ABSTRACT
Nuclear factor-kappaB (NF-kappaB) has been considered as a good target for the treatment of many diseases. Although a lot of NF-kappaB inhibitors have already been reported, many of them have several common problems. Thus, we attempted to identify novel NF-kappaB inhibitors to be unique lead compounds for creating new pharmaceuticals. In the present study, we screened our chemical library for compounds that directly inhibit the DNA binding of NF-kappaB by using fluorescence correlation spectroscopy (FCS). Consequently, we identified a promising compound, 4,6-dichloro-N-phenyl-1,3,5-triazin-2-amine, referred to as NI241. It mediated a dose-dependent inhibition of the DNA binding of NF-kappaB p50. Its analogues also showed dose-dependent inhibition and their inhibitory effects were altered by the substituents on the N-phenyl group. Furthermore, we predicted the binding mode of NI241 with p50 in silico. In this model, NI241 forms three hydrogen bonds with Tyr60, His144, and Asp242 on p50, which are important amino acid residues for the interaction with DNA. These results suggest that NI241 with structural novelty may serve as a useful scaffold for the creation of new NF-kappaB inhibitors by rational optimization.
Subject(s)
Aniline Compounds/chemistry , Aniline Compounds/pharmacology , DNA/metabolism , NF-kappa B/antagonists & inhibitors , NF-kappa B/metabolism , Triazines/chemistry , Triazines/pharmacology , Animals , Binding Sites , Computer Simulation , DNA/chemistry , Mice , Models, Molecular , Molecular Structure , NF-kappa B/chemistry , NF-kappa B p50 Subunit/antagonists & inhibitors , NF-kappa B p50 Subunit/chemistry , NF-kappa B p50 Subunit/metabolism , Protein Binding , Structure-Activity RelationshipABSTRACT
Alveolar epithelial cell death plays a crucial role in the progression of acute lung injury. We have demonstrated up-regulation of Fas expression on alveolar epithelial cells, and soluble Fas ligand secretion from inflammatory cells upon acute lung injury. Here we show that the lipopolysaccharide-stimulated human monocyte cell line THP-1 releases Fas ligand, and that conditioned medium from lipopolysaccharide-stimulated THP-1 cells induces apoptosis of the human pulmonary adenocarcinoma cell line A549. Activation of caspase-3 and -8 is associated with the apoptosis. Gene targeting on Fas in A549 cells by specific small interfering RNA impairs apoptosis induced by conditioned medium from activated THP-1, while that on Fas ligand in THP-1 cells impairs the apoptosis-inducing activity of the conditioned medium produced by lipopolysaccharide-stimulated cells. These results suggest that Fas ligand released by monocytes causes alveolar epithelial cell death through a Fas-dependent apoptotic mechanism in the development of acute lung injury.
