A definite measure of occupancy exposures, seeking with non-radiolabeled in vivo 5-HT2A receptor occupancy and in vitro free fractions.

Unbound drug concentration in the brain would be the true exposure responsible for specific target occupancy. Drug exposures from preclinical are total concentrations of those over/underestimate the clinical dose projection. With the application of mass spectrometry, the current work proposes a definite measure of test drug exposures at serotonin-2A occupancy. The 5-HT2A occupancy of antagonist in the rat brain has determined with non-radiolabeled tracer MDL-100,907 at an optimized dose (3 µg/kg) and treatment time (30 min). Equilibrium dialysis method determines the in vitro free fraction of the test antagonist in untreated rat brain homogenates and plasma. Drug-free fractions derived the unbound concentration (EC50) in plasma and brain at test doses. The corresponding binding affinities (Ki) correlated with the unbound concentrations. Except for quetiapine, the ED50 values in the dose-occupancy curves of antagonists are close and ranged from 1 to 3 mg/kg. The test drug quetiapine, eplivanserin, and clozapine showed high free fractions in plasma, but for ketanserin and olanzapine, the brain free fraction was higher. The correlation between the unbound EC50 of the antagonists and corresponding Ki values was good (r2=0.828). The improved EC50 accuracy with unbound concentrations was 10-250 folds in plasma and 10-170 folds in the brain. Further, the free fractions (fu, plasma/fu, brain) of test drugs had shown a correlation of ∼83% with brain permeability (Ctotal brain/Ctotal plasma), a limiting factor. Thus, correlating the occupancy with unbound exposure and pharmacology would result in an accurate measurement of drug potency and optimizes in selecting the clinical dose.

Simultaneous determination of sarpogrelate and its active metabolite in human plasma by liquid chromatography with tandem mass spectrometry and its application to a pharmacokinetic study.

We established a rapid and simple liquid chromatography with tandem mass spectrometry method for the simultaneous determination of sarpogrelate and its active metabolite, M-1, in human plasma. Sarpogrelate, M-1, and the internal standard, ketanserin, were extracted from a 50 µL aliquot of human plasma by protein precipitation using acetonitrile. Chromatographic separation was performed on a Shim-pack GIS ODS C18 column (100 × 3.0 mm; 3 µm) with an isocratic mobile phase consisting of 10 mM ammonium acetate and acetonitrile (70:30, v/v) at a flow rate of 0.6 mL/min; the total run time was <2.5 min. Mass spectrometric detection was conducted in selected reaction-monitoring mode with positive electrospray ionization at m/z 430.35 → 135.10 for sarpogrelate, m/z 330.30 → 58.10 for M-1, and m/z 395.70 → 188.85 for ketanserin. The linear ranges of concentration for sarpogrelate and M-1 were 1-1000 and 0.5-500 ng/mL, respectively. The coefficient of variation for the assay's precision was ≤9.95%, and the accuracy was 90.6-107%. All analytes were stable under various storage and handling conditions, and no relevant crosstalk and matrix effect was observed. This method was successfully applied to a pharmacokinetic study after oral administration of a 100 mg sarpogrelate tablet to healthy male Korean volunteers.

Simple determination of azasetron in rat plasma by column-switching high-performance liquid chromatography.

For the quantification of azasetron in rat plasma samples, a column-switching HPLC method was developed and validated. Following dilution of plasma samples with mobile phase A (17 mM potassium phosphate buffer (pH 3.0)) and simple protein precipitation by addition of perchloric acid (60%), the mixture was directly injected onto the pre-column. After endogenous plasma substances were eluted to waste, the analyte was transferred to the trap column by switching the system. Then, the analyte was back-flushed to the analytical column for separation with mobile phase B (a 22:78 v/v mixture of acetonitrile and 17 mM potassium phosphate buffer (pH 3.0)) and detected at 250 nm using a photodiode array detector. A linear standard curve was obtained in the concentration range of 10-800 ng/mL with the correlation coefficient (r) of 0.9998. The intra- and inter-day precision and accuracy values for azasetron were in the ranges of 0.3-12.9% and 89.7-101.4%, respectively. The method was valid in terms of specificity, precision, and accuracy. In addition, this efficient analytical method was successfully applied to determine plasma concentrations of azasetron following oral administration of azasetron at a dose of 4.0 mg/kg to rats.

APD791, 3-methoxy-n-(3-(1-methyl-1h-pyrazol-5-yl)-4-(2-morpholinoethoxy)phenyl)benzamide, a novel 5-hydroxytryptamine 2A receptor antagonist: pharmacological profile, pharmacokinetics, platelet activity and vascular biology.

We have evaluated the receptor pharmacology, antiplatelet activity, and vascular pharmacology of APD791 [3-methoxy-N-(3-(1-methyl-1H-pyrazol-5-yl)-4-(2-morpholinoethoxy)phenyl)benzamide] a novel 5-hydroxytryptamine 2A (5-HT(2A)) receptor antagonist. APD791 displayed high-affinity binding to membranes (K(i) = 4.9 nM) and functional inverse agonism of inositol phosphate accumulation (IC(50) = 5.2 nM) in human embryonic kidney cells stably expressing the human 5-HT(2A) receptor. In competition binding assays, APD791 was greater than 2000-fold selective for the 5-HT(2A) receptor versus 5-HT(2C) and 5-HT(2B) receptors, and was inactive when tested against a wide panel of other G-protein-coupled receptors. APD791 inhibited 5-HT-mediated amplification of ADP-stimulated human and dog platelet aggregation (IC(50) = 8.7 and 23.1 nM, respectively). Similar potency was observed for inhibition of 5-HT-stimulated DNA synthesis in rabbit aortic smooth muscle cells (IC(50) = 13 nM) and 5-HT-mediated vasoconstriction in rabbit aortic rings. Oral administration of APD791 to dogs resulted in acute (1-h) and subchronic (10-day) inhibition of 5-HT-mediated amplification of collagen-stimulated platelet aggregation in whole blood. Two active metabolites, APD791-M1 and APD791-M2, were generated upon incubation of APD791 with human liver microsomes and were also indentified in dogs after oral administration of APD791. The affinity and selectivity profiles of both metabolites were similar to APD791. These results demonstrate that APD791 is an orally available, high-affinity 5-HT(2A) receptor antagonist with potent activity on platelets and vascular smooth muscle.

Unbound Fraction of Clozapine Significantly Decreases with Elevated Plasma Concentrations of the Inflammatory Acute-Phase Protein Alpha-1-Acid Glycoprotein.

BACKGROUND: During inflammation, elevated total (unbound plus protein-bound) clozapine plasma concentrations have been observed. Elevated alpha-1-acid glycoprotein concentrations during inflammation are suggested to cause increased plasma clozapine-alpha-1-acid glycoprotein binding, resulting in elevated total clozapine plasma concentrations without significant changes in unbound concentrations. Here, we investigated the association between alpha-1-acid glycoprotein plasma concentrations and clozapine unbound fraction. METHODS: First, 25 and 60 µL of alpha-1-acid glycoprotein solution (20 mg/mL) were added to plasma samples (n = 3) of clozapine users (spiking experiment). Second, the association between alpha-1-acid glycoprotein plasma concentration and clozapine unbound fraction was assessed in patient samples (patient study). Samples were determined by liquid chromatography-tandem mass spectrometry. Data were analyzed with a paired t test (spiking experiment) and an unpaired t test (patient study). RESULTS: The spiking experiment showed significantly lower mean unbound fractions following 25- and 60-µL alpha-1-acid glycoprotein spikes (relative reductions of 28.3%, p = 0.032 and 43.4%, p = 0.048). In the patient study, total clozapine plasma concentrations were 10% higher in elevated (n = 6) compared with normal alpha-1-acid glycoprotein (n = 20) samples [525 µg/L vs. 479 µg/L, mean difference = 47 µg/L (95% confidence interval -217 to 310), p = 0.72]. Elevated alpha-1-acid glycoprotein samples had a 26% lower mean unbound fraction compared with normal samples [1.22% vs. 1.65%, mean difference = -0.43% (95% confidence interval -0.816 to -0.0443), p = 0.03]. CONCLUSIONS: Both the spiking experiment and patient study showed a significant association between elevated alpha-1-acid glycoprotein plasma concentrations and a lower clozapine unbound fraction. Future studies should include clinical data to examine whether this association is clinically relevant, suggesting any clozapine dose adjustments.

Single Administration of HBK-15-a Triple 5-HT1A, 5-HT7, and 5-HT3 Receptor Antagonist-Reverses Depressive-Like Behaviors in Mouse Model of Depression Induced by Corticosterone.

Studies suggest that the blockade of 5-HT1A, 5-HT7, and 5-HT3 receptor may increase the speed of antidepressant response. 1-[(2,6-Dimethylphenoxy)ethoxyethyl]-4-(2-methoxyphenyl)piperazine hydrochloride (HBK-14) and 1-[(2-chloro-6-methylphenoxy)ethoxyethyl]-4-(2-methoxyphenyl)piperazine hydrochloride (HBK-15), dual 5-HT1A and 5-HT7 antagonists, showed significant antidepressant- and anxiolytic-like properties in our previous tests in rodents. In this study, we aimed to investigate their antidepressant potential using mouse model of corticosterone-induced depression. We chose sucrose preference test, forced swim test, and elevated plus maze to determine anhedonic-, antidepressant-, and anxiolytic-like activities. We also evaluated the influence of the active compound on brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) levels in the hippocampus. Moreover, for both compounds, we performed biofunctional (5-HT3 receptor) and pharmacokinetic studies. We found that HBK-14 and HBK-15 were potent 5-HT3 receptor antagonists. HBK-14 (2.5 mg/kg) and HBK-15 (1.25 mg/kg) after intravenous (i.v.) and intraperitoneal (i.p.) administration permeated the blood-brain barrier with brain/plasma ratio lower than 1. The bioavailability of studied compounds after i.p. administration was 15% for HBK-14 and 54% for HBK-15. Chronic administration of HBK-15 (1.25 mg/kg) and fluoxetine (10 mg/kg) protected corticosterone-treated mice from anhedonic-, depressive-, and anxiety-like behaviors, as well as decreases in BDNF and NGF levels in the hippocampus. HBK-14 (2.5 mg/kg) counteracted anxiety-like behaviors in corticosterone-treated mice. Single administration of HBK-15 (1.25 mg/kg) and ketamine (1 mg/kg) reversed depression-like behavior and regulated decreased BDNF level in the hippocampus in corticosterone-treated mice. Our results suggest that simultaneous blockade of serotonergic 5-HT1A, 5-HT7, and 5-HT3 receptors might accelerate antidepressant response.

Optimal metabolite curve fitting for kinetic modeling of 11C-WAY-100635.

UNLABELLED: Many quantitative imaging protocols that make use of a metabolite-corrected arterial input function require the use of a mathematic model to describe the rate of metabolism of the radioligand. Commonly, parametric models are fit to metabolism data and then the fitted model is used to correct the plasma input function. (11)C-WAY 100635 is a rapidly metabolized radioligand used extensively in mapping the 5-hydroxytryptamine receptor 1A system. METHODS: To evaluate the adequacy of fit of 4 metabolite models, we examined data from 92 subjects who received an injection of (11)C-WAY 100635, were imaged with PET, and underwent measurement of total plasma concentration and metabolites. The performance of these models was assessed according to residual plots, as well as fit and information criteria. RESULTS: The study showed that the choice of model has a substantial effect on the resulting estimates of outcome measures. CONCLUSION: Among the models considered, the Hill model provides the best fit across all criteria.

New evidence for oxetorone toxicity.

CONTEXT: Oxetorone is a serotonin antagonist antimigraine drug but literature relating to its toxic properties is poor. The aim of this study is to describe the toxicological profile of oxetorone and to highlight any relationship between clinical and analytical findings. MATERIALS AND METHODS: This is a retrospective and observational study of cases exposure to oxetorone, reported to the Angers Poison and Toxicovigilance Centre between January 2002 and May 2016. Severity was assessed using the Poisoning Severity Score (PSS). Cases where data were incomplete, where oxetorone was deemed not accountable, where clinical signs were linked mainly to a co-ingested drug or where the plasma concentration of oxetorone was negative were all excluded. RESULTS: We included 43 cases of exposure, 31 of whom were suicide attempts. The assumed ingested dose (60-3600 mg) was correlated to severity (rs = 0.45, p = 0.01). Symptoms of moderate severity (PSS2 = drowsiness, hypertonia, myosis, convulsions, arterial hypotension, QRS widening, QTc prolongation) were observed following ingestion of more than 600 mg of oxetorone (median dose =1200 mg) and severe symptoms (PSS 3 = coma, convulsions, QTc prolongation, QRS widening, ventricular tachycardia, arterial hypotension, cardiogenic shock) were observed starting from 1800 mg (median dose =2700 mg). In four cases, a secondary worsening of symptoms 10-48 h following ingestion was observed. Plasma oxetorone was measured in four patients. Severe symptoms were observed in the event of a concentration over 0.3 mg/L and the highest measured serum oxetorone level was delayed by 20-48 h following the ingestion for two cases. CONCLUSIONS: Several clinical and paraclinical parameters strongly point towards membrane-stabilising properties of the molecule and the risk of a delayed occurrence of symptoms or a secondary worsening.

Analgesic effect of acetaminophen in humans: first evidence of a central serotonergic mechanism.

OBJECTIVES: Preclinical studies have suggested that the mechanism of the analgesic action of acetaminophen (INN, paracetamol) is linked to the serotonergic system and that it is inhibited by tropisetron, a 5-hydroxytryptamine type 3 antagonist. The aim of this study was to confirm these findings in humans. METHODS: Twenty-six rapid metabolizers of tropisetron were included in this double-blind crossover study. After ethical approval, at weekly intervals, the subjects took a single oral dose of 1 g acetaminophen combined with either intravenous tropisetron (5 mg), granisetron (3 mg), or placebo (saline solution). For each session, the analgesic effect of acetaminophen was assessed by use of a pain self-evaluation instrument, the Pain Matcher. The pain detection threshold was determined 5 times over the period of the 4 postdosing hours. The area under the curve (0-4 hours) (mean +/- SD) of acetaminophen/tropisetron and the area under the curve of acetaminophen/granisetron were compared with the effect of acetaminophen/placebo. Blood samples for acetaminophen concentration measurements were taken to evaluate a pharmacokinetic interaction. RESULTS: The analgesic effect of acetaminophen/placebo (expressed as the area under the curve of the percentage of the individual pain score reported at baseline along time [% x min]) (2145 +/- 2901 % x min) was totally inhibited by both tropisetron (89 +/- 1747 % x min, P = .007) and granisetron (45 +/- 2020 % x min, P = .002). Acetaminophen concentration was not significantly different when associated with tropisetron (P = .919) or granisetron (P = .309). CONCLUSION: These results clearly show for the first time in humans that the coadministration of tropisetron or granisetron with acetaminophen completely blocks the analgesic effect of acetaminophen. They support the hypothesis that the mechanism of the analgesic action of acetaminophen might involve the serotonergic system. Furthermore, they demonstrate a pharmacodynamic interaction between these 2 types of drugs, which are frequently coadministered, especially in cancer patients.

Development and validation of a rapid 96-well format based liquid-liquid extraction and liquid chromatography-tandem mass spectrometry analysis method for ondansetron in human plasma.

A rapid and sensitive LC-MS/MS method for the quantification of ondansetron was developed and validated. The plasma samples were treated by a semi-automated liquid-liquid extraction (LLE) in 1.2 mL 96-well format micro-tubes. Ondansetron and the internal standard (IS) granisetron were analyzed by combined reversed phase LC-MS/MS, with positive ion electrospray ionization, using multiple reactions monitoring (MRM). The statistical evaluation for this method reveals excellent linearity, accuracy and precision values for the range of concentrations 0.25-40.0 ng/mL. The proposed method enabled the reliable determination of ondansetron in bioequivalence studies after per os administration of a 4 or 8 mg tablet.

Rapid determination of granisetron in human plasma by liquid chromatography coupled to tandem mass spectrometry and its application to bioequivalence study.

A simple, sensitive and rapid method for analysis of granisetron in human plasma, utilizing liquid chromatography tandem mass spectrometry (LC-MS/MS), has been developed and validated to satisfy FDA guidelines for bioanalytical methods. The analyte and internal standard (IS) were isolated from 100microl plasma samples by liquid-liquid extraction (LLE). A Varian 1200l tandem mass spectrometer equipped with an electrospray ionization source was operated in selected reaction monitoring (SRM) mode with the precursor-to-product ion transitions m/z 313.4/138 for granisetron and m/z 270/201 for the IS used for quantitation. The assay exhibited a linear dynamic range of 0.02-20ng/ml for granisetron in human plasma. The lower limit of quantification (LLOQ) was 0.02ng/ml with a relative standard deviation of less than 15%. The mean extraction recovery from spiked plasma samples was 97.9%. The intra-day accuracy of the assay was within 10% of nominal and intra-day precision was better than 15% C.V. A run time of 2.0min for each sample made it possible for high-throughput bioanalysis. The method was employed in a bioequivalence study of two formulations of granisetron hydrochloride 1mg rapidly disintegrating tablets/1mg capsules.

Development and validation of a rapid LC-MS/MS method for simultaneous determination of netupitant and palonosetron in human plasma and its application to a pharmacokinetic study.

A simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was firstly developed and validated for simultaneous determination of netupitant and palonosetron in human plasma using ibrutinib as the internal standard (IS). Following liquid-liquid extraction, the compounds were eluted isocratically on a Phenomenex C18 column (50mm×2.0mm, 3µm) with the mobile phase consisting of acetonitrile and 10mM ammonium acetate buffer (pH 9.0) (89:11, v/v) at the flow rate of 0.3mL/min. The monitored ion transitions were m/z 579.5→522.4 for netupitant, m/z 297.3→110.2 for palonosetron and m/z 441.2→138.1 for IS. Chromatographic run time was 2.5min per injection, which made it possible to analyze more than 300 of samples per day. The assay exhibited a linear dynamic range of 5-1000ng/mL for netupitant and 0.02-10ng/mL for palonosetron in plasma. The values for both within- and between-day precision and accuracy were well within the generally accepted criteria for analytical methods (<15%). Selectivity, linearity, lower limit of quantification (LLOQ), accuracy, precision, stability, matrix effect, recovery and carry-over effect were evaluated for all analytes. The method is simple, rapid, and has been applied successfully to a pharmacokinetic study of netupitant and palonosetron in healthy volunteers.

Sustained release of risperidone from biodegradable microspheres prepared by in-situ suspension-evaporation process.

Risperidone-loaded poly (D,L-lactide-co-glycolide) (PLGA) microspheres were prepared with a suspension-evaporation process with an aqueous suspension containing an in situ-formed aluminum hydroxide inorganic gel (SEP-AL process) and evaluated for encapsulation efficiency, particle size, surface morphology, glass transition temperature, in vitro drug release profile, and in vivo behavior. The SEP-AL microspheres were compared with conventional oil-in-water (O/W) emulsion solvent evaporation method using polyvinylalcohol (PVA) as an emulsifier (CP-PVA process). The microspheres were spherical in shape. DSC measurements showed that risperidone crystallinity was greatly reduced due to the homogeneous distribution of risperidone in PLGA microspheres. In vitro drug release profile from the microspheres showed a sigmoidal pattern of negligible initial burst up to 24h and minimal release (time-lag) for 7 days. After the lag phase, slow release took a place up to 25 days and then rapid release occurred sharply for 1 week. In vivo rat pharmacokinetic profile from the microspheres showed very low blood concentration level at the initial phase (up to 24h) followed by the latent phase up to 21 days. At the 3rd week, main phase started and the blood concentration of the drug increased up to the 5th week, and then gradually decreased. The risperidone-loaded PLGA microspheres produced by SEP-AL process showed excellent controlled release characteristics for the effective treatment of schizophrenia patients.

Phase I/II trial of granisetron: a novel 5-hydroxytryptamine antagonist for the prevention of chemotherapy-induced nausea and vomiting.

A new class of antiemetic agents, the 5-hydroxytryptamine (5-HT3) antagonists, have been shown to possess potent antiemetic properties in the ferret model. We conducted a phase I/II trial of the 5-HT3 antagonist BRL43694 (granisetron) in 24 chemotherapy-naïve patients who were receiving any combination of doxorubicin and/or cisplatin. The first 12 patients received 40 micrograms/kg and the second 12 received 80 micrograms/kg of granisetron intravenously before beginning chemotherapy. Nausea was assessed by a patient-completed visual analogue scale and episodes of retching recorded by the patient and an independent observer. Fifty-two percent of the 22 evaluable patients had no retching or vomiting and 32% had no nausea during the first 24 hours after chemotherapy. Pharmacokinetic measurements were performed. The disposition of granisetron was best described using a two-compartment model. The area under the plasma concentration curve (AUC) was 277 +/- 226 ng.h/mL and 359 +/- 282 ng.h/mL at 40 and 80 micrograms/kg, respectively. The total body clearance was 0.319 +/- 0.315 L/kg/hr and 0.483 +/- 0.504 L/kg/hr at the 40 and 80 micrograms/kg doses. Wide interpatient variation in model independent parameters was observed. There was no suggestion of dose-dependent efficacy at the two dose levels studied. We conclude that granisetron shows promise as a well-tolerated and effective antiemetic. Randomized trials comparing this drug with standard regimens are currently underway.

Effective pharmacotherapy of alcoholic amnestic disorder with fluvoxamine. Preliminary findings.

Ten patients with alcoholic chronic organic brain disease were categorized as having alcohol amnestic disorder, or Korsakoff's psychosis (n = 6), dementia associated with alcoholism (n = 3), or compensated alcoholic liver disease (n = 1). All patients had severe deficits in memory for recently acquired information (episodic memory). Patients with alcohol dementia also showed global intellectual decline, including decreased performance on measures of semantic (knowledge) memory and reduction in levels of cerebrospinal fluid somatostatin. In a 4-week double-blind crossover design, the serotonin-uptake blocker fluvoxamine maleate (100 to 200 mg/d) was found to improve episodic memory in only the patients with alcohol amnestic disorder. These improvements in memory were significantly correlated with reductions in levels of cerebrospinal fluid 5-hydroxyindoleacetic acid, suggesting that facilitation of serotonergic neurotransmission may ameliorate the episodic memory failure in patients with alcohol amnestic disorder.

Pharmacokinetics and bioavailability study of two ondansetron oral soluble film formulations in fasting healthy male Chinese volunteers.

BACKGROUND: Ondansetron oral soluble film is designed to be applied on top of the tongue without requiring water to aid dissolution or swallowing, which is especially fitting for nausea and vomiting patients. PURPOSE: This study was conducted to compare the bioavailability of two 8 mg ondansetron oral soluble film formulations. PATIENTS AND METHODS: This randomized, open-label, two-period crossover study was performed under fasting conditions. A total of ten eligible subjects were randomly assigned at a 1:1 ratio to receive a single 8 mg dose of the test and reference ondansetron oral soluble film formulations, followed by a 1-week washout period and administration of the alternate formulation. The concentrations of ondansetron were assayed using an liquid chromatograph-mass spectrometer/mass spectrometer (LC-MS/MS) method. For analysis of pharmacokinetic properties, including the peak concentration of T max (C max), AUC from time 0 (baseline) to t hours (AUC0- t ), and AUC from baseline to infinity (AUC0-∞), blood samples were obtained at intervals over the 24-hour period after studying drug administration. Tolerability was assessed by monitoring vital signs and laboratory tests (hematology, blood biochemistry, hepatic function, and urinalysis) and by questioning subjects about adverse events. RESULTS: The mean (standard derivation [SD]) relative bioavailability was 96.5 (23.7%). The 90% confidence intervals (CIs) for the log-transformed ratios of C max and AUC0- t were 84.71%-103.28% and 91.38%-108.60%, respectively (P>0.05). Similar results were found for the data without log-transformation. No statistically significant differences were found based on analysis of variance. No significant adverse events occurred or were reported during the study. CONCLUSION: As the 90% CIs based on the differences between the test and reference formulation were within the 80%-125% range for both the C max and AUC0- t , we concluded that the two formulations were bioequivalent with respect to the rate or the extent of absorption. Both formulations are well tolerated.

In vivo occupancy of the 5-HT1A receptor by a novel pan 5-HT1(A/B/D) receptor antagonist, GSK588045, using positron emission tomography.

5-hydroxytryptamine 1 (5-HT1) receptor blockade in combination with serotonin reuptake inhibition may provide a more rapid elevation of synaptic 5-HT compared to serotonin reuptake alone, by blocking the inhibitory effect of 5-HT1 receptor activation on serotonin release. GSK588045 is a novel compound with antagonist activity at 5-HT1A/1B/1D receptors and nanomolar affinity for the serotonin transporter, which was in development for the treatment of depression and anxiety. Here we present the results of an in vivo assessment of the relationship between plasma exposure and 5-HT1A receptor occupancy. Six Cynomolgus monkeys (Macaca fascicularis) were scanned using the PET ligand [(11)C]WAY100635 before and after dosing with GSK588045 (0.03, 0.1 and 0.3 mg/kg 60 min i.v. infusion). Data was quantified using a simplified reference tissue model, with the cerebellar time-activity curve used as an input function. Plasma levels of GSK588045 were measured, and the EC50 of GSK588045 for 5-HT1A receptor occupancy was estimated. An Emax model described the relationship between the GSK588045 plasma concentration and 5-HT1A receptor occupancy data well. EC50 estimates (and 95% confidence intervals) for raphe nuclei and the frontal cortex were 6.99 (2.48 to 11.49) and 7.80 (2.84 to 12.76) ng/ml respectively. GSK588045 dose dependently blocked the signal of the PET ligand [(11)C]WAY100635, confirming its brain entry and occupancy of 5-HT1A receptors in the primate brain. The estimated EC50 at the post-synaptic heteroreceptors and somatodendritic autoreceptors is similar. 5-HT1 receptor blockade by compounds such as GSK588045 may provide a faster alternate mechanism of antidepressant and anxiolytic action than standard SSRI treatment.

Human plasma inhibitors of platelet serotonin uptake and imipramine receptor binding: extraction and heterogeneity.

Human plasma contains a considerable concentration of low molecular weight substances that inhibit, in a dose-dependent manner, both high-affinity imipramine receptor binding and serotonin uptake in platelets. Incubation of plasma with alumina was used to extract and to partly characterize these imipramine-like inhibitors. The human plasma extract inhibited imipramine binding and serotonin uptake with median inhibitory concentration (IC50) of 0.18 +/- 0.1 and 0.36 +/- 0.15 mg/ml, respectively. Imipramine-like activity of the extract was markedly degraded by carboxypeptidase B and leucine aminopeptidase, but was resistant to neurominidase and phospholipases A2, C, and D. The elution profile of the extract after gel chromatography on Bio-Gel P-2 showed two major peaks of serotonin uptake and imipramine binding inhibition and three additional peaks of serotonin uptake inhibitory activity that did not have a significant effect on imipramine binding. The possible mechanism of pharmacological action of the imipramine-like inhibitors and their relation to development of affective illnesses remain to be clarified.

Quantitative analysis for estimating binding potential of the brain serotonin transporter with [11 C]McN5652.

SUMMARY: [11C](+)McN5652 is a selective serotonin reuptake inhibitor with subnanomolar potency for the serotonin transporter, and is currently being used for positron emission tomography studies. However, quantification of the regional [11C](+)McN5652 binding potential in vivo is a controversial issue because of its complex characteristics. The authors examined the regional differences in nonspecific binding and proposed simple methods for estimating the binding potential of [11C](+)McN5652. The regional difference in nonspecific binding was evaluated by the activity ratio of the thalamus compared with the cerebellum for inactive-isomer [11C](-)McN5652 and [11C](+)McN5652 saturation studies. The distribution volume of the thalamus was approximately 1.16 times larger than that of the cerebellum. The thalamus-to-cerebellum distribution volume ratio was estimated by nonlinear least square and graphical methods, with and without arterial input function. The graphical method with k2; without blood sampling was practical and most applicable for estimation of the distribution volume ratio because this method is more stable than the nonlinear least square method in the simulation study. Binding potential estimated with the distribution volume ratio of [11C](+)McN5652 and the correction with distribution volume ratio of [11C](-)McN5652 represent the most reliable parameters for the assessment of serotonin transporter binding.

[3H]paroxetine binding in human platelets in relation to age and sex.

The binding of [3H]paroxetine to serotonin uptake sites in human platelets from 47 individuals between 15 and 95 years of age was investigated. There were no significant age-related changes in maximal binding capacity (Bmax) or apparent binding affinity (Kd). There were no significant differences in binding between males (n = 19) and females (n = 28).