The role of Immune-responsive gene 1 and itaconate in atherosclerotic disease

PROJECT SUMMARY Cardiovascular disease (CVD) is the leading cause of death worldwide and represents a significant economic burden. Atherosclerosis, which is characterized by a build-up of inflammatory lipids and immune cells, is the most common underlying cause of CVD. Early in the pathogenesis of atherosclerosis, LDL-cholesterol particles infiltrate the lining of the artery wall, are modified (e.g., oxidized), and activate inflammatory pathways (e.g., NLRP3 inflammasome) triggering the production of pro-inflammatory cytokines. A failure to resolve inflammation within the artery wall leads to the development of a plaque. The current standard of care includes lipid-lowering drugs such as statins. However, even with successful lipid-lowering, there remains a subset of patients with residual inflammatory risk. These observations led to several large-scale clinical trials (e.g., CANTOS and Lo- DoCo2) that targeted systemic inflammation to treat atherosclerotic CVD. These trials demonstrated that target- ing systemic inflammation reduces adverse cardiovascular events, however, due to an increase in non-cardio- vascular-related deaths, there are currently no FDA-approved anti-inflammatory therapies for atherosclerosis. While targeting inflammation systemically has shown promise, it is important to identify more specific targets and regulators of inflammation within the atherosclerotic plaque to leverage into novel therapies. Research over the past few decades has established that the metabolic state of the immune cell determines its inflammatory ca- pacity. As such, rather than targeting systemic inflammation, we propose to leverage the metabolic regulators of inflammation within the cell. Studies of macrophages stimulated with inflammatory mediators including microbial ligands and cytokines have revealed significant upregulation of the gene Immune-responsive gene 1 (Irg1, also Acod1) and its downstream metabolite itaconate. In addition to being an anti-microbial agent, itaconate controls immune activation and elicits immunological tolerance through (1) regulation of immune cell metabolism, (2) inhibition of late-stage NLRP3 inflammasome activity, and (3) activation of the anti-inflammatory and anti-oxida- tive transcription factors Nuclear factor erythroid 2-related factor 2 and Activating transcription factor 3. While Irg1 and itaconate have been studied in the context of acute inflammation, their role in chronic inflammatory disease, such as atherosclerosis, has not been explored despite sharing many similar pathways. In this proposal, I will define the role of Irg1 and itaconate on immune cell regulation in atherosclerosis, specifi- cally the effects on three major immune cells within the plaque we have found to robustly express Irg1: mono- cytes, macrophages, and neutrophils. Additionally, I will utilize in vivo mouse models of atherosclerosis to test whether IRG1 deficiency exacerbates plaque burden and inflammation. Finally, I will elucidate the utility of 4-OI, a cell-permeable itaconate derivative as a treatment in atherosclerotic CVD. If successful, this will demonstrate for the first time the role of Irg1 and itaconate in chronic inflammation and identify a novel treatment modality for the inflammatory component of atherosclerotic CVD.