Abstract
Highly pathogenic avian influenza (HPAI) H5N1 virus infection is still a potential threat to public health worldwide. While vaccines and antiviral drugs are currently under development, neutralizing antibodies could offer an alternative strategy to prevent and treat H5N1 virus infection. In the present study, we had developed a humanized antibody against H5N1 viruses from mouse-derived hybridoma in order to minimize its immunogenicity for potential clinical application. The humanized antibody hH5M9 was generated by transferring the mouse complementarity determining region (CDR) residues together with four key framework region (FR) residues onto the FR of the human antibody. This humanized antibody exhibited high affinity and specificity comparable to the parental mouse or chimeric counterpart with broad and strong neutralization activity against all H5N1 clades and subclades except for Egypt clades investigated. Furthermore, through epitope mapping we identified a linear epitope on the top region of hemagglutinin (HA) that was H5N1 specific and conserved. Our results for the first time reported a humanized antibody against H5N1 viruses by CDR grafting method. With the expected lower immunogenicity, this humanized antibody was expected to be more efficacious than murine or human-mouse chimeric antibodies for future application in humans.
Highlights
H5N1 virus, one of highly pathogenic avian influenza (HPAI) strains, has caused numerous outbreaks in poultry in Southeast Asia since 1997 [1,2,3], and more recently continues to spread globally
Genes from three complementarity determining region (CDR) of monoclonal antibody H5M9 (mH5M9) VH/variable domain (VL) regions were directly grafted onto human antibody FabOX108 VH/VL frameworks to generate CDR-grafted antibody gene
Four framework region (FR) residues of mH5M9 VH region (Met37, Arg66, Val71, Val109) were retained, which were deduced to influence the conformations of CDRs according to the simulation of three dimensional structure of VH and VL regions (Figure S2)
Summary
H5N1 virus, one of highly pathogenic avian influenza (HPAI) strains, has caused numerous outbreaks in poultry in Southeast Asia since 1997 [1,2,3], and more recently continues to spread globally. These outbreaks are accompanied by the occasional transmission of HPAI H5N1 virus to humans, resulting in a total of 628 cases with 374 deaths in 15 countries since 2003 [4]. It is feasible to induce humoral immunity in humans through preventive vaccination and neutralizing antibody generation to protect against H5N1 virus infection. Passive immune therapies have been further highlighted by transfusion of human convalescent sera leading to a 50% reduction in influenza mortality during the 1918 Spanish influenza pandemic, and more recently by anecdotal reports of treating H5N1 human infection with convalescent sera in China [8,9]
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