Broad, long term objectives:

An adjuvant is an agent that may stimulate the immune system. Adjuvants are added to many vaccines to increase their immunogenicity and efficacy, while having few if any direct effects when given by themselves. There are many adjuvants in widespread use, including oils (commonly used in some veterinary vaccines), aluminium salts (aluminium phosphate and aluminium hydroxide), and virosomes, although precisely how they work is still not entirely understood. Recently, a new lipid A-based adjuvant, monophosphoryl lipid A (MPL), with enhanced humoral and cell-mediated immune responses to protein ligands is being championed by GlaxoSmithKline (GSK) in numerous vaccine trials. MPL is produced using purified from Salmonella minnesota R595 lipid A. Purified native lipid A is detoxified by removal of the phosphate group from the reducing end of the disaccharide backbone of lipid A by acid hydrolysis, producing MPL. The monophosphoryl lipid A is further detoxified by 3-O-deacylation with mild alkaline hydrolysis.

Instead of the chemical treatments described above used in the generation of MPL, our laboratory has undertaken a different approach in the synthesis of MPL-like molecules. Using heterologous expression of various acyltransferases, phosphatases, deaclyases, and glycosyltransferases in different bacterial backgrounds, we have been able to generate numerous lipid A structures with altered proimflammatory responses in humans and murine cells. Individual lipid A structures will be tested for adjuvant potential in a variety of in vitro and in vivo models.

Goal of the specific research proposed will:

i) Identification of acyltransferases, phosphatases, deaclyases, and glycosyltransferases that have the ability to modify lipid A structure in different bacterial backgrounds.

II) Analysis of individual lipid A structures using mass spectrometry and gas chromatography.

III) Analysis of adjuvant potential using a variety of in vitro and in vivo assay systems.