title: In Vitro Assessment of Combinations of Enterovirus Inhibitors against Enterovirus 71.
abstract: Enterovirus 71 (EV-A71) is a major causative pathogen of hand, foot, and mouth disease (HFMD) epidemics. No antiviral therapies are currently available for treating EV-A71 infections. Here, we selected five reported enterovirus inhibitors (suramin, itraconazole [ITZ], GW5074, rupintrivir, and favipiravir) with different mechanisms of action to test their abilities to inhibit EV-A71 replication alone and in combination. All selected compounds have anti-EV-A71 activities in cell culture. The combination of rupintrivir and ITZ or favipiravir was synergistic, while the combination of rupintrivir and suramin was additive. The combination of suramin and favipiravir exerted a strong synergistic antiviral effect. The observed synergy was not due to cytotoxicity, as there was no significant increase in cytotoxicity when compounds were used in combinations at the tested doses. To investigate the potential inhibitory mechanism of favipiravir against enterovirus, two favipiravir-resistant EV-A71 variants were independently selected, and both of them carried an S121N mutation in the finger subdomain of the 3D polymerase. Reverse engineering of this 3D S121N mutation into an infectious clone of EV-A71 confirmed the resistant phenotype. Moreover, viruses resistant to ITZ or favipiravir remained susceptible to other inhibitors. Most notably, combined with ITZ, rupintrivir prevented the development of ITZ-resistant variants. Taken together, these results provide a rational basis for the design of combination regimens for use in the treatment of EV-A71 infections.
authors.affiliation: Pathogen Diagnostic Center, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Yizhuo
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Pathogen Diagnostic Center, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China School of Life Sciences, Shanghai University, Shanghai, China.
authors.forename: Guiming
authors.initials: G
authors.lastname: Li
authors.affiliation: Pathogen Diagnostic Center, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Shilin
authors.initials: S
authors.lastname: Yuan
authors.affiliation: Pathogen Diagnostic Center, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Qianqian
authors.initials: Q
authors.lastname: Gao
authors.affiliation: Pathogen Diagnostic Center, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Ke
authors.initials: K
authors.lastname: Lan
authors.affiliation: Shandong University-Helmholtz Institute of Biotechnology, Shandong University, Qingdao, China altmeyer@sdu.edu.cn gangzou@ips.ac.cn.
authors.forename: Ralf
authors.initials: R
authors.lastname: Altmeyer
authors.affiliation: Pathogen Diagnostic Center, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China altmeyer@sdu.edu.cn gangzou@ips.ac.cn.
authors.forename: Gang
authors.identifier: 0000-0001-9607-7827
authors.initials: G
authors.lastname: Zou
mesh_terms: D000577:Amides / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001665:Binding Sites; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D004338:Drug Combinations; D024882:Drug Resistance, Viral / Q000235:genetics; D004357:Drug Synergism; D029821:Enterovirus A, Human / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D007555:Isoxazoles / Q000494:pharmacology*; D017964:Itraconazole / Q000494:pharmacology*; D062105:Molecular Docking Simulation; D009154:Mutation; D032446:Myoblasts / Q000187:drug effects / Q000821:virology; D010636:Phenols / Q000494:pharmacology; D010649:Phenylalanine / Q000031:analogs & derivatives; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary; D011719:Pyrazines / Q000494:pharmacology; D011760:Pyrrolidinones / Q000494:pharmacology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013498:Suramin / Q000494:pharmacology*; D014633:Valine / Q000031:analogs & derivatives; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D014779:Virus Replication / Q000187:drug effects
chemical_list: D000577:Amides; D000998:Antiviral Agents; D004338:Drug Combinations; D007211:Indoles; D007555:Isoxazoles; D010636:Phenols; D011719:Pyrazines; D011760:Pyrrolidinones; D017361:Viral Nonstructural Proteins; D017964:Itraconazole; D010649:Phenylalanine; D013498:Suramin; C462182:favipiravir; D014633:Valine; C489251:5-iodo-3-((3,5-dibromo-4-hydroxyphenyl)methylene)-2-indolinone; C118874:rupintrivir