Characterizing Metastatic Breast Cancer Heterogeneity Through Post-Mortem Tissue Collection

Breast cancer metastases display significant morphologic and genomic heterogeneity.1–3 Furthermore, there is often significant heterogeneity between breast cancer metastases in a single patient, such that the receptor status, driver mutations, gene expression profile, and tumor microenvironment can change significantly over time and with exposure to therapy.4 Despite this fact, many large-scale genomics studies examining patient-derived tissue have focused on the analysis of primary tumors available at the time of initial diagnosis or surgical resection.5 While informative, analysis of these specimens does not allow for the study of drug resistance mechanisms, nor does it contribute to our understanding of the evolution of metastatic disease. Breast cancer often metastasizes to sites that are technically challenging to biopsy, and consequently, tumor sampling in these situations is limited. When metastatic tissue is obtained, specimens are often small, of limited cellularity, and must be reserved for diagnostic purposes and clinical immunohistochemistry studies. For these reasons, advanced metastatic breast cancer tissue is rarely available for scientific research, and relatively little is known about genomic and cellular alterations present in end-stage metastases. In order to improve therapy for patients with metastatic breast cancer, it is critical to understand the biology of metastatic tumors that ultimately become lethal. The medical autopsy is a traditional tool that pathologists have used for centuries; when coupled with modern advances in genomics, it is an exceptional method for studying the metastatic process and for obtaining tumor samples for analysis. Post-mortem examination allows for the collection of breast cancer metastases from multiple body sites (e.g. bone, liver, lung, brain) in the same patient that can then be analyzed for genomic differences and compared to the primary tumor. Patients that die from metastatic disease often have a large tumor burden at the time of death.6 In this study, metastatic tissue will be banked, and a subset of the collected tissue will be used for NanoString based gene expression profiling. The results of these studies will enhance our understanding of how breast cancer changes during the metastatic process, and how it adapts to grow in different metastatic niches. Importantly for this study, the University of Kentucky Sanders-Brown Center on Aging operates a successful program to conduct short post-mortem interval autopsies in patients with neurodegenerative disease. This successful program will serve as a model for the proposed project. Specific Aims: 1) Establish a tissue donation program for banking breast cancer metastases collected at the time of post-mortem examination. 2) Identify genes whose expression is significantly altered in advanced metastatic breast cancer when compared to the patient’s primary tumor. 3) Compare gene expression between breast cancer metastases at different sites within an individual in order to identify changes associated with metastatic growth in different environments. Objective: In this grant application, I propose to develop a program to bank and analyze end-stage breast cancer metastases. This will be accomplished through the development of a post-mortem tissue donation program that is linked to our institutions biorepository, the Markey Biospecimen Procurement and Translational Pathology Shared Resource Facility (BPTP SRF). My secondary objective is to use a subset of the tissue collected through the tissue donation program to define alterations in gene expression in end-stage breast cancer metastases using NanoString technology. Study Design: Patients with metastatic breast cancer will be identified and consented by clinicians at the University of Kentucky Markey Cancer Center (MCC). At the time of autopsy, a pathologist will supervise the post-mortem examination and document the extent, location, and characteristics of all visible metastatic lesions. Metastases and matched adjacent benign tissue samples will be collected from all metastatic sites that the patient has consented to. We will also obtain access to archival formalin-fixed, paraffin embedded (FFPE) tissue from the primary tumor so that we can compare gene expression in primary versus metastatic disease. Analysis of gene expression will be accomplished using NanoString technology as this method demonstrates superior reproducibility across sample types and will allow for comparison of gene expression levels between the primary tumor (FFPE) and metastatic lesions (fresh frozen). For this study, we plan to consent at least 15 patients and we estimate performing 5 autopsies during the first year of the program. We will collect and bank tumor tissue, benign tissue, blood, and bone marrow. We also have an opportunity to submit tissue from our study to the ORIEN network (Oncology Research Information Exchange Network) for whole-exome sequencing and RNA-seq. In addition to collecting tumor tissue, we plan to bank benign tissue from patients who have undergone extensive chemo and radiation therapy; these tissue samples may prove useful to researchers that study the effects of chemotherapy, for example, taxane-induced peripheral neuropathy.