Inhibition of Ca2+-activated chloride channels by the NSAID meclofenamate for anti-diarrhea

Dysfunctional calcium-activated chloride channels (CaCCs) are implicated in many pathological phenotypes and diseases. The CaCC ANO1/TMEM16A robustly expressed in epithelial cells plays an essential role in regulation of Cl^- secretion and intestinal motility. In this study, we investigated the effects of a nonsteroidal anti-inflammatory drug (NSAID) meclofenamate on ANO1 channel and dextran sulfate sodium (DSS)-induced diarrhea in mice. Meclofenamate inhibits CaCC ANO1 channel in a concentration-dependent manner with an IC₅₀ of 16.2 ± 2.7 μM. Meclofenamate also reduces single-channel open probability without altering the channel conductance. Molecular docking and site-directed mutagenesis demonstrate that residues R515, R535 and E654 are important for meclofenamate-mediated ANO1 inhibition. Selectivity evaluation demonstrates that meclofenamate also inhibits Other CaCCs, including ANO2, ANO6 and Bestrophin-1 with IC₅₀ values ranging from approximately 10 to 20 μM. Further in vivo experiments show that meclofenamate dose-dependently reduces intestinal peristalsis and diarrhea induced by DSS in mice. Altogether, our findings reveal a novel role of meclofenamate in inhibiting CaCC currents and alleviating DSS-induced acute diarrhea, thus holding repurposing potential for therapy of diarrhea or gastrointestinal dysfunction.