Estrogen Receptor B as a therapeutic target in breast cancer stem cells

Abstract

Background: Breast cancer cells with tumor-initiating capabilities (BSCs) are considered to maintain tumor growth and govern metastasis. Hence, targeting BSCs will be crucial to achieve successful treatment of breast cancer. Methods: We characterized mammospheres derived from more than 40 cancer patients and two breast cancer cell lines for the expression of estrogen receptors (ERs) and stem cell markers. Mammosphere formation and proliferation assays were performed on cells from 19 cancer patients and five healthy individuals after incubation with ER-subtype selective ligands. Transcriptional analysis was performed to identify pathways activated in ERb-stimulated mammospheres and verified using in vitro experiments. Xenograft models (n=4 or 5 per group) were used to study the role of ERs during tumorigenesis. Results: We identified an absence of ERa but upregulation of ERb in BSCs associated with phenotypic stem cell markers and responsible for the proliferative role of estrogens. Knockdown of ERb caused a reduction of mammosphere formation in cell lines and in patient-derived cancer cells (40.7%, 26.8%, and 39.1%, respectively). Gene set enrichment analysis identified glycolysis-related pathways (false discovery rate < 0.001) upregulated in ERb-activated mammospheres. We observed that tamoxifen or fulvestrant alone was insufficient to block proliferation of patient-derived BSCs while this could be accomplished by a selective inhibitor of ERb (PHTPP 53.7% in luminal and 45.5% in triple-negative breast cancers). Furthermore, PHTPP reduced tumor initiation in two patient-derived xenografts (75.9% and 59.1% reduction in tumor volume, respectively) and potentiated tamoxifen-mediated inhibition of tumor growth in MCF7 xenografts. Conclusion: We identify ERb as a mediator of estrogen action in BSCs and a novel target for endocrine therapy.