Characterization of a hypoallergenic recombinant Bet v 1 variant as a candidate for allergen-specific immunotherapy
ABSTRACT Recombinant allergens and especially their hypoallergenic variants are promising candidates for a more effective and safer specific immunotherapy.
Physicochemical and immunological characteristics of a folding variant of recombinant Bet v 1 (rBet v 1-FV) were investigated in comparison to natural Bet v 1 (nBet v 1) and the correctly folded recombinant Bet v 1 (rBet v 1-WT) by SDS-PAGE, size exclusion chromatography, multi-angle light scattering, circular dichroism, immunoblotting and enzyme allergosorbent test inhibition assay for detection of IgE reactivity and ELISA with Bet v 1-specific monoclonal antibodies. The functional IgE reactivity of the different Bet v 1 proteins was investigated using basophil activation in terms of CD203c expression and histamine release. T cell reactivity was investigated using T cell lines raised from birch pollen-allergic subjects against nBet v 1. Immunogenicity was investigated in mice.
Physicochemical characterization revealed purity, homogeneity and monomeric properties of rBet v 1-FV. Unlike nBet v 1 and rBet v 1-WT, rBet v 1-FV showed almost no IgE binding in immunoblots. The reduction of allergenicity was further proved by IgE-binding inhibition assays, basophil activation and histamine release. T cell reactivity was completely conserved, as demonstrated by proliferation of Bet v 1-specific T cell lines with multiple epitope specificities. rBet v 1-FV showed strong immunogenicity in mice.
Due to its reduced IgE reactivity and decreased capacity to activate basophils, but retained T cell reactivity and strong immunogenicity, rBet v 1-FV proved to be a very promising candidate for specific immunotherapy in birch pollen-allergic subjects.
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ABSTRACT: This article reviews the literature on allergen modifications and novel routes of delivery for antigen-specific immunotherapy for allergic disease. Allergen modifications include the use of recombinant proteins, combining antigens with infectious carrier proteins, peptide immunotherapy, and genetic vaccines containing the code for allergenic proteins. Novel routes of delivery include oral immunotherapy, intralymphatic immunotherapy, epicutaneous immunotherapy, and oral mucosal immunotherapy. Allergen depot preparations, such as biodegradable, injectable microspheres and sublingual tablets, have also been developed. Current research in immunotherapy for allergic disease has focused on improving efficacy and patient adherence to therapy, while minimizing the risks of serious adverse events.06/2013; 1(2). DOI:10.1007/s40136-013-0009-6