Itaconate and fumarate derivatives exert a dual inhibitory effect on canonical NLRP3 activation in macrophages and microglia

Abstract

The NLRP3 inflammasome is a multi-protein complex that regulates the protease caspase-1 and subsequent interleukin (IL)-1{beta} release from cells of the innate immune system, or microglia in the brain, in response to infection or injury. Derivatives of the metabolites itaconate and fumarate, dimethyl itaconate (DMI), 4-octyl itaconate (4OI) and dimethyl fumarate (DMF), limit both expression of IL-1{beta}, and IL-1{beta} release following NLRP3 inflammasome activation. However, the direct effects of these metabolite derivatives on NLRP3 inflammasome responses in macrophages and microglia require further investigation. Using murine bone marrow-derived macrophages, mixed glia and organotypic hippocampal slice cultures (OHSCs), we demonstrate that DMI and 4OI pre-treatment limited IL-1{beta}, IL-6 and tumor necrosis factor production in response to lipopolysaccharide (LPS) priming, as well as inhibiting subsequent NLRP3 inflammasome activation. DMI, 4OI, DMF and monomethyl fumarate (MMF), another fumarate derivative, also directly inhibited biochemical markers of NLRP3 activation in LPS-primed macrophages, mixed glia and OHSCs, including ASC speck formation, caspase-1 activation, gasdermin D cleavage and IL-1{beta} release. Finally, DMF, an approved treatment for multiple sclerosis, as well as DMI, 4OI and MMF, inhibited NLRP3 activation in macrophages in response to the phospholipid lysophosphatidylcholine, which is used to induce demyelination, suggesting a possible mechanism of action for DMF in multiple sclerosis through NLRP3 inhibition. Together, these findings reveal the importance of immunometabolic regulation for both the priming and activation steps of NLRP3 activation in macrophages and microglia. Furthermore, we highlight itaconate and fumarate derivatives as a potential therapeutic option in NLRP3-driven diseases, including in the brain. Summary statementWe show that itaconate and fumarate derivatives inhibit both the priming and activation steps of NLRP3 inflammasome responses in macrophages and microglia, revealing the importance of immunometabolic NLRP3 regulation.