An attractive strategy to overcome multidrug resistance in cancer chemotherapy is to suppress P-glycoprotein (P-gp), which is a pump overproduced in cancer cells to remove cytotoxic drugs from cells. In the present study, a Ca ²⁺ -permeable channel TRPC5 was found to be overproduced together with P-gp in adriamycin-resistant breast cancer cell line MCF-7/ADM. Suppressing TRPC5 activity/expression reduced the P-gp induction and caused a remarkable reversal of adriamycin resistance in MCF-7/ADM. In an athymic nude mouse model of adriamycin-resistant human breast tumor, suppressing TRPC5 decreased the growth of tumor xenografts. Nuclear factor of activated T cells isoform c3 (NFATc3) was the transcriptional factor that links the TRPC5 activity to P-gp production. Together, we demonstrated an essential role of TRPC5–NFATc3–P-gp signaling cascade in P-gp induction in drug-resistant cancer cells.

A critical challenge for chemotherapy is the development of chemoresistance in breast cancer. However, the underlying mechanisms and validated predictors remain unclear. Extracellular vesicles (EVs) have gained attention as potential means for cancer cells to share intracellular contents. In adriamycin-resistant human breast cancer cells (MCF-7/ADM), we analyzed the role of transient receptor potential channel 5 (TrpC5) in EV formation and transfer as well as the diagnostic implications. Up-regulated TrpC5, accumulated in EVs, is responsible for EV formation and trapping of adriamycin (ADM) in EVs. EV-mediated intercellular transfer of TrpC5 allowed recipient cells to acquire TrpC5, consequently stimulating multidrug efflux transporter P-glycoprotein production through a Ca²⁺- and activated T-cells isoform c3-mediated mechanism and thus, conferring chemoresistance on nonresistant cells. TrpC5-containing circulating EVs were detected in nude mice bearing MCF-7/ADM tumor xenografts, and the level was lower after TrpC5–siRNA treatment. In breast cancer patients who underwent chemotherapy, TrpC5 expression in the tumor was significantly higher in patients with progressive or stable disease than in patients with a partial or complete response. TrpC5-containing circulating EVs were found in peripheral blood from patients who underwent chemotherapy but not patients without chemotherapy. Taken together, we found that TrpC5-containing circulating EVs may transfer chemoresistance property to nonchemoresistant recipient cells. It may be worthwhile to further explore the potential of using TrpC5-containing EVs as a diagnostic biomarker for chemoresistant breast cancer. Significance A critical challenge for chemotherapy is development of chemoresistance, but underlying molecular mechanisms remain unclear. In this study, we found that drug-resistant adriamycin-resistant human breast cancer cells possessed numerous transient receptor potential channel 5 (TrpC5) -containing extracellular vesicles (EVs) on the cell surface. Suppressing TrpC5 expression diminished the formation of EVs. Incubation of drug-sensitive recipient cells with EVs endowed recipients with drug-resistant properties. In both human samples and a mouse model of breast cancer, the expression of TrpC5 proteins was high in the tumor, and the levels of TrpC5-positive EVs were high in the circulation. These data suggest a critical role of TrpC5-containing EVs in the transfer of drug resistance. In the future, monitoring TrpC5-containing EVs in the circulation could potentially be used to predict the clinical outcome of chemotherapy.

Adriamycin is a first-line chemotherapy agent against cancer, but the development of resistance has become a major problem. Although autophagy is considered to be an adaptive survival response in response to chemotherapy and may be associated with chemoresistance, its inducer and the underlying molecular mechanisms remain unclear. Here, we demonstrate that adriamycin up-regulates the both levels of TRPC5 and autophagy, and the increase in autophagy is mediated by TRPC5 in breast cancer cells. Blockade of TRPC5 or autophagy increased the sensitivity to chemotherapy in vitro and in vivo. Notably, we revealed a positive correlation between TRPC5 and the autophagy-associated protein LC3 in paired patients with or without anthracycline-taxane-based chemotherapy. Furthermore, pharmacological inhibition and gene-silencing showed that the cytoprotective autophagy mediated by TRPC5 during adriamycin treatment is dependent on the CaMKKβ/AMPKα/mTOR pathway. Moreover, adriamycin-resistant MCF-7/ADM cells maintained a high basal level of autophagy, and silencing of TRPC5 and inhibition of autophagy counteracted the resistance to adriamycin. Thus, our results revealed a novel role of TRPC5 as an inducer of autophagy, and this suggests a novel mechanism of drug resistance in chemotherapy for breast cancer.

The small conductance and intermediate conductance Ca ²⁺ -activated K ⁺ channels are known to be involved in the endothelium-dependent hyperpolarization. Ca ²⁺ entry into endothelial cells stimulates these channels, causing membrane hyperpolarization in endothelial cells and underlying smooth muscle cells. In the present study, with the use of coimmunoprecipitation and double immunolabeling methods, we demonstrated a physical interaction of transient receptor potential vanilloid 4 (TRPV4) with K _Ca 2.3 in rat mesenteric artery endothelial cells. Acetylcholine and 4α-PDD mainly acted through TRPV4-K _Ca 2.3 pathway to induce smooth muscle hyperpolarization and vascular relaxation. K _Ca 3.1 was also involved in the process but at a much lesser degree than that of K _Ca 2.3. Stimulating TRPV4-K _Ca 2.3 signaling pathway also increased local blood flow in mesenteric beds and reduced systemic blood pressure in anesthetized rats. In streptozotocin-induced diabetic rats, the expression levels of TRPV4 and K _Ca 2.3 were reduced, which could be an underlying reason for the dysfunction of endothelium-dependent hyperpolarization in these animals. These results demonstrated an important physiological and pathological role of TRPV4-K _Ca 2.3 signaling pathway in vascular endothelial cells.

Transient receptor potential vanilloid type 4 (TRPV4) is a Ca²⁺-permeable cation channel that is known to be an osmosensor and thermosensor. Currently, limited evidence shows that TRPV4 plays opposite roles in either promoting or inhibiting cancer development in different cancer types. Furthermore, the precise biological functions and the underlying mechanisms of TRPV4 in carcinogenesis are still poorly understood. In this study, we demonstrated that TRPV4 is upregulated in colon cancer and associated with poor prognosis. Contrary to the reported cell death-promoting activity of TRPV4 in certain cancer cells, TRPV4 positively regulates cell survival in human colon cancer in vitro and in vivo. Inhibition of TRPV4 affects the cell cycle progression from the G1 to S phase through modulating the protein expression of D-type cyclins. Apoptosis and autophagy induced by TRPV4 silencing attenuate cell survival and potentiate the anticancer efficacy of chemotherapeutics against colon cancer cells. In addition, PTEN is activated by inhibition of TRPV4 as indicated by the dephosphorylation and increased nuclear localization. Knockdown of PTEN significantly abrogates TRPV4 silencing induced growth inhibition and recovers the capability of clonogenicity, as well as reduced apoptosis in colon cancer cells. Thus, PTEN regulates the antigrowth effects induced by TRPV4 inhibition through both phosphatase-dependent and independent mechanisms. In conclusion, inhibition of TRPV4 suppresses colon cancer development via activation of PTEN pathway. This finding suggests that downregulation of TPRV4 expression or activity would conceivably constitute a novel approach for the treatment of human colon cancer.