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1.
Nature ; 626(7998): 427-434, 2024 Feb.
Article in English | MEDLINE | ID: mdl-38081299

ABSTRACT

Vesicular monoamine transporter 2 (VMAT2) accumulates monoamines in presynaptic vesicles for storage and exocytotic release, and has a vital role in monoaminergic neurotransmission1-3. Dysfunction of monoaminergic systems causes many neurological and psychiatric disorders, including Parkinson's disease, hyperkinetic movement disorders and depression4-6. Suppressing VMAT2 with reserpine and tetrabenazine alleviates symptoms of hypertension and Huntington's disease7,8, respectively. Here we describe cryo-electron microscopy structures of human VMAT2 complexed with serotonin and three clinical drugs at 3.5-2.8 Å, demonstrating the structural basis for transport and inhibition. Reserpine and ketanserin occupy the substrate-binding pocket and lock VMAT2 in cytoplasm-facing and lumen-facing states, respectively, whereas tetrabenazine binds in a VMAT2-specific pocket and traps VMAT2 in an occluded state. The structures in three distinct states also reveal the structural basis of the VMAT2 transport cycle. Our study establishes a structural foundation for the mechanistic understanding of substrate recognition, transport, drug inhibition and pharmacology of VMAT2 while shedding light on the rational design of potential therapeutic agents.


Subject(s)
Cryoelectron Microscopy , Vesicular Monoamine Transport Proteins , Humans , Binding Sites , Cytoplasm/drug effects , Cytoplasm/metabolism , Ketanserin/chemistry , Ketanserin/metabolism , Ketanserin/pharmacology , Reserpine/chemistry , Reserpine/metabolism , Reserpine/pharmacology , Serotonin/chemistry , Serotonin/metabolism , Substrate Specificity , Tetrabenazine/chemistry , Tetrabenazine/metabolism , Tetrabenazine/pharmacology , Vesicular Monoamine Transport Proteins/antagonists & inhibitors , Vesicular Monoamine Transport Proteins/chemistry , Vesicular Monoamine Transport Proteins/metabolism , Vesicular Monoamine Transport Proteins/ultrastructure
2.
Cell Chem Biol ; 24(7): 870-880.e5, 2017 Jul 20.
Article in English | MEDLINE | ID: mdl-28712745

ABSTRACT

Diacylglycerol kinases (DGKs) are integral components of signal transduction cascades that regulate cell biology through ATP-dependent phosphorylation of the lipid messenger diacylglycerol. Methods for direct evaluation of DGK activity in native biological systems are lacking and needed to study isoform-specific functions of these multidomain lipid kinases. Here, we utilize ATP acyl phosphate activity-based probes and quantitative mass spectrometry to define, for the first time, ATP and small-molecule binding motifs of representative members from all five DGK subtypes. We use chemical proteomics to discover an unusual binding mode for the DGKα inhibitor, ritanserin, including interactions at the atypical C1 domain distinct from the ATP binding region. Unexpectedly, deconstruction of ritanserin yielded a fragment compound that blocks DGKα activity through a conserved binding mode and enhanced selectivity against the kinome. Collectively, our studies illustrate the power of chemical proteomics to profile protein-small molecule interactions of lipid kinases for fragment-based lead discovery.


Subject(s)
Diacylglycerol Kinase/metabolism , Ligands , Adenosine Triphosphate/chemistry , Adenosine Triphosphate/metabolism , Binding Sites , Chromatography, High Pressure Liquid , Diacylglycerol Kinase/chemistry , Diacylglycerol Kinase/genetics , HEK293 Cells , Humans , Isotope Labeling , Ketanserin/chemistry , Ketanserin/metabolism , Peptides/analysis , Protein Binding , Protein Isoforms/chemistry , Protein Isoforms/genetics , Protein Isoforms/metabolism , Proteome/analysis , Recombinant Proteins/biosynthesis , Recombinant Proteins/chemistry , Recombinant Proteins/isolation & purification , Ritanserin/chemistry , Ritanserin/metabolism , Tandem Mass Spectrometry
3.
J Pharmacol Sci ; 133(2): 110-113, 2017 Feb.
Article in English | MEDLINE | ID: mdl-28233634

ABSTRACT

We previously demonstrated that chotosan (CTS), a traditional herbal formula called Kampo medicine, improves diabetes-induced cognitive deficits. In the present study, we investigated the antidepressant-like effects of CTS in mice. The administration of CTS (1.0 g/kg, for 3 days) decreased the immobility time in the forced-swim test, and this decrease was prevented by the prior administration of sulpiride (an antagonist of D2/3 receptors) and WAY100635 (an antagonist of 5-HT1A receptors). None of the treatments tested altered the locomotor activity of mice. These results suggest that CTS exerts antidepressant-like effects through changes in the serotonergic and dopaminergic systems.


Subject(s)
Antidepressive Agents/pharmacology , Dopamine Agents/pharmacology , Drugs, Chinese Herbal/pharmacology , Medicine, Kampo , Serotonin Agents/pharmacology , Animals , Disease Models, Animal , Fenclonine/chemistry , Imipramine/chemistry , Imipramine/pharmacology , Ketanserin/chemistry , Ketanserin/pharmacology , Locomotion , Male , Metergoline/chemistry , Mice , Piperazines/chemistry , Piperazines/pharmacology , Pyridines/chemistry , Pyridines/pharmacology , Sulpiride/chemistry , Sulpiride/pharmacology , Swimming , Yohimbine/chemistry
4.
PLoS One ; 9(11): e109535, 2014.
Article in English | MEDLINE | ID: mdl-25390040

ABSTRACT

Marine sediments can contain B vitamins, presumably incorporated from settled, decaying phytoplankton and microorganisms associated with decomposition. Because B vitamins may be advantageous for the energetically intensive processes of metamorphosis, post-metamorphic growth, and reproduction, we tested several B vitamins to determine if they would stimulate larvae of the deposit-feeding polychaete Capitella teleta to settle and metamorphose. Nicotinamide and riboflavin individually stimulated larvae of C. teleta to settle and metamorphose, generally within 1-2 hours at nicotinamide concentrations as low as 3 µM and riboflavin concentrations as low as 50 µM. More than 80% of the larvae metamorphosed within 30 minutes at a nicotinamide concentration of 7 µM. The pyridine channel agonist pyrazinecarboxamide also stimulated metamorphosis at very low concentrations. In contrast, neither lumichrome, thiamine HCl, pyridoxine HCl, nor vitamin B12 stimulated larvae of C. teleta to metamorphose at concentrations as high as 500 µM. Larvae also did not metamorphose in response to either nicotinamide or pyrazinecarboxamide in calcium-free seawater or with the addition of 4-acetylpyridine, a competitive inhibitor of the pyridine receptor. Together, these results suggest that larvae of C. teleta are responding to nicotinamide and riboflavin via a chemosensory pyridine receptor similar to that previously reported to be present on crayfish chela and involved with food recognition. Our data are the first to implicate B vitamins as possible natural chemical settlement cues for marine invertebrate larvae.


Subject(s)
Ligand-Gated Ion Channels/physiology , Metamorphosis, Biological/drug effects , Niacinamide/chemistry , Polychaeta/drug effects , Polychaeta/embryology , Riboflavin/chemistry , Animals , Biological Assay , Calcium/chemistry , Imidazoles/chemistry , Ketanserin/chemistry , Larva , Pyrazines/chemistry , Pyridines/chemistry , Seawater
5.
Proc Natl Acad Sci U S A ; 111(24): E2492-500, 2014 Jun 17.
Article in English | MEDLINE | ID: mdl-24889602

ABSTRACT

Maternal infection during pregnancy increases the risk for schizophrenia in offspring. In rodent models, maternal immune activation (MIA) yields offspring with schizophrenia-like behaviors. None of these behaviors are, however, specific to schizophrenia. The presence of hallucinations is a key diagnostic symptom of schizophrenia. In mice, this symptom can be defined as brain activation in the absence of external stimuli, which can be mimicked by administration of hallucinogens. We find that, compared with controls, adult MIA offspring display an increased stereotypical behavioral response to the hallucinogen 2,5-dimethoxy-4-iodoamphetamine (DOI), an agonist for serotonin receptor 2A (5-HT2AR). This may be explained by increased levels of 5-HT2AR and downstream signaling molecules in unstimulated MIA prefrontal cortex (PFC). Using manganese-enhanced magnetic resonance imaging to identify neuronal activation elicited by DOI administration, we find that, compared with controls, MIA offspring exhibit a greater manganese (Mn(2+)) accumulation in several brain areas, including the PFC, thalamus, and striatum. The parafascicular thalamic nucleus, which plays the role in the pathogenesis of hallucinations, is activated by DOI in MIA offspring only. Additionally, compared with controls, MIA offspring demonstrate higher DOI-induced expression of early growth response protein 1, cyclooxygenase-2, and brain-derived neurotrophic factor in the PFC. Chronic treatment with the 5-HT2AR antagonist ketanserin reduces DOI-induced head twitching in MIA offspring. Thus, the MIA mouse model can be successfully used to investigate activity induced by DOI in awake, behaving mice. Moreover, manganese-enhanced magnetic resonance imaging is a useful, noninvasive method for accurately measuring this type of activity.


Subject(s)
Amphetamines/chemistry , Contrast Media/chemistry , Hallucinogens/chemistry , Manganese/chemistry , Prefrontal Cortex/physiopathology , Schizophrenia/physiopathology , Animals , Behavior, Animal/drug effects , Brain Mapping , Brain-Derived Neurotrophic Factor/metabolism , Cyclooxygenase 2/metabolism , Disease Models, Animal , Early Growth Response Protein 1/metabolism , Ketanserin/chemistry , Ligands , Magnetic Resonance Imaging , Male , Mice , Mice, Inbred C57BL , Neurons/drug effects , Poly I-C/chemistry , Prefrontal Cortex/drug effects , Receptor, Serotonin, 5-HT2A/metabolism , Risk Factors , Schizophrenia/metabolism
6.
Exp Brain Res ; 230(4): 395-406, 2013 Oct.
Article in English | MEDLINE | ID: mdl-23864045

ABSTRACT

Earlier autoradiographic studies with the 5-HT2 receptor agonist [(125)I](±)DOI in human brain showed unexpected biphasic competition curves for various 5-HT2A antagonists. We have performed similar studies in rat brain regions with selective 5-HT2A (M100907) and 5-HT2C (SB242084) antagonists together with ketanserin and mesulergine. The effect of GTP analogues on antagonist competition was also studied. Increasing concentrations of Gpp(NH)p or GTPγS resulted in a maximal inhibition of [(125)I](±)DOI-specific binding of approximately 50 %. M100907 competed biphasically in all regions. In the presence of 100 µM Gpp(NH)p, M100907 still displaced biphasically the remaining [(125)I](±)DOI binding. Ketanserin showed biphasic curves in some regions and monophasic curves in others. In the latter, Gpp(NH)p evidenced an additional high-affinity site. SB242084 competed biphasically in brainstem nuclei and monophasically in the other regions. In most areas, SB242084 affinities were not notably altered by Gpp(NH)p. Mesulergine competed monophasically in all regions without alteration by Gpp(NH)p. These results conform with the extended ternary complex model of receptor action: receptor exists as an equilibrium of multiple conformations, i.e. ground (R), partly activated (R*) and activated G-protein-coupled (R*G) conformation/s. Thus, [(125)I](±)DOI would label multiple conformations of both 5-HT2A and 5-HT2C receptors in rat brain, and M100907 and ketanserin would recognise these conformations with different affinities.


Subject(s)
Brain/drug effects , Receptor, Serotonin, 5-HT2A/chemistry , Receptor, Serotonin, 5-HT2C/chemistry , Aminopyridines/chemistry , Aminopyridines/pharmacology , Animals , Autoradiography/methods , Brain/metabolism , Ergolines/chemistry , Ergolines/pharmacology , Indoles/chemistry , Indoles/pharmacology , Ketanserin/chemistry , Ketanserin/pharmacology , Male , Rats , Rats, Wistar , Receptor, Serotonin, 5-HT2A/metabolism , Receptor, Serotonin, 5-HT2C/metabolism , Serotonin/metabolism
7.
J Med Chem ; 55(12): 5749-59, 2012 Jun 28.
Article in English | MEDLINE | ID: mdl-22694093

ABSTRACT

Of great interest in recent years has been computationally predicting the novel polypharmacology of drug molecules. Here, we applied an "induced-fit" protocol to improve the homology models of 5-HT(2A) receptor, and we assessed the quality of these models in retrospective virtual screening. Subsequently, we computationally screened the FDA approved drug molecules against the best induced-fit 5-HT(2A) models and chose six top scoring hits for experimental assays. Surprisingly, one well-known kinase inhibitor, sorafenib, has shown unexpected promiscuous 5-HTRs binding affinities, K(i) = 1959, 56, and 417 nM against 5-HT(2A), 5-HT(2B), and 5-HT(2C), respectively. Our preliminary SAR exploration supports the predicted binding mode and further suggests sorafenib to be a novel lead compound for 5HTR ligand discovery. Although it has been well-known that sorafenib produces anticancer effects through targeting multiple kinases, carefully designed experimental studies are desirable to fully understand whether its "off-target" 5-HTR binding activities contribute to its therapeutic efficacy or otherwise undesirable side effects.


Subject(s)
Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Benzenesulfonates/chemistry , Drug Discovery , Pyridines/chemistry , Receptors, Serotonin/metabolism , Serotonin Antagonists/chemistry , Serotonin Antagonists/pharmacology , Antineoplastic Agents/metabolism , Cyproheptadine/chemistry , Cyproheptadine/metabolism , Cyproheptadine/pharmacology , Drug Approval , Ketanserin/chemistry , Ketanserin/metabolism , Ketanserin/pharmacology , Molecular Dynamics Simulation , Niacinamide/analogs & derivatives , Phenylurea Compounds , Protein Conformation/drug effects , Receptors, Serotonin/chemistry , Sequence Homology, Amino Acid , Serotonin Antagonists/metabolism , Sorafenib , United States , United States Food and Drug Administration/legislation & jurisprudence , User-Computer Interface
8.
Nucl Med Biol ; 39(7): 1087-92, 2012 Oct.
Article in English | MEDLINE | ID: mdl-22633218

ABSTRACT

INTRODUCTION: Our aim was the optimization of radiochemical parameters for the microfluidic preparation of [(18)F]altanserin. The four main parameters evaluated were (1) precursor concentration, (2) reaction temperature, (3) bolus flow rate through the microreactor and (4) bolus volume. METHODS: For the determination of optimal reaction conditions within a flow-through microreactor synthesizer, 5-400 µL of precursor and dried [(18)F]fluoride solution were simultaneously pushed through the temperature-controlled reactor (180-220°C) with defined bolus flow rates of 10-60 µL/min. Radiochemical incorporation yields (RCIYs) were examined using a thin layer chromatography (TLC) set-up and radio- high-performance liquid chromatography (HPLC). RESULTS: Optimum reaction parameters for the microfluidic set-up were determined as following: 220°C, 5-10 µL/min pump rate per reactant (10-20 µL/min reaction overall flow rate) and 2mg/mL precursor concentration in the reaction mixture. Applying these optimized conditions, RCIYs of 53.7 ± 7.9 were observed for scaled-up preparations. A positive "bolus effect" was observed: applying higher reaction volume resulted in increased RCIYs. CONCLUSION: This study proved that the reaction bolus volume is an essential parameter influencing the RCIY of [(18)F]altanserin. A possible explanation is the inhomogeneous distribution within the reaction volume probably caused by diffusion at the bolus interface. This important finding should be considered an important variable for the evaluation of all novel radiotracers labeled using a flow-through reactor device.


Subject(s)
Fluorine Radioisotopes , Isotope Labeling/instrumentation , Ketanserin/analogs & derivatives , Microfluidic Analytical Techniques/instrumentation , Ketanserin/chemistry , Radiochemistry , Temperature
9.
PLoS Comput Biol ; 8(4): e1002473, 2012.
Article in English | MEDLINE | ID: mdl-22532793

ABSTRACT

From computational simulations of a serotonin 2A receptor (5-HT(2A)R) model complexed with pharmacologically and structurally diverse ligands we identify different conformational states and dynamics adopted by the receptor bound to the full agonist 5-HT, the partial agonist LSD, and the inverse agonist Ketanserin. The results from the unbiased all-atom molecular dynamics (MD) simulations show that the three ligands affect differently the known GPCR activation elements including the toggle switch at W6.48, the changes in the ionic lock between E6.30 and R3.50 of the DRY motif in TM3, and the dynamics of the NPxxY motif in TM7. The computational results uncover a sequence of steps connecting these experimentally-identified elements of GPCR activation. The differences among the properties of the receptor molecule interacting with the ligands correlate with their distinct pharmacological properties. Combining these results with quantitative analysis of membrane deformation obtained with our new method (Mondal et al, Biophysical Journal 2011), we show that distinct conformational rearrangements produced by the three ligands also elicit different responses in the surrounding membrane. The differential reorganization of the receptor environment is reflected in (i)-the involvement of cholesterol in the activation of the 5-HT(2A)R, and (ii)-different extents and patterns of membrane deformations. These findings are discussed in the context of their likely functional consequences and a predicted mechanism of ligand-specific GPCR oligomerization.


Subject(s)
Cell Membrane/chemistry , Models, Chemical , Models, Molecular , Receptor, Serotonin, 5-HT2A/chemistry , Receptor, Serotonin, 5-HT2A/ultrastructure , Binding Sites , Computer Simulation , Ketanserin/chemistry , Ligands , Lysergic Acid Diethylamide/chemistry , Protein Binding , Protein Conformation , Serotonin/chemistry , Structure-Activity Relationship
10.
PLoS One ; 7(2): e32656, 2012.
Article in English | MEDLINE | ID: mdl-22384279

ABSTRACT

Increased residual platelet reactivity remains a burden for coronary artery disease (CAD) patients who received a coronary stent and do not respond sufficiently to treatment with acetylsalicylic acid and clopidogrel. We hypothesized that serotonin antagonism reduces high on-treatment platelet reactivity. Whole blood impedance aggregometry was performed with arachidonic acid (AA, 0.5 mM) and adenosine diphosphate (ADP, 6.5 µM) in addition to different concentrations of serotonin (1-100 µM) in whole blood from 42 CAD patients after coronary stent placement and 10 healthy subjects. Serotonin increased aggregation dose-dependently in CAD patients who responded to clopidogrel treatment: After activation with ADP, aggregation increased from 33.7 ± 1.3% to 40.9 ± 2.0% in the presence of 50 µM serotonin (p<0.05) and to 48.2 ± 2.0% with 100 µM serotonin (p<0.001). The platelet serotonin receptor antagonist ketanserin decreased ADP-induced aggregation significantly in clopidogrel low-responders (from 59.9 ± 3.1% to 37.4 ± 3.5, p<0.01), but not in clopidogrel responders. These results were confirmed with light transmission aggregometry in platelet-rich plasma in a subset of patients. Serotonin hence increased residual platelet reactivity in patients who respond to clopidogrel after coronary stent placement. In clopidogrel low-responders, serotonin receptor antagonism improved platelet inhibition, almost reaching responder levels. This may justify further investigation of triple antiplatelet therapy with anti-serotonergic agents.


Subject(s)
Blood Platelets/drug effects , Serotonin Antagonists/pharmacology , Serotonin/chemistry , Stents , Ticlopidine/analogs & derivatives , Aged , Aspirin/pharmacology , Clopidogrel , Coronary Artery Disease/drug therapy , Female , Humans , In Vitro Techniques , Ketanserin/chemistry , Light , Male , Middle Aged , Myocardial Infarction/metabolism , Pilot Projects , Platelet Aggregation/drug effects , Platelet Aggregation Inhibitors/pharmacology , Ticlopidine/pharmacology , Treatment Outcome
11.
J Vet Pharmacol Ther ; 35(5): 472-7, 2012 Oct.
Article in English | MEDLINE | ID: mdl-22091605

ABSTRACT

The purpose of this study was to determine the pharmacokinetics (PK) of the 5-HT(2A) receptor antagonist ketanserin in healthy adult horses, and to develop a computational model that could be used to optimize dosing. Plasma concentrations of ketanserin were determined using liquid chromatography with mass spectrometry after single and multiple intravenous administration in the horse. A two-compartment linear pharmacokinetic model described the plasma concentration-time profile of ketanserin after single and multiple doses in healthy horses; the terminal half-life was 11.5 h; steady-state volume of distribution was 10.5 L/kg; AUC was 115 ng · h/mL; and clearance was 0.87 L/h/kg. Model simulations followed by the examination in three healthy horses suggest 0.3 mg/kg q.8 h exhibited linear PK and produced consistent systemic blood concentrations of ketanserin above 3 ng/mL.


Subject(s)
Horses/blood , Ketanserin/pharmacokinetics , Platelet Aggregation Inhibitors/pharmacokinetics , Animals , Area Under Curve , Female , Half-Life , Horses/metabolism , Ketanserin/blood , Ketanserin/chemistry , Molecular Structure , Platelet Aggregation Inhibitors/blood , Platelet Aggregation Inhibitors/chemistry
12.
Chem Biol Drug Des ; 76(4): 361-6, 2010 Oct.
Article in English | MEDLINE | ID: mdl-20636331

ABSTRACT

MH.MZ, MDL 100907, and altanserin are structurally similar 4-benzoyl-piperidine derivatives and are well accommodated to receptor interaction models. We combined structural elements of different high-affinity and selective 5-HT(2A) antagonists, as MH.MZ, altanserin, and SR 46349B, to improve the binding properties of new compounds. Three new derivatives were synthesized with a 4-benzoyl-piperidine moiety as the lead structure. The in vitro affinity of the novel compounds was determined by a [³H]MDL 100907 competition binding assay. The combination of MH.MZ and SR 46349B resulted in a compound (8) with a moderate affinity toward the 5-HT(2A) receptor (K(i) = 57 nm). The remarkably reduced affinity of other compounds (4a), (4b), and (4c) (K(i) = 411, 360 and 356 nm respectively) indicates that MH.MZ can only bind to the 5-HT(2A) receptor with the p-fluorophenylethyl residue in a sterically restricted hydrophobic binding pocket.


Subject(s)
Ligands , Receptor, Serotonin, 5-HT2A/chemistry , Serotonin 5-HT2 Receptor Antagonists/chemistry , Binding, Competitive , Combinatorial Chemistry Techniques , Fluorobenzenes/chemistry , Hydrophobic and Hydrophilic Interactions , Ketanserin/analogs & derivatives , Ketanserin/chemistry , Phenols/chemistry , Piperidines/chemistry , Protein Binding , Pyridines/chemistry , Receptor, Serotonin, 5-HT2A/metabolism
13.
Appl Radiat Isot ; 67(11): 2040-3, 2009 Nov.
Article in English | MEDLINE | ID: mdl-19692252

ABSTRACT

[(18)F]Altanserin, an important PET radioligand for the in vivo imaging of the 5-HT(2A) receptor, was synthesized from its precursor nitro-altanserin in DMF or DMSO at high temperatures of 150 degrees C in an overall radiochemical yield (EOB) of 23-25% after 75min. A new solid phase work-up procedure involving the acidification of the crude reaction mixture and a C18-SepPak-solid phase separation preceded the final HPLC purification. This led to a significantly reduced synthesis time as a result of a stable and early elution from the HPLC column using improved HPLC conditions (MeOH/THF/NaOAc 0.05N pH 5: 27/18/55, flow: 5 mL/min, Symetry Prep 7 microm C18 (Waters)). The synthesis was performed semi-automatically in a modified GE TracerLab synthesis module using an in-house-developed program. The synthesized [(18)F]altanserin was used in our ongoing human and animal PET imaging studies.


Subject(s)
Brain/diagnostic imaging , Brain/metabolism , Fluorine Radioisotopes/pharmacokinetics , Ketanserin/analogs & derivatives , Positron-Emission Tomography/methods , Receptor, Serotonin, 5-HT2A/metabolism , Animals , Fluorine Radioisotopes/chemistry , Humans , Isotope Labeling/methods , Ketanserin/chemistry , Ketanserin/pharmacokinetics , Radiopharmaceuticals/chemical synthesis , Time Factors
14.
Eur J Nucl Med Mol Imaging ; 36(2): 287-93, 2009 Feb.
Article in English | MEDLINE | ID: mdl-18828013

ABSTRACT

PURPOSE: The serotonin 2A (5-HT(2A)) receptor is of interest in several psychiatric and neurological diseases. In the present study we investigated the longitudinal stability of 5-HT(2A) receptors and the stability of the quantification procedure in the elderly in order to be able to study elderly patients with neuropsychiatric diseases on a longitudinal basis. METHODS: [(18)F]-Altanserin PET was used to quantify 5-HT(2A) receptors in 12 healthy elderly individuals at baseline and at 2 years in six volumes of interest. A bolus/infusion protocol was used to achieve the binding potential, BP(P). The reproducibility as assessed in terms of variability and the reliability as assessed in terms of intraclass correlation coefficient (ICC) were used to compare inter- and intraobserver stability and to evaluate the effects of increasing complexity of partial volume (PV) corrections. We also compared the stability of our measurements over 2 years with the stability of data from an earlier study with 2-week test-retest measurements. RESULTS: BP(P) was unaltered at follow-up without the use of PV correction and when applying two-tissue PV correction, test-retest reproducibility was 12-15% and reliability 0.45-0.67 in the large bilateral regions such as the parietal, temporal, occipital and frontal cortices, while orbitofrontal and anterior cingulate cortical regions were less stable. The use of PV correction decreased the variability but also decreased the between-subject variation, thereby worsening the reliability. CONCLUSION: In healthy elderly individuals, brain 5-HT(2A) receptor binding remains stable over 2 years, and acceptable reproducibility and reliability in larger regions and high intra- and interobserver stability allow the use of [(18)F]-altanserin in longitudinal studies of patients with neuropsychiatric disorders.


Subject(s)
Cerebrum/diagnostic imaging , Cerebrum/metabolism , Fluorine Radioisotopes/chemistry , Ketanserin/analogs & derivatives , Receptor, Serotonin, 5-HT2A/metabolism , Aged , Female , Health , Humans , Ketanserin/chemistry , Longitudinal Studies , Male , Middle Aged , Observer Variation , Positron-Emission Tomography/statistics & numerical data , Reproducibility of Results
15.
Appl Radiat Isot ; 64(9): 989-94, 2006 Sep.
Article in English | MEDLINE | ID: mdl-16829074

ABSTRACT

A new type of electrochemical cell with anodic deposition of no-carrier-added [(18)F]fluoride and variable reaction volume has been developed. The reactor is designed for small reaction volumes and non-thermal drying of [(18)F]fluoride. The implementation of this reactor into a complete remotely controlled synthesis device is described for the routine production of [(18)F]altanserin. A radiochemical yield of 23+/-5% was obtained via cryptate-mediated nucleophilic (18)F-fluorination. Batches of up to 6 GBq [(18)F]altanserin, suitable for human application, with a molar activity of >500 GBq/micromol were obtained within 75 min.


Subject(s)
Fluorine Radioisotopes , Isotope Labeling/methods , Ketanserin/analogs & derivatives , Chromatography, High Pressure Liquid , Electrochemistry/instrumentation , Ketanserin/chemistry , Ketanserin/isolation & purification
16.
Biol Pharm Bull ; 23(5): 558-65, 2000 May.
Article in English | MEDLINE | ID: mdl-10823664

ABSTRACT

The in vivo and in vitro cardiovascular effects of the novel 5-HT2A/alpha1/H1 antagonist ketanserin analogues QF 0303B, QF 0307B, QF 0311B, QF 0313B were studied in anaesthetized normotensive rats (ANR) and in isolated rubbed rat aorta (IRRA). In ANR, 0.2 mg x kg(-1) i.v. of each compound produced a rapid, remarkable but short-lasting fall in mean arterial blood pressure (MAP) accompanied by bradycardia. All compounds significantly modified the pressor effects induced by 5-hydroxytryptamine (5-HT) and noradrenaline (NA). In IRRA, the compounds inhibited NA- and 5-HT-induced contractions in a competitive fashion. Furthermore, the analogues displayed lower H1-antagonist activity than ketanserin. Compounds tested showed low 5-HT2B affinity and no activity at muscarinic, nicotinic, or 5-HT3 receptors, nor any marked ability to produce smooth muscle relaxation via calcium entry blockade. There is a significant correlation between hypotension reached and inhibition of the 5-HT-induced pressor responses (but not for NA). A certain degree of correlation was observed between hypotensive effect endurance vs. alpha1-adrenoceptor blockade (but not for serotonin). These results indicate that in this series the brief hypotensive activity in ANR is attributed to a 5-HT1A receptor blockade and the duration of the effect is better attributed to an alpha1 adrenoceptor blockade.


Subject(s)
Antihypertensive Agents/pharmacology , Ketanserin/pharmacology , Receptors, Adrenergic, alpha-1/metabolism , Receptors, Serotonin/metabolism , Adrenergic alpha-Antagonists/pharmacology , Animals , Antihypertensive Agents/chemistry , Aorta , Guinea Pigs , Ileum/drug effects , Ileum/metabolism , Ketanserin/analogs & derivatives , Ketanserin/chemistry , Male , Muscles/drug effects , Muscles/metabolism , Potassium Chloride/pharmacology , Rats , Rats, Sprague-Dawley , Receptor, Serotonin, 5-HT2A , Receptor, Serotonin, 5-HT2B , Receptors, Adrenergic, alpha-1/drug effects , Receptors, Histamine H1/drug effects , Receptors, Histamine H1/metabolism , Receptors, Serotonin/drug effects , Receptors, Serotonin, 5-HT3 , Serotonin Antagonists/pharmacology , Vasoconstriction/drug effects
17.
Bioorg Med Chem ; 6(6): 785-8, 1998 Jun.
Article in English | MEDLINE | ID: mdl-9681143

ABSTRACT

This theoretical analysis shows that the experimentally observed standard Gibbs free energy of binding of a ligand by a receptor can be described by two terms. One term describes the free energy of binding of the drug to the receptor when both are in their lowest energy conformation. The second term gives the difference between the average and the lowest conformational energy of the two species involved. It also follows that all drug molecules having an energy higher than the minimum energy, must have a higher affinity than molecules occurring in the minimum energy conformation, independent of the energy level of the receptor bound conformation.


Subject(s)
Proteins/metabolism , Ketanserin/chemistry , Ligands , Molecular Conformation , Protein Binding , Serotonin Antagonists/chemistry , Thermodynamics
18.
J Biol Chem ; 272(41): 26049-55, 1997 Oct 10.
Article in English | MEDLINE | ID: mdl-9325342

ABSTRACT

The full-length cDNA for the rat recombinant synaptic vesicle monoamine transporter (rVMAT2) containing a COOH-terminal polyhistidine epitope was engineered into baculovirus DNA for expression in Spodoptera frugiperda (Sf9) cells. Using this recombinant baculovirus and cultured Sf9 cells, rVMAT2 has been expressed to high levels and purified to >95% homogeneity using immobilized Ni2+-affinity chromatography followed by lectin (concanavalin A) chromatography. Purified transporter was photolabeled using [125I]-7-azido-8-iodoketanserin ([125I]AZIK) and [125I]2-N-[(3'-iodo-4'-azidophenyl)propionyl]tetrabenazine ([125I]TBZ-AIPP). Both [125I]AZIK and [125I]TBZ-AIPP photoaffinity labeling of purified rVMAT2 were protectable by 10 microM tetrabenazine (TBZ), 10 microM 7-aminoketanserin, and 1 mM concentrations of the transporter substrates dopamine, norepinephrine, and serotonin. Radiolabeled peptides were generated using enzymatic and chemical methods, purified using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and NH2-terminal microsequenced. Radiosequencing of [125I]AZIK-labeled rVMAT2 indicated derivatization of Lys-20 in the NH2 terminus, just prior to putative transmembrane domain 1 (TMD1). [125I]TBZ-AIPP derivatized a segment of rVMAT2 between Gly-408 and Cys-431 in TMD10 and 11. These data implicate juxtaposition of TMD1 and 10/11.


Subject(s)
Affinity Labels/metabolism , Azides/metabolism , Ketanserin/analogs & derivatives , Membrane Glycoproteins/metabolism , Membrane Transport Proteins , Neuropeptides , Neurotransmitter Agents/metabolism , Synaptic Vesicles/metabolism , Tetrabenazine/analogs & derivatives , Affinity Labels/chemistry , Animals , Azides/chemistry , Binding Sites , Biological Transport , Iodine Radioisotopes , Ketanserin/chemistry , Ketanserin/metabolism , Membrane Glycoproteins/chemistry , Models, Molecular , Neurotransmitter Agents/chemistry , Peptide Mapping , Protein Binding , Rats , Recombinant Proteins/metabolism , Spodoptera , Synaptic Vesicles/chemistry , Tetrabenazine/chemistry , Tetrabenazine/metabolism , Vesicular Biogenic Amine Transport Proteins
19.
Eur J Pharmacol ; 306(1-3): 195-210, 1996 Jun 13.
Article in English | MEDLINE | ID: mdl-8813633

ABSTRACT

Molecular modeling techniques were used to build a three-dimensional model of the rat 5-HT2C receptor, which was used to examine receptor interactions for protonated forms of serotonin, ketanserin and ritanserin. Molecular dynamics simulations which were started with the fluoro benzene moiety of ketanserin and ritanserin oriented towards the cytoplasmic side of the receptor model, produced the strongest antagonist-receptor interactions. The fluoro bezene ring(s) of the antagonists interacted strongly with aromatic residues in the receptor model, which predicts slightly different orientations and ligand-receptor interactions of ketanserin and ritanserin at a putative binding site. The model suggests that Asn333 (transmembrane helix 6) is involved in a hydrogen-bonding interaction with ketanserin, but not with ritanserin. The model also also suggests that the position corresponding to Cys362 (transmembrane helix 7) may be an important determinant for specifying 5-HT2A receptor selectivity in ketanserin binding.


Subject(s)
Ketanserin/chemistry , Models, Molecular , Receptors, Serotonin/chemistry , Ritanserin/chemistry , Serotonin Antagonists/chemistry , Serotonin/chemistry , Animals , Ketanserin/metabolism , Models, Chemical , Molecular Conformation , Rats , Receptors, Serotonin/metabolism , Ritanserin/metabolism , Serotonin/metabolism , Serotonin Antagonists/metabolism , Structure-Activity Relationship
20.
Nucl Med Biol ; 23(4): 429-38, 1996 May.
Article in English | MEDLINE | ID: mdl-8832697

ABSTRACT

Starting from the lead structure of ketanserin, a prototypic serotonin (5-HT) antagonist, new oxotechnetium(V) and oxorhenium(V) complexes were synthesized that are able to compete with [3H]ketan-serin in receptor-binding assays. To imitate organic 5-HT2 receptor ligands, fragments of ketanserin were combined with chelate moieties. Neutral compounds of the general formula [MOL1L2] (M = Tc, Re; L1 = HS-CH2CH2-S-CH2CH2-SH, N-(2-mercaptophenyl)salicylideneimine, N-(2-mercaptoethyl)-salicylideneimine, 3-(2-([N,N-bis(2-mercapto-S-ethyl)]-amino)ethyl)-2,4-(1H, 3H)-quinazolinedione and L2 = HS-R with R = subst. alkyl) were prepared by common action of a Tc(V) or Re(V) precursor with a mixture of equimolar amounts of a tridentate ligand L1 and a monodentate thiolate L2 bearing fragments of the lead structure. Lipophilic complexes consisting of a small S4 thiolate/thioether chelate unit, protonable nitrogen-containing spacer, and simple benzyl moiety significantly inhibited the specific binding of [3H]ketan-serin with IC50 values between 10 and 50 nM.


Subject(s)
Organometallic Compounds/metabolism , Organotechnetium Compounds/metabolism , Receptors, Serotonin/metabolism , Rhenium/metabolism , Animals , Binding, Competitive , Brain/metabolism , Brain/ultrastructure , Ketanserin/chemistry , Ketanserin/metabolism , Male , Organometallic Compounds/pharmacokinetics , Organotechnetium Compounds/pharmacokinetics , Rats , Rats, Wistar , Receptor, Serotonin, 5-HT2A , Rhenium/pharmacokinetics , Structure-Activity Relationship , Tissue Distribution
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