Immunogenicity and Protective Efficacy against Murine Tuberculosis of a Prime-Boost Regimen with BCG and a DNA Vaccine Expressing ESAT-6 and Ag85A Fusion Protein

Abstract

Heterologous prime-boost regimens utilizing BCG as a prime vaccine probably represent the best hope for the development of novel tuberculosis (TB) vaccines. In this study, we examined the immunogenicity and protective efficacy of DNA vaccine (pcD685A) expressing the fusion protein of Ag85A and ESAT-6 (r685A) and its booster effects in BCG-immunized mice. The recombinant r685A fusion protein stimulated higher level of antigen-specific IFN-γ release in tuberculin skin test- (TST-) positive healthy household contacts of active pulmonary TB patients than that in TST-negative population. Vaccination of C57BL/6 mice with pcD685A resulted in significant protection against challenge with virulent Mycobacterium tuberculosis H37Rv when compared with the control group. Most importantly, pcD685A could act as a BCG booster and amplify Th1-type cell-mediated immunity in the lung of BCG-vaccinated mice as shown the increased expression of IFN-γ. The most significant reduction in bacterial load of both spleen and lung was obtained in mice vaccinated with BCG prime and pcD685A DNA booster when compared with BCG or pcD685A alone. Thus, our study indicates that pcD685A may be an efficient booster vaccine against TB with a strong ability to enhance prior BCG immunity.

Comparison of BCG prime-DNA booster and rBCG regimens for protection against tuberculosis

“…32 Antigen Ag85A (32kDa) has mycolyltransferase activity and plays an important role in the biogenesis of cell wall of mycobacteria. 33 As promising vaccine candidates for TB, both antigens were designed as the fusion protein r685A in our previous work 15 and we also confirmed that the whole blood cells from TST-positive healthy household contacts stimulated with the protein resulted in higher level of antigenspecific IFN-γ release than TST-negative population. 15 We constructed a DNA vaccine pcD685A expressing the fusion r685A protein and confirmed its protective effect as an effective booster vaccine of BCG.…”

“…33 As promising vaccine candidates for TB, both antigens were designed as the fusion protein r685A in our previous work 15 and we also confirmed that the whole blood cells from TST-positive healthy household contacts stimulated with the protein resulted in higher level of antigenspecific IFN-γ release than TST-negative population. 15 We constructed a DNA vaccine pcD685A expressing the fusion r685A protein and confirmed its protective effect as an effective booster vaccine of BCG. 15 In this study, r685A protein with the same size as the product purified from recombinant E.coli, was confirmed only in the culture supernatant of rBCG::685A strain as demonstrated by western blotting, which indicated that Hsp60 promoter and α-AgSP correctly directed the expression of secreted r685A protein from the rBCG strain.…”

“…15 We constructed a DNA vaccine pcD685A expressing the fusion r685A protein and confirmed its protective effect as an effective booster vaccine of BCG. 15 In this study, r685A protein with the same size as the product purified from recombinant E.coli, was confirmed only in the culture supernatant of rBCG::685A strain as demonstrated by western blotting, which indicated that Hsp60 promoter and α-AgSP correctly directed the expression of secreted r685A protein from the rBCG strain. Although splenocytes from rBCG::685A immunized mice secreted higher level of PPD or r685A protein specific IFN-γ than those of BCG, rBCG::685A did not provide any enhanced protection against M. tuberculosis challenge in the both lung and spleen, when compared with BCG vaccine.…”

“…6,7 To take advantage of the proven efficacy of BCG in children, BCG might first be used for neonatal vaccination as in most countries worldwide. 8 When the effects of BCG start to wane during adolescent, heterologous booster vaccines such as subunit vaccines, [9][10][11] DNA vaccines, [12][13][14][15][16] recombinant adenovirus, 17,18 and modified vaccinia virus, [19][20][21][22] could further be used to extend the protective efficacy of BCG priming. This strategy would benefit a significant proportion of the global population, because about 4 billion people have been immunized with BCG since 1921.…”

“…15 In the present study, a recombinant BCG strain expressing the fusion protein of ESAT-6 and Ag85A (rBCG::685A) was also constructed. The immune response and protective efficacy induced separately by rBCG::685A and BCG prime-pcD685A boost were evaluated and compared in vaccinated C57BL/6 mice.…”