title: Identification of a small-molecule inhibitor of influenza virus via disrupting the subunits interaction of the viral polymerase.
abstract: Assembly of the heterotrimeric influenza virus polymerase complex from the individual subunits PB1, PA, and PB2 is a prerequisite for viral replication, in which the interaction between the C terminal of PA (PAC) and the N-terminal of PB1 (PB1N) may be a desired target for antiviral development. In this study, we compared the feasibility of high throughput screening by enzyme-linked immunosorbent assay (ELISA) and fluorescence polarization assay. Among the two, ELISA was demonstrated to own broader dynamic range so that it was used for screening inhibitors that blocked PAC and PB1N interaction. Several binding inhibitors of PAC-PB1N were identified and subsequently tested for the antiviral efficacy. Apparently, 3-(2-chlorophenyl)-6-ethyl-7-methyl[1,2,4]triazolo[4,3-a]pyrimidin-5-ol, designated ANA-1, was found to be a strong inhibitor of viral polymerase activity and act as a potent antiviral agent against the infections of multiple subtypes of influenza A virus, including H1N1, H3N2, H5N1, H7N7, H7N9 and H9N2 subtypes, in cell cultures. Intranasal administration of ANA-1 protected mice from lethal challenge and reduced lung viral loads in H1N1 virus infected BALB/c mice. Docking analyses predicted that ANA-1 bound to an allosteric site of PAC, which might cause conformational changes thereby disrupting the PAC-PB1N interaction. Overall, our study has identified a novel compound with potential to be developed as an anti-influenza drug.
authors.affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: Shuofeng
authors.initials: S
authors.lastname: Yuan
authors.affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: Hin
authors.initials: H
authors.lastname: Chu
authors.affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: Hanjun
authors.initials: H
authors.lastname: Zhao
authors.affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: Ke
authors.initials: K
authors.lastname: Zhang
authors.affiliation: School of Biological Sciences, Faculty of Science, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: Kailash
authors.initials: K
authors.lastname: Singh
authors.affiliation: School of Biological Sciences, Faculty of Science, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: Billy K C
authors.initials: BK
authors.lastname: Chow
authors.affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: Richard Y T
authors.initials: RY
authors.lastname: Kao
authors.affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China. Electronic address: jiezhou@hku.hk.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China. Electronic address: bzheng@hkucc.hku.hk.
authors.forename: Bo-Jian
authors.initials: BJ
authors.lastname: Zheng
mesh_terms: D000495:Allosteric Site / Q000187:drug effects; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D004285:Dogs; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D061985:Madin Darby Canine Kidney Cells; D051379:Mice; D008807:Mice, Inbred BALB C; D062105:Molecular Docking Simulation; D009975:Orthomyxoviridae / Q000145:classification / Q000187:drug effects* / Q000201:enzymology* / Q000502:physiology; D011485:Protein Binding / Q000187:drug effects; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism*; D054852:Small Molecule Libraries / Q000494:pharmacology; D014764:Viral Proteins / Q000378:metabolism*; D014779:Virus Replication / Q000187:drug effects* / Q000235:genetics
chemical_list: D000998:Antiviral Agents; D054852:Small Molecule Libraries; D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase
keywords: Anti-influenza; Cross-protection; Polymerase assembly