TRPM4 contributes to the formation and outgrowth of Dukes’ stage C colorectal cancer tumor spheroids

Colorectal cancer (CRC) is a leading cause of cancer-related deaths, with disease progression and distant metastasis being the major determinants of mortality. The ion channel transient receptor potential melastatin-4 (TRPM4) has been associated with several types of cancer and has been shown to regulate key cellular functions. This study investigates the role of TRPM4 in two CRC cell lines of Dukes’ stage C, DLD-1 and LoVo, using 2D and 3D models. CRISPR/Cas9-mediated knockout (KO) of TRPM4 led to a complete loss of Ca 2+ -activated non-selective cation channel current (I CAN ) demonstrating that in Dukes’ stage C CRC cells I CAN is exclusively mediated by TRPM4. TRPM4 deficiency in LoVo and in DLD-1 cells increased store-operated Ca 2+ signaling. Loss of TRPM4 resulted in unchanged proliferation, reduced metabolic activity, and migration in DLD-1 cells. LoVo TRPM4 KO cells presented decreased metabolic activity but showed increased proliferation and increased migration, that was associated with higher expression levels of fibronectin 1 (FN1). In 3D models, TRPM4 KO consistently reduced tumor spheroid formation and outgrowth in both cell lines. Gene expression analysis revealed differential regulation of genes related to WNT/β-catenin signaling, epithelial-to-mesenchymal transition (EMT), and cell adhesion upon TRPM4 KO. These findings suggest that TRPM4 supports tumor growth and dissemination in CRC. However, TRPM4 differentially affects expression levels of cancer-related genes, emphasizing the need for personalized approaches when considering TRPM4 as a therapeutic target in CRC.