In vitro and in vivo characterization of a neutral boron-containing thrombin inhibitor

• Published in: The Journal of biological chemistry 1993-03, Vol.268 (7), p.4734-4741
• Main Authors: TAPPARELLI, C, METTERNICH, R, EHRHARDT, C, ZURINI, M, CLAESON, G, SCULLY, M. F, STONE, S. R
• Format: Article
• Language: English
• Subjects: activity; Analytical, structural and metabolic biochemistry; Animals; Antithrombins - chemistry; Antithrombins - pharmacology; Arginine - analogs & derivatives; Biological and medical sciences; Blood Coagulation Tests; boron; Boron - analysis; Bridged Bicyclo Compounds - chemistry; Bridged Bicyclo Compounds - pharmacology; Cells, Cultured; characterization; containing; Dipeptides - chemistry; Dipeptides - pharmacology; Enzymes and enzyme inhibitors; Fundamental and applied biological sciences. Psychology; Hirudins - chemistry; Hirudins - pharmacology; Humans; Hydrolases; inhibitors; Kinetics; Male; Oligopeptides - chemistry; Oligopeptides - pharmacology; Pipecolic Acids - chemistry; Pipecolic Acids - pharmacology; Piperidines - chemistry; Piperidines - pharmacology; Platelet Activation - drug effects; Platelet Aggregation - drug effects; Rats; Rats, Wistar; Sulfonamides; thrombin; Thrombin - pharmacology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)53458-X

Peptide boronic acid derivatives have proven to be very potent inhibitors of serine proteases with boroarginine derivatives being particularly potent thrombin inhibitors. The importance of the charged side chain of arginine has been investigated by synthesizing a derivative in which this side chain has been replaced by a neutral one. This boronic acid derivative, D-benzyloxycarbonyl (Z)-Phe-Pro-methoxypropylglycine-pinanediol (MpgC10H16), inhibited thrombin by a competitive mechanism with an inhibition constant (Ki) of 8.9 nM. In comparison to boroarginine derivatives, Z-D-Phe-Pro-boroMpgC10H16 displayed higher selectivity for thrombin over trypsin (Ki = 1.1 microM) and plasmin (Ki = 15.7 microM). Prolongation of thrombin time and activated partial thromboplastin time were observed with micromolar concentrations of Z-D-Phe-Pro-boroMpgC10H16. In a thrombin-dependent in vitro aggregation assay with human platelets, Z-D-Phe-Pro-boroMpgC10H16 inhibited aggregation with an IC50 of 85 nM. When tested in a thrombin-dependent platelet accumulation model in the rat, a bolus injection of (Z)-D-Phe-Pro-boroMpgC10H16 (0.3-3 mg/kg) inhibited platelet accumulation. Thus, the substitution of the charged guanidino group in the P1 side chain by the neutral methoxy group resulted in a potent and highly selective thrombin inhibitor with an interesting pharmacological profile with in vitro as well as in vivo models.