Role and Mechanism of HER2 Self-Association in Cancer

HER2自关联在癌症中的作用和机制

• 批准号: 7072634
• 负责人: RALF LANDGRAF
• 金额: $ 26.85万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7072634
• 关键词: cell line; cell transformation; fluorescence resonance energy transfer; heregulin; ligands; mass spectrometry; molecular oncology; neoplasm /cancer; neoplastic growth; phosphorylation; polymerase chain reaction; protein structure function; protein tyrosine kinase; receptor expression

DESCRIPTION (provided by applicant): Overexpression of HER2, a receptor tyrosine kinase (RTK) is found in a series of cancers, most notably, 30% of breast cancers and 10% of ovarian cancers. Elevated levels of the receptor are associated with constitutive receptor activation, cellular transformation, and poorer survival of patients in a clinical setting. Activation of RTKs is known to involve dimerization. HER2, which has a potent cytoplasmic kinase domain but fails to bind any ligand by itself, heterodimerizes with other RTKs, preferentially with the kinase-deficient but ligand-binding HER3. The preferred ligand for the heterodimer is heregulin. However, at elevated levels, full-length and membrane-bound HER2 can self-associate ligand independently. The relative contribution of ligand-independent self-association versus an overall increase in ligand-dependent heterodimerization is an unresolved question for HER2 overexpressing cancers. A key objective of our work is to dissect those two reactions on a molecular level. To this end, we will identify and mutate the self association interface of HER2. We found recently that the catalytically inactive HER3, as well as its soluble extracellular domain (ECD), self-associates at low concentrations, a reaction that is disrupted by ligand binding. We will use the strong self-association of HER3 and its high homology with HER2 to identify self association sites in both receptors. This identification relies on computational predictions by 3D cluster analysis as well as alternative mutagenesis and selection approaches. We will apply FRET and DHFR fragment complementation to obtain direct measurements of the extent of receptor association in intact cells. We will use these methods to confirm the identified regions in HER3 and HER2 and evaluate the correlation between the concentration-dependent receptor self-association of HER2 and constitutive signaling and the enhancement of tumor formation. In addition we have evidence that HER3 self-association may modulate heregulin signaling. We will evaluate the possible regulatory role of HER3 self-association and its impact on ligand-dependent and independent signaling by HER2. A better understanding of the specific receptor interactions at elevated expression levels will enhance our understanding of cellular transformation by HER2, help understand and improve the action of drugs like Herceptin, and identify specific and measurable association events as targets for drug development.

描述（由申请人提供）：HER2的过表达，在一系列癌症中发现了受体酪氨酸激酶（RTK），最值得注意的是，30％的乳腺癌和10％的卵巢癌。受体的水平升高与临床环境中患者的组成型受体激活，细胞转化和较差的生存有关。已知RTK的激活涉及二聚化。 HER2具有有效的细胞质激酶结构域，但无法本身结合任何配体，与其他RTK异二聚体优先使用激酶缺陷型但配体粘合的HER3。异二聚体的首选配体是这里策略。但是，在较高的水平下，全长和膜结合的HER2可以独立地自我关联。对于HER2过表达的癌症而言，配体独立于配体的自我联系与配体依赖性异二聚化的总体贡献是一个尚未解决的问题。我们工作的一个关键目的是在分子水平上剖析这两个反应。为此，我们将识别并突变HER2的自我关联界面。我们最近发现，催化无活跃的HER3及其可溶性细胞外结构域（ECD），以低浓度的自求解，这种反应受到配体结合而破坏的反应。我们将使用HER3及其与HER2的高同源性的强烈自我关联来识别两个受体中的自我关联站点。这种识别依赖于3D聚类分析以及替代诱变和选择方法的计算预测。我们将应用FRET和DHFR片段互补，以获得完整细胞中受体缔合程度的直接测量。我们将使用这些方法来确认HER3和HER2中的已鉴定区域，并评估HER2的浓度依赖性受体自我关联与本构信号传导与肿瘤形成的增强之间的相关性。此外，我们有证据表明HER3自我关联可能会调节此处的糖蛋白信号传导。我们将评估HER3自我关联的可能调节作用及其对HER2对配体依赖和独立信号的影响。更好地理解在升高表达水平下的特定受体相互作用将增强我们对HER2的细胞转化的理解，有助于理解和改善诸如Herceptin之类的药物的作用，并确定特定且可测量的关联事件作为药物开发的靶标。