受体酪氨酸激酶 EphB3 的特异性不可逆抑制剂的开发,Journal of the American Chemical Society

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受体酪氨酸激酶 EphB3 的特异性不可逆抑制剂的开发
Journal of the American Chemical Society ( IF 14.4 ) Pub Date : 2016-08-11 , DOI: 10.1021/jacs.6b05483
Alvin Kung , Ying-Chu Chen , Marianne Schimpl ₁ , Feng Ni , Jianfa Zhu , Maurice Turner , Henrik Molina ₂ , Ross Overman ₃ , Chao Zhang

Affiliation

产生促红细胞生成素的人肝细胞癌 (Eph) 受体酪氨酸激酶 (RTK) 调节各种动态细胞事件，包括细胞突出、迁移、增殖和细胞命运决定。Eph 激酶的小分子抑制剂是剖析 Eph 生理和病理作用的宝贵工具。然而，由于家族内的高度同源性，缺乏对单个 Eph 异构体具有选择性的小分子抑制剂。在此，我们报告了单一 Eph 异构体 EphB3 的第一个有效和特异性抑制剂的开发。通过结构生物信息学分析，我们在 EphB3 激酶结构域的铰链区发现了一个半胱氨酸，这是任何其他人类激酶都没有的特征。我们合成并表征了一系列亲电子喹唑啉，以针对 EphB3 中这种独特的反应性特征。一些亲电子喹唑啉在体外和细胞中选择性地和有效地抑制 EphB3。EphB3 与两种喹唑啉复合物的共晶结构证实了蛋白质和抑制剂之间的共价键。创建并使用优化抑制剂的“可点击”版本来验证整个蛋白质组中的特定靶标参与，并探测现有 EphB3 抑制剂靶标参与的程度和动力学。此外，我们证明 EphB3 在近膜区域内的自磷酸化发生在使用特定抑制剂的反式中。这些精巧的特异性抑制剂将促进 EphB3' 的解剖

"点击查看英文标题和摘要"

Development of Specific, Irreversible Inhibitors for a Receptor Tyrosine Kinase EphB3

Erythropoietin-producing human hepatocellular carcinoma (Eph) receptor tyrosine kinases (RTKs) regulate a variety of dynamic cellular events, including cell protrusion, migration, proliferation, and cell-fate determination. Small-molecule inhibitors of Eph kinases are valuable tools for dissecting the physiological and pathological roles of Eph. However, there is a lack of small-molecule inhibitors that are selective for individual Eph isoforms due to the high homology within the family. Herein, we report the development of the first potent and specific inhibitors of a single Eph isoform, EphB3. Through structural bioinformatic analysis, we identified a cysteine in the hinge region of the EphB3 kinase domain, a feature that is not shared with any other human kinases. We synthesized and characterized a series of electrophilic quinazolines to target this unique, reactive feature in EphB3. Some of the electrophilic quinazolines selectively and potently inhibited EphB3 both in vitro and in cells. Cocrystal structures of EphB3 in complex with two quinazolines confirmed the covalent linkage between the protein and the inhibitors. A "clickable" version of an optimized inhibitor was created and employed to verify specific target engagement in the whole proteome and to probe the extent and kinetics of target engagement of existing EphB3 inhibitors. Furthermore, we demonstrate that the autophosphorylation of EphB3 within the juxtamembrane region occurs in trans using a specific inhibitor. These exquisitely specific inhibitors will facilitate the dissection of EphB3's role in various biological processes and disease contribution.

更新日期：2016-08-11

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