Publication: Human monoclonal ScFv that bind to different functional domains of M2 and inhibit H5N1 influenza virus replication
Issued Date
2013
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eng
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Mahidol University
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BioMed Central
Bibliographic Citation
Virology Journal. Vol.10, (2013), 148
Suggested Citation
Tippawan Pissawong, Santi Maneewatch, Kanyarat Thueng-in, Potjanee Srimanote, Fonthip Dong-din-on, Jeeraphong Thanongsaksrikul, Thaweesak Songserm, Pongsri Tongtawe, Kunan Bangphoomi, Wanpen Chaicumpa Human monoclonal ScFv that bind to different functional domains of M2 and inhibit H5N1 influenza virus replication. Virology Journal. Vol.10, (2013), 148. Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/2801
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Title
Human monoclonal ScFv that bind to different functional domains of M2 and inhibit H5N1 influenza virus replication
Abstract
Background: Novel effective anti-influenza agent that tolerates influenza virus antigenic variation is needed. Highly
conserved influenza virus M2 protein has multiple pivotal functions including ion channel activity for vRNP
uncoating, anti-autophagy and virus assembly, morphogenesis and release. Thus, M2 is an attractive target of antiinfluenza
agents including small molecular drugs and specific antibodies.
Methods: Fully human monoclonal single chain antibodies (HuScFv) specific to recombinant and native M2
proteins of A/H5N1 virus were produced from huscfv-phagemid transformed E. coli clones selected from a HuScFv
phage display library using recombinant M2 of clade 1 A/H5N1 as panning antigen. The HuScFv were tested
for their ability to inhibit replication of A/H5N1 of both homologous and heterologous clades. M2 domains
bound by HuScFv of individual E. coli clones were identified by phage mimotope searching and computerized
molecular docking.
Results: HuScFv derived from four huscfv-phagemid transformed E. coli clones (no. 2, 19, 23 and 27) showed
different amino acid sequences particularly at the CDRs. Cells infected with A/H5N1 influenza viruses (both
adamantane sensitive and resistant) that had been exposed to the HuScFv had reduced virus release and
intracellular virus. Phage peptide mimotope search and multiple alignments revealed that conformational epitopes
of HuScFv2 located at the residues important for ion channel activity, anti-autophagy and M1 binding; epitopic
residues of HuScFv19 located at the M2 amphipathic helix and cytoplasmic tail important for anti-autophagy, virus
assembly, morphogenesis and release; epitope of HuScFv23 involved residues important for the M2 activities similar
to HuScFv2 and also amphipathic helix residues for viral budding and release while HuScFv27 epitope spanned
ectodomain, ion channel and anti-autophagy residues. Results of computerized homology modelling and molecular
docking conformed to the epitope identification by phages.
Conclusions: HuScFv that bound to highly conserved epitopes across influenza A subtypes and human pathogenic
H5N1clades located on different functional domains of M2 were produced. The HuScFv reduced viral release and
intracellular virus of infected cells. While the molecular mechanisms of the HuScFv await experimental validation,
the small human antibody fragments have high potential for developing further as a safe, novel and mutation
tolerable anti-influenza agent especially against drug resistant variants.