Suppression of NFAT5-mediated inflammation and chronic arthritis by novel κB-binding inhibitors

Abstract

Abstract Nuclear factor of activated T cells 5 (NFAT5), also known as the tonicity-responsive enhancer-binding protein, is a transcription factor whose DNA binding domain shares structural homology with NF-kB and other members of the NFAT family. Evidence is emerging that NFAT5 is crucial to the pathogenesis of various human diseases, including cancer and arthritis. However, therapeutic agents specifically inhibiting NFAT5 activity are currently unavailable. To discover NFAT5 inhibitors, we screened a library of >40,000 chemicals for the suppression of nitric oxide, a NFAT5 target, through high-throughput screening. We validated anti-NFAT5 activity of the primary hit compounds using an NFAT5-specific reporter and identified the novel NFAT5 inhibitor KRN2, 13-(2-fluoro)-benzyl-berberine and its oral derivative, KRN5. KRN2 suppressed the transcriptional activation of NFAT5 by blocking NF-κB binding to the NFAT5 promoter region, inhibiting NFAT5 mRNA and protein expression in macrophages stimulated with lipopolysaccharide. Interestingly, KRN2 selectively inhibited the expression of pro-inflammatory genes, including Nos2, Tnf, and Il6 without hampering high-salt-induced NFAT5 and its target gene expressions. Moreover, KRN2 and KRN5 ameliorated experimentally induced arthritis in mice, decreasing pro-inflammatory cytokine production. Particularly, orally administered KRN5 was more potent in suppressing arthritis than methotrexate, a commonly used anti-rheumatic drug. Therefore, KRN2 and KRN5 can be potential therapeutic agents in the treatment of chronic arthritis.