Immunotherapy Of House Dust Mite Allergy Research Articles

Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy

AimTo investigate the immunomodulatory potential of a chimera composed of the receptor-binding domain of hemagglutinin 1 (H1s) from Influenza virus and Der p 2 (D2) allergen for allergen-specific immunotherapy of house-dust mite allergy (HDM). Main methodsH1sD2 chimera and D2 allergen were produced by genetic engineering in E. coli. Recombinant antigens were extracted from inclusion bodies by urea, then refolded and purified by immobilized- metal affinity chromatography (IMAC). Purity was verified by 2D-PAGE and secondary structures were assessed by CD spectroscopy. IgE reactivity of H1sD2 and D2 was tested in western blot with sera from 8 persons with clinical history of HDM allergy. Immunogenicity of H1sD2 and D2 were analyzed in Balb/c mice. Cytokine profile was analyzed by ELISA after stimulation of mouse spleen cells with H1sD2 and D2. Leukocyte population abundance of cells isolated from spleen and lymph node was assessed by flow cytometry. Key findingsPurified recombinant proteins H1sD2 (42 kDa) and D2 (15 kDa) revealed well defined secondary structures, and preserved IgE reactive epitopes. Analysis of supernatants of mouse spleen cells after stimulation with H1sD2 and D2, revealed a qualitatively different cytokine profile from H1sD2 immunized mouse cells (increase in IL10). CD8+ cells were decreased in the lymph node of D2 immunized mice, whereas H1sD2 immunization led to an increase of CD8+ cells in both the lymph node and the spleen. SignificanceH1sD2 chimera attenuates Der p 2-inherent Th2 response and directs the immune response toward Th1 and Treg phenotype.

Immunotherapy for house-dust mite allergy.

Since the discovery of house-dust mites (HDM) in the 1960's, allergy immunotherapy trials that used extracts of these mites have been conducted, first by subcutaneous (SCIT) and later by the sublingual (SLIT) route. When reviewed in 2013, published studies of HDM immunotherapy were found to often be characterized by small sample size, widely varying doses, and poorly defined disease severity and outcomes. These trials were thought to to support the efficacy of HDM subcutaneous allergy immunotherapy but the evidence for efficacy of sublingual immunotherapy was less firm. This report will review a large number of well-designed studies reported since 2013, mostly of SLIT, and in particular, of two newly developed HDM sublingual tablets. In addition, other aspects of HDM immunotherapy will be addressed, including use in atopic dermatitis, optimum duration of treatment, evidence for disease modification and use with adjuvants. Seventeen reports on 15 randomized, double-blind, placebo-controlled trials were identified as having been published since the cut-off date of the 2013 systematic review. Twelve of these reported results with the 2 HDM SLIT-tablets. These studies clearly established the appropriate doses and the efficacy and safety of these tablets in treating allergic rhinitis and asthma. Other reports offered support for use of HDM immunotherapy in selected patients with atopic dermatitis, for administration of HDM immunotherapy for 3 to 5 years, for anticipating disease modification after 3-5 years of treatment, and for the use of vitamin D and selected probiotics to enhance its efficacy. HDM SCIT and SLIT-tablet therapy have demonstrated effectiveness in allergic rhinitis and asthma. Appropriate dosing with HDM SLIT-liquid has not been established although a limited number of studies suggest it can be effective as well. HDM SCIT and HDM SLIT share efficacy in allergic rhinitis and asthma, disease modification and the duration of treatment required to produce persisting benefit.

Similar localization of conformational IgE epitopes on the house dust mite allergens Der p 5 and Der p 21 despite limited IgE cross-reactivity.

BackgroundDue to high IgE recognition frequency and high allergenic activity, Der p 5 and Der p 21 are clinically important house dust mite (HDM) allergens. The objective of this study was to characterize the immunodominant IgE epitopes of Der p 5 and Der p 21 responsible for their high allergenic activity.MethodsA panel of 12 overlapping peptides spanning the Der p 5 and Der p 21 sequence were synthesized to search for sequential IgE epitopes by direct testing for allergic patients' IgE reactivity. Peptide‐specific antibodies raised in rabbits were used in inhibition studies for localizing conformational IgE epitopes which were visualized on the surfaces of the allergen structures by molecular modelling. IgE cross‐reactivity between the allergens was investigated by IgE inhibition studies.ResultsImmunodominant IgE epitopes defined by allergic patients' IgE on Der p 5 and Der p 21 were primarily of the conformational, discontinuous type including N‐ and C‐terminal portions of the protein. They could be located on each allergen on one area with similar localization, but despite similar structure of the allergens, no relevant IgE cross‐reactivity could be detected.ConclusionOur study shows that Der p 5 and Der p 21 contain a major conformational IgE epitope‐containing area located on similar portions of their structure, but they lack relevant IgE cross‐reactivity. These data are important for the development of modern allergy vaccines based on defined molecules for allergen‐specific immunotherapy of HDM allergy.

Impact of Dermatophagoides pteronyssinus mite body raw material on house dust mite allergy diagnosis in a Serbian population

House dust mite (HDM) allergy has different clinical and immunological patterns in different geographic regions. The impact of raw material of commercial Dermatophadoides pteronyssinus (Acari: Pyroglyphidae) mite bodies on the quality of allergen extracts for allergy diagnosis in the Serbian population has not been previously evaluated. House dust mite bodies obtained from manufacturers in Europe, South America and Australia were used in the preparation of allergen extracts for in vivo diagnosis and serological analysis in a group of 14 HDM-allergic adults. In the group of mite-allergic patients, there was no statistically significant difference in skin test reactivity (Wilcoxon matched pairs test) among the three HDM body extract preparations. In a CAP inhibition assay, two extracts (A and C) achieved maximum inhibition of >90%, whereas extract B demonstrated a different inhibition slope and lower inhibition potential (80%). However, a remarkable difference in immunoglobulin E reactivity using Western blot analysis with individual patients' sera was observed in one of the preparations (extract B). These findings emphasize the need for the careful selection of starting material for the preparation of HDM diagnostic reagents intended for use in patients from geographically distinct regions as these preparations can have implications on the selection criteria for patient-tailored immunotherapy of HDM allergy.

Reduction of the in vivo allergenicity of Der p 2, the major house-dust mite allergen, by genetic engineering

The major allergen of the house-dust mite Dermatophagoides pteronyssinus, Der p 2, is recognized by approximately 90% of mite-allergic patients. We have produced two recombinant fragments of Der p 2 comprising aa 1–53 and aa 54–129 and a hybrid molecule (aa 54–129 + 1–53), combining the two fragments in inverse order, by genetic engineering. The recombinant Der p 2 derivatives were expressed in E. coli and purified to homogeneity. rDer p 2 derivatives (fragments and hybrid) showed a considerably reduced beta sheet structure and IgE reactivity compared to the Der p 2 wild-type allergen. The allergenic activity of the Der p 2 derivatives was reduced more than tenfold as evaluated in vitro in basophil activation assays and in vivo by skin prick testing of mite-allergic patients. Immunization of mice and rabbits with rDer p 2 derivatives induced Der p 2-specific IgG antibodies, which inhibited the binding of allergic patients’ IgE to Der p 2. Immunization of mice with rDer p 2 derivatives induced less allergenic IgE responses than immunization with rDer p 2. Thus the rDer p 2 derivatives exhibited less in vivo allergenic activity and allergenicity than the Der p 2 allergen but preserved immunogenicity and may hence represent candidates for specific immunotherapy of house-dust mite allergy.

Effects of oral hyposensitization with recombinant Der f2 on immediate airway constriction in a murine allergic model.

Recombinant Der f2 (rDer f2) has recently been developed as a promising allergen for the diagnosis and immunotherapy of house-dust mite allergy, and studies in immunology. The aim of the present study was to evaluate whether oral administration of rDer f2 could suppress an immediate allergic reaction in mice sensitized with mite allergen. We developed a murine allergic model that showed bronchoconstriction after inhalation of rDer f2, and studied the effect of oral administration of rDer f2 on the reaction. Seven week old male A/J mice were intranasally immunized with rDer f2 12 times. Sensitized mice showed anti-rDer f2 immunoglobulin (Ig)E production and immediate airway constriction after inhalation of 10 mg.mL(-1) of rDer f2, as determined by the Konzett-Rössler method. Immunized animals were divided into three groups, and fed phosphate-buffered saline (PBS), 0.1 mg.day(-1), or 1 mg.day(-1) of rDer f2 for 4 weeks, respectively. Seven days after the last feeding, the mice were examined for their immediate response. Animals fed with 1 mg.day(-1) rDer f2 showed significantly reduced bronchoconstriction after inhalation of both 2 mg.mL(-1) and 10 mg.mL(-1) of rDer f2 compared with PBS-fed mice. Similar results were obtained when we examined mice 10 weeks after the last feeding. Reactions in the 0.1 mg.day(-1) rDer f2-fed group also tended to decrease in comparison with PBS-fed animals. Plasma anti-rDer f2 IgE, IgG1, IgG2a, and IgG2b levels were not changed by feeding with rDer f2. We conclude that recombinant Der f2 exhibits both sensitizing and hyposensitizing activities in mice. rDer f2 may be useful in immunotherapy and diagnosis of house-dust mite allergy.

Possible induction of food allergy during mite immunotherapy

Sera of 17 patients receiving immunotherapy for house-dust mite allergy were tested for IgE antibodies against snail and shrimp. Serum samples were taken at the start of immunotherapy and 14-20 months later. While the average IgE response to mite, Der p 1, and Der p 2 did not alter significantly, the average response to snail showed a significant increase. This included two conversions from negative to strongly positive. These novel IgE antibodies against snail were shown to be cross-reactive with mite. Three patients had a positive RAST for shrimp. For one of them, a strong increase of IgE against shrimp (and snail) was observed. In 2/3 snail/shrimp-positive sera, IgE antibodies against the cross-reactive allergen tropomyosin from mite, snail, and shrimp were demonstrated. A clear IgE response to snail (> 10% binding in a snail RAST) was confirmed by a positive skin prick test (SPT) for 6/10 patients. The two patients with antitropomyosin IgE also had a positive SPT for shrimp, and demonstrated the oral allergy syndrome (OAS) after eating shrimp. The observations in this study indicate that house-dust mite immunotherapy is accompanied by the induction of IgE against foods, including tropomyosin-reactive IgE. Food allergy (OAS) was observed in patients that had IgE antibodies against this cross-reactive allergen. In conclusion, induction of IgE during mite immunotherapy might occasionally cause allergy to foods of invertebrate animal origin.