Vaccination with Adjuvanted Recombinant Neuraminidase Induces Broad Heterologous, but Not Heterosubtypic, Cross-Protection against Influenza Virus Infection in Mice

Wohlbold, Teddy John; Nachbagauer, Raffael; Xu, Haoming; Tan, Gene S.; Hirsh, Ariana; Brokstad, Karl Albert; Cox, Rebecca Jane; Palese, Peter; Krammer, Florian

doi:10.1128/mbio.02556-14

Cited by 179 publications

(250 citation statements)

References 42 publications

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“…In addition to HA, a substantial antibody response is also generated against the second major IAV surface glycoprotein, NA. Although present in substantially lower quantities than HA-binding antibodies, NAbinding antibodies can inhibit NA activity, which has been shown to confer protection from infection in vivo (28)(29)(30). Certain HAbinding antibodies have been shown to exhibit NA-inhibiting (NAI) activity through steric hindrance of the NA enzymatic site (28).…”

Section: Haimentioning

confidence: 99%

Epitope specificity plays a critical role in regulating antibody-dependent cell-mediated cytotoxicity against influenza A virus

Tan

Mullarkey

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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157

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The generation of strain-specific neutralizing antibodies against influenza A virus is known to confer potent protection against homologous infections. The majority of these antibodies bind to the hemagglutinin (HA) head domain and function by blocking the receptor binding site, preventing infection of host cells. Recently, elicitation of broadly neutralizing antibodies which target the conserved HA stalk domain has become a promising "universal" influenza virus vaccine strategy. The ability of these antibodies to elicit Fc-dependent effector functions has emerged as an important mechanism through which protection is achieved in vivo. However, the way in which Fc-dependent effector functions are regulated by polyclonal influenza virus-binding antibody mixtures in vivo has never been defined. Here, we demonstrate that interactions among viral glycoprotein-binding antibodies of varying specificities regulate the magnitude of antibody-dependent cell-mediated cytotoxicity induction. We show that the mechanism responsible for this phenotype relies upon competition for binding to HA on the surface of infected cells and virus particles. Nonneutralizing antibodies were poor inducers and did not inhibit antibody-dependent cell-mediated cytotoxicity. Interestingly, anti-neuraminidase antibodies weakly induced antibody-dependent cell-mediated cytotoxicity and enhanced induction in the presence of HA stalk-binding antibodies in an additive manner. Our data demonstrate that antibody specificity plays an important role in the regulation of ADCC, and that cross-talk among antibodies of varying specificities determines the magnitude of Fc receptor-mediated effector functions.T he discovery and ongoing characterization of broadly neutralizing antibodies (bnAbs) that bind to the hemagglutinin (HA) stalk domain of influenza A viruses (IAVs) has galvanized promising new efforts to generate a universal influenza virus vaccine (1). A number of studies have now firmly established that stalk-specific bnAbs, normally present in low quantities, can be boosted substantially in humans following exposure to HA subtypes with antigenically foreign head domains (2-8). Sequential vaccination of animals with chimeric HAs or with "headless" vaccine constructs have effectively recapitulated the boosting of bnAbs observed in humans after exposure to foreign HAs, and also are protective against heterologous and heterosubtypic IAV challenge (9-15). These strategies are now being tested as promising "universal" influenza virus vaccine candidates (16).Whereas traditional strain-specific antibodies neutralize virus by inhibiting receptor binding, stalk-binding bnAbs neutralize virus by distinct postbinding mechanisms (17). A direct comparison of in vitro neutralization has revealed that strain-specific mAbs that inhibit receptor binding tend to be more potent at neutralizing virus compared with monoclonal stalk-binding bnAbs (18, 19); however, this difference is minimized in the context of a polyclonal response (18). In addition to virus neutralization,...

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Section: Haimentioning

confidence: 99%

Epitope specificity plays a critical role in regulating antibody-dependent cell-mediated cytotoxicity against influenza A virus

Tan

Mullarkey

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

157

158

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“…NA inhibition (NI) assays were performed as described in detail earlier (25). In brief, 96-well, flat-bottom, nonsterile Immulon 4 HBX plates (Thermo Scientific) were coated with 150 l of fetuin (Sigma) at a concentration of 50 g/l and refrigerated at 4°C overnight.…”

Section: Fig 1 Anti-h10 and Anti-n8 Antibodies Display Heterologous Cmentioning

confidence: 99%

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

Wohlbold

Chromikova

Tan

et al. 2016

J Virol

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Between 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 antibody in vivo using 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. R ecently, avian influenza A viruses of the H10 subtype have been reported to infect seals and humans and have generated concern over their pandemic potential. Three human cases of H10N8 virus have been reported in China so far, two of which were fatal (1-3). Furthermore, an avian H10N7 strain was found to be the etiological agent responsible for the massive die-off harbor seals in the Baltic Sea, an epidemic that killed more than 10% of the local seal population (4-6). The receptor binding profile of H10 viruses is currently debated (7-12), but the subtype has been proven to cause ...

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“…Within each NA subtype, there are genetically heterologous lineages that are serologically distinct (18,19), suggesting the presence of strain-specific epitopes; however, analyses with polyclonal sera and monoclonal antibodies (MAbs) also demonstrate that NA contains conserved epitopes, in addition to immunodominant strain-specific epitopes (20)(21)(22)(23)(24). A universal epitope in NA has been reported, and a rabbit MAb against this epitope was shown to inhibit multiple subtypes of NA (25,26), although it remains to be investigated whether this epitope elicits an immune response in vivo.…”

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confidence: 99%

Comparative Efficacy of Monoclonal Antibodies That Bind to Different Epitopes of the 2009 Pandemic H1N1 Influenza Virus Neuraminidase

Jiang

Fantoni

Couzens

et al. 2016

J Virol

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Antibodies against the neuraminidase (NA) of influenza virus correlate with resistance against disease, but the effectiveness of antibodies against different NA epitopes has not been compared. In the present study, we evaluated the in vitro and in vivo efficacies of four monoclonal antibodies (MAbs): HF5 and CD6, which are specific to two different epitopes in the NA of 2009 pandemic H1N1 (pH1N1) virus, and 4E9 and 1H5, which are specific to a conserved epitope in the NA of both H1N1 and H5N1 viruses. In the in vitro assays, HF5 and CD6 inhibited virus spread and growth more effectively than 4E9 and 1H5, with HF5 being the most effective inhibitor. When administered prophylactically at 5 mg/kg of body weight, HF5 and CD6 protected ϳ90 to 100% of DBA/2 mice against lethal wild-type pH1N1 virus challenge; however, at a lower dose (1 mg/kg), HF5 protected ϳ90% of mice, whereas CD6 protected only 25% of mice. 4E9 and 1H5 were less effective than HF5 and CD6, as indicated by the partial protection achieved even at doses as high as 15 mg/kg. When administered therapeutically, HF5 protected a greater proportion of mice against lethal pH1N1 challenge than CD6. However, HF5 quickly selected pH1N1 virus escape mutants in both prophylactic and therapeutic treatments, while CD6 did not. Our findings confirm the important role of NA-specific antibodies in immunity to influenza virus and provide insight into the properties of NA antibodies that may serve as good candidates for therapeutics against influenza. IMPORTANCENeuraminidase (NA) is one of the major surface proteins of influenza virus, serving as an important target for antivirals and therapeutic antibodies. The impact of NA-specific antibodies on NA activity and virus replication is likely to depend on where the antibody binds. Using in vitro assays and the mouse model, we compared the inhibitory/protective efficacy of four mouse monoclonal antibodies (MAbs) that bind to different sites within the 2009 pandemic H1N1 (pH1N1) virus NA. The ability of each MAb to protect mice against lethal pH1N1 infection corresponded to its ability to inhibit NA activity in vitro; however, the MAb that was the most effective inhibitor of NA activity selected pH1N1 escape variants in vivo. One of the tested MAbs, which binds to a conserved region in the NA of pH1N1 virus, inhibited NA activity but did not result in escape variants, highlighting its suitability for development as a therapeutic agent.

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Vaccination with Adjuvanted Recombinant Neuraminidase Induces Broad Heterologous, but Not Heterosubtypic, Cross-Protection against Influenza Virus Infection in Mice

Cited by 179 publications

References 42 publications

Epitope specificity plays a critical role in regulating antibody-dependent cell-mediated cytotoxicity against influenza A virus

Epitope specificity plays a critical role in regulating antibody-dependent cell-mediated cytotoxicity against influenza A virus

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

Comparative Efficacy of Monoclonal Antibodies That Bind to Different Epitopes of the 2009 Pandemic H1N1 Influenza Virus Neuraminidase

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