Background:  Tuberculosis (TB) accounts for nearly a third of AIDS deaths worldwide. Vaccines targeting latent TB infection would reduce the global burden of TB since one third of the world’s population has latent TB infection, and each of these individuals has a 10% lifetime risk of developing TB disease. Latent TB infection refers to those who are also HIV-infected and who have a 7 to 10% risk of developing TB disease within 1 year. Ideal vaccine candidates should be able to stimulate CD4 T-cells responses against Mycobacterium TB (Mtb). Since protection has been associated with response to antigens secreted by Mtb in culture filtrate we have identified multiple T-helper epitopes from Mtb secreted proteins to include them in a DNA vaccine construct.

Methods: Using EpiMatrix, a computer algorithm that ranks epitope sequences for the likelihood of binding to any given MHC, we identified promiscuous T-helper epitopes from proteins that were published members of the Mtb-secreted antigen family. Epitopes were validated by ELISpot and proliferation assays. We then used Web-based tools (SignalP and Prosite) to do a full TB genome scan (“GS-1”) to identify putative secreted proteins from Mtb 1551 and H37Rv genomes. A total of 24 epitopes, 23 derived from the pilot study and 1 GS-1 peptide (J), were then cloned into an expression vector separated by a spacer sequence to generate a prototype TB vaccine construct, EPV TB001. HLA DR B*0101 transgenic mice were vaccinated with 100 µg of EPVTB001 or a control plasmid with or without the addition of pIL-15 at day 0, 14, and 28. At day 35 mice were sacrificed and T-cell responses were measured in splenocytes derived from immunized animals as IFN gamma secretion in ELISpot assays.

Results:  ELISpot and proliferation assays showed that Mtb-immune individuals had responses to each of the 24 pilot peptides and to 15 of the 17 “Genome Scan” peptides. Expression of the translated EPV TB001 vaccine construct pseudoprotein was confirmed by Western blot analysis performed in the cell lysates after transient transfection of the DNA construct in 293T cells. In the HLA transgenic mice, epitope-specific T-cell responses were observed to 21 of 22 peptides representing the epitopes inserted in the vaccine constructs when multi-epitope vaccination was combined with pIL-15. 

Conclusions:  DNA vaccine constructs carrying multiple Mtb epitopes are able to induce de novo epitope-specific T-cells responses in vivo when adjuvanted by IL-15. 

Keywords: Vaccine; TB; Epitope