BIOACTIVATION OF DIETARY PHENOLS BY HEMOPROTEINS

这项研究的长期目标是阐明 人类饮食中存在的酚类化合物影响 肿瘤发生正在进行的研究表明， 在许多情况下，细胞色素P450导致烷基酚的形成， 能够共价结合和/或自由基 在细胞中生成。我们现在建议将这项工作扩展到调查 这些反应性代谢物参与肿瘤促进， 最彻底的特征模型，用于探索 在肺中提升。该系统涉及肺肿瘤的增强 通过长期服用食品添加剂丁基化 羟基甲苯（BHT）致癌物启动的小鼠。已经 证明了BHT在靶器官中的代谢对于 促进，并且已知BHT转化为反应性醌型化合物 肺中的代谢物。这些发现引出了一个假设， 取决于两个连续的P450催化氧化， 促进物种，一种强亲电醌甲基化物， 一种或多种导致生长控制中断的关键蛋白质 机制等具体目标如下：（1）确定 代谢在促进的不同反应中的作用- 对肺肿瘤敏感（B+）和促进抗性（B-）小鼠 促进剂BHT。BHT转化为醌甲基化物和其他反应性 代谢物，以及反应性代谢物的解毒，将 在来自B+和B-小鼠的肺组织和细胞中进行检查。(2)探讨 BHT衍生的醌甲基化物在肺细胞中的烷基化靶点， 直接或间接损害细胞间信号传导。蛋白质烷基化， BHT的高反应性醌甲基化物代谢产物将在 从B=和B-小鼠的肺中分离的细胞，以及在致瘤性和 来源于鼠肺的非致瘤细胞系。烷基化模式 将通过放射化学和免疫化学方法进行比较， 通过质谱和微测序鉴定加合物。(3)审查 肺细胞的生化后果和氧化损伤 BHT的反应性代谢产物。从B+和B-小鼠和细胞中分离的细胞 将用BHT的反应性醌型代谢物处理细胞系， 研究细胞毒性、氧化损伤、线粒体抑制 功能和抑制解毒酶。

The long-term goal of this research is to elucidate the underlying mechanisms by which phenolic compounds present in the human diet influence tumorigenesis. Ongoing studies demonstrates that the oxidative metabolism of alkylphenols by cytochromes P450 lead, in many cases, to the formation of quinoid products capable of covalent binding and/or free radical generation in cells. We now propose to extend this work to investigate the involvement of such reactive metabolites in tumor promotion utilizing the most thoroughly characterized model for probing mechanistic aspects of promotion in the lung. This system involves the enhancement of lung tumor development by chronic administration of the food additive butylated hydroxytoluene (BHT) to carcinogen-initiated mice. It has been demonstrated that metabolism of BHT in the target organ is necessary for promotion and it is known that BHT is converted to reactive quinoid metabolites in lung. These findings lead to the hypothesis that promotion depends upon two successive P450 catalyzed oxidations to the ultimate promoting species, a strongly electrophilic quione methide which alkylates one or more critical proteins leading to a disruption of growth control mechanisms. The following specific aims are proposed: (1) Determine the role of metabolism in the differential responsiveness of promotion- sensitive (B+) and promotion-resistant (B-) mice to the lung tumor promoter BHT. Conversion of BHT to a quinone methide and other reactive metabolites, as well as the detoxification of reactive metabolites, will be examined in lung tissues and cells from B+ and B- mice. (2) Investigate alkylation targets of a BHT-derived quinone methide in lung cells that directly or indirectly impair cell-cell signaling. Protein alkylation by a highly reactive quinone methide metabolite of BHT will be examined in cells isolated from the lungs of B= and B- mice, and in tumorigenic and non-tumorigenic cell lines derived from murine lung. Alkylation patterns will be compared by radiochemical and immunochemical methods and selected adducts identified by mass spectrometry and microsequencing. (3) Examine biochemical consequences and oxidative damage in lung cells exposed to reactive metabolites of BHT. Isolated cells from B+ and B- mice and cell lines will be treated with reactive quinoid metabolites of BHT to investigate cytotoxicity, oxidative damage, inhibition of mitochondrial function, and inhibition of enzymes involved in detoxification.