Cas Protein Regulation of EMT in Cancer

描述（由申请人提供）：本提案将基于HEF1升高驱动上皮-间质转化（EMT）的能力，验证肿瘤中HEF1基因上调是癌症转移的重要促成因素的假设。在EMT中，细胞失去了上皮细胞的特征，变得越来越活跃，并获得了侵入周围组织的能力，从而实现了转移。过去一年的一系列研究表明，HEF1蛋白是肿瘤中促转移行为的重要开关。这些研究已经确定HEF1是转移性乳腺腺癌中上调基因的一个小“特征”组成部分，表明HEF1对胶质母细胞瘤的侵袭性很重要，并确定超过30%的转移性黑色素瘤中HEF1上调，并且对转移过程至关重要。自我们的团队于1996年首次描述HEF1基因以来，我们一直致力于剖析该蛋白的信号作用。这项工作已经证明了HEF1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种信号级联反应的活性。在本提案中，我们将解决两个基本问题：1)HEF1过表达的缺失或升高如何构成典型的EMT/转移反应，包括E-cadherin的表达？2) HEF1表达升高是否是乳腺癌转移的重要决定因素？在回答这些问题的具体实验中，Aim 1将使用机制的、基于细胞的分析来研究HEF1如何调节EMT。我们将验证HEF1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HEF1是否是TGF-¿或erbb2诱导EMT的必要中间体。目的2将使用基因定义的HEF1-/-小鼠来验证HEF1表达升高对乳腺癌转移有显著影响的假设。我们将HEF1-/-小鼠与两株发生原发性乳腺肿瘤和肺转移的转基因小鼠（MMTV-PyVT和MMTV-neu）杂交，并确定HEF1的缺失是否限制了转移，以及肿瘤细胞中TGF-¿和erbb2相关信号的表达。最后，Aim 3将询问HEF1升高是否能预测人类乳腺肿瘤的转移特征。这些实验的最终目的是提高人类癌症患者的诊断和治疗。

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.