缺氧复制型溶瘤腺病毒靶向杀伤肝癌栓塞后缺氧肿瘤的实验研究

Tumor necrosis induced by emboliztion is usually imcomplete. Residual hypoxic tumor cells can respond to hypoxia and even progress through hypoxia-inducible factor-1 (HIF-1). Hypoxia replicative oncolytic adenovirus (HYPR-Ad), as a virus selectively toxic towards hypoxic cells, contain a hypoxia response element gene. HYPR-Ad can specifically infect, replicate in and lyse hypoxic/ HIF-active tumor cells, therefore leading to the destruction of the tumor. In this study, we use HYPR-Ad as hypoxic cells killer, and conduct experiments in vitro to evaluate the virus-infected efficiency, selective replication capacity and cytopathic effects in human and animal liver tumor cell lines. We also perform animal experiments using a rabbit liver tumour model to evaluate the therapeutic efficacy and security of HYPR-Ad on hypoxic tumor after embolization, to establish therapeutic regimen, and to clarify the therapeutic mechanism. This study is to introduce HYPR-Ad to interventional therapy for liver cancer and to explore its therapeutic efficacy, protocol and mechanism. Out results could give theoretical and practical evidence for the interventional and targeted gene therapy of liver cancer, and thus contribute to the development of a novel therapeutic strategy for the tumor therapy.

肝癌介入栓塞后肿瘤坏死不彻底，残存的缺氧肿瘤细胞通过缺氧诱导因子-1（HIF-1）对缺氧产生应答继续存活演进。缺氧复制型溶瘤腺病毒（HYPR-Ad）作为一种靶向杀伤缺氧细胞的病毒，携带缺氧反应元件（HRE）基因，可特异性感染缺氧/HIF活化的肿瘤细胞并在其内大量复制，最终导致肿瘤细胞溶解而摧毁肿瘤。本研究旨在采用HYPR-Ad作为缺氧细胞毒剂，在人肝癌和动物肿瘤细胞株中探讨HYPR-Ad对缺氧肿瘤细胞的感染效能、选择性复制能力、细胞溶解及毒性效应；并进一步在肝癌动物模型中评价HYPR-Ad靶向杀伤肝癌栓塞后缺氧肿瘤的作用和安全性，探索其治疗方案，阐明其治疗机理。本研究首次将HYPR-Ad靶向杀伤缺氧肿瘤细胞应用于肝癌介入治疗，探索HYPR-Ad介入靶向治疗肝癌的疗效、治疗方案和治疗机理，将为肝癌介入靶向基因治疗提供新的靶点、方法及实践依据，为肝癌综合介入治疗提供新的策略。

本项目基本按照基金项目申请书计划进度进行。通过体外细胞和体内动物实验证实了介入栓塞后缺氧产生的HIF-1α引起肿瘤组织存活、浸润转移等方面的生物学应答。本研究包装合成了基于HIF-1反应元件（HRE）作为启动子的缺氧复制型溶瘤腺病毒（HYPR-Ad），在肿瘤细胞株和正常肝细胞株上研究HYPR-Ad对缺氧/HIF-1α活化及常氧状态下对宿主细胞的影响。并通过动物实验进一步验证缺氧条件下HYPR-Ad在宿主体内的表达复制下对肝肿瘤的凋亡、坏死的影响，证实了HYPR-Ad通过持续感染介入栓塞后缺氧的肿瘤细胞并在其内大量复制表达蛋白E1A，蛋白E1A通过诱导肿瘤细胞的凋亡、坏死，最终导致肿瘤细胞溶解而摧毁肿瘤。本研究将为在介入栓塞下病毒基于HIF-1α的靶向治疗肿瘤细胞提供理论及实践上的依据，为肝癌的介入治疗和病毒靶向性杀伤的结合治疗提供新的研究方向和治疗手段。