单链抗体导向的慢病毒介导VP22-TK系统对MUC1^+人卵巢癌裸鼠腹腔移植瘤的作用

Anti-tumor effect of lentivirus-mediated MUCI antibody-targeted gene therapy with VP22-TK system on MUC1^+ human ovarian cancer transplanted intraperitoneally in nude mice

目的 探讨抗MUC1单链抗体 (scFv)导向的慢病毒介导的单纯疱疹病毒结构蛋白VP2 2和胸苷激酶 (TK)融合基因及丙氧鸟苷 (GCV)自杀基因系统对MUC1+ 人卵巢上皮癌的特异靶向性生长抑制作用。方法 将慢病毒包装质粒、包膜质粒、转移载体质粒采用磷酸钙沉淀法共转染包装细胞2 93T ,收集病毒上清 ,并建立MUC1+ 人卵巢上皮癌 (3AO)细胞株腹腔移植瘤模型。单药组分为scFv VP2 2 TK +生理盐水 (NS)组、VP2 2 TK +NS组、NS +NS组 ,分别用慢病毒scFv VP2 2 TK、VP2 2 TK或NS1ml腹腔注射 ,继予NS腹腔注射 ;联合用药组分为scFv VP2 2 TK +GCV组、VP2 2 TK +GCV组、NS +GCV组分别应用慢病毒scFv VP2 2 TK、VP2 2 TK及NS腹腔注射 ,2 4h后给予GCV腹腔注射治疗。每组裸小鼠均为 5只。观察各组裸鼠的生存时间及慢病毒的毒性作用。结果 平均生存时间scFv VP2 2 TK +NS组、VP2 2 TK +NS组、NS +NS组、NS +GCV组、VP2 2 TK +GCV组、scFv VP2 2 TK +GCV组分别为 18.4d± 2 .9d、18.8d± 1.5d ,17.6d± 1.1d ,18.5d± 1.6d ,2 4d± 5d和 4 6d± 2 2d ,6组比较 ,差异有显著意义 (χ2 =2 4 .82 ,P =0 .0 0 2 ) ;并且scFv VP2 2 TK +GCV组较VP2 2 TK +GCV组裸鼠平均生存时间明显延长 (χ2 =7.4 3,P =0 .0 0 6 )。

Objective To investigate the anti tumor effect of anti MUCI single chain variable fragment (ScFv) targeted and lentivirus mediated herpes simplex virus structural proten VP22 and thymidinre kinase (TK) therapy on MUC1 + human ovarian epithelial carcinoma tumor in mice transplanted intraperitoneally. Methods Lentiviruses scFv VP22 TK and VP22 TK were constrted Ten female BABL/c mice were injected intraperitoneally with MUC1+ human ovarian epithelial carcinoma cells line 3AO and then pathological examination was done to those mice that died of tumor. A human ovarian epithelial carcinoma model was established in another 30 female mice and they were randomly divided into 6 goups of 5 mice: NS (normal saline)+NS group (injected intraperitoneally with NS once per day for 3 days and then with NS 24 h after once a day for 5 days), VP22 TK+NS group (injected with herpes simplex virus structural proten VP22 and TK once a day for three days and then with NS 24 h after once a day for 5 days), scFv VP22 TK+NS group (injected with scFv VP22 TK and then NS in the same way), NS+ganciclovir (GCV) group ( injected with NS and then with GCV), VP22 TK+GCV group (injected with VP22 TK and then GCV), and scFv VP22 TK+GCV group (injected with scFv VP22 TK and then GCV). The survival time was observed. Ten female nude mice without injection of tumor cells were injected with scFv VP22 TK or VP22 TK, each for 5 mice; 3 weeks later their abdominal organs were examined to observe the effects of lentivirus on organs. Results All of the first ten mice injected with human ovarian epithelial carcinoma cells died of tumor. The mean survival times of the six experimental groups were 18 4 d±2.9 d, 18 8 d±1.5 d, 17 6 d±1 1 d, 18 5 d±1 6 d, 24 d±5 d, and 46 d±22 d respectively with significant differences between the VP22 TK+GCV group and NS+GCV group (χ 2=6.71, P =0.009), between the scFv VP22 TK+GCV group and NS+GCV group (χ 2=9.7, P =0.002), and between the scFv VP22 TK+GCV group and the VP22 TK+GCV group (χ 2=7.43, P = 0.006). Necrosis and apoptosis could be seen in the tumors in the VP22 TK+GCV group and scFV VP22 TK+GCV group. No toxicity was observed in the mice injected with only scFv VP22 TK or VP22 TK. Conclusion The anti MUCI ScFv targeted and lentivirus mediated herpes simplex virus VP22 and thymidinre kinase (TK) gene therapy has a significant anti tumor effect on MUC1+ human ovarian epithelial carcinoma.