A luminescent attenuated SARS-CoV-2 for the identification and validation of drug-resistant mutants DOI

Yao Ma,

Chengjin Ye, Ahmed Magdy Khalil

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: May 13, 2025

ABSTRACT The emergence of SARS-CoV-2 variants has necessitated continuous updating vaccines. In contrast, antivirals remained effective as they target conserved viral proteins that are essential for the life cycle. However, several mutations in may affect efficacy United States (US) Food and Drug Administration (FDA)-approved have been recently identified. Detecting drug-resistant mutants investigating their escape mechanism(s) critical to guide selection antiviral therapies. this study, we constructed an attenuated recombinant (r)SARS-CoV-2 lacking open reading frame (ORF) 3a 7b but expressing nanoluciferase (Nluc), rSARS-CoV-2 Δ3a7b-Nluc, facilitate tracking infection. Using virus, selected main protease (Mpro) inhibitor nirmatrelvir. After passaging Δ3a7b-Nluc 10 times presence increasing concentrations nirmatrelvir, a virus population with enhanced resistance was selected. We identified two non-synonymous (L50F R188G) Mpro, encoded by non-structural protein 5 (NSP5) gene. reverse genetics, generated containing L50F R188G mutations, individually or combination, assessed contribution nirmatrelvir resistance. Our results indicate both involved escaping from Altogether, our demonstrate feasibility using variant identify validate confer FDA-approved drugs without concern conducting gain function (GoF) experiments wild-type (WT) forms SARS-CoV-2. IMPORTANCE Small-molecule used treatment infections. currently US Mpro targeting Information on affecting activity licensed remain limited. developed (Nluc)-expressing ORF (Δ3a7b-Nluc) resistant biosafety concerns associated gain-of-function research reduced sensitivity were validated generation candidate Mpro. These safely overcoming originating adaptation studies WT

Language: Английский

A luminescent attenuated SARS-CoV-2 for the identification and validation of drug-resistant mutants DOI

Yao Ma,

Chengjin Ye, Ahmed Magdy Khalil

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: May 13, 2025

ABSTRACT The emergence of SARS-CoV-2 variants has necessitated continuous updating vaccines. In contrast, antivirals remained effective as they target conserved viral proteins that are essential for the life cycle. However, several mutations in may affect efficacy United States (US) Food and Drug Administration (FDA)-approved have been recently identified. Detecting drug-resistant mutants investigating their escape mechanism(s) critical to guide selection antiviral therapies. this study, we constructed an attenuated recombinant (r)SARS-CoV-2 lacking open reading frame (ORF) 3a 7b but expressing nanoluciferase (Nluc), rSARS-CoV-2 Δ3a7b-Nluc, facilitate tracking infection. Using virus, selected main protease (Mpro) inhibitor nirmatrelvir. After passaging Δ3a7b-Nluc 10 times presence increasing concentrations nirmatrelvir, a virus population with enhanced resistance was selected. We identified two non-synonymous (L50F R188G) Mpro, encoded by non-structural protein 5 (NSP5) gene. reverse genetics, generated containing L50F R188G mutations, individually or combination, assessed contribution nirmatrelvir resistance. Our results indicate both involved escaping from Altogether, our demonstrate feasibility using variant identify validate confer FDA-approved drugs without concern conducting gain function (GoF) experiments wild-type (WT) forms SARS-CoV-2. IMPORTANCE Small-molecule used treatment infections. currently US Mpro targeting Information on affecting activity licensed remain limited. developed (Nluc)-expressing ORF (Δ3a7b-Nluc) resistant biosafety concerns associated gain-of-function research reduced sensitivity were validated generation candidate Mpro. These safely overcoming originating adaptation studies WT

Language: Английский

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