Lise Lund

Allergy Vaccine Engineering: Epitope Modulation of Recombinant Bet v 1 Reduces IgE Binding but Retains Protein Folding Pattern for Induction of Protective Blocking-Antibody Responses

Human type 1 immediate allergic response symptoms are caused by mediator release from basophils and mast cells. This event is triggered by allergens aggregating preformed IgE Abs bound to the high-affinity receptor (FcεRI) on these cells. Thus, the allergen/IgE interaction is crucial for the cascade leading to the allergic and anaphylactic response. Two genetically engineered forms of the white birch pollen major allergen Bet v 1 with point mutations directed at molecular surfaces have been characterized. Four and nine point mutations led to a significant reduction of the binding to human serum IgE, suggesting a mutation-induced distortion of IgE-binding B cell epitopes. In addition, the mutated allergens showed a decrease in anaphylactic potential, because histamine release from human basophils was significantly reduced. Retained α-carbon backbone folding pattern of the mutated allergens was indicated by x-ray diffraction analysis and circular dichroism spectroscopy. The rBet v 1 mutants were able to induce proliferation of T cell lines derived from birch pollen allergic patients. The stimulation indices were similar to the indices of nonmutated rBet v 1 and natural Bet v 1 purified from birch pollen. The ability of anti-rBet v 1 mutant specific mouse IgG serum to block binding of human serum IgE to rBet v 1 demonstrates that the engineered rBet v 1 mutants are able to induce Abs reactive with nonmodified Bet v 1. rBet v 1 mutants may constitute vaccine candidates with improved efficacy/safety profiles for safer allergy vaccination.

Allergenicity, immunogenicity and dose-relationship of three intact allergen vaccines and four allergoid vaccines for subcutaneous grass pollen immunotherapy

Different vaccines containing intact allergens or chemically modified allergoids as active ingredients are commercially available for specific immunotherapy. Allergoids are claimed to have decreased allergenicity without loss of immunogenicity and this is stated to allow administration of high allergoid doses. We compared the allergenicity and immunogenicity of four commercially available chemically modified grass pollen allergoid products with three commercially available intact grass pollen allergen vaccines. The allergenicity was investigated with immunoglobulin (Ig)E‐inhibition and basophil activation assays. Human T cell proliferation and specific IgG‐titres following mouse immunizations were used to address immunogenicity. Furthermore, intact allergen vaccines with different contents of active ingredients were selected to study the influence of the allergen dose. In general, a lower allergenicity for allergen vaccines was clearly linked to a reduced immunogenicity. Compared with the vaccine with the highest amount of intact allergen, the allergoids caused reduced basophil activation as well as diminished immunogenicity demonstrated by reduced T cell activation and/or reduced induction of murine grass‐specific IgG antibodies. Interestingly, intact allergen vaccines with lower content of active ingredient exhibited similarly reduced allergenicity, while immunogenicity was still higher or equal to that of allergoids. The low allergenicity observed for some allergoids was inherently linked to a significantly lower immunogenic response questioning the rationale behind the chemical modification into allergoids. In addition, the linkage between allergenicity, immunogenicity and dose found for intact allergen vaccines and the immunogen as well as allergenic immune responses observed for allergoids suggest that the modified allergen vaccines do not contain high doses of immunologically active ingredients.

Comparison of allergenicity and immunogenicity of an intact allergen vaccine and commercially available allergoid products for birch pollen immunotherapy

Background Specific immunotherapy with intact allergen vaccine is a well‐documented treatment for allergic diseases. Different vaccine formulations are currently commercially available, the active ingredient either being intact allergens or chemically modified allergoids. The rationale behind allergoids is to decrease allergenicity while maintaining immunogenicity. However, data from the German health authorities based on reporting of adverse events over a 10‐year period did not indicate increased safety of allergoids over intact allergens.

Adjuvant effects of aluminium hydroxide‐adsorbed allergens and allergoids – differences in vivo and in vitro

Allergen‐specific immunotherapy (SIT) is a clinically effective therapy for immunoglobulin (Ig)E‐mediated allergic diseases. To reduce the risk of IgE‐mediated side effects, chemically modified allergoids have been introduced. Furthermore, adsorbance of allergens to aluminium hydroxide (alum) is widely used to enhance the immune response. The mechanisms behind the adjuvant effect of alum are still not completely understood. In the present study we analysed the effects of alum‐adsorbed allergens and allergoids on their immunogenicity in vitro and in vivo and their ability to activate basophils of allergic donors. Human monocyte derived dendritic cells (DC) were incubated with native Phleum pratense or Betula verrucosa allergen extract or formaldehyde‐ or glutaraldehyde‐modified allergoids, adsorbed or unadsorbed to alum. After maturation, DC were co‐cultivated with autologous CD4+ T cells. Allergenicity was tested by leukotriene and histamine release of human basophils. Finally, in‐vivo immunogenicity was analysed by IgG production of immunized mice. T cell proliferation as well as interleukin (IL)‐4, IL‐13, IL‐10 and interferon (IFN)‐γ production were strongly decreased using glutaraldehyde‐modified allergoids, but did not differ between alum‐adsorbed allergens or allergoids and the corresponding unadsorbed preparations. Glutaraldehyde modification also led to a decreased leukotriene and histamine release compared to native allergens, being further decreased by adsorption to alum. In vivo, immunogenicity was reduced for allergoids which could be partly restored by adsorption to alum. Our results suggest that adsorption of native allergens or modified allergoids to alum had no consistent adjuvant effect but led to a reduced allergenicity in vitro, while we observed an adjuvant effect regarding IgG production in vivo.

An alternative allergen:adjuvant formulation potentiates the immunogenicity and reduces allergenicity of a novel subcutaneous immunotherapy product for treatment of grass‐pollen allergy

Subcutaneous specific immunotherapy (SCIT) has proven sustained clinical efficacy against allergy. The recommended regimen for SCIT is a gradual updosing over a period of weeks. Commonly, in commercial products for SCIT, the specific allergen is formulated with an adjuvant, most often in the form of aluminium hydroxide (AlOH). It has been shown that allergen‐specific IgG antibodies are induced as a result of successful SIT.