LAUSR

Transient receptor potential melastatin 4 (TRPM4) is a broadly expressed Ca2+ activated monovalent cation channel that contributes to the pathophysiology of several diseases. For this study, we generated stable CRISPR/Cas9 TRPM4 knockout (K.O.) cells from the human prostate cancer cell line DU145 and analyzed the cells for changes in cancer hallmark functions. Both TRPM4-K.O. clones demonstrated lower proliferation and viability rates compared to the parental cells. Migration was also impaired in the TRPM4-K.O. cells. Additionally, analysis of 210 prostate cancer patient tissues demonstrates a positive association between TRPM4 and local/metastatic progression. Moreover, a decreased adhesion rate was detected in the two K.O. clones compared to DU145 cells. Next, we tested three novel TRPM4 inhibitors with whole-cell patch clamp technique for their potential to block TRPM4 current. CBA, NBA and LBA demonstrated strong inhibitory effects on TRPM4 in prostate cancer cells. However, none of these inhibitors demonstrated any TRPM4-specific effect in the cellular assays. To evaluate if the observed effect on migration, viability, and cell cycle is linked to TRPM4 conductivity, we transfected TRPM4-K.O. cells with either TRPM4 wild-type or a dominant-negative mutant, non-permeable to Na+. Our data showed a partial rescue of the viability of cells expressing functional TRPM4, while the pore mutant was not able to rescue this phenotype. For the rescue of cell cycle, TRPM4 ion conductivity was not essential. In conclusion, TRPM4 current partially contributes to viability, migration, cell cycle shift, and adhesion; however, blocking the current is insufficient to prevent its participation in cancer hallmark functions.