Guiding the Immune Response against Influenza Virus Hemagglutinin toward the Conserved Stalk Domain by Hyperglycosylation of the Globular Head Domain

Abstract Due to limitations of current influenza virus vaccines, new vaccines that mediate broad protection and show high efficacy against seasonal and pandemic viruses are urgently needed. The conserved stalk of the viral hemagglutinin has been identified as potential target antigen for this new generation of vaccines. A vaccination strategy based on chimeric hemagglutinin (cHA), which refocuses the immune response toward the stalk domain and the conserved neuraminidase, is currently being tested in clinical trials. Here we discuss how to improve the cHA antigens to generate vaccine candidates that both induce a broad antistalk response and target conserved immunosubdominant epitopes in the head domain of the hemagglutinin. These novel constructs, termed mosaic hemagglutinins, should provide enhanced protection and should be tested in clinical trials to assess their improved potential as universal influenza virus vaccine candidates.

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Protective Immunity Based on the Conserved Hemagglutinin Stalk Domain and Its Prospects for Universal Influenza Vaccine Development

BioMed Research International ◽

10.1155/2014/546274 ◽

2014 ◽

Vol 2014 ◽

pp. 1-7 ◽

Cited By ~ 24

Author(s):

Madhu Khanna ◽

Sachin Sharma ◽

Binod Kumar ◽

Roopali Rajput

Keyword(s):

Influenza Virus ◽

Influenza Vaccine ◽

Natural Infection ◽

Influenza Viruses ◽

Cross Protection ◽

Influenza B ◽

Universal Vaccine ◽

Virus Surface ◽

Head Domain ◽

Globular Head

Influenza virus surface glycoprotein hemagglutinin (HA) is an excellent and chief target thatelicitsneutralizing antibodies during vaccination or natural infection. Its HA2 subunit (stem domain) is most conserved as compared to HA1 subunit (globular head domain). Current influenza vaccine relies on globular head domain that provides protection only against the homologous vaccine strains, rarely provides cross-protection against divergent strains, and needs to be updated annually. There is an urge for a truly universal vaccine that provides broad cross-protection against different subtype influenza A viruses along with influenza B viruses and need not be updated annually. Antibodies against the stem domain of hemagglutinin (HA) are able to neutralize a wide spectrum of influenza virus strains and subtypes. These stem-specific antibodies have great potential for the development of universal vaccine against influenza viruses. In this review, we have discussed the stem-specific cross-reactive antibodies and heterosubtypic protection provided by them. We have also discussed their epitope-based DNA vaccine and their future prospects in this scenario.

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Generation of antibody diversity in the immune response of BALB/c mice to influenza virus hemagglutinin.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.81.10.3180 ◽

1984 ◽

Vol 81 (10) ◽

pp. 3180-3184 ◽

Cited By ~ 305

Author(s):

D. McKean ◽

K. Huppi ◽

M. Bell ◽

L. Staudt ◽

W. Gerhard ◽

...

Keyword(s):

Immune Response ◽

Influenza Virus ◽

Antibody Diversity ◽

Influenza Virus Hemagglutinin

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Disulfide bond formation during the folding of influenza virus hemagglutinin.

The Journal of Cell Biology ◽

10.1083/jcb.118.2.227 ◽

1992 ◽

Vol 118 (2) ◽

pp. 227-244 ◽

Cited By ~ 57

Author(s):

M S Segal ◽

J M Bye ◽

J F Sambrook ◽

M J Gething

Keyword(s):

Influenza Virus ◽

Disulfide Bonds ◽

Intracellular Transport ◽

Cellular Protein ◽

Cysteine Residues ◽

Mutant Proteins ◽

Influenza Virus Hemagglutinin ◽

Stalk Domain ◽

Do So

To study the importance of individual sulfhydryl residues during the folding and assembly in vivo of influenza virus hemagglutinin (HA), we have constructed and expressed a series of mutant HA proteins in which cysteines involved in three disulfide bonds have been substituted by serine residues. Investigations of the structure and intracellular transport of the mutant proteins indicate that (a) cysteine residues in the ectodomain are essential both for efficient folding of HA and for stabilization of the folded molecule; (b) cysteine residues in the globular portion of the ectodomain are likely to form native disulfide bonds rapidly and directly, without involvement of intermediate, nonnative linkages; and (c) cysteine residues in the stalk portion of the ectodomain also appear not to form intermediate disulfide bonds, even though they have the opportunity to do so, being separated from their correct partners by hundreds of amino acids including two or more other sulfhydryl residues. We propose a role for the cellular protein BiP in shielding the cysteine residues of the stalk domain during the folding process, thus preventing them from forming intermediate, nonnative disulfide bonds.

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A retro-inverso peptide analogue of influenza virus hemagglutinin B-cell epitope 91–108 induces a strong mucosal and systemic immune response and confers protection in mice after intranasal immunization

Molecular Immunology ◽

10.1016/s0161-5890(02)00095-0 ◽

2002 ◽

Vol 39 (5-6) ◽

pp. 323-331 ◽

Cited By ~ 22

Author(s):

Tamar Ben-Yedidia ◽

Anne-Sophie Beignon ◽

Charalambos D. Partidos ◽

Sylviane Muller ◽

Ruth Arnon

Keyword(s):

Immune Response ◽

Influenza Virus ◽

B Cell ◽

Cell Epitope ◽

Systemic Immune Response ◽

Peptide Analogue ◽

Intranasal Immunization ◽

Influenza Virus Hemagglutinin ◽

B Cell Epitope

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Unmasking Stem-Specific Neutralizing Epitopes by Abolishing N-Linked Glycosylation Sites of Influenza Virus Hemagglutinin Proteins for Vaccine Design

Journal of Virology ◽

10.1128/jvi.00880-16 ◽

2016 ◽

Vol 90 (19) ◽

pp. 8496-8508 ◽

Cited By ~ 23

Author(s):

Wen-Chun Liu ◽

Jia-Tsrong Jan ◽

Yun-Ju Huang ◽

Ting-Hsuan Chen ◽

Suh-Chin Wu

Keyword(s):

Influenza Virus ◽

Neutralizing Antibody ◽

Influenza Vaccines ◽

Mutant Proteins ◽

Fusion Inhibition ◽

Influenza Virus Hemagglutinin ◽

Glycosylation Sites ◽

Antibody Titers ◽

Virus Strains ◽

Globular Head

ABSTRACTInfluenza virus hemagglutinin (HA) protein consists of two components, i.e., a globular head region and a stem region that are folded within six disulfide bonds, plus several N-linked glycans that produce a homotrimeric complex structure. While N-linked glycosylation sites on the globular head are variable among different strains and different subtypes, N-linked glycosylation sites in the stem region are mostly well conserved among various influenza virus strains. Targeting highly conserved HA stem regions has been proposed as a useful strategy for designing universal influenza vaccines. Since the HA stem region is constituted by an HA1 N-terminal part and a full HA2 part, we expressed a series of recombinant HA mutant proteins with deleted N-linked glycosylation sites in the HA1 stem and HA2 stem regions of H5N1 and pH1N1 viruses. Unmasking N-glycans in the HA2 stem region (H5 N484A and H1 N503A) was found to elicit more potent neutralizing antibody titers against homologous, heterologous, and heterosubtypic viruses. Unmasking the HA2 stem N-glycans of H5HA but not H1HA resulted in more CR6261-like and FI6v3-like antibodies and also correlated with the increase of cell fusion inhibition activity in antisera. Only H5 N484A HA2 stem mutant protein immunization increased the numbers of antibody-secreting cells, germinal center B cells, and memory B cells targeting the stem helix A epitopes in splenocytes. Unmasking the HA2 stem N-glycans of H5HA mutant proteins showed a significantly improvement in the protection against homologous virus challenges but did so to a less degree for the protection against heterosubtypic pH1N1 virus challenges. These results may provide useful information for designing more effective influenza vaccines.IMPORTANCEN-linked glycosylation sites in the stem regions of influenza virus hemagglutinin (HA) proteins are mostly well conserved among various influenza virus strains. Targeting highly conserved HA stem regions has been proposed as a useful strategy for designing universal influenza vaccines. Our studies indicate that unmasking the HA2 stem N-glycans of recombinant HA proteins from H5N1 and pH1N1 viruses induced more potent neutralizing antibody titers against homologous and heterosubtypic viruses. However, only immunization with the H5N1 HA2 stem mutant protein can refocus B antibody responses to the helix A epitope for inducing more CR6261-like/FI6v3-like and fusion inhibition antibodies in antisera, resulting in a significant improvement for the protection against lethal H5N1 virus challenges. These results may provide useful information for designing more effective influenza vaccines.

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Yeast-Derived Avian Influenza Virus Hemagglutinin Protein Induced Immune Response in SPF Chicken

Journal of Animal and Veterinary Advances ◽

10.3923/javaa.2011.999.1002 ◽

2011 ◽

Vol 10 (8) ◽

pp. 999-1002

Author(s):

Hongzhuan Wu ◽

Karyn Scissum-Gu ◽

Narendra K. Singh ◽

Shan-Chia Ou ◽

Joseph J. Giambrone

Keyword(s):

Immune Response ◽

Influenza Virus ◽

Avian Influenza ◽

Avian Influenza Virus ◽

Influenza Virus Hemagglutinin ◽

Hemagglutinin Protein

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Hemagglutinin Stalk- and Neuraminidase-Specific Monoclonal Antibodies Protect against Lethal H10N8 Influenza Virus Infection in Mice

Journal of Virology ◽

10.1128/jvi.02275-15 ◽

2015 ◽

Vol 90 (2) ◽

pp. 851-861 ◽

Cited By ~ 38

Author(s):

Teddy John Wohlbold ◽

Veronika Chromikova ◽

Gene S. Tan ◽

Philip Meade ◽

Fatima Amanat ◽

...

Keyword(s):

Influenza Virus ◽

Monoclonal Antibodies ◽

Virus Infection ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Morbidity And Mortality ◽

Head Domain ◽

Group 2 ◽

Reactive Antibody ◽

Globular Head

ABSTRACTBetween November 2013 and February 2014, China reported three human cases of H10N8 influenza virus infection in the Jiangxi province, two of which were fatal. Using hybridoma technology, we isolated a panel of H10- and N8-directed monoclonal antibodies (MAbs) and further characterized the binding reactivity of these antibodies (via enzyme-linked immunosorbent assay) to a range of purified virus and recombinant protein substrates. The H10-directed MAbs displayed functional hemagglutination inhibition (HI) and neutralization activity, and the N8-directed antibodies displayed functional neuraminidase inhibition (NI) activity against H10N8. Surprisingly, the HI-reactive H10 antibodies, as well as a previously generated, group 2 hemagglutinin (HA) stalk-reactive antibody, demonstrated NI activity against H10N8 and an H10N7 strain; this phenomenon was absent when virus was treated with detergent, suggesting the anti-HA antibodies inhibited neuraminidase enzymatic activity through steric hindrance. We tested the prophylactic efficacy of one representative H10-reactive, N8-reactive, and group 2 HA stalk-reactive antibodyin vivousing a BALB/c challenge model. All three antibodies were protective at a high dose (5 mg/kg). At a low dose (0.5 mg/kg), only the anti-N8 antibody prevented weight loss. Together, these data suggest that antibody targets other than the globular head domain of the HA may be efficacious in preventing influenza virus-induced morbidity and mortality.IMPORTANCEAvian H10N8 and H10N7 viruses have recently crossed the species barrier, causing morbidity and mortality in humans and other mammals. Although these reports are likely isolated incidents, it is possible that more cases may emerge in future winter seasons, similar to H7N9. Furthermore, regular transmission of avian influenza viruses to humans increases the risk of adaptive mutations and reassortment events, which may result in a novel virus with pandemic potential. Currently, no specific therapeutics or vaccines are available against the H10N8 influenza virus subtype. We generated a panel of H10- and N8-reactive MAbs. Although these antibodies may practically be developed into therapeutic agents, characterizing the protective potential of MAbs that have targets other than the HA globular head domain will provide insight into novel antibody-mediated mechanisms of protection and help to better understand correlates of protection for influenza A virus infection.

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