-sectional area or 
greater than 40% tumor.  Where local or regional regulations prevent submission of 
the archival tumor tissue block, a designated number of unstained slides each 
containing the indicated minimum amount of tissue must be submitted (refer to the 
laboratory manual for the required number of slides based on cross sectional area of 
tissue available in the FFPE tissue block) (See Section 5.1.3).
Post-progression biospecimen collection can be performed within 45 days of 
obtaining informed consent for the study.
Post-progression biospecimen collection can be performed within 45 days of the 
last dose of the most recent anti-cancer regimen. 
Age 18 years at the time of informed consent.
Patients who are willing and able to comply with scheduled visits and study 
procedures.

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Evidence of a personally signed and dated informed consent document indicating 
that the patient has been informed of all pertinent aspects of the study. 
4.2. Exclusion Criteria
Patients with any of the following characteristics/conditions will not be included in the study:
Discontinuation of current or most recent anti-cancer therapy due to toxicity and not 
progressive disease.
Initiation of new anti-cancer therapy after PD prior to planned biopsy. 
Any medical condition that, in the investigator’s judgement, unacceptably increases 
risk associated with the tumor biopsy or blood sampling procedures (eg, evidence of 
inadequate wound healing, significant neutropenia or thrombocytopenia, recent 
history of clinically significant bleeding or tumor hemorrhage).  
Cohorts 1, 2 and 3 (NSCLC monotherapy, NSCLC combination, RCC with clear 
cell component) only: Treatment with an anti-PD-1/-L1 agent prior to current or 
most recent anti-PD-1/-L1 therapy.
Cohort 4 (HR+ HER2- breast cancer) only: Treatment with a CDK 4/6 inhibitor 
prior to current or most recent CDK 4/6 inhibitor.
Cohort 5 (castrate-resistant prostate cancer) only: Treatment with an agent that 
blocks adrenal androgen synthesis (eg, abiraterone acetate) and a second-generation 
AR antagonist other than enzalutamide (eg, apalutamide). 
Patients who are investigator site staff members directly involved in the conduct of 
the study and their family members, site staff members otherwise supervised by the 
investigator, or patients who are Pfizer employees, including their family members, 
directly involved in the conduct of the study. 
Other acute or chronic medical or psychiatric condition including recent (within the 
past year) or active suicidal ideation or behavior or laboratory abnormality that may 
increase the risk associated with study participation or may interfere with the 
interpretation of study results and, in the judgment of the Investigator, would make 
the patient inappropriate for entry into this study.
Austria only: Pregnant female patients; breastfeeding female patients. 

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5. STUDY PROCEDURES
5.1. Screening (Study Day -45 to Day 1)
5.1.1. Informed Consent
Informed Consent will be obtained prior to any study-related activities. Consent should be 
obtained as close as possible to the day of the biospecimen collection, optimally the day of, 
but within 45 days before the de novo biopsy or research blood collection (whichever is 
sooner).  
5.1.2. Preliminary Eligibility Assessment
Prior to progressing to archival tumor tissue assessment, a preliminary eligibility assessment 
should be made by confirming the following:

Demographics (Inclusion Criterion 8);

Histological diagnosis of locally advanced or metastatic solid tumors and most recent 
standard of care anti-cancer therapy (Inclusion Criterion 1);

Prior systemic anti-cancer therapy regimens or radiation therapy (Inclusion Criteria 1, 
4c);

New anti-cancer therapy which has not already been started (Exclusion Criterion 2);

Reasons for discontinuation of current or most recent anti-cancer therapy (Exclusion 
Criterion 1).
Patients who have started new anti-cancer therapy prior to performing the de novo tumor 
tissue biopsy or research blood collection are not eligible and should be discontinued.
Screen failures are defined as participants who consent to participate in the clinical study but 
do not subsequently undergo biospecimen collection (ie, de novo tumor biopsy and research 
blood collection are not performed).
Patients are permitted to participate in interventional treatment clinical trials at the same time 
as participating in Study A9001502, with prior Sponsor agreement.
5.1.3. Archival Tumor Tissue Assessment
Assessment of availability and adequacy of an archival, formalin-fixed, paraffin embedded 
(FFPE) tumor tissue block (or equivalent slides; Inclusion Criterion 5) from the most recent 
primary or metastatic tumor biopsy or resection should be performed.
The archival tumor tissue assessment involves inspection by local qualified personnel of the 
available archival FFPE tumor tissue block and an associated H&E-stained slide to confirm 
the following: 1) the presence of tissue that is at least 5 mm2 in cross-sectional area on 
inspection of the cut face of the FFPE block, and 2) the presence of tumor tissue containing 
at least 20-40% tumor by ratio of tumor nuclei to benign nuclei based on review of an 
H&E-stained slide previously generated from the associated FFPE tissue block. 

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The FFPE tissue block must contain sufficient tumor tissue to allow for sectioning of up to 
25 slides each containing tissue sections that are 5 microns thick with a minimum tissue 
cross-sectional area of approximately 5 mm2 and containing a minimum of approximately 
20-40% the sample by ratio of tumor nuclei to benign nuclei.  Fewer slides may be required 
for FFPE tissue blocks containing tissue that is greater than 5 mm2 in cross-sectional area or 
greater than 40% tumor. 
Questions regarding archival tumor tissue sample requirements should be emailed to
Archived tumor tissue from cytological sampling (eg, fine needle aspiration, pleural 
effusion, including FFPE cell pellet material), or from biopsies of bone metastasis that 
require decalcification are not adequate for trial participation and should not be 
submitted.
If an archival FFPE tissue block cannot be provided due to documented local or institutional 
regulations, a designated number of unstained slides each containing tissue sections that are 
5 microns thick must be submitted.  Freshly cut slides should be sectioned as close as 
possible to the post-progression biopsy and must be shipped no more than 21 calendar days 
after sectioning.  The number of slides required is based on the estimated cross-sectional area 
of the available tissue (refer to the laboratory manual for a guide defining the required 
number of slides that must be submitted). 
5.1.4. Confirmation of Eligibility
Prior to Enrollment, medical and oncological history should be reviewed to confirm if there 
have been any changes to systemic anti-cancer therapy regimens or radiation therapy or 
reasons for treatment discontinuation.
Radiographic evidence of PD for the lesion being targeted for the de novo tumor tissue 
biopsy must also be confirmed.
The patient should be discontinued and should not undergo further study-related 
activities (including de novo tumor tissue biopsy or research blood collection) if new 
anti-cancer therapy has already been started following disease progression. The 
participant should be recorded as a screen failure in the case report form (CRF).
5.1.5. Re-Screening
Re-screening is permitted on a case-by-case basis and only with prior agreement from the 
study Sponsor.
5.2. Enrollment (Study Day 1)
Following confirmation of eligibility, the patient may be enrolled in the study and undergo 
study-related procedures.
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5.2.1. Biospecimen Collection 
Biospecimen collection may occur on different days; however, de novo tumor biopsy or 
research blood collection must occur within 45 days of obtaining study Informed Consent.
Day 1 of the study is the date on which the de novo tumor biopsy or research blood collection 
occurs (whichever is sooner, if occurring on different dates). Research blood collection must 
occur ≤5 days before, or ≤1 day (24 hours) after, the de novo biopsy.
5.2.1.1. Archival Tumor Tissue Submission
Eligible archival FFPE tumor tissue (or equivalent) from the most recent primary or 
metastatic tumor biopsy or resection (see Section 5.1.3) should be submitted for analyses. 
Please refer to the laboratory manual for detailed collection, processing, storage, and 
shipment instructions. 
All submitted FFPE tissue blocks (or equivalent slides) will be returned as soon as possible 
after the required number of sections are obtained and analyzed.
5.2.1.2. De Novo Tumor Biopsy
Performing a de novo biopsy should be compatible with the overall patient treatment plan.  
Pre-biopsy clinical and laboratory assessments per institutional guidelines must be completed 
and reviewed prior to biopsy procedure.  It is expected that these pre-biopsy assessments 
include but are not limited to hematology and coagulation profiles.  Safe values for biopsy 
are per institutional standard operational procedures (SOP) and normal lab ranges. 
Anti-coagulation therapy needs to be held prior to the biopsy according to institutional 
guidelines.
De novo tumor biopsies (ie, core needle, excisional) should be conducted according to 
institutional guidelines.  For core biopsies, a minimum 18 gauge core needle and 4 separate 
cores are recommended to ensure, to the greatest extent possible, that the quality and amount 
of tumor tissue obtained is sufficient to enable the intended sample analyses.  Tumor tissue 
from cytologic sampling (eg, fine needle aspiration, pleural effusion, including FFPE cell 
pellet material) is not adequate and should not be submitted.
The de novo tumor biopsy should be processed locally by formalin fixation and paraffin 
embedding, following specific processing guidelines described in the laboratory manual.  
The resulting FFPE tissue block should be shipped in its entirety as part of the tumor tissue 
specimen collection kit (room temperature).  
Any remaining post-progression biopsy material that is not used for molecular analyses as 
part of the study will be returned to the site where the biopsy was conducted within 
approximately 6 months of collection. 
The post-progression de novo biopsy must be obtained within 45 days of the last dose of 
SOC treatment but prior to initiation of any subsequent anti-cancer therapy.

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Post-progression de novo biopsy specimens must be shipped as soon as possible after the 
appropriate processing but no later than 2 weeks after collection to facilitate timely return of 
Next Generation Sequencing analyses.  Please refer to the laboratory manual for detailed 
collection, processing, storage, and shipment instructions. 
There will be no banking of tumor biopsy samples. 
5.2.1.3. Research Blood Specimens 
Research blood specimens will be drawn into separate blood collection tubes as follows:
a.
Three 10-mL tubes of blood will be collected in specially provided tubes containing 
blood stabilizer (ie, Streck tubes, room temperature) for cfDNA analysis using a NGS 
panel approach. 
b. One 8.5-mL blood specimen will be collected in a specially provided tube containing 
blood stabilizer (eg, Paxgene blood DNA tube; Streck tube) for germline DNA 
sequence analysis. Results will be used as a control to compare with tumor DNA 
sequence results in order to identify tumor-specific (ie, somatic) gene alterations, 
and/or to investigate the potential contribution of specific gene alterations to 
mechanisms of resistance.  Remaining DNA material may also be used for 
translational analyses as indicated below (item c).
c.
One 8.5 mL blood specimen will be collected in a specially provided tube containing 
blood stabilizer (eg, Paxgene blood DNA tube; Streck tube) for additional 
translational analyses which may include, but may not be limited to, T and/or B cell 
receptor sequence analysis or epigenetic profiling by NGS.
Research blood collection may be performed at a separate visit (which may be a home visit, 
if appropriate) to the de novo tumor tissue biopsy but must occur ≤5 days before or ≤1 day 
(24 hours) after the de novo biopsy.
The post-progression research blood specimens must be obtained within 45 days of the last 
dose of SOC treatment and prior to initiation of any subsequent anti-cancer therapy. 
The research blood specimens must be shipped no later than one week after collection.  
Please refer to the laboratory manual for detailed collection, processing, storage, and 
shipment instructions. 
There will be no banking of blood samples. 
5.2.2. Additional Medical and Oncological History
Smoking history, prior surgery, duration and best overall response for each therapy should be 
recorded in the CRF.
5.2.3. Pregnancy Testing (Specific to Austria)
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5.3. Post-Biospecimen Collection Follow-Up (≤30 days after receipt of NGS results at 
the provider’s facility)
5.3.1. NGS Results Consultation 
NGS results will be reviewed at a post-biospecimen follow-up consultation visit, which 
should occur within 30 days of receipt of the de novo tumor biopsy or cfDNA NGS results at 
the provider’s facility.  These visits should be conducted in-person; however, phone visits are 
permitted if the patient is unable or unwilling to travel to the facility.
 
  
 
 
5.3.3. Next Anti-Cancer Therapy
The patient’s next anti-cancer therapy following de novo tumor biopsy or cfDNA sample 
collection, and whether it was impacted by the NGS results, will be recorded in the case 
report form (CRF). 
5.4. Patient Withdrawal
Patients may withdraw from the study at any time at their own request, or they may be 
withdrawn at the discretion of the Investigator or Sponsor for safety or behavioral reasons, or 
the inability of the patient to comply with the protocol-required schedule of study visits or 
procedures. 
Reasons for withdrawal from the study may include:
•
Study terminated by Sponsor;
•
Lost to follow-up;
•
Refused further follow-up;
•
Death.
If the patient withdraws from the study, and also withdraws consent for disclosure of future 
information, no further procedures, visits or assessments should be performed, and no 
additional data should be collected.  The sponsor may retain and continue to use any data
collected before such withdrawal of consent.
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Withdrawal of consent: 
Patients should notify the investigator in writing of the decision to withdraw consent from 
post-biospecimen follow-up, whenever possible.  The withdrawal of consent should be 
explained in detail in the medical records by the investigator, as to the reason and entered on 
the appropriate case report form (CRF) page. 
Lost to follow-up: 
All reasonable efforts must be made to locate patients to determine post-biospecimen 
follow-up.  Lost to follow-up is defined by the inability to reach the patient after a minimum 
of 2 documented phone calls, faxes, or e-mails as well as lack of response by the patient to 
1 registered mail letter.  All attempts should be documented in the patient’s medical records. 
6. ASSESSMENTS
Every effort should be made to ensure that the protocol-required procedures, visits and 
assessments are completed as described.  However, it is anticipated that from time to time 
there may be circumstances, outside of the control of the Investigator that may make it 
unfeasible to perform the procedure(s), visits or assessments within the designated times.  In 
these cases, the Investigator will take all steps necessary to ensure the safety and well-being 
of the patient.  The study team will be informed of these incidents in a timely fashion.
For samples being collected and shipped, detailed collection, processing, storage, and 
shipment instructions and contact information will be provided to the investigator site prior 
to initiation of the study in the study laboratory manual.
Patients should follow the approved package insert for any ongoing medications. 
 
 
 
  
 
 
 
 
  
 
 
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6.3. Pregnancy Testing (Specific to Austria)
Pregnancy tests will be performed at the times listed in the Schedule of Activities.  
Pregnancy tests may also be repeated if requested by institutional review boards 
(IRBs)/ethics committees (ECs) or if required by local regulations. 
7. SAFETY
7.1. Adverse Events
An AE is defined as any untoward medical occurrence and can therefore be any unfavorable 
and unintended sign (including an abnormal laboratory finding), symptom, or disease, 
whether or not related to the patient’s participation in the study.
Any AE that occurs from the time the patient undergoes the tumor biopsy and research blood 
collection, whichever occurs first, through and including 24 hours post tumor biopsy and 
blood collection must be recorded.  The investigator is required to assess whether the AE 
may be related to the patient’s participation in the study.
The investigator must pursue and obtain information adequate to determine the outcome of 
the AE and to assess whether it meets the criteria for classification as a research related 
injury requiring immediate notification to Pfizer as described below.
7.1.1. Research Related Injury
Should a patient, in the investigator’s opinion, suffer a medically important research related 
injury caused by their participation in the study, the designated Pfizer clinician or medical 
monitor must be notified immediately by emailing 
A medically important research related injury is any untoward medical occurrence that:
•
Results in death;
•
Is life-threatening (immediate risk of death);
•
Requires inpatient hospitalization or prolongation of existing hospitalization;
•
Results in persistent or significant disability/incapacity (substantial disruption of the 
ability to conduct normal life functions);
•
Results in congenital anomaly/birth defect.
Medical and scientific judgment is exercised in determining whether an injury is an 
important medical event.  An important medical event may not be immediately 
life-threatening and/or result in death or hospitalization.  However, if it is determined that the 
event may jeopardize the patient or may require intervention to prevent one of the other 
outcomes listed in the definition above, the important medical event should be reported as a 
research related injury.
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An investigator may be requested by the designated Pfizer clinician or medical monitor to 
obtain specific additional follow-up information in an expedited fashion.  In general, this will 
include a description of the injury in sufficient detail to allow for a complete medical 
assessment of the case and independent determination of possible causality.  Information on 
other possible causes of the event, such as concomitant treatments, vaccines, and/or illnesses 
must be provided.  In the case of a patient death, a summary of available autopsy findings 
must be submitted as soon as possible to Pfizer or its designated representative.
8. DATA ANALYSIS/STATISTICAL METHODS
Detailed methodology for summary and statistical analyses of the data collected in this study 
will be documented in a statistical analysis plan (SAP), which will be maintained by Pfizer.
This document may modify the plans outlined in the protocol; however, any major 
modifications of the primary endpoint and/or its analysis will also be reflected in a protocol 
amendment.
8.1. Analysis Population
All the endpoints will be analyzed in the evaluable population.  The evaluable population is 
defined as the patients who have both archival tumor tissue and de novo tumor biopsy that is 
sufficient to enable the intended sample analyses.
8.2. Sample Size Determination
The primary objective of this study is to obtain and analyze archival and post-progression 
tumor biopsies to identify molecular markers of resistance to selected standard-of-care 
anti-cancer therapies. 
There is no standard definition of clinically meaningful change in the frequency of gene 
alteration between resistant and sensitive samples.  Therefore, characterization of the 
difference of the frequency of alterations between archival and post-progression tumor in this 
study is considered hypothesis-generating.  The precision of estimates (eg, 95% confidence 
intervals) are provided in Table 2 for different sample sizes (50, 75, 100) and observed 
changes in the frequency of gene alteration (10%, 20% and 30%). This covers the sample 
sizes considered for different tumor types in Table 1. 
Table 2 provides general guidance about the sample size per cohort. The final sample size 
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Table 2.
Sample Size And 95% Score Based Confidence Intervals For Change In The 
Frequency Of Gene Alterations After Disease Progression
Number of patients
Observed Difference
95% CI
100
10%
(6%, 17%)
100
20%
(13%, 29%)
100
30%
(22%, 40%)
75
10%
(5%, 19%)
75
20%
(13%, 30%)
75
30%
(21%, 41%)
50
10%
(2%, 21%)
50
20%
(11%, 33%)
50
30%
(19%, 44%)
8.3. Efficacy Analysis
Formal efficacy analyses will not be performed for this study.
8.4. Safety Analysis
Formal safety analyses will not be performed for this study.
8.5. Analysis of Other Endpoints
8.5.1. Analysis of Primary Endpoint
The primary endpoint is the change in the frequency of gene alterations associated with 
disease progression.  As the goal is to compare the differences in frequency of mutation rate 
between pre-treatment archival and post-progression samples, a score based 95% CI for 
matched pairs will be calculated for each genomic alteration.75,76  Estimates of the frequency 
of mutations, the difference of the frequency of mutations between archival and 
post-progression samples and associated 95% CIs will be generated. 
8.5.2. Analysis of Secondary Endpoints
Summary statistics will be generated for the secondary endpoints.  For the proportion of 
patients with fully evaluable archival and post-progression tumor biopsies, proportions and 
95% CI will be calculated.  For the overall agreement rates of gene alterations between 
post-progression biopsy tissue and blood NGS results, the frequency of mutations detected 
from post-progression biopsy tissue, blood and the corresponding 95% CIs will be generated.  
The concordance between both matched tissue biopsy tissue and blood samples and the 
95% CI will also be calculated to evaluate the agreement.

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8.5.3. Analysis of Asset Specific Secondary Endpoints
Table 3.
Analysis of Asset Specific Secondary Endpoints
Asset
Endpoint
Analysis
PD-1/-L1 
inhibition

Change in the frequency of 
alterations in genes encoding 
HLA, 2-Microglobulin, 
STAT1, JAK1, JAK2, IFN-and 
IFNGR between pre-treatment 
archival and post-progression 
samples.

The frequency of alterations in 
genes encoding HLA, 
2-Microglobulin, STAT1, 
JAK1, JAK2, IFN- and IFNGR 
in cfDNA.

For each alteration, the estimate of the 
frequency and the 95% CI at 
pre-treatment archival and 
post-progression will be generated; A 
score based 95% CI will be generated 
for the difference in frequency between 
pre-treatment archival and 
post-progression samples.

For each alteration in cfDNA, the 
estimate of the frequency and the 
95% CI will be generated. 
CDK4/6 inhibition

Change in the frequency of RB1 
gene alterations between 
pre-treatment archival and 
post-progression samples.

The frequency of RB1 gene 
alterations in cfDNA.

For each alteration, the estimate of the 
frequency and the 95% CI at 
pre-treatment archival and 
post-progression will be generated.  A 
score based 95% CI will be generated 
for the difference in frequency between 
pre-treatment archival and 
post-progression samples.

For each alteration in cfDNA, the 
estimate of the frequency and the 
95% CI will be generated.
AR inhibition

Change in the frequency of AR 
gene alterations between 
pre-treatment archival and 
post-progression samples.

The frequency of AR gene 
alterations in cfDNA.

Changes in the expression of 
nuclear hormone receptors or 
related RNA signatures 
reflecting nuclear receptor 
pathway activity between 
pre-treatment archival and 
post-progression samples.

For each alteration in cfDNA, the 
estimate of the frequency and the 
95% CI at pre-treatment archival and 
post-progression will be generated.  A 
score based 95% CI will be generated 
for the difference in frequency between 
pre-treatment archival and 
post-progression samples.

For each alteration, the estimate of the 
frequency and the 95% CI will be 
generated.

Estimates of mean expression and 
95% CIs at pre-treatment archival and 
post progression will be generated; 
Estimates of the difference in 
expression and the 95% CI between 
pre-treatment archival and 
post-progression samples will be 
generated.
PARP inhibition

Change in the frequency of 
somatic reversion alterations in 
gBRCA mutant allele between 
pre-treatment archival and 
post-progression samples.

For each somatic reversion alteration, 
the estimate of the frequency and the 
95% CI at pre-treatment archival and 
post-progression will be generated.  A 
score based 95% CI will be generated 
for the difference in frequency between 

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Asset
Endpoint
Analysis
pre-treatment archival and 
post-progression samples.
 
 
 
 
 
 
8.6. Interim Analysis
A formal interim analysis will not be performed for this study.  However, multiple snapshots 
of the data will be taken for data review based on accrual (eg, at 25%, 50% and 75% of 
cohort and/or study accrual). 
9. QUALITY CONTROL AND QUALITY ASSURANCE
Pfizer or its agent will conduct periodic monitoring visits during study conduct for studies 
conducted at non-Pfizer investigator sites, to ensure that the protocol and Good Clinical 
Practices (GCPs) are being followed.  The monitors may review source documents to 
confirm that the data recorded on CRFs are accurate.  The investigator and institution will 
allow Pfizer monitors/auditors or its agents and appropriate regulatory authorities direct 
access to source documents to perform this verification.  This verification may also occur 
after study completion.
During study conduct and/or after study completion, the investigator site may be subject to 
review by the Institutional Review Board (IRB)/Ethics Committee (EC,) and/or to quality 
assurance audits performed by Pfizer, or companies working with or on behalf of Pfizer, 
and/or to inspection by appropriate regulatory authorities.
The investigator(s) will notify Pfizer or its agents immediately of any regulatory inspection 
notification in relation to the study.  Furthermore, the investigator will cooperate with Pfizer 
or its agents to prepare the investigator site for the inspection and will allow Pfizer or its 
agent, whenever feasible, to be present during the inspection.  The investigator site and 
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investigator will promptly resolve any discrepancies that are identified between the study 
data and the patient’s medical records.  The investigator will promptly provide copies of the 
inspection findings to Pfizer or its agent.  Before response submission to the regulatory 
authorities, the investigator will provide Pfizer or its agents with an opportunity to review 
and comment on responses to any such findings.
For studies conducted at non-Pfizer investigator sites, it is important that the investigator(s) 
and their relevant personnel are available during the monitoring visits and possible audits or 
inspections and that sufficient time is devoted to the process.
10. DATA HANDLING AND RECORD KEEPING
10.1. Case Report Forms/Data Collection Tools/Electronic Data Record
As used in this protocol, the term CRF should be understood to refer to either a paper form or 
an electronic data record or both, depending on the data collection method used in this study.
A CRF is required and should be completed for each included patient.  The completed 
original CRFs are the sole property of Pfizer and should not be made available in any form to 
third parties, except for authorized representatives of Pfizer or appropriate regulatory 
authorities, without written permission from Pfizer.  The investigator shall ensure that the 
CRFs are securely stored at the study site in encrypted electronic form and will be password 
protected or secured in a locked room to prevent access by unauthorized third parties.
The investigator has ultimate responsibility for the collection and reporting of all clinical, 
safety, and laboratory data entered on the CRFs and any other data collection forms (source 
documents) and ensuring that they are accurate, authentic/original, attributable, complete, 
consistent, legible, timely (contemporaneous), enduring, and available when required.  The 
CRFs must be signed by the investigator or by an authorized staff member to attest that the 
data contained on the CRFs are true.  Any corrections to entries made in the CRFs or source 
documents must be dated, initialed, and explained (if necessary) and should not obscure the 
original entry.
In most cases the source documents are the hospital or the physician's chart.  In these cases, 
data collected on the CRFs must match those charts. 
In some cases, the CRF may also serve as the source document.  In these cases, a document 
should be available at the investigator site and at Pfizer that clearly identifies those data that 
will be recorded on the CRF, and for which the CRF will stand as the source document. 
10.2. Record Retention
To enable evaluations and/or inspections/audits from regulatory authorities or Pfizer, the 
investigator agrees to keep records, including the identity of all participating patients 
(sufficient information to link records, eg, CRFs and hospital records), all original signed 
informed consent documents, copies of all CRFs, safety reporting forms, source documents, 
detailed records of treatment disposition, and adequate documentation of relevant 
correspondence (eg, letters, meeting minutes, and telephone call reports).  The records should 
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(ICH) guidelines, according to local regulations, or as specified in the clinical study 
agreement (CSA), whichever is longer.  The investigator must ensure that the records 
continue to be stored securely for so long as they are retained.
If the investigator becomes unable for any reason to continue to retain study records for the 
required period (eg, retirement, relocation), Pfizer should be prospectively notified.  The 
study records must be transferred to a designee acceptable to Pfizer, such as another 
investigator, another institution, or to an independent third party arranged by Pfizer. 
Investigator records must be kept for a minimum of 15 years after completion or 
discontinuation of the study or for longer if required by applicable local regulations. 
The investigator must obtain Pfizer's written permission before disposing of any records, 
even if retention requirements have been met.
11. ETHICS
11.1. Institutional Review Board/Ethics Committee
It is the responsibility of the investigator to have prospective approval of the study protocol, 
protocol amendments, informed consent documents, and other relevant documents, eg, 
recruitment advertisements, if applicable, from the IRB/EC.  All correspondence with the 
IRB/EC should be retained in the investigator file.  Copies of IRB/EC approvals should be 
forwarded to Pfizer.
The only circumstance in which an amendment may be initiated prior to IRB/EC approval is 
where the change is necessary to eliminate apparent immediate hazards to the patients.  In 
that event, the investigator must notify the IRB/EC and Pfizer in writing immediately after 
the implementation.
11.2. Ethical Conduct of the Study
The study will be conducted in accordance with the protocol, legal and regulatory 
requirements, and the general principles set forth in the International Ethical Guidelines for 
Biomedical Research Involving Human Subjects (Council for International Organizations of 
Medical Sciences 2002), ICH Guideline for Good Clinical Practice, and the Declaration of 
Helsinki.
11.3. Patient Information and Consent
All parties will comply with all applicable laws, including laws regarding the implementation 
of organizational and technical measures to ensure protection of patient personal data.  Such 
measures will include omitting patient names or other directly identifiable data in any 
reports, publications, or other disclosures, except where required by applicable laws. 
The personal data will be stored at the study site in encrypted electronic form and/or paper 
form and will be password protected or secured in a locked room to ensure that only 
authorized study staff have access.  The study site will implement appropriate technical and 
organizational measures to ensure that the personal data can be recovered in the event of 
disaster.  In the event of a potential personal data breach, the study site shall be responsible 

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for determining whether a personal data breach has in fact occurred and, if so, providing 
breach notifications as required by law.
To protect the rights and freedoms of natural persons with regard to the processing of 
personal data, when study data are compiled for transfer to Pfizer and other authorized 
parties, patient names will be removed and will be replaced by a single, specific, numerical 
code, based on a numbering system defined by Pfizer.  All other identifiable data transferred 
to Pfizer or other authorized parties will be identified by this single, patient-specific code.  
The investigator site will maintain a confidential list of patients who participated in the study, 
linking each patient’s numerical code to his or her actual identity.  In case of data transfer, 
Pfizer will maintain high standards of confidentiality and protection of patients’ personal data 
consistent with the Clinical Study Agreement and applicable privacy laws.
The informed consent documents and any patient recruitment materials must be in 
compliance with ICH GCP, local regulatory requirements, and legal requirements, including 
applicable privacy laws.
The informed consent documents used during the informed consent process and any patient 
recruitment materials must be reviewed and approved by Pfizer, approved by the IRB/EC 
before use, and available for inspection.
The investigator must ensure that each study patient is fully informed about the nature and 
objectives of the study, the sharing of data relating to the study and possible risks associated 
with participation, including the risks associated with the processing of the patient’s personal 
data.  The investigator further must ensure that each study patient is fully informed about his 
or her right to access and correct his or her personal data and to withdraw consent for the 
processing of his or her personal data.
The investigator, or a person designated by the investigator, will obtain written informed 
consent from each patient before any study-specific activity is performed.  The investigator 
will retain the original of each patient's signed consent document.
12. PUBLICATIONS BY INVESTIGATORS
Pfizer supports the exercise of academic freedom and has no objection to publication by the 
principal investigator (PI) of the results of the study based on information collected or 
generated by the PI, whether or not the results are favorable to the Pfizer product.  However, 
to ensure against inadvertent disclosure of confidential information or unprotected 
inventions, the investigator will provide Pfizer an opportunity to review any proposed 
publication or other type of disclosure of the results of the study (collectively, “publication”) 
before it is submitted or otherwise disclosed.
The investigator will provide any publication to Pfizer at least 30 days before it is submitted 
for publication or otherwise disclosed.  If any patent action is required to protect intellectual
property rights, the investigator agrees to delay the disclosure for a period not to exceed an 
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The investigator will, on request, remove any previously undisclosed confidential 
information before disclosure, except for any study- or Pfizer product-related information 
necessary to the appropriate scientific presentation or understanding of the study results.
If the study is part of a multicenter study, the investigator agrees that the first publication is 
to be a joint publication covering all investigator sites, and that any subsequent publications 
by the PI will reference that primary publication.  However, if a joint manuscript has not 
been submitted for publication within 12 months of completion or termination of the study at 
all participating sites, the investigator is free to publish separately, subject to the other 
requirements of this section.
For all publications relating to the study, the institution will comply with recognized ethical 
standards concerning publications and authorship, including Section II - “Ethical 
Considerations in the Conduct and Reporting of Research” of the Uniform Requirements for 
Manuscripts Submitted to Biomedical Journals, http://www.icmje.org/index.html#authorship, 
established by the International Committee of Medical Journal Editors.
Publication of study results is also provided for in the CSA between Pfizer and the 
institution.  In this section entitled Publications by Investigators, the defined terms shall have 
the meanings given to them in the CSA.
If there is any conflict between the CSA and any attachments to it, the terms of the CSA 
control.  If there is any conflict between this protocol and the CSA, this protocol will control 
as to any issue regarding treatment of study patients, and the CSA will control as to all other 
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13. REFERENCES

McGranahan N, Swanton C. Cancer Evolution Constrained by the Immune 
Microenvironment. Cell 2017; (170):825-27.

Thorsson V, Gibbs DL, Brown SD, et al. The Immune Landscape of Cancer. Immunity 
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Cancer. Clin Cancer Res 2017; (23):523-35.

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Appendix 1. Abbreviations
This following is a list of abbreviations that may be used in the protocol.
Abbreviation
Term
AE
adverse event
AI
Aromatase inhibitor
ALK
anaplastic lymphoma kinase
AR
Androgen receptor
ARID1A
AT-rich interactive domain-containing protein 1A
AR-V
Androgen receptor splice variant
BRCA
BReast CAncer susceptibility gene
cfDNA
circulating free DNA
CAP
College of American Pathologists
CCND1
Cyclin D1
CCND3
Cyclin D3
CCNE1
cyclin E1 gene
CDK
Cyclin-dependent kinase
CDKN2A
Cyclin-dependent 2A gene
CI
Confidence interval
CLIA
Clinical Laboratory Improvements Amendments
CRF
Case report form
CRISPR
Clustered Regularly Interspaced Short Palindromic Repeats
CRPC
Castrate resistant prostate cancer
CSA
clinical study agreement
CSR
clinical study report
CTC
Circulating tumor cell
CTCAE
Common terminology criteria for adverse events
CTLA4
cytotoxic T-lymphocyte-associated protein 4
DCT
data collection tool
DNA
deoxyribonucleic acid
EC
ethics committee
EGFR
epidermal growth factor receptor
EOT
End of treatment
ER
Estrogen receptor
ESR1
estrogen receptor 1
EZH2
Enhancer of zeste homolog 2
FISH
Fluorescent in situ hybridization
FFPE
Formalin fixed paraffin embedded
FSH
follicle-stimulating hormone
gBRCAm
Germline mutated BRCA
GCP
Good Clinical Practice
GPBP1
GC-Rich Promoter Binding Protein 1
H&E
Hematoxylin and Eosin
HER2
human epidermal growth factor receptor 2

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Abbreviation
Term
HLA
human leukocyte antigen
HR
Hormone receptor
HR DDR
homologous recombination-mediated DNA damage repair
ICH
International Conference on Harmonisation
IFN-
interferon-
IFNGR
interferon-gamma receptor 
IHC
Immunohistochemistry
INK 
inhibitor of the cyclin-dependent kinase
IPRES
innate anti-PD-1 resistance signature
IRB
institutional review board
JAK
janus kinase
K2EDTA
dipotassium ethylenediaminetetraacetic acid
LAG-3
lymphocyte activation gene-3
LOH
Loss of heterozygosity
MAPK
mitogen-activated protein kinase 
MDSC
myeloid derived suppressor cell
MHC
Major histocompatibility
N/A
not applicable
NCI
National Cancer Institute
NEPC
Neuro-endocrine prostate cancer
NGS
Next generation sequencing 
NSCLC 
Non-small cell lung cancer
ORF
Open reading frame
PACL
Protocol administrative clarification letter
PALB2
Partner and localizer of BRCA2 
PARP
Poly (ADP-ribose) polymerase
PARPi
PARP inhibitor
PD
Progressive disease
PD-1
programmed cell death receptor 1
PD-L1 
programmed cell death-ligand 1
PD-1/-L1
programmed cell death receptor 1 or programmed cell 
death-ligand 1
PDx
PD-1/-L1 
PI
principal investigator
PI3K
phosphoinositide 3-kinase
PK 
Pharmacokinetics
PGx
Pharmacogenomics
PgR
Progesterone receptor
PTEN 
phosphatase and tensin homolog
RB1
Retinoblastoma - gene
Rb
Retinoblastoma – gene product
RCC
Renal cell carcinoma
RNA
Ribonucleic acid
ROS1
c-ros oncogene 1

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Abbreviation
Term
SAP
Statistical Analysis Plan
SCCHN
squamous cell carcinoma of the head and neck
SLFN11
Loss of Schlafen
SoA
Schedule of Activities
SOC
Standard of care
SOP
Standard operational procedures
STAT1
Signal transducer and activator of transcription 1
TA
therapeutic area
TCR
T-cell receptor
TIM-3
T-cell immunoglobulin and mucin-domain containing-3
TMB
Tumor mutational burden
WES
Whole Exome Sequencing

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CCI
CCI
CCI
CCI
CCI
CCI

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C
C
I

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CCI
C
C
I

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C
C
I
CCI

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Appendix 4. Country-Specific Requirements
France Contrat Unique
1. GCP Training.
Before enrolling any participants, the investigator and any subinvestigators will 
complete the Pfizer-provided Good Clinical Practice training course (“Pfizer GCP 
Training”) or training deemed equivalent by Pfizer.  Any investigators who later join 
the study will do the same before performing study-related duties.  For studies of 
applicable duration, the investigator and subinvestigators will complete Pfizer GCP 
Training or equivalent every 3 years during the term of the study, or more often if 
there are significant changes to the ICH GCP guidelines or course materials. 
2. Study Intervention.
No participants or third-party payers will be charged for study intervention.
3. Urgent Safety Measures.
In addition, the investigator will inform Pfizer immediately of any urgent safety 
measures taken by the investigator to protect the study participants against any 
immediate hazard, and of any serious breaches of this protocol or of ICH GCP that 
the investigator becomes aware of.
4. Termination Rights.
Pfizer retains the right to discontinue the A9001502 study at any time. There is no 
Investigational Medicinal Product in the study.