T Cell Epitope-Containing Domains of Ragweed Amb a 1 and Mugwort Art v 6 Modulate Immunologic Responses in Humans and Mice

Abstract

BackgroundRagweed (Ambrosia artemisiifolia) and mugwort (Artemisia vulgaris) are the major cause of pollen allergy in late summer. Allergen-specific lymphocytes are crucial for immune modulation during immunotherapy. We sought to generate and pre-clinically characterise highly immunogenic domains of the homologous pectate lyases in ragweed (Amb a 1) and mugwort pollen (Art v 6) for immunotherapy.MethodsDomains of Amb a 1 (Amb a 1α) and Art v 6 (Art v 6α) and a hybrid molecule, consisting of both domains, were designed, expressed in E. coli and purified. Human IgE reactivity and allergenicity were assessed by ELISA and mediator release experiments using ragweed and mugwort allergic patients. Moreover, T cell proliferation was determined. Blocking IgG antibodies and cytokine production in BALB/c mice were studied by ELISA and ELISPOT.ResultsThe IgE binding capacity and in vitro allergenic activity of the Amb a 1 and Art v 6 domains and the hybrid were either greatly reduced or abolished. The recombinant proteins induced T cell proliferative responses comparable to those of the natural allergens, indicative of retained allergen-specific T cell response. Mice immunisation with the hypoallergens induced IL-4, IL-5, IL-13 and IFN-γ production after antigen-specific in vitro re-stimulation of splenocytes. Moreover, murine IgG antibodies that inhibited specific IgE binding of ragweed and mugwort pollen allergic patients were detected.ConclusionAccumulation of T cell epitopes and deletion of IgE reactive areas of Amb a 1 and Art v 6, modulated the immunologic properties of the allergen immuno-domains, leading to promising novel candidates for therapeutic approach.

Diverse and highly cross‐reactive T‐cell responses in ragweed allergic patients independent of geographical region

“…T‐cell reactivity to multiple Amb a 1 epitopes seemed to be more likely in patients with high Amb a 1 IgE (>5 kU/L) in line with the cut‐off for strong T‐cell responses towards mite allergen epitopes (>10 kU/L) suggested by Hinz et al The micro‐heterogeneity of the T‐cell responses in individual donors may explain why various attempts to develop next‐generation AIT vaccines based on a single recombinant Amb a 1 isoform or short peptides derived from such a molecule have failed to progress beyond phase II clinical trials. It may also predict similar challenges for the suggested hybrid molecule combining parts of Amb a 1 and Art v 6 . Moreover, one donor responded only to minor allergen epitopes and other donors responded to multiple epitopes in the minor allergens, indicating that T‐cell responses to minor allergens can be important factors in allergic disease.…”

Allergen Immunotherapy: Current and Future Trends

“…Another hybrid recombinant of ambrosia pectate lyase (rAmb a 1, residues 1174–397) and mugwort (rArt v 6, residues 173–396), as well as a hybrid of both, have been investigated. The recombinant proteins and chimera did not cause the recognition of IgE in patients sensitive to these weeds or the degranulation of preformed mediators or cytokines of the Th2 profile [ 137 ]. Additionally, a fusion protein (rFlaA: Art1hyp), integrated by flagellin A (TLR5 agonist) and hypoallergenic mugwort (Art v1- change in cysteines for serines, altering the epitope for IgE), stimulates the synthesis of subclasses of IgG but not of IgE [ 138 ].…”

DNA vaccines and recombinant allergens with reduced allergenic activity treat allergies

“…Although B cells remain intact during immunotherapy, some studies admitted the induction of allergen-specific IgG to T-epitope vaccines. This fact indicates the presence of preserved antibody epitopes in these vaccines [11,14,20,[24][25][26]. Compared with ASIT and sublingual immunotherapy (SLIT), immunization with immunodominant T-cell epitope peptides promotes the absence of immediate IgEdependent reactions [11,27,28] and provides sustained clinical benefits after a relatively short treatment course [26,29].…”