DNA Double Strand Break Repair in Tobacco Carcinogenesis

描述(由申请人提供)： 尽管有强有力的流行病学数据将烟草烟雾与肺癌联系起来， 烟草化学物质致癌的机制还不是很清楚。 在那些暴露在烟草烟雾中的人中，只有一部分人会患上肺。 癌症提示遗传因素参与易感人群(S) 人口。DNA修复能力的不同被认为是 对易感性的解释。在各种DNA修复途径中，有一条 核苷酸切除修复与烟草致癌物的关系(例如： 苯并[a]芘)已有很好的文献记载。然而，DNA双链断裂 (DSB)修复是另一个重要的修复途径，很少有人用 哺乳动物系统中的烟草化学物质。因为最近的一项研究显示， 肺癌与一种蛋白质活性降低的正相关 参与DNA DSB修复，启动此应用程序是为了确认和 进一步研究DNA DSB修复在烟草致癌中的作用。脱氧核糖核酸 DSB是最具破坏性的DNA损伤形式，可以导致细胞死亡， 突变和转化，如果没有修复或错误修复。一种有效的方式 利用DNA DSB修复缺陷突变体研究DNA DSB修复的作用 暴露于烟草化学物质的细胞系。具体目标1：各种目标 啮齿动物和人类DNA DSB修复缺陷细胞株将被 几种烟草致癌物质和香烟烟雾冷凝物(CSC)，以及 将与野生型细胞在细胞存活率和 突变频率。这是为了验证这样的假设：“哺乳动物细胞利用 DNADSB修复机制或与DNADSB修复相关的蛋白质(S) 烟草致癌物所致DNA损伤的恢复过程“。 建议在细胞系研究的基础上进行分子研究。这些研究 有可能识别出新的烟草敏感亚群。这个 具体目标2的实验是基于我们最近的发现而提出的 肺癌敏感的BALB/c小鼠表现出轻微的DNA DSB修复缺陷。 具体目的2提出对细胞活力、DNA DSB的研究 肺泡巨噬细胞的形成和染色体重排， BALB/c和C57BL/6(对照组)小鼠的淋巴细胞和肺上皮细胞 暴露在烟草致癌物质中。为了实现这一目标，将检验两个假设：(1) 来自BALB/c小鼠的细胞显示出更高水平的染色体错误连接，即 通过一种敏感的早熟染色体凝集(PCC)试验检测 间期染色体结合/不结合原位荧光 杂交(FISH)。(2)敏感度之间有很好的相关性 淋巴细胞和AM和/或肺上皮细胞的数据。这个 拟议的实验不仅提供了对最初的 烟草致癌的步骤，但也有助于提供敏感的诊断 识别肺癌易感人群的工具。

DESCRIPTION (provided by applicant): Despite strong epidemiological data associating tobacco smoke to lung cancer, the mechanism of carcinogenesis by tobacco chemicals is not well understood. Among those exposed to tobacco smoke, only a certain portion develop lung cancer suggesting the involvement of genetic factor(s) in the susceptible population. A difference in DNA repair ability has been suggested for the explanation of the susceptibility. Of the various DNA repair pathways, a relationship between nucleotide excision repair and tobacco carcinogens (e.g. benzo[a]pyrene) has been well documented. However, DNA double strand break (DSB) repair, another important repair pathway, has rarely been studied with tobacco chemicals in mammalian systems. Since a recent study revealed a positive association between lung cancer and the reduced activity of a protein involved in DNA DSB repair, this application is initiated to confirm and extend the study on the role of DNA DSB repair in tobacco carcinogenesis. DNA DSB is the most destructive form of DNA damage and can lead to cell death, mutation and transformation if not repaired or mis-repaired. An efficient way to study the role of DNA DSB repair is to use DNA DSB repair deficient mutant cell lines exposed to tobacco chemicals. Specific Aim 1 addresses: Various rodent and human DNA DSB repair deficient cell lines will be treated with several tobacco carcinogens and cigarette smoke condensate (CSC), and comparisons will be made to wild type cells with regard to cell survival and mutation frequency. This is to test the hypothesis "Mammalian cells utilize DNA DSB repair mechanism or protein(s) associated with DNA DSB repair in the recovery process of DNA damage induced by tobacco carcinogens". Further molecular studies are suggested based on the cell line studies. These studies have the potential to identify a new tobacco sensitive sub-population. The experiments in Specific Aim 2 are proposed based on our recent finding that lung cancer sensitive BALB/c mice showed a mild DNA DSB repair defect. Specific Aim 2 puts forward the studies on the cell viability, DNA DSB formation and chromosome rearrangements in alveolar macrophage (AM), lymphocytes and lung epithelial cells from BALB/c and C57BL/6 (control) mice exposed to tobacco carcinogens. Two hypotheses will be tested in this aim: (1) Cells from BALB/c mice show a higher level of chromosome misrejoining which is detected by a sensitive premature chromosome condensation (PCC) assay in interphase chromosomes combined with/without fluorescence in situ hybridization (FISH)." (2) There is a good correlation between the sensitivity data with lymphocytes and those with AM and/or lung epithelial cells. The proposed experiments not only provide new mechanistic insight on the initial step for tobacco carcinogenesis, but also help furnish sensitive diagnostic tools to identify lung cancer susceptible individuals.