The antibody response to influenza is primarily focused on the head region of the hemagglutinin (HA) glycoprotein, which in turn undergoes antigenic drift, thus necessitating annual updates of influenza vaccines. In contrast, the immunogenically subdominant stem region of HA is highly conserved and recognized by antibodies capable of binding multiple HA subtypes1,2,3,4,5,6. Here we report the structure-based development of an H1 HA stem–only immunogen that confers heterosubtypic protection in mice and ferrets. Six iterative cycles of structure-based design (Gen1–Gen6) yielded successive H1 HA stabilized-stem (HA–SS) immunogens that lack the immunodominant head domain. Antigenic characterization, determination of two HA–SS crystal structures in complex with stem-specific monoclonal antibodies and cryo-electron microscopy analysis of HA–SS on ferritin nanoparticles (H1–SS–np) confirmed the preservation of key structural elements. Vaccination of mice and ferrets with H1–SS–np elicited broadly cross-reactive antibodies that completely protected mice and partially protected ferrets against lethal heterosubtypic H5N1 influenza virus challenge despite the absence of detectable H5N1 neutralizing activity in vitro. Passive transfer of immunoglobulin from H1–SS–np–immunized mice to naive mice conferred protection against H5N1 challenge, indicating that vaccine-elicited HA stem–specific antibodies can protect against diverse group 1 influenza strains.

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We thank A. Taylor, H. Bao, J.P. Todd and C. Chiedi for help with the animal studies, U. Baxa for EM studies, K. Dai and X. Chen for technical support and B. Hartman for manuscript preparation. We thank M.C. Nason for excellent statistical advice and support. We also thank K. Modjarrad and laboratory members for helpful discussions. This work was supported by the Intramural Research Program of the Vaccine Research Center and the Division of Intramural Research, National Institute of Allergy and Infectious Diseases, US National Institutes of Health. X-ray data were collected at the SER-CAT 22-BM beamline at the Advanced Photon Source (APS), Argonne National Laboratory. Use of APS was supported by the US Department of Energy, Basic Energy Sciences, Office of Science under contract no. W-31-109-Eng-38.

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Author notes

    • Patrick M McTamney
    • , Chih-Jen Wei
    • , Daniel Lingwood
    • , Srinivas S Rao
    •  & Gary J Nabel

    Present addresses: MedImmune, Gaithersburg, Maryland (P.M.M.); Sanofi, Cambridge, Massachusetts (C.-J.W. and G.J.N.); Ragon Institute, Cambridge, Massachusetts (D.L.).

    • Hadi M Yassine
    • , Jeffrey C Boyington
    • , Patrick M McTamney
    •  & Chih-Jen Wei

    These authors contributed equally to this work.


  1. Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.

    • Hadi M Yassine
    • , Jeffrey C Boyington
    • , Patrick M McTamney
    • , Chih-Jen Wei
    • , Masaru Kanekiyo
    • , Wing-Pui Kong
    • , Lingshu Wang
    • , Yi Zhang
    • , M Gordon Joyce
    • , Daniel Lingwood
    • , Syed M Moin
    • , Srinivas S Rao
    • , Peter D Kwong
    • , John R Mascola
    • , Gary J Nabel
    •  & Barney S Graham
  2. Structural Informatics Unit, Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.

    • John R Gallagher
    •  & Audray K Harris
  3. BIOQUAL, Inc., Rockville, Maryland, USA.

    • Hanne Andersen
  4. Kanonji Institute, Research Foundation for Microbial Diseases of Osaka University, Kanonji, Japan.

    • Yoshinobu Okuno


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J.C.B., C.-J.W., and G.J.N. developed the concept of the HA–SS; J.C.B. designed the HA–SS constructs; H.M.Y., J.C.B., P.M.M., C.-J.W., G.J.N., J.R.M. and B.S.G. designed the research studies; H.M.Y., J.C.B., M.K., W.-P.K., L.W., J.R.G., M.G.J., Y.Z., P.M.M., C.-J.W., D.L., S.M.M., A.K.H., P.D.K., G.J.N., J.R.M. and B.S.G. performed the research, analyzed data and discussed the results and implications; S.S.R. and H.A. assisted in animal studies; Y.O. isolated and provided C179 mAb; and J.C.B., H.M.Y., P.M.M., C.-J.W., M.K., P.D.K., J.R.M., G.J.N. and B.S.G. wrote the paper.

Competing interests

The authors declare that an intellectual property application has been filed by the US National Institutes of Health on the basis of data presented in this paper.

Corresponding authors

Correspondence to Gary J Nabel or Barney S Graham.

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