Abstract

BackgroundFew somatic mutations have been linked to breast cancer metastasis, whereas transcriptomic differences among primary tumors correlate with incidence of metastasis, especially to the lungs and brain. However, the epigenomic alterations and transcription factors (TFs) which underlie these alterations remain unclear.MethodsTo identify these, we performed RNA-seq, Chromatin Immunoprecipitation and sequencing (ChIP-seq) and Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) of the MDA-MB-231 cell line and its brain (BrM2) and lung (LM2) metastatic sub-populations. We incorporated ATAC-seq data from TCGA to assess metastatic open chromatin signatures, and gene expression data from human metastatic datasets to nominate transcription factor biomarkers.ResultsOur integrated epigenomic analyses found that lung and brain metastatic cells exhibit both shared and distinctive signatures of active chromatin. Notably, metastatic sub-populations exhibit increased activation of both promoters and enhancers. We also integrated these data with chromosome conformation capture coupled with ChIP-seq (HiChIP) derived enhancer-promoter interactions to predict enhancer-controlled pathway alterations. We found that enhancer changes are associated with endothelial cell migration in LM2, and negative regulation of epithelial cell proliferation in BrM2. Promoter changes are associated with vasculature development in LM2 and homophilic cell adhesion in BrM2. Using ATAC-seq, we identified a metastasis open-chromatin signature that is elevated in basal-like and HER2-enriched breast cancer subtypes and associates with worse prognosis in human samples. We further uncovered TFs associated with the open chromatin landscapes of metastatic cells and whose expression correlates with risk for metastasis. While some of these TFs are associated with primary breast tumor subtypes, others more specifically correlate with lung or brain metastasis.ConclusionsWe identify distinctive epigenomic properties of breast cancer cells that metastasize to the lung and brain. We also demonstrate that signatures of active chromatin sites are partially linked to human breast cancer subtypes with poor prognosis, and that specific TFs can independently distinguish lung and brain relapse.

Highlights

  • Few somatic mutations have been linked to breast cancer metastasis, whereas transcriptomic differences among primary tumors correlate with incidence of metastasis, especially to the lungs and brain

  • Identifying the chromatin modification and accessibility landscape of metastatic breast cancer cells To identify epigenomic hallmarks of highly metastatic breast cancer cells, we analyzed the transcriptome and epigenome of cell sub-populations that were in vivo selected from the well-established claudin-low breast cancer cell line MDA-MB-231 (Supplemental Table 1)

  • Putative enhancer regions were defined as distal elements marked with H3K27ac peaks that fall greater than ±2 kb from the nearest transcription start site (TSS) of expressed genes and overlap with an ATAC-seq peak [59, 60]

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Summary

Introduction

Few somatic mutations have been linked to breast cancer metastasis, whereas transcriptomic differences among primary tumors correlate with incidence of metastasis, especially to the lungs and brain. The most prominent molecular subtypes identified to date include the normal-like, luminal A, luminal B, HER2 enriched (referred as HER2), basal-like, and claudin-low tumors [3,4,5]. Many genes associated with metastasis are already differentially expressed in primary tumors This is most notably observed in tumors resected from patients that are at risk for lung or brain relapse [13]. These unique metastatic propensities may be influenced, in part, by primary breast cancer subtype. Specific signatures have been associated with lung and/ or brain metastasis [21, 22], but their molecular underpinning remain unclear

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