ABSTRACT
A series of indole and carbazole based inhibitors of factor Xa (FXa) has been investigated. The most potent compound inhibits FXa with a Ki of 0.2 nM and has 900- and 750-fold selectivity over thrombin and trypsin, respectively.
Subject(s)
Amidines/chemistry , Anticoagulants/chemical synthesis , Anticoagulants/pharmacology , Carbazoles/chemical synthesis , Carbazoles/pharmacology , Factor Xa Inhibitors , Indoles/chemical synthesis , Indoles/pharmacology , Pyridines/chemistry , Carbazoles/chemistry , Chemical Phenomena , Chemistry, Physical , Crystallography, X-Ray , Humans , Indoles/chemistry , Models, Molecular , Molecular Conformation , Structure-Activity Relationship , Thrombin/antagonists & inhibitors , Trypsin Inhibitors/chemical synthesis , Trypsin Inhibitors/pharmacologyABSTRACT
Inhibitors based on the benzimidazole scaffold showed subnanomolar potency against Factor Xa with 500-1000-fold selectivity against thrombin and 50-100-fold selectivity against trypsin. The 2-substituent on the benzimidazole ring had a strong impact on the FXa inhibitory activity. Crystallography studies suggest that the 2-substituent may have a conformational effect favoring the extended binding conformation.
Subject(s)
Anticoagulants/chemical synthesis , Anticoagulants/pharmacology , Benzimidazoles/chemical synthesis , Benzimidazoles/pharmacology , Factor Xa Inhibitors , Benzimidazoles/chemistry , Chemical Phenomena , Chemistry, Physical , Crystallography, X-Ray , Humans , Models, Molecular , Molecular Conformation , Structure-Activity Relationship , Thrombin/antagonists & inhibitors , Trypsin Inhibitors/chemical synthesis , Trypsin Inhibitors/pharmacologyABSTRACT
Double rotational-echo double resonance (double REDOR) has been used to investigate the bound conformations of (13)C,(15)N,(19)F-labeled factor Xa inhibitors to bovine trypsin. Carbon-fluorine dipolar couplings were measured by (13)C{(19)F} REDOR with natural-abundance background interferences removed by (13)C{(15)N} REDOR. The conformations of the bound inhibitors were characterized by molecular dynamics (MD) simulations of binding restrained by double REDOR-determined intramolecular C-F distances. A symmetrical bisamidine inhibitor and an asymmetrical monoamidine-monoamine inhibitor of the same general shape had distinctly different conformations in the bound state. According to the MD models, these differences arise from specific interactions of the amidine and amine groups with the active-site residues of trypsin and nearby water molecules.
Subject(s)
Factor Xa/chemistry , Magnetic Resonance Spectroscopy/methods , Animals , Cattle , Factor Xa/metabolism , Ligands , Macromolecular Substances , Models, Molecular , Protein Binding , Protein Conformation , Structure-Activity Relationship , Trypsin/metabolismABSTRACT
Factor Xa is a serine protease which activates thrombin (factor IIa) and plays a key regulatory role in the blood-coagulation cascade. Factor Xa is, therefore, an important target for the design of anti-thrombotics. Both factor Xa and thrombin share sequence and structural homology with trypsin. As part of a factor Xa inhibitor-design program, a number of factor Xa inhibitors were crystallographically studied complexed to bovine trypsin. The structures of one diaryl benzimidazole, one diaryl carbazole and three diaryloxypyridines are described. All five compounds bind to trypsin in an extended conformation, with an amidinoaryl group in the S1 pocket and a second basic/hydrophobic moiety bound in the S4 pocket. These binding modes all bear a resemblance to the reported binding mode of DX-9065a in bovine trypsin and human factor Xa.
