Cas Protein Regulation of EMT in Cancer

描述（由申请人提供）：本提案将基于HEF 1升高驱动上皮-间质转化（EMT）的能力，检验肿瘤中HEF 1基因上调是癌症转移的重要促成因素的假设。 在EMT中，细胞失去上皮细胞特征，变得越来越能动，并获得侵入周围组织的能力，从而实现转移。 过去一年的一系列研究已经将HEF 1蛋白提名为肿瘤促转移行为的重要开关。 这些研究已经确定HEF 1是转移性乳腺腺癌中上调基因的小“签名”的组分，表明HEF 1对胶质母细胞瘤侵袭性很重要，并确定HEF 1的上调发生在超过30%的转移性黑色素瘤中，并且对于转移过程至关重要。 自从我们的研究小组在1996年首次描述HEF 1基因以来，我们一直在不断地研究这种蛋白质的信号作用。 这项工作已经证明了HEF 1作为支架蛋白或“枢纽”的核心作用，可以增强和协调促进癌症进展的多个信号级联反应的活性。 在这个建议中，我们将解决两个基本问题：1）HEF 1过表达的缺乏或升高如何调节经典的EMT/转移反应，包括E-cadherin的表达？HEF 1的高表达是乳腺癌转移的重要决定因素吗？ 在回答这些问题的具体实验中，Aim 1将使用基于细胞的机械分析来研究HEF 1如何调节EMT。 我们将检验HEF 1通过直接控制E-cadherin在粘附连接处形成稳定结构的能力来促进EMT的假设，并确定HEF 1是否是TGF-β或ErbB 2诱导的EMT的重要中间体。 目的2将使用基因定义的HEF 1-/-小鼠来测试HEF 1表达升高的能力显著促进乳腺癌转移的假设。 我们将HEF 1-/-小鼠与两种转基因小鼠（MMTV-PyVT和MMTV-neu）杂交，这些小鼠会发生原发性乳腺肿瘤和肺转移，并确定缺乏HEF 1是否会限制转移，以及肿瘤细胞中TGF-β和ErbB 2相关信号传导的条件。 最后，目标3将询问HEF 1升高是否预示着人类乳腺肿瘤的转移特征。 这些实验的最终目标是改善人类患者癌症的诊断和治疗。

DESCRIPTION (provided by applicant): This proposal will test the hypothesis that upregulation of the HEF1 gene in tumors is an important contributing factor for cancer metastasis, based on the ability of elevated HEF1 to drive epithelial-mesenchymal transition (EMT). In EMT, cells lose epithelial cell characteristics, become increasingly motile, and acquire the ability to invade surrounding tissue, enabling metastasis. A series of studies within the past year have nominated the HEF1 protein as an essential switch for pro-metastatic behavior in tumors. These studies have identified HEF1 as a component of a small "signature" of genes upregulated in metastasizing breast adenocarcinomas, shown that HEF1 is important for glioblastoma invasiveness, and determined that upregulation of HEF1 occurred in more than 30% of metastatic melanomas and was critical for the metastatic process. Since our group first described the HEF1 gene in 1996, we have worked continually to dissect the signaling role of this protein. This work has demonstrated a central role for HEF1 as a scaffolding protein or "hub" that enhances and coordinates the activity of multiple signaling cascades that promote cancer progression. In this proposal, we will address two basic questions: 1) How does absence or elevation of HEF1 overexpression condition classic EMT/metastatic responses, including expression of E-cadherin? and 2) Is elevated expression of HEF1 an important determinant of breast cancer metastasis? In specific experiments to answer these questions, Aim 1 will use mechanistic, cell-based assays to investigate how HEF1 regulates EMT. We will test the hypothesis that HEF1 promotes EMT by directly controlling the ability of E-cadherin to form stable structures at adherens junctions, and determine whether HEF1 is an essential intermediate in TGF-¿ or ErbB2-induced EMT. Aim 2 will use genetically defined HEF1-/- mice to test the hypothesis that the ability to elevate HEF1 expression contributes significantly to breast cancer metastasis. We will cross HEF1-/- mice to two strains of transgenic mice (MMTV-PyVT and MMTV-neu) that develop primary breast tumors and lung metastases, and determine whether absence of HEF1 limits metastasis, and conditions TGF-¿ and ErbB2-related signaling in tumor cells. Finally, Aim 3 will ask if elevated HEF1 predicts a metastatic profile in human breast tumors. The ultimate goal of these experiments is to improve the diagnosis and treatment of cancer in human patients.