单域抗体类GluN2B谷氨酸位点拮抗剂的构建、优化及其抗脑缺血作用研究

虽然脑缺血已成为威胁人类健康的主要疾患之一，但迄今为止，治疗脑缺血的有效药物极为匮乏，寻找抗脑缺血新药意义重大。NMDA受体是一个重要的抗脑缺血靶点，在基于该靶点开发的化合物中，GluN2B选择性拮抗剂是最有希望用于脑缺血治疗的药物，但现有GluN2B拮抗剂均属非竞争性拮抗剂，具有脱靶效应和活性依赖性的缺点，不利于脑缺血的治疗。本项目拟以GluN2B谷氨酸结合结构域为抗原，采用单域抗体构建技术，构建高特异性的、不受受体活化水平影响的、对GluN2B具有竞争性拮抗作用的单域抗体类GluN2B谷氨酸位点拮抗剂及其与Tat组成的融合蛋白，并评价上述两种抗体类拮抗剂的抗脑缺血作用和不良反应。本项目的成功实施将有助于实现具有竞争性拮抗作用的GluN2B谷氨酸位点拮抗剂零的突破，不仅对脑缺血的治疗，而且对兼具竞争性拮抗作用与亚基选择性的NMDA受体拮抗剂的开发，都具有重要的指导意义。

Cerebral ischemia has become one of the main diseases threatening human health, but so far, there is an extreme lack of effective drugs for cerebral ischemia therapy, so it is of great significance to search for new anti-cerebral ischemia drugs. NMDA receptor is an important therapeutic target for cerebral ischemia. Among the compounds developed based on this target, GluN2B selective antagonists are the most promising drugs for the treatment of cerebral ischemia, but the existing GluN2B antagonists all belong to noncompetitive antagonists, which have the shortcomings of off-target effect and activity dependence, and the above two disadvantages are not conducive to the treatment of cerebral ischemia. In this project, GluN2B glutamate binding domain is used as antigen, and single domain antibody construction technology is used to construct anti-GluN2B single domain antibodies as GluN2B glutamate site antagonists. Also, the fusion proteins with Tat will also be constructed. The above antagonists are not affected by the activation level of NMDA receptors and have the characteristics of high specificity and competitive antagonism. The anti-cerebral ischemia effects and adverse effects of the antagonists will be tested. The successful implementation of this project will help to achieve the first kind of GluN2B glutamate site antagonists, which has the characteristic of competitive antagonism. This is important not only for the treatment of cerebral ischemia, but also for the development of NMDA receptor antagonists with the characteristics of competitive antagonism and subunit selectivity.