抗人MSLN单克隆抗体的构建及体外活性表征  

Preparation and in vitro Bioactivity Characterization of Recombinant Humanized Anti-MSLN Monoclonal Antibody

作　　者：陆颖 朱建伟[1] LU Ying;ZHU Jianwei(gineering Research Center of Cell&Therapeutic Antibody(MOE),School of Pharmacy,Shanghai Jiao Tong University,Shanghai 20)

机构地区：[1]上海交通大学药学院,细胞工程与抗体药物教育部工程研究中心,上海200240

出　　处：《中国医药工业杂志》2024年第6期804-810,共7页Chinese Journal of Pharmaceuticals

基　　金：国家自然科学基金青年项目(82003703)。

摘　　要：由于间皮素(MSLN)在多种实体瘤中广泛表达而在正常间皮细胞上表达较低,可作为实体瘤治疗的重要靶点。研究旨在通过将具有较高亲和力的重组抗人MSLN IgG1型单克隆抗体(rhMSLN mAb)序列构建到哺乳动物细胞表达载体pcDNA3.4中,采用Expi293F细胞系高效表达重组单克隆抗体rhMSLN mAb,并采用表面等离子体共振和流式细胞术对其结合活性进行初步探究,采用乳酸脱氢酶(LDH)释放法鉴定其介导的抗体依赖的细胞毒作用(ADCC)。研究结果表明,rhMSLN mAb在Expi293F细胞中以特定条件进行瞬时表达的产量可达70.2 mg/L,与MSLN重组蛋白的亲和力为7.02 nmol/L,对人胃癌细胞(NCI-N87细胞)ADCC作用的EC50为2.48×10–3μg/mL,为靶向MSLN生物技术药物开发及生产提供了前 期探索性支持。Mesothelin(MSLN)could be served as an important target for immunotherapy for its wide expression in a variety of solid tumors and less expression in normal mesothelial cells.In this study,a recombinant humanized IgGl monoclonal antibody(rhMSLN mAb)sequence against MSLN with high affinity was constructed into the mammalian cell expression vector pcDNA3.4,and efficiently expressed by the Expi293F cell line.Then,the afinity of the antibody was assessed by surface plasmon resonance and flow cytometry at the molecular and cellular levels respectively.Lactate dehydrogenase(LDH)release assay was used to identify the antibody-dependent cellular cytotoxicity(ADCC)mediated by the antibody.The results showed that rhMSLN mAb could yield up to 70.2 mg/L under specific conditions by the Expi293F cell line.The affinity to recombinant MSLN was 7.02 nmol/L.The ECso of ADCC against human gastric carcinoma cell line(NCI-N87 cell)was 2.48x10-3μg/mL.This study provided explorative support for the development and production of biopharmaceuticals targeting MSLN.

关 键 词：间皮素 单克隆抗体 Expi293F细胞 抗体依赖的细胞毒作用 

分 类 号：Q816[生物学—生物工程]