Recombinant T‐cell epitope conjugation: A new approach for Dermatophagoides hypoallergen design

Background Allergen‐specific immunotherapy (AIT) is the only clinical approach that can potentially cure some allergic diseases by inducing immunological tolerance. Dermatophagoides pteronyssinus is considered as the most important source of mite allergens worldwide, with high sensitization rates for the major allergens Der p 1, Der p 2 and Der p 23. The aim of this work is to generate a hypoallergenic hybrid molecule containing T‐cell epitopes from these three major allergens. Methods The hybrid protein termed Der p 2231 containing T‐cell epitopes was purified by affinity chromatography. The human IgE reactivity was verified by comparing those with the parental allergens. The hybrid was also characterized immunologically through an in vivo mice model. Results The hybrid rDer p 2231 stimulated in peripheral blood mononuclear cells (PBMCs) isolated from allergic patients with higher levels of IL‐ 2, IL‐10, IL‐15 and IFN‐γ, as well as lower levels of IL‐4, IL‐5, IL‐13, TNF‐α and GM‐CSF. The use of hybrid molecules as a therapeutic model in D. pteronyssinus allergic mice led to the reduction of IgE production and lower eosinophilic peroxidase activity in the airways. We found increased levels of IgG antibodies that blocked the IgE binding to the parental allergens in the serum of allergic patients. Furthermore, the stimulation of splenocytes from mice treated with rDer p 2231 induced higher levels of IL‐10 and IFN‐γ and decreased the secretion of IL‐4 and IL‐5, when compared with parental allergens and D. pteronyssinus extract. Conclusions rDer p 2231 has the potential to be used in AIT in patients co‐sensitized with D. pteronyssinus major allergens, once it was able to reduce IgE production, inducing allergen‐specific blocking antibodies, restoring and balancing Th1/Th2 immune responses, and inducing regulatory T‐cells. The hybrid rDer p 2231 stimulated in PBMCs isolated from atopic patients, higher levels of IL‐ 2, IL‐10, IL‐15 and IFN‐γ, as well as lower levels of IL‐4, IL‐5, IL‐13, TNF‐α and GM‐CSF. The use of hybrid molecules as a therapeutic model in D. pteronyssinus allergic mice led to the reduction of IgE production and lower eosinophilic peroxidase activity in the airways. We found increased levels of IgG antibodies, which blocked the IgE binding to the parental allergens in serum of atopic patients. Furthermore, the stimulation of splenocytes from mice treated with rDer p 2231 induced higher levels of IL‐10 and IFN‐γ and decreased the secretion of