A new coronavirus (CoV) identified as COVID-19 virus is the etiological agent responsible for the 2019-2020 viral pneumonia outbreak that commenced in Wuhan^1–4. Currently there are no targeted therapeutics and effective treatment options remain very limited. In order to rapidly discover lead compounds for clinical use, we initiated a program of combined structure-assisted drug design, virtual drug screening and high-throughput screening to identify new drug leads that target the COVID-19 virus main protease (M^pro). M^pro is a key CoV enzyme, which plays a pivotal role in mediating viral replication and transcription, making it an attractive drug target for this virus^5,6. Here, we identified a mechanism-based inhibitor, N3, by computer-aided drug design and subsequently determined the crystal structure of COVID-19 virus M^pro in complex with this compound. Next, through a combination of structure-based virtual and high-throughput screening, we assayed over 10,000 compounds including approved drugs, drug candidates in clinical trials, and other pharmacologically active compounds as inhibitors of M^pro. Six of these compounds inhibited M^pro with IC₅₀ values ranging from 0.67 to 21.4 μM. Ebselen also exhibited promising antiviral activity in cell-based assays. Our results demonstrate the efficacy of this screening strategy, which can lead to the rapid discovery of drug leads with clinical potential in response to new infectious diseases for which no specific drugs or vaccines are available.

The COVID-19 RNA-synthesizing machine: Many in the scientific community have mobilized to understand the virus that is causing the global coronavirus disease 2019 (COVID-19) pandemic. Gao et al. focused on a complex that plays a key role in the replication and transcription cycle of the virus. They used cryo–electron microscopy to determine a 2.9-angstrom-resolution structure of the RNA-dependent RNA polymerase nsp12, which catalyzes the synthesis of viral RNA, in complex with two cofactors, nsp7 and nsp8. nsp12 is a target for nucleotide analog antiviral inhibitors such as remdesivir, and the structure may provide a basis for designing new antiviral therapeutics. Science , this issue p. 779

Promising antiviral protease inhibitors: With no vaccine or proven effective drug against the virus that causes coronavirus disease 2019 (COVID-19), scientists are racing to find clinical antiviral treatments. A promising drug target is the viral main protease M ^pro , which plays a key role in viral replication and transcription. Dai et al. designed two inhibitors, 11a and 11b, based on analyzing the structure of the M ^pro active site. Both strongly inhibited the activity of M ^pro and showed good antiviral activity in cell culture. Compound 11a had better pharmacokinetic properties and low toxicity when tested in mice and dogs, suggesting that this compound is a promising drug candidate. Science , this issue p. 1331

Nirmatrelvir, an oral antiviral targeting the 3CL protease of SARS-CoV-2, has been demonstrated to be clinically useful against COVID-19 (refs. ^1,2). However, because SARS-CoV-2 has evolved to become resistant to other therapeutic modalities^3–9, there is a concern that the same could occur for nirmatrelvir. Here we examined this possibility by in vitro passaging of SARS-CoV-2 in nirmatrelvir using two independent approaches, including one on a large scale. Indeed, highly resistant viruses emerged from both and their sequences showed a multitude of 3CL protease mutations. In the experiment peformed with many replicates, 53 independent viral lineages were selected with mutations observed at 23 different residues of the enzyme. Nevertheless, several common mutational pathways to nirmatrelvir resistance were preferred, with a majority of the viruses descending from T21I, P252L or T304I as precursor mutations. Construction and analysis of 13 recombinant SARS-CoV-2 clones showed that these mutations mediated only low-level resistance, whereas greater resistance required accumulation of additional mutations. E166V mutation conferred the strongest resistance (around 100-fold), but this mutation resulted in a loss of viral replicative fitness that was restored by compensatory changes such as L50F and T21I. Our findings indicate that SARS-CoV-2 resistance to nirmatrelvir does readily arise via multiple pathways in vitro, and the specific mutations observed herein form a strong foundation from which to study the mechanism of resistance in detail and to inform the design of next-generation protease inhibitors.

The antineoplastic drug carmofur is shown to inhibit the SARS-CoV-2 main protease (M^pro). Here, the X-ray crystal structure of M^pro in complex with carmofur reveals that the carbonyl reactive group of carmofur is covalently bound to catalytic Cys145, whereas its fatty acid tail occupies the hydrophobic S2 subsite. Carmofur inhibits viral replication in cells (EC₅₀ = 24.30 μM) and is a promising lead compound to develop new antiviral treatment for COVID-19.