Analysis of active and inactive EGFR conformations

活性和非活性 EGFR 构象分析

• 批准号: 7418929
• 负责人: KATHRYN M FERGUSON
• 金额: $ 23.75万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7418929
• 关键词: Affect; Affinity; Agonist; Antibodies; Antineoplastic Agents; Binding; Cell surface; Cetuximab; Characteristics; Clinical Trials; Colon Carcinoma; Colorectal Cancer; Complex; Dimerization; Doctor of Philosophy; Drug Delivery Systems; EGF gene; Epidermal Growth Factor; Epidermal Growth Factor Receptor; Epitopes; ErbB Receptor Family Protein; Erbitux; Extracellular Domain; Fab Immunoglobulins; Family; Fc Receptor; Goals; Growth Factor; Human; Knowledge; Learning; Libraries; Ligand Binding; Ligands; Malignant Neoplasms; Molecular Conformation; Mutation; Personal Satisfaction; Phage Display; Receptor Activation; Receptor Protein-Tyrosine Kinases; Receptor Signaling; Regulation; Relative (related person); Research Personnel; Signal Transduction; Site; Specificity; Structure; X-Ray Crystallography; base; cancer therapy; extracellular; improved; in vivo; inhibiting antibody; insight; member; mutant; novel; prevent; programs; receptor; receptor function; tumor growth

DESCRIPTION (provided by applicant): The Epidermal Growth Factor (EGF) receptor or ErbB family of receptor tyrosine kinases (RTKs) have been implicated in many human cancers. It is well established that antibodies to the extracellular region of the EGF receptor (EGFR) that prevent ligand binding and/or receptor signaling can inhibit tumor growth in vivo. At least 5 such anti-EGFR antibodies are in clinical trials; 1, cetuximab/Erbitux, was FDA-approved to treat advanced colon cancer in February 2004. Recent structural studies of EGFR family extracellular regions have identified a novel mechanism for ligand induced receptor dimerization. The unliganded receptor is found in a characteristic "autoinhibited" configuration in which the receptor dimerization interface is completely occluded in an intramolecular "tether". Ligand binding induces a dramatic conformational change in EGFR that exposes this normally buried extracellular dimerization site, promoting dimerization and receptor activation. We will analyze how particular antibodies against the EGFR extracellular region (sEGFR) can have dramatically different effects. Certain antibodies inhibit EGFR activation, while others alter (or bias) the affinity state of the receptor at the cell surface, sometimes allowing signaling to proceed. Still others specifically impair binding of 1 ligand without affecting binding of another. We hypothesize that these different effects reflect the distinct extracellular conformations stabilized by the particular antibodies. We will analyze antibody binding to sEGFR mutants with defined altered conformations, and use X-ray crystallography to determine structures of Fab:sEGFR complexes. Finally, in an effort to exploit recent structural advances, we will screen phage display libraries for single-chain Fv (scFv) molecules that bind to key epitopes in sEGFR and that we hypothesize will inhibit EGFR in novel ways. We hope to gain structural insight into relationships between extracellular conformation and activity, affinity and specificity of EGFR at the cell surface. The specific aims are: 1. To investigate relationships between EGFR responsiveness and extracellular domain conformation by analyzing the structural consequences of binding antibodies with known effects on in vivo receptor function. 2. To probe ligand specificity in EGFR activation using an antibody that selectively inhibits binding of a subset of EGFR agonists.

描述（由申请人提供）：表皮生长因子（EGF）受体或受体酪氨酸激酶（RTK）的ErbB家族与许多人类癌症有关。已经确定，针对EGF受体（EGFR）的细胞外区域的阻止配体结合和/或受体信号传导的抗体可以在体内抑制肿瘤生长。至少有5种这样的抗EGFR抗体正在进行临床试验; 1种是西妥昔单抗/爱必妥，于2004年2月被FDA批准用于治疗晚期结肠癌。EGFR家族细胞外区的最新结构研究已经确定了配体诱导受体二聚化的新机制。未配体的受体被发现在一个特征性的“自抑制”的配置，其中受体二聚化界面是完全封闭在分子内的“系链”。配体结合诱导EGFR发生显著的构象变化，从而暴露该通常埋藏的细胞外二聚化位点，促进二聚化和受体活化。我们将分析针对EGFR胞外区（sEGFR）的特定抗体如何产生显著不同的效果。某些抗体抑制EGFR活化，而另一些抗体改变（或偏向）细胞表面受体的亲和力状态，有时允许信号传导进行。还有一些特异性地损害1个配体的结合，而不影响另一个配体的结合。我们假设这些不同的效果反映了特定抗体稳定的不同细胞外构象。我们将分析抗体与sEGFR突变体的结合，并使用X射线晶体学来确定Fab：sEGFR复合物的结构。最后，在努力利用最近的结构进展，我们将筛选噬菌体展示库的单链抗体（scFv）分子结合的sEGFR的关键表位，我们假设将抑制EGFR以新的方式。我们希望获得结构洞察细胞外构象和活性之间的关系，在细胞表面的EGFR的亲和力和特异性。具体目标是：1.通过分析已知对体内受体功能有影响的结合抗体的结构结果，研究EGFR反应性与细胞外结构域构象之间的关系。2.使用选择性抑制EGFR激动剂亚组结合的抗体探测EGFR激活中的配体特异性。