Immune responses after immunization with plasmid DNA encoding Bet v 1, themajor allergen of birch pollen

Background: Immunization with plasmid DNA encoding various antigens is a pr omising method in vaccine research. Recent studies also indicate that DNA-b ased 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 Ia, the major a llergen 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 Bet v 1a (pCMV -Bet). In addition, the effect of immunostimulatory DNA sequences was inves tigated by appending CpG motifs to the gene of Bet v 1a, coinjecting CpG-ol igodeoxynucleotides together with the pCMV-Bet construct, or both. IgE and IgG antibody responses, as well as IgG subclasses, were measured by ELISA i n sera after each immunization. IFN-gamma and IL-4 levels were also measure d by ELISA in sera and supernatants of allergen-stimulated spleen cells. Results: The primary humoral response to a single treatment with pCMV-Bet w as very weak, but the reaction could be boosted to higher levels by 2 addit ional injections. On the other hand, proliferation assays of spleen cells a nd measurements of cytokine levels already indicated a cellular response af ter the first injection of plasmid DNA. After 2 immunizations with pCMV-Bet , the ratio of IgG1 to IgG2a pointed to a T-K1 subclass profile. IgE was no t detectable in any group at any time during the immune reaction. According ly, IL-4 levels were markedly reduced in the serum, as well as in the super natants, of stimulated spleen cells. Animals immunized with pCMV-Bet contai ning 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. T he levels of IFN-gamma measured after allergen stimulation of isolated sple en cells were significantly higher in animals immunized with pCMV-Bet plus CpG motifs than with pCMV-Bet alone. Immunization with recombinant Bet v 1a protein elicited a strong T-K2-type response, including IgE production, a high titer of IgG1, and IL-4 production in both serum and supernatants of p roliferation cultures. Conclusion: In contrast to immunization with protein, DNA immunization indu ces a storng T-K1-type response against a relevant inhalant allergen. Our d ata support the concept of developing a novel type of allergen immunotherap y based on plasmid DNA immunization. (J Allergy Clin Immunol 1999:103:107-1 3.)