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Background: Previously we have shown that Ag85B-TB10.4 is a highly efficient vaccine against tuberculosis when delivered in a Th1 inducing adjuvant based on cationic liposomes. Another Th1 inducing adjuvant, which has shown a very promising profile in both preclinical and clinical trials, is IC31. In this study, we examined the potential of Ag85B-TB10.4 delivered in the adjuvant IC31 for the ability to induce protection against infection with Mycobacterium tuberculosis. In addition, we examined if the antigen dose could influence the phenotype of the induced T cells., Methods and Findings: We found that vaccination with the combination of Ag85B-TB10.4 and IC31 resulted in high numbers of polyfunctional CD4 T cells co-expressing IL-2, IFN-gamma and TNF-alpha. This correlated with protection against subsequent challenge with M.tb in the mouse TB model. Importantly, our results also showed that both the vaccine induced T cell response, and the protective efficacy, was highly dependent on the antigen dose. Thus, whereas antigen doses of 5 and 15 microg did not induce significant protection against M.tb, reducing the dose to 0.5 microg selectively increased the number of polyfunctional T cells and induced a strong protection against infection with M.tb. The influence of antigen dose was also observed in the guinea pig model of aerosol infection with M.tb. In this model a 2.5 fold increase in the antigen dose reduced the protection against infection with M.tb to the level observed in non-vaccinated animals., Conclusions/significance: Small changes in the antigen dose can greatly influence the induction of specific T cell subpopulations and the dose is therefore a crucial factor when testing new vaccines. However, the adjuvant IC31 can, with the optimal dose of Ag85B-TB10.4, induce strong protection against Mycobacterium tuberculosis. This vaccine has now entered clinical trials.

Background: Recently we and others have identified CD8 and CD4 T cell epitopes within the highly expressed M. tuberculosis protein TB10.4. This has enabled, for the first time, a comparative study of the dynamics and function of CD4 and CD8 T cells specific for epitopes within the same protein in various stages of TB infection., Methods and Findings: We focused on T cells directed to two epitopes in TB10.4; the MHC class I restricted epitope TB10.4 (3-11) (CD8/10.4 T cells) and the MHC class II restricted epitope TB10.4 (74-88) (CD4/10.4 T cells). CD4/10.4 and CD8/10.4 T cells displayed marked differences in terms of expansion and contraction in a mouse TB model. CD4/10.4 T cells dominated in the early phase of infection whereas CD8/10.4 T cells were expanded after week 16 and reached 5-8 fold higher numbers in the late phase of infection. In the early phase of infection both CD4/10.4 and CD8/10.4 T cells were characterized by 20-25% polyfunctional cells (IL-2(+), IFN-gamma(+), TNF-alpha(+)), but whereas the majority of CD4/10.4 T cells were maintained as polyfunctional T cells throughout infection, CD8/10.4 T cells differentiated almost exclusively into effector cells (IFN-gamma(+), TNF-alpha(+)). Both CD4/10.4 and CD8/10.4 T cells exhibited cytotoxicity in vivo in the early phase of infection, but whereas CD4/10.4 cell mediated cytotoxicity waned during the infection, CD8/10.4 T cells exhibited increasing cytotoxic potential throughout the infection., Conclusions/significance: Our results show that CD4 and CD8 T cells directed to epitopes in the same antigen differ both in their kinetics and functional characteristics throughout an infection with M. tuberculosis. In addition, the observed strong expansion of CD8 T cells in the late stages of infection could have implications for the development of post exposure vaccines against latent TB.

Background: Although CD4 T cells are crucial for defense against M.tb, it is still not clear whether the optimal response against M.tb in fact involves both CD4 and CD8 T cells. To test this, we used a new vaccine strategy that generated a strong balanced T cell response consisting of both CD4 and CD8 T cells., Methods and Findings: To compare CD4 and CD8 responses against Ag85B-TB10.4 (H4), H4 was delivered as a subunit vaccine in cationic liposomes (CAF01), expressed in Ad5 (Ad-H4) or as a heterologous prime boost vaccination. H4/CAF01 induced primarily CD4 T cells and Ad-H4 gave predominantly a CD8 T cell response. In contrast, the heterologous prime boost combination resulted in augmentation of both the CD4 and CD8 response. The majority (>40%) of the CD4 T cells induced by the heterologous prime boost protocol were polyfunctional, and expressed IFN-gamma(+), IL-2(+), and TNF-alpha(+), whereas most of the CD8 T cells expressed IFN-gamma(+) and TNF-alpha(+) and possessed strong cytotoxic potential. The heterologous prime boost protocol also gave an increase in protective efficacy against M.tb challenge compared to H4/CAF01 and Ad-H4. Both the H4 specific CD4 and CD8 T cells were recruited to the site of infection, at the onset of infection. However, compared to CD8 T cells, CD4 T cells showed more extensive recruitment and were the main T cell subset proliferating at the site of infection., Conclusions/significance: Heterologous prime boost based on H4, produced an additive effect on the priming of CD4 and CD8 cells and in terms of the protective capacity of the vaccine, and therefore represent an interesting new vaccine strategy against M.tb. However, CD4 and CD8 T cells respond very differently to live M.tb challenge, in a manner which supports the consensus that CD4 T cells do play the major role during the early stages of an M.tb infection.