Detection of IgE to recombinant Hymenoptera venom allergens has been suggested to improve the diagnostic precision in Hymenoptera venom allergy. However, the frequency of sensitization to the only available recombinant honeybee venom (HBV) allergen, rApi m 1, in patients with HBV allergy is limited, suggesting that additional HBV allergens might be of relevance. We performed an analysis of sensitization profiles of patients with HBV allergy to a panel of HBV allergens. Diagnosis of HBV allergy (n = 144) was based on history, skin test results, and allergen-specific IgE levels to HBV. IgE reactivity to 6 HBV allergens devoid of cross-reactive carbohydrate determinants (CCD) was analyzed by ImmunoCAP. IgE reactivity to rApi m 1, rApi m 2, rApi m 3, nApi m 4, rApi m 5, and rApi m 10 was detected in 72.2%, 47.9%, 50.0%, 22.9%, 58.3%, and 61.8% of the patients with HBV allergy, respectively. Positive results to at least 1 HBV allergen were detected in 94.4%. IgE reactivity to Api m 3, Api m 10, or both was detected in 68.0% and represented the only HBV allergen-specific IgE in 5% of the patients. Limited inhibition of IgE binding by therapeutic HBV and limited induction of Api m 3- and Api m 10-specific IgG4 in patients obtaining immunotherapy supports recent reports on the underrepresentation of these allergens in therapeutic HBV preparations. Analysis of a panel of CCD-free HBV allergens improved diagnostic sensitivity compared with use of rApi m 1 alone, identified additional major allergens, and revealed sensitizations to allergens that have been reported to be absent or underrepresented in therapeutic HBV preparations.

Anti-IgE therapeutics interfere with the ability of IgE to bind to its receptors on effector cells. Here we report the crystal structure of an anti-IgE single-domain antibody in complex with an IgE Fc fragment, revealing how the antibody inhibits interactions between IgE and the two receptors FcεRI and CD23. The epitope overlaps only slightly with the FcεRI-binding site but significantly with the CD23-binding site. Solution scattering studies of the IgE Fc reveal that antibody binding induces a half-bent conformation in between the well-known bent and extended IgE Fc conformations. The antibody acts as functional homolog of CD23 and induces a closed conformation of IgE Fc incompatible with FcεRI binding. Notably the antibody displaces IgE from both CD23 and FcεRI, and abrogates allergen-mediated basophil activation and facilitated allergen binding. The inhibitory mechanism might facilitate strategies for the future development of anti-IgE therapeutics for treatment of allergic diseases.

Insect stings can cause life-threatening IgE-mediated anaphylactic reactions in venom-allergic patients. Although several compounds have already been described as venom allergens, prominent allergen candidates especially in the higher m.w. range have still remained elusive. Tandem mass spectrometry-based sequencing assigned a candidate gene to the most prominent putative high m.w. allergen Api m 5 (allergen C) in honeybee (Apis mellifera) venom and also allowed identification of its homologue Ves v 3 in yellow jacket (Vespula vulgaris) venom. Both proteins exhibit a pronounced sequence identity to human dipeptidyl peptidase IV or CD26. Reactivity of a human IgE mAb verified the presence of these proteins in the venoms. Both proteins were produced in insect cells and characterized for their enzymatic activity as well as their allergenic potential using sera and basophils from insect venom-allergic patients. Both Api m 5 and Ves v 3 were recognized by specific IgE of the majority of patients even in the absence of cross-reactive carbohydrate determinants. Serologic IgE reactivity closely matched activation of human basophils by Api m 5 or Ves v 3, thus underlining their relevance in functional assays. With Api m 5 and Ves v 3, a new pair of homologous allergens becomes available for future clinical applications in diagnosis and therapy that may also contribute to the understanding of molecular mechanisms of insect venoms. Moreover, the patient IgE reactivity together with the cellular activation demonstrates for the first time the relevance of high m.w. allergens in the context of hymenoptera venom allergy.