Mucosal immunogenicity of prion protein fused with heat-labile enterotoxin B subunit

Abstract

The new variant of human Creutzfeldt-Jakob disease is transmitted orally via food contaminated with prions of bovine spongiform encephalopathy from cattle, a fact that demands the development of vaccines, in particular those enhancingmucosal immunity, to prevent the transmission of prions between species. Prions are thought to consist mainly of the proteinase K-resistant isoform of prion protein (PrP) encoded on the host genome. There have been reports showing that the mucosal immunogenicity of a peptide could be enhanced by fusion with the B-subunit of E. coli heat-labile enterotoxin (LTB) or cholera toxin. In the present study, to determine whether PrP can be also immunogenically enhanced by fusion with LTB, we compared the mucosal immunogenicity of LTB-fused mouse (mo) and bovine (bo) PrP (LTB-mo or boPrP) with that of non-fused mo and boPrP by intranasally immunizing C57BL/6 and Balb/c mice three times every two weeksin the presence of recombinant LT as an adjuvant. In the fusion protein, the C-terminal half of mo and boPrP was linked to the C-terminus of LTB by hinge sequence of Gly-Pro-Gly-Pro. The biochemical properties of these fusion proteins were similar to those of LTB, including pentameric protein structure and GM I ganglioside-binding competence. Immune sera were collected one week after the final immunization and subjected to ELISA to detect the PrP-specific antibodies. MoPrP induced only a weak immune response in both mouse lines. No immunogenic enhancement could be detected in the mice immunized with LTB-moPrP. This poor immune response against moPrP and LTB-moPrP is attributable to the self-tolerance of mice to self-antigen. In contrast, boPrP was highly immunogenic inBalb/c mice but not in C57BL/6 mice. This indicates that the antibody response against PrP can differ widely in individuals. However, the fusion with LTB markedly enhanced immonogenicity of boPrP in both mouse lines, producing more than 100-fold IgG and IgA capable of cross-reacting with human and sheep PrPs but not with mo or hamster PrPs. This striking enhancement of the mucosal immunogenicity of boPrP in mice by fusion with LTB points to LTB-fused PrPs as a possible mucosal vaccine to elicit antibodies prophylactic for the oral transmission of prions from one species to another.