Recombinant Hypoallergenic Cat Allergy Vaccines

Abstract

Background: Molecular forms of allergen-specific immunotherapy (AIT) for cat allergy are needed. Fel d 1, Fel d 4, and Fel d 7 are the most important cat allergens. Methods: IgE epitopes of Fel d 4 and Fel d 7 were mapped by blocking allergic patients' IgE binding with allergen peptide-specific antisera. Five recombinant fusion proteins (PreS-Cat 1-PreS-Cat 5) each containing hepatitis B virus (HBV)-derived PreS as an immunological carrier and non-allergenic peptides from the IgE binding sites of Fel d 1, Fel d 4, and Fel d 7 were expressed in Escherichia coli, purified, and characterized by mass spectrometry, circular dichroism (CD), and size exclusion chromatography. ImmunoCAP and basophil activation experiments demonstrated the hypoallergenic activity of PreS-Cat 1–5. The ability of PreS-Cat 1–5 to induce IgE-blocking antibodies in rabbits was compared to three licensed allergen extract-based AIT vaccines. PreS-Cat 1-5-specific IgG antibodies were tested for inhibition of allergen-specific IgE binding and specific basophil activation. T cell activation and induction of specific cytokine secretion by PreS-Cat proteins were compared with cat allergens in PBMC cultures. Results: Recombinant hypoallergenic, biochemically and structurally defined PreS-Cat 1–5 were obtained. Two subcutaneous immunizations of rabbits with PreS-Cat 1–5 induced equal (Fel d 1) or better (Fel d 4 and Fel d 7) antibodies (PreS-Cat 5 > PreS-Cat 1 > PreS-Cat 3) blocking allergic patients' IgE binding to cat allergens than six to fifteen immunizations with allergen extract-based vaccines. PreS-Cat-specific antibodies strongly inhibited specific basophil activation. PreS-Cat 5 > PreS-Cat 1 induced significantly more IL-10 in cultured PBMCs from cat allergic patients than cat allergens. Conclusions: PreS-Cat 5 and PreS-Cat 1 are highly promising molecular vaccine candidates for AIT of cat allergy, combining Fel d 1-, Fel d 4-, and Fel d 7-peptides in single PreS fusion proteins.