Background: Immunization with plasmid DNA encoding various antigens is a promising method in vaccine research. Recent studies also indicate that DNA- based immunization might represent a potential approach in allergen-specific immunotherapy. Objective: In this study we have characterized the immune responses induced by recombinant Bet v 1a and plasmid DNA encoding for Bet v 1a, the major allergen of birch pollen in a mouse system. Methods: Balb/c mice were injected intraperitoneally with recombinant Bet v 1a and intradermally with plasmid DNA encoding for the gene of Bet v 1a (pCMV-Bet). In addition, the effect of immunostimulatory DNA sequences was investigated by appending CpG motifs to the gene of Bet v 1a, coinjecting CpG- oligodeoxynucleotides together with the pCMV-Bet construct, or both. IgE and IgG antibody responses, as well as IgG subclasses, were measured by ELISA in sera after each immunization. IFN-γ and IL-4 levels were also measured by ELISA in sera and supernatants of allergen-stimulated spleen cells. Results: The primary humoral response to a single treatment with pCMV-Bet was very weak, but the reaction could be boosted to higher levels by 2 additional injections. On the other hand, proliferation assays of spleen cells and measurements of cytokine levels already indicated a cellular response after the first injection of plasmid DNA. After 2 immunizations with pCMV-Bet, the ratio of IgG1 to IgG2a pointed to a T(H1) subclass profile. IgE was not detectable in any group at any time during the immune reaction. Accordingly, IL-4 levels were markedly reduced in the serum, as well as in the supernatants, of stimulated spleen cells. Animals immunized with pCMV-Bet containing appended CpG motifs at the 3' end of the Bet v 1a gene and/or with the CpG-ODN GCTAGACGTTAGCGT plus pCMV-Bet displayed reduced humoral responses against Bet v 1a when compared with animals injected with pCMV-Bet alone. The levels of IFN-γ measured after allergen stimulation of isolated spleen cells were significantly higher in animals immunized with pCMV-Bet plus CpG motifs than with pCMV-Bet alone. Immunization with recombinant Bet v la protein elicited a strong T(H2)-type response, including IgE production, a high titer of IgG1, and IL-4 production in both serum and supernatants of proliferation cultures. Conclusion: In contrast to immunization with protein, DNA immunization induces a strong T(H1)-type response against a relevant inhalant allergen. Our data support the concept of developing a novel type of allergen immunotherapy based on plasmid DNA immunization.
Hartl, A., Kiesslich, J., Weiss, R., Bernhaupt, A., Mostböck, S., Scheiblhofer, S., … Thalhamer, J. (1999). Immune responses after immunization with plasmid DNA encoding Bet v 1, the major allergen of birch pollen. Journal of Allergy and Clinical Immunology, 103(1 I), 107–113. https://doi.org/10.1016/S0091-6749(99)70533-6