P53, CHEMICAL CARCINOGEN AND ETHANOL IN ORAL CANCER

有令人信服的证据表明，致癌作用是一个多步骤的过程 并且多种遗传损伤是人类发展癌症所必需的。 在遗传性病变中，p53蛋白功能障碍（由于 p53基因突变或“高危”人乳头瘤病毒感染 [HPV]例如，在一个实施例中，16或18型HPV）是最常见的遗传性 在包括口腔癌的人类癌症中的病症。 尽管有这些 口腔癌细胞中频繁的p53功能障碍，改变p53功能（通过 单独用HPV DNA或突变型p53 cDNA转染）不足以 体外正常人口腔角质形成细胞的肿瘤转化。 因此，p53蛋白功能障碍至少可能是一个早期事件 在口腔癌发生中也是必要的， 其他基因的疾病，将正常细胞转化为肿瘤细胞， 人类口腔 事实上，我们最近的研究表明， 含有可忽略量的野生型（wt）p53蛋白的角质形成细胞 （由于HPV转染）当暴露于 烟草致癌物，但正常的对应物没有。 由于野生型p53蛋白在调节细胞凋亡中起主要作用， 细胞周期停滞，我们假设正常人口腔角质形成细胞 含有野生型p53蛋白修复受损的DNA比口服更有效 具有缺陷性p53功能的角质形成细胞（通过p53基因突变或通过 感染“高危”HPV）。 许多研究表明，细胞 表达野生型p53蛋白的细胞具有建立瞬时延迟的能力 在细胞周期的进展时，暴露于遗传毒性剂。 然而，具有缺陷的p53蛋白的细胞不具有这种能力。 由于细胞周期进程的停滞被认为是必要的， 细胞在复制受损DNA之前修复受损DNA 模板和分离受损染色体，具有缺陷p53的细胞 当暴露于DNA损伤时， 剂. 在拟议的研究中，我们将通过（1）来检验上述假设。 研究致癌物诱导的突变频率：（2）确定 受损DNA的修复;（3）确定化学物质的作用 致癌物，单独或与乙醇结合，对进展的 细胞周期、细胞周期蛋白依赖性激酶（cdks）活性和 主要生长停滞和DNA损伤诱导基因的表达（p53， WAF 1/C1 P1、gadd 45和gadd 153）在正常人口腔角质形成细胞中的表达 正常p53功能，表达HPV-16或HPV-18 E6蛋白的HOK， 表达突变型p53蛋白和HPV永生化的口腔角化细胞。 这些拟议的研究将有助于我们更深入地了解分子 口腔癌的发生机制。

There is compelling evidence that carcinogenesis is a multistep process and multiple genetic lesions are necessary to develop cancer in human. Among the genetic lesions, the dysfunction of p53 protein (because of mutation of p53 gene or infection by "high risk" human papillomaviruses [HPV], e.g., type 16 or 18 HPV) is the most frequently found genetic disorder in human cancers including oral cancer. In spite of such frequent p53 dysfunction in oral cancer cells, alter p53 function (by transfection with HPV DNA or mutant p53 cDNA) alone is not sufficient for neoplastic conversion of normal human oral keratinocytes in vitro. Therefore, the dysfunction of p53 protein may be an early event at least in oral carcinogenesis and also be necessary for subsequent genetic disorders of other genes to convert normal cells to tumor cells in the human oral cavity. In fact, our recent studies show that human oral keratinocytes containing negligible amount of wild-type (wt) p53 protein (because of HPV transfection) convert to tumorigenic cells when exposed to tobacco-carcinogens, but the normal counterpart does not. Inasmuch as wild-type p53 protein plays a major role in the regulation of cell cycle arrest, we hypothesize that normal human oral keratinocytes containing wt p53 protein repair damaged DNA more efficiently than oral keratinocytes with defective p53 function (by mutation of p53 gene or by infection with "high risk" HPV). As demonstrated by many studies, cells expressing wt p53 protein have the ability to establish transient delays in the progression of cell cycle when exposed to genotoxic agents. However, cells with defective p53 protein do not possess such ability. Since the arrest of the cell cycle progression is assumed to be necessary for cells to repair damaged DNA prior to replication of damaged DNA template and segregation of damaged chromosome, cells with defective p53 function may fail to repair the damaged DNA when exposed to DNA damaging agents. In the proposed study, we will test the above hypothesis by (1) investigating the carcinogen-induced mutation frequencies: (2) determining the repair of damaged DNA: and (3) determining the effect of chemical carcinogens, alone or in combination with ethanol, on the progression of cell cycle, the activity of cyclin-dependent kinases (cdks) and the expression of major growth arrest and DNA damage inducible genes (p53, WAF1/C1P1, gadd45, and gadd153) in normal human oral keratinocytes with normal p53 function, HOK expressing HPV-16 or HPV-18 E6 protein, HOK expressing mutant p53 protein, and HPV-immortalized oral keratinocytes. These proposed studies would help us gain more insight into molecular mechanisms of oral carcinogenesis.