Replication capacity and susceptibility of nirmatrelvir-resistant mutants to next-generation Mpro inhibitors in a SARS-CoV-2 replicon system

Chieh-Wen Lo; Omri Kariv; Chenzhou Hao; Karen Anbro Gammeltoft; Jens Bukh; Judith Gottwein; Michael Westberg; Michael Z Lin; Shirit Einav

doi:10.1016/j.antiviral.2024.106022

Replication capacity and susceptibility of nirmatrelvir-resistant mutants to next-generation Mpro inhibitors in a SARS-CoV-2 replicon system

Antiviral Res. 2024 Nov:231:106022. doi: 10.1016/j.antiviral.2024.106022. Epub 2024 Oct 17.

Authors

Chieh-Wen Lo¹, Omri Kariv¹, Chenzhou Hao², Karen Anbro Gammeltoft³, Jens Bukh³, Judith Gottwein³, Michael Westberg⁴, Michael Z Lin⁵, Shirit Einav⁶

Affiliations

¹ Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University, Stanford, CA, USA.
² Department of Neurobiology, Stanford University, Stanford, CA, USA.
³ Copenhagen Hepatitis C Program (CO-HEP), Department of Infectious Diseases, Copenhagen University Hospital, Hvidovre, Denmark; Copenhagen Hepatitis C Program (CO-HEP), Department of Immunology and Microbiology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.
⁴ Department of Neurobiology, Stanford University, Stanford, CA, USA; Department of Chemistry, Aarhus University, Aarhus, Denmark; Interdisciplinary Nanoscience Center (iNANO), Aarhus University, Aarhus, Denmark.
⁵ Department of Neurobiology, Stanford University, Stanford, CA, USA; Department of Bioengineering, Stanford University, Stanford, CA, USA; Department of Chemical and Systems Biology, Stanford University, Stanford, CA, USA.
⁶ Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University, Stanford, CA, USA; Chan Zuckerberg Biohub, San Francisco, CA, USA; Department of Microbiology and Immunology, Stanford University, CA, USA. Electronic address: seinav@stanford.edu.

PMID: 39424074
DOI: 10.1016/j.antiviral.2024.106022

Abstract

There is an ongoing need to expand the anti-SARS-CoV-2 armamentarium to include agents capable of suppressing replication of drug-resistant mutants emerging during monotherapy with approved direct-acting antivirals. Using a subgenomic SARS-CoV-2 replicon system, we studied the RNA replication capacity of nirmatrelvir (NTV)-resistant mutants and their susceptibility to next-generation Mpro inhibitors, including ibuzatrelvir (ITV), ensitrelvir (ETV), and ML2006a4. Our findings revealed that E166V Mpro mutants reduced viral RNA replication, whereas other Mpro mutations retained or increased the replication capacity, suggesting the potential of the latter to dominate under NTV selective pressure. Except for having an advantage against E166A mutants, ITV largely showed the same mutational sensitivity as NTV. ETV was more effective than NTV against E166V mutants but less effective against S144A, E166A, and L167F mutants. ML2006a4 demonstrated the most effective suppression across most mutants (S144A, E166V, S144A + L50F, E166 A/V + L50F, L167F + L50F, and E166A + L167F + L50F). Thus, ML2006a4 represents an attractive investigational candidate against clinically relevant NTV-resistant SARS-CoV-2 mutants.

Keywords: Ensitrelvir; Ibuzatrelvir; ML2006a4; Mpro inhibitors; Nirmatrelvir; Replication; Resistance; SARS-CoV-2.

MeSH terms

Animals
Antiviral Agents* / pharmacology
COVID-19 / virology
COVID-19 Drug Treatment
Chlorocebus aethiops
Drug Resistance, Viral* / genetics
Humans
Indazoles / pharmacology
Lactams
Leucine
Mutation*
Nitriles
Proline
RNA, Viral / genetics
Replicon* / drug effects
Replicon* / genetics
SARS-CoV-2* / drug effects
SARS-CoV-2* / genetics
Vero Cells
Virus Replication* / drug effects

Substances

Antiviral Agents
nirmatrelvir
Indazoles
RNA, Viral
Lactams
Leucine
Nitriles
Proline

Supplementary concepts

SARS-CoV-2 variants