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

Abstract

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 IFNγ, as well as lower levels of IL-4, IL-5, IL-13, TNFα 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 IFNγ 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 protein BTH2 suppresses allergic airway inflammation in a murine model of HDM‐specific immunotherapy

“…Additionally, recent data from our group using other HDM recombinant proteins showed the same pattern of higher IL-10 production in PBMC from allergic donors. 50 BTH2 significantly changed allergic inflammation in mice. The decrease in eosinophils in BTH2-treated mice was a noteworthy result due to these cells' importance for effector mechanisms in allergic responses.…”

“…However, both features can occur in a noncontradictory manner. Additionally, recent data from our group using other HDM recombinant proteins showed the same pattern of higher IL‐10 production in PBMC from allergic donors 50 …”

Biological therapy practice, biomarkers of severe asthma and novel approaches for attaining immunomodulation in upper airway disease

“…Furthermore, the stimulation of splenocytes from mice treated with rDer p 2231 induced higher levels of IL-10 and IFNγ and decreased the secretion of IL-4 and IL-5, when compared to parental allergens and D. pteronyssinus extract. (7).…”

“…The final Editor's Choice article evaluates the allergenicity and immunogenicity of a hybrid molecule containing T‐cell epitopes from three major house dust mite allergens, Der p 1, Derp 2 and Der p 23, as a novel therapeutic option for allergen immunotherapy (AIT) 7 . AIT is the only clinical management of allergies that can control the progression of these immune‐mediated diseases by inducing immunological tolerance 8,9 Therapies with T‐cell‐targeted molecules can significantly impact the safety and efficacy of AIT, making them favourable alternatives to natural extracts.3 A series of clinical trials have already demonstrated that hypoallergenic T‐cell epitope‐based allergens can compose safe and effective therapeutic formulations for AIT.…”

Removal of N-Terminal Peptide Impacts Structural Aspects of an IgE-Reactive Recombinant Der p 5

“…The membrane was blocked with 5% non-fat dried milk diluted in 1× phosphate buffered saline (PBS) plus 3% Tween. Then, the membrane was incubated for 1 h with serum of mice (1:2000) previously sensitized to the extract of D. pteronyssinus, which was obtained from an experimental model performed by the group [37]. After that, incubation was performed with a secondary anti-mouse IgG antibody conjugated with alkaline phosphatase (1:2000, Invitrogen, Carlsbad, CA, USA) for 1 h. The membrane was exposed to a reaction solution of nitro blue tetrazolium (NBT, 0.14 mg/mL, Sigma, St. Louis, MI, USA), 5-bromo-4-chloro-3-indolyl-phosphate (BCIP, 0.07 mg/mL, Sigma, St. Louis, MI, USA), and alkaline phosphatase buffer.…”