The recent development of means to insert genes of other organisms into mycobacteria has made it possible to develop BCG as a vector for vaccines against AIDS. However, in controlled clinical trials, the efficacy of BCG as a vaccine for tuberculosis has varied from 80% protection to no protection at all. The goals of this project are first to understand why only certain BCG preparations are good immunogens and secondly to develop practical procedures for improving the immunogenicity of recombinant HIV- BCG vaccines and to use BCG as an adjuvant for other HIV vaccines. The approach we take is based upon published evidence that several biologic functions including immunogenicity of BCG are strongly influenced by the outermost lipid layer of the organisms which is composed primarily of a p articular glycolipid, trehalose-6,6'-dimycolate(TDM). We developed models for studying the crystalline structures of TDM and relating these structures to biologic activity. A crystalline monolayer of TDM on the surface of microscopic beads was an effective adjuvant. Surfactants and lyophilization were found to disrupt the surface layers of TDM. Since similar surfactants and lyophilization are used in the production of modern BCG vaccines, this suggested that the 'improvement' in manufacturing methods may have altered the critical surface structures of BCG and reduced its immunogenicity. Our previous results also suggest that killed BCG components can be effective adjuvants for cell mediated immunity with chemically modified proteins if surface lipid structures are preserved. This project is designed to characterize the surface lipids of various preparations of BCG with particular emphasis on the crystalline monolayer of TDM and to determine its role in immunogenicity. Lipid analysis together with scanning and transmission electron microscopy will be used to investigate surface lipid structures of BCG grown and treated in ways designed to modify the surface glycolipids. The ability of block copolymer adjuvants to replace or augment the adjuvant effects of natural glycolipids will also be investigated. These results will be correlated with immunogenicity as measured by assays for antibody, cell mediated immunity and protection. This information will then be used in efforts to prepare both live and killed recombinant HIV- BCG vaccines with improved immunogenicity.
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