Broadly Neutralizing Anti-Influenza Virus Antibodies: Enhancement of Neutralizing Potency in Polyclonal Mixtures and IgA Backbones

Abstract

Current influenza virus vaccines rely upon the accurate prediction of circulating virus strains months in advance of the actual influenza season in order to allow time for vaccine manufacture. Unfortunately, mismatches occur frequently, and even when perfect matches are achieved, suboptimal vaccine efficacy leaves several high-risk populations vulnerable to infection. However, the recent discovery of broadly neutralizing antibodies that target the hemagglutinin (HA) stalk domain has renewed hope that the development of "universal" influenza virus vaccines may be within reach. Here, we examine the functions of influenza A virus hemagglutinin stalk-binding antibodies in an endogenous setting, i.e., as polyclonal preparations isolated from human sera. Relative to monoclonal antibodies that bind to the HA head domain, the neutralization potency of monoclonal stalk-binding antibodies was vastly inferior in vitro but was enhanced by several orders of magnitude in the polyclonal context. Furthermore, we demonstrated a surprising enhancement in IgA-mediated HA stalk neutralization relative to that achieved by antibodies of IgG isotypes. Mechanistically, this could be explained in two ways. Identical variable regions consistently neutralized virus more potently when in an IgA backbone compared to an IgG backbone. In addition, HA-specific memory B cells isolated from human peripheral blood were more likely to be stalk specific when secreting antibodies of IgA isotypes compared to those secreting IgG. Taken together, our data provide strong evidence that HA stalk-binding antibodies perform optimally when in a polyclonal context and that the targeted elicitation of HA stalk-specific IgA should be an important consideration during "universal" influenza virus vaccine design. IMPORTANCEInfluenza viruses remain one of the most worrisome global public health threats due to their capacity to cause pandemics. While seasonal vaccines fail to protect against the emergence of pandemic strains, a new class of broadly neutralizing antibodies has been recently discovered and may be the key to developing a "universal" influenza virus vaccine. While much has been learned about the biology of these antibodies, most studies have focused only on monoclonal antibodies of IgG subtypes. However, the study of monoclonal antibodies often fails to capture the complexity of antibody functions that occur during natural polyclonal responses. Here, we provide the first detailed analyses of the biological activity of these antibodies in polyclonal contexts, comparing both IgG and IgA isotypes isolated from human donors. The striking differences observed in the functional properties of broadly neutralizing antibodies in polyclonal contexts will be essential for guiding design of "universal" influenza virus vaccines and therapeutics. I nfluenza A viruses (IAVs) remain one of the most pressing global public health concerns due to their widespread distribution, rapid evolution, and potential for reassortment (1). These traits contribute to the abil...

Sequential Infection in Ferrets with Antigenically Distinct Seasonal H1N1 Influenza Viruses Boosts Hemagglutinin Stalk-Specific Antibodies

“…Recent evidence also highlights the importance of secretory IgA in protecting the respiratory tract against influenza virus infection (44,45). He et al demonstrated that HA stalk-specific MAbs exhibit superior neutralization in vitro when they are expressed in the context of the IgA heavy chain (24). Collectively, these observations support the notion that ferrets sequentially infected with sH1N1 influenza virus exhibited superior protection against novel H1N1 challenge due to an increased abundance of HA stalk-specific Ig.…”

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

“…4B), and total HK/68 NAbinding antibodies (Fig. 4C), as described previously (2,3,18). As expected, IgG antibodies from each donor were capable of inducing ADCC when added to the infected monolayer.…”

“…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, recent work by DiLillo et al (20) has demonstrated that HA stalk-binding antibodies are potent inducers of antibody-dependent cell-mediated cytotoxicity (ADCC), and that this property is essential for optimal protection in vivo.…”

Optimal activation of Fc-mediated effector functions by influenza virus hemagglutinin antibodies requires two points of contact

“…However, the in vitro neutralizing activity of stalk-specific monoclonal antibodies (mAbs) is generally inferior to that of hemagglutination-inhibiting mAbs, and differences in their inhibitory concentrations can approach 2-to 3-log differences (19,20). This disparity in vitro between head-specific and stalkspecific antibodies can be significantly reduced in vivo to approximately fivefold in passive transfer challenge studies (19,21).…”