Co-stimulation With TLR7 Agonist Imiquimod and Inactivated Influenza Virus Particles Promotes Mouse B Cell Activation, Differentiation, and Accelerated Antigen Specific Antibody Production

Abstract

Current influenza vaccines have relatively low effectiveness, especially against antigenically drifted strains, the effectiveness is even lower in the elderly and immunosuppressed individuals. We have previously shown in a randomized clinical trial that the topical application of a toll-like receptor 7 agonist, imiquimod, just before intradermal influenza vaccine could expedite and augment antibody response, including to antigenically-drifted strains. However, the mechanism of this vaccine and imiquimod combination approach is poorly understood. Here, we demonstrated that imiquimod alone directly activated purified mouse peritoneal B cells. When combined with inactivated H1N1/415742Md influenza virus particle (VP) as vaccine, co-stimulation of mouse peritoneal B cells in vitro induced stronger activation, proliferation, and production of virus-antigen specific IgM and IgG. Intraperitoneal injection of a combination of VP and imiquimod (VCI) was associated with an increased number of activated B cells with enhanced expression of CD86 in the mesenteric draining lymph nodes (mesLN) and the spleen at 18 h after injection. Three days after immunization with VCI, mouse spleen showed significantly more IgM and IgG secreting cells upon in vitro re-stimulation with inactivated virus, mouse sera were detected with viral neutralizing antibody. Transfer of these spleen B cells to naïve mice improved survival after lethal dose of H1N1/415742Md challenge. More importantly, the functional response of VCI-induced B cell activation was demonstrated by early challenge with a lethal dose of H1N1/415742Md influenza virus at 3 days after immunization. The spleen and mediastinal lymph nodes (mdLN) in mice immunized with VCI had germinal center formation, and significantly higher number of plasmablasts, plasma cells, and virus-antigen specific IgM and IgG secreting cells at only 3–4 days post virus challenge, compared with those of mice that have received imiquimod, inactivated virus alone or PBS. Serum virus-specific IgG2a, IgG2b, and IgG1 and bronchoalveolar lavage fluid (BALF) virus-specific IgA at 3 or 4 days post challenge were significantly higher in mice immunized with VCI, which had significantly reduced lung viral load and 100% survival. These findings suggested that imiquimod accelerates the vaccine-induced antibody production via inducing rapid differentiation of naïve B cells into antigen-specific antibody producing cells.

Repurposing of Miltefosine as an Adjuvant for Influenza Vaccine

“…The improved antibody response in the MTF-VAC group was associated with a more robust T FH response and a more potent germinal center reaction. This is in contrast with our previous study on the TLR7 agonist, imiquimod, in which there was no difference in the GC reaction between the imiquimod-adjuvanted group and non-adjuvanted group [ 27 ]. T FH cell is the key T cell subtype that interacts with B cells in lymphoid follicles [ 39 ].…”

“…The hemagglutination inhibition assay (HAI), micro-neutralization assay (MN), and enzyme-linked immunosorbent assay (ELISA) assays were performed as previously described [ 13 , 27 ].…”

Distinguishing naive‐ from memory‐derived human B cells during acute responses

“…Similarly, Kwissa et al 21 found that CD14 + CD16 + monocytes, which are induced by dengue virus infection, and by R848 but not CpG, enhance B-cell differentiation, providing another possible explanation for the differential role of monocytes in R848-versus CpG-stimulated B-cell differentiation. The capacity of TLR7/8 agonists to induce B-cell differentiation is also apparent in vivo whereby adjuvanting inactivated influenza vaccine with a TLR7 agonist accelerates B-cell differentiation and antibody secretion in mice 39 and non-human primate neonates. 40 Analogously, inactivated wholevirion influenza vaccine, as opposed to subunit or split virion vaccines, induces TLR7-mediated B-cell activation and increased antibody production in mice.…”

The role of topical imiquimod in melanoma cutaneous metastases: A critical review of the literature

“…The exact mechanism has not been fully understood, but some authors hypothesized a role of the co-stimulation of CD40 ligand-like action expressed by imiquimod cytokines, of the Blimp-1 gene transcription with a strong induction of IgM secretion and of the high capacity of TRL7 to induce naïve B cells into antigen-specific antibody producing cells. 28,29 Adenosine receptor, especially A2A receptor, represents another molecular target of imiquimod. By this, imiquimod inhibits a negative feedback mechanism that normally limits the pro-inflammatory reactions resulting in a synergic effect of TRLs' signal through a maintained production of inflammatory cytokines (Figure 1).…”

“…The result is a strong B cells proliferation, activation and production of antigen specific IgM and IgG. The exact mechanism has not been fully understood, but some authors hypothesized a role of the co‐stimulation of CD40 ligand‐like action expressed by imiquimod cytokines, of the Blimp‐1 gene transcription with a strong induction of IgM secretion and of the high capacity of TRL7 to induce naïve B cells into antigen‐specific antibody producing cells 28,29 …”