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Novel approaches for TB vaccine and treatments : improving BCG vaccine and ‎identification of a potential new drug target

University of British Columbia

Current strategies of TB control include vaccination as a preventive approach and drug-based treatment as post-exposure therapy. In my PhD studies, I aimed to contribute in both directions: improving the current BCG vaccine and identifying and characterizing novel TB drug targets. I developed a novel non-genetic approach for the rapid display of exogenous proteins on mycobacterial cell surfaces as an alternative to DNA-based gene expression for upgrading BCG. A monomeric form of avidin that had the feature of reversible binding to biotin was chosen to generate avidin fusion proteins for surface decoration of biotinylated BCG. Avidin-tagged proteins bound to BCG surface reproducibly and stably with no effect on BCG growth. Thereafter, chimeric proteins corresponding to ovalbumin (OVA) and the M. tuberculosis (M.tb) specific ESAT6 antigen were generated and tested for their immunogenicity in vaccinated mice. BCG-OVA induced an immune response similar to that induced by BCG expressing the same surrogate antigen genetically. Furthermore, BCG decorated with ESAT6 successfully induced the expansion of specific T cell responses in vivo. This technology, therefore, can effectively replace traditional transformation of BCG with antigen-encoding genes and provide a novel platform for rapid evaluation of immunogenic proteins with broad applications in vaccine development. Mycobacterial lipoamide dehydrogenase (LpdC) is involved in the aberrant and prolonged retention of host protein coronin-1A (COR1A) on the phagosomal membrane, which contributes significantly to the inhibition of phagosome maturation, leading to intracellular persistence of M.tb. I constructed and utilized recombinant LpdC protein and ΔLpdC M.tb mutant to characterize LpdC and investigate potential mechanisms of LpdC-COR1A retention. ΔLpdC M.tb showed a decreased growth rate in standard media, induced phagolysosome fusion, and survived less than wild-type M.tb inside macrophages. I also found that LpdC interacted with a series of phosphoinositides (PIPs) and surprisingly aided in the induction of ROS production, which added to the multi-functionality of LpdC. From these studies, I have further characterized mycobacterial LpdC with regard to its role in M.tb persistence and paved ways for further investigations into LpdC’s potential as a novel drug target candidate.

Text

2016-09-08

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/59110

Experimental Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/