聚乙二醇修饰鼠抗人CD3单克隆抗体的研究  

Modification of monoclonal antibody anti-CD3 with mPEG

作　　者：马杭圣[1] 王鸿[1] 孙汉栋 易喻[1] 应国清[1] 

机构地区：[1]浙江工业大学药学院,浙江杭州310032 [2]杭州九源基因工程有限公司,浙江杭州310018

出　　处：《浙江工业大学学报》2008年第6期633-637,共5页Journal of Zhejiang University of Technology

基　　金：浙江省重大科技攻关资助项目(2005C13025)

摘　　要：确定一条聚乙二醇化鼠抗人CD3单克隆抗体和抗体片段的制备工艺路线.以胃蛋白酶酶解并通过凝胶柱分离和纯化得到抗体Fab′,利用PEG-SPA修饰抗体和抗体片段Fab′上的氨基,通过体内和体外T细胞增殖实验,考查修饰后的抗体和抗体片段的免疫活性.较佳的修饰工艺条件为:在pH9.0硼酸缓冲液,反应温度25℃,抗体浓度0.2 mg/mL,抗体和聚乙二醇摩尔比为1∶20,反应3 h.修饰后抗CD3单抗全抗,对T淋巴细胞增殖抑制活性下降了32.28%,抗体Fab′下降了7.96%,PEG化Fab′下降了19.92%.利用聚乙二醇修饰抗体和抗体片段,提高了抗体半衰期,且无细胞毒性,活性损失较小,该技术具有较高的可行性,可极大的提高单克隆抗体尤其是非人源性抗体在临床上的应用.An available technical process of modification of monoclonal antibody of mouse antihuman CD3 and its segment with mPEG were determined. After degradation by pepsin, antibody fragment Fab′ was seprated by gel column and purified by ultrafiltration. And antibody and fragment Fab′ were then modified by PEG-SPA. The immune activity of modified antibody and fragment Fab′ were tested by in vivo and in vitro studies on T cell proliferation. The optimal modification conditions were as following： in the borate buffer （pH 9.0, 25 ℃）, the concentration of the antibody was 0.2 mg/mL, the molar ratio of antibody to mPEG was 1 ： 20, reacted for 3 h. The results showed that the inhibitory activity of modified anti-CD3 monoclonal antibody dropped 32.28%, and that of PEG-modified Fab′ dropped 19.92% while original Fab′ dropped 7. 96%. It is a feasible method to modify monoclonal antibody with mPEG, the half life of the antibody increased with no cytotoxicity and lower activity loss, which greatly improve the clinic application of monoclonal antibody especially non-human type to a great extend.

关 键 词：抗CD3单克隆抗体 聚乙二醇 化学修饰 免疫活性 

分 类 号：R392[医药卫生—免疫学]