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目的探讨抗P-糖蛋白(P-gp)/抗CD3微型双功能抗体介导人T细胞杀伤耐药实体瘤细胞的作用。方法采用抗E-tag亲和层析柱分离纯化抗P-gp/抗CD3微型双功能抗体,十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)进行鉴定;采用51Cr释放实验,测定抗P-gp/抗CD3微型双功能抗体介导的人T细胞体外靶向杀伤活性;采用多药耐药(MDR)细胞系KB/MDR、敏感KB细胞裸鼠移植瘤模型,测定该微型双功能抗体介导的体内靶向杀伤活性。结果纯化的抗P-gp/抗CD3微型双功能抗体,在相对分子质量28 000和26 000处各有1条蛋白带。在抗P-gp/抗CD3微型双功能抗体存在的情况下,激活的T淋巴细胞能够裂解KB/MDR细胞,且随着效靶比的增高而增高。抗P-gp/抗CD3微型双功能抗体联合人T淋巴细胞能有效抑制KB/MDR细胞裸鼠移植瘤的生长,而对敏感KB细胞移植瘤的生长无抑制作用。结论抗P-gp/抗CD3微型双功能抗体在体内`外均能介导人T淋巴细胞杀伤表达P-gp抗原的KB/MDR细胞,是有望用于耐药实体瘤临床治疗的双特异性抗体。
Objective To investigate the effect of anti-P-glycoprotein (P-gp) / anti-CD3 micro-bifunctional antibody on human T cell-mediated drug-resistant solid tumor cells. Methods Anti-P-gp / anti-CD3 micro-bifunctional antibody was purified by anti-E-tag affinity chromatography and identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). 51Cr release assay The anti-P-gp / anti-CD3 micro-bifunctional antibody was used to determine the in vitro cytotoxicity of human T cells. KB / MDR cell lines were used to detect the tumor- Mini-bifunctional antibody-mediated targeted killing activity in vivo. Results The purified anti-P-gp / anti-CD3 micro-bifunctional antibody had one protein band at 28 000 and 26 000 relative molecular mass. In the presence of anti-P-gp / anti-CD3 microbifunctional antibody, activated T lymphocytes can lyse KB / MDR cells and increase with increasing target-to-target ratio. Anti-P-gp / anti-CD3 micro-bifunctional antibody combined with human T lymphocytes can effectively inhibit the growth of KB / MDR nude mice xenografts, but have no inhibitory effect on the growth of xenograft KB cells. Conclusion Anti-P-gp / anti-CD3 micro-bifunctional antibodies can mediate the killing of KB / MDR cells expressing P-gp antigen by human T lymphocytes in vitro and in vivo. antibody.