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A mutant of the major apple allergen, Mal d 1, demonstrating hypo-allergenicity in the target organ by double-blind placebo-controlled food challenge.
Bolhaar, ST, Zuidmeer, L, Ma, Y, Ferreira, F, Bruijnzeel-Koomen, CA, Hoffmann-Sommergruber, K, van Ree, R, Knulst, AC
Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology. 2005;(12):1638-44
Abstract
BACKGROUND Allergen-specific immunotherapy for food allergy has been hindered by severe side-effects in the past. Well-characterized hypo-allergenic recombinant food allergens potentially offer a safe solution. OBJECTIVE To demonstrate hypo-allergenicity of a mutated major food allergen from apple, Mal d 1, in vitro and in vivo. METHODS A mutant of the major apple allergen, Mal d 1, was obtained by site-directed mutagenesis exchanging five amino acid residues. Fourteen patients with combined birch pollen-related apple allergy were included in the study. Hypo-allergenicity of the mutant rMal d 1 (rMal d 1mut) compared with rMal d 1 was assessed by in vitro methods, i.e. RAST (inhibition), immunoblotting and basophil histamine release (BHR) and in vivo by skin prick test and double-blind placebo-controlled food challenge (DBPCFC). RESULTS RAST analysis (n = 14) revealed that IgE reactivity to rMal d 1mut was twofold lower than that of the wild-type molecule (95% confidence interval (CI): 1.7-2.4). RAST inhibition (n = 6) showed a 7.8-fold decrease in IgE-binding potency (95% CI: 3.0-12.6). In contrast to this moderate decrease in IgE-binding potency, the biological activity of rMal d 1mut assessed by SPT and BHR decreased 10-200-fold. Hypo-allergenicity was confirmed by DBPCFC (n = 2) with both recombinant molecules. CONCLUSION A moderate decrease in IgE-binding potency translates into a potent inhibition of biological activity. This is the first study that confirms by DBPCFC that a mutated recombinant major food allergen is clinically hypo-allergenic. This paves the way towards safer immunotherapy for the treatment of food-allergic patients.