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: 6767575
• 负责人: RALF LANDGRAF
• 金额: $ 27.32万
• 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/6767575
• 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（一种受体酪氨酸激酶（RTK））的过表达存在于一系列癌症中，最值得注意的是，30% 的乳腺癌和 10% 的卵巢癌。受体水平升高与组成型受体激活、细胞转化和临床环境中患者生存率较差有关。已知 RTK 的激活涉及二聚化。 HER2 具有有效的胞质激酶结构域，但自身无法结合任何配体，可与其他 RTK 形成异二聚体，优先与缺乏激酶但与配体结合的 HER3 形成异二聚体。异二聚体的优选配体是heregulin。然而，在升高的水平下，全长和膜结合的 HER2 可以独立地自缔合配体。对于 HER2 过表达的癌症来说，不依赖配体的自缔合与配体依赖的异二聚化总体增加的相对贡献是一个尚未解决的问题。我们工作的一个关键目标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 的自关联接口。我们最近发现，催化失活的 HER3 及其可溶性胞外结构域 (ECD) 在低浓度下会发生自缔合，该反应会被配体结合破坏。我们将利用 HER3 的强自关联及其与 HER2 的高度同源性来识别两种受体中的自关联位点。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 片段互补来直接测量完整细胞中受体关联的程度。我们将使用这些方法来确认 HER3 和 HER2 中已识别的区域，并评估 HER2 的浓度依赖性受体自关联和组成性信号传导与肿瘤形成增强之间的相关性。此外，我们有证据表明 HER3 自关联可能调节调蛋白信号传导。我们将评估 HER3 自缔合的可能调节作用及其对 HER2 配体依赖性和独立信号传导的影响。更好地了解表达水平升高时的特定受体相互作用将增强我们对 HER2 引起的细胞转化的理解，有助于理解和改善赫赛汀等药物的作用，并确定特定且可测量的关联事件作为药物开发的目标。