P53, CHEMICAL CARCINOGEN AND ETHANOL IN ORAL CANCER

有令人信服的证据表明致癌是一个多步骤的过程。 而多种基因损伤是人类发生癌症所必需的。 在遗传损伤中，P53蛋白的功能障碍(由于 P53基因突变或“高危”人乳头瘤病毒感染 [HPV]，例如，16型或18型HPV)是最常见的基因 人类癌症中的疾病，包括口腔癌。尽管如此， 口腔癌细胞中频繁的P53功能障碍，改变P53功能(通过 HPV DNA或突变型P53基因的单独转染)不足以 正常人口腔角质形成细胞体外肿瘤转化的研究。 因此，P53蛋白的功能障碍可能至少是一个早期事件 在口腔癌的发生中，也是后续遗传所必需的 正常细胞转化为肿瘤细胞的其他基因紊乱 人类口腔。事实上，我们最近的研究表明，人类的口腔 角质形成细胞含有微不足道的野生型(Wt)P53蛋白 (由于HPV感染)接触时转化为致瘤细胞 烟草是致癌物质，但普通烟草不会。 由于野生型P53蛋白在细胞周期调控中起主要作用 细胞周期停滞，我们假设正常人口腔角质形成细胞 含有wt p53蛋白的DNA修复比口服更有效 具有P53功能缺陷的角质形成细胞(通过P53基因突变或通过 感染“高危”人乳头瘤病毒)。正如许多研究表明的那样，细胞 表达wt p53蛋白能够建立一过性延迟 在细胞周期进程中，当暴露于遗传毒性物质时。 然而，P53蛋白缺陷的细胞不具备这种能力。 因为假定细胞周期进程的停止是必要的 使细胞在复制受损DNA之前修复受损DNA 受损染色体、P53缺陷细胞的模板和分离 当暴露于DNA损伤时，功能可能无法修复受损的DNA 探员们。在拟议的研究中，我们将通过(1)检验上述假设 致癌物诱变频率调查：(2)测定 DNA损伤的修复：以及(3)确定化学物质的作用 致癌物，单独或与乙醇联合使用，对癌症进展的影响 细胞周期、细胞周期蛋白依赖性蛋白激酶(CDKs)活性和细胞周期 主要生长停滞和DNA损伤诱导基因的表达(p53， WAF1/C1P1、GADD45和Gadd153)在口腔角质形成细胞中的表达 P53功能正常，HPV-16或HPV-18 E6蛋白表达 表达突变型P53蛋白和HPV永生化的口腔角质形成细胞。 这些拟议的研究将帮助我们更深入地了解分子 口腔癌变的机制。

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.