A novel therapeutic strategy for allergic diseases based on epitope-paratope blocking

Food allergies are a growing food safety and public health concern that affect up to 8% of children globally, including in Slovenia. That's 1 in 13 children or about 2 students per classroom. Currently, there is no approved drug for treating food allergy and anaphylaxis, nor is there any therapeutic approach to prevent food sensitization, highlighting the critical need for the development of effective primary prevention strategies for food allergy. Strict avoidance of the food allergen is currently the only way to prevent a reaction. Clinical and experimental analyses indicate a pathognomonic role for antigen-induced cross-linking of mast cell-associated lgE leading to rapid mast cell degranulation and release of preformed mediators and enzymes that drive the clinical manifestations associated with food-triggered anaphylaxis. We recently performed biopanning of phage display libraries with anti-peanut lgE's followed by screening and IgE binding experiments. We selected and identified three unique lgE-epitope-like peptides (DHPRFNRDNDVA, DHPRYGP, and DHPRFST) of major peanut allergen Ara h 2. Alignment of these lgE-epitope-like peptides showed overlapping with primary Ara h 2 sequence, indicating that these regions could represent the epitopes of Ara h 2. Immunoblot analyses revealed that the lgE-epitope-like peptides bind to lgE from peanut-allergic individuals. Furthermore, employing Basophil activation assay (BAT), we show that lgE-epitope-like peptides blocked Ara h 2-induced human basophil activation (surface CD63 expression). Given these striking datasets, we hypothesize that synthetic lgE epitope-like peptides can suppress lgE-dependent mast cell activation and suppress lgE-mediated food allergy and anaphylaxis. We plan to test our central hypothesis and accomplish the objectives of this application by pursuing the following specific aims: Aim #1. To assess whether our synthetic lgE-epitope-like peptides can compete and block the binding of Ara h 2 peanut allergen to IgEs paratopes in quantitative ImmunoCAP inhibition assays. Aim #2. To assess our synthetic lgE-epitope-like peptides efficacy in suppression of lgE-mast cell activation (by epitope-paratope blocking) and induction of peanut-induced anaphylaxis in humanized mast cellular model systems. Aim #3. (1) To develop, design, synthesize, and screen a new generation peanut-specific synthetic lgE-epitope-like peptides to optimize for potency, specificity, and drug-like properties (lead optimization) by a bioinformatic approach and using peptide microarray immunoassay; (2) Evaluate the peptide profile of individual peanut-allergic patients and select the most desirable peptide candidates; and (3) demonstrate that these personalized selections of peptides more efficiently suppress lgE-mediated reactions in humanized mast cellular model systems. Validation: For validation of this early-phase proof-of-principle project and applicable protocol for identifying various epitope (peptide) paratope (IgE) interaction, we will select another completely different allergen source, namely major wasp venom allergen Ves v 5. Thus for wasp venom allergen Ves v 5 we will perform biopanning, screening, IgE binding, selection, and mapping of peptides (on a limited scale). We will then generate wasp venom-specific synthetic lgE epitope-like peptides and try to confirm, that also for other allergen sources, peptides can bind to allergen-specific IgE paratopes and suppress effector cell activation.