Arsenic Induced Miotic Arrest Associated Apoptosis

描述（由申请人提供）：砷是世界许多地区饮用水中的天然污染物，是一种已知的人类致癌物，是美国环保署危险化学品清单上的第一名。与慢性砷中毒最常相关的癌症是皮肤的鳞状细胞癌和基底细胞癌。砷是如何致癌的尚不清楚。然而，国家研究委员会关于饮用水中砷的报告得出结论，最可能的作用方式是诱导染色体数量和结构异常。砷的致癌形式砷存在于饮用水中，破坏有丝分裂导致后期延迟和诱导正常二倍体人成纤维细胞和外周血淋巴细胞的非整倍性，并在p53缺陷的人成纤维细胞中诱导有丝分裂阻滞相关凋亡（MAAA）。缺乏p53的人类细胞对亚砷酸盐诱导的MAAA的敏感性表明，亚砷酸盐致癌的机制不同于阳光诱发的皮肤癌，而阳光诱发的皮肤癌中p53突变是一个早期和常见的事件。需要研究的假设是，p53通过激活G2检查站反应来缓解亚砷酸盐诱导的后期阻滞，该反应使细胞周期蛋白B/cdc2失活，并抑制有丝分裂出口网络，使细胞逃脱亚砷酸盐诱导的MAAA。这是预防凋亡砷中毒细胞允许遗传不稳定（非整倍体）有丝分裂破坏后。确定与p53或p53调控基因相互作用的细胞因子，以防止有丝分裂阻滞相关的凋亡，并允许细胞延迟有丝分裂，将为亚砷酸盐的作用方式提供有价值的信息。提出的具体目标是：1.)测定有丝分裂中被亚砷酸盐阻滞的p53（+）和p53（-）细胞G2检查点通路的激活情况；2)。通过过表达和靶向敲除G2检查点蛋白检测G2检查点激活在亚砷酸盐诱导的晚期阻滞中逃逸的作用3)。检测砷相关皮肤肿瘤是p53野生型还是突变型。这些研究结果将确定介导亚砷酸盐诱导的有丝分裂停止释放的参与者，并将为砷诱导的致癌机制提供有价值的信息，为亚砷酸盐作为化疗药物的有用性提供线索，并为有丝分裂破坏药物杀死人类细胞的作用方式提供有价值的信息。

DESCRIPTION (provided by applicant): Arsenic is a natural contaminant of drinking water in many parts of the world, is a known human carcinogen and is #1 on the EPA list of hazardous chemicals. Cancers most often associated with chronic arsenism are squamous and basal cell carcinomas of the skin. How arsenic causes cancer is unknown. However, the National Research Council Report on Arsenic in Drinking Water concluded that the most likely mode of action is induction of numerical and structural chromosomal abnormalities. Arsenite, the carcinogenic form of arsenic found in drinking water, disrupts mitosis causing an anaphase delay and induces aneuploidy in normal diploid human fibroblasts and peripheral blood lymphocytes, and mitotic arrest associated apoptosis (MAAA) in p53 deficient human fibroblasts. The sensitivity of p53 deficient human cells to arsenite induced MAAA suggests that the mechanism of arsenite carcinogenesis is different than sunlight induced skin carcinogenesis in which p53 mutation is an early and common event. The hypothesis to be investigated is that p53 relieves the arsenite-induced anaphase block by activation of the G2 checkpoint response which inactivates cyclin B/cdc2 and derepresses the mitotic exit network and allow the cells to escape arsenite induced MAAA. It is the prevention of apoptosis in arsenic intoxicated cells that allows genetic instability (aneuploidy) after mitotic disruption. Identification of the cellular factors that interact with p53 or the p53 regulated genes to prevent mitotic arrest associated apoptosis and to allow cells to proceed through mitosis with a delay will provide valuable information regarding the mode of action of arsenite. The specific aims proposed are: 1.) Determine activation of the G2 checkpoint pathway in p53(+) and p53(-) cells arrested by arsenite in mitosis; 2.) Test by overexpression and targeted knockdown of G2 checkpoint proteins the role of G2 checkpoint activation in the escape from arsenite induced anaphase block; 3.) Test whether arsenic associated skin tumors are p53 wild type or mutant. The results of these studies will identify players mediating release from arsenite induced mitotic arrest, and will provide valuable information on the mechanism of arsenic induced carcinogenesis, clues to the usefulness of arsenite as a chemotherapeutic agent and valuable information on the mode of action of mitosis disrupting drugs in killing human cells.