Synthesis and hypnotic activity of new 4-thiazolidinone and 2-thioxo-4,5-imidazolidinedione derivatives

Conveniently accessible 4-[(2-(3,4-dimethoxyphenyl)ethyl]-3-thiosemicarbazide (2) was converted to new 1-substituted benzylidene/furfurylidene-4- [2-(3,4-dimethoxyphenyl)ethyl]-3-thiosemicarbazides (3) which furnished 2-(substituted benzylidene/furfurylidene) hydrazono-3-[2-(3,4-dimethoxyphenyl)ethyl]thiazolidin-4-ones (4) and 1-(substituted benzylidene/furfurylidene)-amino -3-[2-(3,4-dimethoxyphenyl)ethyl]-2-thioxo-4,5-imidazolidinedio nes (5) on reaction with chloroacetic acid and oxalyl chloride, respectively. The structure of 5 was confirmed by X-ray diffraction studies performed on 5a. 4 and 5 were evaluated for their potentiating effects on pentobarbital induced hypnosis. Most of the compounds caused remarkable increases in pentobarbital sleeping time.     

Effects of novel synthesized pyridothiazines on various guinea pig heart muscle preparations

Calcium channel blockers have become important tools in the treatment of cardiovascular disorders and other diseases. Hybridization of well established calcium antagonist subclasses was an attempt to optimize their pharmacological profile. The intension of this study was to investigate the electrophysiological properties of MM 10 and MM 11 two newly synthesized compounds structurally closely related to KT-362 (5-[3-[[2-(3,4-dimethoxyphenyl)ethyl]amino]-1-oxopropyl]-2,3,4,5-tetrahydro-1,5-benzothiazepine fumarate) in various isolated guinea pig heart muscle preparations by means of the conventional intracellular microelectrode tech-nique. MM 10 (2,3-dihydro-1-[N-[2-(3,4-dimethoxyphenyl)ethyl]-N-methylaminoacetyl]-1H-pyrido[2,3-b][1,4]thiazine fumarate) and MM 11 (2,3-dihydro-1-[N-[2-(3,4-dimethoxyphenyl)ethyl]-N-methylaminopropionyl]-1H-pyrido[2,3-b][1,4]thiazine fumarate) exerted very similar effects though the action of MM 11 was more pronounced. Whereas action potential amplitude and maximum upstroke velocity (V(max)) in papillary muscle, left atria and spontaneously beating Purkinje fibers was not affected by the compounds in a concentration range from 3 to 30 micromol/l, action potential duration at 90% time to repolarization was significantly prolonged in a concentration-dependent manner. Action potential duration at 20% time to repolarization was decreased in spontaneously beating Purkinje fibers and remained unchanged in papillary muscles and left atria. In sinoatrial nodes both compounds reduced rate of activity, action potential amplitude, maximum upstroke velocity and slope of slow diastolic depolarization while time to repolarization was prolonged. In 3 out of 6 experiments with spontaneously beating Purkinje fibers, MM 11 (30 micromol/l) led to the occurrence of early afterdepolarizations with a take off potential between -50 and -60 mV. All observed effects were completely reversible during washout with drug-free physiological salt solution. From these results it was concluded that both compounds in addition to their calcium antagonistic properties might depress repolarizing potassium currents. In contrast to the mother compound KT-362 they do not seem to affect the fast sodium inward current. Replacement of the benzothiazepine nucleus by a pyridothiazine structure may weaken or even eliminate sodium channel blocking ability. Shortening of the side chain might result in a general loss in activity.     

Synthesis and cytotoxic activity of carboxamide derivatives of benzo[b][1,6]naphthyridines

The reaction of 4-dimethylaminomethylene-6-methyl-4H-pyrano[4,3-b]quinoline-1,3-dione with a range of primary amines gave rise to a series of 2-substituted 6-methyl-1-oxo-1,2-dihydrobenzo[b][1,6]naphthyridine-4-carboxylic acids. The derived 4-N-[2-(dimethylamino)ethyl]carboxamides were tested for growth inhibitory properties against murine P388 leukemia, Lewis lung carcinoma (LLTC), and human Jurkat leukemia cell lines. Most compounds were potent cytotoxins, with some having IC(50) values less than 10 nM. Five were tested in vivo against subcutaneous colon 38 tumors in mice, and a single dose (3.9 mg/kg) proved to be curative for the 2-methyl and 2-(3,4-dimethoxyphenyl) derivatives in this refractory model.     

Effects of novel pyridothiazepines and pyridothiazines on contractility of isolated guinea-pig heart muscle and vascular smooth muscle preparations.

The effects of newly synthesized pyridothiazepines MM 4 (1-[N-[2-(3,4-dimethoxy-phenyl)ethyl]-N-methylaminoacetyl]-1,2,3,4 -tetrahydro-pyrido[2,3-b][1,4]thiazepine fumarate), MM 6 (1-[N-[2-(3,4-dimethoxyphenyl)-ethyl]-N-methylaminopropionyl]-1,2, 3,4-tetrahydro-pyrido[2,3-b][1,4]thiazepine fumarate) and the novel pyridothiazines MM 10 (2,3-dihydro-1-[N-[2-(3,4-dimethoxyphenyl)ethyl]-N-methylaminoacetyl+ ++]-1H-pyrido[2,3-b][1,4]thiazine fumarate) and MM 11 (2,3-dihydro-1-[N-[2-(3,4-dimethoxy-phenyl)ethyl]-N-methylaminopropio nyl]-1H-pyrido[2,3-b][1,4]thiazine fumarate) on the contractility of isolated papillary muscles and aortic preparations of guinea pigs were studied using isometric contraction force measurements. The EC50 values for the negative inotropic effect were 27 micromol/l (MM 4), 19 micromol/l (MM 6), 32 micromol/l (MM 10) and 24 micromol/l (MM 11). In K+-precontracted aortic rings ([K+]o 60 mmol/l), the compounds induced relaxation with EC50 values of 27 micromol/l (MM 4), 24 micromol/l (MM 6), 84 micromol/l (MM 10) and 68 micromol/l (MM 11). Pyridothiazepines as well as pyridothiazines (100 micromol/l) were able to depress norepinephrine bitartrate (NE 10 micromol/l)-induced contraction of aortic rings in a calcium-free solution. It was concluded that the investigated compounds exert calcium antagonistic properties in both cardiac and smooth muscle. This antagonistic effect might be due to the inhibition of transmembrane calcium influx and/or intracellular calcium release.     

Beta 1-selective adrenoceptor antagonists: examples of the 2-[4-[3-(substituted-amino)-2-hydroxypropoxy]phenyl]imidazole class

A series of 2-[4-[3-(substituted-amino)-2-hydroxypropoxy]phenyl]imidazoles is described. The compounds were investigated in vitro for beta-adrenoceptor antagonism, and several examples were found to be selective for the beta 1-adrenoceptor. The structure--activity relationship exhibited by this series of compounds is discussed. (S)-2-[p-[3-[[2-(3,4-dimethoxyphenyl)ethyl]amino]-2-hydroxypropoxy]phenyl]-4-(2 -thienyl)imidazole [(S)-13] was over 100 times more selective than atenolol for the beta 1-adrenergic receptor and has been selected for in-depth studies.     

Regiochemistry and substrate stereoselectivity of O-demethylation of verapamil in the presence of the microsomal fraction from rat and human liver

The oxidative O-demethylation of verapamil (1), a calcium channel antagonist, in the presence of rat and human liver microsomes was examined. By using GC/MS methodology and synthesized regioisomeric standards, we showed that three of the four possible monophenolic metabolites, alpha-[3-([2-(3,4-dimethoxyphenyl)ethyl]methyl-amino) propyl]-3-methoxy-4-hydroxy-alpha-(1-methylethyl)phenyl-acetonitrile (2), alpha-[3-([2-(3,4-methoxyphenyl)ethyl]methyl-amino) propyl]-3-hydroxy-4-methoxy-alpha-(1-methylethyl)phenylaceto nitrile (3), and alpha-[3-([2-(3-methoxy-4-hydroxyphenyl)ethyl]methylamino) propyl]-3,4-dimethoxy-alpha-(1-methylethyl)phenylacetonitrile (4) were formed. The other possible regioisomeric monophenolic metabolite 5 was not observed. Substrate stereoselectivity for the O-demethylation process was determined when pseudoracemic verapamil [equimolar (S)-(-)-verapamil-d6 and (R)-(+)-verapamil-d0] was used as substrate. In the presence of rat liver microsomes, significant substrate stereoselectivity was observed for formation of 4 (S/R ratio 2.28), whereas marginal substrate stereoselectivity was observed in the formation of both 3 and 2 (S/R ratio approximately 0.8). Substrate stereoselectivity for the O-demethylation process in the presence of human liver microsomes was slight and variable (six samples). Quantitatively, the ratio of O-demethylation products obtained (4:2:3) was similar in the presence of rat and human liver microsomes. In both systems, more than one-half of the total O-demethylation occurred in the aromatic ring of the phenethylamine moiety, and of the total O-demethylation process, more para- than meta-O-demethylation was observed. The similarity of regioselectivity for O-demethylation in the presence of rat and human liver microsomes suggests a similar cytochrome P-450 isozyme or set of isozymes may be responsible for the O-demethylation process.     

Beta 1-selective adrenoceptor antagonists: examples of the 2-[4-[3-(substituted amino)-2-hydroxypropoxy]phenyl]imidazole class. 2

An attempt to develop a highly cardioselective beta-adrenoceptor antagonist devoid of intrinsic sympathomimetic activity (ISA) focused on exploring structure-activity relationships around (S)-[p-[3-[[2-(3,4-dimethoxyphenyl)ethyl]amino]-2-hydroxypropoxy] phenyl]-4-(2-thienyl)imidazole. Strategies to reduce or eliminate ISA centered on structural changes that could influence activation of the receptor by the drug itself or by a metabolite. The approaches involved (a) eliminating the acidic imidazole N-H proton, (b) incorporating substituents ortho to the beta-adrenergic blocking side chain, (c) increasing steric bulk around the N-H moiety, (d) decreasing lipophilicity, (e) introducing intramolecular hydrogen bonding involving the imidazole N-H, and (f) displacing the imidazole ring from an activating position by the incorporation of a spacer element. The compounds were investigated in vitro for beta-adrenoceptor antagonism and in vivo for ISA. From these studies, the most successful variation involved the insertion of a spacer between the imidazole and aryl rings. (S)-4-Acetyl-2-[[4-[3-[[2-(3, 4-dimethoxyphenyl)ethyl]amino]-2-hydroxypropoxy]phenyl]methyl] imidazole (S-51) was demonstrated to be highly cardioselective (dose ratio beta 2/beta 1 greater than 9333) and devoid of ISA.     

High-affinity binding of [3H]DTZ323 to the diltiazem-binding site of L-type Ca2+ channels

D-cis-[N-Methyl-3H]-3-(acetyloxy)-5-[2-[[2-(3,4-dimethoxyphenyl)ethyl]-methylamino]ethyl]-2,3-dihydro-2-(4-methoxyphenyl)-1,5-benzothiazepine-4(5H)-one ([3H]DTZ323), a novel 1,5-benzothiazepine radioligand, was characterized in a ligand-receptor binding study. Specific binding of [3H]DTZ323 to rabbit skeletal muscle T-tubule membranes was saturable and reversible. Scatchard analysis indicated a single binding site with a K(d) value of 1.4 and 1.8 nM at 25 and 37 degrees C, respectively. DTZ323 and diltiazem derivatives inhibited specific [3H]DTZ323 binding with a rank order of DTZ323>DTZ417 (quaternary ammonium derivative of DTZ323)>diltiazem>L-cis-DTZ323. The affinity of DTZ323 was 51 times higher than that of diltiazem. [3H]DTZ323 binding was also completely inhibited by verapamil and tetrandrine, thus revealing the unique nature of the diltiazem-binding site. Specific [3H]DTZ323 binding to crude guinea pig ventricular membranes was inhibited by diltiazem, DTZ323 and its derivatives with IC(50) values close to those previously reported for the blockade of L-type Ca(2+) channel currents. These results indicate that [3H]DTZ323 is a potent and selective radioligand for the diltiazem-binding site of L-type Ca(2+) channels.     

Electrophysiological characterization of BRL-32872 in canine Purkinje fiber and ventricular muscle. Effect on early after-depolarizations and repolarization dispersion

Amongst the different pharmacological approaches to the treatment of cardiac arrhythmias, compounds with multiple electrophysiological activities appear to exhibit a reduced adverse effect profile. BRL-32872 (N-(3,4-dimethoxyphenyl)-N-[3[[2-(3,4-dimethoxyphenyl) ethyl] propyl]-4-nitrobenzamide hydrochloride) is a typical example of an antiarrhythmic agent with combined K(+) and Ca(2+) blocking actions. In this study, we investigated the effects of BRL-32872 on early after-depolarizations and on dispersion of repolarization. Action potentials were recorded either in canine cardiac Purkinje fibers alone or in preparations containing both ventricular muscle and the attached Purkinje fibers. In Purkinje fibers, BRL-32872 (0. 3-10 microM) induced a bell-shaped concentration-dependent increase in action potential duration. At 90% of repolarization, the action potential was prolonged at concentrations up to 1 microM and was shortened when the concentration of BRL-32872 was further increased. In all 17 experiments, BRL-32872 did not cause early after-depolarizations in Purkinje fibers. On the contrary, BRL-32872 (3 microM) systematically suppressed early after-depolarizations induced by clofilium (4-chloro-N, N-diethyl-N-heptylbenzenebutanaminium tosylate, 1 microM), a selective inhibitor of the delayed rectifier K(+) current. A similar effect was observed once with 1 microM BRL-32872, a concentration able to prolong Purkinje fiber action potentials. Simultaneous recording of action potentials in ventricular and Purkinje preparations showed that increasing concentrations of BRL-32872 (0. 3-10 microM) induced a limited increase in the difference of repolarization time between the two tissues. The selective K(+) channel inhibitor E-4031 (N-(4-(1-[2-(6-methyl-2-pyridyl) ethyl]-4-piperidyl)-carbonyl] phenyl) methanesulfonamide dihydrochloride dihydrate) exhibited a significant concentration-dependent increase in dispersion of repolarization. We conclude from the present results that the Ca(2+) blocking activity of BRL-32872 (i) prevents the occurrence of early after-depolarizations associated with action potential prolongation and (ii) limits an excessive increase in action potential duration heterogeneity. These electrophysiological features might represent the basis for antiarrhythmic compounds with reduced proarrhythmic profile.     

Antitumor agents. 217. Curcumin analogues as novel androgen receptor antagonists with potential as anti-prostate cancer agents

A number of curcumin analogues were prepared and evaluated as potential androgen receptor antagonists against two human prostate cancer cell lines, PC-3 and DU-145, in the presence of androgen receptor (AR) and androgen receptor coactivator, ARA70. Compounds 4 [5-hydroxy-1,7-bis(3,4-dimethoxyphenyl)-1,4,6-heptatrien-3-one], 20 [5-hydroxy-1,7-bis[3-methoxy-4-(methoxycarbonylmethoxy)phenyl]-1,4,6-heptatrien-3-one], 22 [7-(4-hydroxy-3-methoxyphenyl)-4-[3-(4-hydroxy-3-methoxyphenyl)acryloyl]-5-oxohepta-4,6-dienoic acid ethyl ester], 23 [7-(4-hydroxy-3-methoxyphenyl)-4-[3-(4-hydroxy-3-methoxyphenyl)acryloyl]5-oxohepta-4,6-dienoic acid], and 39 [bis(3,4-dimethoxyphenyl)-1,3-propanedione] showed potent antiandrogenic activities and were superior to hydroxyflutamide, which is the currently available antiandrogen for the treatment of prostate cancer. Structure-activity relationship (SAR) studies indicated that the bis(3,4-dimethoxyphenyl) moieties, the conjugated beta-diketone moiety, and the intramolecular symmetry of the molecules seem to be important factors related to antiandrogenic activity. The data further suggest that the coplanarity of the beta-diketone moiety and the presence of a strong hydrogen bond donor group were also crucial for the antiandrogenic activity, which is consistent with previous SAR results for hydroxyflutamide analogues. When the pharmacophoric elements of dihydrotestosterone (DHT) and compound 4 are superposed, the resulting construct implies that the curcumin analogues may function as a 17alpha-substituted DHT. Compounds 4, 20, 22, 23, and 39 have been identified as a new class of antiandrogen agents, and these compounds or their new synthetic analogues could be developed into clinical trial candidates to control androgen receptor-mediated prostate cancer growth.     

Aminoalkynyldithianes. A new class of calcium channel blockers

Several dithiane derivatives, prepared as intermediates for compounds structurally related to the therapeutically useful antimuscarinic agent oxybutynin, were effective inhibitors of calcium ion induced contraction of guinea pig ileal strips and of KCl-induced calcium entry into neuronal cells. Although the first member of this series, 2-[5-(diethylamino)-3-pentynyl]-1,3-dithiane (2a), was only marginally effective, its condensation product with diphenyl ketone, i.e. 2-[5-(diethylamino)-3-pentynyl]-2-(a,a-diphenyl-a- hydroxymethyl)-1,3-dithiane (3a), demonstrated weak, but significant, calcium channel antagonist activity. As part of a structure-activity relationship (SAR) study, various structural analogues of 2a and 3a were prepared and examined for calcium antagonist properties. In addition to these structural types, ring bridged (tricyclic) congeners of 3, i.e. 4, related bicyclic compounds 5, dehydroxylated derivatives 6, some homologous 2-[[[(N,N-disubstituted-amino)methyl]2- phenyl-1,3-dithianes (7), and a series of 2-[6-[N,N-disubstituted-amino)methyl]-1-hydroxy-1-phenyl- 4-hexynyl]-1,3-dithianes (8) were prepared and studied for calcium channel blocking activity. In general, greatest potency was noted in the tricyclic series 4; however, a definitive SAR could not be established. A structural similarity between several potent calcium antagonists having the structures 7c, 8b, and 8d and the well-known calcium channel blockers verapamil and tiapamil suggests these compounds may act at the same site. Compounds in the other classes (2-6) failed to show clearly defined SAR and their potency differed markedly in two tests for calcium channel antagonist activity. These results may indicate that the dithiane derivatives 2-6 produce their effects in a manner differing from that of the calcium channel antagonists diltiazem, verapamil, and nitrendepine.     

Synthesis of 3-(3,4-dimethoxyphenyl)-1 H-1,2,4-triazole-5-thiol and 2-amino-5-(3,4-dimethoxyphenyl)-1,3,4-thiadiazole derivatives exhibiting anti-inflammatory activity

New 1-acylderivatives of 5-alkylthio-3-(3,4-dimethoxyphenyl)-4H-1,2,4-triazole (5c-f, 6d-f) were synthesized by the acylation of 5-alkylthio-3-(3,4-dimethoxyphenyl)-4H-1,2,4-triazoles (3, 4) with acyl chlorides. The compounds 3, 4 were obtained by the alkylation of 3-(3,4-dimethoxyphenyl)-1H-1,2,4-triazole-5-thiol (2) sodium salt with alkyl iodides. Compound 2 and 2-amino-5-(3,4-dimethoxyphenyl)-1,3,4-thiadiazole (8) were prepared by the treatment of 3,4-dimethoxybenzoylthiosemicarbazide (1) with sodium hydroxide or acetyl chloride (and then sodium hydroxide), respectively. Related 2-acylamino-5-(3,4-dimethoxyphenyl)-1,3,4-thiadiazoles (7c, e, f) were synthesized by the acylation of compound 8 with acyl chlorides. 3-(3,4-Dimethoxyphenyl)-4-phenyl-4,5-dihydro-1H-1,2,4-triazole-5-thione (9) was N-acylated with acyl chlorides or S-methylated with iodomethane to give 1-acyl-3-(3,4-dimethoxyphenyl)-4-phenyl-4,5-dihydro-1H- 1,2,4-triazole-5-thiones (10a, b) or 3-(3,4-dimethoxyphenyl)-5-methylthio-4-phenyl-4H-1,2,4-triazole (11) respectively. The synthesized compounds 6d, 7a, c, 10a, b, 11 exhibit anti-inflammatory activity.     

Comparative cardiac effects of KT-362 and verapamil in isolated heart--correlation to calcium channel current depression.

Direct cardiac effects of KT-362 (5-[3 [[-2-(3,4-dimethoxyphenyl)-ethyl]amino]-1-oxopropyl]-2,3,4,5- tetrahydro-1,5-benzothiazepine fumarate), a drug that may inhibit intracellular calcium mobilization as well as extracellular calcium influx was compared to verapamil. Guinea pig hearts (n = 19) were used to examine the changes in atrial rate, atrioventricular conduction time (AVCT), coronary flow, myocardial oxygen consumption (MVO2), and isovolumetric left ventricular pressure (LVP). Both drugs concentration-dependently and reversibly decreased atrial rate, contractility, and MVO2; AVCT increased during spontaneous rhythm. The increases in AVCT and the incidence of AV dissociation were accentuated during cardiac pacing. Verapamil significantly increased coronary flow, while KT-362 did not. Median effective concentration (EC50) was about 25 times lower for verapamil in depressing LVP and about three times lower in depressing atrial rate and AV conduction. The changes in calcium channel current in voltage-clamped single canine Purkinje cells (n = 6) were also examined. Verapamil (0.3 microM) and KT-362 (7 microM) decreased peak Ca2+ channel current at maximum activation (+10 mV) by 38.1 +/- 8% and 28.6 +/- 6%, respectively, without shifting the current-voltage relationship. This study indicates that verapamil is more potent than KT-362 in depressing contractile function, heart rate, and AV conduction in isolated hearts and calcium current in isolated cardiac Purkinje cells. Moreover, there was a much greater difference between the EC50 for verapamil and that for KT-362 for the depression of indices of contractility (23-30-fold) than for the depression of sinoatrial and atrioventricular nodal function (2.5-4-fold).     

Regiochemistry and enantioselectivity in the oxidative N-dealkylation of verapamil

The oxidative N-dealkylation of verapamil (1), a calcium channel antagonist, was examined in the presence of rat and human liver microsomes by using GC-MS methodology and synthesized regio-isomeric standards. All three possible secondary amine metabolites, N-methyl-4-(3,4-dimethoxyphenyl)-4-cyano-5-methylhexylamine (5), norverapamil (4), and N-methyl-2-(3,4-dimethoxyphenyl)ethylamine (3), were formed as microsomal metabolites. Compound 5 and norverapamil (4) were major products. Substrate stereoselectivity for the N-dealkylation process was determined when pseudoracemic verapamil[equimolar (S)-(-)-verapamil-d6 and (R)-(+):verapamil-d0] was used as substrate. In the presence of rat liver microsomes, a slight enantiomeric preference for the metabolism of (R)-verapamil to secondary amines 3 and 5 (S/R ratio = 0.88 and 0.78, respectively) was observed. In contrast, (S)-verapamil was preferentially metabolized to norverapamil (4) and primary amine 9 (S/R ratio = 1.20 for both). The enantioselectivity for the N-dealkylation process in the presence of human liver microsomes was slight and variable (six samples). Quantitatively, the major N-dealkylation routes in both microsomal systems yielded norverapamil (4) and secondary amine 5. Greater substrate enantioselectivity was observed for the N-dealkylation process in rat liver microsomes than in human liver microsomes. In rat liver microsomal studies, two aliphatic aldehydes (2 and 6) were successfully trapped as their O-methyloximes (7 and 11, respectively) by using methoxylamine. In addition, the alcohols formed from reduction of these aldehydes were observed, due in part to a direct reduction by NADPH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Synthesis and calcium antagonistic activities of fecal metabolites of a new calcium antagonist (+)-(R)-3,4-dihydro-2-[5-methoxy-2-[3-[N-methyl-N- [2-[(3,4-methylenedioxy)phenoxy]ethyl]amino]propoxy]phenyl]-4- methyl-3-oxo-2H-1,4-benzothiazine (SD-3211) in rats

In order to confirm the structure of three fecal metabolites, M-I, M-II and M-III, of a new calcium antagonist, (+)-3,4-dihydro-2-[5-methoxy-2-[3-[N-methyl-N-[2-[(3,4-methylenedioxy) phenoxy]ethyl]amino]propoxy]phenyl]-4-methyl-3-oxo-2H-1,4- benzothiazine (SD-3211), in rats, (+)-3,4-dihydro-2-[5-hydroxy-2-[3-[N-methyl-N-[2- [(3,4-methylenedioxy)-phenoxy]ethyl]amino]propoxy]phenyl]-4-methyl- 3-oxo-2H-1,4-benzothiazine((+)-I), (+)-3,4-dihydro-2-[5-hydroxy-2-[3-[N- [2-[(3,4-methylenedioxy) phenoxy]ethyl]amino] propoxy]-phenyl]-4-methyl-3-oxo-2H-1,4-benzothiazine((+)-II) and (+)-3,4-dihydro-2-[5-methoxy-2-[3-[N-[2-[(3,4-methylenedioxy) phenoxy]ethyl]amino]propoxy]phenyl]-4-methyl-3-oxo-2H-1,4- benzothiazine((+)-III) were synthesized. Compounds (+)-I, (+)-II and (+)-III were identified with the fecal metabolites M-I, M-II and M-III, respectively. The calcium antagonistic activities of (+)-I, (+)-II and (+)-III were examined.     

Cytochromes of the P450 2C subfamily are the major enzymes involved in the O-demethylation of verapamil in humans

The calcium channel blocker verapamil [2,8-bis-(3,4-dimethoxyphenyl)-6-methyl-2-isopropyl-6-azaoctanitrile] undergoes extensive biotransformation in man. We have previously demonstrated cytochrome P450 (CYP) 3A4 and 1A2 to be the enzymes responsible for verapamil N-dealkylation (formation of D-617 [2-(3,4-dimethoxyphenyl)-5-methylamino-2-isopropylvaleronitrile]), and verapamil N-demethylation (formation of norverapamil [2,8-bis(3,4-dimethoxyphenyl)-2-isopropyl-6-azaoctanitrile]), while there was no involvement of CYP3A4 and CYP1A2 in the third initial metabolic step of verapamil, which is verapamil O-demethylation. This pathway yields formation of D-703 [2-(4-hydroxy-3-methoxyphenyl)-8-(3,4-dimethoxyphenyl)-6-methyl-2-isopropyl-6-azaoctanitrile] and D-702 [2-(3,4-dimethoxyphenyl)-8-(4-hydroxy-3-methoxyphenyl)6-methyl-2-isopropyl-6-azaoctanitrile]. The enzymes catalyzing verapamil O-demethylation have not been characterized so far. We have therefore identified and characterized the enzymes involved in verapamil O-demethylation in humans by using the following in vitro approaches: (I) characterization of O-demethylation kinetics in the presence of the microsomal fraction of human liver, (II) inhibition of verapamil O-demethylation by specific antibodies and selective inhibitors and (111) investigation of metabolite formation in microsomes obtained from yeast strain Saccharomyces cerevisiae W(R), that was genetically engineered for stable expression of human CYP2C8, 2C9 and 2C18.In human liver microsomes (n=4), the intrinsic clearance (CL_int), as derived from the ratio of V _max/K_m, was significantly higher for O-demethylation to D-703 compared to formation of D-702 following incubation with racemic verapamil (13.9±1.0 vs 2.4±0.6 ml^*min^{-1 *}g^-1 mean±SD; p<0.05), S-Verapamil (16.8±3.3 vs 2.2±1.2 ml^* mini^*g^-1, p<0.05) and R-verapamil (12.1±2.9 vs 3.6 ±1.3 ml^*min^{-1 *} g^-1; p<0.05), thus indicating regioselectivity of verapamil O-demethylation process. The CL_int of D-703 formation in human liver microsomes showed a modest but significant degree of stereo selectivity (p<0.05) with a S/R-ratio of 1.41±0.17. Anti-LKM2 (anti-liver/kidney microsome) autoantibodies (which inhibit CYP2C9 and 2C19) and sulfaphenazole (a specific CYP2C9 inhibitor) reduced the maximum rate of formation of D-703 by 81.5±4.5% and 45%, that of D-702 by 52.7±7.5% and 72.5%, respectively. Both D-703 and D-702 were formed by stably expressed CYP2C9 and CYP2C18, whereas incubation with CYP2C8 selectively yielded D-703.In conclusion, our results show that enzymes of the CYP2C subfamily are mainly involved in verapamil O-demethylation. Verapamil therefore has the potential to interact with other drugs which inhibit or induce these enzymes.     

Evaluation of radioiodinated 1-[2-(3,4-Dimethoxyphenyl)ethyl]-4-(2-iodophenylpropyl)piperazine as a tumor diagnostic agent with functional sigma receptor imaging by single photon emission computed tomography

Radioiodinated 1-[2-(3,4-dimethoxyphenyl)ethyl]-4-(2-iodophenylpropyl)piperazine (o-BON) and 1-[2-(3,4-dimethoxyphenyl)ethyl]-4-(3-iodophenylpropyl)piperazine (m-BON) were evaluated as single photon emission computed tomography (SPECT) radiopharmaceuticals for tumor imaging by visualization of sigma receptors. In vivo biodistribution studies of [125 I]o-BON and [125 I]m-BON in tumor-bearing mice showed a high tumor uptake and prolonged retention of radiolabeled compounds in the tumor. In contrast with these factors, the blood and muscle accumulations were low, which resulted in a good tumor-to-blood ratio and tumor-to-muscle ratio. In peripheral organs, [125 I]o-BON showed rapid clearance in comparison with [125 I]m-BON. Selective interactions of [125 I]o-BON and [125 I]m-BON with sigma receptors on tumor cell membranes were confirmed by pretreatment experiments with various sigma and other receptor ligands. [125 I]o-BON possesses higher specific binding toward sigma receptors than does [125 I]m-BON; thus, [125 I]o-BON was chosen for further evaluations. High uptake of [125 I]o-BON was observed in various tumors, and a good linear correlation (R2=0.70) was found between accumulation of [125 I]o-BON and the sigma receptor expression level. Furthermore, the accumulation of [125 I]o-BON in tumors reflected their proliferation rate. These results suggest that it is feasible to use radioiodinated o-BON as a marker for measuring the proliferative status associated with sigma receptor expression.     

N-substituted 4beta-methyl-5-(3-hydroxyphenyl)-7alpha-amidomorphans are potent, selective kappa opioid receptor antagonists

In a previous study, we identified (-)-N-[(1R,4S,5S,7R)-5-(3-hydroxyphenyl)-4-methyl-2-(3-phenylpropyl)-2-azabicyclo[3.3.1]non-7-yl]-3-(1-piperidinyl)propanamide (5a, KAA-1) as the first potent and selective kappa opioid receptor antagonist from the 5-(3-hydroxyphenyl)morphan class of opioids. In this study we report an improved synthesis of this class of compounds. The new synthetic method was used to prepare analogues 5b-r where the morphan N-substituent and 7alpha-amido group were varied. Most of the analogues showed sub-nanomolar potency for the kappa opioid receptor and were highly selective relative to the mu and delta opioid receptors. (-)-3-(3,4-Dihydroisoquinolin-2(1H)-yl)-N-{(1R,4S,5S,7R)-5-(3-hydroxyphenyl)-4-methyl-2-[2-(2-methylphenyl)ethyl]-2-azabicyclo[3.3.1]non-7-yl}propanamide (5n, MTHQ) is at least as potent and selective as nor-BNI as a kappa opioid receptor antagonist in the [35S]GTP-gamma-S in vitro functional test.     

Evaluation of WO-2012085582 and WO-2012085583 two identified MABAs: backups to AZD-2115?

These two patent applications each claim combination formulations comprising a single dual-acting muscarinic antagonist/beta 2 agonist (MABA), as a free base or salt, with a second class of drug and the use of these formulations for the treatment of asthma and COPD. The two specified compounds are close analogues of each other and were originally disclosed in the same patent application. They are, respectively, 3-(2-chloro-3-((4-(2-ethylthiazole-4-carbonyl)-1-oxa-4,9-diazaspiro[5.5]undecan-9-yl)methyl)phenethoxy)-N-cyclopentyl-N-(2-(2-(5-hydroxy-3-oxo-3,4-dihydro-2H-benzo[b]{1,4] oxazin-8-yl)ethylamino)ethyl)propanamide and N-butyl-N-(2-(2-(5-hydroxy-3-oxo-3,4-dihydro-2H-benzo[b][l,4]oxazin-8-yl)ethylamino)ethyl)-3-(3-(2-(4-(2-isopropylthiazole-4-carbonyl)-1-oxa-4,9-diazaspiro[5.5]undecan-9-yl)ethyl)phenethoxy)propanamide.