{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,30]],"date-time":"2026-09-30T17:35:35Z","timestamp":1790789735987,"version":"4.1.0"},"reference-count":41,"publisher":"American Chemical Society (ACS)","issue":"10","license":[{"start":{"date-parts":[[2015,8,6]],"date-time":"2015-08-06T00:00:00Z","timestamp":1438819200000},"content-version":"vor","delay-in-days":365,"URL":"https:\/\/pubs.acs.org\/page\/policy\/authorchoice_termsofuse.html"}],"funder":[{"DOI":"10.13039\/100000051","name":"National Human Genome Research Institute","doi-asserted-by":"publisher","award":["U54 HG005031"],"award-info":[{"award-number":["U54 HG005031"]}],"id":[{"id":"10.13039\/100000051","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01 AI088001"],"award-info":[{"award-number":["R01 AI088001"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/na","name":"UWM Research Foundation, Inc.","doi-asserted-by":"publisher","award":["Research Growth Initiative Award"],"award-info":[{"award-number":["Research Growth Initiative Award"]}],"id":[{"id":"10.13039\/na","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2014,10,17]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>The hepatitis C virus (HCV) nonstructural protein 3 (NS3) is both a protease, which cleaves viral and host proteins, and a helicase that separates nucleic acid strands, using ATP hydrolysis to fuel the reaction. Many antiviral drugs, and compounds in clinical trials, target the NS3 protease, but few helicase inhibitors that function as antivirals have been reported. This study focuses on the analysis of the mechanism by which ebselen (2-phenyl-1,2-benzisoselenazol-3-one), a compound previously shown to be a HCV antiviral agent, inhibits the NS3 helicase. Ebselen inhibited the abilities of NS3 to unwind nucleic acids, to bind nucleic acids, and to hydrolyze ATP, and about 1 \u03bcM ebselen was sufficient to inhibit each of these activities by 50%. However, ebselen had no effect on the activity of the NS3 protease, even at 100 times higher ebselen concentrations. At concentrations below 10 \u03bcM, the ability of ebselen to inhibit HCV helicase was reversible, but prolonged incubation of HCV helicase with higher ebselen concentrations led to irreversible inhibition and the formation of covalent adducts between ebselen and all 14 cysteines present in HCV helicase. Ebselen analogues with sulfur replacing the selenium were just as potent HCV helicase inhibitors as ebselen, but the length of the linker between the phenyl and benzisoselenazol rings was critical. Modifications of the phenyl ring also affected compound potency over 30-fold, and ebselen was a far more potent helicase inhibitor than other, structurally unrelated, thiol-modifying agents. Ebselen analogues were also more effective antiviral agents, and they were less toxic to hepatocytes than ebselen. Although the above structure\u2013activity relationship studies suggest that ebselen targets a specific site on NS3, we were unable to confirm binding to either the NS3 ATP binding site or nucleic acid binding cleft by examining the effects of ebselen on NS3 proteins lacking key cysteines.<\/jats:p>","DOI":"10.1021\/cb500512z","type":"journal-article","created":{"date-parts":[[2014,8,6]],"date-time":"2014-08-06T16:41:19Z","timestamp":1407343279000},"page":"2393-2403","source":"Crossref","is-referenced-by-count":90,"title":["Ebselen Inhibits Hepatitis C Virus NS3 Helicase Binding\nto Nucleic Acid and Prevents Viral Replication"],"prefix":"10.1021","volume":"9","author":[{"given":"Sourav","family":"Mukherjee","sequence":"first","affiliation":[{"name":"University of Wisconsin\u2212Milwaukee , , , ,","place":["Milwaukee, Wisconsin, United States, 53211"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Warren S.","family":"Weiner","sequence":"additional","affiliation":[{"name":"University of Kansas , , 2034 Becker Drive , , ,","place":["Lawrence, Kansas, United States, 66047"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chad E.","family":"Schroeder","sequence":"additional","affiliation":[{"name":"University of Kansas , , 2034 Becker Drive , , ,","place":["Lawrence, Kansas, United States, 66047"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Denise S.","family":"Simpson","sequence":"additional","affiliation":[{"name":"University of Kansas , , 2034 Becker Drive , , ,","place":["Lawrence, Kansas, United States, 66047"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alicia M.","family":"Hanson","sequence":"additional","affiliation":[{"name":"University of Wisconsin\u2212Milwaukee , , , ,","place":["Milwaukee, Wisconsin, United States, 53211"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Noreena L.","family":"Sweeney","sequence":"additional","affiliation":[{"name":"University of Wisconsin\u2212Milwaukee , , , ,","place":["Milwaukee, Wisconsin, United States, 53211"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rachel K.","family":"Marvin","sequence":"additional","affiliation":[{"name":"University of Toledo , , , ,","place":["Toledo, Ohio, United States, 43606"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jean","family":"Ndjomou","sequence":"additional","affiliation":[{"name":"University of Wisconsin\u2212Milwaukee , , , ,","place":["Milwaukee, Wisconsin, United States, 53211"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rajesh","family":"Kolli","sequence":"additional","affiliation":[{"name":"University of Wisconsin\u2212Milwaukee , , , ,","place":["Milwaukee, Wisconsin, United States, 53211"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dragan","family":"Isailovic","sequence":"additional","affiliation":[{"name":"University of Toledo , , , ,","place":["Toledo, Ohio, United States, 43606"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Frank J.","family":"Schoenen","sequence":"additional","affiliation":[{"name":"University of Kansas , , 2034 Becker Drive , , ,","place":["Lawrence, Kansas, United States, 66047"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David N.","family":"Frick","sequence":"additional","affiliation":[{"name":"University of Wisconsin\u2212Milwaukee , , , ,","place":["Milwaukee, Wisconsin, United States, 53211"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2014,8,6]]},"reference":[{"key":"2026090206115448400_cit1","doi-asserted-by":"crossref","first-page":"1001","DOI":"10.1038\/nrd2424","article-title":"The design of drugs for HIV and HCV","volume":"6","author":"De Clercq","year":"2007","journal-title":"Nat. Rev.\nDrug Discovery"},{"key":"2026090206115448400_cit2","first-page":"1","article-title":"The hepatitis\nC virus NS3 protein: a model RNA helicase and potential drug target","volume":"9","author":"Frick","year":"2007","journal-title":"Curr. Issues Mol. Biol."},{"key":"2026090206115448400_cit3","first-page":"237","article-title":"Hepatitis C virus NS3 helicase inhibitors","volume-title":"Hepatitis C: Antiviral Drug Discovery and Development","author":"Belon","year":"2011"},{"key":"2026090206115448400_cit4","doi-asserted-by":"crossref","first-page":"6072","DOI":"10.1128\/JVI.02495-05","article-title":"Dissociation of a MAVS\/IPS-1\/VISA\/Cardif-IKKepsilon\nmolecular complex from the mitochondrial outer membrane by hepatitis\nC virus NS3-4A proteolytic cleavage","volume":"80","author":"Lin","year":"2006","journal-title":"J. Virol."},{"key":"2026090206115448400_cit5","doi-asserted-by":"crossref","first-page":"2992","DOI":"10.1073\/pnas.0408824102","article-title":"Immune evasion by\nhepatitis C virus NS3\/4A protease-mediated cleavage of the Toll-like\nreceptor 3 adaptor protein TRIF","volume":"102","author":"Li","year":"2005","journal-title":"Proc. Natl.\nAcad. Sci. U.S.A."},{"key":"2026090206115448400_cit6","doi-asserted-by":"crossref","first-page":"521","DOI":"10.1073\/pnas.0913380107","article-title":"Three conformational\nsnapshots of the hepatitis C virus\nNS3 helicase reveal a ratchet translocation mechanism","volume":"107","author":"Gu","year":"2010","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"2026090206115448400_cit7","doi-asserted-by":"crossref","first-page":"89","DOI":"10.1016\/S0969-2126(98)00010-0","article-title":"Hepatitis C virus\nNS3 RNA helicase domain with a bound oligonucleotide: the crystal\nstructure provides insights into the mode of unwinding","volume":"6","author":"Kim","year":"1998","journal-title":"Structure"},{"key":"2026090206115448400_cit8","doi-asserted-by":"crossref","first-page":"8607","DOI":"10.1093\/nar\/gks623","article-title":"Identification\nand analysis of hepatitis C virus NS3 helicase inhibitors using nucleic\nacid binding assays","volume":"40","author":"Mukherjee","year":"2012","journal-title":"Nucleic Acids Res."},{"key":"2026090206115448400_cit9","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1073\/pnas.0912966107","article-title":"Unbiased probing of the entire hepatitis\nC virus life cycle identifies clinical compounds that target multiple\naspects of the infection","volume":"107","author":"Gastaminza","year":"2010","journal-title":"Proc. Natl. Acad.\nSci. U.S.A."},{"key":"2026090206115448400_cit10","doi-asserted-by":"crossref","first-page":"9294","DOI":"10.1073\/pnas.0503596102","article-title":"Robust\nhepatitis C virus infection in vitro","volume":"102","author":"Zhong","year":"2005","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"2026090206115448400_cit11","doi-asserted-by":"crossref","first-page":"2748","DOI":"10.1038\/ncomms3748","article-title":"Mechanism of inhibition\nof Mycobacterium tuberculosis antigen 85 by ebselen","volume":"4","author":"Favrot","year":"2013","journal-title":"Nat. Commun."},{"key":"2026090206115448400_cit12","doi-asserted-by":"crossref","first-page":"183","DOI":"10.1021\/cb400485k","article-title":"High-throughput\nscreening using the differential radial capillary action of ligand\nassay identifies ebselen as an inhibitor of diguanylate cyclases","volume":"9","author":"Lieberman","year":"2014","journal-title":"ACS Chem. Biol."},{"key":"2026090206115448400_cit13","doi-asserted-by":"crossref","first-page":"245","DOI":"10.1016\/j.antiviral.2012.08.006","article-title":"Fluorescent primuline derivatives\ninhibit hepatitis C virus NS3-catalyzed RNA unwinding, peptide hydrolysis\nand viral replicase formation","volume":"96","author":"Ndjomou","year":"2012","journal-title":"Antiviral Res."},{"key":"2026090206115448400_cit14","doi-asserted-by":"crossref","first-page":"67","DOI":"10.1177\/108705719900400206","article-title":"A simple\nstatistical parameter for\nuse in evaluation and validation of high throughput screening assays","volume":"4","author":"Zhang","year":"1999","journal-title":"J. Biomol. Screen."},{"key":"2026090206115448400_cit15","doi-asserted-by":"crossref","first-page":"1153","DOI":"10.1016\/0306-3623(95)00003-J","article-title":"Molecular\nactions of ebselen\u2500an antiinflammatory antioxidant","volume":"26","author":"Schewe","year":"1995","journal-title":"Gen. Pharmacol."},{"key":"2026090206115448400_cit16","doi-asserted-by":"crossref","first-page":"27456","DOI":"10.1074\/jbc.M113.495101","article-title":"Ebsulfur is a benzisothiazolone\ncytocidal inhibitor targeting the trypanothione reductase of Trypanosoma brucei","volume":"288","author":"Lu","year":"2013","journal-title":"J. Biol. Chem."},{"key":"2026090206115448400_cit17","doi-asserted-by":"crossref","first-page":"3319","DOI":"10.1021\/jm300021v","article-title":"Optimization of\npotent hepatitis C virus NS3 helicase inhibitors isolated from the\nyellow dyes thioflavine S and primuline","volume":"55","author":"Li","year":"2012","journal-title":"J.\nMed. Chem."},{"key":"2026090206115448400_cit18","doi-asserted-by":"crossref","first-page":"23311","DOI":"10.1074\/jbc.M301283200","article-title":"ATP binding\nmodulates the nucleic\nacid affinity of hepatitis C virus helicase","volume":"278","author":"Levin","year":"2003","journal-title":"J. Biol. Chem."},{"key":"2026090206115448400_cit19","doi-asserted-by":"crossref","first-page":"851","DOI":"10.1016\/j.jmb.2009.03.059","article-title":"Fuel specificity of the hepatitis C virus NS3 helicase","volume":"388","author":"Belon","year":"2009","journal-title":"J. Mol. Biol."},{"key":"2026090206115448400_cit20","doi-asserted-by":"crossref","first-page":"3950","DOI":"10.1128\/JVI.77.7.3950-3961.2003","article-title":"Hepatitis C virus\nNS3 ATPases\/helicases from different genotypes exhibit variations\nin enzymatic properties","volume":"77","author":"Lam","year":"2003","journal-title":"J. Virol."},{"key":"2026090206115448400_cit21","doi-asserted-by":"crossref","first-page":"9400","DOI":"10.1128\/jvi.71.12.9400-9409.1997","article-title":"Mutational analysis\nof the hepatitis C virus RNA helicase","volume":"71","author":"Kim","year":"1997","journal-title":"J. Virol."},{"key":"2026090206115448400_cit22","doi-asserted-by":"crossref","first-page":"96","DOI":"10.1038\/nature08960","article-title":"Chemical\ngenetics strategy identifies an HCV NS5A inhibitor\nwith a potent clinical effect","volume":"465","author":"Gao","year":"2010","journal-title":"Nature"},{"key":"2026090206115448400_cit23","doi-asserted-by":"crossref","first-page":"3764","DOI":"10.1073\/pnas.0915117107","article-title":"A cell protection screen\nreveals potent inhibitors of multiple stages of the hepatitis C virus\nlife cycle","volume":"107","author":"Chockalingam","year":"2010","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"2026090206115448400_cit24","doi-asserted-by":"crossref","first-page":"3731","DOI":"10.1128\/AAC.00662-13","article-title":"Interrogating\na hexokinase-selected small-molecule\nlibrary for inhibitors of Plasmodium falciparum hexokinase","volume":"57","author":"Harris","year":"2013","journal-title":"Antimicrob. Agents Chemother."},{"key":"2026090206115448400_cit25","first-page":"154","article-title":"Exploring the mode of action of ebselen\nin Trypanosoma brucei hexokinase inhibition","volume":"3","author":"Joice","year":"2013","journal-title":"Int. J. Parasitol.: Drugs Drug Resist."},{"key":"2026090206115448400_cit26","doi-asserted-by":"crossref","first-page":"2935","DOI":"10.1016\/j.bmcl.2009.04.074","article-title":"Discovery\nof a novel HCV helicase inhibitor by a de novo drug design approach","volume":"19","author":"Kandil","year":"2009","journal-title":"Bioorg. Med. Chem. Lett."},{"key":"2026090206115448400_cit27","doi-asserted-by":"crossref","first-page":"701","DOI":"10.1099\/0022-1317-80-3-701","article-title":"Characterization\nand mutational analysis of the helicase and NTPase activities of hepatitis\nC virus full-length NS3 protein","volume":"80","author":"Wardell","year":"1999","journal-title":"J. Gen. Virol."},{"key":"2026090206115448400_cit28","doi-asserted-by":"crossref","first-page":"471","DOI":"10.1038\/nchembio.925","article-title":"Reversible\ntargeting of noncatalytic cysteines with\nchemically tuned electrophiles","volume":"8","author":"Serafimova","year":"2012","journal-title":"Nat. Chem. Biol."},{"key":"2026090206115448400_cit29","doi-asserted-by":"crossref","first-page":"1529","DOI":"10.4155\/fmc.10.237","article-title":"Observations\non screening-based research and some concerning trends in the literature","volume":"2","author":"Baell","year":"2010","journal-title":"Future Med. Chem."},{"key":"2026090206115448400_cit30","doi-asserted-by":"crossref","first-page":"1248","DOI":"10.1016\/j.bcp.2013.08.028","article-title":"The early research and development of ebselen","volume":"86","author":"Parnham","year":"2013","journal-title":"Biochem. Pharmacol."},{"key":"2026090206115448400_cit31","doi-asserted-by":"crossref","first-page":"2417","DOI":"10.1056\/NEJMoa1013086","article-title":"Telaprevir\nfor retreatment of HCV infection","volume":"364","author":"Zeuzem","year":"2011","journal-title":"N. Engl. J.\nMed."},{"key":"2026090206115448400_cit32","doi-asserted-by":"crossref","first-page":"1207","DOI":"10.1056\/NEJMoa1009482","article-title":"Boceprevir for previously\ntreated chronic HCV genotype 1 infection","volume":"364","author":"Bacon","year":"2011","journal-title":"N.\nEngl. J. Med."},{"key":"2026090206115448400_cit33","doi-asserted-by":"crossref","first-page":"1673","DOI":"10.1021\/jm401507s","article-title":"Discovery and development of simeprevir\n(TMC435), a HCV NS3\/4A protease inhibitor","volume":"57","author":"Rosenquist","year":"2014","journal-title":"J.\nMed. Chem."},{"key":"2026090206115448400_cit34","doi-asserted-by":"crossref","first-page":"1878","DOI":"10.1056\/NEJMoa1214853","article-title":"Sofosbuvir\nfor previously untreated chronic hepatitis C infection","volume":"368","author":"Lawitz","year":"2013","journal-title":"N. Engl. J. Med."},{"key":"2026090206115448400_cit35","doi-asserted-by":"crossref","first-page":"463","DOI":"10.1016\/B978-0-12-396546-2.00021-8","article-title":"Identification and\nanalysis of inhibitors targeting the hepatitis C virus NS3 helicase","volume":"511","author":"Hanson","year":"2012","journal-title":"Methods Enzymol."},{"key":"2026090206115448400_cit36","doi-asserted-by":"crossref","first-page":"223","DOI":"10.1007\/978-1-60327-355-8_16","article-title":"A method to simultaneously monitor\nhepatitis C virus NS3 helicase and protease activities","volume":"587","author":"Frick","year":"2010","journal-title":"Methods Mol. Biol."},{"key":"2026090206115448400_cit37","doi-asserted-by":"crossref","first-page":"2964","DOI":"10.1021\/jo00273a035","article-title":"Expedient synthesis of ebselen and related compounds","volume":"54","author":"Engman","year":"1989","journal-title":"J. Org. Chem."},{"key":"2026090206115448400_cit38","doi-asserted-by":"crossref","first-page":"851","DOI":"10.1016\/0968-0896(96)00053-3","article-title":"2,5-Diarylisothiazolone: novel inhibitors of cytokine-induced\ncartilage destruction","volume":"4","author":"Wright","year":"1996","journal-title":"Bioorg. Med. Chem."},{"key":"2026090206115448400_cit39","doi-asserted-by":"crossref","first-page":"3501","DOI":"10.1021\/jo060070+","article-title":"Novel alternative\nfor the N\u2013N bond formation through a PIFA-mediated oxidative\ncyclization and its application to the synthesis of indazol-3-ones","volume":"71","author":"Correa","year":"2006","journal-title":"J. Org. Chem."},{"key":"2026090206115448400_cit40","doi-asserted-by":"crossref","first-page":"6151","DOI":"10.1021\/bi4006495","article-title":"Aurintricarboxylic\nacid modulates the affinity of hepatitis C virus NS3 helicase for\nboth nucleic acid and ATP","volume":"52","author":"Shadrick","year":"2013","journal-title":"Biochemistry"},{"key":"2026090206115448400_cit41","doi-asserted-by":"crossref","first-page":"433","DOI":"10.2144\/000112834","article-title":"Monitoring helicase\nactivity with molecular beacons","volume":"45","author":"Belon","year":"2008","journal-title":"BioTechniques"}],"container-title":["ACS Chemical Biology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pubs.acs.org\/doi\/pdf\/10.1021\/cb500512z","content-type":"application\/pdf","content-version":"vor","intended-application":"unspecified"},{"URL":"https:\/\/pubs.acs.org\/acbcct\/article-pdf\/9\/10\/2393\/15726033\/cb500512z.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/pubs.acs.org\/acbcct\/article-pdf\/9\/10\/2393\/15726033\/cb500512z.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,2]],"date-time":"2026-09-02T10:12:08Z","timestamp":1788343928000},"score":1,"resource":{"primary":{"URL":"https:\/\/pubs.acs.org\/acbcct\/article\/9\/10\/2393\/1479306\/Ebselen-Inhibits-Hepatitis-C-Virus-NS3-Helicase"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2014,8,6]]},"references-count":41,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2014,8,6]]},"published-print":{"date-parts":[[2014,10,17]]}},"URL":"https:\/\/doi.org\/10.1021\/cb500512z","relation":{},"ISSN":["1554-8929","1554-8937"],"issn-type":[{"value":"1554-8929","type":"print"},{"value":"1554-8937","type":"electronic"}],"subject":[],"published":{"date-parts":[[2014,8,6]]}}}