Mechanisms of Met Induced Hepatocytes Survival

描述（由申请人提供）：逃避细胞凋亡是肝肿瘤和其他恶性肿瘤来源的癌细胞的主要特征。肝细胞生长因子受体（HGFR）被称为Met，是在促进肝细胞存活和增殖中起重要作用的跨膜受体酪氨酸激酶（RTK），并且其在包括肝细胞癌（HCC）的各种人类癌组织和细胞系中过表达。了解HCC发展的分子机制是NIDDK的一个关键举措（即，肝病研究行动计划）。通过结构-功能研究，我们发现人Met胞内结构域的C-末端含有一个新的串联的Caspase-3切割位点，我们将其命名为Met Caspase Decoy Site（MCDS）。它具有以下序列：DNAD（DEVD（TRPASFWETS（（表示胱天蛋白酶切割位点）。我们在该A2更新申请中要测试的中心假设是，由于具有独特的胱天蛋白酶底物诱饵基序（MCDS），人Met的细胞质尾区起到“诱饵”的作用，并通过在切割过程中形成稳定的过渡态中间体来捕获胱天蛋白酶-3的活性位点。这导致Caspase-3活性的抑制、凋亡抵抗和HCC的促进。国家的最先进的分子生物化学和生物学的方法，计划测试这一有趣的假设。在目的1中，我们将使用功能丧失和功能获得方法研究Met Caspase诱饵位点（MCDS）在Caspase-3抑制和促进肝细胞存活和肝癌发生中的功能作用。在目标2中，我们将评估参与MCDS-Caspase-3相互作用和Caspase-3抑制的分子机制。在目标3中，我们将确定MCDS功能是否取决于Met的激活状态。从这些研究中，我们应该收集大量的洞察力，以了解一种以前从未描述过的新的促生存机制。 公共卫生相关性：肝细胞癌（HCC）是最致命的癌症之一（仅次于胰腺癌）。对于HCC不存在有效的治疗，因为这些肿瘤因对主要通过诱导凋亡（或程序性细胞死亡）杀死细胞的化疗剂具有抗性而臭名昭著。我们已经发现，Met是一种致癌蛋白质，它会劫持导致细胞死亡的细胞凋亡机制（称为半胱天冬酶的专门酶），从而有助于癌细胞的存活和长寿。我们将研究Met抑制caspase的分子机制。总的来说，我们提出的研究将建立新的见解肝癌生长和细胞存活的分子机制。它们可能为合理的药物设计（即模拟Met胱天蛋白酶诱饵位点的药物）开辟途径，以治疗从肝炎到HCC的肝脏疾病。

DESCRIPTION (provided by applicant): Escape from apoptosis is a cardinal feature of cancer cells derived from liver tumors and other malignancies. The Hepatocyte Growth Factor Receptor (HGFR) known as Met is a transmembrane receptor tyrosine kinase (RTK) that plays an important role in promoting hepatocyte survival and proliferation, and it is overexpressed in various human carcinoma tissues and cell lines including hepatocellular carcinoma (HCC). Understanding the molecular mechanisms of HCC development is a key NIDDK initiative (i.e., the Action Plan for Liver Disease Research). Using structure-function studies, we discovered that the C-terminal end of human Met's intracellular domain harbors a novel tandem Caspase-3 cleavage site which we named the Met Caspase Decoy Site (MCDS). It has the following sequence: DNAD(DEVD(TRPASFWETS ((denotes the caspase cleavage site). Our central hypothesis to be tested in this A2 Renewal Application is that, by virtue of harboring a unique caspase substrate decoy motif (MCDS), the cytoplasmic tail of human Met functions as a 'bait' and traps the active site of Caspase-3 by forming a stable transition state intermediate during the cleavage process. This results in inhibition of Caspase-3 activity, apoptosis resistance and promotion of HCC. State-of-the-art molecular biochemical and biological approaches are planned to test this intriguing hypothesis. In Aim 1, we will investigate the functional role of the Met Caspase Decoy Site (MCDS) in Caspase-3 inhibition and promotion of hepatocyte survival and hepatocarcinogenesis using loss-of-function and gain-of-function approaches. In Aim 2, we will assess the molecular mechanisms involved in MCDS-Caspase-3 interaction and Caspase-3 inhibition. In Aim 3, we will determine whether MCDS function depends on the activation status of Met. From these studies, we should gather substantial insight into a novel pro-survival mechanism never before described. PUBLIC HEALTH RELEVANCE: Hepatocellular carcinoma (HCC) is the one of most lethal forms of cancer (only second to pancreatic adenocarcinoma). No effective treatment for HCC exists as these tumors are notorious for being resistant to chemotherapeutic agents which kill cells mainly via induction of apoptosis (or programmed cell death). We have discovered that Met, a cancer-causing protein, highjacks the cellular apoptotic machinery (specialized enzymes called caspases) that causes cell death hence contributing to survival and longevity of cancer cells. We will look at the molecular mechanisms of caspase inhibition by Met. Collectively our proposed studies will establish novel insights into the molecular mechanisms of HCC growth and cell survival. They may open avenues for rational drug design (i.e. drugs that mimic Met caspase decoy site) to treat liver diseases ranging from hepatitis to HCC.