Evaluation of Immune Responses to a DNA Vaccine Encoding Ag85a-Cfp10 Antigen of Mycobacterium tuberculosis in an Animal Model

Many studies indicate that the Bacillus Calmette-Guérin (BCG) vaccine has low protective efficacy, especially in endemic areas. There are several factors in this assessment, such as genetic diversity of BCG strains, pre-exposure to environmental mycobacteria, and variations in host immune responses. Currently, more than 200 new vaccine candidates have been proposed, such as recombinant BCG, DNA, and subunit vaccines. However, none of them are superior to BCG. Nevertheless, several approaches are considered to reduce the cases of tuberculosis infection. The aim of the present study was to evaluate the capability of the Ag85a-cfp10 fusion protein as a new chimeric protein in stimulating immune responses. A DNA vaccine encoding Ag85a-cfp10 fusion protein was constructed in the previous study. The expression of Ag85a-cfp10 fusion protein in host cells was confirmed by the RT-PCR method. Six pathogen-free female mice were injected intramuscularly at a total concentration of 100 µg/mL three times at two-week intervals. The BCG and the control groups received BCG and PBS vaccines, respectively. One month after the final immunization, mice were killed and their splenocytes were cultured in RPMI medium supplemented with 1% antibiotics and 10% serum. Four cytokines including IL-4, IL-12, TGF-β, and IFN-γ were measured in the culture supernatant using the ELISA test. RT-PCR analysis showed that Ag85a-cfp10 recombinant vector is able to replicate in eukaryotic cells and produce mRNA. The vaccinated groups were compared to the control group, showing induction of high levels of cytokine production. Some reports depicted that DNA vaccines are able to induce humoral and cellular immune responses both in animal models and humans. Therefore, in the current study, the immune response was induced in mice, which were inoculated with recombinant expression plasmid, pcDNA3.1 (+)-Ag85a-cfp10. We showed that this recombinant vector can stimulate mycobacterial specific modulating cytokines. Nonetheless, analysis appeared that this vaccine is unable to stimulate cell mediated immunity, however, still further studies are needed in future.