单抗401与AQP4相互作用分子机制及其对NMO疾病模型治疗效果的研究

视神经脊髓炎（NMO）是一种高致盲率的中枢神经系统脱髓鞘疾病，约50%的NMO患者首发症状为视神经炎，临床无特异性治疗手段。NMO的发病机制为B细胞产生的AQP4-IgG与星形胶质细胞终足上的AQP4蛋白相互作用，介导星形胶质细胞的死亡，引发脱髓鞘病变，最终导致NMO发病。申请人团队研究发现AQP4蛋白胞外结构域3个Loop区中的Loop C对于AQP4-IgG与AQP4结合至关重要，同时研发了特异性全片段保护Loop C的单抗401，但401与AQP4之间相互作用的分子机制，对髓鞘保护效果如何依然未知。基于前期研究，本项目拟先对401进行体外重组表达纯化，体外重组表达纯化人源AQP4蛋白，将401和AQP4共结晶，进行X光衍射，解析结构，阐明401与AQP4分子相互作用机制；然后建立NMO动物模型，通过行为学评分的方式验证其对髓鞘的保护活性，为401应用于临床治疗打下坚实的研究基础。

Neuromyelitis Optica (NMO) is a demyelinating disease of the central nervoussystem with high incidence of blindness. The pathogenesis of NMO is that, theinteraction between AQP4-IgG produced by B cells and AQP4 protein on the end footof astrocytes mediates the death of astrocytes, inducing demyelination of opticnerves. The applicant's team found that loop C, one of the three loop regions ofthe extracellular domain of AQP4 protein, is crucial for the binding betweenAQP4-IgG and AQP4. The applicant's team also developed monoclonal antibody 401with specific full-segment loop protection. However, the molecular mechanism ofinteraction between 401 and AQP4, and its effect on myelin protection are stillunknown. Based on previous studies and discoveries, the team intends to conductfurther study on 401 and AQP4 in this project to mainly clairify two questions,the interaction mechanism between 401 and AQP4 and the verification of theprotective effect of the interaction between 401 and AQP4. Firstly, the team willconduct in vitro recombinant expression and purification of 401, and in vitrorecombinant expression and purification of human AQP4 protein. Then, theartificial 401 and AQP4 protein will be crystallized together , be X-raydiffracted to solve the structure, to clarify the interaction mechanism between401 and AQP4. Also, the team will establish an NMO animal model to verify theprotective activity between 401 and AQP4 on myelin sheath of by means ofbehavioral score, laying a solid research foundation for the application of 401 inclinical treatment.