Immunogenicity and Protective Efficacy of a Fusion Protein Vaccine Consisting of Antigen Ag85B and HspX against Mycobacterium tuberculosis Infection in Mice

Abstract

Subunit vaccines have the potential advantage to boost Mycobacterium bovis Bacillus Calmette‐Guérin (BCG)‐primed immunity in adults. However, most candidates are antigens highly expressed in replicating bacilli but not in dormant or persisting bacilli, which exist during Mycobacterium tuberculosis infection. We constructed M. tuberculosis fusion protein Ag85B‐Mpt64190–198‐HspX (AMH) and Ag85B‐Mpt64190–198‐Mtb8.4 (AMM), which consist of Ag85B, the 190–198 peptide of Mpt64, HspX (Rv2031c) and Mtb8.4 (Rv1174c), respectively. AMH and/or AMM were mixed with adjuvants composed of dimethyl‐dioctyldecyl ammonium bromide and BCG polysaccharide nucleic acid (DDA‐BCG PSN) to construct subunit vaccines. Mice were immunized thrice with Ag85B, AMH and AMM vaccines and the immunogenicity of the fusion protein vaccines was determined. Then, mice were primed with BCG and boosted twice with Ag85B, AMH, AMM and AMM + AMH vaccines, respectively, followed by challenging with M. tuberculosis virulent strain H37Rv, and the immune responses and protective effects were measured. It was found that mice immunized with AMH vaccine generated high levels of antigen‐specific cell‐mediated responses. Compared with the group injected only with BCG, the mice boosted with AMM, AMH and AMM + AMH produced higher levels of Ag85B‐specific IgG1 and IgG2a and IFN‐γ‐secreting T cells upon Ag85B and Mycobacterium tuberculosis purified protein derivative (PPD) stimulation. It is interesting that only mice boosted with AMM + AMH had significantly lower bacterial count in the lungs than those receiving BCG, whereas mice boosted with AMH or AMM did not. The results suggest that AMH consisting of HspX, the antigen highly expressed in dormant bacilli, could be combined with antigens from replicating bacilli to enhance BCG primed immunity so as to provide better protection against both growing and non‐growing bacteria that occur during the infection process.

Immunogenicity of a Fusion Protein Containing Immunodominant Epitopes of Ag85C, MPT51, and HspX from Mycobacterium tuberculosis in Mice and Active TB Infection

“…2). These antibody classes were similarly induced to levels comparable to those induced by other fusion protein constructs, such as the Ag85B-MPT64 190–198 -HspX construct that was used as vaccine booster for BCG in mice or Mtb10.4-HspX (MH) emulsified in DDA–TDM for use as subunit vaccine [26]–[27].…”

Subunit Vaccine Consisting of Multi-Stage Antigens Has High Protective Efficacy against Mycobacterium tuberculosis Infection in Mice

“…We found that mice receiving EAMM plus MH had significantly lower bacterial counts in the lungs and spleens compared with EAMM or MH alone. In addition, our previous study also found that fusion protein AMH containing dormancy-related antigen could enhance the effect of AMM [41]. MH and AMH alone did not confer obvious protective effect, but when they were combined with EAMM or AMM they improved the protective effect of EAMM and AMM significantly.…”

“…Up to now, our lab has evaluated 9 vaccine candidates based on 9 fusion proteins including AMM [36], Ag85B-MPT64 190-198 -HspX (AMH) [41], EAMM, MH [35], ESAT6-Ag85B, ESAT6-Mtb8.4, Mtb10.4-Ag85B, ESAT6-RpfE, and Ag85B. We found that EAMM provided the most effective protection against M.…”

HspX‐mediated protection against tuberculosis depends on its chaperoning of a mycobacterial molecule

“…10, 11 Until recently, the majority of subunit vaccines were based on antigens expressed during the early stages of M. tuberculosis infection; however, to target protection during latency, several current studies have incorporated the latency-associated antigen, 16-kDa α-crystallin protein (also called HspX, acr and Rv2031c) in heterologous prime-boost strategies using protein HspX subunit vaccines, DNA vaccines expressing HspX and recombinant BCG strains overexpressing HspX. 12, 13, 14, 15 …”