Estrogen receptor 1 chromatin profiling in human breast tumors reveals high inter-patient heterogeneity with enrichment of risk SNPs and enhancer activity at most-conserved regions.

Estrogen Receptor 1 (ESR1; also known as ERα, encoded by ESR1 gene) is the main driver and prime drug target in luminal breast cancer. ESR1 chromatin binding is extensively studied in cell lines and a limited number of human tumors, using consensi of peaks shared among samples. However, little is known about inter-tumor heterogeneity of ESR1 chromatin action, along with its biological implications. Here, we use a large set of ESR1 ChIP-seq data from 70 ESR1+ breast cancers to explore inter-patient heterogeneity in ESR1 DNA binding to reveal a striking inter-tumor heterogeneity of ESR1 action. Of note, commonly shared ESR1 sites show the highest estrogen-driven enhancer activity and are most engaged in long-range chromatin interactions. In addition, the most commonly shared ESR1-occupied enhancers are enriched for breast cancer risk SNP loci. We experimentally confirm SNVs to impact chromatin binding potential for ESR1 and its pioneer factor FOXA1. Finally, in the TCGA breast cancer cohort, we can confirm these variations to associate with differences in expression for the target gene. Cumulatively, we reveal a natural hierarchy of ESR1-chromatin interactions in breast cancers within a highly heterogeneous inter-tumor ESR1 landscape, with the most common shared regions being most active and affected by germline functional risk SNPs for breast cancer development.

Histone acetylation dynamics modulates chromatin conformation and allele-specific interactions at oncogenic loci.

In cancer cells, enhancer hijacking mediated by chromosomal alterations and/or increased deposition of acetylated histone H3 lysine 27 (H3K27ac) can support oncogene expression. However, how the chromatin conformation of enhancer-promoter interactions is affected by these events is unclear. In the present study, by comparing chromatin structure and H3K27ac levels in normal and lymphoma B cells, we show that enhancer-promoter-interacting regions assume different conformations according to the local abundance of H3K27ac. Genetic or pharmacological depletion of H3K27ac decreases the frequency and the spreading of these interactions, altering oncogene expression. Moreover, enhancer hijacking mediated by chromosomal translocations influences the epigenetic status of the regions flanking the breakpoint, prompting the formation of distinct intrachromosomal interactions in the two homologous chromosomes. These interactions are accompanied by allele-specific gene expression changes. Overall, our work indicates that H3K27ac dynamics modulates interaction frequency between regulatory regions and can lead to allele-specific chromatin configurations to sustain oncogene expression.

Endonuclease FEN1 Coregulates ERα Activity and Provides a Novel Drug Interface in Tamoxifen-Resistant Breast Cancer.

Estrogen receptor α (ERα) is a key transcriptional regulator in the majority of breast cancers. ERα-positive patients are frequently treated with tamoxifen, but resistance is common. In this study, we refined a previously identified 111-gene outcome prediction-classifier, revealing FEN1 as the strongest determining factor in ERα-positive patient prognostication. FEN1 levels were predictive of outcome in tamoxifen-treated patients, and FEN1 played a causal role in ERα-driven cell growth. FEN1 impacted the transcriptional activity of ERα by facilitating coactivator recruitment to the ERα transcriptional complex. FEN1 blockade induced proteasome-mediated degradation of activated ERα, resulting in loss of ERα-driven gene expression and eradicated tumor cell proliferation. Finally, a high-throughput 465,195 compound screen identified a novel FEN1 inhibitor, which effectively blocked ERα function and inhibited proliferation of tamoxifen-resistant cell lines as well as ex vivo-cultured ERα-positive breast tumors. Collectively, these results provide therapeutic proof of principle for FEN1 blockade in tamoxifen-resistant breast cancer. SIGNIFICANCE: These findings show that pharmacologic inhibition of FEN1, which is predictive of outcome in tamoxifen-treated patients, effectively blocks ERα function and inhibits proliferation of tamoxifen-resistant tumor cells.

EZH2 oncogenic mutations drive epigenetic, transcriptional, and structural changes within chromatin domains.

Chromatin is organized into topologically associating domains (TADs) enriched in distinct histone marks. In cancer, gain-of-function mutations in the gene encoding the enhancer of zeste homolog 2 protein (EZH2) lead to a genome-wide increase in histone-3 Lys27 trimethylation (H3K27me3) associated with transcriptional repression. However, the effects of these epigenetic changes on the structure and function of chromatin domains have not been explored. Here, we found a functional interplay between TADs and epigenetic and transcriptional changes mediated by mutated EZH2. Altered EZH2 (p.Tyr646* (EZH2Y646X)) led to silencing of entire domains, synergistically inactivating multiple tumor suppressors. Intra-TAD gene silencing was coupled with changes of interactions between gene promoter regions. Notably, gene expression and chromatin interactions were restored by pharmacological inhibition of EZH2Y646X. Our results indicate that EZH2Y646X alters the topology and function of chromatin domains to promote synergistic oncogenic programs.