Three sesquiterpene lactones suppress lung adenocarcinoma by blocking TMEM16A-mediated Ca2+-activated Cl- channels.

Transmembrane protein TMEM16A, which encodes calcium-activated chloride channel has been implicated in tumorigenesis. Overexpression of TMEM16A is associated with poor prognosis and low overall survival in multiple cancers including lung adenocarcinoma, making it a promising biomarker and therapeutic target. In this study, three structure-related sesquiterpene lactones (mecheliolide, costunolide and dehydrocostus lactone) were extracted from the traditional Chinese medicine Aucklandiae Radix and identified as novel TMEM16A inhibitors with comparable inhibitory effects. Their effects on the proliferation and migration of lung adenocarcinoma cells were examined. Whole-cell patch clamp experiments showed that these sesquiterpene lactones potently inhibited recombinant TMEM16A currents in a concentration-dependent manner. The half-maximal concentration (IC50) values for three tested sesquiterpene lactones were 29.9 ± 1.1 µM, 19.7 ± 0.4 µM, and 24.5 ± 2.1 µM, while the maximal effect (Emax) values were 100.0% ± 2.8%, 85.8% ± 0.9%, and 88.3% ± 4.6%, respectively. These sesquiterpene lactones also significantly inhibited the endogenous TMEM16A currents and proliferation, and migration of LA795 lung cancer cells. These results demonstrate that mecheliolide, costunolide and dehydrocostus lactone are novel TMEM16A inhibitors and potential candidates for lung adenocarcinoma therapy.

Cepharanthine, a novel selective ANO1 inhibitor with potential for lung adenocarcinoma therapy.

Anoctamin-1 (ANO1), also known as transmembrane protein 16A (TMEM16A), is identified as a Ca2+-activated Cl- channel that is expressed in many organs and tissues. It is involved in numerous major physiological functions and especially in tumor growth. By screening 530 natural compounds, we identified cepharanthine as a potent blocker of ANO1 channels with an IC50 of 11.2 ± 0.9 µM and Emax of 92.7 ± 1.7%. The Lys384, Arg535, Thr539, and Glu624 in ANO1 are critical for the inhibitory effect of cepharanthine. Similar to its effect on ANO1, cepharanthine inhibits ANO2, the closest analog of TMEM16A. In contrast, up to 30 µM of cepharanthine showed limited inhibitory effects on recombinant ANO6 and bestrophin-1-encoded Ca2+-activated Cl- currents, but it showed no effects on endogenous volume-regulated anion currents (VRAC). Cepharanthine could also potently suppress endogenous ANO1 currents, significantly inhibit cell proliferation and migration, and induce apoptosis in LA795 lung adenocarcinoma cells. Moreover, animal experiments have shown that cepharanthine can dramatically inhibit the growth of xenograft tumors in mice. The high specificity provided by cepharanthine could be an important foundation for future studies of the physiological role of ANO1 channels, and these findings may reveal a new mechanism of its anticancer effect.

Identification of evodiamine and rutecarpine as novel TMEM16A inhibitors and their inhibitory effects on peristalsis in isolated Guinea-pig ileum.

The transmembrane member 16A (TMEM16A)-encoded Ca2+-activated Cl- channel (CaCC) is expressed in interstitial cells of Cajal (ICCs) and involved in the generation of the slow-wave currents of gastrointestinal (GI) smooth muscles. TMEM16A modulators have been shown to positively or negatively regulate the contraction of gastrointestinal smooth muscle. Therefore, targeting the pharmacological modulation of TMEM16A may represent a novel treatment approach for gastrointestinal dysfunctions such as constipation and diarrhoea. In this study, evodiamine and rutecarpine were extracted from the traditional Chinese medicine Evodia rutaecarpa and identified as novel TMEM16A inhibitors with comparable inhibitory effects. Their effects on intestinal peristalsis were examined. Whole-cell patch clamp results show that evodiamine and rutecarpine inhibited TMEM16A Cl- currents in CHO cells. The half-maximal inhibition values (IC50) of evodiamine and rutecarpine on TMEM16A Cl- currents were 11.8 ± 1.3 µΜ and 9.2 ± 0.4 µM, and the maximal effect values (Emax) were 95.8 ± 5.1% and 99.1 ± 1.6%, respectively. The Lys384, Thr385, and Met524 in TMEM16A are critical for evodiamine and rutecarpine's inhibitory effects. Further functional studies show that both evodiamine and rutecarpine can significantly suppress the peristalsis in isolated guinea-pig ileum. These findings demonstrate that evodiamine and rutecarpine are new TMEM16A inhibitors and support the regulation effect of TMEM16A modulators on gastrointestinal motility.