Despite advances in the diagnosis and therapy of breast cancer, issues continue to drive morbidity and mortality, especially tumor cells that persist after chemotherapy and contribute to recurrence and metastasis. The presence of such persister cells is particularly relevant for triple negative breast cancer (TNBC) because it harbors a relatively high frequency of cells with properties of cancer stem cells (CSCs) that resist standard-of-care therapies. My interest is understanding the nature of persister cells in TNBC and identifying novel mechanisms that contribute to persistence and stemness that can be exploited to improve therapy. In my first project, I identified a calcium channel TRPC6 that is enriched specifically in CSCs, including cells that are quiescent and persist after chemotherapy, and that has a causal role in promoting resistance. The mechanism by which it functions in this capacity involves its ability to regulate integrin α6 mRNA splicing. Specifically, TRPC6-mediated Ca2+ entry represses the epithelial splicing factor epithelial splicing regulatory protein 1 (ESRP1), which enables expression of the integrin α6B splice variant. This integrin splice variant has been implicated in sustaining breast CSCs. TRPC6 and α6B function in tandem to facilitate stemness and persistence by activating TAZ and, consequently, repressing Myc. Therapeutic targeting of TRPC6 using a specific chemical inhibitor sensitizes TNBC cells, organoids, and patient-derived xenografts (PDX) to chemotherapy by impeding the splicing of α6 integrin mRNA and inducing Myc. These data revealed a Ca2+-dependent mechanism of chemotherapy-induced persistence, which is amenable to therapy, that involves integrin mRNA splicing.
In my second project, I extended my investigation of TRPC6 to cell metabolism based on my observation that TRPC6 inhibition sensitized TNBC cell lines and organoids to ferroptosis, a form of cell death that involves iron-dependent lipid peroxidation of cell membranes. This observation piqued my interest because there is evidence that the ability to resist ferroptosis facilitates metastasis. In pursuit of the mechanism by which TRPC6 promotes ferroptosis resistance, I discovered that it maintains a level of reduced glutathione (GSH) that is sufficient to buffer oxidative stress. More specifically, I observed that the TRPC6-driven quiescent population has a low biosynthetic demand that enables GSH levels to be maintained by intracellular cysteine biosynthesis without relying on extracellular uptake through the xCT amino acid transporter. Based on these data, I propose that TRPC6 facilitates metastasis by enabling metastatic cells to resist ferroptosis. In summary, this thesis details how a specific calcium channel (TRPC6) contributes to the function of persister/CSCs in TNBC to promote therapy resistance and metastasis, and that it can be targeted in vivo to improve the therapy of TNBC.