ATHERO-FLUXTargeting novel lipid
pathways for treatment
of cardiovascular disease
The Athero-Flux workflow enables close integration between the Academic and SME capacities. The planned Academic and SME partnership will feed potential targets into the SME pipeline. The Academia will provide knowledge and experience in SL biology and state-of-the-art in vitro and in vivo models of atherosclerosis. The SMEs will in turn offer high-throughput lipidomics and screening technologies as well as LNA validation tools with a proven translational pipeline.
WP1 and WP2 form the TARGET IDENTIFICATION PHASE
WP1 will optimise new lipidomics platforms (“Flux”) to define molecular species of SLs and their kinetics over time, truly encompassing the complexity and the variety of SL molecular structures. WP1 will clarify mechanisms through which some specific SLs are increased in this category of patients (e.g. specific blocks or enhancements along the metabolic pathway).
WP2 will complement WP1 in target identification by employing whole genome siRNA screening to define the metabolic and signalling pathways affected by SLs in general. This approach will be coupled with the lipidomics tools described in WP1 and with phosphoproteomics approaches to study the signalling pathways affected by SL metabolism and signalling blockade. Due to the strong relationships between SL classes and cellular signalling and inflammation, WP2 will generate key new knowledge on the metabolic and cellular consequences of SL blockade and will identify a number of potential therapeutic targets within SL metabolism (e.g. enzymes) and associated signalling pathways (e.g. signalling intermediates and transcription factors). Systems biology analysis of the data (WP1, WP2) will generate a list of targets to be further evaluated and validated in remaining WPs.
WP3 and WP4 comprise the TARGET EVALUATION PHASE
WP3 will evaluate the promising hits from WP1 and WP2 and delve deeper into the biological significance of the hits and their relevance to the pathogenesis of atherosclerosis. Various in vitro assays will be deployed and/or developed to evaluate the hits in human and murine culture systems with different cell types relevant to atherosclerosis: atheroma cell cultures, hepatocytes, macrophages and endothelial cells. Lipidomics and proteomics will be utilised here as described above. Moreover, we will employ innovative high-resolution technology such as CyTOF allowing detection of at least 40 (and theoretically up to 100) cellular epitopes and activated pathways in a single cell. Human biobanks of atherosclerotic tissues will be utilised to compare changes in cells and serum with changes in the plaques.
WP4 will employ state-of-the-art animal models of atherosclerosis to study in vivo the effect of systemic factors (e.g. hyperlipidemia and gut microbiota) on the biodistribution of SL species in serum and vascular tissues using a combination of imaging and lipidomics. The outcome of these interactions on atherosclerosis development will be assessed. Moreover, the impact of changes in SL structure will be assessed in atherosclerosis using genetically modified mice.
WP5 will analyse the best hits from the TARGET IDENTIFICATION AND EVALUATION PHASE and produce LNA oligonucleotides for use in in vitro and in vivo models of atherosclerosis. The consequence of reducing the expression of selected targets (e.g. enzymes involved in SL production or rate-limiting steps in the associated signalling) on atherosclerosis development will be quantified. Lipidomics, imaging, CyTOF and other traditional technologies will be used to monitor changes in SL production in serum and tissues, in inflammatory signalling and other features of atherosclerosis such as foam cell formation and production of extracellular matrix.
In addition to the 5 research WPs the dissemination of the results and the successful direction of the project will be ensured by two complementary WPs dedicated to Management (WP6) and Dissemination (WP7).