Role of Sprouty 2 in Hepatocellular Carcinoma

描述（由申请人提供）： Sprouty的四种蛋白（Spry 1，2，3，4）抑制受体酪氨酸激酶（RTK）的生物学作用，从而调节各种器官的发育。由于它们作为RTK介导的细胞增殖和迁移的抑制剂的作用， 特别是Spry 2与许多癌症有关。因此，现在已经确定，Spry 2蛋白水平在乳腺癌、肺癌、前列腺癌和肝细胞癌（HCC）中降低。该提案的共同PI表明，在患者来源的HCC肿瘤中，Spry 2水平的显著降低与患者预后不良相关。此外，在这些HCC的一个亚组中，Spry 2水平的降低与Nedd 4 -1水平的增加呈负相关，PI的实验室已经表明Nedd 4 - 1是E3泛素连接酶之一，其使Spry 2靶向其进行泛素化， 降解PI的实验室还发现，Spry 2蛋白水平可以通过脯氨酰羟基化和von Hippel Lindau蛋白（pVHL）相关的E3连接酶进行调节。其他人以及我们表明pVHL蛋白水平在HCC亚群中升高。PI实验室的其他发现表明，Spry 2也是SUMO化的，这种翻译后修饰增加了其细胞含量。鉴于这些发现，本提案将检验的中心假设是， HCC中的Spry 2含量通过Nedd 4 -1和/或pVHL介导的泛素化和降解或通过其SUMO化的改变来调节，并且HCC中Spry 2含量的降低提高了RTK信号传导，从而增强了肿瘤的形成和生长。为了解决这一假设，我们将确定破坏内源性Spry 2/Nedd 4 -1相互作用是否增强Spry 2含量，从而增强其抑制小鼠中HCC细胞迁移和增殖以及HCC形成的能力。我们还将确定干扰Spry 2与Spry 2和pVHL的相互作用是否会增强其细胞含量，并增强其抑制HCC形成的能力。最后，我们将阐明Spry 2 SUMO化的机制，确定SUMO化是否稳定Spry 2并调节其在HCC细胞中的生物学作用，并确定Spry 2 SUMO化在HCC肿瘤中的状态。我们的研究不仅将确定Spry 2水平调节的新机制，还将为HCC治疗策略的新可能性提供见解。此外，由于Spry 2蛋白水平在乳腺癌、前列腺癌和肺癌中降低，这些癌症折磨着男性和女性退伍军人，我们的研究也将对这些疾病产生更广泛的影响。

DESCRIPTION (provided by applicant): Abstract The four Sprouty proteins (Spry1, 2, 3, & 4) inhibit the biological actions of receptor tyrosine kinases (RTKs) and thereby regulate the development of various organs. Because of their role as inhibitors RTK- mediated cell proliferation and migration, decrease in Spry proteins, especially Spry2 has been implicated in a number of cancers. Thus, it is now well established that Spry2 protein levels are decreased in breast, lung, prostate and hepatocellular carcinomas (HCC). The co-PI of this proposal has shown that in patient-derived HCC tumors, marked decreases in Spry2 levels correlated with poor patient outcome. Moreover, in a subset of these HCCs, the decreased Spry2 levels inversely correlated with increased levels of Nedd4-1, which the PI's lab has shown is one of the E3 ubiquitin ligases that ubiquitylates Spry2 targeting it for degradation. The PI's lab has also found that Spry2 protein levels can be regulated by prolyl hydroxylation and von Hippel Lindau protein (pVHL)- associated E3 ligase. Others as well as we show that pVHL protein levels are elevated in subsets of HCCs. Additional findings from the PI's lab show that Spry2 is also SUMOylated and this post-translational modification increases its cellular content. Given these findings, the central hypothesis that this proposal will test is that Spry2 content in HCCs is regulated by Nedd4-1- and/or pVHL- mediated ubiquitylation and degradation or via alterations in its SUMOylation and that decreased Spry2 content in HCC elevates RTK signaling, thereby augmenting tumor formation and growth. To address this hypothesis, we will determine whether disrupting the endogenous Spry2/Nedd4-1 interactions enhance Spry2 content and, therefore its ability to inhibit migration and proliferation of HCC cells and HCC formation in mice. We will also determine whether interfering with Spry2 interactions with Spry2 and pVHL enhances its cellular content and also augments its ability to suppress HCC formation. Finally, we will elucidate the mechanisms by which Spry2 is SUMOylated, determine whether SUMOylation stabilizes Spry2 and regulates its biological actions in HCC cells, and also determine the status of Spry2 SUMOylation in HCC tumors. Our studies will not only identify novel mechanisms by which Spry2 levels are regulated but also provide insights into newer possibilities of therapeutic strategies for HCCs. Moreover, since Spry2 protein levels are decreased in breast, prostate, and lung cancer that afflict male and female Veterans, our studies will also have a broader impact in these diseases.