具有人鼠交叉结合活性的抗IL-1R3抗体的筛选与鉴定  

Screening and Identification of Antibodies with Cross-binding Activity to Human and Mouse IL-1R3

作　　者：秦慧 郁心蕊 刘娇 古丽赛娜·恰尔谢 崔靖敏 王茜 杜鹏 周春阳[1] QIN Hui;YU Xinrui;LIU Jiao;Gulisaina Qiaerxie;CUI Jingmin;WANG Xi;DU Peng;ZHOU Chunyang(School of Pharmacy,North Sichuan Medical College,Institute of Pharmaceutical Research,North Sichuan Medical College,Nanchong 637000,China;Institute of Biotechnology,Academy of Military Medical Science,Academy of Military Science,Beijing 100071,China;School of Basic Medical Sciences and Life Sciences,Hainan Medical University,Haikou 571158,China;School of Medical Instrumentation,Shenyang Pharmaceutical University,Benxi 117004,China)

机构地区：[1]川北医学院药学院川北医学院药物研究所,南充637000 [2]军事科学院军事医学研究院生物工程研究所,北京100071 [3]海南医学院基础医学与生命科学学院,海口571158 [4]沈阳药科大学医疗器械学院,本溪117004

出　　处：《中国生物工程杂志》2024年第2期14-24,共11页China Biotechnology

摘　　要：目的:IL-1受体辅助蛋白(IL-1R3)是炎症调控的潜在新靶点。筛选获得具有人鼠交叉结合活性的抗IL-1R3抗体,为炎症干预机制探究与新药开发奠定基础。方法:基于对人源和鼠源IL-1R3氨基酸序列与结构的比对,采用交替包被人源和鼠源IL-1R3的策略对噬菌体抗体库进行筛选;将获得的抗体可变区基因克隆到真核表达载体,制备抗体样品;在分子和细胞水平对候选抗体的结合活性与功能活性进行评价;采用重新配对抗体轻链与重链的策略获得新抗体,尝试改善其特性。结果:筛选获得5个具有人鼠交叉结合活性的抗IL-1R3抗体,其中4个对人源和鼠源IL-1R3的亲和力相当,均在10^(-7)mol/L水平;2个溶解度较好的抗体在细胞水平上具有阻断活性;重新配对候选抗体的轻链与重链,获得1个溶解度明显提升且结合活性有所改善的新抗体。结论:优选获得2个具有人鼠交叉结合活性的抗IL-1R3抗体AET1907和4H6L,为后续深入探索奠定了物质基础。Objective:Interleukin-1 receptor accessory protein(IL-1R3)is a potential new target for inflammation regulation.The aim of this study is to obtain anti-IL-1R3 antibodies with cross-binding activity to human and mouse IL-1R3 and to lay the foundation for the new drug development and pharmacodynamic mechanism research of a novel inflammation intervention strategy.Methods:Based on the sequence and structure alignment of human and mouse IL-1R3,a phage-displayed human single-chain(scFv)antibody library was challenged by alternately coating immune-tubes with human or mouse IL-1R3.The variable region genes of the resulting antibodies were cloned into eukaryotic expression vectors to prepare antibodies.The binding activities of the candidate antibodies were determined by ELISA and SPR and their functional activities were evaluated by cell assay.A strategy of re-pairing the light and heavy chains of these antibodies was then followed to obtain new antibodies with improved properties.Results:Five antibodies with cross-binding activity to human and mouse IL-1R3 were identified,four of which showed comparable affinity to both human and mouse IL-1R3,approximately 10^(-7) mol/L.Two candidate antibodies with favorable solubility effectively blocked IL-1R3-mediated activities in the ex vivo functional assay.A new antibody with re-paired light and heavy chains showed improved binding activity and apparently increased solubility.Conclusions:By introducing antibodies AET1907 and 4H6L with comparable human-mouse IL-1R3 cross-binding activity,this study has laid the groundwork for further investigation.

关 键 词：IL-1R3 抗体 噬菌体展示 抗体库 交叉结合活性 溶解度 

分 类 号：Q342.4[生物学—遗传学]