Novel adjuvants in allergen-specific immunotherapy: where do we stand?

Abstract

Type I hypersensitivity, or so-called type I allergy, is caused by Th2-mediated immune responses directed against otherwise harmless environmental antigens. Currently, allergen-specific immunotherapy (AIT) is the only disease-modifying treatment with the potential to re-establish clinical tolerance towards the corresponding allergen(s). However, conventional AIT has certain drawbacks, including long treatment durations, the risk of inducing allergic side effects, and the fact that allergens by themselves have a rather low immunogenicity. To improve AIT, adjuvants can be a powerful tool not only to increase the immunogenicity of co-applied allergens but also to induce the desired immune activation, such as promoting allergen-specific Th1- or regulatory responses. This review summarizes the knowledge on adjuvants currently approved for use in human AIT: aluminum hydroxide, calcium phosphate, microcrystalline tyrosine, and MPLA, as well as novel adjuvants that have been studied in recent years: oil-in-water emulsions, virus-like particles, viral components, carbohydrate-based adjuvants (QS-21, glucans, and mannan) and TLR-ligands (flagellin and CpG-ODN). The investigated adjuvants show distinct properties, such as prolonging allergen release at the injection site, inducing allergen-specific IgG production while also reducing IgE levels, as well as promoting differentiation and activation of different immune cells. In the future, better understanding of the immunological mechanisms underlying the effects of these adjuvants in clinical settings may help us to improve AIT.

Keywords: CpG; Th1/Th2 responses; adjuvant; allergen-specific immunotherapy; flagellin; mannan; type I hypersensitivity; virus-like particle.

Figure 1

Boosting of immune responses by first- and second-generation adjuvants. First-generation adjuvants can adsorb soluble allergens through electrostatic interactions with the allergens hydroxyl groups to form insoluble aggregates (A). Micron-sized adjuvant-allergen aggregates can (I) induce activation of immune cells, which further recruit and activate APCs, (II) be directly taken up by APCs, or (III) act as depots to effectively enhance allergen-specific antibody production. In contrast, second-generation adjuvants activate immune cells by triggering the activation of APCs via their PRRs (B). Here, the binding of PAMPs (as second-generation adjuvants) to PRRs can induce transcriptional changes in APCs, leading to increased expression of co-stimulatory molecules and cytokine secretion. In addition, combining second-generation adjuvants with allergens can induce effective allergen uptake by APCs which then present allergen-derived peptides in the context of the adjuvant-induced APC activation. This results in effective T cell activation, thus triggering adaptive immune responses. For more information, see text. OH, hydroxyl group; MCT, microcrystalline tyrosine; NLRP3, NOD-, LRR- and pyrin domain-containing protein 3; PAMPs, pathogen-associated molecular patterns; DAMPs, damage-associated molecular patterns; APC, antigen-presenting cells; PRR, pathogen recognition receptor; Ag, antigen.

Figure 2

Adjuvants currently studied to improve AIT. The types of adjuvants can be classified based on their carrier or immunostimulatory capabilities. Here, liposomes serve as carriers, while virus-like particles, alum, and O/W emulsions have both carrier function and immune stimulatory characteristics. Besides viral components (the T-cell epitope derived from the hepatitis B virus PreS or the HIV-trans-activator of transcription (TAT)), TLR-ligands (flagellin, CpG, and MPLA), and carbohydrate-based adjuvants (QS-21, glycan, and mannan) have immunomodulatory properties triggering different kinds of immune responses. The described properties of these adjuvants to induce either Th1 or Th2 responses are indicated below the respective adjuvant. For more information, see text. AS, adjuvant system, alum: aluminum hydroxide; O/W, oil-in-water; Pres, HBV-derived T-cell epitope; TAT, HIV type 1 trans-activating regulatory protein; TLR, “Toll”-like receptor; MPLA, monophosphoryl lipid A. Figure modified after (17).

Figure 3

Types of VLPs investigated pre-clinically in AIT and their immunological effects. VLPs have been applied in pre-clinical studies to improve the treatment of allergies either as empty particles, packaging immune-stimulatory CpG-motifs, displaying either cytokines or allergen(s) on their surface, co-displaying both allergen(s) and immune-modulating proteins on their surface, packaging allergen(s) inside the VLPs, or mixed with allergen(s). VLPs were shown to thymus-independently activate naïve B cells, resulting in production of potentially neutralizing antibodies while also promoting the activation of plasmacytoid DCs (pDCs) (–94). These APCs in turn favor the induction of regulatory- and Th1 cells which suppress the allergy-causing Th2 cells. Together these effects of VLPs may result in reduced mast cell activation and allergic inflammation [reviewed in (90)]. (p)DC, (plasmacytoid) dendritic cell; VLP, virus-like particle; IgG/A/E, immunoglobulin G/A/E.

Figure 4

Structure of alpha- and beta-glucans. Depicted the difference between alpha- and beta-(D)-glucose, the different types of glycosidic bonds, and some simplified exemplary alpha- and beta-glucan structures.

Figure 5

Immune modulatory properties of non-oxidized mannan:allergoid conjugates. Non-oxidized mannan:allergoid conjugates were shown to reduce skin prick test reactivity compared to either grass pollen allergens or grass pollen-based allergoids (right side). Immunologically, the mannan:allergoid conjugates taken up via DC-SIGN and mannose receptor induced higher secretion of IL-6 and IL-10 as well as surface expression of PD-L1 paralleled by an increase in mTOR-dependent glycolysis (left side). In in vivo analyses, mannan:allergoid conjugates stimulated the differentiation of CD4⁺CD25^highFOXP3⁺ T_reg cells, paralleled by increased pollen-specific IgG_2a/IgE ratios. In grass pollen allergic patients, the mannan allergoid conjugates induced lower skin prick test reactivity compared to either non-modified allergens or the non-conjugated allergoids alone. Results summarized according to (127). For more information see text. DC-SIGN, dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin; FOXP3, forkhead box protein 3; Mannose R, mannose receptor; mTOR, mammalian target of rapamycin.

Figure 6

Immune modulating properties of flagellin:allergen fusion proteins. Flagellin:allergen fusion proteins combining the TLR5- and NLRC4-ligand flagellin and different allergens into a single molecule were shown to strongly activate both myeloid DCs and macrophages, resulting in the secretion of both pro- and anti-inflammatory cytokines and the expression of co-stimulatory molecules. The activated APCs promoted the induction of Th1 responses both in vitro and in vivo while also efficiently suppressing Th2 responses in an IL-10-dependent manner. This pro-tolerogenic phenotype of the APCs was critically dependent on a metabolic shift towards increased glycolytic activity termed the Warburg effect. The fusion protein consisting of flagellin A and the major birch pollen allergen Bet v 1 was also shown to strongly activate epithelial cells, resulting in the secretion of IL-6 and the chemokines CCL2 and CCL20. The activated epithelial cells furthermore produced prostaglandin E2 (PGE2) which was shown to modulate DC responses towards the fusion protein. Finally, B cells stimulated with the fusion protein were shown to form a CD19⁺CD1d⁺CD24⁺CD38⁺IgM⁺ subpopulation with regulatory properties that secreted anti-inflammatory IL-10, produced allergen-binding IgG_1&3 antibodies and suppressed allergen-specific Th2 responses. For more information see text. PGE2, Prostaglandin E2.

Figure 7

Immunological effects of CpG ODNs as adjuvants in mice and men. CpG ODNs can activate DCs and macrophages via TLR9, resulting in increased uptake of co-applied allergens and the differentiation of both allergen-specific Th1 and T_reg cells. In mice the mixture of Bet v 1 and CpG ODN was shown to predominantly induce IFN-γ secreting Th1 cells and the production of Bet v 1-specific IgG_2a antibodies. Furthermore, CpG ODNs can thymus-independently activate either B-1 or marginal zone B cells, promoting the secretion of antibodies and pro-inflammatory cytokines. Likely because of this strong capacity of CpG ODNs to activate multiple immune cells they were also reported to increase serum levels of TNF-α that were associated with toxic shock in mice. For more information see text. IDO, indoleamine 2,3-dioxygenase.