分泌表达的抗ErbB2单链抗体与重构型人半胱氨酸蛋白水解酶3融合蛋白对SKBr3细胞的靶向杀伤作用  

Targeted tumor supprssion by a secreted fusion protein consisting of anti-erbB2 antibody and reversed caspase-3 to SKBr3 cells

作　　者：张立红[1] 贾林涛[1] 于翠娟[1] 曲萍[2] 董海龙[2] 赵晶[1] 许彦鸣[1] 王成济[1] 杨安钢[1] 

机构地区：[1]第四军医大学生物化学与分子生物学教研室,西安710032 [2]第四军医大学病理学教研室,西安710032

出　　处：《中华医学杂志》2003年第7期564-568,共5页National Medical Journal of China

基　　金：国家"八六三"高技术研究发展计划基金资助项目( 2 0 0 1AA2 1710 1) ;国家杰出青年科学基金资助项目 ( 3 99Z5 0 3 6) ;军队杰出中青年人才研究基金资助项目 ( 98J0 0 9)

摘　　要：目的　探讨分泌表达的抗erbB2单链抗体与重构型人半胱氨酸蛋白水解酶 (caspase 3)融合蛋白对ErbB2抗原阳性肿瘤细胞的靶向杀伤作用。方法　将重构型人caspase 3基因亚克隆入pCMV e2 3scFv PEII PEIII相应位点 ,构建表达分泌型的抗erbB2单链抗体与重构型人caspase 3融合蛋白基因的真核表达载体 pCMV e2 3scFv PEII revcasp 3,转染人T淋巴瘤细胞系Jurkat,筛选并建系 ,ELISA检测培养上清中融合蛋白的分泌表达以及对人乳腺癌细胞SKBr3生长的抑制作用 ,并将重组质粒经脂质体包裹后肌肉注射荷瘤裸鼠研究其对肿瘤的抑制作用 ,间接免疫荧光检测重构型人caspase 3蛋白在瘤体的靶向分布。结果　融合蛋白基因可通过Jurkat细胞分泌表达并杀伤SKBr3细胞 ,肌肉注射重组质粒明显延长荷瘤裸鼠生存时间 (延长 72 % )和抑制荷瘤裸鼠肿瘤生长 (抑制率为77% )。间接免疫荧光染色显示重构型人Caspase 3融合蛋白具有靶向分布。结论　分泌表达的抗erbB2单链抗体与重构型人Caspase 3融合蛋白靶向诱导SKBr3细胞死亡。Objective To investigate the targeted killing effect to SKBr3 cells due to the expression of a secreted fusion protein consisting of anti-erbB2 antibody and reversed caspase-3. Methods A recombinant plasmid pCMV-e23scFv-PEII-revcasp 3 was constructed by subcloning reversed caspase-3 gene to the downstream of anti-erbB2 antibody and transfected into Jurkat cells. The cell lines which secreted expressing fusion protein stably were selected. The fusion protein in media was detected by ELISA and the media was used to culture human breast cancer SKBr3 cells. The recombinant plasmids with liposomes was administrated to BALB/C nude mouses bearing SKBr3 tumor by intramuscular injection. The targetting effect of the recombinant fusion protein caspase-3 was detected by indirect immunofluorescence staining. Results Fusion protein can be expressed and secreted by Jurkat cells stably and kill SKBr3 cells. Significant prolonged survival time (prolonged by 72%) and inhibition of tumor growth in vivo (withan inhibition ratio of 77%) were seen in the group administered with recombinant plasmids. Indirect immunofluorescence staining showsed that the recombinant fusion protein caspase-3 has targetting effect. Conclusion Secreted expression of the fusion protein consisting of anti-erbB2 antibody and reversed caspase-3 can targetedly induce SKBr3 cells to death.

关 键 词：分泌表达 抗ErbB2单链抗体 融合蛋白 SKBr3细胞 靶向杀伤作用 CASPASE-3 肿瘤 基因疗法 

分 类 号：R346[医药卫生—基础医学]