BRD8 is a therapeutic vulnerability for overcoming resistance to dual ER/HER2 blockade therapy in HR+/HER2+ breast cancer

Hormone receptor (HR)-positive, HER2-positive breast cancers often develop resistance to endocrine and anti-HER2 therapies due to the heterogeneous expression of estrogen receptor (ER) and HER2 and the crosstalk of these growth-promoting pathways. However, how anti-HER2 agents activate ER and other growth-promoting pathways remains unknown. Single-cell RNA sequencing of BT474 breast cancer cells identified Bromodomain Containing Protein 8 (BRD8), an acetyl-lysine reader protein in the histone acetylase EP400 complex, as a pivotal mediator to activate ER in response to neratinib treatment. BRD8 expression was rapidly induced by various anti-HER2 agents (neratinib, lapatinib, and trastuzumab), and its depletion disrupted the crosstalk between ER and HER2 signaling pathways and rendered HR+/HER2+ cells and tumor organoids more sensitive to anti-HER2 agents. BRD8, ER, and ER target genes are co-induced by neratinib in single-nucleus (sn) RNA sequencing of a patient-derived xenograft (PDX). Concomitantly, SnATAC-seq reveals that the activated genes share open chromatin regions enriched in transcription factors (TF) binding motifs of ER, forkhead box (FOX), and ETS family. Since EP400 enhances H2AZ deposition and acetylation on chromatin, we performed H2AZ and H2AZac ChIP-sequencing in the presence or absence of BRD8 and neratinib treatment. Upon neratinib treatment, BRD8 activates ER, FOX, and ETS target genes through modulating H2AZac deposition and chromatin decompaction. This finding coincides with RNA-sequencing, where BRD8 promotes cell growth in an ER-dependent and independent manner. In line with these findings, patients who responded poorly to the anti-HER2 therapies exhibited higher BRD8 gene signatures. Furthermore, BRD8 knockdown ablates the ER and HER2 signaling crosstalk and re-sensitizes neratinib-resistant HR+/HER2+ cells to neratinib. Together, this work not only explains why ER signaling is activated upon anti-HER2 therapies but also identifies BRD8 as a druggable vulnerability in HR+/HER2+ breast cancer.