Characterization of the Composition and Biological Activity of the Venom from Vespa bicolor Fabricius, a Wasp from South China

We analyzed, for the first time, the major components and biological properties of the venom of Vespa bicolor, a wasp from South China. Using HPLC and SDS-PAGE, combined with LC–MS/MS, MALDI-TOF-MS, and NMR data to analyze V. bicolor venom (VBV), we found that VBV contains three proteins (hyaluronidase A, phospholipase A1 (two isoforms), and antigen 5 protein) with allergenic activity, two unreported proteins (proteins 5 and 6), and two active substances with large quantities (mastoparan-like peptide 12a (Vb-MLP 12a), and 5-hydroxytryptamine (5-HT)). In addition, the antimicrobial activity of VBV was determined, and results showed that it had a significant effect against anaerobic bacteria. The minimum inhibitory concentration and minimum bactericidal concentration for Propionibacterium acnes were 12.5 µg/mL. Unsurprisingly, VBV had strong antioxidant activity because of the abundance of 5-HT. Contrary to other Vespa venom, VBV showed significant anti-inflammatory activity, even at low concentrations (1 µg/mL), and we found that Vb-MLP 12a showed pro-inflammatory activity by promoting the proliferation of RAW 264.7 cells. Cytotoxicity studies showed that VBV had similar antiproliferative effects against all tested tumor cell lines (HepG2, Hela, MCF-7, A549, and SASJ-1), with HepG2 being the most susceptible. Overall, this study on VBV has high clinical importance and promotes the development of Vespa bicolor resources.

HOUSE DUST MITE ALLERGENS: FROM NATIVE EXTRACTS TO MODIFIED PREPARATIONS

The growth of allergic diseases dictates the need to develop new forms of therapeutic allergens with high immunogenic and low allergenic activity. For many years, our laboratory has been developing drugs for the diagnosis and treatment of house dust mites (HDM) allergies. The purpose of this study is to summarize the results of the five-year development of therapeutic preparations of HDM allergens. During this period, we obtained the following forms of therapeutic allergens: a granular sublingual dosage form of a mixed allergen from Dermatophagoides pteronyssinus (Der.p) and Dermatophagoides farinaе (Der.f) and a succinylated monomeric HDM allergoid Der.p. The physicochemical and immunobiological properties of the obtained preparations were studied by methods: electrophoresis in PAGE in the presence of SDS-sodium, micropoint immunoblot, ELISA, inhibition of the binding reaction of allergen-specific IgE in the sera of patients. The research results showed that the obtained preparations have a reduced allergenic and increased immunogenic activity in comparison with native extracts. The created forms of mite allergens can be further used to treat patients sensitized to HDM of the genus Dermatophagoides.

IgE Epitopes of the House Dust Mite Allergen Der p 7 Are Mainly Discontinuous and Conformational

BackgroundSeveral studies indicate that Der p 7 is an important and clinically relevant allergen of Dermatophagoides pteronyssinus which should be included in vaccines for treatment of house dust mite (HDM) allergy. Aim of this study was to characterize the IgE epitopes of Der p 7.MethodsRecombinant Der p 7 was expressed and purified, analyzed for fold by circular dichroism and tested for its allergenic activity by basophil activation. Seven overlapping, surface-exposed peptides (P1–P7) with a length of 27 to 37 amino acids, which spanned the Der p 7 sequence, were synthesized and tested for IgE reactivity and allergenic activity by basophil activation assay. Carrier-bound peptides were studied for their ability to induce allergen-specific IgG antibodies in rabbits. Peptide-specific antibodies were used to inhibit allergic patients` IgE binding to Der p 7 by ELISA for mapping of IgE epitopes.ResultsrDer p 7 showed high allergenic activity comparable with Der p 5, Der p 21, and Der p 23. None of the seven tested peptides showed any IgE reactivity or allergenic activity when tested with HDM- allergic patients indicating lack of sequential IgE epitopes on Der p 7. IgE inhibition experiments using anti-peptide specific IgGs and molecular modeling enabled us to identify discontinuous, conformational IgE epitopes of Der p 7.Conclusion and Clinical RelevanceIgE epitopes of Der p 7 belong to the conformational and discontinuous type whereas sequential Der p 7 peptides lack IgE reactivity. It should thus be possible to construct hypoallergenic vaccines for Der p 7 based on carrier-bound allergen peptides.

Allergological Importance of Invertebrate Glutathione Transferases in Tropical Environments

Glutathione-S transferases (GSTs) are part of a ubiquitous family of dimeric proteins that participate in detoxification reactions. It has been demonstrated that various GSTs induce allergic reactions in humans: those originating from house dust mites (HDM), cockroaches, and helminths being the best characterized. Evaluation of their allergenic activity suggests that they have a clinical impact. GST allergens belong to different classes: mu (Blo t 8, Der p 8, Der f 8, and Tyr p 8), sigma (Bla g 5 and Asc s 13), or delta (Per a 5). Also, IgE-binding molecules belonging to the pi-class have been discovered in helminths, but they are not officially recognized as allergens. In this review, we describe some aspects of the biology of GST, analyze their allergenic activity, and explore the structural aspects and clinical impact of their cross-reactivity.

Regulatory Requirements for the Quality of Allergen Products for Allergen Immunotherapy of Food Allergy

Purpose of Review Medicinal products for allergen immunotherapy (AIT) of food allergies have gained enormous momentum in recent years. With this new class of products entering marketing authorization procedures, compliance to regulatory requirements becomes a critical element. Here, an overview is provided on specific requirements and aspects concerning the quality control and manufacturing of these products. Recent Findings Recent developments in the field of AIT for food allergies are divers, including products for oral, epicutaneous, and subcutaneous application, most notably targeting egg, milk, and peanut allergy. As the source materials for food AIT product are typically produced for food consumption and not for medicinal purposes, unique challenges arise in the manufacturing processes and controls of these medicinal products. Individual approaches are needed to assure acceptable quality, including control of relevant quantitative and qualitative characteristics. Major characteristics for quality verification include determination of protein content, total allergenic activity, and major allergen content. The applied manufacturing processes need to be established such that relevant process parameters are kept within justified limits and consistency of produced batches is assured. Summary Allergen products for food AIT present specific challenges with respect to quality aspects that differentiate them from other commonly available AIT products. While established regulation is available and provides clear guidance for most aspects, other issues require consideration of new and individual settings relevant here. Consequently, as experience grows, respective amendments to currently available guidance may be needed.

DNA vaccines and recombinant allergens with reduced allergenic activity treat allergies

This review is intended to familiarize readers with major novel directions of developing allergy vaccines, their structure, as well as the mechanisms of forming a new immunological response in the course of the treating immunoglobulin E (IgE)-mediated allergic diseases. Currently, science offers a huge variety of new experimental forms of recombinant allergens with reduced allergenic activity and increased immunogenicity, or vice-versa, immune tolerance. Often, the mechanisms of their effect on the immune system are not fully understood. Scientific publications, including reviews covering this topic, allowed us identifying top priority areas in the development of allergy vaccines: recombinant hypoallergenic allergen derivatives, T cell epitope-based allergy vaccines, and B cell epitope-based allergy vaccines. In addition, the review discusses use of deoxyribonucleic acid (DNA) vaccines. Immunotherapy with DNA vaccines is the newest and least studied method of treating allergic diseases.

Allergy vaccines for specific immunotherapy

Allergen-specific immunotherapy (ASIT) has been used for more than a hundred years to treat patients with IgEmediated allergic diseases. The most common allergens have been obtained using molecular cloning technology in the past two decades. To increase the safety of immunotherapy, a large group of genetically modified allergens with reduced allergenic activity was created. The mechanism of action of these therapeutic allergens differs from natural allergen extracts, and more research is needed to understand how desensitization occurs in each case. The objective of this review is to introduce readers to new therapeutic allergy vaccines and their structural modification features as well as immunological effects on the body. To achieve this objective, we have analyzed and systematized the experimental developments presented in the literature on the main directions of creating new allergy vaccines: hypoallergenic derivatives of recombinant allergens, T cell epitope-based allergy vaccines and B cell epitope-based allergy vaccines, DNA vaccines.Summing up the results of the research presented in the literature, it is necessary to note the high heterogeneity of designs used to achieve the high efficiency of the developed therapeutic allergens. All allergy vaccines presented in the review solve the tasks set by the researchers: in experimental animal models they induce immunogenicity or tolerance, in clinical trials they reduce the symptoms of allergic reactions. The the effectiveness of the proposed medicinal products is quite high but its evaluation requires further long-term preclinical and clinical trials to confirm the safety and harmlessness of the created allergy vaccines.

ALLERGENIC ACTIVITY AND DANGER OF INDUSTRIAL DUSTS OF DRY PRODUCTS OF COW'S MILK PROCESSING

The purpose of the work was to establish in a model experiment the allergenic activity and danger of the extracts obtained from the dust of dry products of cow's milk processing (DPMP), containing complexes of soluble whey (WMP) or casein milk proteins (CMP), as a stage of hygienic regulation of the content of dust DPMP in the air of the working area. Experiments on albino guinea pigs sensitized by the intradermal injection of standard doses of WMP and СМР solutions into the ear revealed the development of severe allergic reactions in the animals of the experimental groups with the prevalence of mixed mechanisms of immediate anaphylactic and delayed cell-mediated types. According to the criteria for the classification of industrial allergens, the WMP and СМР complexes have a strong allergenic activity and are differentiated to the 1-st class of allergenic hazard, which determines the classification of the DPMP dust containing them as extremely dangerous industrial allergens. This is confirmed by the established high levels of indicators of allergic-diagnostic reactions in vivo and in vitro when testing sensitized WMP and СМР animals with a solution of skim milk powder dust, indicating the presence of antigenic determinants of whey and casein milk proteins in it and a real ability to form cross-allergic reactions in the body of workers to dust from all dry milk processing products containing these proteins.

Identification and Physicochemical Characterization of a New Allergen from Ascaris lumbricoides

To analyze the impact of Ascaris lumbricoides infection on the pathogenesis and diagnosis of allergic diseases, new allergens should be identified. We report the identification of a new Ascaris lumbricoides allergen, Asc l 5. The aim of this study was to evaluate the physicochemical and immunological features of the Asc l 5 allergen. We constructed an A. lumbricoides cDNA library and Asc l 5 was identified by immunoscreening. After purification, rAsc l 5 was physicochemically characterized. Evaluation of its allergenic activity included determination of Immunoglobulin E (IgE) binding frequency (in two populations: 254 children and 298 all-age subjects), CD203c based-basophil activation tests (BAT) and a passive cutaneous anaphylaxis (PCA) mouse model. We found by amino acid sequence analysis that Asc l 5 belongs to the SXP/RAL-2 protein family of nematodes. rAsc l 5 is a monomeric protein with an alpha-helical folding. IgE sensitization to rAsc l 5 was around 52% in general population; positive BAT rate was 60%. rAsc l 5 induced specific IgE production in mice and a positive PCA reaction. These results show that Asc l 5 has structural and immunological characteristics to be considered as a new allergen from A. lumbricoides.