Estradiol induces bone osteolysis in triple–negative breast cancer via its membrane–associated receptor ERα36

Abstract

Abstract Triple–negative breast cancer (TNBC) is thought to be an estradiol–independent, hormone therapy–resistant cancer because of lack of estrogen receptor alpha 66 (ERα66). We identified a membrane–bound splice variant, ERα36, in TNBC cells that responds to estrogen (E2) and may contribute to bone osteolysis. We demonstrated that the MDA-MB-231 TNBC cell line, which expresses ERα36 similarly to MCF7 cells, is responsive to E2, forming osteolytic tumors in vivo. MDA-MB-231 cells activate osteoclasts in a paracrine manner. Conditioned media (CM) from MDA-MB-231 cells treated with bovine serum albumin–bound E2 (E2-BSA) increased activation of human osteoclast precursor cells; this was blocked by addition of anti–ERα36 antibody to the MDA-MB-231 cultures. Osteoclast activation and bone resorption genes were elevated in RAW 264.7 murine macrophages following treatment with E2-BSA–stimulated MDA-MB-231 CM. E2 and E2-BSA increased phospholipase C (PLC) and protein kinase C (PKC) activity in MDA-MB-231 cells. To examine the role of ERα36 signaling in bone osteolysis in TNBC, we used our bone–cancer interface mouse model in female athymic homozygous Foxn1nu mice. Mice with MDA-MB-231 tumors and treated with tamoxifen (TAM), E2, or TAM/E2 exhibited increased osteolysis, cortical bone breakdown, pathologic fracture, and tumor volume; the combined E2/TAM group also had reduced bone volume. These results suggest that E2 increased osteolytic lesions in TNBC through a membrane–mediated PLC/PKC pathway involving ERα36, which was enhanced by TAM, demonstrating the role of ERα36 and its membrane–associated signaling pathway in bone tumors. This work suggests that ERα36 may be a potential therapeutic target in patients with TNBC.