D1.221 - Identification of IgE epitopes of major ragweed pollen allergens (Amb a 1 and Amb a 11)

Ragweed pollen is a major cause of allergic diseases in Europe. Among the 12 known ragweed pollen allergens, Amb a 1 and Amb a 11 are the major allergens sensitizing 90% and 60% patients. Allergen-specific immunotherapy (AIT) is the only disease-modifying treatment for allergy, but its widespread use is limited by variability and safety concerns of allergen extracts and long treatment regimens. Recombinant peptide-based allergen vaccines targeting B-cell epitopes of allergens represent a promising alternative. This study aimed to identify and functionally characterize the novel IgE-binding B-cell epitopes of Amb a 1 and Amb a 11.

In silico epitope prediction was performed using multiple independent algorithms assessing hydrophilicity, flexibility, surface accessibility, beta-turns, and potential epitope localization. Selected peptides (35–36 amino acids) were conjugated to keyhole limpet hemocyanin (KLH) and used to immunize BALB/c female mice. Antibody responses were evaluated by ELISA. Functional activity was assessed by competitive ELISA measuring inhibition of patient’s IgE (from sera of 20 ragweed-allergic patients) binding to whole extract or recombinant Amb a 1 using anti-peptide sera from immunized mice.

9 peptides from Amb a 1 and 7 peptides from Amb a 11 were identified as potential IgE epitopes. Competitive ELISA revealed significant inhibition (average 30%) of patients’ IgE binding for peptides A1P2, A1P5, A1P7, and A1P8 (Amb a 1), and bulg and A11P1 (Amb a 11) (p ≤ 0.05) (figure 1.A.). Peptides A1P7 and A1P8 overlapped with previously described epitopes, whereas peptides A1P5 and A11P1 were firstly identified as IgE-epitopes. Additionally, several Amb a 1.02-derived peptides induced IgG-antibodies, which were capable to neutralize IgE binding to another isoform Amb a 1.01 (figure 1.B.). This fact indicated conservative structure of identified epitopes.

The peptides that have been identified as IgE-epitopes exhibit high immunogenicity, the ability to block IgE, and cross‑reactivity supporting their potential as components of a recombinant peptide‑based AIT vaccine for ragweed pollen allergy.