Suppression of NF-κB activity by sulfasalazine is mediated by direct inhibition of IκB kinases α and β

Abstract

Background & Aims: Activation of NF-KB/Rel has been implicated in the pathogenesis of inflammatory bowel disease (IBD). Various drugs used in the treatment of IBD, such as glucocorticoids, 5-aminosalicylic acid, and sulfasalazine, interfere with NF-KB/Rel signaling. The aim of this study was to define the molecular mechanism by which sulfasalazine inhibits NF-KB activation. Methods: The effects of sulfasalazine and its moieties on NF-KB signaling were evaluated using electromobility shift, transfection, and immune complex kinase assays. The direct effect of sulfasalazine on IKB kinase (IKK) activity was investigated using purified recombinant IKK-a and - proteins. Results: NF-KB/Rel activity induced by tumor necrosis factor a, 12-0-tetradecanoylphorbol-13-acetate, or overexpression of NF-KB-inducing kinase, IKK-a, IKK-, or constitutively active IKK-a and IKK- mutants was inhibited dose dependently by sulfasalazine. Sulfasalazine inhibited tumor necrosis factor -induced activation of endogenous IKK in Jurkat T cells and SW620 colon cells, as well as the catalytic activity of purified IKK-a and IKK- in vitro. In contrast, the moieties of suifasalazine, 5-aminosalicylic acid, and sulfapyridine or 4-aminosalicylic acid had no effect. Activation of extracellular signal-related kinase (ERK) 1 and 2, c-Jun-N-terminal kinase (JNK) 1, and p38 was unaffected by sulfasalazine. The decrease in substrate phosphorylation by IKK-a and - is associated with a decrease in autophosphorylation of IKKs and can be antagonized by excess adenosine triphosphate. Conclusions: These data identify sulfasalazine as a direct inhibitor of IKK-a and - by antagonizing adenosine triphosphate binding. The suppression of NF-KB activation by inhibition of the IKKs contributes to the wellknown anti-inflammatory and immunosuppressive effects of sulfasalazine. © 2000 by the American Gastroenterological Association.