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Development of myosin light chain kinase-inhibitors for anti-secretory and anti-ulcer drugs.

开发用于抗分泌和抗溃疡药物的肌球蛋白轻链激酶抑制剂。

基本信息

批准号：
10557219
负责人：
URUSHIDANI Tetsuro
金额：
$ 3.78万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10557219/
关键词：
myosin light chain kinase gastric secretion calmodulin enzyme inhibitor rabbit cytoskeleton

项目摘要

This work was conducted to search a potential application of myosin light chain kinase inhibitors for anti-ulcer drugs with new mechanism of action. In the present study, we obtained data showing that parietal cell contains a new type of myosin kinase and its new substrate, and they suggested a unique regulation was conducted in the cell. As a candidate substrate for the kinase, we identified a new protein of 100 kDa. This protein was purified, partially sequenced, and suggested to be a rabbit homologue of Mena, a cytoskeletal protein. Mena has been reported to be involved in the actin polymerization as well as interacting with SH3-domain via its proline-rich domain. As the regulation of Mena by phosphorylation has not been studied yet, this work is the first time to postulate this protein as a new drug target. In order to find out a lead compound that inhibit the enzyme, we screened a group of compounds supplied by Kyowa Hakko Kogyo Co. Ltd., as well as the known various types of inhibitors, using isolated rabbit gastric glands and partially purified enzyme. This enzyme did not depend on calcium-calmodulin, showing clear difference from conventional myosin light chain kinases. It is neither sensitive to conventional calmodulin-antagonists nor usual kinase inhibitors. Among the compounds from Kyowa Hakko, some compounds, suggested to be bound to ATP-binding site, showed some inhibitory activity in the purified enzyme. Using isolated glands, two of them showed inhibitory activity on the acid secretion (IC50 was about 10μM). These results are promising that clinically applicable drugs could be developed using these as lead compounds.

本研究旨在寻找肌球蛋白轻链激酶抑制剂作为抗溃疡药物的潜在应用。在本研究中，我们获得的数据表明，壁细胞含有一种新型的肌球蛋白激酶及其新的底物，这表明在细胞中进行了独特的调节。作为激酶的候选底物，我们鉴定了一个100 kDa的新蛋白。这种蛋白质被纯化，部分测序，并建议是一个兔同源的Mena，细胞骨架蛋白。已报道Mena参与肌动蛋白聚合以及通过其富含脯氨酸的结构域与SH 3-结构域相互作用。由于Mena通过磷酸化的调节尚未被研究，这项工作是第一次假设该蛋白质作为新的药物靶点。为了找到抑制该酶的先导化合物，我们筛选了一组由Kyowa Hakko Kogyo Co. Ltd.提供的化合物，以及已知的各种类型的抑制剂，使用分离的兔胃腺和部分纯化的酶。这种酶不依赖于钙-钙调蛋白，与传统的肌球蛋白轻链激酶有明显的不同。它对常规钙调素拮抗剂和常用激酶抑制剂均不敏感。从协和发酵中得到的化合物中，有一些化合物对纯化的酶有一定的抑制活性，这些化合物可能与ATP结合位点结合。使用离体腺体，其中两个显示出对酸分泌的抑制活性（IC 50约为10μM）。这些结果是有希望的，临床上适用的药物可以使用这些作为先导化合物开发。