Modulation of ErbB Signaling

项目描述（由申请人提供）：本项目的总体目标是阐明ErbB信号成分的调控机制，并探索这些分子作为癌症治疗的分子靶点。SHP2蛋白酪氨酸磷酸酶（PTPase）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，SHP2如何介导这些反应尚不清楚。研究表明，在表皮生长因子（EGF）刺激的细胞中，Gab1是关键的SHP2调节因子，SHP2激活Src和Ras，但SHP2激活Src和Ras的机制尚未明确。我们假设了两种shp2介导的Src激活模型，并将在Specific Aim i中进行评估。我们还将分析Src在egf诱导的Ras和Erk激活中的作用。在Specific Aim 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Gab1， Gab2和SHP2在egf诱导的细胞反应中的作用。其中包括Src和Akt/PKB激活、Ras-MAP激酶激活的时间调控、基因表达、paxillin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Gab1和相关的pleckstrin-homology （PH）结构域是对抗人类癌症中ErbB信号和PTEN突变的新分子靶点。这一假设将在特异性Aim III中进行评估，使用强力霉素诱导的诱饵构建物在体外稳定的癌细胞系和肿瘤异种移植中进行。这些研究将定义一个主要的ErbB和SHP2信号传导机制，极大地推进我们对Gab蛋白功能的理解，揭示恶性信号传导分子干预的新靶点，并刺激进一步开发针对这些信号传导成分的新抑制剂用于癌症治疗。

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.