Abstract
Vaccines remain the most effective tool to prevent infectious diseases. Here, we introduce an in vitro booster vaccination approach that relies on antigen-dependent activation of human memory B cells in culture. This stimulation induces antigen-specific B cell proliferation, differentiation of B cells into plasma cells, and robust antibody secretion after a few days of culture. We validated this strategy using cells from healthy donors to retrieve human antibodies against tetanus toxoid and influenza hemagglutinin (HA) from H1N1 and newly emergent subtypes such as H5N1 and H7N9. Anti-HA antibodies were cross-reactive against multiple subtypes, and some showed neutralizing activity. Although these antibodies may have arisen as a result of previous influenza infection, we also obtained gp120-reactive antibodies from non-HIV-infected donors, indicating that we can generate antibodies without prior antigenic exposure. Overall, our novel approach can be used to rapidly produce therapeutic antibodies and has the potential to assess the immunogenicity of candidate antigens, which could be exploited in future vaccine development.
Highlights
B lymphocytes (B cells) play a critical role in adaptive immunity, providing protection from pathogens through the production of specific antibodies
In vitro stimulation of memory B cells by particulate CpG in a B cell receptor (BCR)-dependent manner Previous studies have demonstrated that the in vitro stimulation of human B cells by TLRs is an efficient way of inducing the activation and proliferation of these cells, irrespective of BCR specificity (Bernasconi et al, 2003; Pinna et al, 2009). These findings have been exploited as an efficient way of retrieving human monoclonal antibodies in vitro (Traggiai et al, 2004).We hypothesized that the delivery of a TLR ligand via BCR-mediated internalization might represent a way of exclusively activating B cells with a particular BCR specificity.To determine the best stimulatory conditions for such activation, we first took advantage of an anti–κ-chain antibody to target CpG to the BCR, because 60% of circulating human B cells bear this BCR chain; the rest bear a λ chain.We coated streptavidin polystyrene nanoparticles with a mixture of biotinylated anti-κ antibody and the TLR ligand CpG, as we have previously described in mice (Eckl-Dorna and Batista, 2009)
To determine whether we could use these nanoparticles in vitro to selectively activate only those memory B cells expressing a κ-chain BCR, CellTrace Violet–labeled human memory B cells obtained from healthy donors were cultured in the presence of a cytokine cocktail with nanoparticles coated with either anti-κ and CpG or CpG alone; soluble CpG; or no stimulant as a control. 6 d after stimulation, flow cytometry was used to measure B cell proliferation by the dilution of the CellTrace Violet
Summary
B lymphocytes (B cells) play a critical role in adaptive immunity, providing protection from pathogens through the production of specific antibodies. B cells recognize and respond to pathogen-derived antigens through surface B cell receptors (BCRs). The second is to mediate antigen uptake and processing, leading to antigen presentation to T cells within the MHC class II context and full activation of the B cells (Lanzavecchia, 1985). BCR-mediated antigen internalization has been shown to facilitate the presentation of lipid antigens in the context of CD1d, which can result in the recruitment of iNKT cell help (Barral et al, 2008; Leadbetter et al, 2008) or the transport of TLR agonists, resulting in TLR7 or TLR9 signaling (Marshak-Rothstein, 2006; Hou et al, 2011).
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