Stalling the Enemy: Targeting Nsp13 for Next-Generation SARS-CoV-2 Antivirals

DOI record: { "DOI": "10.3390/ijms27062587", "ISSN": [ "1422-0067" ], "URL": "http://dx.doi.org/10.3390/ijms27062587", "abstract": "<jats:p>The SARS-CoV-2 public health challenges have highlighted the urgent need for coronavirus-targeting life-saving therapeutics. Given the emergence of drug-resistant strains, the development of antivirals against viral proteins beyond the commonly targeted main protease or RNA-dependent RNA polymerase is critical. The SARS-CoV-2 nonstructural protein 13 (nsp13) is a highly conserved RNA helicase and an essential component of the viral replication–transcription complex (RTC). It unwinds double-stranded RNA to facilitate viral transcription and replication, making it a strong target for drug development. To identify nsp13 inhibitors, we used an ultra-high-throughput nucleic acid unwinding assay to screen a library of FDA-approved drugs and bioactive compounds. We identified forty inhibitors with IC50 values ranging from 1.4 to 10 μM. Ten were further selected for biochemical and biophysical characterization. Four of these are bound to nsp13 without interacting with the nucleic acid substrate and without inhibiting the ATPase activity of nsp13. Hydrogen–deuterium exchange coupled with Mass Spectrometry (HDX-MS) studies show compound binding causes differential exchange in two regions of nsp13. Furthermore, these compounds have antiviral activity against infectious SARS-CoV-2 in multiple cell lines, with cytotoxicity affecting, in some cases, the apparent antiviral effect. Future optimization efforts could help develop therapeutics against SARS-CoV-2 and other potential coronavirus threats.</jats:p>", "alternative-id": [ "ijms27062587" ], "author": [ { "ORCID": "https://orcid.org/0000-0002-4002-4475", "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "authenticated-orcid": false, "family": "Castro", "given": "Jose M.", "sequence": "first" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Slack", "given": "Ryan L.", "sequence": "additional" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Ong", "given": "Yee T.", "sequence": "additional" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Zhang", "given": "Huanchun", "sequence": "additional" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Gifford", "given": "Levi B.", "sequence": "additional" }, { "affiliation": [ { "name": "The Herbert Wertheim Scripps Institute for Biomedical Innovation and Technology, University of Florida Health, Jupiter, FL 33458, USA" } ], "family": "Courouble", "given": "Valentine V.", "sequence": "additional" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Aiken", "given": "Riley M.", "sequence": "additional" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Shankar", "given": "Vishal", "sequence": "additional" }, { "affiliation": [ { "name": "The Herbert Wertheim Scripps Institute for Biomedical Innovation and Technology, University of Florida Health, Jupiter, FL 33458, USA" } ], "family": "O’Leary", "given": "Timothy R.", "sequence": "additional" }, { "affiliation": [ { "name": "The Herbert Wertheim Scripps Institute for Biomedical Innovation and Technology, University of Florida Health, Jupiter, FL 33458, USA" } ], "family": "Griffin", "given": "Patrick R.", "sequence": "additional" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Lan", "given": "Shuiyun", "sequence": "additional" }, { "affiliation": [ { "name": "Department of Pharmacology and Chemical Biology, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Emory Chemical Biology Discovery Center (ECBDC), Emory University, Atlanta, GA 30322, USA" } ], "family": "Du", "given": "Yuhong", "sequence": "additional" }, { "affiliation": [ { "name": "Department of Pharmacology and Chemical Biology, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Emory Chemical Biology Discovery Center (ECBDC), Emory University, Atlanta, GA 30322, USA" } ], "family": "Fu", "given": "Haian", "sequence": "additional" }, { "affiliation": [ { "name": "Center for ViroScience and Cure, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA" }, { "name": "Children’s Healthcare of Atlanta, Atlanta, GA 30329, USA" } ], "family": "Sarafianos", "given": "Stefan G.", "sequence": "additional" } ], "container-title": "International Journal of Molecular Sciences", "container-title-short": "IJMS", "content-domain": { "crossmark-restriction": false, "domain": [] }, "created": { "date-parts": [ [ 2026, 3, 11 ] ], "date-time": "2026-03-11T15:32:09Z", "timestamp": 1773243129000 }, "deposited": { "date-parts": [ [ 2026, 3, 11 ] ], "date-time": 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