The metabolism and disposition of 14C-indolidan, a potent and orally active cardiotonic agent, in four species of laboratory animals

1. The metabolism and disposition of 14C-indolidan, a potent, orally-active positive inotrope with vasodilator properties, has been studied after single dose oral administration to rats, mice, dogs and monkeys. 2. Excretion of 14C in all 4 species was mostly via the urine, largely as parent drug together with two other major metabolites. 3. The two metabolites have been isolated and identified, by mass spectroscopy and 1H-n.m.r., as a dehydro-compound, with a double bond in the pyridazanone ring, and a hydroxylated derivative of the parent drug. 4. Plasma t 1/2 values, based on 14C, were 14 h in dog, 5 h in mouse and 8 h in monkey. Plasma t 1/2 of parent drug, by h.p.l.c. was 10 h in dog, approx. 5 h in rodents, and 8 h in monkeys. 5. Tissue distribution in rats showed no accumulation in any tissue; 14C concn. in all tissues were indistinguishable from background 48 h after dosage. 14C peaked at 6-8 h for most tissues but in blood and plasma, 14C was maximal 1 h after dosing.     

Cardiovascular pharmacology of RS-1893, an orally active cardiotonic agent with arterial and venous vasodilator actions

RS-1893, 2-[2-chloro-4-(2,3,4,5-tetrahydro-3-oxo-6-pyridazinyl)]-phenoxy-N- (2-(2-morpholinoethyl)-acetamide, is a newly synthesized compound whose structure is different from that of cardiac glycosides and beta-stimulants. The in vitro cardiotonic action of RS-1893 was about 3 times more potent than that of milrinone. This action is most likely due to inhibition of phosphodiesterase-III, as has been suggested for many other cardiotonic agents. In pentobarbital anesthetized dogs, RS-1893 (1-30 micrograms/kg, i.v.) produced dose dependent increases in left ventricular dP/dtmax and cardiac output and caused decreases in blood pressure and total peripheral resistance with a relatively small increase in heart rate. Central venous pressure decreased markedly, suggesting venous vasodilation. The in vivo cardiotonic action of RS-1893 was 3 times more potent than that of milrinone and was not affected in the presence of a large dose of propranolol. Oral administration of RS-1893 (0.03 and 0.1 mg/kg) also produced a dose-related increase in cardiac contractility in conscious beagles. The increase in LVdP/dtmax reached a maximum in 1-3 hr after administration and lasted for more than 8 hr. Thus, RS-1893 appeared to be an orally active cardiotonic agent with vasodilator properties, probably acting on both arterioles and veins.     

The metabolic and pharmacokinetic disposition of mebendazole in the rat

The metabolism and pharmacokinetics of mebendazole was studied in rats using [2'-3H]-mebendazole (biologically stable; specific activity 383.9 (mCi/mMol) and [2-14C]-mebendazole (specific activity 2.57 mCi/mMol). Analyses were performed by high pressure liquid chromatography and liquid scintillation spectrometry. About 85% of an intravenous dose was eliminated with the bile and the remainder with the urine. The majority of the dose was recovered as conjugated metabolites. The major metabolite (methyl-5(6)-(alpha-hydroxybenzyl)-2-benzimidazole carbamate) accounted for about 77% of the total recovered and 99% of it was conjugated. Anaerobic metabolism studies conducted in vitro with intestinal microorganisms obtained from rats indicated that metabolism of mebendazole did not occur in the gut, but that the intestinal microflora was able to hydrolyse conjugated metabolites which were eliminated with the bile. Mebendazole was found to have a biphasic elimination profile after intravenous administration. Its terminal plasma elimination half-life was 3.2 hours and its re-distribution half-life was 0.4 hour. After oral administration, as a solution in aqueous dimethyl sulphoxide, a bioavailability of 53% was obtained.     

Details of mode and mechanism of action of denopamine, a new orally active cardiotonic agent with affinity for beta 1-receptors

We investigated the detailed mode and mechanism of action of denopamine in canine right ventricular muscle. When administered in single doses, denopamine produced a positive inotropic effect (PIE) and increased cyclic AMP levels. Concentration--response curves for both variables were sigmoid. The maximum PIE of denopamine was almost the same as that produced by isoproterenol. However, the maximum increase in cyclic AMP caused by denopamine was approximately 65% of that attained with isoproterenol. When administered cumulatively, concentration--PIE curves for denopamine were bell-shaped, ascending at 10(-7) to 10(-6) M, reaching a maximum at approximately 3 X 10(-6) M which was approximately 75% of that attained with single administrations, and descending at higher concentrations. The bell-shaped curve, which was computer-fitted, suggested that denopamine behaves as an agonist with pD2 of 6.12 and as an antagonist with pD2 of 4.50. The PIE of denopamine was antagonized by atenolol (pA2 = 7.66) but not by ICI 118,551 (less than 10(-7) M), indicating that denopamine is a selective beta 1-receptor agonist. The PIE of denopamine was augmented by 3-isobutyl-1-methyl-xanthine. The PIE and increase in cyclic AMP level caused by 3 X 10(-6) M denopamine were abolished by 10(-6) M atenolol or 3 X 10(-6) M carbachol. From these results we concluded that the PIE of denopamine is derived from the mechanism of action as a selective beta 1-receptor partial agonist. We suggested the close coupling of the beta 1-receptor-adenylate cyclase-cyclic AMP system to positive inotropy.     

The metabolic disposition of acifran, a new antihyperlipidemic agent, in rats and dogs

The metabolic disposition of the antihyperlipidemic agent acifran (AY-25, 712) was determined in rats and dogs. The synthesis of 14C-labelled acifran is described. Serum levels of 14C and acifran were measured in rats and dogs after p.o. and i.v. administration of 14C-acifran at a dose of 10 mg/kg. Over 80% of the 14C in serum was due to acifran. The drug was rapidly absorbed and the pharmacokinetics, unaffected by increasing the dose or by daily multiple doses, were characterized by a two-compartment open model. Food reduced the bioavailability of acifran by 27% in the dog. About 65% of the dose was absorbed in rats, and at least 88% in dogs. The elimination t 1/2 of acifran from serum was 1.5 h in the rat and 3 h in the dog. Acifran was partially bound to serum proteins, man greater than rat greater than dog; the drug was found to displace protein-bound warfarin in rat and dog, but not in human serum. Radioactivity did not tend to accumulate in tissues, except for the kidney, where the 14C concentration was five times higher than in the serum; elimination of 14C from all the tissues was similar to that from serum. Most of the absorbed dose was excreted in the urine. Acifran did not undergo enterohepatic circulation in the rat. Virtually all the urinary 14C in both species was due to the unchanged compound. In conclusion, the disposition of acifran was similar in rats and dogs. The drug was rapidly absorbed and eliminated, and underwent no detectable biotransformation. There was no tissue retention and excretion was mainly in the urine.     

Purine nucleoside phosphorylase inhibitor BCX-1777 (Immucillin-H)--a novel potent and orally active immunosuppressive agent.

Patients with purine nucleoside phosphorylase (PNP) deficiency present a selective T-cell immunodeficiency. Inhibitors of PNP are, therefore, of interest as potential T-cell selective immunosuppressive agents. BCX-1777 is a potent inhibitor of PNP from various species including human, mouse, rat, monkey and dog, with IC50 values ranging from 0.48 to 1.57 nM. BCX-1777, in the presence of 2'-deoxyguanosine (dGuo, 3-10 microM), inhibits human lymphocyte proliferation activated by various agents such as interleukin-2 (IL-2), mixed lymphocyte reaction (MLR) and phytohemagglutinin (PHA) (IC50 values < 0.1-0.38 microM). BCX-1777 is a 10-100-fold more potent inhibitor of human lymphocyte proliferation than other known PNP inhibitors like PD141955 and BCX-34. Nucleotide analysis of human lymphocytes indicate that inhibition of proliferation by BCX-1777 correlates with dGTP levels in the cells. BCX-1777 has excellent oral bioavailability (63%) in mice. At a single dose of 10 mg/kg in mice, BCX-1777 elevates dGuo to approximately 5 microM. BCX-1777 was not effective in mouse T-cell models such as delayed type hypersensitivity (DTH) and splenomegaly because mouse T-cells do not accumulate dGTP as do human T-cells. However, in the human peripheral blood lymphocyte severe combined immunodeficiency (hu-PBL-SCID) mouse model, BCX-1777 was effective in prolonging the life span 2-fold or more. This is the first known example of a PNP inhibitor that elevates dGuo in mice similar to the levels observed in PNP-deficient patients. Furthermore, these dGuo levels are also required for in vitro T-cell inhibition by BCX-1777. Thus, BCX-1777 represents a novel class of selective immunosuppressive agents that could have therapeutic utility in various T-cell disorders.     

The non-psychoactive cannabis constituent cannabidiol is an orally effective therapeutic agent in rat chronic inflammatory and neuropathic pain

Cannabidiol, the major psycho-inactive component of cannabis, has substantial anti-inflammatory and immunomodulatory effects. This study investigated its therapeutic potential on neuropathic (sciatic nerve chronic constriction) and inflammatory pain (complete Freund's adjuvant intraplantar injection) in rats. In both models, daily oral treatment with cannabidiol (2.5-20 mg/kg to neuropathic and 20 mg/kg to adjuvant-injected rats) from day 7 to day 14 after the injury, or intraplantar injection, reduced hyperalgesia to thermal and mechanical stimuli. In the neuropathic animals, the anti-hyperalgesic effect of cannabidiol (20 mg/kg) was prevented by the vanilloid antagonist capsazepine (10 mg/kg, i.p.), but not by cannabinoid receptor antagonists. Cannabidiol's activity was associated with a reduction in the content of several mediators, such as prostaglandin E(2) (PGE(2)), lipid peroxide and nitric oxide (NO), and in the over-activity of glutathione-related enzymes. Cannabidiol only reduced the over-expression of constitutive endothelial NO synthase (NOS), without significantly affecting the inducible form (iNOS) in inflamed paw tissues. Cannabidiol had no effect on neuronal and iNOS isoforms in injured sciatic nerve. The compound's efficacy on neuropathic pain was not accompanied by any reduction in nuclear factor-kappaB (NF-kappaB) activation and tumor necrosis factor alpha (TNFalpha) content. The results indicate a potential for therapeutic use of cannabidiol in chronic painful states.     

Metabolism of denopamine, a new cardiotonic agent, in the rat and dog

The metabolism of denopamine, (R)-(-)-1-(p-hydroxyphenyl)-2-[(3,4-dimethoxyphenethyl)amino] ethanol, a new, orally active, selectively inotropic cardiotonic agent, was studied in the rat and dog. Animals were given single oral doses of 5 mg/kg of denopamine labeled with 14C. Denopamine was metabolized in the rat and dog by several pathways including conjugation, side chain oxidation, and ring hydroxylation followed by O-methylation. Rats excreted the drug in the urine almost entirely as unchanged drug and its phenolic O-glucuronide whereas in the dog, the major metabolites were the phenolic O-glucuronide, the alcoholic O-glucuronide, and the phenolic O-sulfate of denopamine and the phenolic O-glucuronide of 3-methoxydenopamine. Demethylation, which has been shown to be a major metabolic pathway in man, and side chain oxidation were minor pathways in the rat and dog. Furthermore, a high degree of stereoselective resistance of the alcoholic O-glucuronide of denopamine to hydrolysis by beta-glucuronidase was observed.     

The metabolic disposition of C-labelled carmoisine in the rat after oral and intravenous administration

The absorption, distribution and excretion of the red azo dye carmoisine (Ext. D & C No. 10) was studied in male rats. [14C]Carmoisine was administered in a dose of 200 mg/kg (25 microCi) by gavage or in the same dose (200 mg/kg; 3 microCi) by intravenous injection, and radioactivity was measured in blood, tissue, faeces and urine at different times after dosing. After oral administration of the dye, no radioactivity was detected in the brain, adipose tissue, muscle, testes, spleen or lung, and recovery of the administered activity in faeces and urine was almost complete by 32 hr. The radioactivity profile of the blood indicated rapid but poor absorption of [14C]carmoisine, a maximum radioactivity content corresponding to 0.01% of the dose per ml of blood being reached within 10 min. The decay curve for 14C radioactivity in the blood after iv injection of [14C]carmoisine indicated rapid distribution to the tissues and could be described in terms of a two-compartment mathematical model. The highest levels of radioactivity occurred in the gastro-intestinal tract and liver after the injection but after 24 hr no radioactivity was detectable in these or other tissues. All the radioactivity was recovered in the faeces and urine in the 24 hr following iv injection, the 79% of the dose present in faeces indicating active excretion of the dye and its metabolites in the bile and poor reabsorption from the intestine. The bioavailability of [14C]carmoisine, calculated from the blood-radioactivity curves after oral and iv administration, was less than 10%.     

The murine disposition and pharmacokinetics of the antineoplastic agent, diaziquone (NSC 182986)

We have studied the murine disposition and pharmacokinetics of diaziquone (AZQ), a new aziridinylbenzoquinone antitumor agent that is currently undergoing clinical trials. 14C-AZQ, dissolved in dimethylacetamide/0.01M phosphate buffer, pH 6.5, 5:95 (v/v), was administered iv to mice at a dosage of 6 mg/kg (18 mg/m2). After injection, mice were killed and plasma and organs were obtained and analyzed for total radioactivity and for chloroform-extractable 14C. Both total plasma radioactivity and chloroform-extractable plasma 14C declined very rapidly. By 3 hr, chloroform-extractable 14C was about 0.2% of its initial concentration. Within 30 min after injection, more radioactivity remained in the aqueous phase than was extracted into chloroform. When analyzed by TLC, all chloroform-extractable radioactivity in plasma as well as in brain, heart, liver, and skeletal muscle co-chromatographed with parent AZQ. Tissue distribution of 14C was extensive. 14C concentrations (dpm/g tissue wet weight) were initially greatest in heart and lung, but by 2 min after injection, were greatest in kidney, and by 10 min were second greatest in liver. Substantial amounts of total and chloroform-extractable 14C were found in brains. By 3 hr after injection, total 14C in all tissues exceeded total 14C in plasma. The apparent volume of distribution of AZQ was approximately 1 ml/g and the total body clearance of AZQ was 0.9 ml/min or 130 ml/min/m2. Extraction of tissues with chloroform showed conversion of AZQ into non-chloroform-extractable metabolites. This conversion was reproduced in vitro by mouse liver homogenate at 37 degrees C but not at 0 degree C.     

Discovery and development of GS 4104 (oseltamivir): an orally active influenza neuraminidase inhibitor

Rational drug design utilizing available X-ray crystal structures of sialic acid analogues bound to the active site of influenza virus neuraminidase has led to the discovery of a series of potent carbocyclic influenza neuraminidase inhibitors. From this series, GS 4104 (oseltamivir, TAMIFLU ) has emerged as a promising antiviral for the treatment and prophylaxis of human influenza infection. This article will summarize the design, discovery, and development of oseltamivir as an oral therapeutic to treat influenza infection.     

The metabolic disposition of C-labelled Ponceau 4R in the rat, mouse and guinea-pig

The absorption, metabolism and excretion of ¹⁴C-labelled Ponceau 4R has been studied in the rat, mouse and guinea-pig. Following administration of a single oral dose of 0·5 or 50 mg/kg body weight substantially all of the dose was excreted in the urine and faeces within 72 hr, with the majority being accounted for in the faeces. In all three species, naphthionic acid was the major urinary metabolite, whereas in the faeces naphthionic acid, 7-hydroxy-8-aminonaphthalene-1,3-disulphonic acid and unchanged dye were found. Pretreating male rats with unlabelled Ponceau 4R in the diet (50 mg/kg/day) for 28 days prior to dosing with the ¹⁴C-labelled colouring had no effect on the route of excretion or the time taken to eliminate the majority of the label. Following a single dose of ¹⁴C-labelled colouring to previously untreated rats, mice and guinea-pigs or to rats pretreated as above, no marked accumulation of radioactivity in any tissue was found, although tissue levels of radioactivity at 72 hr after dosing were higher in the pretreated rats than in those that were not pretreated. Pregnant rats eliminated a single oral dose of ¹⁴C-Iabelled colouring at a similar rate to non-pregnant females; however, some retention of radioactivity in the foetuses was found. In studies of absorption from isolated loops of small intestine containing 50, 500 or 5000 ppm Ponceau 4R, no significant absorption was detected in rats, but some absorption was seen in mice at the lowest concentration, and in the guinea-pig at the two higher concentrations.     

The disposition and toxicology of retinyl methyl ether in rats dosed orally

In disposition studies, retinyl methyl ether (RME) was administered to rats in oral doses of 10 or 40 mg/kg. For the high dose, RME was eliminated from plasma with a terminal half-life of 19.5 hr but for the low dose the terminal phase could not be determined. For both doses, the concentrations of RME in the tissues examined (liver, spleen, adrenals, and mammary glands) were greater than those in plasma. In the adrenals of rats given the low dose, concentrations were as much as 10- to 100-fold higher. Concentrations of RME in the mammary gland, a site for chemopreventive activity, were also relatively high (about 1000 ng/g for the low dose and about 4000 ng/g for the high dose), and there was an elimination phase with a half-life of 63-81 hr. After administration of RME, the concentration of retinyl esters in the liver did not increase, and no retinyl esters were detected in the mammary gland. For toxicology studies, rats were administered 20, 40, and 80 mg/kg of RME or retinyl acetate (ROAc) daily for 28 days. The toxic effects of RME were similar to those of ROAc. At equivalent mg/kg doses, weight gain depressions, bone fractures, elevations in serum triglycerides, anemia, elevations in cholesterol in females, and reductions in serum albumin were similar.(ABSTRACT TRUNCATED AT 250 WORDS)     

(Pyridylcyanomethyl)piperazines as orally active PAF antagonists.

A series of (pyridylcyanomethyl)piperazines was prepared and evaluated for PAF-antagonist activity. Compounds were tested in vitro in a PAF-induced platelet aggregation assay and in vivo in a PAF-induced hypotension test in normotensive rats. Oral activity was ascertained through a PAF-induced mortality test in mice. The main structure-activity trends of the series were established. Activity was mainly found in four skeletons: 1-acyl-4-(3-pyridylcyanomethyl)piperazine, 1-acyl-4-(4-pyridylcyanomethyl)piperazine, 1-acyl-4-(3-pyridylcyanomethyl)piperidine, and 1-acyl-4-cyano-4-(3-pyridylamino)piperidine. The acyl substituents, diphenylacetyl and 3,3-diphenylpropionyl, provided the most active compounds, and the introduction of an amine or hydroxy group in the 3,3-diphenylpropionyl substituent led to further improvement in oral activity. As a result, three of the most active compounds (100, 114, and 115) have been selected for further pharmacological development.     

The disposition of the hypolipidemic agent, o-(N-phthalimido)acetophenone, in Sprague-Dawley rats

Disposition studies of the potent experimental hypolipidemic agent, o-(N-phthalimido)acetophenone, were conducted in the laboratory rat. Intravenous administration of the drug demonstrated a declining biphasic plasma concentration-time curve suggesting a rapid distribution into tissues. Less than 5% of the intravenous dose was recovered in urine and feces after 5 days. Oral administration of the drug showed that the maximum blood levels were attained within 15 min. After 48 hr, 92% of the orally administered 14C dose was eliminated either via urine (60%) or feces (40%). 14C in a 0-24-hr urine collection after oral administration showed that urinary radioactivity was composed of 22% of the parent compound o-(N-phthalimido) acetophenone, a benzoic acid metabolite (22%) o-(N-phthalimido)benzoic acid, and an amic acid metabolite (56%) N-(o-acetophenone)phthalamic acid. The two metabolites were unconjugated and possessed less hypolipidemic activity than the parent drug.     

The metabolic disposition of 14C-ciramadol in humans

Twelve subjects received single 15 mg oral doses of 14C-ciramadol. Excretion of the dose occurred almost entirely by the renal route (93.5 +/- 11.7 (S.D.)% of the dose), and only 0.7 +/- 0.6% of the dose was recovered in faeces indicating that absorption was essentially complete. More than 90% of the amount recovered in urine was excreted within 24 h after dosing. Unchanged drug accounted for 43.9 +/- 6.5% of the dose, while a phenolic glucuronide conjugate was the only major urinary metabolite accounting for a further 37.9 +/- 7.8%. A second glucuronide that was conjugated with the alicyclic ring was also identified but constituted only 2.3 +/- 0.6% of the dose. Concentrations of radioactivity in plasma reached a peak at 2 h after dosing and declined with a terminal disposition half life of 4.9 h. Only ciramadol and the aryl-O-glucuronide were detected in substantial amounts in plasma. Renal clearance of ciramadol amounted to 298 +/- 54 ml/min suggesting tubular secretion in addition to glomerular filtration.     

The metabolic disposition of 14C-labelled Ponceau 4R in the rat,mouse and guinea-pig

The absorption, metabolism and excretion of ¹⁴C-labelled Ponceau 4R has been studied in the rat, mouse and guinea-pig. Following administration of a single oral dose of 0·5 or 50 mg/kg body weight substantially all of the dose was excreted in the urine and faeces within 72 hr, with the majority being accounted for in the faeces. In all three species, naphthionic acid was the major urinary metabolite, whereas in the faeces naphthionic acid, 7-hydroxy-8-aminonaphthalene-1,3-disulphonic acid and unchanged dye were found. Pretreating male rats with unlabelled Ponceau 4R in the diet (50 mg/kg/day) for 28 days prior to dosing with the ¹⁴C-labelled colouring had no effect on the route of excretion or the time taken to eliminate the majority of the label. Following a single dose of ¹⁴C-labelled colouring to previously untreated rats, mice and guinea-pigs or to rats pretreated as above, no marked accumulation of radioactivity in any tissue was found, although tissue levels of radioactivity at 72 hr after dosing were higher in the pretreated rats than in those that were not pretreated. Pregnant rats eliminated a single oral dose of ¹⁴C-Iabelled colouring at a similar rate to non-pregnant females; however, some retention of radioactivity in the foetuses was found. In studies of absorption from isolated loops of small intestine containing 50, 500 or 5000 ppm Ponceau 4R, no significant absorption was detected in rats, but some absorption was seen in mice at the lowest concentration, and in the guinea-pig at the two higher concentrations.     

Tolvaptan, an orally active vasopressin V(2)-receptor antagonist - pharmacology and clinical trials

Tolvaptan is an orally effective nonpeptide arginine vasopressin (AVP) V(2)-receptor antagonist synthesized by Otsuka Pharmaceutical Co., Ltd. In in vitro receptor-binding studies, tolvaptan blocked the binding of [(3)H]AVP to human V(2) receptors with 29-fold greater selectivity than that for V(1a) receptors, and showed no inhibition of V(1b) receptors. Tolvaptan inhibited not only the binding of [(3)H]AVP but also the AVP-induced production of cyclic AMP in human V(2)-receptor-expressing HeLa cells. In addition, tolvaptan has no intrinsic V(2) receptor agonistic effect. In in vivo studies, tolvaptan showed marked aquaresis in healthy and diseased animals. In rat models with acute and chronic hyponatremia, tolvaptan improved hyponatremia, resulting in the prevention of death, and improved organ water retention. Tolvaptan reduced cardiac preload without unfavorable effects on renal functions, systemic hemodynamics, or circulating neurohormones in dogs with heart failure (HF). Furthermore, in animal models of human polycystic kidney disease (PKD), tolvaptan showed a decrease in kidney weight as well as in cyst and fibrosis volume. In clinical trials including the "ACTIV in CHF" study, tolvaptan in addition to standard therapy increased fluid loss resulting in decreased body weight, and improved edema and serum sodium without affecting blood pressure, heart rate, or renal functions in patients with HF. In patients with hyponatremia, treatment with tolvaptan without fluid restriction appeared to be more effective than fluid restriction alone at correcting hyponatremia without an increase in adverse events. A phase III trial EVEREST is currently being conducted to evaluate the long-term efficacy and safety of tolvaptan in hospitalized patients with severe HF. In conclusion, tolvaptan offers the possibility of a useful therapy in hyponatremia, congestive heart failure, and various other diseases that are associated with volume overload. Furthermore, tolvaptan is also expected to be effective in the treatment of PKD.     

Cardiovascular effects of the novel cardiotonic agent DPI 201-106 in the anaesthetized rat

DPI 201-106 (4-[3-(4-diphenylmethyl-1-piperazinyl)-2-hydroxypropoxy]-1H-indole -2- carbonitrile) was given intravenously to anaesthetized male rats. DPI caused an increase in left ventricular dP/dt (LV dP/dt), giving a significant increase at 0.03 mumol/kg. At this dose DPI had no effect on either mean arterial pressure (MAP) or heart rate (HR). At higher doses, MAP decreased transiently. At 0.3 and 1 mumol/kg, HR was decreased. The results indicate that DPI produces positive inotropic and negative chronotropic effects in the anaesthetized rat.