Inhibitory effect of T-614 on tumor necrosis factor-alpha induced cytokine production and nuclear factor-kappaB activation in cultured human synovial cells.

Abstract

To investigate the mechanism of the immunosuppressive effect of T-614 [N-(3-formylamino-4-oxo-6-phenoxy-4H-chromen-7-yl)methanesulfonamide], a new antirheumatic drug whose clinical efficacy has been determined for the treatment of patients with rheumatoid arthritis (RA). RA synovial fibroblast-like cells were cultured with tumor necrosis factor-alpha (TNF-alpha, 10 ng/ml) in the presence or absence of T-614. After incubation, cytokine production was measured by ELISA. Expression of interleukin 6 (IL-6) and IL-8 mRNA was examined by real-time quantitative reverse transcriptase-polymerase chain reaction analysis and TNF-alpha induced nuclear factor-kappaB (NF-kappaB) activation was observed using immunostaining with an antibody against NF-kappaB p65. T-614 suppressed TNF-alpha induced production of IL-6, IL-8, and monocyte chemoattractant protein 1, and also reduced the accumulation of IL-6 and IL-8 mRNA in a concentration dependent manner. T-614 interfered with the TNF-alpha induced translocation of NF-kappaB to the nucleus from the cytoplasm. Inhibition of NF-kappaB activation and transcription of proinflammatory cytokines by T-614 contributes to its clinical antirheumatic effect.