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A high-affinity CDR-grafted antibody against influenza A H5N1 viruses recognizes a conserved epitope of H5 hemagglutinin.

Xiong F, Xia L, Wang J, Wu B, Wang D, Yuan L, Cheng Y, Zhu H, Che X, Zhang Q, Zhao G, Wang Y - PLoS ONE (2014)

Bottom Line: While vaccines and antiviral drugs are currently under development, neutralizing antibodies could offer an alternative strategy to prevent and treat H5N1 virus infection.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.

View Article: PubMed Central - PubMed

Affiliation: School of Life Science and Technology, Tongji University, Shanghai, China ; Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center at Shanghai, Shanghai, China.

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.

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Generation of chimeric cH5M9 and CDR-grafted hH5M9 antibodies.(A) Amino acid sequences of the VH regions of mouse antibody mH5M9, human antibody FabOX108 and CDR-grafted antibody hH5M9. Amino acid residues were listed according to the convention of Kabat et al. [31]. Residues shown by underline in frameworks were deemed essential for maintaining the combining sites of mH5M9. Dashes indicated residues that were identical in mH5M9, FabOX108 and hH5M9 whereas gaps denoted amino acid residues missing at those positions. (B) SDS-PAGE analysis of purified cH5M9 and hH5M9 under reducing conditions. mH5M9 was used as the positive control.
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pone-0088777-g001: Generation of chimeric cH5M9 and CDR-grafted hH5M9 antibodies.(A) Amino acid sequences of the VH regions of mouse antibody mH5M9, human antibody FabOX108 and CDR-grafted antibody hH5M9. Amino acid residues were listed according to the convention of Kabat et al. [31]. Residues shown by underline in frameworks were deemed essential for maintaining the combining sites of mH5M9. Dashes indicated residues that were identical in mH5M9, FabOX108 and hH5M9 whereas gaps denoted amino acid residues missing at those positions. (B) SDS-PAGE analysis of purified cH5M9 and hH5M9 under reducing conditions. mH5M9 was used as the positive control.

Mentions: To generate a human-mouse chimeric antibody, the variable regions of both light chain (VL) and heavy chain (VH) from mH5M9 were subcloned into IgG expression cassette vectors IFH and IFL, respectively. IFH-VH and IFL-VL vectors were co-expressed in 293F cells, yielding chimeric H5M9 (cH5M9) (Figure 1B).


A high-affinity CDR-grafted antibody against influenza A H5N1 viruses recognizes a conserved epitope of H5 hemagglutinin.

Xiong F, Xia L, Wang J, Wu B, Wang D, Yuan L, Cheng Y, Zhu H, Che X, Zhang Q, Zhao G, Wang Y - PLoS ONE (2014)

Generation of chimeric cH5M9 and CDR-grafted hH5M9 antibodies.(A) Amino acid sequences of the VH regions of mouse antibody mH5M9, human antibody FabOX108 and CDR-grafted antibody hH5M9. Amino acid residues were listed according to the convention of Kabat et al. [31]. Residues shown by underline in frameworks were deemed essential for maintaining the combining sites of mH5M9. Dashes indicated residues that were identical in mH5M9, FabOX108 and hH5M9 whereas gaps denoted amino acid residues missing at those positions. (B) SDS-PAGE analysis of purified cH5M9 and hH5M9 under reducing conditions. mH5M9 was used as the positive control.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3928294&req=5

pone-0088777-g001: Generation of chimeric cH5M9 and CDR-grafted hH5M9 antibodies.(A) Amino acid sequences of the VH regions of mouse antibody mH5M9, human antibody FabOX108 and CDR-grafted antibody hH5M9. Amino acid residues were listed according to the convention of Kabat et al. [31]. Residues shown by underline in frameworks were deemed essential for maintaining the combining sites of mH5M9. Dashes indicated residues that were identical in mH5M9, FabOX108 and hH5M9 whereas gaps denoted amino acid residues missing at those positions. (B) SDS-PAGE analysis of purified cH5M9 and hH5M9 under reducing conditions. mH5M9 was used as the positive control.
Mentions: To generate a human-mouse chimeric antibody, the variable regions of both light chain (VL) and heavy chain (VH) from mH5M9 were subcloned into IgG expression cassette vectors IFH and IFL, respectively. IFH-VH and IFL-VL vectors were co-expressed in 293F cells, yielding chimeric H5M9 (cH5M9) (Figure 1B).

Bottom Line: While vaccines and antiviral drugs are currently under development, neutralizing antibodies could offer an alternative strategy to prevent and treat H5N1 virus infection.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.

View Article: PubMed Central - PubMed

Affiliation: School of Life Science and Technology, Tongji University, Shanghai, China ; Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Center at Shanghai, Shanghai, China.

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.

Show MeSH
Related in: MedlinePlus