ABSTRACT
By the screening of a combinatorial library for inhibitors of nitric oxide (NO) formation by the inducible isoform of nitric oxide synthase (iNOS) using a whole-cell assay, 2-(imidazol-1-yl)pyrimidines were identified. Compounds were found to inhibit the dimerization of iNOS monomers, thus preventing the formation of the dimeric, active form of the enzyme. Optimization led to the selection of the potent, selective, and orally available iNOS dimerization inhibitor, 21b, which significantly ameliorated adjuvant-induced arthritis in a rat model. Analysis of the crystal structure of the 21b--iNOS monomer complex provided a rationalization for both the SAR and the mechanism by which 21b blocks the formation of the protein--protein interaction present in the dimeric form of iNOS.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/chemical synthesis , Benzodioxoles/chemical synthesis , Imidazoles/chemical synthesis , Nitric Oxide Synthase Type II/metabolism , Pyrimidines/chemical synthesis , Animals , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Arthritis, Experimental/therapy , Benzodioxoles/chemistry , Benzodioxoles/pharmacology , Cell Line , Chlorocebus aethiops , Crystallography, X-Ray , Dimerization , Imidazoles/chemistry , Imidazoles/pharmacology , Male , Models, Molecular , Pyrimidines/chemistry , Pyrimidines/pharmacology , Rats , Rats, Inbred LewABSTRACT
Compound 1 was identified by high throughput screening as a novel, potent, non-amidine factor Xa inhibitor with good selectivity against thrombin and trypsin. A series of modifications of the three aromatic groups of 1 was investigated. Substitution of chlorine or bromine for fluorine on the aniline ring led to the discovery of subnanomolar factor Xa inhibitors. Positions on the anthranilic acid ring that can accommodate further substitution were also identified.
Subject(s)
Factor Xa Inhibitors , Thiophenes/pharmacology , ortho-Aminobenzoates/pharmacology , Animals , Anticoagulants/chemical synthesis , Anticoagulants/pharmacology , Cattle , Heterocyclic Compounds/pharmacology , Humans , Indicators and Reagents , Kinetics , Prothrombin Time , Serine Proteinase Inhibitors/pharmacology , Structure-Activity Relationship , Thiophenes/chemistry , Thrombin/antagonists & inhibitors , Trypsin Inhibitors/chemical synthesis , Trypsin Inhibitors/pharmacology , ortho-Aminobenzoates/chemistryABSTRACT
A novel series of diaryloxypyridines have been designed as selective nanomolar factor Xa (fXa) inhibitors for use as anticoagulants. In this paper, we describe our efforts to identify an additional interaction and a replacement for the distal amidine group that binds in the S3/S4 pocket of fXa. Introduction of a hydroxyl group para to the proximal amidine group increases the potency vs fXa by 1-2 orders of magnitude, which is the result of a hydrogen bond to Ser195 of the catalytic triad. A methyl imidazoline and a dimethylamide are good alternatives for the second amidine. These substitutions have increased the selectivity vs the related serine proteases trypsin and thrombin. The synthesis, in vitro activity, and hypothetical modes of binding to fXa based on trypsin crystallographic data are outlined.
