An In Vivo Human-Plasmablast Enrichment Technique Allows Rapid Identification of Therapeutic Influenza A Antibodies

Abstract

Recent advances enabling the cloning of human immunoglobulin G genes have proven effective for discovering monoclonal antibodies with therapeutic potential. However, these antibody-discovery methods are often arduous and identify only a few candidates from numerous antibody-secreting plasma cells or plasmablasts. We describe an in vivo enrichment technique that identifies broadly neutralizing human antibodies with high frequency. For this technique, human peripheral blood mononuclear cells from vaccinated donors are activated and enriched in an antigen-specific manner for the production of numerous antigen-specific plasmablasts. Using this technology, we identified four broadly neutralizing influenza A antibodies by screening only 840 human antibodies. Two of these antibodies neutralize every influenza A human isolate tested and perform better than the current anti-influenza A therapeutic, oseltamivir, in treating severe influenza infection in mice and ferrets. Furthermore, these antibodies elicit robust in vivo synergism when combined with oseltamivir, thus highlighting treatment strategies that could benefit influenza-infected patients.

Relating influenza virus membrane fusion kinetics to stoichiometry of neutralizing antibodies at the single-particle level

“…In contrast, stem-binding antibodies recognize an epitope region that is highly conserved between influenza strains and possess a broad neutralization capacity across many viral subtypes (19,(22)(23)(24)(25)(26) or even across groups (19,27,28). We recently demonstrated that binding of HA by the broadly neutralizing, stem-binding antibodies CR6261 (group 1-specific) and CR8020 (group 2-specific) (22)(23)(24) results in inhibition of HA-mediated viral membrane fusion (29).…”

A Broadly Neutralizing Human Monoclonal Antibody Directed against a Novel Conserved Epitope on the Influenza Virus H3 Hemagglutinin Globular Head

“…Such therapeutic efficacy in preclinical models has been demonstrated previously for other MAbs directed against influenza virus HA (10, 12, 13, 17), suggesting that anti-HA MAbs may be effective therapeutics for treating influenza virus infection. These preclinical data also raise the possibility that the anti-HA MAbs such as D1-8 may be more effective than oseltamivir either alone or in combination with oseltamivir, particularly late in infection (17,41). Anti-HA MAbs have been shown to mediate antiviral activity via multiple mechanisms in preclinical models (in vitro and in vivo), including direct virus neutralization as well as mechanisms dependent on MAb Fc effector function (e.g., antibody-dependent cell-mediated cytotoxicity or complement-dependent cytotoxicity) (13).…”

“…Some of these previously described MAbs target the globular head region of HA (26,27,30,31), while others target the more conserved stalk structure (13,15,17). However, only a few have demonstrated activity against diverse H3 viruses (Ͼ5 viruses), and all such MAbs target the HA stalk structure (13,15,17).…”

“…Several MAbs have been described that neutralize H3 viruses, and some also exhibit cross-reactivity with other subtypes of influenza virus (e.g., H1 strains) (13,15,17,26,27,30,31). Some of these previously described MAbs target the globular head region of HA (26,27,30,31), while others target the more conserved stalk structure (13,15,17).…”

“…Therefore, there remains a significant unmet medical need for new therapies to treat influenza, particularly ones that are effective in high-risk populations (e.g., the elderly) at the time of hospitalization. Recently, a number of preclinical studies with animal models have demonstrated that anti-influenza virus hemagglutinin (HA) monoclonal antibodies (MAbs) have protective activity when administered late in infection, suggesting the potential utility of these agents for treating severe influenza virus infection (10)(11)(12)(13)(14)(15)(16)(17).…”