Apoptosis Modulation in Prodrug Activation Gene Therapy

描述(申请人提供)：旁观者细胞毒性效应，定义为肿瘤内局部产生的激活的前药对周围肿瘤细胞的杀伤作用的延伸，鉴于目前基因传递技术的限制，只允许用治疗性基因转导一小部分肿瘤细胞，这是基因导向的酶前体药物疗法(GDEPT)的一个关键特征。最近在Waxman博士的实验室进行的研究表明，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。该建议的主要目的是1)研究杆状病毒caspase抑制剂p35在基于细胞色素P450的GDEPT临床前模型中增强癌症化疗药物环磷酰胺的旁观者杀伤效应的实用性；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系统中发现的增强的旁观者活性是否可广泛应用于其他GDEPT系统，如单纯疱疹病毒胸苷激酶联合更昔洛韦和大肠杆菌胞嘧啶脱氨酶与5-氟胞嘧啶联合应用，这两种系统都通过不同的机制发挥其杀瘤和/或旁观者效应。根据这一建议获得的结果可能为在临床上实施这一基因治疗策略提供有价值的信息。

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.