Sex Hormone Estrone Sulfate Serendipitously Downregulates Intercellular Tunneling Nanotube Conduits, EMT, and Migration in Breast and Pancreatic Cancer Cells

Aim: Estrone (E1) is of particular interest as most breast cancers are diagnosed after menopause, the period in which E1 becomes the dominant estrogen. Triple-negative breast cancer (TNBC) is a highly invasive cancer that lacks hormone receptors for estrogen (ER), progesterone (PR), and HER2 (human epidermal growth factor receptor-2). TNBC management remains clinically challenging despite recent advances in targeted and immunotherapeutic approaches. This study investigated the effects of the estrogen hormone estrone sulfate (E1S) on cancer cells. Here, E1S engages in non-classical biased G protein-coupled estrogen receptor (GPER) signaling in TNBC MDA-MB-231 cells, estrogen receptor-positive MCF-7 breast cancer cells, and pancreatic PANC-1 cancer cells, which express estrogen, progesterone, and androgen receptors. Methods: We performed sialidase and colocalization assays in breast MDA-MB-231 and MCF-7 cells and pancreatic PANC-1 cancer cells. An epithelial-to-mesenchymal (EMT) transition, a tunneling nanotube assay, and a scratch wound assay were conducted to elucidate the cellular behaviors of E1S. We used a RawBlue reporter macrophage cell line to measure NFκB activity using the secreted embryonic alkaline phosphatase (SEAP) assay, and we measured cellular viability and metabolic activity using the AlamarBlue assay. Results: E1S induced NMBR/MMP9/Neu1 signaling in MDA-MB-231, MCF-7, PANC-1 and RawBlue cells. E1S increased E-cadherin expression, reduced N-cadherin and vimentin expression, decreased tunneling nanotube density, and reduced scratch wound migration. E1S increased SEAP activity, suggestive of increased nuclear factor-κB (NF-κB) and activator protein-1 (AP-1)-mediated transcription. E1S treatment did not affect cell viability and metabolic activity in triple-negative MDA-MB-231 breast cells using the AlamarBlue assay in a dose-dependent manner. Conclusions: E1S binding to GPER potentiates a biased heterodimerization with the GPCR neuromedin B receptor (NMBR) to activate matrix metalloproteinase 9 (MMP9) and neuraminidase 1 (Neu1), a signaling paradigm with previously established receptor tyrosine kinase (RTK) and Toll-like (TLR) receptors. Biased, temporal GPER signaling has significant implications for breast and pancreatic cancer cellular behavior, which may be leveraged to inform therapeutic strategies for aggressive forms of breast and pancreatic cancers. This research study is mechanistic in nature and may have potential translational implications.