Microbiology and immunology 2016 8 17() doi 10.1111/1348-0421.12410
DNA-based vaccine is a promising candidate for immunization and induction of a T cell-focused protective immune response against infectious pathogens such as Mycobacterium tuberculosis (M. tb). To induce multi-functional T response against multi-TB antigens, a multi-epitope DNA vaccine and a "protein backbone grafting" designing method is adopted to graft 5 discontinuous T cell epitopes into HSP65 scaffold protein of M. tb for enhancement of epitope processing and immune presentation. A DNA plasmid with 5 T cell epitopes derived from ESAT-6, Ag85B, MTB10.4, PPE25 and PE19 proteins of H37Rv strain of M. tb genetically inserted into HSP65 backbone was constructed and designated as pPES. After confirmation of its in vitro expression efficiency, pPES DNA was intramuscularly injected to C57BL/6 mice with 4 doses of 50µg DNA followed by mycobacterial challenge 4 weeks after the final immunization. It was found that pPES DNA injection maintained the ability of HSP65 backbone to induce specific serum IgG. ELISPOT assay demonstrated that pPES epitope-scaffold construct was significantly more potent to induce IFN-γ(+) T response to 5 T cell epitopes protein than other DNA constructs (with epitopes alone or with epitopes series connected to HSP65), especially in multi-functional-CD4(+) T response. It also enhanced granzyme B(+) CTL and IL-2(+) CD8(+) T response. In consistency, significantly improved protection against Mycobacterium bovis BCG challenge was achieved by pPES injection compared to other DNA constructs. Taken together, HSP65 scaffold grafting strategy for multi-epitope DNA vaccine represents a successful example of rational protein backbone engineering design and could prove useful in TB vaccine design.