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Mast cell activators are a novel class of mucosal vaccine adjuvants. The polymeric compound, Compound 48/80 (C48/80), and cationic peptide, Mastoparan 7 (M7) are mast cell activators that provide adjuvant activity when administered by the nasal route. However, small molecule mast cell activators may be a more cost-efficient adjuvant alternative that is easily synthesized with high purity compared to M7 or C48/80. To identify novel mast cell activating compounds that could be evaluated for mucosal vaccine adjuvant activity, we employed high-throughput screening to assess over 55,000 small molecules for mast cell degranulation activity. Fifteen mast cell activating compounds were down-selected to five compounds based onin vitroimmune activation activities including cytokine production and cellular cytotoxicity, synthesis feasibility, and selection for functional diversity. These small molecule mast cell activators were evaluated forin vivoadjuvant activity and induction of protective immunity against West Nile Virus infection in BALB/c mice when combined with West Nile Virus envelope domain III (EDIII) protein in a nasal vaccine. We found that three of the five mast cell activators, ST101036, ST048871, and R529877, evoked high levels of EDIII-specific antibody and conferred comparable levels of protection against WNV challenge. The level of protection provided by these small molecule mast cell activators was comparable to the protection evoked by M7 (67%) but markedly higher than the levels seen with mice immunized with EDIII alone (no adjuvant 33%). Thus, novel small molecule mast cell activators identified by high throughput screening are as efficacious as previously described mast cell activators when used as nasal vaccine adjuvants and represent next-generation mast cell activators for evaluation in mucosal vaccine studies.

Food allergy is a potentially fatal disease affecting 8% of children and has become increasingly common in the past two decades. Despite the prevalence and severe nature of the disease, the mechanisms underlying sensitization remain to be further elucidated. The Collaborative Cross is a genetically diverse panel of inbred mice that were specifically developed to study the influence of genetics on complex diseases. Using this panel of mouse strains, we previously demonstrated CC027/GeniUnc mice, but not C3H/HeJ mice, develop peanut allergy after oral exposure to peanut in the absence of a Th2-skewing adjuvant. Here, we investigated factors associated with sensitization in CC027/GeniUnc mice following oral exposure to peanut, walnut, milk, or egg. CC027/GeniUnc mice mounted antigen-specific IgE responses to peanut, walnut and egg, but not milk, while C3H/HeJ mice were not sensitized to any antigen. Naïve CC027/GeniUnc mice had markedly lower total fecal IgA compared to C3H/HeJ, which was accompanied by stark differences in gut microbiome composition. Sensitized CC027/GeniUnc mice had significantly fewer CD3+ T cells but higher numbers of CXCR5+ B cells and T follicular helper cells in the mesenteric lymph nodes compared to C3H/HeJ mice, which is consistent with their relative immunoglobulin production. After oral challenge to the corresponding food, peanut- and walnut-sensitized CC027/GeniUnc mice experienced anaphylaxis, whereas mice exposed to milk and egg did not. Ara h 2 was detected in serum collected post-challenge from peanut-sensitized mice, indicating increased absorption of this allergen, while Bos d 5 and Gal d 2 were not detected in mice exposed to milk and egg, respectively. Machine learning on the change in gut microbiome composition as a result of food protein exposure identified a unique signature in CC027/GeniUnc mice that experienced anaphylaxis, including the depletion of Akkermansia. Overall, these results demonstrate several factors associated with enteral sensitization in CC027/GeniUnc mice, including diminished total fecal IgA, increased allergen absorption and altered gut microbiome composition. Furthermore, peanuts and tree nuts may have inherent properties distinct from milk and eggs that contribute to allergy.

The development of mucosally administered vaccines remains a goal of many researchers who desire to develop a needle-free method of immunization that can induce antigen-specific immune responses in both systemic and mucosal tissues. The coadministration of adjuvants is often required to maximize the induction of protective immunity following mucosal vaccination, especially when subunit immunogens are used. A major class of mucosal vaccine adjuvants and modulators includes substances that can activate the innate immune system. Innate immune system activators that exhibit mucosal vaccine adjuvant and modulator activity will be discussed, including proinflammatory cytokines, nanoemulsions, mast cell activators, and toll-like receptor ligands.

Mast cells (MCs) are gaining recognition as key initiators and coordinators of host inflammatory and immune responses to various microbial pathogens. Their presence in mucosal tissues and skin make them one of the first immune cells to make contact with incoming pathogens. MCs can be readily activated at infection sites and release their collection of prestored and de novo synthesized mediators to initiate immune responses. Additionally, since MCs are in close proximity to blood vessels, they help to augment immune cell mobilization from the circulation to the infection site. However, not all MC reactions are beneficial, and if MCs become overactivated, as is observed with chronic or systemic infections, pathogenic sequelae ensue. In view of the powerful immunomodulatory role of MCs, there is growing interest in targeting these cells to either boost immunity or diminish harmful inflammatory outcomes of severe or chronic infection.

Mast cells are an important cell type of the innate immune system that when activated, play a crucial role in generating protective innate host responses after bacterial and viral infection. Additionally, activated mast cells influence lymph node composition to regulate the induction of adaptive immune responses. The recognition that mast cells play a beneficial role in host responses to microbial infection and induction of adaptive immunity has provided the rationale to evaluate mast cell activators for use as antimicrobials or vaccine adjuvants. This review summarizes the role of mast cell activators in antimicrobial responses while also discussing the use of different classes of mast cell activators as potent vaccine adjuvants that enhance the induction of protective immune responses.

Mast cells (MCs) are known to regulate innate and adaptive immunity. MC activators have recently been described as safe and effective vaccine adjuvants. However, many currently known MC activators are inadequate for in vivo applications, and research on identifying novel MC activators is limited. In this study, we identified novel MC activators by employing high-throughput screening (HTS) assays using approximately 55,000 small molecules. Data sets obtained by the primary HTS assays were statistically evaluated using quality control rules and the B-score calculation, and compounds with B-scores of > 3.0 were chosen as mast cell activators (hits). These hits were re-evaluated with secondary and tertiary HTS assays, followed by further statistical analysis. From these hits, we selected 15 compounds that caused degranulation in murine and human MCs with potential for flexible chemical modification for further study. Among these 15 compounds, ST101036, ST029248 and ST026567 exhibited higher degranulation potency than other hit compounds in both human and mouse MCs. Additionally, the 15 compounds identified promote de novo synthesis of cytokines and induce the release of eicosanoids from human and mouse MCs. High-throughput screening enabled us to identify small molecule MC activators with unique properties that may be useful as vaccine adjuvants.