Metabolism-Directed Optimization of 3-Aminopyrazinone Acetamide Thrombin Inhibitors. Development of an Orally Bioavailable Series Containing P1 and P3 Pyridines

Abstract

Recent efforts in the field of thrombin inhibitor research have focused on the identification of compounds with good oral bioavailability and pharmacokinetics. In this manuscript we describe a metabolism-based approach to the optimization of the 3-(2-phenethylamino)-6-methylpyrazinone acetamide template (e.g., 1) which resulted in the modification of each of the three principal components (i.e., P1, P2, P3) comprising this series. As a result of these studies, several potent thrombin inhibitors (e.g., 20, 24, 25) were identified which exhibit high levels of oral bioavailability and long plasma half-lives.

BIOACTIVATION OF THE 3-AMINO-6-CHLOROPYRAZINONE RING IN A THROMBIN INHIBITOR LEADS TO NOVEL DIHYDRO-IMIDAZOLE AND IMIDAZOLIDINE DERIVATIVES: STRUCTURES AND MECHANISM USING 13C-LABELS, MASS SPECTROMETRY, AND NMR

“…Both I-B and I-C were prepared at Merck Research Laboratories (Labeled Compound Synthesis group, Rahway, NJ) and were determined to have a purity of Ͼ99% by NMR. (Burgey et al, 2003b) of 5 mg/kg (10 mg/ml in DMSO) to bile duct-cannulated male Sprague-Dawley rats (n ϭ 3). Specimens of bile and urine were collected over ice at different time points for 48 h postdosing to quantify the in vivo metabolites.…”

“…For metabolite profiling, [ 14 C]-labeled I was administered i.v. at a pharmacologically relevant dose (Burgey et al, 2003b) of 5 mg/kg (10 mg/ml in DMSO) to bile duct-cannulated male Sprague-Dawley rats (n ϭ 3). Specimens of bile and urine were collected over ice at different time points for 48 h postdosing to quantify the in vivo metabolites.…”

“…Both therapeutic agents suffer from significant limitations: low-molecular-weight heparin requires either continuous intravenous infusion or subcutaneous injection, and warfarin has a delayed onset of action, narrow therapeutic index, and a high potential for drug-drug interactions. Hence, a small-molecule thrombin inhibitor represents an alternative target in the search for orally administered anticoagulants that could offer a safer alternative to existing therapies for the treatment and prevention of thromboembolic disorders (Vacca, 2000;Burgey et al, 2003b).is a potent, selective and orally bioavailable thrombin inhibitor from the amino-pyrazinone acetamide series (Burgey et al., 2003a,b). In this study, we provide details of the in vivo biotransformation of I in rats.…”

“…Triethylamine (77 ml, 550 mmol) was added to a suspension of glycine HCl salt (38.4 g, 276 mmol) in methylene chloride (300 ml), and the mixture was stirred at room temperature for 1 h. Ethyl chlorooxoacetate (30 ml, 275 mmol) was then introduced, and the resulting mixture was stirred at room temperature for 18 h. Water (25 ml) was added and layers were separated. Organic layer was dried over MgSO 4 (5 g), filtered, and concentrated to afford ethyl N-(ethylcarboxymethyl)oxamate (42.2 g, 75%).A solution of ethyl N-(ethylcarboxymethyl)oxamate (203 mg, 1 mmol), 2-(2-amino-1,1-difluoroethyl)pyridine (316 mg, 1 mmol) (Burgey et al, 2003b), triethylamine (0.5 ml), and 2-propanol (3 ml) was stirred at room temperature for 18 h then purified by flash chromatography on silica gel (30% ethyl acetate in hexane) to give a white solid (110 mg, 35%), which was dissolved in methanol (5 ml) and 1 M NaOH solution (5 ml). After stirring at room temperature for 2 h, solvent was removed under vacuum to give a light yellow solid.…”

“…Dimethylformamide (2 ml) was added and a solution was formed. To this solution, 1,2-dichloroethane (191 mg, 1 mmol), 1-hydroxybenzotriazole hydrate (159 mg, 1 mmol), triethylamine (1 ml), and 2-(aminomethyl)-3-fluoropyridine (50 mg, 0.25 mmol) (Burgey et al, 2003b) were added, and the mixture was stirred at room temperature for 16 h. Reverse-phase HPLC purification (Zorbax RX C18, MeCN:water:trifluoroacetic acid ϭ 12:88:0.1) yielded M3 (66 mg, 66%) as a white solid. …”