Development of the 4th generation calcium antagonists based on molecular revealing the binding sites in calcium channels

基于分子揭示钙通道结合位点开发第四代钙拮抗剂

• 批准号: 08557138
• 负责人: NAKAYAMA Hitoshi
• 金额: $ 3.46万
• 依托单位: Kumamoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08557138/
• 关键词: Calcium antagonist; photoaffinity labeling; drug binding site; structural biology; new compounds; drug design; 光アフィニティラベリング; 心筋Caチャンネル; 精密合成化学; 抗ペプチド抗体; 薬理活性

In this project, we aim to reveal binding sites for calcium antagonists in cardiac calcium channels by photoaffinity labeling and the results are to be applied to design and synthesis of new drugs. New findings are as follows. (1) We identified the dihydropyridine sites in cardiac channel, which were identical to the skeletal muscle counterpart but some amino acids were substituted. The substitution can be interpreted to dominate the difference of binding affinity in two calcium channels. (2) Benzothiazepine binding sites were identified in segment 5 and 6 in repeat IV by photoaffinity labeling using azidobutyryl clentiazem with a rather small-sized photoprobe. The recent report by foreign investigaters where two labeled sites in repeat III and IV must be a wrong conclusion resulted from using a bulky photoprobe with long side chain. (3) We revealed the binding site of a new typed calcium antagonist of 1, 4-benzothiazine structure I which is homologous to benzothiazepine 2 but the identified sites were different. The results can explain the pharmacological difference between two drugs. (4) Three dimentional structures of 1 and 2 are not ovelapped by x-ray crystallographic analysis. It suggests that we will design and synthesis of improved calcium antagonists which are derived from 2 as a lead compound.

在这个项目中，我们旨在通过光性标记揭示心脏钙通道中钙拮抗剂的结合位点，结果将应用于新药的设计和合成。新发现如下。 （1）我们确定了心脏通道中的二氢吡啶位点，这些位点与骨骼肌对应物相同，但取代了一些氨基酸。可以解释该替代是在两个钙通道中主导结合亲和力的差异。 （2）使用偶氮二烯酰胺克伦蒂亚Zem具有相当小的光探针，在重复IV的第5和6节中鉴定了苯并噻唑的结合位点。外国调查员的最新报告是，重复III和IV中两个标记的位点必须是错误的结论，这是由于使用具有长侧链的笨重的光探针而产生的。 （3）我们揭示了与苯锡氮杂2同源的1个4-苯甲噻嗪结构I的新型钙拮抗剂的结合位点，但所鉴定的位点不同。结果可以解释两种药物之间的药理差异。 （4）1和2的三维结构并非通过X射线晶体学分析而被遮盖。这表明我们将设计和合成改进的钙拮抗剂，这些钙拮抗剂源自2作为铅化合物。

项目成果

K.Anzai,et al.: "Effects of hydroxyl radical and sulfhydryl reagents on the open probability of the purified cardiac ryanodine receptor." Biochem.Biophys.Res.Commun.249. 938-942 (1998)

K.Anzai 等人：“羟基自由基和巯基试剂对纯化的心脏兰尼碱受体开放概率的影响”。

H.Nakayama and A.Kuniyasu: "Identification of the binding sites for calcium channel antagonists" Jpn.Heart J.37. 643-650 (1996)

H.Nakayama 和 A.Kuniyasu：“钙通道拮抗剂结合位点的鉴定”Jpn.Heart J.37。

H.Yabana,et al.: "Identification of the benzothiazepine binding sites within the α1 subunt of calcium channel by photpaffinity Labeling" J.Biol.Chem.(印刷中). (1997)

H. Yabana 等人：“通过光亲和标记鉴定钙通道 α1 亚基内的苯并硫氮杂卓结合位点”J. Biol。

A.Kuniyasu,et al.: "A new scorpion toxin which stimulates the Ca^<2+> release channel activity of the skeletal muscle ryanodine receptor." Biochem.J.339(in press). (1999)

A.Kuniyasu,et al.：“一种新的蝎子毒素，可刺激骨骼肌兰尼碱受体的 Ca^2 释放通道活性。”

A.Kuniyasu,et al.: "Identification of the binding region for a new calcium antagonist within the α1 subunit of calcium channels" J.Biol.Chem.(印刷中). (1997)

A. Kuniyasu 等人：“钙通道 α1 亚基内新钙拮抗剂的结合区域的鉴定”J. Biol。（出版中）。