Expanding the utilization of binding pockets proves to be effective for noncovalent small molecule inhibitors against SARS-CoV-2 Mpro

Abstract

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in millions of deaths and continues to pose serious threats to global public health. The main protease (M^pro) of SARS-CoV-2 is crucial for viral replication and its conservation, making it an attractive drug target. Here, we employed a structure-based drug design strategy to develop and optimize novel inhibitors targeting SARS-CoV-2 M^pro. By fully exploring occupation of the S1, S2, and S3/S4 binding pockets, we identified eight promising inhibitors with half-maximal inhibitory concentration (IC₅₀) values below 20 nM. The cocrystal structure of M^pro with compound 10 highlighted the crucial roles of the interactions within the S3/S4 pockets in inhibitor potency enhancement. These findings demonstrated that expanding the utilization of these binding pockets was an effective strategy for developing noncovalent small molecule inhibitors that target SARS-CoV-2 M^pro. Compound 4 demonstrated outstanding in vitro antiviral activity against wild-type SARS-CoV-2 with an EC₅₀ of 9.4 nM. Moreover, oral treatment with compounds 1 and 9 exhibited excellent antiviral potency and substantially ameliorated virus-induced tissue damage in the lungs of Omicron BA.5-infected K18-human ACE2 (K18-hACE2) transgenic mice, indicating that these novel noncovalent inhibitors could be potential oral agents for the treatment of COVID-19.