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目的：单纯疱疹病毒Ⅰ型胸苷激酶（ＨＳＶ－ｔｋ）基因治疗恶性胶质瘤体内外试验。方法：分子克隆及真核细胞基因转染技术构建逆转录病毒（ＲＶ）载体ｐＭＶ７（ｔｋ）及ＰＡ３１７ｔｋ包装细胞系；体外不同比例混合鼠Ｃ６胶质瘤细胞与ＰＡ３１７ｔｋ细胞，在ＧＣＶ（Ｇａｎｃｉｃｌｏｖｉｒ）作用下观察细胞存活率；建立ＳＤ大鼠颅内Ｃ６胶质瘤模型（种植５×１０５Ｃ６细胞），治疗组第３天原位注射５×１０６ＰＡ３１７ｔｋ细胞，５天后腹腔给予ＧＣＶ（３０ｍｇ／ｋｇ．ｄ），ＭＲＩ全程监测肿瘤消长，观察病症及存活期，并行病理检查。结果：在ＧＣＶ０．１～１０１μｇ／ｍｌ浓度范围内，Ｃ６细胞存活率随ＰＡ３１７ｔｋ细胞混入比例增加而逐渐减低（Ｐ＜０．００１），并具有ＧＣＶ剂量依赖性（Ｐ＜０．０１）；体内试验治疗组病症轻，生存期延长，ＭＲＩ表明治疗组肿瘤体积较对照组明显减小（Ｐ＜０．０１），１个月时病理检查见肿瘤细胞消失，代之以小胶质细胞增生并形成坏死囊。结论：应用本实验室构建的ＲＶ载体ｐＭＶ７（ｔｋ）及其ＰＡ３１７ｔｋ包装细胞系治疗恶性胶质瘤是一种有效及具有前途的治疗方法。
OBJECTIVE: Herpes simplex virus type I thymidine kinase (HSV-tk) gene was used to treat malignant glioma in vitro and in vivo. METHODS: Molecular cloning and eukaryotic gene transfection techniques were used to construct the retroviral (RV) vectors pMV7(tk) and PA317tk packaging cell lines; different ratios of mixed rat C6 glioma cells and PA317tk cells in vitro were used for GCV (Ganciclovir). The survival rate of the cells was observed. An intracranial C6 glioma model was established in SD rats (planted with 5×105C6 cells). The treatment group was injected orally with 5×106 PA317tk cells on the third day, and GCV was given intraperitoneally 5 days later (30 mg/kg.d). The MRI monitored tumor growth throughout the course of the study, observed the condition and survival, and performed pathological examinations. RESULTS: The survival rate of C6 cells gradually decreased with increasing proportion of PA317tk cells in the range of GCV 0.1-101 μg/ml (P<0.001), and had GCV dose-dependent (P<0.01); The experimental treatment group showed milder disease and longer survival time. MRI showed that the tumor volume in the treatment group was significantly smaller than that in the control group (P<0.01). At 1 month, the pathological examination showed that the tumor cells disappeared and replaced with microglial cell proliferation. Formation of necrotic sac. Conclusion: The use of the RV vector pMV7(tk) and its PA317tk packaging cell line constructed in our laboratory to treat malignant gliomas is an effective and promising treatment method.