Therapeutic And Protective Effects Of Mycobacterium Tuberculosis Antigen Rv3425 And Fusion Antigen Ag85B - Rv3425 In Tuberculosis Vaccine

Tuberculosis (TB), caused by Mycobacterium tuberculosis, continues to be a major culprit of morbidity and mortality throughout the world. About one third of the world’s population are, or have been infected with Mycobacterium tuberculosis. In the 20th century, with the assistance of Bacillus Calmette-Guerin (BCG) and anti-TB drugs, TB burden has been reduced. However, the emergence of HIV and drug-resistant TB aggravated tuberculosis situation. But worse still, most anti-TB drugs used for clinical treatment are accompanied by long-term-therapy and serious side effect. This, in return, results in more incidence of drug-resistant tb and relapse. Furthermore, the only available vaccine BCG confers inconsistent efficacy, with protection varying from 0 to 80% in different countries. Hence, It is urgent to find novel therapeutic methods and new Tb vaccines.According to these two solutions, our research is divided into two parts:1. Immunotherapy against TB infection in mice with recombinant lentivirus that expresses fusion antigen Ag85B-Rv3425:For the first time, lentivirus was served as delivery vehicle in therapeutic vaccine against tuberculosis. Fusion gene of Ag85B-Rv3425 (A3) was introduced into pLenti6.3 vector and all plasmids were packaged to produce recombinant lentivirus A3-Len. An in vivo investigation into the titer of A3-specific antibody IgG was carried out on mice immunized with a single injection of recombinant lentivirus into foot pad. The result demonstrates that lentivirus can mediate the expression of A3 in vivo, which, immediately, is presented to immune system to initiate immune response. Moreover, mice received single injection of recombinant lentivirus display an increase in the proportion of CD4+ and CD8+ T cells as well as Thl immune response and an enhanced IFN-gamma and IL-2 production.During a therapeutic trial, mice were infected with H37Rv via intravenous challenge.4 weeks after infection, a single dose of recombinant lentivirus was given by foot pad injection. After a 8-week of lentivirus immunotherapy, bacterial load of the lung is decreased by 2/3 while body weight compared with that of mice immunized with PBS. These results indicate that recombinant lentivirus A3-Len is a very promising candidate for tuberculosis immunotherapy.2. Protection against tuberculosis challenge by a recombinant BCG prime followed by a protein boost vaccination strategy:Recombinant BCG that overexpressing fusion antigen A3 serves as a candidate for tuberculosis vaccine. Here we employed heterologous prime-boost strategy to seek a more effective combination. Our data suggests that mice received A3-BCG prime followed by Rv3425 protein boost vaccination perform a long-lasting CD4+cell proliferation and an increase ratio of CD19+CD27+memory B cell.To evaluate protection level, immunized mice were challenged via intravenous injection of H37Rv. Intriguingly, Rv3425 boost after A3-BCG prime strategy provides the highest protection level with a significant decline of bacterial load in lung and spleen in contrast to BCG or A3-BCG vaccination. This allows A3-BCG/Rv3425 to be a vaccine candidate with great promise.In summary, we evaluated the immune function of Ag85B and Rv3425 in therapy and protection effect and confirmed two promising candidate vaccines for immunotherapy and protection.