A multistage-polyepitope vaccine protects against Mycobacterium tuberculosis infection in HLA-DR3 transgenic mice

Mycobacterium tuberculosis (Mtb) is responsible for almost 2 million deaths annually. BCG, currently the only TB vaccine, induces variable protection and does not protect against reactivation of latent TB. Thus, efficient vaccines to supplement BCG are required urgently. Since Mtb's proteome differs qualitatively and quantitatively during bacterial replication stages from that expressed during dormancy, improved TB vaccines should drive immune responses to Mtb antigens expressed during multiple stages of infection. Consequently, such "multistage" vaccines should be composed of (immunodominant) antigens expressed during different phases of Mtb infection. As a concept multistage vaccine, we constructed a polyepitope by fusing five HLA-DR3-restricted T-cell epitopes derived from different Mtb proteins either expressed highly by replicating bacteria (Ag85B, hsp65, 19 kDa lipoprotein), or abundantly expressed by dormant bacilli and recognized preferentially by TST(+) individuals (hsp16, Rv1733c). PBMC of HLA-DR3(+) but not HLA-DR3(-) cured TB patients and TST(+) individuals responded well to the multistage-polyepitope in vitro. The in vivo immunogenicity and protective efficacy of the multistage-polyepitope were analyzed using HLA-DR3 transgenic mice lacking endogenous murine class II as a model. Immunization with the multistage-polyepitope adjuvanted with CpG generated high IgG levels as well as polyfunctional CD4(+) T-cells producing IFN-γ, TNF and IL-2, specific for these HLA-DR3-restricted epitopes. Importantly, multistage-polyepitope immunization reduced the number of bacilli in the lungs after Mtb challenge when administered as prophylactic vaccine. Given the extensive repertoire of potential Mtb antigens available for immune recognition, the data of our model demonstrate the potential of multistage-polyepitope vaccines to protect against TB.