慢病毒介导的双自杀基因对大肠肿瘤细胞的靶向杀伤作用

目的 研究慢病毒介导的KDR启动子驱动CD/TK双自杀基因系统对大肠肿瘤细胞选择性杀伤作用.方法 构建的重组FGW-KDRP-CD/TK慢病毒载体在293T细胞包装扩增后,获得病毒颗粒,体外感染表达KDR的LOVO细胞株和不表达KDR的LS174T细胞株,以RT-PCR方法检测转基因细胞CDglyTK的表达,然后给予不同浓度的前药5-FC及GCV处理,观察该体系对不同细胞株的杀伤效应.结果 重组体对各细胞株的感染率相似,RT-PCR方法检测发现:除LS174T细胞外,LOVO细胞有目的基因CDglyTK的表达.LOVO与LS174T细胞株对前药敏感性有显著性差异(P＜0.001),双自杀基因的疗效优于任一单自杀基因的疗效(P＜0.001).结论 KDR启动子可调控双自杀基因系统选择性杀伤表达KDR的大肠肿瘤细胞.

Targeted killing of colorectal tumor cells by lentiviral constructs containing CD/TK suicide genes and KDR promoter

Objective To investigate the selective killing of colorectal tumor cells by lentivirus-mediated double suicide gene under the regulation of KDR promoter.Methods 293T packaging cells were transfected with the plasmid FGW-KDRP-CD/TK to obtain the infectious viruses.KDR-expressing LoVo cells and LS174T cells that did not produce KDR were transfected with the recombinant virus,and the transfection efficiency was evaluated by the fluorecence microscope.RT-PCR was employed to examine the expression of CDglyTK.After treatment of the cells with 5-FC and GCV,the killing effects on the two cell lines were evaluated.Results The recombinant construct showed similar infection rate of the two cell lines.RT-PCR demonstrated that CDglyTK gene was expressed only in LoVo cells infected with FGW-KDRP-CD/TK but not in LS147T cells,and the sensitivity of the two cell lines to the prodrugs was significantly different(P<0.001).The killing effect of the double suicide gene was much stronger than that of single suicide gene administered(P<0.001).Conclusion The double suicide gene driven by KDR promoter has specific killing effect on the KDR-expressing colorectal tumor cells.