Structure-Based Design of Eph Receptor Antagonists

描述（由申请人提供）：Eph受体是最大的蛋白酪氨酸激酶家族。Eph受体及其蛋白配体ephrin之间的蛋白-蛋白相互作用以及随后的双向信号传导最近被证明在病理性血管生成以及癌症进展和转移中发挥重要作用。最近对EphB2/ephrin-B2复合物x射线晶体结构的研究使得基于结构设计小肽和拟肽物作为蛋白-蛋白相互作用抑制剂成为可能。对受体-配体复合物的x射线晶体结构的检查发现了受体-配体界面上的一个“热点”，一个由六个氨基酸组成的GHL环深入到Eph受体表面的通道中。对简单二硫链环肽的初步研究表明，含有环残基的短构象限制性环肽可以以亚微摩尔亲和力与EphB2受体结合。本研究设计了新的环状肽类似物和新的肽模拟物，以验证以下两个假设：1)更好的构象限制，更准确地模拟G-HL环与Eph受体结合的构象，可能提供更好的亲和力；2)基于两个亚类ephrin不同保守序列的分子可能导致发现亚类特异性小分子拮抗剂（也可能是激动剂）。两个构象限制性环肽类似物已经在我们的实验室合成，我们的合作者正在评估。本项目的具体目的是：1)确定已合成的构象限制性环肽相对于EphA2和EphA3受体对EphB2和EphB1受体的受体结合亲和力，并将构象限制性环肽中的残基修饰为ephrin- a中常见的残基，以期获得epha特异性的小分子拮抗剂；2)以d -葡萄糖为支架，结合被认为与EphB受体结合至关重要的基团，合成和评估拟肽物；3)合成并评价含有d -葡萄糖支架和环三肽的杂化结构，进一步限制构象，提高对EphB受体的亲和力。成功的拮抗剂可以作为阐明ephdn和Eph在各种生理和病理过程中的功能的药理学探针，并作为潜在的抗血管生成和抗癌药物用于治疗癌症。

DESCRIPTION (provided by applicant): Eph receptors are the largest family of protein tyrosine kinases. The protein-protein interaction between Eph receptors and their protein ligand ephrins and the subsequent bi-directional signaling were recently shown to play an important role in pathological angiogenesis as well as in cancer progression and metastasis. The recent solution of the X-ray crystal structure of EphB2/ephrin-B2 complex made it possible to perform structure-based design of small peptides and peptidomimetics as inhibitors of the protein-protein interaction. Examination of the X-ray crystal structure of the receptor-ligand complex identified a "hot spot" in the receptor-ligand interface, a GHL loop consisting of six amino acids protruding deep into a channel on the Eph receptor surface. Preliminary studies with simple disulfide-linked cyclic peptides indicate that short conformationally restricted cyclic peptides containing the loop residues could bind to the EphB2 receptor with submicromolar affinity. New cyclic peptide analogues and new peptidomimetics were designed in this proposal to test the following two hypotheses: i) better conformational restriction mimicking more accurately the conformation of the G-HL loop as bound to the Eph receptor might offer even better affinity, and ii) molecules based on the distinct conserved sequences of two subclass ephrins might lead to the discovery of subclass-specific small molecule antagonists (possibly also agonists). Two of the conformationally restricted cyclic peptide analogues were already synthesized in our laboratory and are being evaluated by our collaborators. The specific aims of this project are: 1) to determine the receptor binding affinity towards EphB2 and EphB1 receptors of the conformationally restricted cyclic peptides already synthesized, relative to EphA2 and EphA3 receptors, and to modify residues in the conformationally restricted cyclic peptides to those commonly found in ephrin-As in the hope of achieving EphA-specific small molecule antagonists; 2) to synthesize and evaluate peptidomimetics using D-glucose as a scaffold incorporating groups believed to be critical in binding to EphB receptors; and 3) to synthesize and evaluate a hybrid structure containing the D-glucose scaffold and a cyclic tripeptide in efforts to further restrict the conformation and increase the affinity towards EphB receptors. Successful antagonists can be used as pharmacological probes for the elucidation of ephdn and Eph functions in various physiological and pathological processes and as potential antiangiogenic and anticancer agents for the treatment of cancer.