Ca2+激活氯离子通道（CaCCs）在哺乳动物细胞广泛表达，种类多样，具多种重要生理功能。除TMEM16A和B外，其它类型CaCCs分子身份未明。选择性调节剂是研究CaCCs重要的药理工具。申请人前期建立了中草药组分库，利用CaCCs功能测定模型获得了大量活性组分和化合物。本项目拟继续开展：活性化合物的分离；在细胞和组织上应用细胞生物学和电生理手段对化合物的选择性、作用机制和分子药理学进行研究；在腹泻和便秘模型小鼠上对化合物的药理学和毒理学进行分析；结合基因敲除小鼠验证选择性调节TMEM16A的表达和活性对腹泻和便秘的治疗作用。本项目获得的选择性强、亲和力高、体内活性好的CaCCs调节剂不但对于阐明CaCCs在肠道液体分泌和肠动力中的作用机制具有重要的理论意义，为区分不同CaCCs活性及发现新型CaCC提供分子探针，而且还能为以CaCCs为靶标的分泌性腹泻和慢性便秘治疗提供先导化合物。

Abstract: Calcium-activated Cl- channels (CaCCs) are widely expressed in intestinal epithelial and nonepithelial cell types, where they facilitate epithelial fluid secretion and regulate smooth muscle contraction. Apart from TMEM16A and TMEM16B, the molecular identities of other CaCCs remain unclear. Selective CaCCs modulators (including activators and inhibitors) provide powerful pharmacological tools for CaCCs functional characterization and identification of novel CaCCs. Currently there are few selective CaCCs modulators available. Because of the low specificity, chemical instability and/or poor in vivo activity, none of the modulators identified so far was proved to fully meet the needs of CaCCs functional studies in vivo. Previously, we identified numerous active fractions by high throughput screening that could modulate CaCCs chloride channel activities from a library of 40,000 small molecule fractions that generated from 500 most commonly used Chinese herbs. More than forty active compounds from the positive fractions have been identified by further fractionation and structure determination. On the basis of our current work, we propose a four-year project to identify and pharmacologically characterize high-affinity TMEM16A and enterocyte CaCC modulators that can selectively modulate CaCCs chloride channel activities in vivo. The proposed work includes: completion of isolation and structural determination of single compound modulators from the current positive fractions with CaCC modulatory activities; completion of characterization of the selectivity, action mechanisms and molecular pharmacology of the active compounds by molecular biological and electrophysiological methods; completion of primary pharmacological studies of the active compounds on live intestinal tissues and mouse models of secretory diarrhea and chronic idiopathic constipation. The CaCCs modulators identified in this project will provide not only useful probes in characterizing the channel gating mechanisms, physiological and pathological functions of CaCCs, but also lead compounds in the therapy of CaCCs-related intestinal disorders including secretory diarrhea and idiopathic chronic constipation.