Identification of Determinants Required for Agonistic and Inverse Agonistic Ligand Properties at the ADP Receptor P2Y12
Identification of Determinants Required for Agonistic and Inverse Agonistic Ligand Properties at the ADP Receptor P2Y12
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DOI：
10.1124/mol.112.082198
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发表时间：
2013-01-01
影响因子：
3.6
通讯作者：
Schoeneberg, Torsten
中科院分区：
文献类型：
--
作者：
Schmidt, Philipp;Ritscher, Lars;Schoeneberg, Torsten
ADP受体P2 Y（12）属于G蛋白偶联受体（GPCR）超家族，其激活触发血小板聚集。因此，强效拮抗剂（如氯吡格雷）在预防和治疗血栓栓塞事件方面具有高度临床相关性。P2 Y（12）在体外显示出升高的基础活性，因此，与拮抗剂相比，反向激动剂可能在治疗上是有益的。仅描述了少数P2 Y（12）的反向激动剂。为了扩展这种有限的化学空间并提高对反向激动剂-受体相互作用的结构决定因素的理解，本研究使用野生型（WT）人P2 Y（12）和28种组成型活性突变体筛选了嘌呤化合物文库的前导结构。结果表明，ATP和ATP衍生物是P2 Y的激动剂（12）。在P2 Y（12）处的效力为2-（甲硫基）-ADP > 2-（甲硫基）-ATP > ADP > ATP。确定了激动性配体活性所需的决定因素。分子对接研究揭示了ATP衍生物的结合口袋，其与人P2 Y中的跨膜螺旋3、5、6和7邻接（12），其中Y-105、E-188、R-256、Y-259和K-280在配体相互作用中发挥特别重要的作用。2 '-脱氧核糖的3'-OH处的N-甲基-邻氨基苯甲酰修饰产生具有反向激动剂活性的配体（mant-deoxy-ATP [dATP]，mant-deoxy-ADP）。在WT人P2 Y（12）和一半组成型活性P2 Y（12）突变体中发现mant-dATP的反向激动剂活性。这项研究表明，除了ADP和ATP，其他ATP衍生物不仅是P2 Y（12）的配体，而且是激动剂。ATP内核糖的修饰可导致ATP衍生配体的反向活性。
The ADP receptor P2Y(12) belongs to the superfamily of G protein-coupled receptors (GPCRs), and its activation triggers platelet aggregation. Therefore, potent antagonists, such as clopidogrel, are of high clinical relevance in prophylaxis and treatment of thromboembolic events. P2Y(12) displays an elevated basal activity in vitro, and as such, inverse agonists may be therapeutically beneficial compared with antagonists. Only a few inverse agonists of P2Y(12) have been described. To expand this limited chemical space and improve understanding of structural determinants of inverse agonist-receptor interaction, this study screened a purine compound library for lead structures using wild-type (WT) human P2Y(12) and 28 constitutively active mutants. Results showed that ATP and ATP derivatives are agonists at P2Y(12). The potency at P2Y(12) was 2-(methylthio)-ADP > 2-(methylthio)-ATP > ADP > ATP. Determinants required for agonistic ligand activity were identified. Molecular docking studies revealed a binding pocket for the ATP derivatives that is bordered by transmembrane helices 3, 5, 6, and 7 in human P2Y(12), with Y-105, E-188, R-256, Y-259, and K-280 playing a particularly important role in ligand interaction. N-Methyl-anthraniloyl modification at the 3'-OH of the 2'-deoxyribose leads to ligands (mant-deoxy-ATP [dATP], mant-deoxy-ADP) with inverse agonist activity. Inverse agonist activity of mant-dATP was found at the WT human P2Y(12) and half of the constitutive active P2Y(12) mutants. This study showed that, in addition to ADP and ATP, other ATP derivatives are not only ligands of P2Y(12) but also agonists. Modification of the ribose within ATP can result in inverse activity of ATP-derived ligands.