We performed an investigation of the in?vitro and in?vivo effects of a purified allergen chemically conjugated to a novel 8-OH modified adenine as an adjuvant.
Purified group 2 major allergen from house dust mite chemically conjugated to 4-(6-amino-9-benzyl-8-hydroxy-9H-purin-2-ylsulfanyl)-butyric acid succinimidyl ester was analyzed by using mass spectrometry. The adduct (nDer p 2-Conj) was assayed for Toll-like receptor activation on transfected HEK293 cells, stimulation of innate cells, and effects on the functional phenotype of specific T-cell lines and clones by means of flow cytometry, real-time PCR, and expression of TH-related transcription factors. Lung cells and sera of nDer p 2-Conj-sensitized C57Bl/6 mice were studied by means of cytology, histology, real-time PCR, and ELISA.
nDer p 2-Conj stimulated IL-12 and IFN-¦Á production from monocytes and plasmacytoid dendritic cells, respectively, retaining the ability to trigger Toll-like receptor 7 exclusively, and expanded human allergen-specific lymphocytes with reduced ability to produce TH2-related cytokines and increased IFN-¦Ã levels, as based on GATA-3/T-bet expression. In?vivo adduct-sensitized mice exhibited reduced eosinophil infiltration and IL-13 expression in the airways, IFN-¦Ã upregulation together with IgE downregulation, and an increase in allergen-specific IgG2a levels in sera. The conjugate exhibited reduced ability to activate human Fc¦ÅRI+ cells without inducing TH17 cells or autoantibodies.
The codelivery of an allergen with a modified adenine as a conjugate inducing modulatory cytokines from innate cells redirects in?vitro and in?vivo pathogenic TH2 responses without eliciting harmful effects.