Investigation of TRPM4 in Prostate Cancer Cells with Novel Small Molecule Inhibitors

Abstract

Transient receptor potential melastatin 4 (TRPM4) is a monovalent cation channel, mainly conducting Na+ and K+. TRPM4 is widely expressed, although its expression is most prominent in prostate and colon. TRPM4 protein and RNA expression levels are increased in prostate cancer compared to normal prostate tissue and TRPM4 was recently identified as a cancer driver gene in androgen-insensitive prostate cancer. TRPM4 is activated by Ca2+ and plays an important role in intracellular Ca2+ signaling through the initiation of an important negative feedback mechanism of store-operated calcium entry (SOCE). As a universal second messenger calcium is regulating multiple biological processes. Therefore, it is important that calcium homeostasis is tightly regulated. However, in several cancers, SOCE and its signaling components are dysregulated, contributing to several cancer hallmarks such as increased proliferation and enhanced migration. Sodium influx via TRPM4 depolarizes the membrane and by that reduces the driving force for further calcium influx. We hypothesize that TRPM4, as negative regulator of SOCE, plays a role in migration and proliferation of prostate cancer cells. We recently successfully evaluated a new TRPM4 blocker, 4-Chloro-2-(2-(2-chlorophenoxy) acetamido) benzoic acid (CBA), as a potent inhibitor of TRPM4 current. CBA blocks endogenous TRPM4 currents in LNCaP cells in a low micromolar range. Furthermore, LNCaP cells treated with CBA showed decreased viability. Additionally, two additional small molecule inhibitors were tested on the same cell line whereof one showed strong negative effect on the viability of LNCaP cells. Furthermore, DU145 cells treated with CBA demonstrated slight decrease in migration compared to vehicle treated cells. Stable TRPM4-K.O. cells were used to further study the role of TRPM4 in prostate cancer migration and proliferation. In line with our previous data using the blockers, DU145 TRPM4-K.O. cells displayed decreased migration and proliferation.