A novel hyper-immunogenic low virulent BCG vaccine against tuberculosis

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SUMMARY Human morbidity and mortality due to tuberculosis (TB), caused by the bacterial pathogen Mycobacterium tuberculosis (Mtb), continue to be of significant health concern throughout the world. Bacille Calmette-Guerin (BCG) still remains the only approved vaccine against TB. Although BCG is considered safe and partially effective against extra-pulmonary childhood TB, its ability to protect against childhood and adult pulmonary TB is still questionable. In addition, there is a concern that BCG does not induce long lasting immune responses in the immunized individuals. However, our findings with Mtb- and BCG- derived recombinant vaccines strongly suggest that BCG can be improved to be more efficacious against TB. The goal of this proposal is to improve BCG by rationally deleting genes. We plan to sequentially delete three genes of BCG namely sapM, zmp1 and nuoG, through homologous recombination, to result in a triple knockout BCG (BCG-TKO) strain. These genes encode proteins that enable BCG to evade host immune response by preventing phagosome-lysosomal fusion, autophagy, apoptosis and other related processes in the antigen presenting cells (APCs) such as dendritic cells and macrophages. We hypothesize that deletion of these genes will allow the BCG-TKO strain to be efficiently processed by APCs, which will lead to increased antigen presentation to the immune cells and enhanced in vivo immunogenicity and efficacy. Additionally, we anticipate that deletion of these genes will reduce the virulence of the BCG, thus making the BCG-TKO to be HIV safe. We plan to accomplish our goal with three aims: 1) construct a BCG-TKO strain through homologous recombination, 2) analyze the immunogenicity and safety of the BCG-TKO strain in a SCID mouse model, and 3) investigate the efficacy of the BCG-TKO in the regular and humanized mouse models with and without HIV infection. Overall, we expect that this proposal will produce a highly efficacious, third generation BCG vaccine against TB that will be suitable for administration even in HIV infected infants.