Department of Pharmaceutical and Biomedical Sciences, University of Georgia, College of Pharmacy, 240 W. Green St, Athens, GA 30602, USA.
Department of Biochemistry and Molecular Biology, University of Georgia, 120 Green St, Athens, GA 30602, USA; Institute of Bioinformatics, University of Georgia, 120 Green St, Athens, GA 30602, USA.
Bioorg Med Chem. 2018 Mar 15;26(6):1167-1173. doi: 10.1016/j.bmc.2017.08.051. Epub 2017 Sep 5.
Although EGFR is a highly sought-after drug target, inhibitor resistance remains a challenge. As an alternative strategy for kinase inhibition, we sought to explore whether allosteric activation mechanisms could effectively be disrupted. The kinase domain of EGFR forms an atypical asymmetric dimer via head-to-tail interactions and serves as a requisite for kinase activation. The kinase dimer interface is primarily formed by the H-helix derived from one kinase monomer and the small lobe of the second monomer. We hypothesized that a peptide designed to resemble the binding surface of the H-helix may serve as an effective disruptor of EGFR dimerization and activation. A library of constrained peptides was designed to mimic the H-helix of the kinase domain and interface side chains were optimized using molecular modeling. Peptides were constrained using peptide "stapling" to structurally reinforce an alpha-helical conformation. Peptide stapling was demonstrated to notably enhance cell permeation of an H-helix derived peptide termed EHBI2. Using cell-based assays, EHBI2 was further shown to significantly reduce EGFR activity as measured by EGFR phosphorylation and phosphorylation of the downstream signaling substrate Akt. To our knowledge, this is the first H-helix-based compound targeting the asymmetric interface of the kinase domain that can successfully inhibit EGFR activation and signaling. This study presents a novel, alternative targeting site for allosteric inhibition of EGFR.
虽然 EGFR 是一个备受关注的药物靶点,但抑制剂耐药性仍然是一个挑战。作为激酶抑制的替代策略,我们试图探索变构激活机制是否能够有效地被破坏。EGFR 的激酶结构域通过头到尾相互作用形成一个非典型的不对称二聚体,是激酶激活的必要条件。激酶二聚体界面主要由一个激酶单体的 H 螺旋和第二个单体的小 lobe 形成。我们假设,设计成类似于 H 螺旋结合表面的肽可能成为有效破坏 EGFR 二聚化和激活的物质。设计了一个约束肽文库来模拟激酶结构域的 H 螺旋,使用分子建模优化了界面侧链。使用肽“订书钉”来约束肽,以增强其α螺旋构象。肽订书钉显著提高了一种称为 EHBI2 的 H 螺旋衍生肽的细胞通透性。通过细胞测定,EHBI2 进一步显示可显著降低 EGFR 活性,如 EGFR 磷酸化和下游信号底物 Akt 的磷酸化所测量的。据我们所知,这是第一个靶向激酶结构域不对称界面的基于 H 螺旋的化合物,可以成功抑制 EGFR 的激活和信号转导。本研究提出了一种新的、替代的变构抑制 EGFR 的靶点。