重组腺病毒驱动KDR-CDglyTK融合基因系统对MCF-7细胞及血管内皮细胞的靶向杀伤作用

南方医科大学学报 ›› 2004, Vol. 24 ›› Issue (12): 1346-1349,1358.

重组腺病毒驱动KDR-CDglyTK融合基因系统对MCF-7细胞及血管内皮细胞的靶向杀伤作用

苏国强¹, 黄宗海¹, 刘志锋², 郭爱华², 俞金龙¹, 厉周¹, 宋彗娟¹

1. 南方医科大学珠江医院, 广东, 广州, 510282;
2. 南方医科大学病理生理教研室, 广东, 广州, 510515

出版日期:2004-12-20 发布日期:2004-12-20
基金资助:
收稿日期:2004-4-28。
基金项目:国家863计划项目(2001AA217171);广东省自然科学基金项目(013072);
作者简介:苏国强(1966- ),男,南方医科大学在读博士研究生,副主任医师,电话:020-61643211,E-mail:suguoqiang66@yahoo.com.cn

Adenovirus-mediated CDglyTK fusion gene system driven by KDR promoter selectively kills MCF-7 breast cancer cells and vascular endothelial cells

SU Guo-qiang¹, HUANG Zong-hai¹, LIU Zhi-feng², GUO Ai-hua², YU Jin-long¹, LI Zhou¹, SONG Hui-juan¹

1. 南方医科大学珠江医院, 广东, 广州, 510282;
2. 南方医科大学病理生理教研室, 广东, 广州, 510515

Online:2004-12-20 Published:2004-12-20

摘要/Abstract

摘要： 目的探讨腺病毒介导血管内皮生长因子受体(KDR)启动子驱动的CDglyTK融合基因体系对人乳腺癌细胞株MCF-7及血管内皮细胞(ECV304)选择性杀伤作用。方法将质粒pAdEasy-KDR-CDglyTK在293细胞内包装、扩增后,体外感染表达KDR的MCF-7、ECV304细胞株和对照组不表达KDR的LS174T细胞株,并给予不同浓度的前药5-FC和/或GCV,观察该体系对不同细胞株的杀伤效应及其旁观者效应。并通过流式细胞仪检测前药对瘤细胞周期的影响。结果所得病毒滴度为2.0×10¹²pfu/ml。3种细胞的感染率相似,其感染率随腺病毒滴度的递增而增加,当感染复数(MOI)为200时,所有细胞株均达约100%感染。以MOI为100的重组体分别感染各细胞株表现出对前药的不同敏感性:表达KDR的MCF-7细胞和ECV304细胞对前药具有较高的敏感性,二者敏感性差异无显著性意义(P=1.00);与前2者相比,不表达KDR的LS174T细胞对前药不敏感(P<0.001)。CDglyTK融合基因的疗效优于任一单自杀基因(P<0.001)。将感染腺病毒的细胞与未感染细胞以不同混合培养,观察到该体系明显的旁观者效应。流式细胞术检测表明该体系抑制MCF-7细胞DNA的合成,表现为G₁期细胞比率增多及S期细胞减少(P<0.001)。结论 KDR基因启动子调控的CDglyTK融合基因体系可选择性杀伤乳腺癌MCF-7细胞及血管内皮细胞。

Abstract: Objective To observe the selective killing of MCF-7 human breast cancer cells and vascular endothelial ECV304 cells by adenovirus (Ad)-mediated double suicide gene driven by KDR promoter. Methods The plasmid pAdEasy-KDR- CDglyTK was transfected into 293 packaging cells for amplification of the infectious Ad, which were then used to infect KDR-producing ECV304 and MCF-7 cells and LS174T cells that did not produce KDR. The infected cells were treated with 5-FC and/or GCV at different doses to observe the cell-killing and bystander effects of AdKDR-CdglyTK. The cell cycle changes were also detected by flow cytometry. Results The Ad at the titer of 2.0×10¹²pfu/ml was obtained after the amplification, whose infection rates of the cells were similar, but could be increased gradually with the multiplicity of infection (MOI) till reaching 100% with the MOI of 200. The infected cells exhibited different sensitivities to the two prodrugs: ECV304 cells and MCF-7 cells infected with Ad-KDR-CDglyTK showed similar high sensitivity to the prodrugs (P=1.00), whereas the infected LS174T cells appeared to be less sensitive (P<0.001). The killing effect of CD/TK fusion gene on the target cells was much stranger than that of either suicide gene (P<0.001), but all exhbiting considerable bystander effect. In addition, the cell cycle of MCF-7 cells was arrested at G₁ phase. Conclusions CD/TK fusion gene system driven by KDR promoter can selectively kill KDR-expressing vascular endothelial cells and MCF-7 cells.

中图分类号:

R730.59

苏国强¹, 黄宗海¹, 刘志锋², 郭爱华², 俞金龙¹, 厉周¹, 宋彗娟¹. 重组腺病毒驱动KDR-CDglyTK融合基因系统对MCF-7细胞及血管内皮细胞的靶向杀伤作用[J]. 南方医科大学学报, 2004, 24(12): 1346-1349,1358.

SU Guo-qiang¹, HUANG Zong-hai¹, LIU Zhi-feng², GUO Ai-hua², YU Jin-long¹, LI Zhou¹, SONG Hui-juan¹. Adenovirus-mediated CDglyTK fusion gene system driven by KDR promoter selectively kills MCF-7 breast cancer cells and vascular endothelial cells[J]. Journal of Southern Medical University, 2004, 24(12): 1346-1349,1358.

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