Ligand metabolites in plasma during PET-studies with the 11C-labelled dopamine antagonists,raclopride, SCH 23390 and N-methylspiroperidol

The ¹¹C-labelled radioligands raclopride, SCH 23390 and N-methyl-spiroperidol are well established for examination of central dopamine receptors by positron emission tomography (PET). Thin layer chromatography was used to examine the composition of radioactivity in plasma after intravenous injection of any of these three ligands into human subjects. For all three ligands there was a considerable metabolism during the time of a PET-experiment but to a different degree. Forty-two minutes after injection (¹¹C)raclopride was unchanged to 76 per cent, (¹¹C)N-methylspiroperidol to 57 per cent and (¹¹C) SCH 23390 only to 13 per cent. A time curve for ligand metabolism is necessary for compartmental analysis with an arterial input function. The considerable ligand metabolism demonstrated shortly after i.v. injection motivates further the identification of main metabolites to evaluate if they pass the blood-brain barrier and bind to the receptors.     

Evaluation of SCH 39166 as PET ligand for central D1 dopamine receptor binding and occupancy in man

SCH 39166 is the first selective D₁-dopamine receptor antagonist developed for clinical trials in schizophrenia. SCH 39166 was evaluated as a radioligand for PET, labeled with¹¹C, and as a D₁-dopamine receptor antagonist after single oral doses in healthy men. After intravenous injection of [¹¹C]SCH 39166 distribution of radioactivity in brain grossly reflected D₁-dopamine receptor density. The putamen to cerebellum ratio at equilibrium was low (1.54±0.18 SD), which makes [¹¹C]SCH 39166 less suitable as a radioligand for applied PET studies. Saturability of specific binding was demonstrated after IV injection of [¹¹C]SCH 39166 with low specific radioactivity. Stereospecificity of binding was examined using the stereoisomer [¹¹C]SCH 39165. D₁-Receptor occupancy was demonstrated with [¹¹C]SCH 39166 2 h after administration of single oral doses of unlabeled SCH 39166 to each of three healthy subjects (25, 100 and 400 mg). There was a substantial reduction of specific [¹¹C]SCH 39166 uptake in the putamen after all doses. Single oral doses of 100 mg induced approximately 70% D₁-dopamine receptor occupancy in the basal ganglia, which should be sufficient to investigate the antipsychotic potential of D₁-dopamine receptor antagonism in clinical studies.     

Selective D1- and D2-dopamine receptor blockade both induces akathisia in humans — a PET study with [11C]SCH 23390 and [11C]raclopride

Pharmacological effects were recorded and time course for receptor binding in brain was followed by positron emission tomography after IV injection of the selective D₁-dopamine receptor antagonist SCH 23390 in four healthy subjects in doses of 310–810 µg. Akathisia, the syndrome of motor restlessness, appeared after the three highest doses. The akathisia was transient and occurred only when [¹¹C]SCH 23390 binding in the basal ganglia was at a high level with a central D₁-dopamine receptor occupancy of 45–59%. The D₂-dopamine receptor antagonist [¹¹C]raclopride was injected IV into 20 healthy subjects and 13 schizophrenic patients. Akathisia appeared in 14 healthy subjects and 7 patients and coincided with maximal [¹¹C]raclopride binding in the basal ganglia. The findings for [¹¹C]raclopride and [¹¹C]SCH 23390 are the first demonstration of a relationship between time courses for radioligand binding in the human brain and simultaneously induced pharmacological effects.     

Characterization of C-11 or I-123 Labelled β-CIT-FP and β-CIT-FE Metabolism Measured in Monkey and Human Plasma. Identification of Two Labelled Metabolites with HPLC

β-CIT-FP and β-CIT-FE which are novel fluoroalkyl derivatives of 2β-carbomethoxy-3β-(4-iodophenyl)tropane (β-CIT), have recently been labelled with¹¹C or¹²³I and used for imaging the dopamine transporter with positron emission tomography (PET) or single photon emission tomography (SPET). The metabolite pattern of these novel tracers was studied with HPLC in plasma in monkeys and in urine and plasma in humans. Four labelled metabolites were found of which two were identified, the¹²³I-labelled free acid and¹¹C- or¹²³I-labelled nor-β-CIT. The latter was detected only in small amounts (<4 per cent). Furthermore, a polar metabolite and an unknown lipophilic¹¹C- or¹²³I-labelled metabolite were found. The amount of this unidentified labelled metabolite of [¹¹C]β-CIT-FP or [¹¹C]β-CIT-FE in monkey plasma was 21±8 per cent and 16±9 per cent at 60 min after injection, respectively. In conclusion, four types of labelled metabolites were found after injection of¹¹C- or¹²³I-labelled β-CIT-FP and β-CIT-FE in monkeys and humans of which one may hamper the quantitation of the dopamine transporter by PET or SPET.     

Synthesis and PET evaluation of (R)‐[S‐methyl‐11C]thionisoxetine,a candidate radioligand for imaging brain norepinephrine transporters

Introduction: (R)‐3‐(2‐(methylthio)phenoxy)‐N‐methyl‐3‐phenylpropan‐1‐amine [(R)–thionisoxetine; 1 ] is a potent inhibitor of the norepinephrine transporter (NET). We aimed to label 1 with carbon‐11 (t_1/2 = 20.4 min) for evaluation as a radioligand for imaging NET in living brain with positron emission tomography (PET). Methods: Methyl 3‐(2‐((R)‐3‐(methylamino)‐1‐phenylpropoxy)phenylthio)‐propanoate (MPPP) and 1 were each prepared from o‐hydroxythiophenol in three steps. Treatment of MPPP with potassium t‐butoxide and [¹¹C]methyl iodide in tetrahydrofuran gave [S‐methyl‐¹¹C]thionisoxetine ([¹¹C] 1 ), which was purified with HPLC. The distribution of radioactivity in brain after intravenous injection of [¹¹C] 1 into cynomolgus monkey was followed with PET and the appearance of radiometabolites in plasma monitored with radio‐HPLC. Results: [¹¹C] 1 was obtained in high yield from [¹¹C]methyl iodide. Of the radioactivity injected into monkey, 2.4% entered brain. Ratios of radioactivity in thalamus, mesencephalon, occipital cortex and caudate to that in cerebellum at 93 min were 1.3, 1.2, 1.2 and 1.1, respectively. The radioactivity in plasma corresponding to unchanged radioligand decreased to 53% at 45 min, with the remainder represented by hydrophilic radiometabolites. Conclusions: MPPP is an effective precursor for ¹¹C‐methylation to [¹¹C] 1 , suggesting that the S‐γ‐propionic acid methyl ester protecting group may have wider value in the ¹¹C‐labeling of aryl methyl sulfides. However, the relatively low ratios of radioactivity to the cerebellum together with an unexpected accumulation of radioactivity in the caudate, makes [¹¹C] 1 an unpromising NET radioligand. Copyright © 2006 John Wiley & Sons, Ltd.     

Metabolism of [3H]epinephrine in rabbit fetal tissues

[³H]epinephrine was injected into rabbit fetuses through the uterine wall on day 26 of gestation and its metabolism was studied in fetal tissues. [¹⁴C]metanephrine (MN) was synthesized using a soluble rat liver preparation and it, along with [¹⁴C]epinephrine, was used as recovery markers for the quantitation of metabolites of [³H]epinephrine. The radiochemical purity of MN, the predominant metabolite isolated from the fetal liver, was achieved by obtaining a constant [³H]/[¹⁴C] after sequential thin-layer chromatography. Thirty min after the injection of labeled hormone. [³H]epinephrine comprised 25 per cent of the radioactivity in the serum and 30 per cent in the heart, while less than 5 per cent was found in the lung, liver, kidney and intestine. Unconjugated MN comprised 45 per cent of the [³H] in the serum, 63 per cent in the heart and lung and 47, 46 and 40 per cent in the liver, kidney and intestine respectively. Conjugated MN is only found in liver, kidney and intestine and accounts for 35–40 per cent of the tissue radioactivity. Enzymatic hydrolysis of conjugates with β-glucuronidase in the presence and absence of sulfatase indicates that MN is present as a glucuronide conjugate. O-methylated, deaminated metabolites accounted for 27 per cent of the radioactivity in the serum, 28 per cent in the lung and between 7 and 16 per cent in the heart, liver, kidney and intestine. Thus, in the fetal rabbit, O-methylation is the predominant route of metabolism of injected [³H]epinephrine, followed by extensive glucuronide conjugation in the fetal liver, kidney and intestine.     

Physiologic disposition of [3H]epinephrine in the rabbit fetus: Effect of promethazine

[³H]epinephrine was injected into fetal rabbits through the uterine wall on day 26 of gestation in order to study placental transfer, and the uptake, retention and metabolism of the hormone in fetal tissues. Less than 0.1 per cent of the administered [³H] was found in maternal blood or within uninjected littermates. Accumulation of [³H]/g of tissue was greatest in the fetal kidney, liver and intestine, and less in the heart and lung. Levels of [³H] in the fetal serum, heart, lung and placenta were comparable in value at 5 and 30 min, but all fell by 60 min. In contrast, the fetal kidney and intestine show a 2- to 4-fold accumulation of [³H] in 30 min as compared to 5 min, while the fetal liver exhibits a 10-fold accumulation over 30 and 60 min. [³H] metabolites present in fetal serum at 30 min, expressed as a per cent of the total radioactivity, are 45 per cent for metanephrine and 27 per cent for O-methylated deaminated products, while 25 per cent is present as unmetabolized [³H]epinephrine. O-methylated metabolites are predominant in the liver, where 85 per cent of the radioactivity comprises metanephrine in conjugated and unconjugated forms. Administration of promethazine to the doe prior to injection of [³H]epinephrine into rabbit fetuses resulted in a 40–64 per cent decrease in accumulation of [³H] in the fetal brain, liver, lung, kidney, intestine and heart. The pattern of metabolites in the serum of promethazine-treated fetuses shows an increase in the per cent of [³H] in the O-methylated, deaminated fraction from 27 to 37 per cent. While the total per cent of O-methylated metabolite is not altered in the liver of treated fetuses, metanephrine is predominantly present in the conjugated form. The liver of the rabbit fetus is an important site for accumulation of catecholamine metabolites and is pharmacologically sensitive to promethazine administered to the mother.     

Binding of [3H]SCH 39166 to Human Post Mortem Brain Tissue

Abstract: The dopamine D₁ antagonist SCH 39166 was labelled with tritium and used for in vitro binding and autoradiography using human post mortem brain tissue. Competition studies on tissue from human nucleus caudatus showed that SCH 23390 inhibited the binding of [³H]SCH 39166 biphasically. The non-specific binding of [³H]SCH 39166 in both nucleus caudatus and cerebellum was lower after the addition of SCH 23390 or SCH 39166 than after flupentixol (10 μM). Autoradiography showed specific [³H]SCH 39166 binding in the caudate nucleus and putamen in the brain sections. The binding of [³H]SCH 39166 in the medial part of the caudate nucleus was very dense and similar to that obtained with [³H]SCH 23390, which was used as a reference ligand. Dense binding of [³H]SCH 39166 was also found in cortical regions, and binding was also obtained in the cerebellum and in the hippocampus. Addition of flupentixol (10 μM) abolished some but not all the binding of [³H]SCH 39166. The binding of [³HJSCH 39166 to caudate and putamen was not totally abolished by the addition of excess SCH 23390, while excess SCH 39166 diminished the binding of [³H]SCH 23390 in all regions. The present study indicates that [³H]SCH 39166, similar to [³H]SCH 23390, binds to dopamine D₁ receptors in the human brain. It is concluded that [³H]SCH 39166 has a slightly different binding pattern than [³H]SCH 23390, which can be due to labelling of one or two additional binding site(s) pharmacologically unrelated to dopamine D₁ receptors.     

Catalepsy produced by striatal microinjections of the D1 dopamine receptor antagonist SCH 23390 in neonatal rats

Systemic injection of the D₁ dopamine receptor antagonist SCH 23390 produces catalepsy that is of lesser magnitude in neonatal than in adult rats. The present experiments were conducted in order to determine if SCH 23390 would produce catalepsy in neonatal rats following intrastriatal injection and if the ontogenetic pattern of catalepsy induced by intrastriatal SCH 23390 would be similar to the pattern observed with systemic injections. Rat pups (11 or 28 days of age) were microinjected unilaterally with SCH 23390 (0.2, 1, 5 or 10 μg) and tested for catalepsy using the forepow-on-horizontal-bar test. The results demonstrated that robust catalepsy occurred at both ages following intrastriatal injection and that catalepsy induced by 5 μg SCH 23390 was of lesser magnitude in 11-day-olds than in 28-day-olds. A separate study assessed the distribution of [³H]SCH 23390 (5 μg) following intrastriatal injection in 28-day-olds. Results of the distribution study indicated that [³H]SCH 23390 was localized primarily within the striatum. Taken together, these results suggest that the striatal mechanisms for catalepsy produced by D₁ receptor blockade are present, but not fully mature, in preweanling rat pups.     

In vivo binding of the dopamine-1 receptor PET tracers [11C]NNC112 and [11C]SCH23390: a comparison study in individuals with schizophrenia

Rationale

A deficit in dopamine-1 (D₁) receptor function in the prefrontal cortex is suggested to play a role in the cognitive dysfunction observed in patients with schizophrenia. However, the results from positron emission tomography imaging studies of D₁ receptor levels in individuals with schizophrenia are mixed.

Objectives

The aim of this investigation was to determine whether the in vivo characteristics of the different D₁ receptor tracers used in previous reports, [¹¹C]SCH23390 and [¹¹C]NNC112, may have contributed to these discrepancies reported in the literature.

Methods

Eight patients with schizophrenia and 12 healthy control subjects were scanned with both [¹¹C]SCH23390 and [¹¹C]NNC112.

Results

[¹¹C]SCH23390 and [¹¹C]NNC112 binding potentials in both patients and control subjects were compared and no tracer by diagnosis interactions were observed.

Conclusions

The results of this study suggest that differences in the binding of [¹¹C]SCH23390 and [¹¹C]NNC112 observed in previous studies are not due to differences in the in vivo behavior of these tracers.     

“In-loop” carbonylation—A simplified method for carbon-11 labelling of drugs and radioligands

Transition-metal mediated carbonylation with ¹¹C-labelled carbon monoxide ([¹¹C]CO) is a versatile method for introducing ¹¹C (t_1/2 = 20.3 min) into drugs and radioligands for subsequent use in positron emission tomography (PET). The aim of the current study was to perform the ¹¹C-carbonylation reaction on the interior surface of a stainless-steel loop used for high performance liquid chromatography (HPLC). In the experimental setup, cyclotron produced ¹¹C-labelled carbon dioxide ([¹¹C]CO₂) was converted to [¹¹C]CO by reduction over heated Molybdenum and swept into an HPLC loop pre-charged with the appropriate reaction mixture. Following a 5 min reaction, the radiochemical purity (RCP) and the trapping efficiency (TE) of the reaction mixture was determined. After optimization, [¹¹C]N-Benzylbenzamide was obtained in quantitative radiochemical yield (RCY) following a 5 min reaction at room temperature. The methodology was further applied to label [¹¹C]benzoic acid (RCP≥99%, TE>91%), [¹¹C]methyl benzoate (RCP≥99%, TE>93%) and [¹¹C]phthalide (RCP≥99%, TE>88%). A set of pharmaceuticals was finally radiolabelled using non-optimized conditions. Excellent yields were obtained for the histamine-3 receptor radioligand [¹¹C]AZ13198083, the oncology drug [¹¹C]olaparib and the dopamine D2 receptor radioligand [¹¹C]raclopride, whereas a moderate yield was observed for the high-affinity dopamine D2 receptor radioligand [¹¹C]FLB457. The presented “in-loop” process proved efficient for diverse ¹¹C-carbonylations, providing [¹¹C]amides, [¹¹C]esters and [¹¹C]carboxylic acids in moderate to excellent RCYs. Based on the advantages associated with performing the radiolabelling step as an integrated part of the purification system, this methodology may become a valuable addition to the toolbox of methodologies used for ¹¹C-carbonylation of drugs and radioligands for PET.     

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.     

Effects of the substituted (S)-3-phenylpiperidine (-)-OSU6162 on PET measurements of [11C]SCH23390 and [11C]raclopride binding in primate brains

The substituted (S)-3-phenylpiperidine (-)-OSU6162 belongs to a novel class of functional modulators of dopaminergic systems. In vivo, (-)-OSU6162 has a unique stabilising profile on dopaminergic functions. In vitro this compound exhibits low affinity for the dopamine D2 receptor, but due to its similarity to neuroleptics on brain dopaminergic neurochemistry and different postsynaptic effects it has been characterised as a preferential dopamine autoreceptor antagonist. To further clarify the effects of (-)-OSU6162 on the postjunctional nigrostriatal dopaminergic system, dopamine receptor binding was measured in rhesus monkeys (Macaca mulatta) by positron emission tomography (PET) using the D1 and D2 dopamine receptor radioligands [11C]SCH23390 and [11C]raclopride respectively, before and during continuous intravenous infusions of(-)-OSU6162. Additionally, the test-retest variability of sequential [11C]SCH23390 scans was estimated. Following the administration of (-)-OSU6162, [11C]raclopride binding in striatum was dose-dependently decreased with a 76% reduction occurring after 3.0 mg/kg per h continuous infusion. Whereas (-)-OSU6162 in the lower doses had no effect on [11C]SCH23390 binding, the highest dose, 3.0 mg/kg per h, increased [11C]SCH23390 binding, which may indicate a potentiating effect on D1 dopamine receptor mediated functions. Thus, in contrast to the conditions in vitro, (-)-OSU6162 produces a high displacement of raclopride from D2 receptors in vivo.     

Administration of L-3,4-dihydroxyphenylalanine to rats after complete hepatectomy-II. Excretion of metabolites

The excretion of metabolites of l-[3-¹⁴C]dihydroxyphenylalanine (l-[3-¹⁴C]DOPA) was studied after its injection into rats with complete hepatectomies and into control rats. Approximately 60 per cent of the dose (11 mg/kg; 20 μCi) of injected [¹⁴C]DOPA was excreted in urine in 24 hr by the control rats, and 11 per cent in bile. Although a similar percentage of the dose (69.4 per cent) was excreted by the hepatectomized rats into urine, excretion was at a slower rate. Decarboxylation of injected [¹⁴C]DOPA within 24 hr appeared to be as great in the hepatectomized rats as in the controls, but metabolism of 3,4-dihydroxyphenylethylamine (dopamine, DA) to norepinephrine was less. In the operated animals most of the DA was metabolized to 3,4-dihydroxyphenylacetic acid or to homovanillic acid. Little radioactivity was present in tissues at 24 hr after injection of l-DOPA into control rats or into hepatectomized rats; however, some radioactivity appeared to be bound to protein in some tissues in both groups of rats.     

Dopamine D1 receptors labelled with [3H]SCH23390 in rabbit cerebral cortex and neostriatum Equilibrium binding,kinetics and selectivity

Summary The binding characteristics of the novel benzazepine compound SCH23390 were studied using membrane preparations from rabbit cerebral cortex (CTX) and neostriatum (CPU; caudate putamen). The association kinetics of [³H]SCH23390 to membranes from CTX and CPU were rapid, while the dissociation kinetics were extremely slow and only around 40–60% of the binding was displaced two hours after the addition of either S(+)-butaclamol or 30 volumes of buffer. The saturation curves revealed that [³HISCH23390 bound with high affinity in both tissues, with densities of 133 fmol/mg protein for CTX (K_d 25°C = 0.31 nM) and 664 fmol/mg protein for CPU (K_d = 0.13 nM). The specificity of binding to the cortical D₁ receptor was verified in competition experiments with a variety of dopaminergic agents. The rank order of potency of these compounds was consistent with the pharmacology of the dopaminergic D₁ site. All competition curves were better fitted to a one-site model with Hill coefficients around one, indicating that [³H]SCH23390 was binding to a single cortical site. The stereoselectivity of the cortical [³H]SCH23390 binding site could be demonstrated by the use of enantiomer pairs of dopaminergic drugs. This study provides compelling evidence that [³H]SCH23390 binds to dopamine D₁ receptors in the neostriatum and cerebral cortex of the rabbit. Send offprint requests to T. A. Reader at the above address     

The fate of [C]l-3,4-dihydroxyphenylalanine in isolated perfused rat hearts

The fate of [¹⁴C]l-3,4-dihydroxyphenylalanine (¹⁴C-levodopa) in isolated perfused rat hearts has been studied. Seventy per cent of the perfused [¹⁴C]levodopa was recovered intact, while 15 per cent was recovered as dopa metabolites. Only 2 per cent of the perfused radioactivity was retained by the heart tissue. Deaminated derivatives accounted for the majority of the metabolites (7.2 per cent of perfused dose) although 3-O-methyl dopa was found in substantial quantities (1.6 per cent). The absence of significant tissue accumulation of norepinephrine-¹⁴C leads to the speculation that amine synthesis from perfused dopa occurs at an extraneuronal site.     

Dopamine depletion and in vivo binding of PET D1 receptor radioligands: implications for imaging studies in schizophrenia.

Recent positron emission tomography (PET) studies have assessed the level of dopamine (DA) D1 receptors in the prefrontal cortex (PFC) in patients with schizophrenia and have generated contradictory findings. In the PFC of patients with schizophrenia, the binding potential (BP) of [11C]NNC 112 has been reported as increased, while the BP of [11C]SCH 23390 was reported as decreased or unchanged. In this study, the effect of acute and subchronic DA depletion on the in vivo binding of [11C]NNC 112 and [3H]SCH 23390 was evaluated in rats. Acute DA depletion did not affect [11C]NNC 112 in vivo binding, but paradoxically decreased [3H]SCH 23390 in vivo binding. Subchronic DA depletion was associated with increased [11C]NNC 112 in vivo binding and decreased [3H]SCH 23390 in vivo binding. Together, these data demonstrate that the in vivo binding of these radiotracers is differentially affected by changes in endogenous DA tone, and suggest that alterations in the binding of these tracers in the PFC of patients with schizophrenia might reflect changes in D1 receptors secondary to sustained deficit in prefrontal DA function.     

11C-SCH 39166, a selective ligand for visualization of dopamine-D1 receptor binding in the monkey brain using PET

The new selective D₁-dopamine receptor antagonist SCH 39166 was labelled with the positron emitting isotope¹¹C and used as ligand for visualization of dopamine-D₁ receptor binding in Cynomolgus monkeys by PET. After intravenous administration of the ligand a marked uptake of radioactivity was recorded in the D₁-dopamine receptor-rich striatum and neocortex but not in the dopamine receptor-poor cerebellum. The uptake of radioactivity in striatum and neocortex was markedly displaced after the intravenous injection of a high dose of the D₁-dopamine receptor antagonist SCH 23390 but not after the 5-HT₂ receptor antagonist ketanserine.¹¹C-SCH 39166 should be a useful tool to explore D₁-dopamine receptor characteristics in the living human brain by PET.     

The metabolism of iodine-123 labelled 3-(5-cyclopropyl-1,2,4-oxadiazo-3-yl)-7-iodo-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5-a][1,4]benzodiazepine (NNC 13–8241) measured in human plasma is only minor

The imidazobenzodiazepine derivate ¹²³I-labelled 3-(5-cyclopropyl-1,2,4-oxadiazo-3-yl)-7-iodo-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5-a][1,4]-benzodiazepine (NNC 13–8241) is a partial benzodiazepine agonist and binds with a high affinity to the benzodiazepine receptor. The favourable kinetic properties indicate that [¹²³I]NNC 13–8241 is a promising SPET ligand. In the present study, an extensive examination of the metabolite pattern of [¹²³I]NNC 13–8241 in plasma and urine from seven healthy subjects was performed using gradient HPLC. After injection of [¹²³I]NNC 13–8241 into human beings, only one radioactive metabolite was found in plasma 3–300 min post injection. This polar metabolite eluted together with the solvent front fraction. The proportion of unchanged [¹²³I]NNC 13–8241 was 82–86 per cent during the entire study. In addition, two other radioactive metabolites were found in urine. The first metabolite was lipophilic and eluted slightly before the parent compound [¹²³I]NNC 13–8241. The second metabolite eluted slightly after solvent front peak. The amount of unchanged [¹²³I]NNC 13–8241 in human urine was 9 per cent and the solvent front fraction contained 72 per cent of total radioactivity. No detectable radioactivity appeared with the same retention time as synthetisized nor-NNC 13–8241 in plasma or urine, which excludes the possibility of in vivo demethylation of [¹²³I]NNC 13–8241. In conclusion, the [¹²³I]NNC 13–8241 was found to have only a minor metabolism, which favours its use as a SPET tracer for quantitation of the benzodiazepine receptor. © 1998 John Wiley & Sons, Ltd.     

Development of novel PET probes, [18F]BCPP‐EF, [18F]BCPP‐BF,and [11C]BCPP‐EM for mitochondrial complex 1 imaging in the living brain

We developed three novel positron‐emission tomography (PET) probes, 2‐tert‐butyl‐4‐chloro‐5‐{6‐[2‐(2[¹⁸F]fluoroethoxy)‐ethoxy]‐pyridin‐3‐ylmethoxy}‐2H‐pyridazin‐3‐one ([¹⁸F]BCPP‐EF), 2‐tert‐butyl‐4‐chloro‐5‐[6‐(4‐[¹⁸F]fluorobutoxy)‐pyridin‐3‐ylmethoxy]‐2H‐pyridazin‐3‐one ([¹⁸F]BCPP‐BF), and 2‐tert‐butyl‐4‐chloro‐5‐{6‐[2‐(2‐[¹¹C]methoxy‐ethoxy)‐ethoxy]‐pyridin‐3‐ylmethoxy}‐2H‐pyridazin‐3‐one ([¹¹C]BCPP‐EM), for quantitative imaging of mitochondrial complex 1 (MC‐1) activity in vivo. These three PET probes were successfully labeled by nucleophilic [¹⁸F]fluorination or by [¹¹C]methylation of their corresponding precursor with sufficient radioactivity yield, good radiochemical purity, and sufficiently high specific radioactivity for PET measurement. The specificity of these probes for binding to MC‐1 was assessed with rotenone, a specific MC‐1 inhibitor, by a rat brain slice imaging method in vitro. Rat whole‐body imaging by small‐animal PET demonstrated that all probes showed high uptake levels in the brain as well as in the heart sufficient to image them clearly. The rank order of uptake levels in the brain and the heart just after injection was as follows: high in [¹⁸F]BCPP‐BF, intermediate in [¹¹C]BCPP‐EM, and low in [¹⁸F]BCPP‐EF. The kinetics of [¹⁸F]BCPP‐EF and [¹¹C]BCPP‐EM provided a reversible binding pattern, whereas [¹⁸F]BCPP‐BF showed nonreversible accumulation‐type kinetics in the brain and heart. Metabolite analyses indicated that these three compounds were rapidly metabolized in the plasma but relatively stable in the rat brain up to 60 min post‐injection. The present study demonstrated that [¹⁸F]BCPP‐EF could be a useful PET probe for quantitative imaging of MC‐1 activity in the living brain by PET.