SK&F 96365 (1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenylethyl]-1H- imidazole hydrochloride) stimulates phosphoinositide hydrolysis in human U373 MG astrocytoma cells

SK&F 96365 (1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenylethyl]-1H-imi dazole hydrochloride) stimulated the accumulation of [3H]inositol monophosphates ([3H]IP1) in human U373 MG astrocytoma cells prelabelled with [3H]inositol (EC50 15 +/- 1 microM, Hill coefficient 3.8 +/- 0.4). SK&F 96365-induced accumulation of [3H]IP1 increased linearly with time, but there was no initial rapid formation of [3H]IP3. SK&F 96365 also stimulated [3H]IP1 accumulation in human HeLa cells, but only to a small extent in slices of rat cerebral cortex and guinea-pig cerebellum. SK&F 96365-induced accumulation of [3H]IP1 in U373 MG cells increased as extracellular Ca2+ was increased from nominally zero to 4 mM, but there was no evidence that SK&F 96365 induced any marked entry of Ca2+ into cells; only an inhibition of store-refilling-induced Ca2+ entry was apparent. Further, the response to SK&F 96365 was additive with that to the Ca2+ ionophore ionomycin. Depolarization of the cells with raised K+ produced only a small stimulation of phosphoinositide hydrolysis. SK&F 96365 caused the release of Ca2+ from intracellular stores in U373 MG cells (EC50 26 +/- 14 microM), but thapsigargin induced only a small accumulation of [3H]IP1. Miconazole, another N-substituted imidazole, also stimulated [3H]IP1 accumulation in U373 cells.     

Studies on anti-platelet agents. II. Synthesis and platelet-inhibitory activity of 5-methyl-4-(3-pyridyl)-2-(substituted benzimidazol-5-yl)imidazoles

A series of 5-methyl-4-(3-pyridyl)-2-(substituted benzimidazol-5-yl)imidazole derivatives was synthesized and tested for anti-platelet and vasodilatory activities. Some compounds were found to have potent activities and low acute toxicity. In particular, 5-methyl-4-(3-pyridyl)-2-(7-chloro-6-methoxy-2- methylbenzimidazol-5-yl)imidazole (26) and 5-methyl-4-(3-pyridyl)-2-(7-chloro-3-methoxy-2-methylbenzimidazol- 5-yl)imidazole (33) exhibited 63% or 51% inhibition at a dose of 10 mg/kg for anti-patelet activity ex vivo in rats, respectively, while they showed no toxicity even at 180 or 100 mg/kg, respectively. Compound 33 also exhibited potent vasodilatory activity (ED50 = 11 micrograms/ml). Enzyme studies on these imidazoles showed that the novel imidazoles inhibit some enzymes which are involved in the platelet aggregation cascade such as cyclooxygenase, phosphodiesterase (PDE), and thromboxane A2 synthetase. The enzyme assay also suggested that the inhibitory activity on PDE may account for the vasodilatory activity of these imidazoles.     

Retention mechanism of imidazoles in connective tissue. I. Binding to elastin

To elucidate the retention mechanism of drugs with imidazole moiety in the connective tissue, the retention form and site of [2-14C]imidazole and 2-methyl[2-14C]imidazole were studied after intravenous administration to rats (3 micromol/kg body weight). The aorta, which is representative of the connective tissue, retained considerable radioactivity after dosing for both the imidazoles. It was observed that most of the aortic radioactivity came from the irreversibly bound fraction with elastin and that this was in close agreement with the microautoradiographic observation that showed that the retention of radioactivity occurred near the elastic fiber in the aorta. Pretreatment of rats with SKF525-A significantly increased the irreversible binding of radioactivity from the imidazoles in aorta, whereas neither phenobarbital nor 3-methylcholanthrene increased the binding. Regarding the urinary metabolite profile, the excretion of intact form significantly increased by SKF525-A pretreatment for imidazole, and an increasing tendency was also observed for 2-methylimidazole. However, no in vitro irreversible binding of imidazoles to aortic tissue was observed after incubating at physiological pH and temperature. These findings indicate that the retention of drugs with imidazole moiety in the connective tissue is largely attributable to irreversible binding between the imidazole moiety and elastin, and that the binding may be mediated through cytochrome P450-independent biotransformation.     

Activation of two types of Ca2+-permeable nonselective cation channel by endothelin-1 in A7r5 cells

1. In A7r5 cells loaded with the Ca2+ indicator fura-2, we examined the effect of a Ca2+ channel blocker SK&F 96365 on increases in intracellular free Ca2+ concentrations ([Ca2+]i) and Mn2+ quenching of fura-2 fluorescence by endothelin-1 (ET-1). Whole-cell patch-clamp was also performed. 2. Higher concentrations (> or = 10 nM) of ET-1 (higher [ET-1]) evoked a transient peak and a subsequent sustained elevation in [Ca2+]i: removal of extracellular Ca2+ abolished only the latter. A blocker of L-type voltage-operated Ca2+ channel (VOC) nifedipine at 1 microM reduced the sustained phase to about 50%, which was partially sensitive to SK&F 96365 (30 microM). 3. Lower [ET-1] (< or = 1 nM) evoked only a sustained elevation in [Ca2+]i which depends on extracellular Ca2+. The elevation was partly sensitive to nifedipine but not SK&F 96365. 4. In the presence of 1 microM nifedipine, higher [ET-1] increased the rate of Mn2+ quenching but lower [ET-1] had little effect. 5. In whole-cell recordings, both lower and higher [ET-1] induced inward currents at a holding potential of -60 mV with linear I-V relationships and reversal potentials close to 0 mV. The current at lower [ET-1] was resistant to SK&F 96365 but was abolished by replacement of Ca2+ in the bath solution with Mn2+. The current at higher [ET-1] was abolished by the replacement plus SK&F 96365. 6. In a bath solution containing only Ca2+ as a movable cation, ET-1 evoked currents: the current at lower [ET-1] was sensitive to Mn2+, whereas that at higher [ET-1] was partly sensitive to SK&F 96365. 7. These results indicate that in addition to VOC, ET-1 activates two types of Ca2+-permeable nonselective cation channel depending on its concentrations which differ in terms of sensitivity to SK&F 96365 and permeability to Mn2+.     

Syntheses and evaluation of benzodiazaborine compounds against M. tuberculosis H37Rv in vitro

The synthesis of six benzo[e]diazaborine compounds and thier in vitro evaluation against M. tuberculosis H37Rv is described. The compounds 1,2-dihydro-1-hydroxy-2-phenyl-2,4,1-benzo[e]diazaborin-3(4H)-one, 4, and 1,2-dihydro-1-hydroxy-2-(3-pyridyl)-2,4,1-benzo[e]diazaborin-3(4H) - thione, (5), showed the greatest inhibitory activity.     

Studies on substances with antiblastic activity. IX. Anthramycin and analogs. IX. Synthesis of 7-methoxy-8-hydroxy-10,11-dihydro-5H-pyrrolo[2,1-c] [1,4] benzodiazepine and other compounds related to oxotomaymycin

The synthesis of some pyrrolobenzodiazepine derivatives related to oxotomaymycin, an antibiotic recovered together with tomaymycin from fermentation broths of Streptomyces achromogenes var. tomaymycetics, is described. Reaction between 2-nitro-4-benzyloxy-5-methoxybenzylbromide and pyrrole-2-carboxyaldehyde afforded 1-(2-nitro-4-benzyloxy-5-methoxybenzyl)pyrrole-2-carboxyaldehyde. Catalytic reduction of this compound with hydrogen in the presence of Pd/C gave 10,11-dihydro-8-hydroxy-7-methoxy-5H-pyrrolo[2.1-c] [1,4]benzodiazepine. Amides obtained from condensation between 2-nitro-4-benzyloxy-5-methoxybenzoic acid chloride and proline or hydroxyproline were reduced catalytically to 2,3-dihydro-8-hydroxy-7-methoxy-1H-pyrrolo [2,1-c] [1,4]benzodiazepine-5,11 (10H, 11aH)-dione and its 2-hydroxyderivative respectively. The synthesis of 10,11-dihydro-8-hydroxy-9-methoxy-5-pyrrolo [2,1-c] [1,4]benzodiazepine is also reported.     

The Rho GTPases in macrophage motility and chemotaxis

The present study was carried out to clarify the role of nonselective cation channels as a Ca2+ entry pathway in the contraction and the increase in [Ca2+]i induced by endothelin- in endothelium-denuded rat thoracic aorta rings, and their suppression by nitric oxide (NO). In Ca2+-free medium, the endothelin-1-induced contraction was suppressed to about 20% of control values, although the increase in [Ca2+]i became negligible. The contraction and the increase in [Ca2+]i monitored by fura 2 fluorescence were unaffected by a blocker of L-type voltage-operated Ca2+ channels nifedipine. A blocker of nonselective cation channels 1-[beta-[3-(4-methoxyphenyl)propoxyl]-4-methoxyphenethyl]-1H-imida zole . HCl(SK&F 96365) suppressed the endothelin-1-induced contraction and increase in [Ca2+]i to the level similar to that after removal of extracellular Ca2+. SK&F 96365 had no further effect on the endothelin-1-induced contraction in the absence of extracellular Ca2+. The endothelin-1-induced contraction and increase in [Ca2+]i were abolished by a donor of NO sodium nitroprusside. The effects of another NO donor 3-morpholinosydnonimine (SIN-1) were also tested and yielded essentially similar results to those for sodium nitroprusside on the endothelin-1-induced contraction. Furthermore, the inhibitory effects of sodium nitroprusside could be blocked with a guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ) at 30 microM. These findings suggest that Ca2+ entry through nonselective cation channels but not voltage-operated Ca2+ channels plays a critical role in the endothelin-1-induced increase in [Ca2+]i and the resulting contraction and that inhibition by NO of the endothelin-1-induced contraction is mainly the result of blockade of Ca2+ entry through these channels.     

A case of primary splenic large cell lymphoma with a t(9;14)(p13;q32)

Appropriately substituted 2,3-dihydro-7H-thiazolo[3,2-a]pyrimidin-7-ones 9-12 and 18 were considered as annulated analogues of HEPT (1-[(2-hydroxyethoxy)methyl]-6-(phenylthio)-thymine), and some of these compounds were also found active against HIV-1, the most active one being 2,3-dihydro-5-[(3,5-dimethylphenyl)methyl]-3-ethoxy-6-ethyl-7H- thiazolo[3,2-a]pyrimidin-7-one (10b). S-Alkylation of 5-alkyl-6-(arylmethyl)-2-thiouracils 1-4 was performed with 2-bromoacetaldehyde acetals to furnish the S-[bis(alkoxy)ethyl] derivatives 5-8 and with allyl bromide to furnish S-allyl derivatives 17. The target compounds 9-12 were obtained by an N1 regioselective intramolecular cyclization reaction of silylated 5-8 using trimethylsilyl trifluoromethanesulfonate (TMS triflate) as the catalyst. Treatment of the S-allyl derivatives 17 with bromine in dry methylene chloride afforded the 3-(bromomethyl) derivatives 18.     

High-performance liquid chromatography study of stereospecific microsomal enzymes catalysing the reduction of a potential cytostatic drug, oracin. Interspecies comparison

One of the main metabolites of oracin (I) ?6-[2-(2-hydroxyethyl)aminoethyl]-5,11-dioxo-5,6-dihydro-11H-indeno[1,2- c] isoquinoline?, a potential cytostatic drug, is 11-dihydrooracin (II) ?(+),(-)-6-[2-(2-hydroxyethyl)aminoethyl]-5-oxo-11-hydroxy-5,6-dihydro-1 1H- indeno[1,2-c]isoquinoline?, a metabolite formed by the reduction of oracin's pro-chiral centre on C 11. This metabolite has been found in all laboratory species in vitro and in vivo and it constitutes the main metabolite in man. The stereospecificity of reducing enzymes participating in the oracin biotransformation pathway was investigated using microsomal preparations from standard laboratory animals. Enzyme stereospecificity has been defined as preferential formation by the enzyme of the (+) or (-) stereoisomer of II. Significant interspecies differences were observed in the stereospecificity of the respective biotransformation enzymes. HPLC quantitative determinations of both enantiomers were performed using a Chiralcel OD-R column as chiral stationary phase with excellent resolution and stability.     

New indole and pyridazinoindole analogs--synthesis and study as inhibitors of phosphodiesterases and as inhibitors of blood platelet aggregation

This paper presents the synthesis of new indole, pyridazino[4,5-b]-indole, and pyridazino[4,5-a]indole analogs as well as a study of their "in vitro" activity as inhibitors of different phosphodiesterases isolated from dog cardiac tissue, dog aorta, and bovine platelets; the study of their activity as inhibitors of platelet aggregation in guinea pig whole blood, with ADP and arachidonic acid (AA) as pro-aggregants, is also included. The selected compounds 8-benzyloxy-3,4-dihydro-1-(3,4,5-trimethoxy)benzylideneaminopyridazin o[4,5- b]indole 14g, and 8-benzyloxy-4-[(3,4-dimethyl)pyrazolyl]pyridazino[4,5-b]indo le 20 present an interesting profile as potential inodilators, with a complementary, beneficial activity as inhibitors of the aggregation, activities which could possibly be related to the inhibition of the PDE's. Among the other compounds studied, 8-benzyloxy-3,4-dihydro-1-[4-(methyl)piperazino]acetamidopyrida zino[4,5- b]indol-4-one 16c and 8-benzyloxy-3,4-dihydro-1-[4-(2- methoxyphenyl)piperazino]acetamidopyridazino[4,5-b]indol-4-o ne 16f stood out as inhibitors of platelet aggregation, with a mechanism that could possibly be related to the AA cascade.     

5-aryl-2,3-dihydro-5H-imidazo[2,1-a]isoindol-5-ols. A novel class of anorectic agents

A series of 5-aryl-2,3-dihydro-5H-imidazo[2,1-a]isoindol-5-ols (IV), prepared by the LiA1H4 reduction of the corresponding 9b-aryl-1,2,3,9b-tetrahydro-5H-imidazo[2,1-a]isoindol-5-ones (II), was evaluated for suppression of food consumption in rats. One member of this series, 5-p-chlorophenyl-2,3-dihydro-5H-imidazo[2,1-a]isoindol-5-ol (6, mazindol), was evaluated in squirrel and capuchin monkeys and found to have anorexic activity approximately equal to d-amphetamine.     

Influence of some novel N-substituted azoles and pyridines on rat hepatic CYP3A activity

A series of N-substituted heteroaromatic compounds structurally related to clotrimazole was synthesized, and the effects of these compounds on ethosuximide clearance in rats were determined as a measure of their abilities to induce cytochrome P4503A (CYP3A) activity. Ethosuximide clearance and in vitro erythromycin N-demethylase activity were shown to correlate. In this series, imidazole or other related heteroaromatic "head groups" were linked to triphenylmethane or other phenylmethane derivatives. Within the series, it was found that 1-triphenylmethane-substituted imidazoles elicited the greatest increase in CYP3A activity, and that among the triphenylmethyl-substituted imidazoles, the highest activities were achieved by the substitution of F- or Cl- in either the meta or para position of one of the phenyl rings. Diphenylmethyl-substituted pyridine was effectively devoid of activity. Compounds eliciting the largest increase in CYP3A activity (viz. 1-[(3-fluorophenyl)diphenylmethyl]imidazole, 1-[(4-fluorophenyl)diphenylmethyl]imidazole, and 1-[tri-(4-fluorophenyl)methyl]imidazole) produced little or no increase in ethoxyresorufin O-dealkylase (EROD) activity (i.e. CYP1A), whereas benzylimidazole, which elicited only a small increase in CYP3A activity, produced an almost 9-fold increase in CYP1A activity. For a series of eleven compounds exhibiting a wide range of influence on CYP3A activity, a positive correlation was found between ethosuximide clearance and hepatic CYP3A mRNA levels.     

Activation of chemically diverse procarcinogens by human cytochrome P-450 1B1

A human cytochrome P-450 (P450) 1B1 cDNA was expressed in Saccharomyces cerevisiae and the microsomes containing P450 1B1 were used to examine the selectivity of this enzyme in the activation of a variety of environmental carcinogens and mutagens in Salmonella typhimurium TA1535/pSK1002 or NM2009 tester strains, using the SOS response as an end point of DNA damage. We also determined and compared these activities of P450 1B1 with those catalyzed by recombinant human P450s 1A1 and 1A2, which were purified from membranes of Escherichia coli. The carcinogenic chemicals tested included 27 polycyclic aromatic hydrocarbons and their dihydrodiol derivatives, 17 heterocyclic and aryl amines and aminoazo dyes, three mycotoxins, two nitroaromatic hydrocarbons, N-nitrosodimethylamine, vinyl carbamate, and acrylonitrile. Among the three P450 enzymes examined here, P450 lB1 was found to have the highest catalytic activities for the activation of 11,12-dihydroxy-11,12-dihydrodibenzo[a,l]pyrene, 1,2-dihydroxy-1,2-dihydro-5-methylchrysene, (+)-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene, 11,12-dihydroxy-11,12-dihydrobenzo[g]chrysene, 3,4-dihydroxy-3,4-dihydrobenzo[c]phenanthrene, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole, 2-aminoanthracene, 3-methoxy-4-aminoazobenzene, and 2-nitropyrene. P450 1B1 also catalyzed the activation of 2-amino-3,5-dimethylimidazo[4,5-f]quinoline, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, 2-amino-3-methylimidazo[4,5-f]quinoline, 2-aminofluorene, 6-aminochrysene and its 1,2-dihydrodiol, (-)-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene, 1,2-dihydroxy-1,2-dihydrochrysene, 1,2-dihydroxy-1,2-dihydro-5,6-dimethylchrysene, 2,3-dihydroxy-2,3-dihydrofluoranthene, 3,4-dihydroxy-3,4-dihydro-7,12-dimethylbenz[a]anthracene, and 6-nitrochrysene to appreciable extents. However, P450 1B1 did not produce genotoxic products from benzo[a]pyrene, trans- 3,4-dihydroxy-3,4-dihydrobenzo[a]anthracene, trans-8,9-dihydroxy-8,9-dihydrobenzo[a]anthracene, 7,12-dimethylbenz[a]anthracene and its cis-5,6-dihydrodiol, 5-methylchrysene, 11,12-dihydroxy-11,12-dihydro-3-methylcholanthrene, 1,2-dihydroxy-1,2-dihydro-6-methylchrysene, benzo[c]phenanthrene, 2-amino-6-methyldipyridol[1,2-a:3',2'-d]imidazole, 2-acetylaminofluorene, benzidine, 2-naphthylamine, aflatoxin B1, aflatoxin G1, sterigmatocystin, N-nitrosodimethylamine, vinyl carbamate, or acrylonitrile in this assay system. P450 1B1 is expressed constitutively in extrahepatic organs, including fetal tissue samples, and is highly inducible in various organs by 2,3,7,8-tetrachlorodibenzo-p-dioxin and related compounds in experimental animal models. Thus, activation of procarcinogens by P450 lB1 may contribute to human tumors of extrahepatic origin.     

Differential expression of extracellular matrix and cell adhesion molecules in the olfactory nerve and glomerular layers of adult rats

A series of five 6,7-disubstituted 1,4-dihydro-2,3-quinoxalinediones was prepared, two of which are known microbial flavin metabolites and three of which are potential flavin metabolites. Four of the five compounds inhibited specific binding of [3H]-amino-3-hydroxy-5-methyl-4-isoxazolepropanoic acid ([3H]AMPA), [3H]kainic acid, and [3H]6-cyano-1,4-dihydro-7-nitro-2,3-quinoxalinedione ([3H]CNQX) in rat brain homogenate fractions, with IC50 values in the low micromolar range (the fifth compound competed only with [3H]CNQX). Two of the compounds were moderately potent AMPA antagonists in an in vitro functional test.     

Cross-linking of proteins by Maillard processes--model reactions of D-glucose or methylglyoxal with butylamine and guanidine derivatives

Advanced Maillard reaction in proteins leads to formation of covalently cross-linked aggregates the chemical nature of which is largely unknown. From model reactions of methylglyoxal and butylamine with creatine or alpha-N-acetyl-L-arginine, one main product each was isolated. These two compounds were identified, on the basis of unequivocal spectroscopic evidence, as 2-[(5-butylimino-4-methyl-4,5-dihydro-1H-2-imidazolyl)(methyl)amin o]acetic acid and 2-acetylamino-5-[(5-butylimino-4-methyl-4,5-dihydro-1H-2-imidazoly l)amino]pentanoic acid, respectively. Using D-glucose instead of methylglyoxal, two main products each were obtained from reaction with the respective guanidine derivative. The spectroscopic data definitively establish the formation of the diastereoisomeric 2-[(4-butyl-6,7-dihydroxy-4,5,6,7,8,8a-hexahydroimidazo[4,5-b]a zepin-2-yl)(methyl)amino]acetic acid from the reaction with creatine, and of the diastereoisomeric 2-acetylamino-5-[(4-butyl-6,7-dihydroxy-4,5,6,7,8,8a-hexahydroimidazo [4,5-b]azepin-2-yl)amino]pentanoic acid from the reaction with alpha-N-acetyl-L-arginine. All products were isolated in fairly good yield and represent 1:1:1 adducts of the respective reaction partners. Formation of these compounds thus constitutes an efficient reaction pathway for linking primary amines to guanidine derivatives. It seems justified, therefore, to expect cross-linking of proteins by action of reducing carbohydrates to proceed analogously.     

The in vivo pharmacological profile of the novel glycoprotein IIb/IIIa antagonist, SK&F 106760

The in vivo pharmacological profile of SK&F 106760 [N alpha-acetyl-cyclo(S,S)-cysteinyl-N alpha-methylarginyl-glycyl-aspartyl-penicillamine-amide], a novel, potent glycoprotein IIb/IIIa (GPIIb/IIIa) antagonist has been investigated. In conscious dogs, SK&F 106760 (0.3-3 mg/kg i.v.) produced a dose-related inhibition of ex vivo whole blood platelet aggregation induced by collagen (5 micrograms/ml) with complete inhibition being produced for 5, 90 and 165 min after administration of 0.3, 1 and 3 mg/kg i.v., respectively. Plasma levels of SK&F 106760 were measured by high-performance liquid chromatography after i.v. bolus administration of 1 mg/kg. An initial alpha-disposition phase with a T1/2 of 11 +/- 6 min was followed by a longer terminal beta-elimination phase with a T1/2 of 66 +/- 12 min, which accounted for 79 +/- 9% of the total area under the plasma concentration-time curve. The apparent steady-state volume of distribution was 259 +/- 26 ml/kg and the plasma clearance was 3.4 +/- 0.8 ml/min/kg. The plasma concentration of SK&F 106760 at which collagen-induced ex vivo whole blood aggregation was inhibited by 50% was estimated to be 593 +/- 52 nM. After intraduodenal and intrajejunal administration of 3 mg/kg, SK&F 106760 had a bioavailability of 3 to 6% and produced a peak inhibition of ex vivo platelet aggregation of 40 to 50%. In anesthetized dogs, SK&F 106760 (0.3-3.0 mg/kg i.v.) produced a complete inhibition of platelet-dependent coronary artery thrombosis, with a dose-related duration of action.(ABSTRACT TRUNCATED AT 250 WORDS)     

Novel 5-hydroxytryptamine (5-HT3) receptor antagonists. I. Synthesis and structure-activity relationships of conformationally restricted fused imidazole derivatives

We prepared a novel series of conformationally restricted fused imidazole derivatives 4b, 4c and 4d (possessing 4,5,6,7-tetrahydroimidazo[4,5-c] pyridine and substituted 4,5,6,7-tetrahydro-1H-benzimidazole for 4b, 5,6,7,8-tetrahydroimidazo[1,2-a]pyridine for 4c and 5,6,7,8-tetrahydroimidazo[1,5-a]pyridine for 4d as a basic amine part and (2-methoxyphenyl)aminocarbonyl group as an aromatic-carbonyl part). Their activities were then evaluated as an 5-hydroxytryptamine (5-HT3) receptor antagonist which may be useful for the treatment of irritable bowel syndrome (IBS) as well as for nausea and vomiting associated with cancer chemotherapy. The most potent compound was N-(2-methoxyphenyl)-4,5,6, 7-tetrahydro-1H-benzimidazole-5-carboxamide 14 in this series with an ID50 value of 0.32 microgram/kg on the von Bezold-Jarisch reflex in rats and an IC50 value of 0.43 microM on the isolated colonic contraction in guinea pig, approximately ten and two times more potent than ondansetron 1, respectively. The structure activity relationships (SAR) study suggested that the high potency of 14 may be attributed to the suitable position and direction of the N-C-N centroid in the conformationally restricted imidazole ring against the planar (2-methoxyphenyl)aminocarbonyl part in the binding of 14 to the receptor.     

Geranylgeranyl pyrophosphate synthase encoded by the newly isolated gene GGPS6 from Arabidopsis thaliana is localized in mitochondria

A number of new 1,4-benzodiazepin-2-one-based gastrin/CCK-B receptor antagonists related to the archetypal analogue L-365,260, and more closely to the recently reported compound YM022, have been synthesized and evaluated for biological activity. The compounds were screened for their ability to inhibit the binding of [125I]CCK-8 to gastrin/CCK-B receptors prepared from rat brains and that of [3H]L-364,718 to CCK-A receptors from rat pancreas, and were shown to be potent and selective ligands for the gastrin/CCK-B receptor. Functional studies in vivo demonstrated the compounds to be antagonists of the receptor as evidenced by their ability to inhibit pentagastrin-induced gastric acid secretion in anesthetized rats. More extensive evaluation in vivo included determination of ED50 values in the rat acid secretion model for selected compounds and an examination of the effect of these compounds on pentagastrin-induced gastric acid secretion in Heidenhain pouch dogs following oral and intravenous administration. Two compounds, i.e. (3R)-N-[1-[(tert-butylcarbonyl)methyl]-2,3-dihydro-2-oxo-5-(2-pyri dyl) -1H-1,4-benzodiazepin-3-yl]-N'-[3-(methylamino)phenyl]urea, 15c (YF476), and (3R)-N-[1-[(tert-Butylcarbonyl)methyl]-2,3-dihydro-2-oxo-5- (2-pyridyl)-1H-1,4-benzodiazepin-3-yl]-N'-[3-(dimethylamino)phenyl ]urea hydrochloride, 15d, showed potent dose-dependent effects in both models with the former showing excellent oral bioavailability and an ED50 of 21nmol/kg po in dogs. 15c is currently under clinical investigation for the treatment of gastro-oesophagal reflux disease (GORD).     

Amino acid residue 200 on the alpha1 subunit of GABA(A) receptors affects the interaction with selected benzodiazepine binding site ligands

Mutant alph1 subunits of the GABA(A) receptor were coexpressed in combination with the wild-type beta2 and gamma2 subunits in human embryonic kidney (HEK) 293 cells. The binding properties of various benzodiazepine site ligands were determined by displacement of ethyl-8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5a]-[1,4]benzodia zepine-3-carboxylate ([3H]Ro 15-1788). The mutation G200E led to a decrease in zolpidem and 3-methyl-6-[3-(trifluoromethyl)phenyl]-1,2,4-triazolo[4,3-b]pyridazine (CL 218872) affinity amounting to 16- and 8-fold. Receptors containing a conservative T206V substitution showed a 41- and 38-fold increase in methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM) and CL 218872 affinity combined with a decrease in diazepam and zolpidem affinity, amounting to 7- and 10-fold. Two mutations, Q203A and Q203S showed almost no effects on the binding of benzodiazepine site ligands, indicating that this residue is not involved in the binding of benzodiazepines and related compounds.     

Synthesis and pharmacological activities of ethyl 5-cyano-1,6-dihydro-6-oxo-2-(2,3,4-pyridyl)-3-pyridinecarboxylates and derivatives

The synthesis of ethyl esters of 5-cyano-1,6-dihydro-6-oxo-3-pyridinecarboxylic acids carrying as 2-substituent the 2-,3- or 4-pyridyl group is described. By alkaline hydrolysis followed by acidification, these esters gave the corresponding carboxylic acids, which were decarboxylated to 1,2-dihydro-2-oxo-6-(2,3,4-pyridyl)-3-pyridinecarbonitriles. As milrinone analogues, the above compounds were tested on contractile activity and frequency rate of spontaneously beating atria from reserpine-treated guinea-pigs. Ethyl 5-cyano-1,6-dihydro-6-oxo-2-(2-pyridyl)-3-pyridinecarboxylate showed an appreciable positive inotropic activity, although inferior to that of milrinone; moreover, some other compounds bearing the above 2-substitution pattern showed interesting antiinflammatory, analgesic and hypotensive activity.