A Balancing Act: Role of HNF4α/β-catenin in Hepatobiliary Development and Cholangiocarcinoma Formation

项目摘要/摘要 胆管细胞癌(CC)是世界上最致命的癌症之一，其五年存活率为 肝内胆管细胞癌(ICC)约30%，肝外胆管细胞癌(ECC)约50%。这是一个 罕见的癌症，每年在美国影响大约8,500名患者(5,500名ICC，3000名ECC)。通常癌症会发展成 来自调节细胞生长和分化的信号之间的不平衡。虽然在这方面的大多数进展 癌症的治疗是通过抑制细胞增殖，重新建立细胞分化来进行的 最终治愈了某些形式的白血病。在这份提案中，我们将重点关注 肝细胞核因子4α(HNF4α)是肝脏分化的主要调节因子，Wnt/β- 连环蛋白信号通路，促进细胞增殖。我们将使用斑马鱼进行研究，这已经 由于活体成像、时间分辨率和化学筛选方法作为模型生物的优势 在啮齿动物模型中不可用或不可行。重要的是，斑马鱼表现出疾病的形态和 类似于哺乳动物的病理生理学。我们假设HNF4α/β-连环蛋白的相互作用是重要的 在肝胆发育中发挥重要作用的细胞命运在发育和 胆管细胞癌形成。我们的第一个目标是表征和定义HNF4α/β-连环蛋白的重要性 与肝胆发育有关的相互作用。我们最近产生了新奇的HNF4α基因敲除 斑马鱼和正在产生一种热休克诱导的HNF4α过度表达的斑马鱼。 这些模型将使我们不仅能够确定HNF4α在肝胆发育中的作用，而且在 结合Wnt/β-连环蛋白调节的遗传和药理学模型，将使我们能够表征 这两条路径之间的相互作用。我们将使用这些突变体和化学调节剂来 描述肝胆发育过程中的肝脏生长、形态和生理特征，并评估 正常肝和胆管细胞命运的微扰。我们的第二个目标是确定HNF4α/β的角色- 连环蛋白在胆管癌细胞形成中的相互作用。我们将首先描述肿瘤的发病率、进展、 以及突变导致HNF4α/β-连环蛋白途径失调的斑马鱼的死亡率。此外， 我们将利用遗传和药物手段重建正常的HNF4α/β-连环蛋白相互作用和 观察肿瘤负荷的变化。这些研究不仅将揭示胆管细胞癌的新机制 发病机制，也将使我们能够为这种毁灭性的治疗开发新的靶点 疾病。

PROJECT SUMMARY/ABSTRACT Cholangiocarcinoma (CC) is among the deadliest cancers worldwide, with a five-year survival rate of ~30% in intrahepatic cholangiocarcinoma (ICC) and ~50% in extrahepatic cholangiocarcinoma (ECC). It is a rare cancer, affecting ~8,500 (5,500 ICC, 3000 ECC) patients in the U.S. each year. Often cancers develop from an imbalance between signals that regulate cellular growth and differentiation. While most advances in cancer therapy have been made by inhibition of cell proliferation, the re-establishment of cell differentiation has resulted in the cure of certain forms of leukemia. In this proposal, we will focus on the interaction between hepatocyte nuclear factor 4 alpha (HNF4α), the master regulator of hepatic differentiation, and the Wnt/β- catenin signaling pathway, which enhances cell proliferation. We will use zebrafish for our studies, which has advantages as a model organism because of in vivo imaging, time resolution, and chemical screening methods not available or feasible in rodent models. Importantly, zebrafish display disease morphology and pathophysiology that is similar to mammals. We hypothesize that an HNF4α/β-catenin interaction is important in hepatobiliary development playing an important role cell fate during both development and cholangiocarcinoma formation. Our first aim is to characterize and define the importance of HNF4α/β-catenin interaction in regard to hepatobiliary development. We have recently generated novel HNF4α knockout zebrafish and are in the process of generating a heat shock-inducible HNF4α over-expression zebrafish. These models will allow us to not only define the role of HNF4α in hepatobiliary development, but, in combination with genetic and pharmacologic models of Wnt/β-catenin modulation, will allow us to characterize an interaction between these two pathways. We will use these mutants and and chemical modulators to characterize liver growth, morphology, and physiology during hepatobiliary development and assess perturbations in normal hepatic and biliary cell fate. Our second aim is to determine a role for the HNF4α/β- catenin interaction in cholangiocarcinoma formation. We will first characterize tumor incidence, progression, and mortality in zebrafish with mutations resulting in dysregulation of the HNF4α/β-catenin pathway. Further, we will utilize both genetic and pharmacologic means to re-establish a normal HNF4α/β-catenin interaction and observe changes in tumor burden. These studies will not only reveal novel mechanisms of cholangiocarcinoma pathogenesis, but also will allow us to develop new targets for therapeutic development for this devastating disease.