Immune cells undergo rapid and extensive metabolic changes during inflammation. In addition to meeting the energetic and biosynthetic demands, metabolites can also function in cell signaling. Itaconate rapidly accumulates to high levels in myeloid cells under infectious and non-infectious conditions, and it binds to and regulates the function of diverse proteins to influence metabolism, oxidative response, epigenetic modification and gene expression intrinsically, and to signal GPCR after secretion. Administration of itaconate protects against inflammatory diseases and blocking itaconate production enhances antitumor immunity in preclinical models. In this Review, we revisit itaconate metabolism and its regulation, discuss its target proteins and mechanisms, and provide a rationale for developing itaconate-based therapeutics to treat inflammatory diseases and targeting itaconate production for cancer immunotherapy.