PET Imaging of VMAT2 with the Novel Radioligand [18F]FE-DTBZ-d4 in Nonhuman Primates: Comparison with [11C]DTBZ and [18F]FE-DTBZ.

The vesicular monoamine transporter type 2 (VMAT2) is believed to be responsible for the uptake of monoamines into the vesicles of the synaptic terminals. Two VMAT2 radioligands [11C]DTBZ and [18F]FP-DTBZ have been used to assess the degree of nigrostriatal deficit in Parkinson's disease (PD) using positron emission tomography (PET). [18F]FE-DTBZ-d4, the nondeuterated analogue of [18F]FE-DTBZ showed similar imaging properties with better stability against defluorination. Therefore, [18F]FE-DTBZ-d4 draws attention to be investigated as an imaging marker for VMAT2 in the brain. The aim of this study was to investigate the brain kinetics and quantification of [18F]FE-DTBZ-d4 in nonhuman primates (NHPs), with comparison to [11C]DTBZ and [18F]FE-DTBZ. Radiolabeling was successfully achieved either by one-step 11C-methylation or by a two-step fluorine-18 nucleophilic substitution reaction. The stability and radiochemical yield were analyzed with high-performance liquid chromatography (HPLC). Three female cynomolgus monkeys were included in the study and underwent a total of 12 positron emission tomography (PET) measurements. Each monkey was examined with each tracer. In addition, two pretreatment and one displacement PET measurements with tetrabenazine (2.0 mg/kg) were performed for [18F]FE-DTBZ-d4. All PET measurements were conducted using a high-resolution research tomograph (HRRT) system. Radiometabolites were measured in monkey plasma using gradient radio-HPLC. [18F]FE-DTBZ-d4 (SUV: 4.28 ± 1.01) displayed higher brain uptake compared to both [18F]FE-DTBZ (SUV: 3.43 ± 0.54) and [11C]DTBZ (SUV: 3.06 ± 0.32) and faster washout. Binding potential (BPND) values of [18F]FE-DTBZ-d4 in different brain regions (putamen: 5.5 ± 1.4; caudate: 4.4 ± 1.1; midbrain: 1.4 ± 0.4) were higher than those of [11C]DTBZ and [18F]FE-DTBZ. [18F]FE-DTBZ showed faster radiometabolism in plasma compared to [11C]DTBZ and [18F]FE-DTBZ-d4. [18F]FE-DTBZ-d4 is a suitable radioligand for quantification of VMAT2 in the nonhuman primate brain, with better imaging properties than [11C]DTBZ and [18F]FE-DTBZ. A preliminary comparison suggests that [18F]FE-DTBZ-d4 has increased stability against defluorination compared to the nondeuterated analogue.

A Nonhuman Primate PET Study: Measurement of Brain PDE4 Occupancy by Roflumilast Using (R)-[11C]Rolipram.

PURPOSE: Phosphodiesterase 4 (PDE4) inhibition in the brain has been reported to improve cognitive function in animal models. Therefore, PDE4 inhibitors are one of key targets potential for drug development. Investigation of brain PDE4 occupancy would help to understand the effects of PDE4 inhibition to cognitive functions. Roflumilast is a selective phosphodiesterase type 4 (PDE4) inhibitor used clinically for severe chronic obstructive pulmonary disease, but the effects to the brain have not been well investigated. In this study, we aimed to investigate whether roflumilast entered the brain and occupied PDE4 in nonhuman primates. PROCEDURES: Positron emission tomography (PET) measurements with (R)-[11C]rolipram were performed at baseline and after intravenous (i.v.) administration of roflumilast (3.6 to 200 µg/kg) in three female rhesus monkeys. Arterial blood samples were taken to obtain the input function. Protein binding was measured to obtain the free fraction (fp) of the radioligand. Total distribution volume (VT) and VT/fp were calculated as outcome measures from two tissue compartment model. Lassen plot approach was taken to estimate the target occupancy. RESULTS: The brain uptake of (R)-[11C]rolipram decreased after roflumilast administration. PDE 4 occupancy by roflumilast showed dose- and plasma concentration-dependent increase, although PDE4 occupancy did not reach 50 % even after the administration of up to 200 µg/kg of roflumilast, regardless of outcome measures, VT or VT/fp. CONCLUSIONS: This PET study showed that the brain PDE4 binding was blocked to a certain extent after i.v. administration of clinical relevant doses of roflumilast in nonhuman primates. Further clinical PET evaluation is needed to understand the relationship between PDE4 inhibition and potential improvement of cognitive function in human subjects.

Development of [ Carbonyl-11C]AZ13198083, a Novel Histamine Type-3 Receptor Radioligand with Favorable Kinetics.

The histamine subtype-3 receptor (H3R) is implicated in a range of central nervous system disorders, and several radioligands have been developed for H3R positron emission tomography imaging. However, a limitation of currently used PET radioligands for H3R is the slow binding kinetics in high density brain regions. To address this, we herein report the development of three novel candidate H3R radioligands, namely, [ carbonyl-11C]AZ13153556 ([ carbonyl-11C]4), [ carbonyl-11C]AZD5213([ carbonyl-11C]5), and [ carbonyl-11C]AZ13198083 ([ carbonyl-11C]6), and their subsequent preclinical evaluation in nonhuman primates (NHP). Radioligands [ carbonyl-11C]4-6 were produced and isolated in high radioactivity (>1000 MBq), radiochemical purity (>99%), and moderate molar activity (19-28 GBq/µmol at time of injection) using a palladium-mediated 11C-aminocarbonylation protocol. All three radioligands showed high brain permeability as well as a regional brain radioactivity distribution in accordance with H3R expression (striatum > cortex > cerebellum). [ Carbonyl-11C]6 displayed the most favorable in vivo kinetics and brain uptake, with an early peak in the striatal time-activity curve followed by a progressive washout from the brain. The specificity and on-target kinetics of [ carbonyl-11C]6 were next investigated in pretreatment and displacement studies. After pretreatment or displacement with 5 (0.1 mg/kg), a uniformly low distribution of radioactivity across the NHP brain was observed. Collectively, this work demonstrates that [ carbonyl-11C]6 is a promising candidate for H3R imaging in human subjects.

Development of two fluorine-18 labeled PET radioligands targeting PDE10A and in vivo PET evaluation in nonhuman primates.

INTRODUCTION: Phosphodiesterase 10A (PDE10A) is a member of the PDE enzyme family that degrades cyclic adenosine and guanosine monophosphates (cAMP and cGMP). Based on the successful development of [11C]T-773 as PDE10A positron emission tomography (PET) radioligand, in this study our aim was to develop and evaluate fluorine-18 analogs of [11C]T-773. METHODS: [18F]FM-T-773-d2 and [18F]FE-T-773-d4 were synthesized from the same precursor used for 11C-labeling of T-773 in a two-step approach via 18F-fluoromethylation and 18F-fluoroethylation, respectively, using corresponding deuterated synthons. A total of 12 PET measurements were performed in seven non-human primates. First, baseline PET measurements were performed using High Resolution Research Tomograph system with both [18F]FM-T-773-d2 and [18F]FE-T-773-d4; the uptake in whole brain and separate brain regions, as well as the specific binding and tissue ratio between putamen and cerebellum, was examined. Second, baseline and pretreatment PET measurements using MP-10 as the blocker were performed for [18F]FM-T-773-d2 including arterial blood sampling with radiometabolite analysis in four NHPs. RESULTS: Both [18F]FM-T-773-d2 and [18F]FE-T-773-d4 were successfully radiolabeled with an average molar activity of 293 ± 114 GBq/µmol (n=8) for [18F]FM-T-773-d2 and 209 ± 26 GBq/µmol (n=4) for [18F]FE-T-773-d4, and a radiochemical yield of 10% (EOB, n=12, range 3%-16%). Both radioligands displayed high brain uptake (~5.5% of injected radioactivity for [18F]FM-T-773-d2 and ~3.5% for [18F]FE-T-773-d4 at the peak) and a fast washout. Specific binding reached maximum within 30 min for [18F]FM-T-773-d2 and after approximately 45 min for [18F]FE-T-773-d4. [18F]FM-T-773-d2 data fitted well with kinetic compartment models. BPND values obtained indirectly through compartment models were correlated well with those obtained by SRTM. BPND calculated with SRTM was 1.0-1.7 in the putamen. The occupancy with 1.8 mg/kg of MP-10 was approximately 60%. CONCLUSIONS: [18F]FM-T-773-d2 and [18F]FE-T-773-d4 were developed as fluorine-18 PET radioligands for PDE10A, with the [18F]FM-T-773-d2 being the more promising PET radioligand warranting further evaluation.

Positron emission tomography measurement of brain MAO-B inhibition in patients with Alzheimer's disease and elderly controls after oral administration of sembragiline.

PURPOSE: In Alzheimer's disease (AD), increased metabolism of monoamines by monoamine oxidase type B (MAO-B) leads to the production of toxic reactive oxygen species (ROS), which are thought to contribute to disease pathogenesis. Inhibition of the MAO-B enzyme may restore brain levels of monoaminergic neurotransmitters, reduce the formation of toxic ROS and reduce neuroinflammation (reactive astrocytosis), potentially leading to neuroprotection. Sembragiline (also referred as RO4602522, RG1577 and EVT 302 in previous communications) is a potent, selective and reversible inhibitor of MAO-B developed as a potential treatment for AD. METHODS: This study assessed the relationship between plasma concentration of sembragiline and brain MAO-B inhibition in patients with AD and in healthy elderly control (EC) subjects. Positron emission tomography (PET) scans using [11C]-L-deprenyl-D2 radiotracer were performed in ten patients with AD and six EC subjects, who received sembragiline each day for 6-15 days. RESULTS: At steady state, the relationship between sembragiline plasma concentration and MAO-B inhibition resulted in an Emax of ∼80-90 % across brain regions of interest and in an EC50 of 1-2 ng/mL. Data in patients with AD and EC subjects showed that near-maximal inhibition of brain MAO-B was achieved with 1 mg sembragiline daily, regardless of the population, whereas lower doses resulted in lower and variable brain MAO-B inhibition. CONCLUSIONS: This PET study confirmed that daily treatment of at least 1 mg sembragiline resulted in near-maximal inhibition of brain MAO-B enzyme in patients with AD.

Brain PET measurement of PDE10A occupancy by TAK-063, a new PDE10A inhibitor, using [11 C]T-773 in nonhuman primates.

Because phosphodiesterase 10A (PDE10A) degrades both cyclic adenosine monophosphate and cyclic guanosine monophosphate and is distributed mainly in the striatum, PDE10A inhibitors have been considered to potentially be useful therapeutic agents for psychiatric and neurodegenerative diseases such as schizophrenia and Huntington's disease. We measured striatal PDE10A occupancy by TAK-063, a newly developed compound with high affinity and selectivity for PDE10A, using PET with [(11)C]T-773 in nonhuman primates. Two 123-min dynamic PET measurements were performed on three female rhesus monkeys, once at baseline and again after intravenous administration of different doses of TAK-063 (0.2-1.6 mg/kg). Total distribution volume (V(T)) was calculated with a two-tissue compartment model using metabolite-corrected plasma input. Although the in vitro autoradiography did not show high specific binding to [(11)C]T-773 in the cerebellum, V(T) in the cerebellum decreased after TAK-063 treatment. The specific binding to PDE10A (V(S)) was calculated as the difference of the V(T) between the target regions and the cerebellum. PDE10A occupancy was calculated as the percent change of V(S). The average PDE10A occupancy of the caudate nucleus and putamen was 35.2% at 0.2 mg/kg and 83.2% at 1.6 mg/kg. In conclusion, this nonhuman primate PET study demonstrated that [(11)C]T-773 is useful to estimate the PDE10A occupancy by TAK-063 in the striatum although there is in vivo interaction of the uptake between [(11)C]T-773 and TAK-063 in the cerebellum. These results warrant further clinical occupancy study for TAK-063.

Discovery and Preclinical Validation of [(11)C]AZ13153556, a Novel Probe for the Histamine Type 3 Receptor.

UNLABELLED: The histamine type 3 receptor (H3) is a G protein-coupled receptor implicated in several disorders of the central nervous system. Herein, we describe the radiolabeling and preclinical evaluation of a candidate radioligand for the H3 receptor, 4-(1S,2S)-2-(4-cyclobutylpiperazine-1-carbonyl)cyclopropyl]-N-methyl-benzamide (5), and its comparison with one of the frontrunner radioligands for H3 imaging, namely, GSK189254 (1). Compounds 1 and 5 were radiolabeled with tritium and carbon-11 for in vitro and in vivo imaging experiments. The in vitro binding of [(3)H]1 and [(3)H]5 was examined by (i) saturation binding to rat and nonhuman primate brain tissue homogenate and (ii) in vitro autoradiography on tissue sections from rat, guinea pig, and human brain. The in vivo binding of [(11)C]1 and [(11)C]5 was examined by PET imaging in mice and nonhuman primates. Bmax values obtained from Scatchard analysis of [(3)H]1 and [(3)H]5 binding were in good agreement. Autoradiography with [(3)H]5 on rat, guinea pig, and human brain slices showed specific binding in regions known to be enhanced in H3 receptors, a high degree of colocalization with [(3)H]1, and virtually negligible nonspecific binding in tissue. PET measurements in mice and nonhuman primates demonstrated that [(11)C]5 binds specifically and reversibly to H3 receptors in vivo with low nonspecific binding in brain tissue. Whereas [(11)C]1 showed similar binding characteristics in vivo, the binding kinetics appeared faster for [(11)C]5 than for [(11)C]1. CONCLUSIONS: [(11)C]5 has suitable properties for quantification of H3 receptors in nonhuman primate brain and has the potential to offer improved binding kinetics in man compared to [(11)C]1.

Development of a series of novel carbon-11 labeled PDE10A inhibitors.

Phosphodiesterase 10A (PDE10A) is a member of the PDE family of enzymes that degrades cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP). Our aim was to label a series of structurally related PDE10A inhibitors with carbon-11 and evaluate them as potential positron emission tomography (PET) radioligands for PDE10A using nonhuman primates. The series consisted of seven compounds based on the 3-(1H-pyrazol-5-yl)pyridazin-4(1H)-one backbone. These compounds were selected from the initial larger library based on a number of parameters such as affinity, selectivity for hPDE10A in in vitro tests, lipophilicity, and on the results of multidrug resistance protein 1 (MDR1)-LLCPK1 and the parallel artificial membrane permeability assays. Seven radioligands (KIT-1, 3, 5, 6, 7, 9, and 12) were radiolabeled with carbon-11 employing O-methylation on the hydroxyl moiety using [(11)C]methyl triflate. In vivo examination of each radioligand was performed using PET in rhesus monkeys; analysis of radiometabolites in plasma also was conducted using HPLC. All seven radioligands were labeled with high (>90%) incorporation of [(11)C]methyl triflate into their appropriate precursors and with high specific radioactivity. Carbon-11 labeled KIT-5 and KIT-6 showed high accumulation in the striatum, consistent with the known anatomical distribution of PDE10A in brain, accompanied by fast washout and high specific binding ratio. In particular [(11)C]KIT-6, named [(11)C]T-773, is a promising PET tool for further examination of PDE10A in human brain.

Evaluation of a novel PDE10A PET radioligand, [(11) C]T-773, in nonhuman primates: brain and whole body PET and brain autoradiography.

Phosphodiesterase 10A (PDE10A) is considered to be a key target for the treatment of several neuropsychiatric diseases. The characteristics of [(11) C]T-773, a novel positron emission tomography (PET) radioligand with high binding affinity and selectivity for PDE10A, were evaluated in autoradiography and in nonhuman primate (NHP) PET. Brain PET measurements were performed under baseline conditions and after administration of a selective PDE10A inhibitor, MP-10. Total distribution volume (VT ) and binding potential (BPND ) were calculated using various kinetic models. Whole body PET measurements were performed to calculate the effective dose of [(11) C]T-773. Autoradiography studies in postmortem human and monkey brain sections showed high accumulation of [(11) C]T-773 in the striatum and substantia nigra which was blocked by MP-10. Brain PET showed high accumulation of [(11) C]T-773 in the striatum, and the data could be fitted using a two tissue compartment model. BPND was approximately 1.8 in the putamen when the cerebellum was used as the reference region. Approximately 70% of PDE10A binding was occupied by 1.8 mg/kg of MP-10. Whole body PET showed high accumulation of [(11) C]T-773 in the liver, kidney, heart, and brain in the initial phase. The radioligand was partly excreted via bile and the gastrointestinal tract, and partly excreted through the urinary tract. The calculated effective dose was 0.007 mSv/MBq. In conclusion, [(11) C]T-773 was demonstrated to be a promising PET radioligand for PDE10A with favorable brain kinetics. Dosimetry results support multiple PET measurements per person in human studies. Further research is required with [(11) C]T-773 in order to test the radioligand's potential clinical applications.

Large Variation in Brain Exposure of Reference CNS Drugs: a PET Study in Nonhuman Primates.

BACKGROUND: Positron emission tomography microdosing of radiolabeled drugs allows for noninvasive studies of organ exposure in vivo. The aim of the present study was to examine and compare the brain exposure of 12 commercially available CNS drugs and one non-CNS drug. METHODS: The drugs were radiolabeled with (11)C (t 1/2 = 20.4 minutes) and examined using a high resolution research tomograph. In cynomolgus monkeys, each drug was examined twice. In rhesus monkeys, a first positron emission tomography microdosing measurement was repeated after preadministration with unlabeled drug to examine potential dose-dependent effects on brain exposure. Partition coefficients between brain and plasma (KP) were calculated by dividing the AUC0-90 min for brain with that for plasma or by a compartmental analysis (VT). Unbound KP (KP u,u) was obtained by correction for the free fraction in brain and plasma. RESULTS: After intravenous injection, the maximum radioactivity concentration (C max, %ID) in brain ranged from 0.01% to 6.2%. For 10 of the 12 CNS drugs, C max, %ID was >2%, indicating a preferential distribution to brain. A lower C max, %ID was observed for morphine, sulpiride, and verapamil. K P ranged from 0.002 (sulpiride) to 68 (sertraline) and 7 of 13 drugs had KP u,u close to unity. For morphine, sulpiride, and verapamil, K P u,u was <0.3, indicating impaired diffusion and/or active efflux. Brain exposure at microdosing agreed with pharmacological dosing conditions for the investigated drugs. CONCLUSIONS: This study represents the largest positron emission tomography study on brain exposure of commercially available CNS drugs in nonhuman primates and may guide interpretation of positron emission tomography microdosing data for novel drug candidates.

Quantitative Analysis of ¹8F-(E)-N-(3-Iodoprop-2-Enyl)-2ß-Carbofluoroethoxy-3ß-(4'-Methyl-Phenyl) Nortropane Binding to the Dopamine Transporter in Parkinson Disease.

UNLABELLED: (18)F-(E)-N-(3-iodoprop-2-enyl)-2ß-carbofluoroethoxy-3ß-(4'-methyl-phenyl) nortropane ((18)F-FE-PE2I) is a recently developed radioligand for the in vivo quantification of the dopamine transporter (DAT) in the striatum and substantia nigra (SN). The aim of this study was to examine the suitability of (18)F-FE-PE2I as a tool for imaging the nigrostriatal pathway in Parkinson disease (PD) with PET. METHODS: Ten PD patients (9 men and 1 woman; mean age ± SD, 60 ± 9 y; Hoehn and Yahr, 1-2; Unified Parkinson Disease Rating Scale motor, 18.9 ± 6.7) and 10 controls (9 men and 1 woman; mean age ± SD, 60 ± 7 y) were included. PET measurements with (18)F-FE-PE2I were conducted for 93 min using the High-Resolution Research Tomograph. Venous blood was drawn to compare protein binding, parent fraction, and radiometabolite composition in PD patients and controls. Regions of interest for the caudate, putamen, ventral striatum, SN, and cerebellum were drawn on coregistered MR images. The outcome measure was the binding potential (BP(ND)) estimated with the simplified reference tissue model and the Logan graphical analysis, using the cerebellum as a reference region. Time stability of BP(ND) was examined to define the shortest acquisition protocol for quantitative studies. The wavelet-aided parametric imaging method was used to obtain high-resolution BP(ND) images to compare DAT availability in the striatum and SN in PD patients and control subjects. Group differences were assessed with the unpaired t test (P < 0.05). RESULTS: Parent, radiometabolite fractions, plasma concentration, and cerebellar uptake of (18)F-FE-PE2I did not differ significantly between PD patients and controls. Stable estimates of BP(ND) (<8% of the 93-min value) were obtained with the simplified reference tissue model using approximately 66 min of data. BP(ND) values in PD patients were significantly lower than those in controls (P < 0.05) in the caudate (2.54 ± 0.79 vs. 3.68 ± 0.56), putamen (1.39 ± 1.04 vs. 4.41 ± 0.54), ventral striatum (2.26 ± 0.93 vs. 3.30 ± 0.46), and SN (0.46 ± 0.20 vs. 0.68 ± 0.15). CONCLUSION: (18)F-FE-PE2I is clearly a suitable radioligand for DAT quantification and imaging of the nigrostriatal pathway in PD. Similar metabolism in controls and PD patients, suitability of the cerebellum as a reference region, and accuracy of quantification using approximately 66 min of PET data are advantages for noninvasive and simplified imaging protocols for PD studies. Finally, DAT loss in PD can be measured in both the striatum and the SN, supporting the utility of (18)F-FE-PE2I as an imaging tool of the nigrostriatal pathway.

Molecular Imaging of PDE10A Knockout Mice with a Novel PET Radiotracer: [(11)C]T-773.

PURPOSE: [(11)C]T-773 is a new radioligand for positron emission tomography (PET) targeting the phosphodiesterase 10A enzyme (PDE10A). PDE10A is highly expressed in the striatum by medium spiny neurons, and it has been demonstrated to be involved in the regulation of striatal signaling through the reduction of medium spiny neuronal sensitivity towards glutamatergic excitation. PDE10A is associated with Parkinson's disease and different neuropsychiatric disorders such as Huntington's disease, obsessive-compulsive disorders (OCD) and schizophrenia. Studies have indicated that the inhibition of PDE10A may represent a novel therapeutic approach to the treatment of the aforementioned diseases characterized by the reduced activity of medium spiny neurons. An appropriate PET radioligand for PDE10A would help to facilitate drug development and drug evaluation. PROCEDURES: We have evaluated the [(11)C]T-773 ligand in PDE10A knockout mice (heterozygous [HET] and homozygous [HOM]) as well as in normal control animals (WILD) with PET. RESULTS: The regional percent standardized uptake values (%SUV; mean ± SD) in the striatum were 48.2 ± 1.0 (HOM), 63.6 ± 5.3 (HET) and 85.1 ± 6.3 (WILD). Between each animal group the striatal %SUV values were significantly different (p < 0.0001). The striatal BPND values (mean ± SD) were 0.0 ± 0.0 (HOM), 0.14 ± 0.07 (HET) and 0.56 ± 0.15 (WILD). The BPND values were significantly lower in homozygous and heterozygous animals compared to wild type (p < 0.0001). CONCLUSIONS: The novel PDE10A radioligand [(11)C]T-773 shows increased signals with higher levels of PDE10A and acceptable binding in the striatum in control animals compared to knockout mice.

Positron emission tomography imaging of the 18-kDa translocator protein (TSPO) with [18F]FEMPA in Alzheimer's disease patients and control subjects.

PURPOSE: Imaging of the 18-kDa translocator protein (TSPO) is a potential tool for examining microglial activation and neuroinflammation in early Alzheimer's disease (AD). [(18)F]FEMPA is a novel high-affinity second-generation TSPO radioligand that has displayed suitable pharmacokinetic properties in preclinical studies. The aims of this study were to quantify the binding of [(18)F]FEMPA to TSPO in AD patients and controls and to investigate whether higher [(18)F]FEMPA binding in AD patients than in controls could be detected in vivo. METHODS: Ten AD patients (five men, five women; age 66.9 ± 7.3 years; MMSE score 25.5 ± 2.5) and seven controls (three men, four women; age 63.7 ± 7.2 years, MMSE score 29.3 ± 1.0) were studied using [(18)F]FEMPA at Turku (13 subjects) and at Karolinska Institutet (4 subjects). The in vitro binding affinity for TSPO was assessed using PBR28 in a competition assay with [(3)H]PK11195 in seven controls and eight AD patients. Cortical and subcortical regions of interest were examined. Quantification was performed using a two-tissue compartment model (2TCM) and Logan graphical analysis (GA). The outcome measure was the total distribution volume (V T). Repeated measures analysis of variance was used to assess the effect of group and TSPO binding status on V T. RESULTS: Five AD patients and four controls were high-affinity binders (HABs). Three AD patients and three controls were mixed-affinity binders. V T estimated with Logan GA was significantly correlated with V T estimated with the 2TCM in both controls (r = 0.97) and AD patients (r = 0.98) and was selected for the final analysis. Significantly higher V T was found in the medial temporal cortex in AD patients than in controls (p = 0.044) if the TSPO binding status was entered as a covariate. If only HABs were included, significantly higher V T was found in the medial and lateral temporal cortex, posterior cingulate, caudate, putamen, thalamus and cerebellum in AD patients than in controls (p < 0.05). CONCLUSION: [(18)F]FEMPA seems to be a suitable radioligand for detecting increased TSPO binding in AD patients if their binding status is taken into account.

Amphetamine decreases α2C-adrenoceptor binding of [11C]ORM-13070: a PET study in the primate brain.

BACKGROUND: The neurotransmitter norepinephrine has been implicated in psychiatric and neurodegenerative disorders. Examination of synaptic norepinephrine concentrations in the living brain may be possible with positron emission tomography (PET), but has been hampered by the lack of suitable radioligands. METHODS: We explored the use of the novel α2C-adrenoceptor antagonist PET tracer [(11)C]ORM-13070 for measurement of amphetamine-induced changes in synaptic norepinephrine. The effect of amphetamine on [(11)C]ORM-13070 binding was evaluated ex vivo in rat brain sections and in vivo with PET imaging in monkeys. RESULTS: Microdialysis experiments confirmed amphetamine-induced elevations in rat striatal norepinephrine and dopamine concentrations. Regional [(11)C]ORM-13070 receptor binding was high in the striatum and low in the cerebellum. After injection of [(11)C]ORM-13070 in rats, mean striatal specific binding ratios, determined using cerebellum as a reference region, were 1.4±0.3 after vehicle pretreatment and 1.2±0.2 after amphetamine administration (0.3mg/kg, subcutaneous). Injection of [(11)C]ORM-13070 in non-human primates resulted in mean striatal binding potential (BP ND) estimates of 0.65±0.12 at baseline. Intravenous administration of amphetamine (0.5 and 1.0mg/kg, i.v.) reduced BP ND values by 31-50%. Amphetamine (0.3mg/kg, subcutaneous) increased extracellular norepinephrine (by 400%) and dopamine (by 270%) in rat striata. CONCLUSIONS: Together, these results indicate that [(11)C]ORM-13070 may be a useful tool for evaluation of synaptic norepinephrine concentrations in vivo. Future studies are required to further understand a potential contribution of dopamine to the amphetamine-induced effect.

Determination of plasma protein binding of positron emission tomography radioligands by high-performance frontal analysis.

Positron emission tomography (PET) is an imaging technique based on the use of radioligands labeled with short lived radionuclides, such as (11)C (t½=20.4min) and (18)F (t½=109.8min), which as a consequence often requires rapid plasma protein binding analysis methods. In addition, PET radioligands can suffer from non-specific binding to the membrane when ultrafiltraion, which is the most commonly used method for measuring protein binding in PET, is employed. In this study a high-performance frontal analysis (HPFA) method based on incorporation of a gel filtration column (discovery(®) BIO GFC 100, 50mm×4.6mm, 5µm, 100Å) into a radio-LC system with phosphate buffered saline (PBS, pH 7.4) at a flow rate of 3ml/min as mobile phase was developed and investigated for four PET radioligands. The minimum injection volume (MIV) of plasma, which is a crucial factor in HPFA, was determined to be 200µl (human), 500µl (monkey), 700µl (human) and 1000µl (monkey) for these four radioligands. The MIV values increased as a higher fraction of the radioligand was present in the protein-free form. The protein binding results obtained were in good agreement with ultrafiltration and the method did not suffer from non-specific binding. The short analysis time (<12min) allowed multiple protein binding measurements during time course of a human [(11)C]PBR28 PET study.

(18)F-MCL-524, an (18)F-Labeled Dopamine D2 and D3 Receptor Agonist Sensitive to Dopamine: A Preliminary PET Study.

UNLABELLED: PET has been used to examine changes in neurotransmitter concentrations in the living brain. Pioneering PET studies on the dopamine system have used D2 and D3 receptor (D2/D3) antagonists such as (11)C-raclopride. However, more recently developed agonist radioligands have shown enhanced sensitivity to endogenous dopamine. A limitation of available agonist radioligands is that they incorporate the short-lived radioisotope (11)C. In the current study, we developed the (18)F-labeled D2/D3 receptor agonist (R)-(-)-2-(18)F-fluoroethoxy-N-n-propylnorapomorphine ((18)F-MCL-524). METHODS: In total, 10 PET measurements were conducted on 5 cynomolgus monkeys. Initially, the binding of (18)F-MCL-524 was compared with that of (11)C-MNPA in 3 monkeys. Second, the specificity of (18)F-MCL-524 binding was examined in pretreatment studies using raclopride (1.0 mg/kg) and d-amphetamine (1.0 mg/kg). Third, a preliminary kinetic analysis was performed using the radiometabolite-corrected arterial input function of the baseline studies. Finally, 2 whole-body PET measurements were conducted to evaluate biodistribution and radiation dosimetry after intravenous injection of (18)F-MCL-524. RESULTS: (18)F-MCL-524 entered the brain and provided striatum-to-cerebellum ratios suitable for reliable quantification of receptor binding using the multilinear reference tissue model. Mean striatal nondisplaceable binding potential (BPND) values were 2.0 after injection of (18)F-MCL-524 and 1.4 after (11)C-MNPA. The ratio of the BPND values of (18)F-MCL-524 and (11)C-MNPA was 1.5 across striatal subregions. After administration of raclopride and d-amphetamine, the (18)F-MCL-524 BPND values were reduced by 89% and 56%, respectively. Preliminary kinetic analysis demonstrated that BPND values obtained with the 1-tissue- and 2-tissue-compartment models were similar to values obtained with the multilinear reference tissue model. Estimated radiation doses were highest for gallbladder (0.27 mSv/MBq), upper large intestine (0.19 mSv/MBq), and small intestine (0.17 mSv/MBq). The estimated effective dose was 0.035 mSv/MBq. CONCLUSION: The (18)F-labeled agonist (18)F-MCL-524 appears suitable for quantification of D2/D3 receptor binding in vivo, and the results encourage extension to human studies. The longer half-life of (18)F makes (18)F-MCL-524 attractive for studies on modulation of the dopamine concentration-for example, in combination with simultaneous measurement of changes in blood-oxygen-level-dependent signal using bimodal PET/functional MRI.

Positron emission tomography imaging of 5-hydroxytryptamine1B receptors in Parkinson's disease.

Impairment of the central serotonin system in Parkinson's disease (PD) has been shown postmortem and in vivo with positron emission tomography (PET). The aim of this PET study was to examine and compare the availability of the 5-hydroxytryptamine (5-HT)(1B)-receptor subtype in patients with PD and age-matched control subjects. Twelve control subjects and 12 PD patients were examined with PET using the 5-HT(1B)-radioligand [(11)C]AZ10419369. In PD patients, 5-HT(1B)-receptor availability in the right orbitofrontal cortex was lower than in control subjects. A statistically significant negative correlation between 5-HT(1B)-receptor availability and age was obtained for the right temporal cortex in control subjects and for the right midbrain and left parahippocampal gyrus in PD patients. The lower regional 5-HT(1B)-receptor availability is in line with previous studies showing a decrease of serotonin imaging markers in PD and corroborates a role of the serotonin system in the pathophysiology of PD. The demonstrated age effect on 5-HT(1B) receptors suggest a physiologic and PD-related decline of serotonin function, indicating the importance of controlling for age in clinical studies.

Synthesis, radiolabeling, and in vivo pharmacokinetic evaluation of the amyloid beta radioligand [11C]AZD4694 in nonhuman primates.

PURPOSE: [(18)F]AZD4694 (2-(2-(18)F-fluoro-6-(methylamino)-3-pyridyl)benzofuran-5-ol) is a radioligand suitable for imaging of amyloid beta deposits in the living human brain using positron emission tomography (PET). Here, we report the preparation and pharmacokinetic profile of its carbon-11 (t1/2 = 20.4 min) labeled isotopolog [(11)C]AZD4694 and compare [(11)C]AZD4694 with the hitherto most widely applied amyloid PET radioligand [(11)C]Pittsburgh Compound B (PiB). PROCEDURES: The immediate unlabeled precursor to [(11)C]AZD4694 was prepared in a four-step convergent synthesis. Subsequent N-(11)C-methylation of this precursor with [(11)C]methyl iodide yielded [(11)C]AZD4694, which after isolation and formulation was injected into cynomolgus monkeys. The radioactivity in nonhuman primate brain following injection of [(11)C]AZD4694 and [(11)C]PiB was measured using PET. RESULTS: [(11)C]AZD4694 was prepared in a 60 % incorporation yield. In a head to head comparison with [(11)C]PiB, it appeared that [(11)C]AZD4694 displayed slightly lower nonspecific binding in white matter than [(11)C]PiB as well as more rapid pharmacokinetics in the brain. CONCLUSIONS: The advantageous pharmacokinetic profile and low nonspecific binding render [(11)C]AZD4694 a promising PET radioligand for imaging of amyloid beta in the human brain with PET.

Characterization of [(11)C]Cimbi-36 as an agonist PET radioligand for the 5-HT(2A) and 5-HT(2C) receptors in the nonhuman primate brain.

Only recently the first successful serotonin 2A (5-HT2A) receptor agonist PET radioligands have been described, with [(11)C]Cimbi-36 reported as the most promising in the pig brain so far. Agonist radioligands may target specifically the G protein-coupled state of the receptors and thereby provide a more meaningful assessment of available receptors than antagonist radioligands. In the current study we characterized [(11)C]Cimbi-36 receptor binding in the primate brain. On five experimental days, a total of 14 PET measurements were conducted in three female rhesus monkeys. On each day, PET measurements were conducted after intravenous injection of [(11)C]Cimbi-36 during baseline conditions and after intravenous infusion of the 5-HT2 receptor antagonist ketanserin (n=3) or the 5-HT2C receptor antagonist SB 242084 (n=2). On four of the experimental days an additional baseline PET measurement was conducted after injection of [(11)C]MDL 100907. All PET measurements were performed for 2h in a HRRT PET system and arterial blood was obtained for measurement of the [(11)C]Cimbi-36 input function. Quantification of [(11)C]Cimbi-36 receptor binding was performed using kinetic and graphical analysis. After injection of [(11)C]Cimbi-36 the regional distribution of radioactivity in brain was in accordance with the known 5-HT2 receptor distribution. The two-tissue compartment model was superior for the description of the time-radioactivity curves of all examined brain regions. BPND values obtained with reference tissue models correlated with corresponding values obtained with kinetic modeling. Administration of ketanserin decreased the binding in all brain regions but did not affect the cerebellar distribution volume. The BPND of [(11)C]Cimbi-36 was 56±8% of [(11)C]MDL 100907 across cortical regions, but higher in other brain regions including choroid plexus. After administration of SB 242084, [(11)C]Cimbi-36 binding was nearly completely inhibited in the choroid plexus, partly reduced in several subcortical regions (e.g. hippocampus), but not affected in the cortical regions. In conclusion, the receptor binding of [(11)C]Cimbi-36 can be quantified using kinetic modeling and the cerebellum was found to be a suitable reference region. The difference between [(11)C]Cimbi-36 and [(11)C]MDL 100907 binding in the choroid plexus is related to 5-HT2C receptor binding of [(11)C]Cimbi-36. [(11)C]Cimbi-36 is the first agonist radioligand suitable for examination of 5-HT2A receptors in the cortical regions and of 5-HT2C receptors in the choroid plexus of the primate brain.

Simultaneous determination of protein-free and total positron emission tomography radioligand concentrations in plasma using high-performance frontal analysis followed by mixed micellar liquid chromatography: application to [11C]PBR28 in human plasma.

A two-dimensional liquid chromatographic (LC) system was developed for the determination of protein-free and total (free + bound forms) positron emission tomography (PET) radioligand concentrations in plasma by direct plasma injection. The unbound PET radioligand was first analyzed by high-performance frontal analysis using a short gel-filtration column and phosphate buffered saline solution as the mobile phase. All the collected effluent from the gel-filtration column was then transferred to the second dimension consisting of a monolithic C-18 column and mixed (anionic and nonionic surfactants) micellar eluent for determination of the total PET radioligand concentration. The simultaneous analysis of protein binding and radiometabolism of [(11)C]PBR28 was completed within 11 min without any pretreatment of plasma and employing a single analytical system. This system allowed highly sensitive analysis of total [(11)C]PBR28 with a limit of detection (LOD) of 1.6 becquerel (Bq). The LOD for the determination of unbound [(11)C]PBR28 was 21 Bq. Finally, simultaneous measurements of protein binding and radiometabolism of [(11)C]PBR28 in human plasma were achieved for up to 50 min after radioligand administration.