A new coronavirus (SARS-CoV-2) has been identified as the etiologic agent for the COVID-19 outbreak. Currently, effective treatment options remain very limited for this disease; therefore, there is an urgent need to identify new anti-COVID-19 agents. In this study, we screened over 6,000 compounds that included approved drugs, drug candidates in clinical trials, and pharmacologically active compounds to identify leads that target the SARS-CoV-2 papain-like protease (PLpro). Together with main protease (M^pro), PLpro is responsible for processing the viral replicase polyprotein into functional units. Therefore, it is an attractive target for antiviral drug development. Here we discovered four compounds, YM155, cryptotanshinone, tanshinone I and GRL0617 that inhibit SARS-CoV-2 PLpro with IC₅₀ values ranging from 1.39 to 5.63 μmol/L. These compounds also exhibit strong antiviral activities in cell-based assays. YM155, an anticancer drug candidate in clinical trials, has the most potent antiviral activity with an EC₅₀ value of 170 nmol/L. In addition, we have determined the crystal structures of this enzyme and its complex with YM155, revealing a unique binding mode. YM155 simultaneously targets three “hot” spots on PLpro, including the substrate-binding pocket, the interferon stimulating gene product 15 (ISG15) binding site and zinc finger motif. Our results demonstrate the efficacy of this screening and repurposing strategy, which has led to the discovery of new drug leads with clinical potential for COVID-19 treatments.

一种新的冠状病毒（SARS-CoV-2）已被确定为 COVID-19 爆发的病原体。目前，这种疾病的有效治疗选择仍然非常有限；因此，迫切需要确定新的抗 COVID-19 药物。在这项研究中，我们筛选了 6,000 多种化合物，包括已批准的药物、临床试验中的候选药物和药理活性化合物，以确定靶向 SARS-CoV-2 类木瓜蛋白酶 (PLpro) 的先导化合物。PLpro与主蛋白酶 (M ^pro ) 一起负责将病毒复制酶多蛋白加工成功能单元。因此，它是抗病毒药物开发的一个有吸引力的目标。在这里，我们发现了四种化合物 YM155、隐丹参酮、丹参酮 I 和 GRL0617，它们以 IC _{50抑制 SARS-CoV-2 PLpro}值范围为 1.39 至 5.63 μmol/L。这些化合物在基于细胞的测定中也表现出很强的抗病毒活性。YM155是临床试验中的抗癌候选药物，具有最强的抗病毒活性，EC ₅₀值为170 nmol/L。此外，我们已经确定了这种酶的晶体结构及其与 YM155 的复合物，揭示了一种独特的结合模式。YM155 同时靶向 PLpro 上的三个“热点”，包括底物结合袋、干扰素刺激基因产物 15 (ISG15) 结合位点和锌指基序。我们的结果证明了这种筛选和再利用策略的有效性，这导致发现了具有 COVID-19 治疗临床潜力的新药物先导。