Understanding EGF receptor activation by growth factors and oncogenic mutations

了解生长因子和致癌突变对 EGF 受体的激活

• 批准号: 9321880
• 负责人: KATHRYN M FERGUSON
• 金额: $ 53.57万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-18 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9321880
• 关键词: Address; Affinity; Allosteric Regulation; Amphiregulin; Binding; Biochemical; Biochemistry; Biophysics; Blood - brain barrier anatomy; Brain; Brain Neoplasms; Cells; Characteristics; Clinical; Communication; Complex; DTR gene; Data; Deuterium; Dimerization; Disease; Dissociation; Epidermal Growth Factor Receptor; Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor; Epiregulin; Erlotinib; Extracellular Domain; Family; Family member; Glioblastoma; Goals; Growth Factor; Hydrogen; Kinetics; Lead; Length; Ligand Binding; Ligands; Link; Malignant - descriptor; Malignant Neoplasms; Malignant neoplasm of lung; Mass Spectrum Analysis; Membrane; Methods; Missense Mutation; Modeling; Molecular Conformation; Mutate; Mutation; Oncogenic; Pattern; Pharmaceutical Preparations; Phosphotransferases; Problem Solving; Property; Receptor Activation; Receptor Protein-Tyrosine Kinases; Receptor Signaling; Regulation; Reporting; Research; Roentgen Rays; Role; Signal Transduction; Specificity; Structure; Tail; Techniques; Technology; Testing; Therapeutic Intervention; Time; Variant; base; betacellulin; dimer; epidermal growth factor receptor VIII; epigen; extracellular; inhibitor/antagonist; insight; kinase inhibitor; lapatinib; preference; public health relevance; receptor; resistance mutation; response; targeted treatment; trafficking

 DESCRIPTION (provided by applicant): The epidermal growth factor receptor (EGFR) is often described as a `prototypic' receptor tyrosine kinase (RTK), and its activation is typically considered to occur through straightforward growth factor-induced receptor dimerization. With such a simple model, however, it is difficult to explain how different activating ligands such as TGFa, betacellulin, HB-EGF, epiregulin, epigen, and amphiregulin can elicit distinct cellular signals through the same receptor - possibly through different patterns of autophosphorylation or through trafficking differences. Moreover, a simple dimerization model cannot explain how mutationally activated forms of EGFR in lung cancer (with kinase mutations) and glioblastoma (with extracellular missense mutations or deletions as in EGFRvIII) would have distinct inhibitor sensitivities, as has been reported. These observations and many others suggest that EGFR is a complex allosterically regulated receptor, which we know binds ligands with negative cooperativity. We are therefore focused on understanding allosteric regulation of this receptor in detail, with a view to understanding EGFR signaling specificity and identifying which oncogenic mutations promote sensitivity to which classes of EGFR inhibitors. These studies may also suggest new strategies for targeted therapeutics. To gain insight into the communication of allosteric changes across the membrane, we propose structural and biochemical studies of EGFR activated by its different ligands - or by oncogenic mutations - using a variety of techniques. Importantly, we will also analyze the receptor's conformational dynamics using hydrogen/deuterium exchange mass spectrometry (HX-MS). HX-MS studies will be performed for isolated extracellular regions of wild-type and mutated receptors and for intact EGFR activated by ligands or mutations - to provide insight into conformational fluctuations in the extracellular domain, kinase domain, and linking regions. We will determine how ligand binding or oncogenic mutation alters the conformation, dynamics, and extent of activation of the intact EGFR, and how these changes are propagated across the membrane. Our preliminary data also suggest that HX-MS analyses will provide valuable insight into the role of the disordered carboxy-terminal tail, and how it might contribute to signaling specificity. Our overall goal is t bridge the current gulf between the structural information about EGFR family members, our understanding of their transmembrane signaling properties in cells, and mechanisms of activation (and inhibition) in cancer. Our Specific Aims address two main questions: 1 how do activated complexes of EGFR induced by its seven different ligands differ in structure, activity, and dynamics? 2 How do oncogenic forms of EGFR found in glioblastoma compare in structure, activity, and dynamics to the ligand-activated receptor - and how do differences lead to altered kinase inhibitor selectivity?

 描述（由适用提供）：表皮生长因子受体（EGFR）通常被描述为“原型”受体酪氨酸激酶（RTK），通常认为其激活是通过直接生长因子诱导的受体二聚化而发生的。但是，使用如此简单的模型，很难解释不同的激活配体，例如TGFA，Betacellulin，HB -EGF，Epiregulin，Epigen和Amphiregulin如何通过同一受体引起不同的细胞信号 - 可能通过自动磷酸化或通过不同模式或通过差异。此外，一个简单的维度模型无法解释肺癌中的EGFR的突变激活形式（具有激酶突变）和胶质母细胞瘤（如EGFRVIII中的细胞外错义突变或缺失）如何具有明显的抑制剂敏感性。这些观察结果和许多其他观察结果表明，EGFR是一种复杂的变构受体，我们知道，它结合了具有负配位的配体。因此，我们专注于理解该受体的变构调节，以了解EGFR信号传导特异性并确定哪些致癌突变促进了对EGFR抑制剂的敏感性。这些研究还可能提出针对性治疗的新策略。为了深入了解整个膜上变构变化的通信，我们提出了使用多种技术通过其不同的配体或致癌突变激活的EGFR的结构和生化研究。重要的是，我们还将使用氢/氘交换质谱法（HX-MS）分析受体的构象动力学。将对野生型和突变受体的分离细胞外区域以及由配体或突变激活的完整EGFR进行HX-MS研究，以洞悉无序的羧基末端尾尾和连接区域的作用。我们将确定配体结合或致癌突变如何改变完整EGFR激活的构象，动力学和程度，以及这些变化如何在整个膜上传播。我们的初步数据还表明，HX-MS分析将为无序的羧基末端尾巴的作用提供宝贵的见解，以及它如何促进信号特异性。我们的总体目标是T桥桥接有关EGFR家族成员的结构信息，我们对细胞中的跨膜信号传导特性的理解以及癌症激活（和抑制）的结构信息之间的当前海湾。我们的具体目的解决了两个主要问题：1由七种不同的结构，活性和动力学不同的配体引起的EGFR的活化复合物如何？ 2在胶质母细胞瘤中发现的EGFR的致癌形式如何比较结构，活性和动力学与配体激活的受体 - 差异如何导致激酶抑制剂的选择性改变？