IgG1, IgG4 and IgE Antibody Reactivity to Mutant Forms of the Major Dust Mite Allergen Lep d 2 among Atopic and Nonatopic Subjects Naturally Exposed to Lepidoglyphus destructor

Abstract

Background: Lepidoglyphus destructor is a common dust mite causing IgE-mediated sensitization. The major allergen, Lep d 2, has previously been cloned and expressed as several double Cys to Ser mutants with the purpose of producing hypoallergenic variants for immunotherapy. Our aim was to investigate the reactivity pattern of IgG1, IgG4 and IgE antibodies to wild-type (wt) rLep d 2 and four mutants among atopic and nonatopic subjects in relation to sensitization and exposure to L. destructor. Methods: Inhibition and sandwich ELISA were used to compare IgG1, IgG4 and IgE antibody reactivities to rLep d 2 variants in serum of 20 atopic and 18 nonatopic farmers naturally exposed to L. destructor. A group of 22 urban subjects served as controls. Results: Atopic farmers demonstrated correlating IgE and IgG4 levels to rLep d 2(wt) (r_s = 0.70; p < 0.0001) which were significantly (p < 0.0001) higher than those of nonatopic farmers and urban controls. No IgG4 antibodies were detected in nonatopic farmers despite chronic allergen exposure. A parallel reactivity pattern of IgE and IgG4 to all rLep d 2 mutants was observed. The mutant lacking all 3 disulfide bonds, rLep d 2.6Cys, demonstrated neither any IgE nor IgG4 reactivity. In contrast, IgG1 antibodies had a different reactivity pattern and were detected among most subjects irrespective of atopy, exposure to L. destructor or disulfide impairments in rLep d 2. Moreover, IgG1 levels to rLep d 2(wt) and rLep d 2.6Cys correlated (n = 60; r_s = 0.65; p < 0.0001). Conclusions: IgE/IgG4 Ab to rLep d 2 were restricted to atopic farmers and demonstrated parallel recognition patterns of conformational epitopes. In contrast, IgG1 antibodies are ubiquitously found and mainly recognize sequential structures. The observed isotypic difference and interindividual variation in antibody specificities among atopic and nonatopic subjects imply careful investigation of hypoallergenic variants destined for immunotherapy.