Role of Age in Liver Cancer

描述(申请人提供)：年龄是导致肝癌的主要危险因素之一，它是世界上第五种最常见的癌症，也是导致癌症相关死亡的第三大常见原因。尽管癌症研究取得了重大进展，但在肝癌的早期发现和治疗方面进展甚微，导致预后不良。在过去的15年里，我的实验室一直在进行研究，以确定随着年龄的增长而改变的关键信号通路，这些信号通路使肝细胞更容易发生肝癌。其中一条途径是转录因子CCAAT/增强子结合蛋白αC/EBPA在S193处的过度磷酸化。我们已经成功地建立了C/EBPA-S193D敲门小鼠(S193D)，它模拟了年龄相关的C/EBPA的ph-S193亚型，并发现S193D小鼠肝部分切除后的肝脏增殖完全受到抑制。令我们惊讶的是，我们发现S193D小鼠的肝脏肿瘤发生得比WT小鼠快得多。此外，我们还发现，gankyrin与C/EBPA的S193D和ph-S193亚型特异地相互作用并降解。这一应用的主要假设是，甘结蛋白的激活和C/EBPA的降解是肝癌发生发展的关键调控途径。我们最近的研究表明，法尼类X受体(FXR)及其下游靶标SHP对甘结蛋白有抑制作用。FXR/SHP基因的缺失导致甘结蛋白的激活，导致C/EBPA的降解，并在12-17个月龄时形成自发性肝肿瘤。具体目标1将阐述FXR/SHP途径抑制gankyrin的机制。为此，我们计划完成以下工作：a)研究单个FXR和SHP KO小鼠是否激活gankyrin，b)检测新生、2个月、6个月和12个月龄FXR/SHP DKO小鼠的gankyrin激活情况，以确定gankyrin被激活的年龄，c)确定参与gankyrin激活的依赖于FXR的转录因子，以及d)通过检测不同年龄的人肝癌标本中的gankyrin来验证gankyrin-C/EBPA通路的激活。特异性目标2将检测FXR的激活是否抑制肝肿瘤。我们将通过FXR特异性配体GW4064激活FXR，并检测这种激活是否会在DEN介导的致癌过程中降低肝癌的发病率/数量。特异性目标3将研究抑制gankyrin-C/EBP1通路是否能阻止肝肿瘤的形成。我们计划使用以下方法：a)在WT DEN处理的小鼠和FXR/SHP小鼠中通过siRNA抑制gankyrin的表达，并检查这种抑制是否会减少肝脏肿瘤的发生率/数量，以及b)开发阻断gankyrin与C/EBP1结合的方法。因此，拟议的研究将为基于甘草蛋白的肝癌预防提供新的方法。 公共卫生相关性：这个项目调查了肝癌发生的分子基础。尽管癌症研究取得了重大进展，但迫切需要开发新的方法来早期发现和有效处理肝癌。我们将探讨肝癌激活甘结蛋白和降解肿瘤抑制因子Ph-S193亚型C/EBP1的机制，为肝癌预防治疗方法的研究奠定基础。

DESCRIPTION (provided by applicant): Age is one of the major risk factors for development of the liver cancer, which is the fifth most common cancer and the third most common cause of cancer related death in the world. Despite significant advances in cancer research, little progress has been made in early detection and treatment of liver cancer leading to poor prognosis. Over the past 15 years, my laboratory has pursued the studies to determine the key signaling pathways that are changed by aging and that make liver cells more susceptible to development of liver cancer. One of these pathways is the hyper-phosphorylation of a transcription factor, CCAAT/Enhancer Binding protein alpha, C/EBPa, at S193. We have successfully generated C/EBPa-S193D knockin mice (S193D) which mimic the age-associated ph-S193 isoform of C/EBPa and found that liver proliferation after partial hepatectomy is completely inhibited in S193D mice. To our surprise, we found that the development of liver tumors in S193D mice occurs much faster than in WT mice. Furthermore, we found that gankyrin specifically interacts with and degrades the S193D and ph-S193 isoforms of C/EBPa. The main hypothesis of the application is that the activation of gankyrin and following degradation of C/EBPa is a key regulatory pathway in development of liver cancer. Our recent studies show that farnesoid X receptor (FXR) and its down-stream target SHP suppress gankyrin. The deletion of FXR/SHP leads to activation of gankyrin resulting in the degradation of C/EBPa and the formation of spontaneous liver tumors at age between 12-17 months. Specific Aim 1 will address mechanisms by which FXR/SHP pathway represses gankyrin. In this aim, we plan to accomplish the following: a) Investigate if single FXR and SHP KO mice activate gankyrin, b) Examine the activation of gankyrin in newborn, 2 mo, 6 mo and 12 mo old FXR/SHP DKO mice in order to determine the age at which gankyrin is activated, c) Identify FXR-dependent transcription factors that are involved in the activation of gankyrin, and d) Verify the activation of gankyrin- C/EBPa pathway by measuring gankyrin in human HCC samples obtained from patients of different ages. Specific Aim 2 will examine if the activation of FXR inhibits liver tumor. We will activate FXR by FXR-specific ligand GW4064 and examine if this activation will reduce the incidence/number of liver tumors in DEN- mediated carcinogenesis. Specific Aim 3 will investigate if the inhibition of gankyrin-C/EBP1 pathway will prevent liver tumor formation. We plan to use the following approaches: a) Inhibit the expression of gankyrin by siRNA in both WT DEN-treated mice and FXR/SHP mice and examine if this inhibition will reduce the incidence/number of liver tumors, and b) Develop approaches to block gankyrin binding to C/EBP1. Thus, the proposed studies will provide novel gankyrin-based approaches for the prevention of liver cancer. PUBLIC HEALTH RELEVANCE: This project investigates molecular basis for the development of cancer in the liver. Despite significant advances in cancer research, there is an urgent need to develop new approaches for the early detection and effective management of liver cancer. We will determine mechanisms by which liver cancer activates gankyrin and degrades tumor suppressor ph-S193 isoform of C/EBP1 with the goal to provide basement for the generation of therapeutic approaches to prevent liver cancer.