Recombinant BCG Overexpressing phoP-phoR Confers Enhanced Protection against Tuberculosis

Abstract

The live tuberculosis vaccine Mycobacterium bovis BCG (Bacille Calmette-Guérin) comprises a number of genetically distinct substrains. In BCG-Prague, phoP of the PhoP-PhoR two-component system is a pseudogene due to a single insertion mutation. We hypothesized that this mutation partially accounts for the low immunogenicity of BCG-Prague observed in the 1970s. In this study, we showed that complementation with the M. bovis allele of phoP restored BCG-Prague’s immunogenicity. Furthermore, we showed that overexpression of the M. bovis allele of phoP-phoR in BCG-Japan, a strain already containing a copy of phoP-phoR, further enhanced immunogenicity and protective efficacy. Vaccination of C57BL/6 mice with the recombinant strain rBCG-Japan/PhoPR induced higher levels of interferon-γ (IFN-γ) production by CD4+ T cells than that with the parental BCG. Guinea pigs vaccinated with rBCG-Japan/PhoPR were better protected against challenge with Mycobacterium tuberculosis than those immunized with the parental BCG, showing significantly longer survival time, reduced bacterial burdens, and less severe pathology. Taken together, our study has identified a genetic modification that could be generally applied to generate new recombinant BCG vaccines.

Preclinical Progress of Subunit and Live Attenuated Mycobacterium tuberculosis Vaccines: A Review following the First in Human Efficacy Trial

“…Guinea pigs subcutaneously vaccinated with rBCG-Japan had a reduced bacterial load after 8 weeks compared to unvaccinated guinea pigs (lung p < 0.05; spleen n.s.) and increased survival by 133% over the parental BCG strain after M. tb H37Rv infection [35]. This study suggested that recombinant BCG overexpressing phoP-phoR is a promising candidate to replace BCG.…”

“…A major hindrance is the lack of a known immune correlate of protection in animals and humans [116]. As evidenced by many clinical and preclinical studies, robust immunogenicity does not always translate to improved protective efficacy [26,35,102]. Another hurdle is the lack of an animal model that faithfully recapitulates the spectrum of human TB disease including latent TB infection (LTBI) [117].…”

“…PhoP-PhoR is a two-component transcriptional regulator of a number of genes in M. tb, including two T cell antigens (Ag85A, PPE18) [35]. A loss of function mutation of phoP from BCG-Prague results in lower immunogenicity compared to other BCG strains [35]. BCG-Japan, a strain containing phoP-phoR, was engineered to overexpress phoP-phoR and evaluated for its safety, immunogenicity and protection in mice and guinea pigs.…”

After 100 Years of BCG Immunization against Tuberculosis, What Is New and Still Outstanding for This Vaccine?

“…( 4) Another refinement to the use of RD1 was recently reported in the BCG::ESAT6-PE25SS strain, which secretes full-length ESAT-6 via the ESX-5 secretion system, and reduced M. tuberculosis H37Rv loads in lungs as well lung pathology in intratracheallyvaccinated C57BL/6 more than parental BCG [52], so it will be interesting to see further development of this vaccine candidate. (5) BCG::phoPR, an interesting candidate based on the fact that the phoPR genes from BCG Pasteur were introduced into BCG Japan to restore it from its natural point mutations, and which prolonged survival of guinea pigs infected with M. tuberculosis H37Rv compared with parental BCG [53]. Whether this specific recombinant BCG strain will undergo further development or will be the basis of possible restoration of gene function in other rBCG vaccine candidates remains to be seen.…”

Vaccines Against Tuberculosis: Problems and Prospects (Review)

“…Speaking of the genetic characteristics of BCG strains, it is interesting to consider one of them in more detail. In research on the BCG-Prague strain, a phoP gene mutation was detected in the form of a single nucleotide insert, which led to the destruction of the C-terminal DNA-binding domain [ 11 , 12 ]. This mutation is specific for BCG-Prague and is not found in other BCG strains [ 13 ].…”