Moriuchi S, Oligino T, Krisky D, Marconi P, Fink D, Cohen J, Glorioso J C
Department of Molecular Genetics, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
Cancer Res. 1998 Dec 15;58(24):5731-7.
Past studies have documented the promise of herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) suicide gene therapy as a potential antitumor treatment. HSV-TK converts the pro-drug ganciclovir (GCV) into a toxic nucleotide analogue, the incorporation of which into cellular DNA blocks cell proliferation. In this report, we have examined the hypothesis that the effectiveness of HSV-TK suicide gene therapy can be enhanced by coexpression of the antitumor cytokine human tumor necrosis factor-alpha (TNF-alpha) from the same replication-defective HSV-1 vector. In vitro testing demonstrated that TNF-alpha expression from this vector potentiated the killing of both TNF-alpha-sensitive L929 tumor cells and TNF-alpha-resistant U-87 MG cells in the presence of GCV. Furthermore, treatment of established intradermal L929 tumors in vivo with the TNF-alpha/TK vector and GCV resulted in prolonged animal survival compared with treatment with parental HSV-TK vector in the presence or absence of GCV. Treatment of intracerebral U-87 MG tumors showed a clear benefit of TK therapy, but a significant further increase in survival using the TNF-alpha vector could not be demonstrated. We found that potentiation of cell killing in vitro required intracellular TNF-alpha because purified protein added to the culture medium of cells infected with HSV-TK vector failed to have the same effect. Accordingly, potentiation in vivo should depend on efficient infection, but immunohistochemical analysis indicated that virus administration by U-87 MG intratumoral injection was inadequate, resulting in an estimated <1% infection of all tumor cells. Moreover, the majority of infected tumor cells were localized at the tumor margin. Together, these results suggest that TNF-enhanced tk gene therapy should provide a useful treatment for TNF-alpha-sensitive tumors and perhaps also for TNT-alpha-resistant tumors if vector delivery can be improved to increase the percentage of transduced tumor cells.
过去的研究已证明单纯疱疹病毒1型（HSV-1）胸苷激酶（TK）自杀基因疗法有望成为一种潜在的抗肿瘤治疗方法。HSV-TK将前体药物更昔洛韦（GCV）转化为一种有毒的核苷酸类似物，该类似物掺入细胞DNA会阻止细胞增殖。在本报告中，我们检验了这样一种假设，即通过同一复制缺陷型HSV-1载体共表达抗肿瘤细胞因子人肿瘤坏死因子-α（TNF-α），可增强HSV-TK自杀基因疗法的有效性。体外试验表明，在存在GCV的情况下，该载体表达的TNF-α增强了对TNF-α敏感的L929肿瘤细胞和TNF-α耐药的U-87 MG细胞的杀伤作用。此外，与在有或无GCV情况下用亲本HSV-TK载体治疗相比，用TNF-α/TK载体和GCV治疗体内已形成的皮内L929肿瘤可延长动物存活时间。对脑内U-87 MG肿瘤的治疗显示TK疗法有明显益处，但使用TNF-α载体未能证明存活时间有显著进一步延长。我们发现体外增强细胞杀伤作用需要细胞内TNF-α，因为添加到感染HSV-TK载体的细胞培养基中的纯化蛋白没有相同效果。因此，体内增强作用应取决于有效感染，但免疫组化分析表明，通过U-87 MG瘤内注射给予病毒不足，导致估计所有肿瘤细胞的感染率<1%。此外，大多数被感染的肿瘤细胞位于肿瘤边缘。总之，这些结果表明，如果能改进载体递送以增加转导肿瘤细胞的百分比，TNF增强的tk基因疗法应为TNF-α敏感肿瘤提供一种有用的治疗方法，也许对TNF-α耐药肿瘤也有用。
