NS-398 (N-(2-cyclohexyloxy-4-nitrophenyl)methane sulfonamide) is a COX-2 inhibitor wih anti-inflammatory activity.

COX-2:3.8 μM

NS-398 inhibits COX-2 enzyme activity in a concentration dependent manner, the IC50 being 3.8 μM, whereas NS-398 at 100 μM has no effect on COX-1 activity[1]. At 10 μM, NS-398 treatment results in increased production of COX-2 and the pro-inflammatory cytokine. NS-398 (10 μM) induces apoptosis in LNCaP cells, but not in the more aggressive, androgen-unresponsive C4-2b cells. The C4-2b cells are observed to continue to proliferate when treated with NS-398 and continues to retain malignant phenotype characteristics. NS-398 treatment results in C4-2b cell differentiation into an unusual neuroendocrinelike cell. These neuroendocrine-like cells produces both epithelial (cytokeratin 18 and prostate specific antigen) and neuronal (neuron-specific enolase and chromogranin A) proteins. Furthermore, this C4-2b cellular response to NS-398 is mediated by NF-kB transcription factor activation. NS-398 induces NF-kB and down-regulates Ikβ-α protein expression in LNCaP C4-2b cells[2].

NS398 could inhibit Cox-2 expression induced by acoustic injury and could attenuate noise-induced hearing threshold shifts and cochlear hair cell loss. The inhibition of Cox-2 by NS398 could attenuate Noise-induced hearing loss(NIHL)and related hair cell damage.[3].

1×106 cells are plated in six well cluster plates with 2 ml of medium for 24 h. At time point 0, medium was removed, cells were carefully washed with phosphate buffered saline (PBS) and serum free(SF), phenol-red free medium containing 0.5 μg/ml BSA was added. Incubations are continued for an additional 72 h with or without increasing NS-398 concentrations. NS-398 stocks (5 mM) are dissolved in 0.1% dimethylsulfoxide (DMSO). Cells and culture medium are harvested at 24 h time intervals. Dead cells are removed by gentle washing with PBS and cell number determined by direct counting using trypan blue dye exclusion to identify viable cells. DMSO (0.1%) is added to control cultures.(Only for Reference)