Synthesis and in vitro biological evaluation of carbon-11-labeled quinoline derivatives as new candidate PET radioligands for cannabinoid CB2 receptor imaging

Cannabinoids have been recently proposed as a new family of potential antitumor agents, and cannabinoid receptor 2 (CB2) is believed to be over-expressed in tumor cells. This study was designed to develop new radioligands for imaging of CB2 receptor in cancer using biomedical imaging technique positron emission tomography (PET). Carbon-11-labeled 2-oxoquinoline and 2-chloroquinoline derivatives, [¹¹C]6a–d and [¹¹C]9a–d, were prepared by O-[¹¹C]methylation of their corresponding precursors using [¹¹C]CH₃OTf under basic conditions and isolated by a simplified solid-phase extraction (SPE) method in 40–50% radiochemical yields based on [¹¹C]CO₂ and decay corrected to end of bombardment (EOB). The overall synthesis time from EOB was 15–20 min, the radiochemical purity was >99%, and the specific activity at end of synthesis (EOS) was 111–185 GBq/μmol. Radioligand binding assays indicated compounds 6f, 6b, and 9f display potent in vitro binding affinities with nanomolar K_i values and at least 100–2000-fold selectivity for CB2.     

Radionuclide labeling and evaluation of candidate radioligands for PET imaging of histone deacetylase in the brain

Histone deacetylases (HDACs) regulate gene expression by inducing conformational changes in chromatin. Ever since the discovery of a naturally occurring HDAC inhibitor, trichostatin A (TSA) stimulated the recent development of suberoylanilide (SAHA, Zolinza®), HDAC has become an important molecular target for drug development. This has created the need to develop specific in vivo radioligands to study epigenetic regulation and HDAC engagement for drug development for diseases including cancer and psychiatric disorders. 6-([¹⁸F]Fluoroacetamido)-1-hexanoicanilide ([¹⁸F]FAHA) was recently developed as a HDAC substrate and shows moderate blood–brain barrier (BBB) permeability and specific signal (by metabolic trapping/or deacetylation) but rapid metabolism. Here, we report the radiosynthesis of two carbon-11 labeled candidate radiotracers (substrate- and inhibitor-based radioligand) for HDAC and their evaluation in non-human primate brain. PET studies showed very low brain uptake and rapid metabolism of both labeled compounds but revealed a surprising enhancement of brain penetration by F for H substitution when comparing one of these to [¹⁸F]FAHA. Further structural refinement is needed for the development of brain-penetrant, metabolically stable HDAC radiotracers and to understand the role of fluorine substitution on brain penetration.     

Synthesis, in vitro characterization, and radiolabeling of reboxetine analogs as potential PET radioligands for imaging the norepinephrine transporter

Six new (S,S)-enantiomers of reboxetine derivatives were synthesized and their binding affinities were determined via competition binding assays in cells expressing the human norepinephrine transporter (NET), serotonin transporter (SERT) or dopamine transporter (DAT). All six compounds prepared exhibit high affinity for the NET (K(i)相似文献   

Synthesis of potent and selective dopamine D(4) antagonists as candidate radioligands

A series of dopamine D(4) antagonists was synthesized and evaluated as potential candidates for development as positron emission tomography (PET) radioligands. All new compounds display high affinity and selectivity for the D(4) receptors and compounds 5b, 5d, and 5e were identified as candidates for radioligand development.     

Synthesis of carbon-11-labeled aminoalkylindole derivatives as new candidates of cannabinoid receptor radioligands for PET imaging of alcohol abuse

Carbon-11-labeled aminoalkylindole derivatives (1-butyl-7-[¹¹C]methoxy-1H-indol-3-yl)(naphthalene-1-yl)methanone ([¹¹C]3), 1-butyl-7-[¹¹C]methoxy-3-(naphthalene-1-ylmethyl)-1H-indole ([¹¹C]5), and 1-butyl-7-[¹¹C]methoxy-3-(naphthalene-2-yl)-1H-indole ([¹¹C]8) were prepared by O-[¹¹C]methylation of their corresponding precursors with [¹¹C]CH₃OTf under basic condition (2 N NaOH) and isolated by a simplified solid-phase extraction (SPE) method in 50–60% radiochemical yields based on [¹¹C]CO₂ and decay corrected to end of bombardment (EOB). The overall synthesis time from EOB was 23 min, the radiochemical purity was >99%, and the specific activity at end of synthesis (EOS) was 185–555 GBq/μmol.     

Synthesis and evaluation of 11C-labeled coumarin analog as an imaging probe for detecting monocarboxylate transporters expression

Upregulated monocarboxylate transporters (MCTs) in tumors are considered diagnostic imaging targets. Herein, we synthesized the positron emission tomography probe candidates coumarin analogs 2 and 3, and showed 55 times higher affinity of 2 for MCTs than a representative MCT inhibitor. Whereas [¹¹C]2 showed low tumor accumulation, probably due to adduct formation with plasma proteins, [¹¹C]2 showed high initial brain uptake, suggesting that the scaffold of 2 has properties that are preferable in imaging probes for the astrocyte–neuron lactate shuttle. Although further optimization of 2 is required, our findings can be used to inform the development of MCT-targeted imaging agents.     

Synthesis and neuropharmacological evaluation of R(-)-N-alkyl-11-hydroxynoraporphines and their esters

We synthesized several N-substituted-11-hydroxynoraporphines and their esters of varying chain length, evaluated their binding affinity at dopamine (DA) receptor sites in rat caudate-putamen membranes, and quantified their effects on motor activity in normal adult male rats. The 11-hydroxyaporphines showed similar neuropharmacological properties to the corresponding 10,11-catecholaporphines. At moderate doses, their esters proved to have more prolonged behavioral actions and superior oral bioavailability.     

Synthesis and biodistribution of [(11)C]R116301, a promising PET ligand for central NK(1) receptors

N1-(2,6-Dimethylphenyl)-2-(4-{(2R,4S)-2-benzyl-1-[3,5-di(trifluoromethyl)[carbonyl-(11)C]benzoyl]hexahydro-4-pyridinyl}piperazino)acetamide ([(11)C]R116301) was prepared and evaluated as a potential positron emission tomography (PET) ligand for investigation of central neurokinin(1) (NK(1)) receptors. 1-Bromo-3,5-di(trifluoromethyl)benzene was converted in three steps into 3,5-di(trifluoromethyl)[carbonyl-(11)C]benzoyl chloride, which was reacted with N1-(2,6-dimethylphenyl)-2-{4-[(2R,4S)-2-benzylhexahydro-4-pyridinyl]piperazino}acetamide providing [(11)C]R116301 in 45-57% decay-corrected radiochemical yield. The total synthesis time, from end of bombardment (EOB) to the formulated product, was 35 min. Specific activity (SA) was 82-172 GBq/micromol (n=10) at the end of synthesis. N1-([4-(3)H]-2,6-Dimethylphenyl)-2-(4-{(2R,4S)-2-benzyl-1-[3,5-di(trifluoromethyl)benzoyl]hexahydro-4-pyridinyl}piperazino)acetamide ([(3)H]R116301) was also synthesized (SA: 467 GBq/mmol). The B(max) for [(3)H]R116301 measured in vitro on Chinese hamster ovary cell membranes stably transfected with the human NK(1) receptor was 19.10+/-1.02 pmol/mg protein with an apparent dissociation constant of 0.08+/-0.01 nM. Ex vivo, in vivo and in vitro autoradiography studies with [(3)H]R116301 in gerbils demonstrated a preferential accumulation of the radioactivity in the striatum, olfactory tubercule, olfactory bulb and locus coeruleus. In vivo, the biodistribution of [(11)C]R116301 in gerbils revealed that the highest initial uptake is in the lung, followed by the liver and kidney. In the brain, maximum accumulation was found in the olfactory tubercules (1.10+/-0.08 injected dose (ID)/g 20 min post injection (p.i.)) and the nucleus accumbens (1.00+/-0.12ID/g 10 min p.i.). Tissue/cerebellum concentration ratios for striatum and nucleus accumbens increased with time due to rapid uptake followed by a slow wash out (1.29 and 1.64, respectively, 30 min p.i.). A tissue to cerebellum ratio of 1.33 and 1.62 was also observed for olfactory bulb and olfactory tubercules, respectively (20 min p.i.). In summary, [(11)C]R116301 appears to be a promising radioligand suitable for the visualization of NK(1) receptors in vivo using PET.     

Synthesis and biological evaluation of novel carbon-11-labelled analogues of citalopram as potential radioligands for the serotonin transporter

Three serotonin reuptake inhibitors where the 5-cyano group in citalopram [1-(3-dimethylamino-propyl)-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-carbonitrile (1)] was replaced with a methyl, acetyl and piperidinyl carbonyl group, respectively, were synthesized. In a Stille reaction applying [(11)C]methyl iodide the labelled compound [5-methyl-(11)C][3-[1-(4-fluorophenyl)-5-methyl-1,3-dihydroisobenzofuran-1-yl]-propyl]-dimethylamine ([(11)C]-2) was synthesized in 60-90% radiochemical yield. [5-carbonyl-(11)C][1-[1-(3-dimethylaminopropyl)-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-yl]-1-piperidin-1-yl-methanone] ([(11)C]-3) was synthesized in 62% radiochemical yield in a palladium mediated cross-coupling reaction utilizing [(11)C]carbon monoxide. The specific activity of [(11)C]-2 was highly dependent on whether the corresponding trimethyltin or tributyltin precursor was applied. In ex vivo rodent studies compound [(11)C]-2 exhibited a good blood-brain barrier (BBB) penetration whereas [(11)C]-3 did not. The brain distribution of [(11)C]-2 was investigated in a non-human primate using PET. There was a rapid uptake of radioactivity into the brain. Accumulation of the radiotracer was in agreement with the known distribution of serotonin transporters. The maximal thalamus to cerebellum ratio of 1.3 was reached after 85 min and the specific binding was partly blocked after pre-treatment with citalopram. Thus, [(11)C]-2 does not exhibit appropriate properties as radioligand for visualization of the serotonin transporter in vivo.     

Synthesis and evaluation of indolinyl- and indolylphenylacetylenes as PET imaging agents for beta-amyloid plaques

Two new phenylacetylene derivatives, 5-((4-(2-(2-(2-fluoroethoxy)ethoxy)ethoxy)phenyl)ethynyl)indoline 8 and 5-((4-(2-(2-(2-fluoroethoxy)ethoxy)ethoxy)phenyl)ethynyl)-1H-indole 14, targeting β-amyloid (Aβ) plaques have been prepared. In vitro binding carried out in tissue homogenates prepared from postmortem AD brains with [¹²⁵I]IMPY (6-iodo-2-(4′-dimethylamino-)phenyl-imidazo[1,2-a]pyridine) as the radioligand indicated good binding affinities (K_i = 4.0 and 1.5 nM for 8 and 14, respectively). Brain penetration of the corresponding radiofluorinated ligands, evaluated in the normal mice, showed good initial brain penetration (4.50 and 2.43% ID/g (injected dose/gram) for [¹⁸F]8 and [¹⁸F]14 at 2 min after injection) with moderate to low washout rates from the brain (1.71% ID/g at 2 h and 2.10% ID/g at 3 h, respectively). Autoradiography and homogenate binding studies demonstrated the high specific binding of [¹⁸F]14 to the Aβ plaques; however, [¹⁸F]8 showed low specific binding. These preliminary results identified that indolylphenylacetylene, 14, may be a good lead for further structural modification to develop a useful Aβ plaque imaging agent.     

Synthesis and evaluation of new 1-oxa-8-azaspiro[4.5]decane derivatives as candidate radioligands for sigma-1 receptors

We report the design, synthesis, and evaluation of a series of 1-oxa-8-azaspiro[4.5]decane and 1,5-dioxa-9-azaspiro[5.5]undecane derivatives as selective σ₁ receptor ligands. All seven ligands exhibited nanomolar affinity for σ₁ receptors (K_i(σ₁) = 0.47 – 12.1 nM) and moderate selectivity over σ₂ receptors (K_i(σ₂)/ K_i(σ₁) = 2 – 44). Compound 8, with the best selectivity among these ligands, was selected for radiolabeling and further evaluation. Radioligand [¹⁸F]8 was prepared via nucleophilic ¹⁸F-substitution of the corresponding tosylate precursor, with an overall isolated radiochemical yield of 12–35%, a radiochemical purity of greater than 99%, and molar activity of 94 – 121 GBq/μmol. Biodistribution studies of [¹⁸F]8 in mice demonstrated high initial brain uptake at 2 min. Pretreatment with SA4503 resulted in significantly reduced brain-to-blood ratio (70% − 75% at 30 min). Ex vivo autoradiography in ICR mice demonstrated high accumulation of the radiotracer in σ₁ receptor-rich brain areas. These findings suggest that [¹⁸F]8 could be a lead compound for further structural modifications to develop potential brain imaging agents for σ₁ receptors.     

Synthesis and C-11 labeling of three potent norepinephrine transporter selective ligands ((R)-nisoxetine, lortalamine, and oxaprotiline) for comparative PET studies in baboons

Three potent antidepressants, (R)-nisoxetine, lortalamine, and oxaprotiline, with high affinity and high selectivity for the norepinephrine transporter (NET) were synthesized and radiolabeled with C-11 via [11C]methylation. The reference compounds and their corresponding normethyl precursors were synthesized via multi-step synthetic approaches. The radiochemical syntheses were performed by simple alkylation of the corresponding normethyl precursors with no-carrier-added [11C]CH3I in DMF. After HPLC purification, (R)-[N-11CH3]nisoxetine, [11C]lortalamine, and [11C]oxaprotiline were obtained in 63-97% radiochemical yields, whereas (R)-[O-11CH3]nisoxetine was obtained in 23-29% radiochemical yields due to substantial formation of the undesired N-[11C]methylated byproduct (64-70%). These C-11 labeled tracers allowed us to carry out comparative studies of NET in baboons with positron emission tomography (PET) and evaluate their potential as PET tracers for imaging brain NET.     

Synthesis and biological evaluation of a series of flavone derivatives as potential radioligands for imaging the multidrug resistance-associated protein 1 (ABCC1/MRP1)

Multidrug resistance (MDR) is one of the major problems affecting the treatment of cancer. In vivo visualization and quantification of MDR proteins would be of great value to better select the therapeutic strategy. Six flavone-based compounds were synthesized and evaluated for their cytotoxic activity and MDR-reversing capacity using hMRP1 or hMDR1 overexpressing cell lines for in vitro assays. All the flavone derivatives were highly selective for hMRP1-expressing cell lines. These derivatives each used at 4muM (a non-cytotoxic concentration) enhance significantly the sensitivity of hMRP1-mediated MDR cell line toward doxorubicin toxicity. Their MDR-reversing capacity suggests that, in particular, the 4'-fluoroalkyloxy and 4'-iodo apigenin derivatives are potential new radiopharmaceuticals to visualize in vivo MRP1-mediated MDR phenomenon by PET or SPECT.     

Synthesis and biological evaluation of 2beta,3alpha-(substituted phenyl)nortropanes as potential norepinephrine transporter imaging agents

A series of 2beta,3alpha-(substituted phenyl)nortropanes was synthesized and evaluated in vitro for human monoamine transporters. All compounds studied in this series exhibited nanomolar potency for the norepinephrine transporter (NET). Radiolabeling and nonhuman primate microPET brain imaging studies were performed with the most promising compound, [(11)C]1, to determine its utility as a NET imaging agent. Despite high in vitro affinity for the human NET, the high uptake of [(11)C]1 in the caudate and putamen excludes its use as an in vivo PET imaging agent for the NET.     

Facile radiosynthesis of new carbon-11-labeled propanamide derivatives as selective androgen receptor modulator (SARM) radioligands for prostate cancer imaging

The androgen receptor (AR) is an attractive target for the treatment and molecular imaging of prostate cancer. New carbon-11-labeled propanamide derivatives were first designed and synthesized as selective androgen receptor modulator (SARM) radioligands for prostate cancer imaging using the biomedical imaging technique positron emission tomography (PET). The target tracers, (S)-N-(4-cyano-3-(trifluoromethyl)phenyl)-2-hydroxy-3-(2-[¹¹C]methoxyphenoxy)-2-methylpropanamide ([¹¹C]8a), (S)-2-hydroxy-3-(2-[¹¹C]methoxyphenoxy)-2-methyl-N-(4-nitro-3-(trifluoromethyl)phenyl)propanamide ([¹¹C]8e), (S)-N-(4-cyano-3-(trifluoromethyl)phenyl)-2-hydroxy-3-(4-[¹¹C]methoxyphenoxy)-2-methylpropanamide ([¹¹C]8c) and (S)-2-hydroxy-3-(4-[¹¹C]methoxyphenoxy)-2-methyl-N-(4-nitro-3-(trifluoromethyl)phenyl)propanamide ([¹¹C]8g), were prepared by O-[¹¹C]methylation of their corresponding precursors, (S)-N-(4-cyano-3-(trifluoromethyl)phenyl)-2-hydroxy-3-(2-hydroxyphenoxy)-2-methylpropanamide (9a), (S)-2-hydroxy-3-(2-hydroxyphenoxy)-2-methyl-N-(4-nitro-3-(trifluoromethyl)phenyl)propanamide (9b), (S)-N-(4-cyano-3-(trifluoromethyl)phenyl)-2-hydroxy-3-(4-hydroxyphenoxy)-2-methylpropanamide (9c) and (S)-2-hydroxy-3-(4-hydroxyphenoxy)-2-methyl-N-(4-nitro-3-(trifluoromethyl)phenyl)propanamide (9d), with [¹¹C]CH₃OTf under basic conditions and isolated by a simplified C-18 solid-phase extraction (SPE) method in 55 ± 5% (n = 5) radiochemical yields based on [¹¹C]CO₂ and decay corrected to end of bombardment (EOB). The overall synthesis time from EOB was 23 min, the radiochemical purity was >99%, and the specific activity at end of synthesis (EOS) was 277.5 ± 92.5 GBq/μmol (n = 5).     

Synthesis and evaluation of novel tropane derivatives as potential PET imaging agents for the dopamine transporter

A novel series of tropane derivatives containing a fluorinated tertiary amino or amide at the 2β position was synthesized, labeled with the positron-emitter fluorine-18 (t(1/2)=109.8 min), and tested as potential in vivo dopamine transporter (DAT) imaging agents. The corresponding chlorinated analogs were prepared and employed as precursors for radiolabeling leading to the fluorine-18-labeled derivatives via a one-step nucleophilic aliphatic substitution reaction. In vitro binding results showed that the 2β-amino compounds 6b, 6d and 7b displayed moderately high affinities to DAT (K(i)<10nM). Biodistribution studies of [(18)F]6b and [(18)F]6d showed that the brain uptakes in rats were low. This is likely due to their low lipophilicities. Further structural modifications of these tropane derivatives will be needed to improve their in vivo properties as DAT imaging agents.     

Synthesis of novel neutrophil-specific imaging agents for Positron Emission Tomography (PET) imaging

A neutrophil-specific peptide, cinnamoyl-F(D)LF(D)LFK (cFLFLFK), was conjugated consecutively with a polyethylene glycol moiety (3.4 K) and 2,2′,2″,2-(1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrayl)tetraacetic acid (DOTA) to form cFLFLFK-PEG-DOTA. After ⁶⁴Cu labeling, Positron Emission Tomography (PET) imaging was successfully able to detect mouse lung inflammation.     

Synthesis of 68Ga-labeled DOTA-nitroimidazole derivatives and their feasibilities as hypoxia imaging PET tracers

The imaging of hypoxia is important for therapeutic decision making in various diseases. (68)Ga is an important radionuclide for positron emission tomography (PET), and its usage is increasing, due to the development of the (68)Ge/(68)Ga-generator. In the present study, the authors synthesized two nitroimidazole derivatives by conjugating nitroimidazole and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) via an amide bond (4) and a thiourea bond (5). Both derivatives were labeled with (68)Ga with high labeling efficiency and were stable after labeling. The low partition coefficients (logP) of (68)Ga-4 (-4.6) and (68)Ga-5 (-4.5) demonstrated the hydrophilic natures of the derivatives, and both showed higher uptake in cancer cell lines cultured under hypoxic condition than under normoxic condition. However, (68)Ga-5 showed higher liver uptake than (68)Ga-4 in a biodistribution study due to higher lipophilicity. In an animal PET study, (68)Ga-4 showed higher standard uptake values (SUV) in tumors than (68)Ga-5 in mice xenografted with CT-26 mouse colon cancer cells.     

Synthesis and preliminary evaluation of [(18)F]-fluoro-(2S)-Exaprolol for imaging cerebral beta-adrenergic receptors with PET

Cerebral beta-adrenergic receptors (beta-ARs) are of interest in several disorders including Parkinson's disease, Alzheimer's disease and in particular major depressive disorder. Development of a positron emission tomography (PET) ligand for imaging beta-ARs would allow the quantification of these receptors in the living human brain so as to better understand both the pathophysiology of depression and how to improve treatment. Currently there are no radioligands suitable for this purpose. In an attempt to achieve this goal, we prepared [(18)F]-labeled (2S)-1-(1-fluoropropan-2-ylamino)-3-(2-cyclohexylphenoxy)propan-2-ol (fluoro-Exaprolol; (2S)-1). Radiolabeling with fluorine-18 was accomplished via preparation of a precursor containing a tosyl leaving group (10), and utilizes the 2-oxazolidinone group to simultaneously protect both the amine and hydroxy groups. The oxazolidinone was readily removed with lithium aluminum hydride following a nucleophilic [(18)F]-fluoride for tosyl displacement to prepare [(18)F]-(2S)-1 in 31% radiochemical yield (uncorrected for decay), with >98% radiochemical purity in <1h. The specific activity of the formulated product was 927 mCi/micromol and the log P (pH 7.4) was 2.97. Preliminary biological evaluations in conscious rats indicated that [(18)F]-(2S)-1 had good brain uptake for imaging (0.8-1.3% injected dose/gram (% ID/g) of wet tissue, 5 min post-injection of the radiotracer) with a slow washout (>0.5% ID/g at 60 min post-injection) in all brain regions. Pharmacological challenges indicate that the binding is largely non-specific, as administration of Propranolol, authentic (2S)-1, or WAY 100635 prior to injection of [(18)F]-(2S)-1 did not block uptake of the radiotracer. These results indicate that [(18)F]-(2S)-1 is not a suitable candidate for PET imaging of cerebral beta-ARs.