Abstract
Tuberculosis causes the deaths of 1. 8 million people per year and imposes a significant health burden on infected people. Tuberculosis presents many challenges in terms of both treatment and infection prevention. The chemotherapy course lasts for 6 months and there are increasing numbers of drug-resistant infections. The current BCG vaccination fails to control adult pulmonary disease and can cause infections in susceptible individuals. To address the deficiencies in protection induced by BCG vaccination research has been directed towards enhancement or replacement vaccine strategies. Mycobacterial Hsp70 has been shown to chaperone peptides in vivo and in vitro and has immunostimulatory activity, promoting maturation of antigen-presenting cells and the release of proinflammatory cytokines. This project has sought to harness the immunostimulatory activity and the peptide binding capacity of Hsp70 to deliver a range of mycobacterial peptides to the host immune system as a novel vaccine to cobacteria. This study showed that mycobacterial Hsp70 can be purified from BCG in commyplex with a range of different peptides and vaccination with the Hsp70-peptide complexes induced activation and IFNy production by CD4+ T-cells in addition to the secretion of IL-2, IFNy and TNFɑ. Conversely, vaccination with Hsp70 alone did not induce significant T-cell activation or cytokine production. Overexpression of Hsp70 in BCG did not enhance mycobacterial antigen-specific T-cell activation or cytokine production compared to vaccination with the BCG parent strain. Overexpression of both Hsp70 and GroEL in BCG did not induce a mycobacterial antigen-specific T-cell response. Recombinant Mycobacterium vaccae were engineered to express M. tuberculosis alpha-crystallin 2 (Acr2) and the fusion protein construct Hsp70-Acr2. Vaccination with M. vaccae that overexpressed Acr2 resulted in Acr2-specific IFNy production by CD4+ T-cells. M. vaccae that expressed Hsp70-Acr2 fusion protein had raised numbers of T-cells that expressed IFNy and IL4 for all culture conditions but lacked an Acr2-specific T-cell response. The research presented here suggests that the effect of mycobacterial Hsp70 on splenic T-cell function appears to be dependent upon the context of the Hsp70 and antigen-presenting-cell interaction, that over-expression of Hsp70 by mycobacterial species results in a mixed Type 1/Type 2 T-cell response, and that Acr2 and Hsp70-peptide complexes represent good heat shock protein candidates for further vaccine development.