Generation of hypoallergenic DNA vaccines by forced ubiquitination: Preventive and therapeutic effects in a mouse model of allergy

Background

Hypoallergenic immunotherapy of type I allergies aims at inducing T-cell immunity while avoiding cross-linking of pre-existing IgE. DNA-based immunotherapy depends on the recruitment of antigen-specific T_H1 cells and therefore has to provide the whole repertoire of T-cell epitopes. Ubiquitination offers a general approach for the production of hypoallergenic DNA vaccines.

Objective

A DNA-based vaccine encoding the major birch pollen allergen Bet v 1 stably linked to ubiquitin was evaluated for its antiallergic potential in a BALB/c mouse model of allergy.

Methods

Plasmid DNA was applied to mice before (preventive) or after (therapeutic) sensitization with recombinant Bet v 1. In the preventive setting, mice were exposed to aerosolized allergen in addition. Cytokine production was monitored via ELISPOT and Luminex. IgG₁, IgG_2a, and IgE subclass antibody titers were determined by ELISA. In vitro antigen-specific cross-linking of IgE was measured in a degranulation assay. Bronchoalveolar lavages were analyzed for leukocyte subsets as well as for IFN-γ and IL-5, and paraffin sections of lungs were examined for mucus production and endothelial damage.

Results

Prevaccination with ubiquitinated Bet v 1–stimulated T_H1-biased immune responses with concomitant suppression of functional IgE, reduction of eosinophil counts in bronchoalveolar lavages, and alleviation of lung pathology, and could also suppress an ongoing IgE response in a therapeutic setting.

Conclusion

The data clearly demonstrate that hypoallergenic DNA vaccines encoding ubiquitin fusion constructs induce effective antiallergic immune responses.

Clinical implications

Ubiquitination of allergen gene vaccines eliminates the risk of IgE cross-linking, thereby meeting the safety requirements for clinical applications.

Key words: Type I allergy, DNA vaccine, genetic immunization, gene therapy, ubiquitin, IgE, bronchoalveolar lavage, eosinophils, lung pathology

Abbreviations used: BAL, Bronchoalveolar lavage, pcDNA Luc, Plasmid vector expressing firefly luciferase, pCMV, Empty plasmid vector, pCMV-β, Plasmid vector expressing β-galactosidase, pCMV-Bet, Plasmid vector expressing Bet v 1a, pCMV-ubi-β, Plasmid vector expressing ubiquitinated β-galactosidase, pCMV-ubi-Bet, Plasmid vector expressing ubiquitinated Bet v 1a, RBL, Rat basophil leukemia, SIT, Specific immunotherapy