Modulation of ErbB Signaling

描述（由申请人提供）：本项目的总体目标是阐明ErbB信号传导组分的调控机制，并探索这些分子作为癌症治疗的分子靶点。SHP 2蛋白酪氨酸磷酸酶（PTP 2）介导ErbB诱导的Erk丝裂原活化蛋白（MAP）激酶活化，并参与细胞骨架重组和细胞运动。然而，SHP 2如何介导这些反应还不清楚。研究表明，在表皮生长因子（EGF）刺激的细胞中，Gab 1是SHP 2的关键调节因子，Src和Ras被SHP 2激活，但SHP 2激活Src和Ras的机制尚未确定。假设了两种SHP 2介导的Src激活模型，并将在特定目标I中进行评价。Src在EGF诱导的Ras和Erk激活中的参与也将被分析。在特定目标II中，将开发用于基因敲低的新型小干扰RNA（siRNA）技术，并用于分析Gab 1，Gab 2和SHP 2在EGF诱导的细胞反应中的作用。这些包括Src和Akt/PKB激活，Ras-MAP激酶激活的时间调节，基因表达，桩蛋白去磷酸化，以及细胞生长和转移特性。基于我们的初步观察，我们假设Gab 1和相关的pleckstrin-homology（PH）结构域是拮抗ErbB信号传导和人类癌症中PTEN突变的新分子靶点。将在Specific Aim III中使用多西环素诱导型诱饵构建体在体外稳定的癌细胞系和肿瘤异种移植物中评估该假设。这些研究将定义一个主要的ErbB和SHP 2信号传导机制，大大推进我们对Gab蛋白功能的理解，发现恶性信号传导分子干预的新靶点，并刺激针对这些信号传导成分的新抑制剂的进一步开发。

DESCRIPTION (provided by applicant): The overall goals of this project are to elucidate regulatory mechanisms of ErbB signaling components and to explore these molecules as molecular targets for cancer therapy. SHP2 protein tyrosine phosphatase (PTPase) mediates ErbB-induced Erk mitogen-activated protein (MAP) kinase activation and is involved in cytoskeletal reorganization and cell motility. However, how SHP2 mediates these responses is not well understood. It has been shown that Gab1 is a key SHP2 regulator in epidermal growth factor (EGF)-stimulated cells and that Src and Ras are activated by SHP2, but the mechanisms by which SHP2 activates Src and Ras have not been defined. Two models for SHP2-mediated Src activation are postulated and will be evaluated in Specific Aim I. The involvement of Src in EGF-induced Ras and Erk activation also will be analyzed. In Specific Aim II, the novel small interfering RNA (siRNA) technique for gene knockdown will be developed and used to analyze the role of Gab1, Gab2, and SHP2 in EGF-induced cellular responses. These include Src and Akt/PKB activation, temporal regulation of Ras-MAP kinase activation, gene expression, paxillin dephosphorylation, and cell growth and metastatic properties. Based on our preliminary observations, we postulate that Gab1 and related pleckstrin-homology (PH) domains are novel molecular targets for antagonizing ErbB signaling and PTEN mutation in human cancers. This hypothesis will be evaluated in Specific Aim III using doxycycline-inducible decoy constructs in stable cancer cell lines in vitro and in tumor xenografts. These studies will define a major ErbB and SHP2 signaling mechanism, greatly advance our understanding of the function of Gab proteins, uncover novel targets for molecular intervention of malignant signaling, and stimulate further development of new inhibitors targeting these signaling components for cancer treatment.