title: In vitro evolution of allergy vaccine candidates, with maintained structure, but reduced B cell and T cell activation capacity.
abstract: Allergy and asthma to cat (Felis domesticus) affects about 10% of the population in affluent countries. Immediate allergic symptoms are primarily mediated via IgE antibodies binding to B cell epitopes, whereas late phase inflammatory reactions are mediated via activated T cell recognition of allergen-specific T cell epitopes. Allergen-specific immunotherapy relieves symptoms and is the only treatment inducing a long-lasting protection by induction of protective immune responses. The aim of this study was to produce an allergy vaccine designed with the combined features of attenuated T cell activation, reduced anaphylactic properties, retained molecular integrity and induction of efficient IgE blocking IgG antibodies for safer and efficacious treatment of patients with allergy and asthma to cat. The template gene coding for rFel d 1 was used to introduce random mutations, which was subsequently expressed in large phage libraries. Despite accumulated mutations by up to 7 rounds of iterative error-prone PCR and biopanning, surface topology and structure was essentially maintained using IgE-antibodies from cat allergic patients for phage enrichment. Four candidates were isolated, displaying similar or lower IgE binding, reduced anaphylactic activity as measured by their capacity to induce basophil degranulation and, importantly, a significantly lower T cell reactivity in lymphoproliferative assays compared to the original rFel d 1. In addition, all mutants showed ability to induce blocking antibodies in immunized mice.The approach presented here provides a straightforward procedure to generate a novel type of allergy vaccines for safer and efficacious treatment of allergic patients.
authors.affiliation: Department of Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Ola B
authors.initials: OB
authors.lastname: Nilsson
authors.forename: Justus
authors.initials: J
authors.lastname: Adedoyin
authors.forename: Claudio
authors.initials: C
authors.lastname: Rhyner
authors.forename: Theresa
authors.initials: T
authors.lastname: Neimert-Andersson
authors.forename: Jeanette
authors.initials: J
authors.lastname: Grundström
authors.forename: Kurt D
authors.initials: KD
authors.lastname: Berndt
authors.forename: Reto
authors.initials: R
authors.lastname: Crameri
authors.forename: Hans
authors.initials: H
authors.lastname: Grönlund
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology; D001491:Basophils / Q000276:immunology; D002415:Cats; D003888:Desensitization, Immunologic / Q000379:methods*; D004926:Escherichia coli / Q000378:metabolism; D005260:Female; D015723:Gene Library; D006801:Humans; D006967:Hypersensitivity / Q000276:immunology* / Q000628:therapy; D007073:Immunoglobulin E / Q000737:chemistry; D008213:Lymphocyte Activation; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009154:Mutation; D019151:Peptide Library; D017386:Sequence Homology, Amino Acid; D013601:T-Lymphocytes / Q000276:immunology
chemical_list: D019151:Peptide Library; D007073:Immunoglobulin E