Kristian Tage Hansen

PET examination of [11C]NNC 687 and [11C]NNC 756 as new radioligands for the D1-dopamine receptor

The benzazepines NNC 687 and NNC 756 have in animal studies been described as selective D1-dopamine receptor antagonists. Both compounds have been labeled with11C for examination by positron emission tomography (PET). In the present study central receptor binding was studied in monkeys and healthy men. After IV injection of both radioligands in Cynomolgus monkeys radioactivity accumulated markedly in the striatum, a region with a high density of D1-dopamine receptors. This striatal uptake was displaced by high doses of the selective D1-antagonist SCH 23390 (2 mg/kg) but not by the 5HT2-antagonist ketanserin (1.5 mg/kg) or the selective D2-antagonist raclopride (3 mg/kg). The cortical uptake after injection of [11C]NNC 687 was not reduced in displacement experiments with ketanserin. The cortical uptake of [11C]NNC 756 was reduced in displacement and protection experiments with ketanserin by 24–28% (1.5 mg/kg), whereas no reduction could be demonstrated on striatal uptake. In healthy males both compounds accumulated markedly in the striatum. For [11C]NNC 687 the ratio of radioactivity in the putamen to cerebellum was about 1.5. For [11C]NNC 756 the ratio was about 5. This ratio of 5 for [11C]NNC 756 is the highest obtained so far for PET radioligands for the D1-dopamine receptor.

Influence of Drugs Interacting with CYP3A4 on the Pharmacokinetics, Pharmacodynamics, and Safety of the Prandial Glucose Regulator Repaglinide

The object of this study was to analyze drug interactions between repaglinide, a short‐acting insulin secretagogue, and five other drugs interacting with CYP3A4: ketoconazole, rifampicin, ethinyloestradiol/levonorgestrel (in an oral contraceptive), simvastatin, and nifedipine. In two open‐label, two‐period, randomized crossover studies, healthy subjects received repaglinide alone, repaglinide on day 5 of ketoconazole treatment, or repaglinide on day 7 of rifampicin treatment. In three open‐label, three‐period, randomized crossover studies, healthy subjects received 5 days of repaglinide alone; 5 days of ethinyloestradiol/levonorgestrel, simvastatin, or nifedipine alone; or 5 days of repaglinide concomitant with ethinyloestradiol/levonorgestrel, simvastatin, or nifedipine. Compared to administration of repaglinide alone, concomitant ketoconazole increased mean AUC0‐∞ for repaglinide by 15% and mean Cmax by 7%. Concomitant rifampicin decreased mean AUC0‐∞ for repaglinide by 31% and mean Cmax by 26%. Concomitant treatment with CYP3A4 substrates altered mean AUC0–5h and mean Cmax for repaglinide by 1% and 17% (ethinyloestradiol/levonorgestrel), 2% and 27% (simvastatin), or 11% and 3% (nifedipine). Profiles of blood glucose concentration following repaglinide dosing were altered by less than 8% by both ketoconazole and rifampicin. In all five studies, most adverse events were related to hypoglycemia, as expected in a normal population given a blood glucose regulator. The safety profile of repaglinide was not altered by pretreatment with ketoconazole or rifampicin or by coadministration with ethinyloestradiol/levonorgestrel. The incidence of adverse events increased with coadministration of simvastatin or nifedipine compared to either repaglinide or simvastatin/nifedipine treatment alone. No clinically relevant pharmacokinetic interactions occurred between repaglinide and the CYP3A4 substrates ethinyloestradiol/levonorgestrel, simvastatin, or nifedipine. The pharmacokinetic profile of repaglinide was altered by administration of potent inhibitors or inducers, such as ketoconazole or rifampicin, but to a lesser degree than expected. These results are probably explained by the metabolic pathway of repaglinide that involves other enzymes than CYP3A4, reflected to some extent by a small change in repaglinide pharmacodynamics. Thus, careful monitoring of blood glucose in repaglinide‐treated patients receiving strong inhibitors or inducers of CYP3A4 is recommended, and an increase in repaglinide dose may be necessary. No safety concerns were observed, except a higher incidence in adverse events in patients receiving repaglinide and simvastatin or nifedipine.

Xanomeline: A selective muscarinic agonist for the treatment of Alzheimer's disease

Xanomeline is a novel muscarinic receptor agonist relatively devoid of parasympathomimetic side effects. Xanomeline had high affinity for muscarinic receptors and much lower affinity for a variety of other neuronal receptors in radioligand binding assays. Functional studies in cell lines transfected with the muscarinic receptor subtypes demonstrated that xanomeline had higher potency and efficacy for m1 and m4 receptors than m2, m3, and m5 receptor subtypes. Similarly, in isolated tissue studies, xanomeline had higher potency and efficacy for M1 receptors in rabbit vas deferens than at M2 receptors in guinea pig atria or M3 receptors in guinea pig bladder. Secretion of soluble amyloid precursor protein from m1 cell lines was potently stimulated by xanomeline. In vivo, xanomeline robustly stimulated phosphoinositide hydrolysis in brain, consistent with m1 agonism. Xanomeline produced modest increases in brain acetylcholine levels and did not produce bradycardia, suggesting little, if any, m2 agonist activity in vivo. Additionally, xanomeline did not induce nonselective cholinergic agonist side effects such as tremor, hypothermia and salivation. In animal behavior studies, xanomeline reduced locomotion and blocked memory deficits that were induced by a muscarinic antagonist in a passive avoidance paradigm. Xanomeline was found to be safe and reasonably well tolerated in safety studies in humans. In a placebo controlled double blind clinical trial of 6 months duration, xanomeline halted cognitive decline in patients with Alzheimers disease. Furthermore, behavioral symptoms associated with Alzheimers disease such as hallucinations, delusions and vocal outbursts were significantly decreased by xanomeline treatment. Additional clinical trials are under way to assess the novel therapeutic effects of xanomeline. Drug Dev. Res. 40:158–170, 1997.

Pig hepatocytes as an in vitro model to study the regulation of human CYP3A4: prediction of drug-drug interactions with 17α-ethynylestradiol

The objective of this study was to provide evidence of the validity of pig hepatocytes as a model to study the regulation of human CYP3A4 with special emphasis on drug-drug interactions. Thirteen different drugs were incubated with primary monolayer cultures of pig hepatocytes (n = 4). The study included both drugs reported to cause drug interactions in the clinic with 17 alpha-ethynylestradiol (EE2), other drugs metabolized by CYP3A4, and drugs not reported to cause any problems. Effect of the drug exposure to pig hepatocytes was determined by immunodetection using a monoclonal human CYP3A4 antibody and measurement of 6 beta-hydroxylation of testosterone and 2-hydroxylation of 17 alpha-ethynylestradiol (EE2), both reactions known to be catalyzed by CYP3A4 in humans. Data were compared to data from human hepatocytes and to reported observations of drug-drug interactions in the clinic. The drugs known to be inducers of CYP3A4 in humans significantly increased a CYP isoform in pigs catalyzing 6 beta-hydroxylation of testosterone and 2-hydroxylation of EE2, whereas drugs not reported to have clinical interactions with EE2 had no or only marginal effect. Induction by the drugs known to be inducers of CYP3A4 increased with drug exposure time and the CYP3A4 activity, represented by testosterone 6 beta-hydroxylation, was highest at 72 h for the investigated induction periods (24, 48 and 72 h), except for dexamethasone where the effect peaked after 24 h. Induction of the 2-hydroxylation of EE2 correlated well with the increase in 6 beta-hydroxylation of testosterone (except for sulphinpyranzone) and the increase in the protein level of CYP3A detected by a monoclonal human CYP3A4 antibody, thus confirming the 2-hydroxylation of EE2 in pigs as being biotransformed by a CYP isoform presumably belonging to the CYP3A subfamily as in humans. In conclusion, these results indicate that pig hepatocytes may be a valuable model to mimic the regulation of human CYP3A4.

[11C]NNC 687 and [11C]NNC 756, dopamine D-1 receptor ligands. Preparation, autoradiography and PET investigation in monkey.

NNC 687 and NNC 756 [(+)-5-(2,3-dihydrobenzofuran-7-yl)-7-hydroxy-3-methyl- 8-nitro-2,3,4,5-tetrahydro-1H-3-benzazepine and (+)-8-chloro-5-(2,3-dihydrobenzofuran-7-yl)-7-hydroxy-3-methyl-2,3,4,5- tetrahydro-1H-3-benzazepine] are two new potent dopamine D-1 receptor antagonists. [11C]NNC 687 and [11C]NNC 756 were both prepared by N-methylation of the corresponding desmethyl compounds with [11C]methyl iodide. The reactions were performed in acetone with subsequent normal-phase semi-preparative HPLC and resulting in 50-60% radiochemical yield (from EOB and decay-corrected) with a total synthesis time of 30-35 min and a radiochemical purity higher than 99%. The specific radioactivity obtained at time of injection was about 1500 Ci/mmol (55 GBq/mumol). Autoradiographic examination of [11C]NNC 687 and [11C]NNC 756 binding in post-mortem human brain sections showed specific binding in the striatum, a region with high density of dopamine D-1 receptors. PET examination of the radioligands in a Cynomolgus monkey demonstrated accumulation of radioactivity predominantly in the striatum. The ratio between radioactivities in the striatum and the cerebellum was about 2 and 8 for [11C]NNC 687 and [11C]NNC 756 after 60 min. [11C]NNC 756 should have potential as PET ligand for examination of central dopamine D-1 receptors in man.

Prodrugs as drug delivery systems. 77. Phthalidyl derivatives as prodrug forms for amides, sulfonamides, carbamates and other NH-acidic compounds

Abstract A series of N -phthalidyl derivatives of various carboxamides, sulfonamides, a carbamate and a urea has been prepared with the aim of assessing their potential as prodrug forms for NH-acidic compounds. The hydrolysis of the compounds was studied in aqueous solution at various pH values and in the presence of human plasma. The degradation was shown to take place by hydrolytic opening of the lactone ring with the formation of an N -hydroxyalkyl intermediate which quickly decomposed to the parent NH-acidic compound and phthalaldehydic acid. This hydrolysis was catalyzed by plasma enzymes. The phthalidyl derivative of a primary sulfonamide behaved differently as it was extremely unstable in aqueous solution, the decomposition proceeding via an elimination-addition mechanism. It is concluded that in contrast to unstable linear N -acyloxyalkyl derivatives, phthalidyl derivatives in which the ester function is incorporated in a lactone group may be useful as prodrug forms for primary as well as secondary amides and other similar NH-acidic compounds, the exception being primary sulfonamides.

Enhanced bioavailability of a new class of dopamine D-1 antagonists following oral administration of their carbamic acid ester prodrugs to dogs

Using specific high-pressure liquid chromatography (HPLC) methods, the pharmacokinetics of NNC 0112 and NNC 0756 were studied in mongrel dogs. Both compounds have a low oral bioavailability of 5.4 ± 3.6% (mean ± SD, n = 4) and 6.0 ± 0.5% (n = 4), respectively, due to a large first-pass metabolism. Both compounds were rapidly cleared from the body with a total body clearance of 26.0–26.5 ml/min per kg, and terminal elimination half-lives of approx. 2 h. The high first-pass metabolism could be reduced using various mono- and disubstituted carbamate ester prodrugs previously characterized in vitro. The isopropyl monosubstituted carbamate ester increased the oral bioavailability of NNC 0112 3-fold, whereas the N,N-dimethyl-substituted carbamate ester improved the bioavailability to approx. 20%. The dimethyl- and diethyl-substituted carbamate esters of NNC 0756 increased the bioavailability to 30 and 14%, respectively, and a novel prodrug form based on the methyl ester of N-methylalanine improved the oral bioavailability of NNC 0756 approx. 3-fold.

Hepatic uptake and biliary excretion of the neuroprotectant 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo-(F)-quinoxaline in rat liver. Involvement of an organic anion transport system.

The hepatic uptake and biliary excretion of the neuroprotective drug 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo-(f)-quinoxaline (NBQX) was studied in rat liver. In the isolated single-pass perfused rat liver preparation NBQX was infused in protein-free Krebs-Henseleit bicarbonate buffer at input concentrations ranging from 0.5 to 15 microM. The hepatic uptake could be characterized as a pseudo first-order unidirectional process with an apparent half-life of 5.8 min. Significantly higher concentrations of NBQX were measured in bile compared to perfusate with a maximal ratio at the lowest input concentration (approx. 2400-fold). Hepatic uptake and biliary excretion of NBQX exhibited saturation at increasing input concentrations, indicating an active transport mechanism. The uptake process could be described by Michaelis-Menten kinetics resulting in a Km,uptake of 2.2 microM. The corresponding maximal uptake rate (Vmax,uptake) was calculated to be 103 nmol/min. The maximal biliary excretion rate was estimated to be 58 nmol/min. The rate-limiting factor in the overall hepatic elimination was thus biliary excretion. Co-infusion with the uricosuric drug probenecid resulted in significantly decreased hepatic uptake and biliary excretion. These data suggest that NBQX is eliminated by an organic anion transport system in rat liver which is sensitive to probenecid.