Hev b 6.01 is a major allergen of natural rubber latex with sensitization of 70–86% of latex glove-allergic subjects. Recently, we mapped the immunodominant T cell sites of Hev b 6.01 to the highly IgE-reactive hevein (Hev b 6.02) domain. Hev b 6.01 contains 14 cysteine residues with multiple disulphide bridges stabilizing tertiary conformation. With the goal of a standardized specific immunotherapy we developed hypoallergenic Hev b 6.01 mutants by site-directed mutagenesis of selected cysteine residues (3, 12, 17, and 41) within the Hev b 6.02 domain. Peptides corresponding to the Hev b 6.02 domain of two of the mutants were also synthesized. These mutants and peptide variants showed markedly decreased or ablated latex-allergic patient serum IgE binding by immunoblotting and ELISA. Basophil activation testing confirmed markedly decreased activation with successive cysteine substitutions of the mutants and complete abrogation with the Hev b 6.02 (Cys 3, 12, 17, 41 Ala) peptide. Retention of T cell reactivity is crucial for effective specific immunotherapy and all mutants and peptide variants maintained their latex-specific T cell reactivity. The ablated allergenicity but retained T cell reactivity of the Hev b 6.02 (Cys 3, 12, 17, 41 Ala) peptide suggests this peptide is a suitable candidate for inclusion in a latex immunotherapy preparation.

Background: Bermuda grass pollen (BGP) is an increasingly important seasonal aeroallergen in Australia and other subtropical and temperate regions. BGP shares minimal allergenic cross-reactivity with pollens of rye grass or other Pooideae grasses often used for desensitization regimens in grass pollen allergy. Current allergen immunotherapy is seldom used in asthmatic patients due to IgE-mediated side effects. Since clinically effective immunotherapy is linked with altered allergen-specific T cell response, characterisation of human T cell reactivity to Cyn d 1, the major B cell allergen of BGP, should permit the design of effective and safe immunotherapy for BGP allergy. Methods: Short-term BGP-specific CD4+ T cell lines were established from peripheral blood of 14 BGP-sensitive patients before and after conventional 50% BGP and 50% 7-grass mix subcutaneous specific allergen immunotherapy (SIT). T cell diversity of antigen specificity and function was assessed by proliferation and cytokine production to BGP, Cyn d 1 and Cyn d 1 peptides. Results: Three highly immunogenic regions of Cyn d 1 were identified in 13/14 patients pre-SIT: Cyn d 1 (109–128), (181–209) and (217–241). The SIT regimen was clinically efficacious. Following SIT, decreased proliferation to BGP, Cyn d 1 and Cyn d 1 peptides was observed with a marked decrease in the IL-5:IFN-γ ratio. Conclusions: Cyn d 1 is a major T cell allergen of BGP. Decreased Cyn d 1-specific IL-5 dominant T cell responses were observed in association with clinically effective treatment with the 50% BGP and 50% 7-grass mix. Identified dominant T cell regions of Cyn d 1 should facilitate safer vaccine development for BGP-induced asthma in addition to rhinitis.

Background: Knowledge of dominant T cell epitopes of major allergens recognized by allergic individuals is required to improve efficacy and safety of allergen immunotherapy. Rye grass pollen (RGP) is the most important source of seasonal aeroallergens in temperate climates and Lol p 1 and Lol p 5 are the two major IgE‐reactive allergens. This study aimed to characterize the T cell response to these allergens using a large panel of RGP‐sensitive individuals. Methods: Short‐term RGP‐specific T cell lines (TCL) were generated from 38 RGP‐sensitive subjects and stimulated with Lol p 1 and/or Lol p 5 allergens and synthetic 20‐mer peptides. Proliferative responses were determined by 3 H‐thymidine uptake and IL‐5 and IFN‐γ in culture supernatants analysed by ELISA. Results: Of 17 subjects tested for reactivity to both allergens 16 (94%) responded to Lol p 1 and/or Lol p 5, establishing these as major T cell‐reactive allergens. Sites of T cell reactivity were spread throughout the allergen molecules but regions of high reactivity were found. For Lol p 1 these spanned residues 19–38, 109–128, 154–173, 190–209, and for Lol p 5 37–56, 100–119, 145–164, 154–173, 190–209, 217–236 and 226–245. IL‐5 and IFN‐γ were produced by T cells cultured with proliferation‐inducing peptides. Conclusions: T cell responses to RGP major allergens have been extensively characterized, providing fundamental information for developing T cell‐targeted immunotherapy for RGP allergy.