去唾液酸糖蛋白受体单链抗体靶向蜂毒肽的体外抑瘤效应研究  

Expression of Anti-ASGPR ScFv Targeting Melittin and Its Anti-tumor Efficacy in Vitro

作　　者：曹利民[1] 赵晓蓉[1] 叶庆[1] 雷萍[1] 吴砂[1] 代维[1] 朱慧芬[1] 朱荫昌[2] 司进[2] 沈关心[1] 

机构地区：[1]华中科技大学同济医学院免疫学系,武汉430030 [2]江苏省寄生虫病防治研究所,无锡214064

出　　处：《医药导报》2006年第7期639-641,共3页Herald of Medicine

基　　金：国家重点基础研究发展计划基金(基金编号:2002CB513109);卫生部科研基金(基金编号:WKJ2004-2-013)

摘　　要：目的探讨抗去唾液酸糖蛋白受体(ASGPR)单链抗体C1与蜂毒肽(Melittin)重组蛋白C1M靶向抑制肝癌细胞的效果。方法将含C1M/pGC的XL1-Blue转化菌通过IPTG诱导表达。表达产物用Ni2+螯合柱亲和纯化,免疫组织化学技术分析重组蛋白C1M的抗原结合能力,四氮唑盐(MTT)法检测C1M对肝癌细胞株HepG2的增殖抑制作用。结果原核表达质粒C1M/pGC在XL1-Blue中获得有效表达;表达产物以可溶性形式存在;纯化的C1M相对分子量为29.4×103;免疫组化结果表明C1M能有效识别去唾液酸糖蛋白受体,与HepG2(C1M)共培养3d,细胞生长抑制率达92.0%。结论在大肠埃希菌中成功表达C1M,位于C端的Melittin蜂毒肽能有效抑制肿瘤细胞的生长,为体内研究Melittin的靶向抗肝癌效果奠定了基础。Objective To explore the cytolytic efficacy on liver cancercell line HepG2 of the recombinant protein C1M formed by Melittin fusing to an anti-ASGPR single-chain Fv antibody （ C1 ）. Methods A single colony of E. coli XL1-Blue containing plasmid pGC-C1M was inoculated in LB broth, then diluted 1/100 into 1 000 mL LB broth and induced with 1 mmol·L^-1 IPTG. The recombinant C1M was purified with Ni^2＋ chelating HiTrap HP column. Its antigen-binding ability was evaluated with immnunohistochemistry, and growth inhibitor rate of tumor was analysed by MTT. Results SDS-PAGE showed that the purified recombinant protein C1M was expressed in E. coli as soluble style with a molecular weight of about 29.4 × 10^3. The result of immnunohistochemistry showed that C1M could effectively recognized ASGPR. On the third day of culturing HepG2 with C1M,the cellular growth inhibiting rate reached 92.0%. Conclusion Successful expression of C1M was achieved in E. coli,and Melittin at its C end can effectively inhibit the growth of tumor cells.

关 键 词：蜂毒肽 重组免疫毒素 单链抗体 肝癌细胞 

分 类 号：R966[医药卫生—药理学]