ABSTRACT
There is a clinical need for direct-acting antivirals targeting SARS-CoV-2, the coronavirus responsible for the COVID-19 pandemic, to complement current therapeutic strategies. The main protease (Mpro) is an attractive target for antiviral therapy. However, the vast majority of protease inhibitors described thus far are peptidomimetic and bind to the active-site cysteine via a covalent adduct, which is generally pharmacokinetically unfavorable. We have reported the optimization of an existing FDA-approved chemical scaffold, perampanel, to bind to and inhibit Mpro noncovalently with IC50s in the low-nanomolar range and EC50s in the low-micromolar range. Here, we present nine crystal structures of Mpro bound to a series of perampanel analogs, providing detailed structural insights into their mechanism of action and structure-activity relationship. These insights further reveal strategies for pursuing rational inhibitor design efforts in the context of considerable active-site flexibility and potential resistance mechanisms.
Subject(s)
Antiviral Agents/chemistry , Coronavirus 3C Proteases/chemistry , Protease Inhibitors/chemistry , Pyridones/chemistry , SARS-CoV-2/enzymology , Antiviral Agents/pharmacology , COVID-19/virology , Catalytic Domain , Coronavirus 3C Proteases/antagonists & inhibitors , Coronavirus 3C Proteases/metabolism , Drug Design , Molecular Dynamics Simulation , Molecular Structure , Nitriles , Protease Inhibitors/pharmacology , Protein Conformation , Protein Multimerization , SARS-CoV-2/drug effects , SARS-CoV-2/physiology , COVID-19 Drug TreatmentABSTRACT
Effective control of COVID-19 requires antivirals directed against SARS-CoV-2. We assessed 10 hepatitis C virus (HCV) protease-inhibitor drugs as potential SARS-CoV-2 antivirals. There is a striking structural similarity of the substrate binding clefts of SARS-CoV-2 main protease (Mpro) and HCV NS3/4A protease. Virtual docking experiments show that these HCV drugs can potentially bind into the Mpro substrate-binding cleft. We show that seven HCV drugs inhibit both SARS-CoV-2 Mpro protease activity and SARS-CoV-2 virus replication in Vero and/or human cells. However, their Mpro inhibiting activities did not correlate with their antiviral activities. This conundrum is resolved by demonstrating that four HCV protease inhibitor drugs, simeprevir, vaniprevir, paritaprevir, and grazoprevir inhibit the SARS CoV-2 papain-like protease (PLpro). HCV drugs that inhibit PLpro synergize with the viral polymerase inhibitor remdesivir to inhibit virus replication, increasing remdesivir's antiviral activity as much as 10-fold, while those that only inhibit Mpro do not synergize with remdesivir.
Subject(s)
Antiviral Agents/pharmacology , COVID-19 Drug Treatment , Coronavirus Papain-Like Proteases/antagonists & inhibitors , SARS-CoV-2/drug effects , SARS-CoV-2/enzymology , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Alanine/analogs & derivatives , Alanine/pharmacology , COVID-19/virology , Cell Culture Techniques , Cell Line , Coronavirus Papain-Like Proteases/metabolism , Drug Repositioning/methods , Drug Synergism , Hepacivirus/drug effects , Hepatitis C/drug therapy , Humans , Molecular Docking Simulation , Molecular Dynamics Simulation , Protease Inhibitors/pharmacology , Virus Replication/drug effectsABSTRACT
The COVID-19 pandemic caused by SARS-CoV-2 requires rapid development of specific therapeutics and vaccines. The main protease of SARS-CoV-2, 3CL Mpro, is an established drug target for the design of inhibitors to stop the virus replication. Repurposing existing clinical drugs can offer a faster route to treatments. Here, we report on the binding mode and inhibition properties of several inhibitors using room temperature X-ray crystallography and in vitro enzyme kinetics. The enzyme active-site cavity reveals a high degree of malleability, allowing aldehyde leupeptin and hepatitis C clinical protease inhibitors (telaprevir, narlaprevir, and boceprevir) to bind and inhibit SARS-CoV-2 3CL Mpro. Narlaprevir, boceprevir, and telaprevir are low-micromolar inhibitors, whereas the binding affinity of leupeptin is substantially weaker. Repurposing hepatitis C clinical drugs as COVID-19 treatments may be a useful option to pursue. The observed malleability of the enzyme active-site cavity should be considered for the successful design of specific protease inhibitors.