RV-HSV-TK/GCV自杀基因系统治疗宫颈癌的实验研究(英文)  

Experi mental study of the RV-HSV-TK/GCVsuicide gene therapy systemin cervix cancer

作　　者：陈道桢[1] 刘璐[2] 薛文群[1] 赵琪[3] 鲁晓萱[2] 樊启英[1] 黄学文 

机构地区：[1]南京医科大学附属无锡妇幼保健院中心实验室,江苏无锡214002 [2]东南大学医学院放射免疫中心,江苏南京210009 [3]无锡市第一人民医院检验科,江苏无锡214042 [4]上海华东疗养院检验科,江苏无锡214042

出　　处：《现代医学》2005年第5期281-285,共5页Modern Medical Journal

基　　金：thesocialdevelopmentprojectofJiangsuprovincescienceandtechnologybureau,China(Proj.No.BS2003614),nationalnatu-ralsciencefund(No.304770500)

摘　　要：目的探讨单纯疱疹病毒胸苷激酶/丙氧鸟苷(HSV-TK/GCV)自杀基因系统对人宫颈癌细胞系Hela体外及体内的杀伤作用及其产生的旁观者效应。方法采用脂质体转染法将GINaTK载体转入包装细胞PA317。取病毒上清液感染人宫颈癌细胞Hela,得到带有HSV-TK基因的Hela/TK细胞,并将其分别用于体外和体内实验。体外实验中,Hela/TK细胞对丙氧鸟苷(GCV)的敏感性和旁观者效应采用四甲基偶氮唑盐(MTT)法测定。动物试验中分别在BALB/C小鼠的右腋窝皮下按不同比例注射Hela/TK细胞和Hela细胞,然后给予GCV治疗。逆转录聚合酶链反应(RT-PCR)检测肿瘤组织中HSV-TK基因的表达情况。结果转染HSV-TK基因的Hela/TK细胞表现出比亲本Hela细胞更强的杀伤肿瘤效应。体外实验结果显示,当Hela/TK细胞数占混合细胞10%时,低浓度(10μg.ml-1)的GCV就可将50%左右的肿瘤细胞杀死。体内实验结果显示,GCV可明显抑制Hela/TK细胞在BALB/C小鼠体内的肿瘤形成。经GCV治疗后,Hela/TK组和混合细胞组肿瘤体积分别较对照组肿瘤体积缩小52.8%和69.4%(均P<0.001)。小鼠的平均生存期也显著延长(P<0.001)。结论逆转录病毒可介导HSV-TK基因转入人宫颈癌细胞Hela并获稳定表达,HSV-TK/GCV自杀基因系统在体内外对宫颈癌细胞均有杀伤作用,且存在明显的旁观者效应。Objective To study in vitro and in vivo killing effect and the bystander effect of HSV-TK/GCV suicide gene system on cervix cancer cell line Hela. Methods GINaTK retroviral vector containing HSV-TK gene was transduced into PA317 packaging cell by lipofectin. Cervix cancer cell line Hela was infected by high titer viral supernatant. Hela/TK cells and Hela cells were used in in vitro and in vivo study. In in vitro study, sensitivity of Hela/TK cells to GCV and bystander effect were observed by MTT test. In in vivo study, Hela/TK cells and Hela cells were injected s.c. into flanking BALB/C mice, GCV was administrated i. p., RT-PCR was applied to detect the expression of HSV-TK gene. Statistical analysis of the data was performed by using the ANOVA（analysis of variance）.Results The Hela cells transfered with GINaTK gene displayed a higher antitumor effect than parent cells. In in vitro study, when the ratio of Hela/TK cells reached 10%, the tumor cell-killing proportion was 50%. In in vivo study, all mice developed tumors in 7 days after tumor cells implanted,the ratio of tumors formation was 100%. GCV could suppress tumor formation of the Hela/TK cells. After mice treated with GCV, the median tumor volume of mice implanted with Hela/TK cells and mice with cells mixed was decreased to 52.8 %（ P ＜ 0.001 ） and 69.4% （ P ＜ 0.001 ） respectively compared with the control tumors. Their median survival was significantly prolonged （ P ＜ 0. 001）. Conclusion The test showed that the HSV-TK gene can be transducted into cervix cancer cell Hela under the mediation of retrovirus and be stable expressed, HSV-TK/GCV suicide gene therapy system could improve the antitumoral efficiency. The bystander effect could be observated in HSV-TK/GCV system in in vitro and in vivo.

关 键 词：宫颈癌 自杀基因治疗 单纯疱疹病毒胸苷激酶基因 旁观者效应 逆转录病毒载体 

分 类 号：Q789[生物学—分子生物学] R737.33[医药卫生—肿瘤]