Vol. 282, Issue 1, 459-466, 1997

The Natural Product Hymenialdisine Inhibits Interleukin-8 Production in U937 Cells by Inhibition of Nuclear Factor-B

John J. Breton and Marie C. Chabot-Fletcher

Department of Immunopharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania

The nuclear factor-B (NF-B) family of transcription factors have been implicated in the inducible expression of genes involved in inflammatory and immune responses. As such, a specific inhibitor of NF-B would be a useful therapeutic agent in a variety of inflammatory disorders. The marine natural product hymenialdisine was evaluated as an inhibitor of NF-B in U937 cells. U937 cells were transfected with either a luciferase reporter plasmid containing the human immunodeficiency virus long terminal repeat or the interleukin-8 (IL-8) core promoter, both of which are activated by NF-B. Hymenialdisine caused a concentration-dependent decrease in luciferase production from both reporters when the cells were stimulated with tumor necrosis factor-, lipopolysaccharide or phorbol myristate acetate. An electrophoretic mobility shift assay confirmed its activity by inhibiting DNA binding of NF-B. Hymenialdisine was shown to be a selective inhibitor of NF-B in that it had no effect on the binding of other transcription factors to their DNA concensus motifs; these included activator protein-1, CCAAT/enhancer binding protein and Sp1. Functional studies showed hymenialdisine to be an inhibitor of IL-8 production and IL-8 mRNA formation in the U937 cell. Investigation into the mechanism of action of hymenialdisine showed that it was not due to inhibition of protein kinase C because the selective protein kinase C inhibitor RO 32-0432 was inactive against tumor necrosis factor--stimulated luciferase and IL-8 production. The compound also had no effect on IB or IB phosphorylation and degradation. Thus, hymenialdisine is a potent inhibitor of NF-B and IL-8 production in U937 cells.