Testing the Specificity of Compounds Designed to Inhibit CPT1A in T Cells

In response to antigen and costimulation, T cells undergo a series of metabolic transitions that fulfill the biosynthetic demands of clonal expansion, differentiation, and effector function. Following antigen clearance, the oxidation of long-chain fatty acids (LCFAO) has been implicated in the transition from effector to central memory T cells. However, studies demonstrating a role for LCFAO in memory T-cell development have largely relied on the use of etomoxir (ETO), a small molecule inhibitor of the long-chain fatty acid transporter CPT1A. Understanding how the depletion of nutrients including LCFA that might occur in tumor microenvironments affects T-cell proliferation, differentiation, and function has important implications for tumor immunotherapy. Here, we combine the analysis of posttranscriptional gene silencing with extracellular flux assays to determine if etomoxir exerts nonspecific effects on oxidative metabolism. The off-target effects of ETO that we describe highlight the challenges of using pharmacologic inhibitors in loss-of-function approaches in T cells.