Developmental pathways, environmental agents, and epigenetics in liver disease

描述(申请人提供)：肝细胞癌是世界上第五大最常见的癌症，每年造成约60万人死亡。酗酒和丙型肝炎病毒(丙型肝炎病毒)感染会导致炎症和慢性肝病，包括肝硬化，并协同增加患肝癌的风险。肝硬变是一种明确的癌前病变，但它在表观基因组水平的分子基础还没有被研究过。在许多国家，酒精消费正在上升，每年导致约180万人死亡。全球约有1.7亿人感染了丙型肝炎病毒，美国有400万人感染了丙型肝炎病毒，使其成为美国最常见的血液传播疾病。大多数肝癌(85%)的病因可能比任何其他癌症都更清楚，而且都是由环境因素或人类行为(如酒精、丙型肝炎病毒)引起的，但我们对这些制剂如何在30年内发挥致癌作用仍有许多不清楚的地方，具有显著的临床变异性。没有确凿的证据表明酒精或丙型肝炎病毒直接介导了基因损伤。然而，越来越多的证据表明，肝病期间早期和渐进性的表观遗传学变化，以及饮食和炎症在扰乱表观基因组中的作用。根据我们小组和其他人的研究，调控发育的基因似乎是包括肝癌在内的癌症表观遗传缺陷的优先靶点之一。对发育途径的表观遗传调控的全面分析从未在正常或疾病的肝脏中进行过，这限制了我们对人类肝病病因中表观遗传变化的全面范围和时间的了解，以及这些变化如何作为预后标志来改善治疗。我们假设，慢性酒精暴露和/或丙型肝炎病毒感染引起的发育途径的表观遗传学改变在肝病、丙型肝炎病毒诱导的细胞损伤和肝细胞癌的发生机制中起关键作用。我们提出了四个目标来解决这一假设。在目标1中，我们将通过将一个大的肝组织库与一个专注于发育调节基因的定制平铺阵列相结合，对酒精和/或丙型肝炎引起的肝硬变或肝癌患者的人肝组织进行表观遗传学分析。在目标2中，我们将使用细胞培养系统对未经处理和酒精处理/丙型肝炎病毒感染的原代肝细胞进行表观遗传学分析。在目标3中，我们将用灵敏和定量的分析来确认我们的阵列数据，并研究乙醇/丙型肝炎病毒调节模型发育基因的表观遗传控制的机制，以及发育基因的异常表达如何调节细胞生长。最后，在目标4中，我们将研究发育基因的表观遗传标记/表达如何与肝病进展过程中的临床和病理变化相关。这些研究的结果有望极大地提高我们对肝细胞癌表观遗传学的理解，为治疗和诊断/预后提供新的靶点，并提供关于普遍存在的娱乐药物和广泛存在的感染性因素如何影响表观基因组的新信息。

DESCRIPTION (provided by applicant): Hepatocellular carcinoma (HCC) is the 5th most common cancer worldwide, causing an estimated 600,000 deaths/yr. Alcohol abuse and hepatitis C virus (HCV) infection cause inflammation and chronic liver diseases, including cirrhosis, and synergistically increase risk for HCC. Cirrhosis is a well-defined precancerous lesion, yet its molecular underpinnings at the level of the epigenome have not been examined. Alcohol consumption is rising in many countries and results in ~1.8 million deaths/yr. Approximately 170 million people worldwide are infected with HCV, 4 million in the United States, making it the most common blood-borne disease in the U.S. The etiologic agents responsible for the majority of HCCs (>85%) are perhaps better understood than for any other cancer, and all are due to environmental agents or human behaviors (e.g. alcohol, HCV), yet there is much that we do not understand about how these agents exert their carcinogenic effects over >30 years with significant clinical variability. There is no definitive evidence for alcohol or HCV directly mediating genetic damage. There is mounting evidence, however, for early and progressive epigenetic changes during liver disease and for a role of diet and inflammation in disrupting the epigenome. Based on studies from our group and others, genes regulating development appear to be one of the preferential targets of epigenetic defects in cancers, including HCC. A comprehensive analysis of the epigenetic regulation of developmental pathways has never been performed in normal or diseased liver, limiting our knowledge of the full extent and timing of epigenetic changes in the etiology of human liver disease and how these changes may be used as prognostic markers to improve treatment. We hypothesize that epigenetic changes in developmental pathways caused by chronic alcohol exposure and/or HCV infection play a key role in the pathogenesis of liver disease, HCV-induced cellular damage, and HCC development. We propose four aims to address this hypothesis. In aim 1 we will epigenetically profile human liver tissues from patients with cirrhosis or HCC caused by alcohol and/or HCV by coupling a large liver tissue bank with a custom tiling array focused on developmental regulatory genes. In aim 2 we will epigenetically profile untreated and alcohol treated/HCV infected primary hepatocytes using a cell culture system. In aim 3 we will confirm our array data with sensitive and quantitative assays and examine the mechanisms by which ethanol/HCV modulate epigenetic control of model developmental genes and how aberrant expression of developmental genes regulates cell growth. Finally, in aim 4 we will examine how epigenetic marks/expression of developmental genes relates to clinical and pathological changes during liver disease progression. Results from these studies are expected to greatly enhance our understanding of the epigenetic etiology of HCC, provide new targets for therapy and diagnosis/prognosis, and yield novel information on how a ubiquitous recreational drug and a widespread infectious agent impact the epigenome.