College of Pharmacy, Yonsei Institute of Pharmaceutical Sciences, Yonsei University, 85 Songdogwahak-ro, Yeonsu-gu, Incheon 21983, Korea.
Int J Mol Sci. 2021 Nov 7;22(21):12037. doi: 10.3390/ijms222112037.
Anoctamin1 (ANO1), a calcium-activated chloride channel, is frequently overexpressed in several cancers, including oral squamous cell carcinoma (OSCC). OSCC is a highly aggressive cancer and the most common oral malignancy. ANO1 has been proposed as a potential candidate for targeted anticancer therapy. In this study, we performed a cell-based screening to identify novel regulators leading to the downregulation of ANO1, and discovered cinobufagin, which downregulated ANO1 expression in oral squamous cell carcinoma CAL-27 cells. ANO1 protein levels were significantly reduced by cinobufagin in a dose-dependent manner with an IC value of ~26 nM. Unlike previous ANO1 inhibitors, short-term (≤10 min) exposure to cinobufagin did not alter ANO1 chloride channel activity and ANO1-dependent intestinal smooth muscle contraction, whereas long-term (24 h) exposure to cinobufagin significantly reduced phosphorylation of STAT3 and mRNA expression of ANO1 in CAL-27 cells. Notably, cinobufagin inhibited cell proliferation of CAL-27 cells expressing high levels of ANO1 more potently than that of ANO1 knockout CAL-27 cells. In addition, cinobufagin significantly reduced cell migration and induced caspase-3 activation and PARP cleavage in CAL-27 cells. These results suggest that downregulation of ANO1 by cinobufagin is a potential mechanism for the anticancer effect of cinobufagin in OSCC.
Anoctamin1（ANO1），一种钙激活氯离子通道，在包括口腔鳞状细胞癌（OSCC）在内的多种癌症中经常过表达。OSCC 是一种高度侵袭性的癌症，也是最常见的口腔恶性肿瘤。ANO1 已被提议作为靶向抗癌治疗的潜在候选药物。在这项研究中，我们进行了基于细胞的筛选，以鉴定导致ANO1 下调的新型调节剂，并发现了华蟾酥毒基，它可下调口腔鳞状细胞癌 CAL-27 细胞中的 ANO1 表达。华蟾酥毒基以剂量依赖性方式显著降低 ANO1 蛋白水平，IC 值约为 26 nM。与先前的 ANO1 抑制剂不同，华蟾酥毒基短期（≤10 分钟）暴露不会改变 ANO1 氯离子通道活性和 ANO1 依赖性肠平滑肌收缩，而长期（24 小时）暴露则显著降低 CAL-27 细胞中 STAT3 的磷酸化和 ANO1 的 mRNA 表达。值得注意的是，华蟾酥毒基抑制表达高水平 ANO1 的 CAL-27 细胞的增殖作用强于 ANO1 敲除的 CAL-27 细胞。此外，华蟾酥毒基可显著减少 CAL-27 细胞的迁移，并诱导 CAL-27 细胞中 caspase-3 的激活和 PARP 的裂解。这些结果表明，华蟾酥毒基下调 ANO1 可能是其在 OSCC 中抗癌作用的潜在机制。
