In vivo evaluation of a microtubule PET ligand, [11C]MPC-6827, in mice following chronic alcohol consumption.

BACKGROUND: Excessive alcohol consumption is a global health burden and requires a better understanding of its neurobiology. A lower density of brain microtubules is found in alcohol-related human brain disease postmortem and in rodent models of chronic alcohol consumption. Here, we report in vivo imaging studies of microtubules in brain using our recently reported Positron Emission Tomography (PET) tracer, [11C]MPC-6827, in chronic alcohol-consuming adult male C57BL/6 J mice and control mice. METHODS: In vivo PET imaging studies of [11C]MPC-6827 (3.7 ± 0.8 MBq) were performed in two groups of adult male mice: (1) water-consuming control mice (n = 4) and (2) mice that consumed 20% alcohol (w/v) for 4 months using the intermittent 2-bottle choice procedure that has been shown to lead to signs of alcohol dependence. Dynamic 63 min PET images were acquired using a microPET Inveon system (Siemens, Germany). PET images were reconstructed using the 3D-OSEM algorithm and analyzed using VivoQuant version 4 (Invicro, MA). Tracer uptake in ROIs that included whole brain, prefrontal cortex (PFC), liver and heart was measured and plotted as %ID/g over time (0-63 min) to generate time-activity curves (TACs). RESULTS: In general, a trend for lower binding of [11C]MPC-6827 in the whole brain and PFC of mice in the chronic alcohol group was found compared with control group. No group difference in radiotracer binding was found in the peripheral organs such as liver and heart. CONCLUSIONS: This pilot study indicates a trend of loss of microtubule binding in whole brain and prefrontal cortex of chronic alcohol administered mice brain compared to control mice, but no loss in heart or liver. These results indicate the potential of [11C]MPC-6827 as a PET ligand for further in vivo imaging investigations of AUD in human.

Progress in PET Imaging of Neuroinflammation Targeting COX-2 Enzyme.

Neuroinflammation and cyclooxygenase-2 (COX-2) upregulation are associated with the pathogenesis of degenerative brain diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), epilepsy, and a response to traumatic brain injury (TBI) or stroke. COX-2 is also induced in acute pain, depression, schizophrenia, various cancers, arthritis and in acute allograft rejection. Positron emission tomography (PET) imaging allows for the direct measurement of in vivo COX-2 upregulation and thereby enables disease staging, therapy evaluation and aid quantifying target occupancy of novel nonsteroidal anti-inflammatory drugs or NSAIDs. Thus far, no clinically useful radioligand is established for monitoring COX-2 induction in brain diseases due to the delay in identifying qualified COX-2-selective inhibitors entering the brain. This review examines radiolabeled COX-2 inhibitors reported in the past decade and identifies the most promising radioligands for development as clinically useful PET radioligands. Among the radioligands reported so far, the three tracers that show potential for clinical translation are, [11CTMI], [11C]MC1 and [18F]MTP. These radioligands demonstrated BBB permeablity and in vivo binding to constitutive COX-2 in the brain or induced COX-2 during neuroinflammation.

Radiosynthesis and evaluation of [18F]FMTP, a COX-2 PET ligand.

BACKGROUND: The upregulation of cyclooxygenase-2 (COX-2) is involved in neuroinflammation associated with many neurological diseases as well as cancers of the brain. Outside the brain, inflammation and COX-2 induction contribute to the pathogenesis of pain, arthritis, acute allograft rejection, and in response to infections, tumors, autoimmune disorders, and injuries. Herein, we report the radiochemical synthesis and evaluation of [18F]6-fluoro-2-(4-(methylsulfonyl)phenyl)-N-(thiophen-2-ylmethyl)pyrimidin-4-amine ([18F]FMTP), a high-affinity COX-2 inhibitor, by cell uptake and PET imaging studies. METHODS: The radiochemical synthesis of [18F]FMTP was optimized using chlorine to fluorine displacement method, by reacting [18F]fluoride/K222/K2CO3 with the precursor molecule. Cellular uptake studies of [18F]FMTP was performed in COX-2 positive BxPC3 and COX-2 negative PANC-1 cell lines with unlabeled FMTP as well as celecoxib to define specific binding agents. Dynamic microPET image acquisitionwas performed in anesthetized nude mice (n = 3), lipopolysaccharide (LPS) induced neuroinflammation mice (n = 4), and phosphate-buffered saline (PBS) administered control mice (n = 4) using a Trifoil microPET/CT for a scan period of 60 min. RESULTS: A twofold higher binding of [18F]FMTP was found in COX-2 positive BxPC3 cells compared with COX-2 negative PANC-1 cells. The radioligand did not show specific binding to COX-2 negative PANC-1 cells. MicroPET imaging in wild-type mice indicated blood-brain barrier (BBB) penetration and fast washout of [18F]FMTP in the brain, likely due to the low constitutive COX-2 expression in the normal brain. In contrast, a ~ twofold higher uptake of the radioligand was found in LPS-induced mice brain than PBS treated control mice. CONCLUSIONS: Specific binding to COX-2 in BxPC3 cell lines, BBB permeability, and increased brain uptake in neuroinflammation mice qualifies [18F]FMTP as a potential PET tracer for studying inflammation.

Synthesis, in vitro and in vivo evaluation of 11C-O-methylated arylpiperazines as potential serotonin 1A (5-HT1A) receptor antagonist radiotracers.

BACKGROUND: Serotonin 1A (5-HT1A) receptors are implicated in the pathogenesis of several psychiatric and neurodegenerative disorders motivating the development of suitable radiotracers for in vivo positron emission tomography (PET) neuroimaging. The gold standard PET imaging agent for this target is [carbonyl-11C]WAY-100635, labeled via a technically challenging multi-step reaction that has limited its widespread use. While several antagonist and agonist-based PET radiotracers for 5-HT 1A receptors have been developed, their clinical translation has been hindered by methodological challenges and/or and non-specific binding. As a result, there is continued interest in the development of new and more selective 5-HT1A PET tracers having a relatively easier and reliable radiosynthesis process for routine production and with favorable metabolism to facilitate tracer-kinetic modeling. The purpose of the current study was to develop and characterize a radioligand with suitable characteristics for imaging 5-HT1A receptors in the brain. The current study reports the in vitro characterization and radiosyntheses of three candidate 5-HT1A receptor antagonists, DF-100 (1), DF-300 (2) and DF-400 (3), to explore their suitability as potential PET radiotracers. RESULTS: Syntheses of 1-3 and corresponding precursors for radiolabeling were achieved from isonicotinic, picolinic acid or picolino nitrile. In vitro binding studies demonstrated nanomolar affinity of the compounds for 5-HT1A receptors. Binding of 1-3 for other biogenic amines, neurotransmitter receptors, and transporters was negligible with the exception of moderate affinities for α1-adrenergic receptors (4-6-fold less potent than that for 5-HT1A receptor). Radioligands [11C]1-3 were efficiently prepared by 11C-O-methylation of the corresponding phenolic precursor in non-decay corrected radiochemical yields of 7-11% with > 99% chemical and radiochemical purities. Dynamic PET studies in rats demonstrated negligible brain uptake of [11C]1 and [11C]2. In contrast, significant brain uptake of [11C]3 was observed with an early peak SUV of 4-5. However, [11C]3 displayed significant off-target binding attributed to α1-adrenergic receptors based on regional distribution (thalamus>hippocampus) and blocking studies. CONCLUSION: Despite efficient radiolabeling, results from PET imaging experiments limit the application of [11C]3 for in vivo quantification of 5-HT1A receptors. Nevertheless, derivatives of compound 3 may provide a scaffold for alternative PET radiotracers with improved selectivity for 5-HT 1A receptors or α1-adrenergic receptors.

In vivo comparison of N-11CH3 vs O-11CH3 radiolabeled microtubule targeted PET ligands.

Altered dynamics of microtubules (MT) are implicated in the pathophysiology of a number of brain diseases. Therefore, radiolabeled MT targeted ligands that can penetrate the blood brain barrier (BBB) may offer a direct and sensitive approach for diagnosis, and assessing the clinical potential of MT targeted therapeutics using PET imaging. We recently reported two BBB penetrating radioligands, [11C]MPC-6827 and [11C]HD-800 as specific PET ligands for imaging MTs in brain. The major metabolic pathway of the above molecules is anticipated to be via the initial labeling site, O-methyl, compared to the N-methyl group. Herein, we report the radiosynthesis of N-11CH3-MPC-6827 and N-11CH3-HD-800 and a comparison of their in vivo binding with the corresponding O-11CH3 analogues using microPET imaging and biodistribution methods. Both O-11CH3 and N-11CH3 labeled MT tracers exhibit high specific binding and brain. The N-11CH3 labeled PET ligands demonstrated similar in vivo binding characteristics compared with the corresponding O-11CH3 labeled tracers, [11C]MPC-6827 and [11C]HD-800 respectively.

In vivo PET Imaging of [11C]CIMBI-5, a 5-HT2AR Agonist Radiotracer in Nonhuman Primates.

PURPOSE: 5-HT2AR exists in high and low affinity states. Agonist PET tracers measure binding to the active high affinity site and thus provide a functionally relevant measure of the receptor. Limited in vivo data have been reported so far for a comparison of agonist versus antagonist tracers for 5-HT2AR used as a proof of principle for measurement of high and low affinity states of this receptor. We compared the in vivo binding of [11C]CIMBI-5, a 5-HT2AR agonist, and of the antagonist [11C]M100907, in monkeys and baboons. METHODS: [11C]CIMBI-5 and [11C]M100907 baseline PET scans were performed in anesthetized male baboons (n=2) and male vervet monkeys (n=2) with an ECAT EXACT HR+ and GE 64-slice PET/CT Discovery VCT scanners. Blocking studies were performed in vervet monkeys by pretreatment with MDL100907 (0.5 mg/kg, i.v.) 60 minutes prior to the scan. Regional distribution volumes and binding potentials were calculated for each ROI using the likelihood estimation in graphical analysis and Logan plot, with either plasma input function or reference region as input, and simplified reference tissue model approaches. RESULTS: PET imaging of [11C]CIMBI-5 in baboons and monkeys showed the highest binding in 5-HT2AR-rich cortical regions, while the lowest binding was observed in cerebellum, consistent with the expected distribution of 5-HT2AR. Very low free fractions and rapid metabolism were observed for [11C]CIMBI-5 in baboon plasma. Binding potential values for [11C]CIMBI-5 were 25-33% lower than those for [11C]MDL100907 in the considered brain regions. CONCLUSION: The lower binding potential of [11C]CIMBI-5 in comparison to [11C]MDL100907 is likely due to the preferential binding of the former to the high affinity site in vivo in contrast to the antagonist,  [11C]MDL100907, which binds to both high and low affinity sites.

Radiosynthesis and evaluation of [11C]CMP, a high affinity GSK3 ligand.

Dysfunction of GSK3 is implicated in the etiology of many brain, inflammatory, cardiac diseases, and cancer. PET imaging would enable in vivo detection and quantification of GSK3 and can impact the choice of therapy, allow non-invasive monitoring of disease progression and treatment effects. In this report, the synthesis and evaluation of a high affinity GSK3 ligand, [11C]2-(cyclopropanecarboxamido)-N-(4-methoxypyridin-3-yl)isonicotinamide, ([11C]CMP, (3), (IC50 = 3.4 nM, LogP = 1.1) is described. [11C]CMP was synthesized in 25 ±â€¯5% yield by radiomethylating the corresponding phenolate using [11C]CH3I. The radioligand exhibited modest uptake in U251 human glioblastoma cell lines with ∼50% specific binding. MicroPET studies in rats indicated negligible blood-brain barrier (BBB) penetration of [11C]CMP, despite its high affinity and suitable logP value for BBB penetration. However, administration of cyclosporine prior to [11C]CMP injection showed significant improvement in brain radioactivity uptake and the tracer binding. This finding indicates that [11C]CMP might be a P-gp efflux substrate and therefore has some limitations for routine in vivo PET evaluations in brain.

Molecular docking, dynamics simulations and 3D-QSAR modeling of arylpiperazine derivatives of 3,5-dioxo-(2H,4H)-1,2,4-triazine as 5-HT1AR agonists.

Serotonin receptor, 5-HT1AR, agonists and partial agonists have established drug candidates for psychiatric and neurologic disorders. Recently, we reported the synthesis and evaluation of arylpiperazine derivatives of 3,5-dioxo-(2H,4H)-1,2,4-triazine as 5-HT1AR ligands. Herein, we generated a homology model of the receptor and docked the ligands against it, predicted the stability of the receptor model and complexes by molecular dynamics and generated a 3D-QSAR model for the arylpiperazine derivatives of 3,5-dioxo-(2H,4H)-1,2,4-triazine. The model suggests the hydrophobic part that arises from the aromatic region and the electron withdrawing parts play a vital role in the agonist activity of the lead molecules.

In vivo evaluation of [11C]TMI, a COX-2 selective PET tracer, in baboons.

Overexpression of Cyclooxygenase-2 (COX-2) enzyme is associated with the pathogenesis of inflammation, cancers, stroke, arthritis, and neurological disorders. Because of the involvement of COX-2 in these diseases, quantification of COX-2 expression using Positron Emission Tomography (PET) may be a biological marker for early diagnosis, monitoring of disease progression, and an indicator of effective treatment. At present there is no target-specific or validated PET tracer available for in vivo quantification of COX-2. The objective of this study is to evaluate [11C]TMI, a selective COX-2 inhibitor (Ki ≤ 1 nM) in nonhuman primates using PET imaging. PET imaging in baboons showed that [11C]TMI penetrates the blood brain barrier (BBB) and accumulates in brain in a somewhat heterogeneous pattern. Metabolite analyses indicated that [11C]TMI undergoes no significant metabolism of parent tracer in the plasma for baseline scans, however a relative faster metabolism was found for blocking scan. All the tested quantification approaches provide comparable tracer total distribution volume (VT) estimates in the range of 3.2-7 (mL/cm3). We observed about 25% lower VT values in blocking studies with meloxicam, a nonselective COX-2 inhibitor, compared to baseline [11C]TMI binding. Our findings indicate that [11C]TMI may be a suitable PET tracer for the quantification of COX-2 in vivo. Further experiments are needed to confirm the potential of this tracer in COX-2 overexpressing models for brain diseases.

In Vivo Brain Imaging, Biodistribution, and Radiation Dosimetry Estimation of [11C]Celecoxib, a COX-2 PET Ligand, in Nonhuman Primates.

COX-2 selective inhibitors (COXIBs) are non-steroidal anti-inflammatory drugs (NSAIDs), with fewer side effects compared with non-selective NSAIDs, and are used for the treatment of arthritis, headaches, and other inflammatory diseases of the brain and peripheral tissues. Radiolabeled COXIBs may permit positron emission tomography (PET) imaging of COX-2 localization and activity in diseases, enable monitoring of inflammatory processes, and determine target occupancy of COX-2 activity by NSAIDs, thus, accelerating the development of novel CIXIBs. We synthesized [11C]celecoxib, one of the COXIBs and a prescription drug, and here report its in vivo uptake in the brain, whole body biodistribution, and radiation dosimetry in baboons using PET. Brain imaging experiments were performed in one baboon and whole body PET scans were performed in triplicates in two male baboons using an ECAT ACCEL (Siemens Medical Solutions, Inc. Knoxville) under anesthetic conditions. PET studies in baboons show that [11C]celecoxib penetrates the blood brain barrier (BBB) and accumulates in the brain, followed by a washout of radioactivity. The liver has the highest residence time and the gallbladder is the critical organ for [11C]celecoxib. Organ Level Internal Dose Assessment (OLINDA) estimates indicate that the maximum permissible single study dosage of [11C]celecoxib in humans is 1110 MBq (30 mCi) for both males and females under the 21 CFR 361.1 dose limit for research subjects.

Radiosynthesis and in vivo evaluation of [11C]MOV as a PET imaging agent for COX-2.

Radiosynthesis and in vivo evaluation of [11C]4-[5-(4-methylphenyl)-3-(trifluoromethyl)-1H-pyrazol-1-yl]benzenesulfonamide (methoxy analogue of valdecoxib, [11C]MOV), a COX-2 inhibitor, was conducted in rat and baboon. Synthesis of the reference standard MOV (3), and its desmethyl precursor 2 for radiolabeling were performed using 1,2-diphenylethan-1-one as the starting material in five steps with 15% overall yield. Radiosynthesis of [11C]MOV was accomplished in 40 ±â€¯10% yield and >99% radiochemical purity by reacting the precursor 2 in dimethyl formamide (DMF) with [11C]CH3I followed by removal of the dimethoxytrityl (DMT) protective group using trifluroacetic acid. PET studies in anesthetized baboon showed very low uptake and homogeneous distribution of [11C]MOV in brain. The radioligand underwent rapid metabolism in baboon plasma. MicroPET studies in male Sprague Dawley rats revealed [11C]MOV binding in lower thorax. The tracer binding in rats was partially blocked in heart and duodenum by the administration of 1 mg/kg oral dose of COX-2 inhibitor valdecoxib.

Radiosynthesis and Evaluation of [11C]HD-800, a High Affinity Brain Penetrant PET Tracer for Imaging Microtubules.

Microtubules (MTs) are highly abundant throughout the cytoskeleton, and their dysfunction is implicated in the pathogenesis of malignancies, various neurodegenerative disorders, and brain injuries. Validated radiotracers reported so far for MTs are [11C]paclitaxel, [18F]fluoropaclitaxel, and [11C]docetaxel; however, they are well-characterized substrates of efflux transporters and consequently have poor uptake into the brain due to minimal blood brain barrier (BBB) penetration. PET imaging of MT expression requires radiolabeled BBB penetrating MT ligands, and it may offer a direct and more sensitive approach for early diagnosis, monitoring disease progression, and treatment effects in brain diseases and assessing the clinical potential of targeted therapeutics and treatments. We have identified N-(4-methoxyphenyl)-N-5-dimethylfuro[2,3-d]pyrimidin-4-amine (HD-800) as a high affinity and selective colchicine site tubuline inhibitor amenable to radiolabel with C-11, a positron emitting isotope. HD-800 and desmethyl-HD-800 were synthesized in one step with 75% and 80% yields respectively from commercial synthons. The radiosynthesis of [11C]HD-800 was achieved in 45 ± 5% yield at EOS. Ex vivo biodistribution binding data of [11C]HD-800 indicate that the radioligand penetrated the BBB and it was retained in brain with 75% specific binding. Apart from the brain, specific binding was observed in muscle (55%), heart (50%), lungs (43%), blood (37%), and pancreas (30%). MicroPET imaging in mice showed excellent binding in brain that was blocked by preadministration of unlabeled HD-800 and a colchicine site binding MT ligand MPC-6827. The above results indicate that [11C]HD-800 may be a suitable PET ligand for the in vivo quantification of MT inside and outside the brain.

Radiosynthesis and in Vivo Evaluation of [11C]MPC-6827, the First Brain Penetrant Microtubule PET Ligand.

Abnormalities of microtubules (MTs) are implicated in the pathogenesis of many CNS diseases. Despite the potential of an MT imaging agents, no PET ligand is currently available for in vivo imaging of MTs in the brain. We radiolabeled [11C]MPC-6827, a high affinity MTA, and demonstrated its specific binding in rat and mice brain using PET imaging. Our experiments show that [11C]MPC-6827 has specific binding to MT in brain, and it is the first MT-binding PET ligand.

Radiosynthesis and evaluation of IGF1R PET ligand [11C]GSK1838705A.

Radiosynthesis and evaluation of [11C]GSK1838705A in mice using microPET and determination of specificity in human GBM UG87MR cells are described herein. The radioligand was synthesized by reacting desmethyl-GSK1838705A with [11C]CH3I using GE FX2MeI module in ∼5% yield (EOS), >95% radiochemical purity and a specific activity of 2.5±0.5Ci/µmol. MicroPET imaging in mice indicated that [11C]GSK1838705A penetrated blood brain barrier (BBB) and showed retention of radiotracer in brain. The radioligand exhibited high uptake in U87MG cells with >70% specific binding to IGF1R. Our experiments suggest that [11C]GSK-1838705A can be a potential PET radiotracer for the in vivo quantification of IGF1R expression in GBM and other brain tumors.

Radiosynthesis and in Vivo Evaluation of [11C]A1070722, a High Affinity GSK-3 PET Tracer in Primate Brain.

Dysfunction of glycogen synthase kinase 3 (GSK-3) is implicated in the etiology of Alzheimer's disease, Parkinson's disease, diabetes, pain, and cancer. A radiotracer for functional positron emission tomography (PET) imaging could be used to study the kinase in brain disorders and to facilitate the development of small molecule inhibitors of GSK-3 for treatment. At present, there is no target-specific or validated PET tracer available for the in vivo monitoring of GSK-3. We radiolabeled the small molecule inhibitor [11C]1-(7-methoxy- quinolin-4-yl)-3-(6-(trifluoromethyl)pyridin-2-yl)urea ([11C]A1070722) with high affinity to GSK-3 (Ki = 0.6 nM) in excellent radiochemical yield. PET imaging experiments in anesthetized vervet/African green monkey exhibited that [11C]A1070722 penetrated the blood-brain barrier (BBB) and accumulated in brain regions, with highest radioactivity binding in frontal cortex followed by parietal cortex and anterior cingulate, and with the lowest bindings found in caudate, putamen, and thalamus, similarly to the known distribution of GSK-3 in human brain. Our studies suggest that [11C]A1070722 can be a potential PET radiotracer for the in vivo quantification of GSK-3 in brain.

In vivo evaluation of IGF1R/IR PET ligand [18F]BMS-754807 in rodents.

In vivo evaluation of [18F]BMS-754807 binding in mice and rats using microPET and biodistribution methods is described herein. The radioligand shows consistent binding characteristics, in vivo, in both species. Early time frames of the microPET images and time activity curves of brain indicate poor penetration of the tracer across the blood brain barrier (BBB) in both species. However, microPET experiments in mice and rats show high binding of the radioligand outside the brain to heart, pancreas and muscle, the organs known for higher expression of IGF1R/1R. Biodistribution analysis 2h after injection of [18F]BMS-754807 in rats show negligible [18F]defluorination as reflected by the low bone uptake and clearance from blood. Overall, the data indicate that [18F]BMS-754807 can potentially be a radiotracer for the quantification of IGF1R/IR outside the brain using PET.

In vivo evaluation of [18F]FECIMBI-36, an agonist 5-HT2A/2C receptor PET radioligand in nonhuman primate.

We recently reported the radiosynthesis and in vitro evaluation of [18F]-2-(4-bromo-2,5-dimethoxyphenyl)-N-(2-(2-fluoroethoxy)benzyl)ethanamine, ([18F]FECIMBI-36) or ([18F]1), an agonist radioligand for 5HT2A/2C receptors in postmortem samples of human brain. Herein we describe the in vivo evaluation of [18F]FECIMBI-36 in vervet/African green monkeys by PET imaging. PET images show that [18F]FECIMBI-36 penetrates the blood-brain barrier and a low retention of radioactivity is observed in monkey brain. Although the time activity curves indicate a somehow heterogeneous distribution of the radioligand in the brain, the low level of [18F]FECIMBI-36 in brain may limit the use of this tracer for quantification of 5-HT2A/2C receptors by PET.

Autoradiographic Evaluation of [(18)F]FECUMI-101, a High Affinity 5-HT1AR Ligand in Human Brain.

[(18)F]FECUMI-101 ([(18)F]1) is a 5HT1AR ligand demonstrating specific binding in brain regions corresponding to the distribution of 5-HT1AR in baboons. However, we detected moderate uptake of [(18)F]1 in baboon thalamus, a brain region lacking 5-HT1AR. We sought to investigate the relative binding of [(18)F]1 to 5-HT1AR, α1R, and 5-HT7R in vitro. Using autoradiography in human brain sections, specific binding of [(18)F]1 to 5-HT1AR was confirmed. However, [(18)F]1 also showed 26% binding to α1R in PFC. The hippocampal formation exhibited 51% and 92% binding of [(18)F]1 to α1R and 5-HT1AR, respectively. Thalamus and cerebellum showed very little binding. There is no measurable specific binding of [(18)F]1 to 5-HT7R and no effect of temperature on [(18)F]1 specific binding to 5-HT1AR or α1R. These results indicate that, while [(18)F]FECUMI-101 is not a completely selective 5-HT1AR ligand for receptor quantification, it may be useful for occupancy measurements of drugs acting at 5-HT1AR in vivo.

Radiosynthesis and in Vivo Evaluation of Neuropeptide Y5 Receptor (NPY5R) PET Tracers.

Neuropeptide Y receptor type 5 (NPY5R) is a G-protein coupled receptor (GPCR) that belongs to the subfamily of neuropeptide receptors (NPYR) that mediate the action of endogenous neuropeptide Y (NPY). Animal models and preclinical studies indicate a role for NPY5R in the pathophysiology of depression, anxiety, and obesity and as a target of potential therapeutic drugs. To better understand the pathophysiological involvement of NPY5R, and to measure target occupancy by potential therapeutic drugs, it would be advantageous to measure NPY5R binding in vivo by positron emission tomography (PET). Four potent and selective NPY5R antagonists were radiolabeled via nucleophilic aromatic substitution reactions with [(18)F]fluoride. Of the four radioligands investigated, PET studies in anesthetized baboons showed that [(18)F]LuAE00654 ([(18)F]N-[trans-4-({[4-(2-fluoropyridin-3-yl)thiazol-2-yl]amino}methyl)cyclohexyl]propane-2-sulfonamide) penetrates blood brain barrier (BBB) and a small amount is retained in the brain. Slow metabolism of [(18)F]LuAE00654 was observed in baboon plasma. Blocking studies with a specific NPY5R antagonist demonstrated up to 60% displacement of radioactivity in striatum, the brain region with highest NPY5R binding. Our studies suggest that [(18)F]LuAE00654 can be a potential PET radiotracer for the quantification and occupancy studies of NPY5R drug candidates.

Synthesis and in vitro evaluation of [18F]FECIMBI-36: A potential agonist PET ligand for 5-HT2A/2C receptors.

Radiosynthesis and in vitro evaluation of [(18)F]-2-(4-bromo-2,5-dimethoxyphenyl)-N-(2-(2-fluoroethoxy)benzyl)ethanamine, ([(18)F]FECIMBI-36) or ([(18)F]1), a potential agonist PET imaging agent for 5-HT2A/2C receptors is described. Syntheses of reference standard 1 and the corresponding des-fluoroethyl radiolabeling precursor (2) were achieved with 75% and 65% yields, respectively. In vitro pharmacology assay of FECIMBI-36 by [(3)H]-ketanserin competition binding assay obtained from NIMH-PDSP showed high affinities to 5-HT2AR (Ki = 1nM) and 5-HT2CR (Ki=1.7 nM). Radiolabeling of FECIMBI-36 was achieved from the boc-protected precursor 2 using [(18)F]-fluoroethyltosylate in presence of Cs2CO3 in DMSO followed by removal of the protective group. [(18)F]1 was isolated using RP-HPLC in 25 ± 5% yield, purity > 95% and specific activity 1-2Ci/µmol (N = 6). In vitro autoradiography studies demonstrate that [(18)F]1 selectively label 5-HT2A and 5-HT2C receptors in slide-mounted sections of postmortem human brain using phosphor imaging. Our results indicate the potential of [(18)F]1 for imaging 5-HT2A/2C receptors in the high affinity state in vivo using PET imaging.