Genetic Analysis in Blood and Tumor Samples From Patients With Advanced or Metastatic Estrogen Receptor Positive and HER2 Negative Breast Cancer Receiving Palbociclib and Endocrine Therapy

Study Overview
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Brief Summary
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This research trial studies genetic profiles in blood and tumor samples from patients with estrogen receptor positive and HER2 negative breast cancer that has spread to other places in the body who are receiving palbociclib and endocrine therapy. Examining the genetic changes associated with the cancer and comparing the genetic material from the cancer tissue with the genetic material found in the blood may help doctors to develop customized treatment for breast cancer.

Detailed Description
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PRIMARY OBJECTIVES: I. To identify novel genomic variants and pathways associated with baseline, as well as the change in serum thymidine kinase (TK1) levels (measured pre-registration, after two cycles, and at disease progression) among women with advanced hormone receptor (HR) positive breast cancer treated with palbociclib and endocrine therapy (ET). SECONDARY OBJECTIVES: I. To generate patient derived xenograft (PDX)s and organoids from tumor biopsies obtained at pre-registration, at 2 months, and at disease progression, to test novel endocrine therapy combinations. (PDX Aim) II. To test whether PDX engraftment is associated with progression free survival. (PDX Aim) Sequencing Aims III. to describe changes in the genomic landscape of HR-positive HER2-negative advanced breast cancer by comparing sequencing data (deoxyribonucleic acid [DNA] and ribonucleic acid [RNA]) from preregistration archival breast primary tumors (when available), with tumor biopsies obtained preregistration, after two months on study, and at disease progression on palbociclib and ET. IV. To evaluate whether genomic variants and pathways (measured using DNA and RNA sequencing at pre-registration and after two months of treatment) are associated with 1) PFS; 2) baseline and changes in serum TK1. (Sequencing Aim) V. To tabulate the number of patients who have targetable mutations found in the preregistration tumor biopsy and to record their subsequent treatment regimen following progression on palbociclib and ET. (Sequencing Aim) VI. To compare and contrast the actionable genomic alterations found in cfDNA or CTC derived DNA that are not found in tumor tissue. (Sequencing Aim) VII. To assess whether genetic alterations known to be associated with endocrine monotherapy resistance are seen in circulating tumor- DNA (ctDNA) and circulating tumor cell (CTC) derived DNA. (Sequencing Aim) VIII.To determine whether use of a 3D micro cancer model, combined with sequencing data from tumor biopsies, ctDNA and PDXs collected at the time of disease progression, can be used to personalize future treatment options for PROMISE participants. (Sequencing Aim) IX. To compare the changes in Ki67 levels (by immunohistochemistry [IHC]) and serum TK1 levels after 2 months of treatment. (Proliferative biomarker aim) X. To assess whether the change in Ki67 and serum TK1 levels after 2 months of treatment with palbociclib and ET is associated with progression free survival (PFS) outcomes. (Proliferative biomarker aim) XI. To assess whether high baseline serum TK1 levels (> 200 Du/L) are associated with PFS outcomes. (Proliferative biomarker aim) XII. To assess whether an increase in TK1 between 2 months and 12 months is associated with PFS. (Proliferative biomarker aim) XIII. To assess the change in EMT marker expression (e.g. Vimentin, SLUG and E-cadherin) and serum TK1 after two months of treatment with palbociclib and ET. (EMT aim) XIV. To determine the association between the expression of EMT markers and Ki67 levels after two months of treatment with palbociclib and ET. (EMT aim) XV. To determine the association between the expression of EMT markers after two months of treatment with palbociclib and ET, and PFS outcomes. (EMT aim) XVI. To assess whether there is a change in the level of tumor infiltrating lymphocytes (TILs) observed in archival tumor, at pre-registration, at 2 months, and at disease progression on palbociclib and ET. (Immune aim) XVII. To assess the change in TILs (including CD8, PD-L1, and FOXP3) after 2 months of treatment with palbociclib and ET. (Immune aim) XVIII. To assess the association between TILs and serum TK1 levels at preregistration, at 2 months, and at disease progression on palbociclib and ET. (Immune aim) XIX. To characterize and evaluate changes in the immune landscape of HR+ HER2 -MBC using serial biopsies obtained pre-registration, at 2 months, and at disease progression (as well as archived primary tumor when available) using digital spatial profiling (DSP) (NanoString GeoMx platform) . (Immune aim) XX. To assess, by use of mass cytometry (CyTOF), whether i) peripheral blood immune markers obtained pretreatment are associated with either TK1 or PFS and ii) whether the changes in peripheral blood immune markers after 2 cycles of treatment are associated with changes in serum TK1 levels. (Immune aim) XXI. To discover putative mechanistic connections underlying bacteria-drug interactions in all patients. (Microbiome aim) XXII. To identify the biomolecular features within the gut (stool) microbiome and its association with the pharmacokinetics and pharmacodynamics of letrozole and palbociclib. (Microbiome aim) XXIII. To discover novel predictive biomarkers of resistance to CDK4/6i and ET and/or to find biomarkers that add additional predictive power to other promising biomarkers such as TK1 by performing the following: (Metabolic, proteomic and lipidomic aim) XXIV. A metabolomics analysis (>5,000 metabolites sampled through untargeted mass spectrometry at Metabolon) of paired serum samples obtained pre-registration and after 2 months on palbociclib and ET. XXIVa. An Olink-based cytokines panel (Explore panel of 1,536 proteins) of paired serum samples obtained pre-registration and after 2 months on palbociclib and ET. XXIVb. A lipidomics analysis (platform TBC) of paired serum samples obtained pre-registration and after 2 months on palbociclib and ET. XXV. To correlate the changes in the metabolites, cytokines and lipids to the preregistration genomic tumor characteristics, serial Ki67 levels and PFS outcomes. (Metabolic, proteomic and lipidomic aim) XXVI. To assess the change in serum TK1 levels after 2 months, and at disease progression, on palbociclib and ET, and its association with the preregistration CD44high/CD24/low/ERlow cancer stem cell-like phenotype, including the following: total and phosphorylated AURKA, breast cancer stemness biomarkers (CD44, CD24, ALDH1) EMT transcription factors (SMAD5, SOX2), cyclin E, cyclin A and phosphorylated Retinoblastoma (Rb). (Other aim) XXVII. To examine shifts in the populations of epithelial like CTCs and stem cell like CTCs during the course of endocrine therapy with palbociclib. (Other aim) XXVIII. To assess CTC expression of ER, HER2, and other markers of endocrine resistance. (Other aim) OUTLINE: Patents undergo collection of blood and stool samples at baseline, 7 days after letrozole monotherapy treatment, and at completion of each cycle, urine samples at baseline and completion of each cycle, and saliva samples at baseline. Patients also undergo collection of blood and urine samples at disease progression. Biopsy samples are analyzed for genetic profile via genome sequencing and ribonucleic acid (RNA) sequencing. Biopsy samples are also used for the generation of xenograft mice model. After completion of study, patients are followed up every 6 months for 7 years.

Official Title
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Prospective Study to Evaluate the Role of Tumor Sequencing in Women Receiving Palbociclib for Advanced Hormone Receptor (HR)-Positive, Breast Cancer (PROMISE)

Conditions
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Advanced Breast Carcinoma, Locally Advanced Breast Carcinoma, Metastatic Breast Carcinoma, Recurrent Breast Carcinoma, Stage III Breast Cancer AJCC v7, Stage IIIA Breast Cancer AJCC v7, Stage IIIB Breast Cancer AJCC v7, Stage IIIC Breast Cancer AJCC v7, Stage IV Breast Cancer AJCC v6 and v7

Intervention / Treatment
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* Procedure: Biopsy
* Procedure: Biospecimen Collection
* Other: Laboratory Biomarker Analysis


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: PRE-REGISTRATION INCLUSION CRITERIA Women who have disease that is amenable to biopsy and agree to undergo a standard of care and /or research biopsy Note: If a standard of care biopsy was recently obtained =< 2 months of pre-registration, eligible patients should agree to a research biopsy of recurrent or metastatic breast cancer prior to the start of protocol treatment to collect additional core samples for research purposes Patients must satisfy one of the following criteria for prior therapy: First line setting: No prior endocrine therapy in the metastatic setting with no more than one prior line of chemotherapy in the advanced/metastatic setting Second line setting: Progression on one prior line of endocrine based therapy monotherapy either in the adjuvant or advanced/metastatic setting; either one or two prior lines of chemotherapy in the advanced setting are allowed Note: Patients receiving bisphosphonate or denosumab therapy prior to registration may continue at the same intervals used prior to study registration First line therapy setting only: The intention to begin palbociclib and letrozole as treatment for locally advanced or metastatic breast cancer Second line therapy setting only: The intention to begin palbociclib and fulvestrant as treatment for metastatic breast cancer (after progression on first line endocrine therapy) Note: Patients who are to receive second line endocrine therapy are allowed to remain on their most recent treatment (tamoxifen or an aromatase inhibitor during the pre-registration period as well as after registration while awaiting insurance approval for the use of palbociclib Measurable disease as defined by Response Evaluation Criteria in Solid Tumors (RECIST) criteria or bone only disease are eligible. Note: Those patients with both non-measurable disease and bone metastases are eligible Note: Patients are not allowed to begin a new systemic anti-cancer therapy during pre-registration with the exception of bisphosphonate or denosumab; palliative radiation to lesions that will not be biopsied or used for assessing disease response (target lesions) is allowed during pre-registration No current evidence of visceral crisis History of central nervous system metastasis are allowed provided they have been treated (i.e., surgery, radiation, and/or radiosurgery) >= 12 weeks prior to pre-registration and have stable neurologic function, including no requirement for medication(s) to control symptoms for at least 2 weeks; Note: patients with known leptomeningeal disease are not eligible Women who are premenopausal must agree to begin or continue an leutinizing hormone releasing hormone (LHRH) agonist (goserelin preferred) NOTE: A woman is considered premenopausal if menses has occurred in the last 12 months prior to preregistration and both serum and follicle stimulating hormone (FSH) levels are not in the laboratory's reference range for postmenopausal females Eastern Cooperative Oncology Group (ECOG) performance status: 0, 1, or 2 Able to swallow oral formulation of drugs Signed and dated informed consent document for study participation Willing to submit tissue, blood, stool, and saliva and urine for required correlative research REGISTRATION INCLUSION CRITERIA Histologic confirmation from the pre-registration biopsy of either locally advanced or metastatic breast cancer that is ER-positive and HER2 -negative Note: ER-positive disease is defined as >= 10% nuclear staining; HER2-negative disease per American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) guidelines, one of the following must apply: 0 or 1+ by immunohistochemistry (IHC) and not amplified by in situ hybridization (ISH) 0 or 1+ by IHC and ISH not done 2+ by IHC and not amplified by ISH or IHC not done and not amplified by ISH Absolute neutrophil count (ANC) >= 1500/mm^3 (=< 14 days prior to registration) Platelet count >= 100,000/mm^3 (=< 14 days prior to registration) Hemoglobin >= 9.0 g/dL (=< 14 days prior to registration) Total bilirubin =< 1.5 x upper limit of normal (ULN), (=< 3 x ULN if Gilbert's disease) (=< 14 days prior to registration) Aspartate transaminase (AST) =< 3 x ULN (=< 5 x ULN if liver metastases present) (=< 14 days prior to registration) Creatinine =< 1.5 x ULN (=< 14 days prior to registration) Willing to return to enrolling institution for follow-up (during the active monitoring phase of the study) Toxicities related to all prior anticancer therapies must have resolved or stabilized, apart from alopecia and peripheral neuropathy; Note: Peripheral neuropathy which has resolved to =< grade 2 toxicity is acceptable Exclusion Criteria: PRE-REGISTRATION EXCLUSION CRITERIA History of metastatic ER negative or HER2 positive breast cancer Prior treatment in the metastatic setting with everolimus, or any agent whose mechanism of action is to inhibit the PI3K-mTOR pathway Uncontrolled intercurrent illness including, but not limited to: Ongoing or active infection Symptomatic congestive heart failure Unstable angina pectoris Uncontrolled symptomatic cardiac arrhythmia Uncontrolled hypertension (defined as blood pressure > 160/90) Other active second malignancy other than non-melanoma skin cancers =< 3 years of pre-registration; Note: a second malignancy is not considered active if all treatment for that malignancy is completed and the patient has been disease-free for =< 3 years prior to pre-registration Prior hematopoietic stem cell or bone marrow transplantation =< 3 years of pre-registration Known hypersensitivity to palbociclib, letrozole, fulvestrant, goserelin (if applicable) or to any of their excipients Known to be pregnant and planning to continue nursing REGISTRATION EXCLUSION CRITERIA No tumor identified on biopsy or insufficient tumor cells to obtain ER or HER2 status Any of the following therapies prior to registration: Chemotherapy =< 2 weeks Immunotherapy =< 2 weeks Biologic therapy =< 2 weeks Monoclonal antibodies =< 2 weeks Note: Denosumab is exempt from this requirement and can be started at any time prior to initiation of palbociclib and letrozole/fulvestrant without the need to delay protocol therapy Radiation therapy =< 2 weeks Note: Palliative radiation therapy to lesions that will not be biopsied for this study or used for assessing disease response (target lesions) is permitted during the pre-registration period and does not count towards the 2 week window Anti HER2 or other targeted therapy =< 2 weeks The following patients are not eligible Pregnant women Nursing women Women of childbearing potential who are unwilling to employ adequate contraception Women who have already commenced treatment with an aromatase inhibitor (first line setting) or fulvestrant (second line setting) prior to screening for this study or any prior palbociclib

Ages Eligible for Study
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Minimum Age: 18 Years

Sexes Eligible for Study
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Female

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Ancillary-Correlative (genetic profile analysis)<br>Patents undergo collection of blood and stool samples at baseline, 7 days after letrozole monotherapy treatment, and at completion of each cycle, urine samples at baseline and completion of each cycle, and saliva samples at baseline. Patients also undergo collection of blood and urine samples at disease progression. Biopsy samples are analyzed for genetic profile via genome sequencing and RNA sequencing. Biopsy samples are also used for the generation of xenograft mice model. | Procedure: Biopsy<br>* Undergo tumor biopsy<br>* Other names: Bx;Procedure: Biospecimen Collection<br>* Undergo collection of blood, urine, stool, and saliva<br>Other: Laboratory Biomarker Analysis<br>* Correlative studies<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Bioinformatics analysis | Next-generation sequencing data will be used to identify variants associated with the progression free survival. Pathology analysis will also be performed. | Up to 3 years | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Ki67 and TK1 changes | Spearman rank correlation coefficients will be used to assess the relationship between tumor ki67 levels and serum TK1 levels prior to the start of treatment and after 2 months of treatment with palbocic. | after 2 months of treatment | 
| Changes in EMT markers (including Vimentin, SLUG and E-cadherin) and tumor infiltrating lymphocytes (TILs) (including CD8, PD-L1, and FOXP3) | Wilcoxon signed rank tests will be used to assess the fold changes in EMT and TILs after 2 cycles of treatment. Benjamini-Hochberg procedure will be used to control false postive rate. | after 2 months of treatment | 
| Changes in serum TK1 levels | Wilcoxon rank sum tests will be used to assess whether a given element of the CD44high/CD24/low/estrogen receptor (ER) low cancer stem cell-like phenotype differ between those whose TK1 levels fell below 200 after 2 cycles of treatment and those whose TK1 levels remained above 200 after 2 cycles of treatment. | After 2 months of treatment | 
| Change in phenotype of Ki67 and serum TK1 levels | The parameter estimates from fitting a univariate Cox model to these data will be used to obtain an estimate of the hazard ratio and its corresponding 95% confidence interval. | After 2 months of treatment | 
| Differences between those with and without a blood draw taken | A Wilcoxon rank sum test will be used to assess whether baseline TK1 levels differ among those who discontinue treatment prior to the 2 month blood draw and those who do not. | After 2 months of treatment |