Effects of chemopreventive agents on DNA damage response

描述(由申请人提供)： 苯并[a]芘(B[a]P)是一种典型的不完全燃烧产生的多环芳烃，是香烟烟雾和烧焦食物中的主要致癌物。苯并[a]磷等多环芳烃是主要的环境污染物，暴露范围广泛。高活性和致突变的B[a]P代谢物(+)-7R，8S-dihydroxy-9S，10R-epoxy-7，8，9，10-tetrahydrobenzo[a]pyrene(BPDE)被认为在人类肺癌的发生中起着重要的作用。化学预防是一种相对较新且前景看好的预防癌症的策略。染料木素、姜黄素和白藜芦醇等化学预防药物具有多个分子靶点，影响多个信号通路，对肺癌、乳腺癌、前列腺癌和结肠癌具有强大的抗增殖作用。近年来，这三种成分作为癌症化学保护剂受到了极大的关注。关于染料木素、白藜芦醇和姜黄素对多环芳烃诱导的DNA损伤和修复反应的化学预防作用的研究很少。核苷酸切除修复的全局基因组修复部分由p53诱导来调节，以响应DNA损伤。P53在DNA损伤后以剂量和时间依赖的方式被诱导。低剂量的BPDE具有致突变性，但不能诱导P53介导的DNA损伤反应。我们的初步结果表明，联合使用白藜芦醇和姜黄素可以通过低剂量的BPDE诱导P53的表达。我们假设，化学预防药物可能通过诱导P53激活整体基因组DNA修复过程，使细胞对低剂量遗传毒物诱导的DNA损伤做出反应，从而实现更有效的DNA修复。本研究旨在探讨化学保护剂对BPDE诱导的人肺细胞DNA损伤反应的影响。我们的目标是确定化学预防药物(单独和联合使用)对P53介导的(1)细胞周期进程和(2)DNA修复诱导的调节的机制和作用。实验方法将包括通过蛋白质印迹法检测人肺细胞中的P53和受调控的信号蛋白，以及通过32P后标记分析来测量BPDE诱导的DNA加合物的形成和修复率。这一结果可能会阐明化学预防机制，并提供更深入的了解在更具生理意义的环境暴露剂量下DNA损伤和修复的机制。

DESCRIPTION (provided by applicant): Benzo[a]pyrene (B[a]P) is a prototypical polycyclic aromatic hydrocarbon produced by incomplete combustion and is a major carcinogen present in cigarette smoke and charred foods. PAH such as B[a]P are major environmental pollutants and exposure is widespread. The highly reactive and mutagenic B[a]P metabolite (+)-7R,8S-dihydroxy-9S,10R-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) is believed to play a significant role in human lung carcinogenesis. Chemoprevention is a relatively new and promising strategy to prevent cancer. Chemopreventive agents such as genistein, curcumin and resveratrol have a number of molecular targets, impact several signalling pathways and have strong antiproliferation effects on lung, breast, prostate and colon cancers. These three constituents have received a great deal of attention recently as cancer chemoprotective agents. Little work has been done to investigate the chemoprevention effects of genistein, resveratrol and curcumin with respect to PAH-induced DNA damage and repair responses. The global genomic repair component of nucleotide excision repair is regulated by p53 induction in response to DNA damage. P53 is induced in a dose and time-dependent manner after DNA damage. Low dose BPDE which is known to be mutagenic fails to induce the p53 mediated response to DNA damage. Our preliminary results suggest that pretreatment with a combination of resveratrol and curcumin enables p53 induction by low dose BPDE exposure. We hypothesize that chemopreventive agents may "sensitize" the cell to respond to low dose genotoxicant induced DNA damage through induction of p53 which activates global genomic DNA repair process allowing more efficient DNA repair. This study aims to investigate the effects of chemoprevention agents on cellular responses to BPDE-induced DNA damage in human lung cells. We aim to determine mechanism and effects of chemopreventive agents (singly and in combination) on p53 mediated regulation of (1) cell cycle progression and (2) DNA repair induction. Experimental approaches will include western blot to measure p53 and regulated signaling proteins in human lung cells and measure BPDE-induced DNA adduct formation and repair rate by 32P-post-labeling analyses. The results will likely illuminate a chemoprevention mechanism and also provide a deeper understanding of the mechanism of DNA damage and repair under a more physiologically relevant environmental exposure dose.