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Broadly protective murine monoclonal antibodies against influenza B virus target highly conserved neuraminidase epitopes

Abstract

A substantial proportion of influenza-related childhood deaths are due to infection with influenza B viruses, which co-circulate in the human population as two antigenically distinct lineages defined by the immunodominant receptor binding protein, haemagglutinin. While broadly cross-reactive, protective monoclonal antibodies against the haemagglutinin of influenza B viruses have been described, none targeting the neuraminidase, the second most abundant viral glycoprotein, have been reported. Here, we analyse a panel of five murine anti-neuraminidase monoclonal antibodies that demonstrate broad binding, neuraminidase inhibition, in vitro antibody-dependent cell-mediated cytotoxicity and in vivo protection against influenza B viruses belonging to both haemagglutinin lineages and spanning over 70 years of antigenic drift. Electron microscopic analysis of two neuraminidase–antibody complexes shows that the conserved neuraminidase epitopes are located on the head of the molecule and that they are distinct from the enzymatic active site. In the mouse model, one therapeutic dose of antibody 1F2 was more protective than the current standard of treatment, oseltamivir, given twice daily for six days.

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Fig. 1: In vitro binding of IBV anti-NA mAbs.
Fig. 2: Negative-stain electron microscopy analysis of NA structures reveals binding footprints for 1F2 and 4F11.
Fig. 3: In vivo efficacy of IBV anti-NA mAbs.
Fig. 4: Non-neutralizing IBV anti-NA mAbs reduce viral lung titres in mice, activate ADCC, inhibit activity of a drug-resistant IBV and demonstrate superior effectiveness to oseltamivir.

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Acknowledgements

The authors thank I. Margine for pilot studies, M. Rajendran Marilyne Panis and R. Nachbagauer for their assistance in the deep sequencing analysis of IBV mutants and competition ELISAs, A. Hirsh for producing recombinant neuraminidase proteins and N. Bouvier for her instructions regarding oral gavaging. We thank A. Hurt (WHO Influenza Collaborating Centre For Reference And Research On Influenza, Melbourne, Australia) and E. Govorkova (St. Jude Children's Hospital, Memphis, TN) for providing NA-inhibitor resistant influenza B virus isolates. This work was funded by NIAID grants R01 AI117287 (to F.K.) and U19 AI109946 (to P.P. and F.K.).

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T.J.W., K.A.P., S.S. and F.K. designed experiments and wrote the manuscript. T.J.W., K.A.P., V.C. and P.M. performed experiments. J.T. and F.A. assisted with experiments. F.A. and G.S.T. generated reagents. T.J.W., K.A.P., V.C., V.F., J.T., E.K., B.R.t., P.P., S.S. and F.K. analysed and interpreted data.

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Correspondence to Florian Krammer.

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The Icahn School of Medicine at Mount Sinai has filed patents regarding use of the described mAbs as therapeutics (application no. 62/483,262). T.J.W., P.P. and F.K. are named as inventors on the application.

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Wohlbold, T.J., Podolsky, K.A., Chromikova, V. et al. Broadly protective murine monoclonal antibodies against influenza B virus target highly conserved neuraminidase epitopes. Nat Microbiol 2, 1415–1424 (2017). https://doi.org/10.1038/s41564-017-0011-8

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