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Characterization of a P ar j 1/ P ar j 2 mutant hybrid with reduced allergenicity for immunotherapy of P arietaria allergy
Author(s) -
Bonura A.,
Passantino R.,
Costa M. A.,
Montana G.,
Melis M.,
Luisa Bondì M.,
Butteroni C.,
Barletta B.,
Corinti S.,
Felice G. Di,
Colombo P.
Publication year - 2012
Publication title -
clinical and experimental allergy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.462
H-Index - 154
eISSN - 1365-2222
pISSN - 0954-7894
DOI - 10.1111/j.1365-2222.2011.03938.x
Subject(s) - immunogenicity , basophil activation , immunoglobulin e , recombinant dna , allergy , antibody , microbiology and biotechnology , immunology , western blot , allergen , immunotherapy , immune system , sensitization , allotype , monoclonal antibody , biology , basophil , chemistry , biochemistry , gene
Summary Background P arietaria pollen is one of the major cause of pollinosis in the southern E urope. Specific immunotherapy is the only treatment able to modify the natural outcome of the disease restoring a normal immunity against allergens. Methods We designed a recombinant molecule ( P j ED loop1) comprised of genetic‐engineered variants of the major allergens of the P arietaria pollen ( P ar j 2/ P ar j 1). Purity and chemical–physical properties of the derivative were analysed by RP ‐ HPLC chromatography and Photon Correlation Spectroscopy. Immunological activity was evaluated by means of W estern blotting, ELISA inhibition and PBMC proliferation assay in 10 P arietaria allergic patients. Basophil activation was studied in six subjects. The immunogenicity of the hybrid was studied looking at the immune responses induced in a mouse model of sensitization. Results The P j ED loop1 hybrid was produced as a purified recombinant protein with high stability in solution. W estern blot, ELISA inhibition and basophil activation test showed that the P j ED loop1 displays a remarkable reduced IgE binding and anaphylactic activity. CD 3 reactivity was conserved in all patients. Mice immunization with the r P j ED loop1 induced antibodies and T cell responses comparable to that obtained by the wild type allergens. Such antibodies shared the specificities to r P ar j 1 and r P ar j 2 with human IgE antibodies. Conclusion Our results demonstrated that a mutant hybrid expressing genetically engineered forms of the major P . judaica allergens displayed reduced allergenicity and retained T cell reactivity for the induction of protective antibodies in vaccination approaches for the treatment of P arietaria pollinosis.