Prevention of EtOH-induced promotion of hepatocarcinogenesis by genistein/soy

描述（由申请人提供）：肝细胞癌（HCC）现在是世界上第三大癌症死亡原因。酒精是HCC的一个众所周知的危险因素，并建议占所有HCC病例的32-45%。由于约7%的美国成年人滥用或依赖酒精，酒精诱导的HCC是一个严重的公共卫生问题。尽管其重要性，饮酒导致HCC的机制仍然不完全清楚。乙醇（EtOH）的主要作用似乎是作为一种肿瘤促进剂，增加肿瘤的发展速度，肿瘤的发展是由于暴露于饮食和环境致癌物，如酒精饮料和香烟烟雾中发现的亚硝胺或通过肝炎感染。我们的实验室已经证明了大鼠在EtOH处理后的肝脏修复（肝细胞增殖增加）伴随着b-连环蛋白的激活。此外，我们的初步数据表明，乙醇在小鼠中作为肿瘤促进剂，也与b-连环蛋白激活一致。这些新的数据表明，阻断EtOH刺激Wnt-b-连环蛋白信号传导可能是酒精相关HCC的有效预防机制。大豆产品大豆分离蛋白（SPI）和大豆植物化学染料木黄酮已被证明可以阻断包括乳腺和结肠在内的几种组织中Wnt-b-连环蛋白信号的激活。我们将检验以下假设：1）致癌物二乙基亚硝胺（DEN）和ETOH将通过增加的转录和通过增加的典型Wnt信号传导而相加地激活B-连环蛋白，（二）将膳食蛋白质从酪蛋白改变为SPI或在膳食中补充与大豆中发现的水平相似的染料木黄酮，将通过阻断EtOH通过诱导Wnt信号转导激活来防止EtOH促进肿瘤发生。可溶性FRZ蛋白抑制剂。为了做到这一点，我们将1）在我们的肿瘤促进的小鼠模型中检查SPI/染料木黄酮与EtOH共施用相对于EtOH单独处理减少肿瘤/小鼠数量的能力; 2）使用小鼠报告系统（TOP-GAL）和来自HCC患者的存档样品来证实EtOH诱导b-连环蛋白和与肝细胞中增殖标志物共定位;和3）通过使用靶向微阵列、蛋白质印迹、免疫组织化学和免疫沉淀分析基因表达，确定DEN/EtOH诱导b-连环蛋白的分子机制，以及与膳食染料木素/SPI的相互作用。 公共卫生相关性：肝细胞癌是世界上第三大癌症死亡原因，酗酒现在占病例的32-45%。这是一个重大的公共卫生问题，但酒精导致HCC的机制尚不完全清楚，也没有临床策略来降低酒精消费者患HCC的风险。我们的动物数据将Wnt-b-连环蛋白信号传导的激活与乙醇促进肝癌联系起来，并表明饮食因素如大豆/染料木黄酮（已被建议抑制-b-连环蛋白激活）可能会阻止这些作用。酒精和大豆/染料木黄酮对肝脏Wnt-b-连环蛋白信号级联作用的分子研究有望为肿瘤促进的共同途径提供基本见解。

DESCRIPTION (provided by applicant): Hepatocellular carcinoma (HCC) is now the world's third leading cause of cancer mortality. Alcohol is a well known risk factor for HCC and is suggested to account for 32-45% of all HCC cases. Since about 7% of the adult U.S. population abuses or is dependent on alcohol, alcohol-induced HCC is a serious public health problem. Despite its importance, the mechanisms whereby alcohol consumption causes HCC remain incompletely understood. The major role of ethanol (EtOH) appears to be to act as a tumor promoter, increasing the rate of development of tumors initiated as a result of exposure to dietary and environmental carcinogens such as nitrosamines found in alcoholic beverages and cigarette smoke or via hepatitis infection. Our laboratory has demonstrated liver repair (increased hepatocyte proliferation) after EtOH treatment in rats accompanied by activation of b-catenin. Moreover, our preliminary data suggest that EtOH acts as a tumor promoter in mice, also coincident with b-catenin activation. These novel data suggest that blockage of EtOH stimulation of Wnt- b-catenin signaling might be an effective prevention mechanism for alcohol related HCC. The soy product soy protein isolate (SPI) and the soy phytochemical genistein have been shown to block activation of Wnt-b-catenin signaling in several tissues including breast and colon. We will test the hypotheses that 1) The carcinogen diethylnitrosamine (DEN) and ETOH will act additively to activate b- catenin through increased transcription and via increased canonical Wnt signaling, 2) Changing the dietary protein from casein to SPI or supplementation of the diet with genistein at levels similar to those found in soy will prevent EtOH promotion of tumorigenesis by blocking EtOH activation of Wnt signaling through induction of soluble FRZ protein inhibitors. To do this we will 1) examine the ability of SPI/genistein co-administration with EtOH to reduce the number of tumors/mouse relative to EtOH treatment by itself in our mouse model of tumor promotion; 2) use a mouse reporter system (TOP-GAL) and archived samples from HCC patients to confirm EtOH induction of b-catenin and co-localization with proliferation markers in hepatocytes; and 3) determine the molecular mechanisms underlying DEN/EtOH induction of b-catenin, and interactions with dietary genistein/SPI, by analyzing gene expression using targeted microarrays, Western blotting, immunohistochemistry and immunoprecipitation. PUBLIC HEALTH RELEVANCE: Hepatocellular carcinoma is the world's third leading cause of cancer mortality and alcohol abuse now accounts for 32-45% of cases. This is a major public health issue and yet the mechanisms whereby alcohol causes HCC are incompletely understood and there are no clinical strategies to reduce risk of HCC in alcohol consumers. Our animal data link activation of Wnt-b-catenin signaling with promotion of liver cancer by ethanol and suggest that dietary factors such as soy/genistein which have been suggested to inhibit-b-catenin activation, might prevent these effects. Molecular studies of alcohol and soy/genistein actions on the liver Wnt- b-catenin signaling cascade are expected to provide fundamental insights into common pathways underlying tumor promotion.