1) To predict by computer modelling functionally relevant domains of apoA-I and apoA-II. 2) To utilise existing and generate new mutant forms of apoA-I and apoA-II that will be used to study the functions of these proteins. 3) To map the domains and residues of apoA-I and apoA-II involved in binding to the scavenger SR-BI type receptor or other cell receptors. 4) To map the domains and residues of apoA-I required for binding to lipids and activation of LCAT efflux of cholesterol from cells as well as the domains of apoA-II required for displacement of apoA-I from the surface of HDL. 5) To establish by transgenic methodologies how mutations that alter specific functions in vitro affect the functions or the concentrations of HDL in vivo and their impact on the development of atherosclerosis. 6) To generate and utilise commercially apoA-I and apoA-II derivatives of pharmacological importance for treatment or prevention of cardiovascular and other diseases.
Epidemiological and genetic data combined with recent transgenic experiments suggest that increased apoA-I and HDL levels protect from atherosclerosis. In contrast, low HDL levels predispose to coronary artery disease (CAD), a leading cause of mortality world-wide. Existing knowledge suggests that the protective functions of HDL are mediated by apoA-I whereas apoA-II antagonises the functions of apoA-I. The partners will use interdisciplinary approaches and state-of-the-art structural molecular and biochemical methodologies and their complementary expertise to elucidate the functions of apoA-I and apoA-II in vitro and in vivo.
Computer modelling of apoA-I and apoA-II domains and existing functional information will be utilised to generate new variant forms of apoA-I and apoA-II. Existing and new mutants of apoA-I and apoA-II forms will be purified on a large scale and the proteins will be utilised for functional analysis in vitro. These analyses will aim at mapping of the functionally relevant domains of apoA-I and apoA-II which are involved in high affinity binding to cell receptors (high affinity binding site on HepG2 cells and SR-BI), binding to lipids and activation of LCAT, efflux of cholesterol from cells.
Transgenic mouse models will also be utilised for functional analyses of apoA-I and apoA-II in vivo. This will involve generation of transgenic mice expressing selected variant forms of apoA-I and apoA-II and analysis of the in vivo functions of the mutant proteins including their capacity to predispose or protect from atherosclerosis.
Commercial exploitation of apoA-I and apoA-II derivatives will involve utilisation of apoA-I and apoA-II derivatives which have improved biological functions as lead compounds to develop drugs to protect from cardiovascular disease by the industrial partner Glaxo Welcome.
Successful completion of the proposed work is expected to yield a wealth of information on the biological mechanisms associated with the functions of apoA-I, apoA-II, HDL and their receptors which are relevant to the protection from CAD. This information will open new avenues which may lead to the development of new pharmaceuticals for the prevention and/or treatment of cardiovascular disease.
Apolipoprotein A-I, apolipoprotein A-II, HDL, CAD, A-I receptor, transgenic mice, scavenger SR-B1 receptor, reverse cholesterol transport, signal transduction, LCAT activation.
Funding SchemeCSC - Cost-sharing contracts