GENE THERAPY STRATEGIES TO RADIOSENSITIZE HUMAN TUMOR CELLS

该项目的总体目标是进一步发展基因治疗与放射治疗相结合治疗人类癌症的战略。该项目是该项目前一个供资期具体目标2的直接延伸，并以过去5年支助期间取得的广泛进展和初步数据为基础。待开发的主要策略涉及"基因置换"，即将野生型肿瘤抑制基因转染到缺乏这些基因的肿瘤细胞中。这种类型的某些基因恢复细胞凋亡倾向和/或恢复正常的细胞周期检查点控制，因此，预期具有放射增敏能力。因此，在本文中待检查的gone治疗载体将具有双重作用，因为当作为单一药剂使用时，它们将具有抗肿瘤活性，此外，还具有放射增敏性质。已经产生了一种特定的载体，将进行详细测试：腺病毒介导的mda7（Ad-mda7）。具体而言，我们建议回答以下问题：1）Ad-mda7功能优先放射增敏人肿瘤细胞在体外？细胞将在体外生长和处理，克隆形成存活将是终点。2)Ad-mda7对肿瘤微血管内皮细胞的放射增敏作用在这里，我们建议测试内皮细胞生长在体外放射增敏的mda7蛋白。细胞将在体外生长和处理，克隆形成存活将是终点。3)Ad-mda7对人异种移植瘤的放射增敏作用在上述目的1中显示为放射增敏的人肿瘤细胞系将作为异种移植肿瘤在裸鼠中生长， 体内用Ad-mda7和放射治疗。肿瘤生长延迟将是终点。4)Ad-mda7对肿瘤细胞放射增敏的分子机制是什么？了解这种基因治疗策略所赋予的放射增敏作用的分子基础将使我们能够在未来设计更好的基因治疗载体。因此，在这个目标下，我们建议检查那些特定的分子途径的细胞凋亡，信号转导和DNA修复已知的受这种载体，可以解释放射增敏。通过这项研究获得的知识将有望导致开发新的，更有效的策略，结合基因治疗和放射治疗人类癌症。

The overall goal of this project is to further develop the strategy of gene therapy combined with radiotherapy for the treatment of human cancers. This project is a direct extension of specific aim 2 of the prior funding period of this project and is based on extensive progress and preliminary data generated during the last 5 years of support. The primary strategies to be developed involve "gene replacement", i.e. wild-type tumor suppressor genes will be transfected into tumor cells deficient in these genes. Certain genes of this type restore apoptosis propensity and/or restore normal cell cycle checkpoint control and would, therefore, be expected to have radiosensitizing abilities. Thus, the gone therapy vectors to be examined in this context will have dual effects in that they will have antitumor activity when used as single agents and, in addition, have radiosensitizing properties. A specific vector has already been generated that will be tested in detail: adenoviral-mediated mda7 (Ad-mda7). Specifically we propose to answer the following questions: 1) Does Ad-mda7 function to preferentially radiosensitize human tumor cells in vitro? The cells will be grown and treated in vitro and clonogenic survival will be the endpoint. 2) Does Ad-mda7 function to radiosensitize tumor microvascular endothelial cells? Here, we propose to test endothelial cells growing in vitro for radiosensitization by mda7 protein. Cells will be grown and treated in vitro and clonogenic survival will be the endpoint. 3) Does Ad-mda7 function to radiosensitize human xenograft tumorsin vivo? The human tumor cell lines shown to be radiosensitized in aim 1 above will be grown as xenograft tumors in nude mice and treated with Ad-mda7 and radiation in vivo. Tumor growth delay will be the endpoint. 4) What is the molecular mechanism by which Ad-mda7 radiosensitizes human tumor cells? Understanding the molecular basis for the radiosensitization imparted by this gene therapy strategy will enable us to design better gene therapy vectors in the future. Thus, under this aim, we propose to examine those specific molecular pathways of apoptosis, signal transduction and DNA repair known to be affected by this vector that may explain radiosensitization. Knowledge gained through this research will hopefully lead to the development of new, more-effective strategies for combining gene therapy and radiotherapy for the treatment of humar cancer.