The coronavirus disease 2019 (COVID-19) pandemic has necessitated the development of antiviral agents against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The main protease (Mpro) is a promising target for...

The coronavirus disease 2019 (COVID-19) pandemic has necessitated the development of antiviral agents against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). 3C-like protease (3CLpro) is a promising target for COVID-19 treatment. Here, we report a new class of covalent inhibitors of 3CLpro that possess chlorofluoroacetamide (CFA) as a cysteine-reactive warhead. Based on an aza-peptide scaffold, we synthesized a series of CFA derivatives in enantiopure form and evaluated their biochemical efficiency. The data revealed that 8a (YH-6) with the R configuration at the CFA unit strongly blocks SARS-CoV-2 replication in infected cells, and its potency is comparable to that of nirmatrelvir. X-ray structural analysis showed that YH-6 formed a covalent bond with Cys145 at the catalytic center of 3CLpro. The strong antiviral activity and favorable pharmacokinetic properties of YH-6 suggest its potential as a lead compound for the treatment of COVID-19.

The pandemic of COVID-2019 has urged the development of antiviral agents against its causative pathogen SARS-CoV-2. The main protease (Mpro), a cysteine protease essential for viral replication, is a promising protein target. Here we report an irreversible SARS-CoV-2 Mpro inhibitor possessing chlorofluoroacetamide (CFA) as the warhead for covalent modification of Mpro. Ugi multi-component reaction employing chlorofluoroacetic acid allowed rapid generation of CFA derivatives, of which diastereomers displayed significantly different inhibitory activity against Mpro. We established a practical protocol for the optical resolution of chlorofluoroacetic acid, which enable the isolation of the stereoisomers of the best CFA compound. Kinetic analysis revealed that (R)-CFA is crucial for both binding affinity and the rate of irreversible inactivation of Mpro. Our findings highlight the prominent influence of the CFA chirality on the covalent modification of cysteine, and provide the basis for improving the potency and selectivity in the development of novel CFA-based covalent inhibitors.

The pandemic of COVID-2019 has urged the development of antiviral agents against its causative pathogen SARS-CoV-2. The main protease (Mpro), a cysteine protease essential for viral replication, is a promising protein target. Here we report an irreversible SARS-CoV-2 Mpro inhibitor possessing chlorofluoroacetamide (CFA) as the warhead for covalent modification of Mpro. Ugi multi-component reaction employing chlorofluoroacetic acid allowed rapid generation of CFA derivatives, of which diastereomers displayed significantly different inhibitory activity against Mpro. We established a practical protocol for the optical resolution of chlorofluoroacetic acid, which enable the isolation of the stereoisomers of the best CFA compound. Kinetic analysis revealed that (R)-CFA is crucial for both binding affinity and the rate of irreversible inactivation of Mpro. Our findings highlight the prominent influence of the CFA chirality on the covalent modification of cysteine, and provide the basis for improving the potency and selectivity in the development of novel CFA-based covalent inhibitors.

The pandemic of coronavirus disease 2019 (COVID-19) has urgently necessitated the development of antiviral agents against severe acute respiratory syndrome coronavirus 2 (SARSCoV-2). The 3C-like protease (3CLpro) is a promising target for COVID-19 treatment. Here, we report the new class of covalent inhibitors for 3CLpro possessing chlorofluoroacetamide (CFA) as a cysteine reactive warhead. Based on the aza-peptide scaffold, we synthesized the series of CFA derivatives in enantiopure form and evaluated their biochemical efficiencies. The data revealed that 8a (YH-6) with R configuration at the CFA unit strongly blocks the SARS-CoV-2 replication in the infected cells and this potency is comparable to that of nirmatrelvir. The X-ray structural analysis shows that 8a (YH-6) forms a covalent bond with Cys145 at the catalytic center of 3CLpro. The strong antiviral activity and sufficient pharmacokinetics property of 8a (YH-6) suggest its potential as a lead compound for treatment of COVID-19.