Hypoallergenic mutants of Ole e 1, the major olive pollen allergen, as candidates for allergy vaccines

Authors


Correspondence:
Eva Batanero, Departamento de Bioquímica y Biología Molecular, Facultad de Química, Universidad Complutense de Madrid, 28040 Madrid, Spain.
E-mail: ameva@bbm1.ucm.es

Summary

Background The C-terminal region of Ole e 1, a major allergen from olive pollen, is a dominant IgE-reactive site and offers a target for site-directed mutagenesis to produce variants with reduced IgE-binding capability.

Objective To evaluate in vitro and in vivo the immunogenic properties of three engineered derivatives of Ole e 1.

Methods One point (Y141A) and two deletion (135Δ10 and 140Δ5) mutants were generated by site-directed mutagenesis of Ole e 1-specific cDNA and produced in Pichia pastoris. Ole e 1 mutants were analysed for IgE reactivity by ELISA using sera from olive pollen-allergic patients. Their allergenicity was also investigated in both a mouse model of allergic sensitization and in basophil activation assays. IgG1 response was assayed by immunoblotting and competitive ELISA. T cell reactivity was evaluated by proliferation assays and cytokine production in splenocyte cultures.

Results The 135Δ10 mutant showed the strongest reduction in the IgE-binding capability of sera from olive pollen-allergic patients. Rat basophil leukaemia assays identified the deletion mutant 135Δ10 as the variant with the lowest β-hexosaminidase-releasing capacity. Furthermore, the same 135Δ10 mutant induced the lowest IgE levels in a BALB/c mouse model of sensitization. All Ole e 1 mutants retained their allergen-specific T cell reactivity. Immunization of mice with the mutants induced IgG1 antibodies, which cross-reacted with Ole e 1 and Ole e 1-like allergens from ash, lilac and privet pollens. The ability of the human IgE to block the binding of anti-Ole e 1 mutant-specific mouse IgG1 antibodies to natural Ole e 1 demonstrated that Ole e 1 mutants are able to induce in vivo antibodies reactive to the natural allergen.

Conclusion The 135Δ10 mutant with reduced allergenicity, intact T cell reactivity and capacity to induce blocking antibodies could provide a suitable candidate vaccine for efficient and safer therapy of olive pollen allergy.

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