Comparative assessment of (18) F-Mefway as a serotonin 5-HT1A receptor PET imaging agent across species: Rodents, nonhuman primates, and humans.

We have developed (18) F-trans-Mefway ((18) F-Mefway) for positron emission tomography (PET) imaging studies of serotonin 5-HT1A receptors which are implicated in various brain functions. Translation of imaging the 5-HT1A receptor in animal models to humans will facilitate an understanding of the role of the receptor in human brain disorders. We report comparative brain distribution of (18) F-Mefway in normal mice, rats, monkeys, and healthy human volunteers. Mefway was found to be very selective, with subnanomolar affinity for the 5-HT1A receptor. Affinities of >55 nM were found for all other human-cloned receptor subtypes tested. Mefway was found to be a poor substrate (>30 µM) for the multidrug resistance 1 protein, suggesting low likelihood of brain uptake being affected by P-glycoprotein. Cerebellum was used as a reference region in all imaging studies across all species due to the low levels of (18) F-Mefway binding. Consistent binding of (18) F-Mefway in cortical regions, hippocampus, and raphe was observed across all species. (18) F-Mefway in the human brain regions correlated with the known postmortem distribution of 5-HT1A receptors. Quantitation of raphe was affected by the resolution of the PET scanners in rodents, whereas monkeys and humans showed a raphe to cerebellum ratio of approximately 3. (18) F-Mefway appears to be an effective 5-HT1A receptor imaging agent in all models, including humans. (18) F-Mefway therefore may be used to quantify 5-HT1A receptor distribution in brain regions for the study of various CNS disorders. J. Comp. Neurol. 524:1457-1471, 2016. © 2015 Wiley Periodicals, Inc.

Evaluation of serotonin 5-HT(1A) receptors in rodent models using [¹8F]mefway PET.

INTRODUCTION: Serotonin 5-HT(1A) receptors have been investigated in various CNS disorders, including epilepsy, mood disorders, and neurodegeneration. [¹8F]Mefway (N-{2-[4-(2'-methoxyphenyl)piperazinyl]ethyl}-N-(2-pyridyl)-N-(cis/trans-4'-[¹8F]fluoromethylcyclohexane)-carboxamide) has been developed as a suitable positron emission tomography (PET) imaging agent for these receptors. We have now evaluated the suitability of [¹8F]trans-mefway in rat and mouse models using PET and computerized tomography (CT) imaging and corroborated with ex vivo and in vitro autoradiographic studies. METHODS: Normal Sprague-Dawley rats and Balb/C mice were used for PET/CT imaging using intravenously injected [¹8F]trans-mefway. Brain PET data were coregistered with rat and mouse magnetic resonance imaging template and regional distribution of radioactivity was quantitated. Selected animals were used for ex vivo autoradiographic studies to confirm regional brain distribution and quantitative measures of binding, using brain region to cerebellum ratios. Binding affinity of trans-mefway and WAY-100635 was measured in rat brain homogenates. Distribution of [¹8F]trans-4-fluoromethylcyclohexane carboxylate ([¹8F]FMCHA), a major metabolite of [¹8F] trans-mefway, was assessed in the rat by PET/CT. RESULTS: The inhibition constant, K(i) for trans-mefway was 0.84 nM and that for WAY-100635 was 1.07 nM. Rapid brain uptake of [¹8F]trans-mefway was observed in all rat brain regions and clearance from cerebellum was fast and was used as a reference region in all studies. Distribution of [¹8F]trans-mefway in various brain regions was consistent in PET and in vitro studies. The dorsal raphe was visualized and quantified in the rat PET but identification in the mouse was difficult. The rank order of binding to the various brain regions was hippocampus > frontal cortex > anterior cingulate cortex > lateral septal nuclei > dorsal raphe nuclei. CONCLUSION: [¹8F]trans-Mefway appears to be an effective 5-HT(1A) receptor imaging agent in rodents for studies of various disease models.

Synthesis and evaluation of 18F-FE-PEO in rodents: an 18F-labeled full agonist for opioid receptor imaging.

UNLABELLED: We have investigated the opioid receptor (OR) agonist (20R)-4,5-α-epoxy-6-(2-(18)F-fluoroethoxy)-3-hydroxy-α,17-dimethyl-α-(2-phenyleth-1-yl)-6,14-ethenomorphinan-7-methanol ((18)F-FE-PEO) as a candidate OR PET ligand. This tracer is attractive because it combines (18)F labeling, is suited to the slow kinetics of high-affinity ligands, and has agonist binding, which has been shown to be more sensitive to changes in OR occupation than is antagonist binding. METHODS: Agonist potency and off-target binding were investigated in vitro, and autoradiographic studies on rat brain sections were used to assess binding patterns. Quantification of the tracer in vivo was investigated using small-animal PET in rats with blood sampling. RESULTS: (18)F-FE-PEO was obtained by direct nucleophilic radiofluorination and subsequent deprotection with a yield of 28% ± 15%, a specific activity of 52-224 MBq/nmol, and a radiochemical purity of more than 97% (90 min from end of bombardment). In vitro studies showed it to be a full agonist ligand, which selectively binds to OR with high affinity, although it is not selective to a single OR subtype (inhibition constant, 0.4-1.6 nM across OR subtypes). Autoradiography binding patterns were consistent with the known distribution of OR, although nondisplaceable signal typically constituted one third of the signal in OR-dense regions. Although metabolites were present in blood (∼40% of plasma radioactivity was nonparent 3 h after injection), no significant metabolite fraction was found in brain tissue, aiding PET quantification. A plasma input 2-tissue-compartment model provided good fits to the PET data, and regional distribution volumes from the latter correlated well with those from Logan plot analysis (r(2) = 0.98). The cerebellum had the lowest distribution volume, but the time-activity curve data could not be adequately fitted with a 1-tissue-compartment model. Reference tissue models using the cerebellum as the reference region did not provide good fits to the data, so blood-based kinetic analysis is recommended. CONCLUSION: As the first (18)F-labeled OR agonist ligand, (18)F-FE-PEO is a useful addition to the existing OR ligand portfolio.

Synthesis and biologic evaluation of a novel serotonin 5-HT1A receptor radioligand, 18F-labeled mefway, in rodents and imaging by PET in a nonhuman primate.

UNLABELLED: Serotonin 5-HT1A receptors have been implicated in disorders of the central nervous system and, therefore, are being studied by PET. Efforts are under way to improve in vivo stability of 5-HT1A agents currently in human use (11C-labeled N-(2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl-N-(2-pyridinyl)cyclohexanecarboxamide [11C-WAY-100635], 4-(2'-methoxyphenyl)-1-[2'-(N-2''-pyridinyl)-p-18F-fluorobenzamido]ethylpiperazine [18F-MPPF], and 18F-labeled trans-4-fluoro-N-(2-[4-(2-methoxyphenyl)piperazin-1-yl)ethyl]-N-(2-pyridyl)cyclohexanecarboxamide [18F-FCWAY]). We have synthesized N-{2-[4-(2-methoxyphenyl)piperazinyl]ethyl}-N-(2-pyridyl)-N-(4-18F-fluoromethylcyclohexane)carboxamide (18F-mefway), which contains a 18F on a primary carbon to make the compound more stable to defluorination. METHODS: Radiosynthesis of 18F-mefway was performed in a single tosylate for 18F-fluoride exchange. In vitro binding studies on rat brain slices using 18F-mefway were read on a phosphor imager. Monkey PET studies were performed on a whole-body PET scanner. RESULTS: Binding affinity (inhibitory concentration of 50% [IC50]) of mefway was 26 nmol/L and was comparable to that of WAY-100635, 23 nmol/L. Yields of 18F-mefway were 20%-30% in specific activities of 74-111 GBq/micromol at the end of synthesis. In vitro binding of 18F-mefway in the hippocampus (Hp), colliculus (Co), cortex (Ctx), and other brain regions-with limited binding in the cerebellum (Cer)--was observed, with ratios of Hp/Cer = 82.3, Co/Cer = 45.8, and Ctx/Cer = 40. Serotonin displaced 18F-mefway from various brain regions with IC50 values in the range of 169-243 nmol/L. PET studies in a rhesus monkey showed 18F-mefway binding in the fontal cortex (FC), temporal cortex (TC) including hippocampus, raphe (Rp), and other brain regions, with ratios of FC/Cer = 9.0, TC/Cer = 10, and Rp/Cer = 3.3. Plasma analysis indicated the presence of approximately 30% of 18F-mefway at 150-180 min after injection. CONCLUSION: The high ratios in specific brain regions such as the hippocampus suggest that 18F-mefway has potential as a PET agent for 5HT1A receptors.