Here we present WIDOCK, a virtual screening protocol that supports the selection of diverse electrophiles as covalent inhibitors by incorporating ligand reactivity towards cysteine residues into AutoDock4. WIDOCK applies the reactive docking method (Backus et al. in Nature 534:570–574, 2016) and extends it into a virtual screening tool by introducing facile experimental or computational parametrization and a ligand focused evaluation scheme together with a retrospective and prospective validation against various therapeutically relevant targets. Parameters accounting for ligand reactivity are derived from experimental reaction kinetic data or alternatively from computed reaction barriers. The performance of this docking protocol was first evaluated by investigating compound series with diverse warhead chemotypes against KRAS^G12C, MurA and cathepsin B. In addition, WIDOCK was challenged on larger electrophilic libraries screened against OTUB2 and NUDT7. These retrospective analyses showed high sensitivity in retrieving experimental actives, by also leading to superior ROC curves, AUC values and better enrichments than the standard covalent docking tool available in AutoDock4 when compound collections with diverse warheads were investigated. Finally, we applied WIDOCK for the prospective identification of covalent human MAO-A inhibitors acting via a new mechanism by binding to Cys323. The inhibitory activity of several predicted compounds was experimentally confirmed and the labelling of Cys323 was proved by subsequent MS/MS measurements. These findings demonstrate the usefulness of WIDOCK as a warhead-sensitive, covalent virtual screening protocol.

在这里，我们介绍了 WIDOCK，这是一种虚拟筛选协议，通过将配体对半胱氨酸残基的反应性纳入 AutoDock4，支持选择不同的亲电试剂作为共价抑制剂。WIDOCK 应用反应对接方法（Backus et al. in Nature 534:570–574, 2016），并通过引入简便的实验或计算参数化和以配体为重点的评估方案以及回顾性和前瞻性验证将其扩展为虚拟筛选工具各种治疗相关的靶点。考虑配体反应性的参数来自实验反应动力学数据，或者来自计算的反应势垒。该对接协议的性能首先通过研究具有针对 KRAS 的不同弹头化学型的化合物系列进行评估^G12C、MurA 和组织蛋白酶 B。此外，WIDOCK 在针对 OTUB2 和 NUDT7 筛选的更大的亲电子文库中受到挑战。这些回顾性分析显示了在检索实验活性物质方面的高灵敏度，与 AutoDock4 中可用的标准共价对接工具相比，在研究具有不同弹头的化合物集合时，还产生了更好的 ROC 曲线、AUC 值和更好的富集。最后，我们将 WIDOCK 应用于通过与 Cys323 结合的新机制起作用的共价人 MAO-A 抑制剂的前瞻性鉴定。实验证实了几种预测化合物的抑制活性，随后的 MS/MS 测量证明了 Cys323 的标记。这些发现证明了 WIDOCK 作为弹头敏感的共价虚拟筛选协议的有用性。