IA类磷脂酰肌醇3激酶α（PI3Kα）是一个重要的药物靶点，因为它是人类癌症中最常见的突变蛋白之一。然而，目前市场上或正在开发中的小分子抑制剂由于缺乏选择性存在安全问题。因此，需要其他化学支架或独特的催化激酶抑制机制。在这里，我们报道了野生型PI3Kα的冷冻电子显微镜结构，即p110α和p85α的二聚体，与三种亲和力不同且对激酶活性无抑制作用的Y形配体[化合物16（cpd16），化合物17（cpd17）和化合物18（cpd18）]结合。与ATP竞争性抑制剂不同，cpd17采用了一种Y形构象，其中一个ARM插入由R770和W780形成的结合口袋，另一个ARM以与ATP磷酸尾不同的角度卡在ATP结合口袋中。这种特殊的相互作用引发了与未受配体结合的蛋白类似的PI3Kα构象。这些观察结果也适用于两个同分异构体（cpd16和cpd18）。对这些Y形配体的进一步分析揭示了立体或位置化学修饰引起的差异结合亲和力的结构基础。我们的研究结果可能为设计针对PI3Kα的治疗药物提供不同的方向。

Class IA phosphoinositide 3-kinase alpha (PI3Kα) is an important drug target because it is one of the most frequently mutated proteins in human cancers. However, small molecule inhibitors currently on the market or under development have safety concerns due to a lack of selectivity. Therefore, other chemical scaffolds or unique mechanisms of catalytic kinase inhibition are needed. Here, we report the cryo-electron microscopy structures of wild-type PI3Kα, the dimer of p110α and p85α, in complex with three Y-shaped ligands [cpd16 (compound 16), cpd17 (compound 17), and cpd18 (compound 18)] of different affinities and no inhibitory effect on the kinase activity. Unlike ATP-competitive inhibitors, cpd17 adopts a Y-shaped conformation with one arm inserted into a binding pocket formed by R770 and W780 and the other arm lodged in the ATP-binding pocket at an angle that is different from that of the ATP phosphate tail. Such a special interaction induces a conformation of PI3Kα resembling that of the unliganded protein. These observations were confirmed with two isomers (cpd16 and cpd18). Further analysis of these Y-shaped ligands revealed the structural basis of differential binding affinities caused by stereo- or regiochemical modifications. Our results may offer a different direction toward the design of therapeutic agents against PI3Kα.