Immunosuppressive Leflunomide Metabolite (A77 1726) Blocks TNF-Dependent Nuclear Factor-κB Activation and Gene Expression

Abstract Leflunomide is a novel immunosuppressive and antiinflammatory agent currently being tested for treatment of autoimmune diseases and transplant rejection. NF-κB is a transcription factor activated in response to a wide variety of inflammatory stimuli, including TNF, but whether leflunomide blocks NF-κB activation is not known. In the present report we demonstrate that treatment of a human T cell line (Jurkat) with leflunomide blocks TNF-mediated NF-κB activation in a dose- and time-dependent manner, with maximum inhibition at 5–10 μM. Inhibition was not restricted to TNF-induced activation, because leflunomide also inhibited NF-κB activation induced by other inflammatory agents, including phorbol ester, LPS, H2O2, okadaic acid, and ceramide. Leflunomide blocked the degradation of IκBα and subsequent nuclear translocation of the p65 subunit, steps essential for NF-κB activation. This correlated with inhibition of dual specificity-mitogen-activated protein kinase kinase as well as an Src protein tyrosine kinase, p56lck, by leflunomide. Reducing agents did not reverse the effect of leflunomide. Leflunomide also suppressed the TNF-activated NF-κB-dependent reporter gene expression. Our results thus indicate that leflunomide is a potent inhibitor of NF-κB activation induced by a wide variety of inflammatory stimuli, and this provides the molecular basis for its anti-inflammatory and immunosuppressive effects.