Apoptosis Modulation in Prodrug Activation Gene Therapy

描述（由申请人提供）：旁观者细胞毒性效应，定义为将肿瘤内局部产生的活化前药的杀伤作用扩展到周围的肿瘤细胞，是癌症基因导向酶前药治疗（GDEPT）的一个关键特征，因为当前基因传递技术的局限性只允许一小部分肿瘤细胞被治疗性基因转导。Waxman博士的实验室最近进行的研究表明，在单层培养系统中，通过将抗凋亡因子p35引入大鼠胶质肉瘤9L细胞中，基于细胞色素p450的GDEPT与抗癌前药环磷酰胺联合使用，可以显著增强旁观者效应。本提案的主要目标是：1)研究杆状病毒半胱氨酸酶抑制剂p35在增强肿瘤化疗药物环磷酰胺在基于细胞色素p450的GDEPT临床前模型中的旁观者杀伤作用；2)使用复制病毒辅助系统在人类肿瘤异种移植模型中实施这种基因治疗。3)确定p35在P450 GDEPT系统中增强的旁观者活性是否可以广泛应用于其他GDEPT系统，如单纯疱疹病毒胸苷激酶与更昔洛韦联用和大肠杆菌胞嘧啶脱氨酶与5-氟胞嘧啶联用，两者通过不同的机制发挥其肿瘤杀伤和/或旁观者作用。在这个建议下获得的发现可能为在临床上实施这种基因治疗策略铺平道路提供有价值的信息。

DESCRIPTION (provided by applicant): The bystander cytotoxic effect, defined as the extension of the killing effects of an activated prodrug produced locally within a tumor to surrounding tumor cells, is a key feature of gene-directed enzyme prodrug therapy (GDEPT) for cancer, given that the limitation of current technologies for gene delivery allows for only a small percentage of tumor cells to be transduced with a therapeutic gene. Recent studies conducted in Dr. Waxman's laboratory suggest that a significantly enhanced bystander effect can be achieved for cytochrome P450-based GDEPT in combination with the anti-cancer prodrug cyclophosphamide by introduction of an anti-apoptotic factor, p35, into rat gliosarcoma 9L cells in a monolayer culture system. The major goals of this proposal are 1) to investigate the utility of a baculoviral caspase inhibitor, p35, in augmenting the bystander killing effect of the cancer chemotherapeutic drug cyclophosphamide in a preclinical model of cytochrome P450-based GDEPT, 2) to implement this gene therapy in a human tumor xenograft model using a replicating viral helper system, and 3) to determine whether the enhanced bystander activity seen with p35 in the P450 GDEPT system can be broadly applied to other GDEPT systems, such as herpes simplex virus thymidine kinase in combination with ganciclovir and E. coli cytosine deaminase in combination with 5-flurocytosine, both of which exert their tumor killing and/or bystander effect via distinct mechanisms. Findings obtained under this proposal may provide valuable information for paving the way of implementing this gene therapeutic strategy in the clinic.