RNA-Seq of ERa-positive breast cancer cell lines to measure the interplay of fulvestrant treatment and hypoxic conditions

Estrogen receptor alpha (ERa) and hypoxia-inducible factors (HIFs) are key regulators of transcriptional programmes in breast cancer. To investigate their interplay, we performed RNA sequencing on ERa-positive breast cancer cell lines (MCF-7 and T-47D) following treatment with the selective estrogen receptor degrader fulvestrant and/or exposure to hypoxic conditions (1% O2). Cells were cultured for 96 hours in the presence or absence of 100 nM fulvestrant, with the final 48 hours spent under normoxia or hypoxia. RNA was extracted from biological triplicates and sequenced using Illumina short-read technology (=20 million reads/sample). Differential gene expression analysis revealed distinct transcriptional programmes regulated by ERa and hypoxia, as well as a large set of overlapping genes modulated by both conditions. Fulvestrant-induced ERa degradation suppressed canonical estrogen-responsive genes (e.g. GREB1, TFF1) and upregulated EPAS1 (HIF-2a). Hypoxia induced well-characterised HIF targets (e.g. CA9, VEGFA).