Dissociated glucocorticoids with anti-inflammatory potential repress interleukin-6 gene expression by a nuclear factor-kappa B-dependent mechanism

Synthetic glucocorticoids (GCs) remain among the most effective agents for the management of chronic inflammatory diseases. However, major side effect s severely limit their therapeutic use. Physiologic and therapeutic activit ies of GCs are mediated by a nuclear receptor belonging to a superfamily of ligand-inducible transcription factors that, in addition to directly regul ating their cognate gene programs, can also mutually interfere with other s ignaling pathways. We recently identified selective ligands of the glucocor ticoid receptor that dissociate transactivation from activator protein 1 tr ansrepression, and most importantly retain in vivo anti-inflammatory activi ty. To further document the mechanisms of action sustaining the observed in vivo activity, we report here on the interference of dissociated GCs with nuclear factor kappa B (NF-kappa B)-driven gene activation. We show that di ssociated GCs repress tumor necrosis factor-induced interleukin-6 gene expr ession by an NF-kappa B-dependent mechanism, without changing the expressio n level of inhibitor kappa B. The DNA-binding activity of induced NF-kappa B also remained unchanged after stimulation of cells with the various compo unds. Evidence for a direct nuclear mechanism of action was obtained by ana lysis of cell lines constitutively expressing a fusion protein between the DNA-binding domain of the yeast Gal4 protein and the transactivating p65 su bunit of NF-kappa B, which was able to efficiently repress a Gal4-dependent luciferase reporter gene upon addition of the dissociated compounds. We th erefore conclude that, in addition to dissociating transactivation from act ivator protein 1 transrepression, dissociated GCs mediate inhibition of NF- kappa B signaling by a mechanism that is independent of inhibitor kappa B i nduction.