Synthesis, characterization, and Ca2+ antagonistic activity of diltiazem metabolites.

Diltiazem is a calcium antagonist widely used in the treatment of angina and hypertension. The contributions of metabolites of diltiazem to the vasorelaxant effects of diltiazem were investigated. The synthesis and spectroscopic characterization of eight major cis-diltiazem metabolites are described. Three of the compounds--N, O-didemethylated metabolite (21), O-demethylated metabolite (22), and diltiazem N-oxide (27)--have been recently reported and have not previously been synthesized. The identities of all eight synthetic metabolites have been verified with samples obtained from human urine using combined LC-MS/MS. The Ca2+ antagonistic activities of diltiazem and its metabolites (except 27) were studied on hamster aorta preparations depolarized with KCl. The order of potencies (IC50 +/- SE, microM) is as follows: diltiazem (0.98 +/- 0.47) greater than 17 (2.46 +/- 0.38) greater than or equal to 23 (3.27 +/- 1.02) greater than 26 (20.2 +/- 10.5) greater than 22 (40.4 +/- 15.4) greater than or equal to 25 (45.5 +/- 18.1) greater than 21 (112.2 +/- 33.2) greater than or equal to 24 (126.7 +/- 24.2). Structure-activity relationships are also discussed.

Erythrocyte adenosine transport. A rapid screening test for cardiovascular drugs.

An erythrocyte (RBC) model based on whole blood was used to investigate the effect of cardiovascular drugs on the uptake of adenosine in vitro. Fresh whole blood obtained from healthy volunteers was allowed to equilibrate with various concentrations (5-1000 microM) of a tested agent. (2-3H)-Adenosine was used as a substrate, and the reaction was terminated after 2 sec of incubation at room temperature by rapid addition of a "Stopping Solution" which was a mixture of erythro-9-(2-hydroxy-3-nonyl)adenine, dipyridamole, and EDTA. The mixture was centrifuged (1760 g, 4 degrees C, 10 min), and the radioactivity of an aliquot of the supernatant was determined by a scintillation counter. The results showed that dipyridamole was the most potent agent tested (IC50 = 0.2 microM). Amongst the calcium antagonists studied, isradipine was most potent, followed by verapamil, clentiazem, diltiazem, and then nifedipine. The racemates of two metabolites of diltiazem, MX and MB, were more potent than the parent drug. The antiarrhythmic agents, amiodarone and sotalol, the two new lipid peroxidation inhibitors, U-74389F and U-78517F, and the anxiolytic agent, alprazolam, were as active as verapamil. The beta-receptor antagonist propranolol and the angiotensin converting enzyme (ACE) inhibitor, enalapril, were practically inactive. In addition, the model was stereoselective such that the S(-)-enantiomer of verapamil was considerably more potent than the R(+)-antipote, whereas d(+)-sotalol was practically inactive compared to racemic sotalol.

Functional modification of indole binding site with indomethacin congeners.

Indomethacin and four congeners were shown by equilibrium dialysis to interfere with the binding of L-tryptophan at the primary indole binding site on defatted human serum albumin. Incubation of albumin with 1-(p-bromoacetamidobenzovl)- and 1-(m-bromoacetamidobenzoyl)-5-methoxy-2-methyl-3-indolylacetic acid functionally modified this site, decreasing the availability of the site for the binding of L-tryptophan. Incubation in the presence of a large excess of L-tryptophan resulted in the partial protection of the site from modification by 1-(m-bromoacetamidobenzoyl)-5-methoxy-2-methyl-3-indolylacetic acid.

A partially rigid butyrophenone analog with extended conformation.

The synthesis and in vitro dopamine D2 receptor binding of a conformationally restricted analog of 4-piperidinobutyrophenone is described. 2-(2-Piperidinoethyl)-1-tetralone displaces [3H]spiperone from the rat striatal dopamine receptor with an IC50 of 1.2 X 10(-4)mol/l.

2-substituted tetralin derivatives as conformationally restricted butyrophenone analogs.

Five related derivatives of 2-(2-piperidinoethyl)tetralin have been synthesized as partially rigid analogs of the neuroleptic butyrophenones. Their in vitro dopamine receptor binding properties and in vivo inhibition of dexamphetamine-induced locomotor activity are described. 1-Phenyl-2-(2-piperidinoethyl)-3,4-dihydronaphthalene hydrochloride (8) is about 1% as potent as haloperidol.

Dibenzothiazepinones as potential calcium channel antagonists. I.

Certain dibenzo[a,d]cycloheptenes and dibenzo[b,e]thiepins (e.g., 1 and 2) substituted in the 5 and 11-positions, respectively, are potent calcium channel antagonists. It has previously been shown that potency increases with a decrease in the angle between the planes of the flanking aromatic rings, as computed by semi-empirical AM1 calculations with geometry minimization. In the present study, we evaluated a series of dibenzo[b,f]-1,4-thiazepinones (10-14) as potential bioisosteres of these compounds having a slightly more acute angle of flexure. X-Ray diffraction reveals the crystal structure of the parent dibenzo[b,f]-1,4-thiazepinone 4 to have a flexure angle of 108.4 degrees. Due to electron delocalization, the amide moiety in the seven-membered ring adopts a planar conformation. The synthesis of a series of 10-[omega-[4-(4-fluorophenyl)piperazin-1-yl]alkyl]dibenzo[b,f]-1,4 - thiazepin-11(10H)-ones (10-14), and results of their evaluation for calcium channel antagonistic activity on hamster aorta, are presented.

Steady-state plasma concentrations of diltiazem and its metabolites in patients and healthy volunteers.

Diltiazem (DTZ) is a calcium antagonist widely used in the treatment of angina and hypertension. It is extensively metabolized in humans via N-demethylation, O-demethylation, deacetylation, and oxidative deamination, yielding a host of metabolites, some of which have potent pharmacological properties. After our initial identification of O-desmethyl DTZ (Mx) and N,O-didesmethyl DTZ (MB) as major metabolites of DTZ and our subsequent of identification of their chemical synthesis, an improved high-performance liquid chromatography assay was developed to determine the plasma concentrations of DTZ and seven of its major basic metabolites, including the previously unquantitated Mx and MB. The system consisted of a C18 analytical column protected by a C18 cartridge guard column and a variable wavelength ultraviolet detector set at 237 nm. The mobile phase was a mixture of methanol, 0.04 M ammonium acetate, and acetonitrile (38:36:26) containing 0.08% triethylamine, with final pH of the mobile phase adjusted to 7.5. The system was operated at room temperature isocratically at a flow rate of 1.2 ml/min. Using verapamil as an internal standard, DTZ and the basic metabolites in plasma were determined in young healthy volunteers (n = 21) and in patients with ischemic heart disease (n = 19) at steady state after repeated oral doses of 60 mg DTZ four times daily. Preliminary results show that steady-state plasma concentrations of DTZ and its metabolites were higher in the older patients than in young healthy subjects (p < 0.05).

Molecular geometries of dibenzothiazepinone and dibenzoxazepinone calcium antagonists.

A number of dibenzothiazepinones and dibenzoxazepinones have been designed, synthesized and evaluated as calcium antagonists. Molecular geometries of these dibenzotricyclic calcium antagonists have been studied using X-ray crystallography, molecular modeling and two-dimensional NMR spectroscopy. X-Ray diffraction reveals dibenzothiazepinone 1 and dibenzoxazepinone 2 to have, respectively, flexure angles of 108 degrees and 116.9 degrees between the two benzene rings. The molecular mechanics-optimized geometry of dibenzothiazepinone 1 shows a 7 degrees smaller flexure angle than the X-ray crystallographic result, while that of dibenzoxazepinone 2 has an angle only 2 degrees smaller than the X-ray result. AM1 and ab initio calculations show that the side chains can affect the geometry of the tricyclic nucleus and both 1 and 2 have negative electrostatic potentials around the bridged portion of the tricyclics. Two-dimensional NOESY NMR spectroscopy supports the extended geometry of the 6 carbon spacer as obtained from X-ray crystallography and molecular mechanics calculations. Vasorelaxation properties among these compounds appear to be relatively insensitive to the flexure angle and to chain length. Vasorelaxation is profoundly influenced by the nature of the basic terminal moiety.