PROTEASE INHIBITORS AND RADIATION TRANSFORMATION

电离辐射是一种普遍的致癌物质， 人类和动物模型中恶性表型的发展 系统. 细胞体外转化体系的建立 人类和上皮起源将有助于确认和扩展 已经用啮齿动物成纤维细胞得到的结果。 使用immortalized 人表皮角质形成细胞在体外，我们已经证明了诱导 通过DNA肿瘤病毒的协同作用产生恶性表型， 电离辐射 这一进程涉及多个事件。 首先， 收购无限的增长潜力，由于感染 AD 12-SV 40杂合病毒。 剩下的步骤包括连续曝光 电离辐射。 这种辐射诱导的转化导致了 形态学改变，锚定独立生长和形成 低分化的鳞状细胞癌的实验。 这些观察结果表明，永生化的人表皮角质形成细胞 将提供一个良好的体外模型系统，研究辐射 致癌作用及其蛋白酶抑制剂的抑制作用。 因此，委员会认为， 本提案的目的是评估该系统作为一种可重复的， 研究蛋白酶抑制剂在细胞凋亡中作用的体外模型 抑制人上皮系统中的辐射致癌作用。 此外，使用消减cDNA表达文库，鉴定细胞的 基因或原癌基因，这些基因或原癌基因在肿瘤发生过程中被激活或失活。 用蛋白酶抑制剂处理后抑制转化。 最后，我们建议通过二维电泳鉴定 作为内源性蛋白酶底物的蛋白种类， 已经被蛋白酶抑制剂灭活了 这些研究将为蛋白酶的作用提供分子基础 抑制辐射诱发恶性转化的抑制剂 体外培养的人类细胞。 此外，它们还可以提供分子基础， 为有风险的人类受试者开发化学预防疗法。

Ionizing radiation is a universal carcinogen which has been implicated in the development of malignant phenotypes in both human and animal model systems. The development of an in vitro transformation system using cells of human and epithelial origin would be useful for confirming and extending the results already obtained with rodent fibroblasts. Using immortalized human epidermal keratinocytes in vitro, we have demonstrated the induction of a malignant phenotype by the concerted action of a DNA tumor virus and ionizing radiation. this process involved multiple events. First, was the acquisition of unlimited growth potential as a result of infection with the AD12-SV40 hybrid virus. the remaining steps involved successive exposures to ionizing radiation. This radiation-induces transformation resulted in morphological alterations, anchorage-independent growth and the formation of poorly differentiated squamous cell carcinomas in athymic mice. These observations indicate that immortalized human epidermal keratinocytes will provide a good in vitro model system to study both radiation carcinogenesis and its suppression by protease inhibitors. Accordingly, the goal of this proposal is to evaluate this system as a reproducible in vitro model for the study of the role of protease inhibitors in the suppression of radiation carcinogenesis in a human epithelial system. Further, using subtracted cDNA expressions libraries, identify the cellular genes or proto-oncogenes that have been activated or inactivated during the suppression of transformation following treatment with protease inhibitors. Finally, we propose to identify by 2-dimensional electrophoresis the protein species which are the substrates of the endogenous proteases that have been inactivated by the protease inhibitors. The studies will provide a molecular basis for the role of protease inhibitors in the suppression of radiation-induces malignant transformation of human cells in vitro. further, they may provide a molecular basis for the development of chemo-preventative therapies for human subjects at risk.