Eph Kinase Signaling in Prostate Cancer

描述（由申请人提供） 该提案的长期目标是表征分子机制 前列腺上皮细胞迁移的潜在有效抑制 受体酪氨酸激酶。细胞迁移涉及几个关键步骤 在肿瘤转移期间;对于前列腺癌， 在体外的运动性和体内的转移潜力之间。使用PC-3细胞作为 在模型系统中，申请人的实验室发现EphA 2激酶激活 有效抑制对纤连蛋白的趋触运动和趋化运动 向表皮和肝细胞生长因子的运动性。的抑制作用 与整合素亲和力的抑制和去磷酸化相关 和粘着斑激酶（FAK）的催化失活， 酪氨酸激酶主要参与细胞运动调节。EphA2 该复合物与FAK物理结合，并在EphA 2 activation. Shp-2蛋白酪氨酸磷酸酶（PTP-Sh）在细胞内呈一过性表达， 被招募到激活的EphA 2，并代表一个候选人PTP 3 参与FAK去磷酸化。这些发现定义了一种新的信号传导， 负性调节前列腺的趋触性和趋化性的途径 上皮细胞本提案的重点是检验FAK 是EphA 2激酶的下游效应子。在具体目标1中，各区域和 EphA 2和FAK上负责它们相互作用的位点将使用 缺失和定点诱变。EphA 2/FAK缺陷突变体 关联将用于评估 与细胞迁移B phA 2激酶的负调控相关。 特异性目的2旨在研究FAK去磷酸化的作用， 使用分子和遗传方法在EphA 2信号传导中的Shp-2募集。 鉴于细胞运动在前列腺癌转移中的基本作用， 完成拟议的研究将揭示自然存在的 信号通路，关闭细胞迁移，并可能导致新的 预防和治疗前列腺癌转移的策略。

DESCRIPTION (Provided by the applicant) The long-term goal of this proposal is to characterize the molecular mechanisms underlying potent inhibition of prostatic epithelial cell migration by Eph receptor tyrosine kinases. Cell migration is involved in several critical steps during tumor metastasis; for prostate cancer there is a direct correlation between motility in vitro and metastatic potential in vivo. Using PC-3 cells as a model system, the applicant's laboratory found that EphA2 kinase activation potently inhibited both haptotactic motility toward fibronectin and chemotactic motility toward epidermal and hepatocyte growth factors. The inhibitory effects were correlated with suppression of integrin affinity, and dephosphorylation and catalytic inactivation of focal adhesion kinase (FAK), a cytoplasmic tyrosine kinase critically involved in cell motility regulation. EphA2 physically associated with FAK and the complex was dissociated upon EphA2 activation. Shp-2 protein tyrosine phosphatase (PTPase) was transiently recruited to the activated EphA2, and represented one candidate PTPase implicated in FAK dephosphorylation. These findings defined a novel signaling pathway that negatively regulates haptotaxis and chemotaxis of prostatic epithelial cells. The focus of this proposal is to test the hypothesis that FAK is a downstream effector of EphA2 kinase. In Specific Aim 1, the regions and sites on EphA2 and FAK responsible for their interaction will be mapped using deletion and site-directed mutagenesis. Mutants deficient in EphA2/FAK association will e used to assess the functional significance of the association in the negative regulation of cell migration b phA2 kinase. Specific Aim 2 is designed to investigate the role of FAK dephosphorylation and Shp-2 recruitment in EphA2 signaling using molecular and genetic approaches. Given the fundamental role of cell motility in prostate cancer metastasis, completion of the proposed studies will shed light on a naturally-existing signaling pathway that turns off cell migration, and may lead to novel strategies in preventing and treating prostate cancer metastasis.