腺病毒介导的HSV-tk基因治疗膀胱癌的研究

Adenovirus mediated HSV-tk gene therapy for experimental human bladder cancer

目的观察腺病毒介导的单纯疱疹病毒胸苷激酶基因(HSVtk)联合更昔洛韦(GCV)治疗膀胱癌的效果。方法构建膀胱癌SCaBER移植瘤模型,观察肿瘤注射重组腺病毒(Ad.tk)结合GCV治疗移植瘤的变化;免疫组织化学法(SP法)检测肿瘤内微血管密度(MVD)和增殖细胞核抗原(PCNA)表达;PCR检测肿瘤及脏器内tk基因和腺病毒E1区表达。结果肿瘤内注射5×108pfu的Ad.tk/GCV可使肿瘤缩小或消失,治疗组肿瘤平均重量为0.21g,对照组分别为1.63g(HSVtk/PBS组)、1.69g(PBS/GCV组)、1.72g(PBS组),治疗组与对照组比较差异有显著性(P<0.01)。PCNA染色治疗组肿瘤细胞呈弱阳性(21.43%),对照组呈强阳性,分别为71.44%、65.81%、74.63%,差异有显著性(P<0.01)。MVD在治疗组明显低于对照组(P<0.01)。结论HSVtk/GCV系统治疗膀胱癌效果显著,其作用机制包括干扰DNA合成。

Objective To evaluate the feasibility and efficacy of suicide gene therapy using adnovirus mediated herpes simplex virus thymidine kinase gene combined with ganciclovir as a potential therapeutic approach in murine subcutaneous bladder cancer models.Methods A replication deffective adenovirus vector containing toxic HSV tk gene under the transcriptional control of CMV promoter was used.Subcutaneous bladder cancer was established with 1×10 6 human bladder cancer cells in BALB/c nude mice.Intratumoral injection of Ad.CMV tk (5×10 8 plaque forming units,PFU) in combination with GCV (20 mg/kg body weight every day,i.p.,bid×10 days) was administered in vivo for the determination of treatment efficacy in separately controlled experiments.Results In vivo experiments demonstrated greater inhibition in SCaBER tumor growth for the animals treated with Ad. tk/GCV therapy ( P <0.01).The microvessel density and PCNA were significantly decreased as compared with other control animals revealed by immunohistochemical methods ( P <0.01).The expression of HSV tk gene was determined by PCR.Derect intratumoral injection with Ad.CMV tk followed by GCV also resulted in significantly tumor growth inhibition over the MMC group ( P <0.01).Conclusion Suicide gene therapy using Ad.CMV tk/GCV provides an effective therapy in an experimental murine subcutaneous bladder cancer by significantly inhibiting tumor growth and improving efficacy over MMC treatment,and may become a potential protocol for poor prognosis bladder cancer for clinical use.