Severe acute respiratory syndrome coronavirus replication inhibitor that interferes with the nucleic acid unwinding of the viral helicase.
Identifieur interne : 001C54 ( Main/Exploration ); précédent : 001C53; suivant : 001C55Severe acute respiratory syndrome coronavirus replication inhibitor that interferes with the nucleic acid unwinding of the viral helicase.
Auteurs : Adeyemi O. Adedeji [États-Unis] ; Kamalendra Singh ; Nicholas E. Calcaterra ; Marta L. Dediego ; Luis Enjuanes ; Susan Weiss ; Stefan G. SarafianosSource :
- Antimicrobial agents and chemotherapy [ 1098-6596 ] ; 2012.
Descripteurs français
- KwdFr :
- ARN double brin (antagonistes et inhibiteurs), ARN double brin (génétique), ARN viral (antagonistes et inhibiteurs), ARN viral (génétique), Antiviraux (pharmacologie), Bibliothèques de petites molécules (pharmacologie), Cellules HEK293, Cinétique, Concentration inhibitrice 50, Conformation d'acide nucléique (), Escherichia coli (génétique), Helicase (antagonistes et inhibiteurs), Helicase (métabolisme), Humains, Protéines recombinantes (métabolisme), Protéines virales (antagonistes et inhibiteurs), Protéines virales (métabolisme), Réplication virale (), Survie cellulaire (), Syndrome respiratoire aigu sévère (traitement médicamenteux), Syndrome respiratoire aigu sévère (virologie), Transfert d'énergie par résonance de fluorescence, Triazoles (pharmacologie), Virus du SRAS (), Virus du SRAS (enzymologie), Virus du SRAS (génétique).
- MESH :
- antagonistes et inhibiteurs : ARN double brin, ARN viral, Helicase, Protéines virales.
- enzymologie : Virus du SRAS.
- génétique : ARN double brin, ARN viral, Escherichia coli, Virus du SRAS.
- métabolisme : Helicase, Protéines recombinantes, Protéines virales.
- pharmacologie : Antiviraux, Bibliothèques de petites molécules, Triazoles.
- traitement médicamenteux : Syndrome respiratoire aigu sévère.
- virologie : Syndrome respiratoire aigu sévère.
- Cellules HEK293, Cinétique, Concentration inhibitrice 50, Conformation d'acide nucléique, Humains, Réplication virale, Survie cellulaire, Transfert d'énergie par résonance de fluorescence, Virus du SRAS.
English descriptors
- KwdEn :
- Antiviral Agents (pharmacology), Cell Survival (drug effects), DNA Helicases (antagonists & inhibitors), DNA Helicases (metabolism), Escherichia coli (genetics), Fluorescence Resonance Energy Transfer, HEK293 Cells, Humans, Inhibitory Concentration 50, Kinetics, Nucleic Acid Conformation (drug effects), RNA, Double-Stranded (antagonists & inhibitors), RNA, Double-Stranded (genetics), RNA, Viral (antagonists & inhibitors), RNA, Viral (genetics), Recombinant Proteins (metabolism), SARS Virus (drug effects), SARS Virus (enzymology), SARS Virus (genetics), Severe Acute Respiratory Syndrome (drug therapy), Severe Acute Respiratory Syndrome (virology), Small Molecule Libraries (pharmacology), Triazoles (pharmacology), Viral Proteins (antagonists & inhibitors), Viral Proteins (metabolism), Virus Replication (drug effects).
- MESH :
- chemical , antagonists & inhibitors : DNA Helicases, RNA, Double-Stranded, RNA, Viral, Viral Proteins.
- chemical , genetics : RNA, Double-Stranded, RNA, Viral.
- chemical , metabolism : DNA Helicases, Recombinant Proteins, Viral Proteins.
- chemical , pharmacology : Antiviral Agents, Small Molecule Libraries, Triazoles.
- drug effects : Cell Survival, Nucleic Acid Conformation, SARS Virus, Virus Replication.
- drug therapy : Severe Acute Respiratory Syndrome.
- enzymology : SARS Virus.
- genetics : Escherichia coli, SARS Virus.
- virology : Severe Acute Respiratory Syndrome.
- Fluorescence Resonance Energy Transfer, HEK293 Cells, Humans, Inhibitory Concentration 50, Kinetics.
Abstract
Severe acute respiratory syndrome (SARS) is a highly contagious disease, caused by SARS coronavirus (SARS-CoV), for which there are no approved treatments. We report the discovery of a potent inhibitor of SARS-CoV that blocks replication by inhibiting the unwinding activity of the SARS-CoV helicase (nsp13). We used a Förster resonance energy transfer (FRET)-based helicase assay to screen the Maybridge Hitfinder chemical library. We identified and validated a compound (SSYA10-001) that specifically blocks the double-stranded RNA (dsRNA) and dsDNA unwinding activities of nsp13, with 50% inhibitory concentrations (IC(50)s) of 5.70 and 5.30 μM, respectively. This compound also has inhibitory activity (50% effective concentration [EC(50)] = 8.95 μM) in a SARS-CoV replicon assay, with low cytotoxicity (50% cytotoxic concentration [CC(50)] = >250 μM), suggesting that the helicase plays a still unidentified critical role in the SARS-CoV life cycle. Enzyme kinetic studies on the mechanism of nsp13 inhibition revealed that SSYA10-001 acts as a noncompetitive inhibitor of nsp13 with respect to nucleic acid and ATP substrates. Moreover, SSYA10-001 does not affect ATP hydrolysis or nsp13 binding to the nucleic acid substrate. SSYA10-001 did not inhibit hepatitis C virus (HCV) helicase, other bacterial and viral RNA-dependent RNA polymerases, or reverse transcriptase. These results suggest that SSYA10-001 specifically blocks nsp13 through a novel mechanism and is less likely to interfere with the functions of cellular enzymes that process nucleic acids or ATP. Hence, it is possible that SSYA10-001 inhibits unwinding by nsp13 by affecting conformational changes during the course of the reaction or translocation on the nucleic acid. SSYA10-001 will be a valuable tool for studying the specific role of nsp13 in the SARS-CoV life cycle, which could be a model for other nidoviruses and also a candidate for further development as a SARS antiviral target.
DOI: 10.1128/AAC.00957-12
PubMed: 22733076
Affiliations:
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Dediego</name></author><author><name sortKey="Enjuanes, Luis" sort="Enjuanes, Luis" uniqKey="Enjuanes L" first="Luis" last="Enjuanes">Luis Enjuanes</name></author><author><name sortKey="Weiss, Susan" sort="Weiss, Susan" uniqKey="Weiss S" first="Susan" last="Weiss">Susan Weiss</name></author><author><name sortKey="Sarafianos, Stefan G" sort="Sarafianos, Stefan G" uniqKey="Sarafianos S" first="Stefan G" last="Sarafianos">Stefan G. 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Adedeji</name><affiliation wicri:level="2"><nlm:affiliation>Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, University of Missouri School of Medicine, Columbia, Missouri, USA.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Christopher S. Bond Life Sciences Center, Department of Molecular Microbiology and Immunology, University of Missouri School of Medicine, Columbia, Missouri</wicri:regionArea><placeName><region type="state">Missouri (État)</region></placeName></affiliation></author><author><name sortKey="Singh, Kamalendra" sort="Singh, Kamalendra" uniqKey="Singh K" first="Kamalendra" last="Singh">Kamalendra Singh</name></author><author><name sortKey="Calcaterra, Nicholas E" sort="Calcaterra, Nicholas E" uniqKey="Calcaterra N" first="Nicholas E" last="Calcaterra">Nicholas E. Calcaterra</name></author><author><name sortKey="Dediego, Marta L" sort="Dediego, Marta L" uniqKey="Dediego M" first="Marta L" last="Dediego">Marta L. Dediego</name></author><author><name sortKey="Enjuanes, Luis" sort="Enjuanes, Luis" uniqKey="Enjuanes L" first="Luis" last="Enjuanes">Luis Enjuanes</name></author><author><name sortKey="Weiss, Susan" sort="Weiss, Susan" uniqKey="Weiss S" first="Susan" last="Weiss">Susan Weiss</name></author><author><name sortKey="Sarafianos, Stefan G" sort="Sarafianos, Stefan G" uniqKey="Sarafianos S" first="Stefan G" last="Sarafianos">Stefan G. Sarafianos</name></author></analytic><series><title level="j">Antimicrobial agents and chemotherapy</title><idno type="eISSN">1098-6596</idno><imprint><date when="2012" type="published">2012</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Antiviral Agents (pharmacology)</term><term>Cell Survival (drug effects)</term><term>DNA Helicases (antagonists & inhibitors)</term><term>DNA Helicases (metabolism)</term><term>Escherichia coli (genetics)</term><term>Fluorescence Resonance Energy Transfer</term><term>HEK293 Cells</term><term>Humans</term><term>Inhibitory Concentration 50</term><term>Kinetics</term><term>Nucleic Acid Conformation (drug effects)</term><term>RNA, Double-Stranded (antagonists & inhibitors)</term><term>RNA, Double-Stranded (genetics)</term><term>RNA, Viral (antagonists & inhibitors)</term><term>RNA, Viral (genetics)</term><term>Recombinant Proteins (metabolism)</term><term>SARS Virus (drug effects)</term><term>SARS Virus (enzymology)</term><term>SARS Virus (genetics)</term><term>Severe Acute Respiratory Syndrome (drug therapy)</term><term>Severe Acute Respiratory Syndrome (virology)</term><term>Small Molecule Libraries (pharmacology)</term><term>Triazoles (pharmacology)</term><term>Viral Proteins (antagonists & inhibitors)</term><term>Viral Proteins (metabolism)</term><term>Virus Replication (drug effects)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ARN double brin (antagonistes et inhibiteurs)</term><term>ARN double brin (génétique)</term><term>ARN viral (antagonistes et inhibiteurs)</term><term>ARN viral (génétique)</term><term>Antiviraux (pharmacologie)</term><term>Bibliothèques de petites molécules (pharmacologie)</term><term>Cellules HEK293</term><term>Cinétique</term><term>Concentration inhibitrice 50</term><term>Conformation d'acide nucléique ()</term><term>Escherichia coli (génétique)</term><term>Helicase (antagonistes et inhibiteurs)</term><term>Helicase (métabolisme)</term><term>Humains</term><term>Protéines recombinantes (métabolisme)</term><term>Protéines virales (antagonistes et inhibiteurs)</term><term>Protéines virales (métabolisme)</term><term>Réplication virale ()</term><term>Survie cellulaire ()</term><term>Syndrome respiratoire aigu sévère (traitement médicamenteux)</term><term>Syndrome respiratoire aigu sévère (virologie)</term><term>Transfert d'énergie par résonance de fluorescence</term><term>Triazoles (pharmacologie)</term><term>Virus du SRAS ()</term><term>Virus du SRAS (enzymologie)</term><term>Virus du SRAS (génétique)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="antagonists & inhibitors" xml:lang="en"><term>DNA Helicases</term><term>RNA, Double-Stranded</term><term>RNA, Viral</term><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>RNA, Double-Stranded</term><term>RNA, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>DNA Helicases</term><term>Recombinant Proteins</term><term>Viral Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="pharmacology" xml:lang="en"><term>Antiviral Agents</term><term>Small Molecule Libraries</term><term>Triazoles</term></keywords><keywords scheme="MESH" qualifier="antagonistes et inhibiteurs" xml:lang="fr"><term>ARN double brin</term><term>ARN viral</term><term>Helicase</term><term>Protéines virales</term></keywords><keywords scheme="MESH" qualifier="drug effects" xml:lang="en"><term>Cell Survival</term><term>Nucleic Acid Conformation</term><term>SARS Virus</term><term>Virus Replication</term></keywords><keywords scheme="MESH" qualifier="drug therapy" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" qualifier="enzymologie" xml:lang="fr"><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="enzymology" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Escherichia coli</term><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ARN double brin</term><term>ARN viral</term><term>Escherichia coli</term><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Helicase</term><term>Protéines recombinantes</term><term>Protéines virales</term></keywords><keywords scheme="MESH" qualifier="pharmacologie" xml:lang="fr"><term>Antiviraux</term><term>Bibliothèques de petites molécules</term><term>Triazoles</term></keywords><keywords scheme="MESH" qualifier="traitement médicamenteux" xml:lang="fr"><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="virologie" xml:lang="fr"><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="virology" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Fluorescence Resonance Energy Transfer</term><term>HEK293 Cells</term><term>Humans</term><term>Inhibitory Concentration 50</term><term>Kinetics</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Cellules HEK293</term><term>Cinétique</term><term>Concentration inhibitrice 50</term><term>Conformation d'acide nucléique</term><term>Humains</term><term>Réplication virale</term><term>Survie cellulaire</term><term>Transfert d'énergie par résonance de fluorescence</term><term>Virus du SRAS</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Severe acute respiratory syndrome (SARS) is a highly contagious disease, caused by SARS coronavirus (SARS-CoV), for which there are no approved treatments. We report the discovery of a potent inhibitor of SARS-CoV that blocks replication by inhibiting the unwinding activity of the SARS-CoV helicase (nsp13). We used a Förster resonance energy transfer (FRET)-based helicase assay to screen the Maybridge Hitfinder chemical library. We identified and validated a compound (SSYA10-001) that specifically blocks the double-stranded RNA (dsRNA) and dsDNA unwinding activities of nsp13, with 50% inhibitory concentrations (IC(50)s) of 5.70 and 5.30 μM, respectively. This compound also has inhibitory activity (50% effective concentration [EC(50)] = 8.95 μM) in a SARS-CoV replicon assay, with low cytotoxicity (50% cytotoxic concentration [CC(50)] = >250 μM), suggesting that the helicase plays a still unidentified critical role in the SARS-CoV life cycle. Enzyme kinetic studies on the mechanism of nsp13 inhibition revealed that SSYA10-001 acts as a noncompetitive inhibitor of nsp13 with respect to nucleic acid and ATP substrates. Moreover, SSYA10-001 does not affect ATP hydrolysis or nsp13 binding to the nucleic acid substrate. SSYA10-001 did not inhibit hepatitis C virus (HCV) helicase, other bacterial and viral RNA-dependent RNA polymerases, or reverse transcriptase. These results suggest that SSYA10-001 specifically blocks nsp13 through a novel mechanism and is less likely to interfere with the functions of cellular enzymes that process nucleic acids or ATP. Hence, it is possible that SSYA10-001 inhibits unwinding by nsp13 by affecting conformational changes during the course of the reaction or translocation on the nucleic acid. SSYA10-001 will be a valuable tool for studying the specific role of nsp13 in the SARS-CoV life cycle, which could be a model for other nidoviruses and also a candidate for further development as a SARS antiviral target.</div></front></TEI><affiliations><list><country><li>États-Unis</li></country><region><li>Missouri (État)</li></region></list><tree><noCountry><name sortKey="Calcaterra, Nicholas E" sort="Calcaterra, Nicholas E" uniqKey="Calcaterra N" first="Nicholas E" last="Calcaterra">Nicholas E. Calcaterra</name><name sortKey="Dediego, Marta L" sort="Dediego, Marta L" uniqKey="Dediego M" first="Marta L" last="Dediego">Marta L. Dediego</name><name sortKey="Enjuanes, Luis" sort="Enjuanes, Luis" uniqKey="Enjuanes L" first="Luis" last="Enjuanes">Luis Enjuanes</name><name sortKey="Sarafianos, Stefan G" sort="Sarafianos, Stefan G" uniqKey="Sarafianos S" first="Stefan G" last="Sarafianos">Stefan G. Sarafianos</name><name sortKey="Singh, Kamalendra" sort="Singh, Kamalendra" uniqKey="Singh K" first="Kamalendra" last="Singh">Kamalendra Singh</name><name sortKey="Weiss, Susan" sort="Weiss, Susan" uniqKey="Weiss S" first="Susan" last="Weiss">Susan Weiss</name></noCountry><country name="États-Unis"><region name="Missouri (État)"><name sortKey="Adedeji, Adeyemi O" sort="Adedeji, Adeyemi O" uniqKey="Adedeji A" first="Adeyemi O" last="Adedeji">Adeyemi O. Adedeji</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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