¹¹C-labeled analogs of indomethacin esters and amides for brain cyclooxygenase-2 imaging: radiosynthesis, in vitro evaluation and in vivo characteristics in mice

There is great potential in the use of positron emission tomography (PET) and suitable radiotracers for the study of cyclooxygenase type 2 (COX-2) enzyme in living subjects. In the present study, we prepared and evaluated five 11C-labeled ester and amide analogs derived from indomethacin as potential PET imaging agents for the in vivo visualization of the brain COX-2 enzyme. Five 11C-labeled COX-2 inhibitors, with different lipophilicities and moderate COX-2 inhibitory activity, were prepared by treatment of the corresponding O-desmethyl precursors with [11C]methyl triflate and purified by HPLC (radiochemical yields of 55-71%, radiochemical purity of >93%, and the specific activities of 22-331 GBq/μmol). In mice, radioactivity in the brain for all radiotracers was low, with very low brain-to-blood ratios. A clear inverse relationship was observed between brain uptake at 1 min postinjection and the lipophilicity (experimental log P?.?) of the studied 11C-radiotracers. Pretreatment of mice with cyclosporine A to block P-glycoproteins caused a significant increase in brain uptake of radioactivity following injection of the 11C-radiotracer compared to control. HPLC analysis showed that each radiotracer was rapidly metabolized, and a few metabolites, which were more polar than the original radiotracers, were found in both plasma and brain. No specific binding of the tracers towards the COX-2 enzyme in the brain was clearly revealed by in vivo blocking study. Further structural refinement of the tracer agent is necessary for better enhancement of brain uptake and for sufficient metabolic stability.     

A prodrug of NMDA/glycine site antagonist, L-703,717, with improved BBB permeability: 4-acetoxy derivative and its positron-emitter labeled analog

4-Acetoxy derivative (1) of L-703,717, a high-affinity (IC50=4.5 nM) antagonist for the glycine site of NMDA receptors, was synthesized and its brain uptake was examined using a carbon-11 labeled analog ([11C]1). Initial radioactivity in the brain after intravenous injection of [11C]1 was a 2-fold that of [11C]L-703,717 in mice. Rapid bioconversion of [11C]1 into [11C]L-703,717 was demonstrated by metabolite analyses of rat brain after [11C]1 injection. Ex vivo autoradiography of [11C]1 in rat brain showed the same cerebellar localization of radioactivity as [11C]L-703,717. These results indicate that 1 is a promising pharmacological tool as a prodrug of L-703,717 with improved BBB permeability.     

Synthesis of C-labeled imipramine and its biodistribution in mice: a potential tracer for positron emission tomography

A tricyclic antidepressant, C-labeled imipramine was synthesized by N-methylation of desipramine with 11CH3I to assist in the imaging of the human imipramine receptor by positron emission tomography. The radiochemical yield after purification of 11C-imipramine by high performance liquid chromatography was 28-63% at a specific activity of 26-53 Ci/mmol. The time required for synthesis, including purification was 30 min from the end of 11CH3I trapping. The organ distribution of 11C-imipramine was investigated in mice at various times after i.v. injection. The main accumulation of radioactivity was in the kidney, followed by the lung and the heart. In the brain, the radioactivity levels in the hypothalamus and striatum were the highest and remained constant, differentiating them from other portions of the brain. Furthermore, the result of a binding assay with 3H-labeled imipramine suggested that the regional distribution of 11C-imipramine in the same mouse brain correlated to that of the high affinity imipramine binding site.     

Synthesis of C-4 substituted nicotine derivatives via an N-acylpyridinium salt of (S)-nicotine

[reaction: see text] A variety of novel nicotine derivatives were prepared from (S)-nicotine via a two-step sequence. Addition of a cuprate reagent to an N-acylpyridinium salt of nicotine, followed by aromatization with elemental sulfur, afforded C-4 substituted nicotines in moderate to high yield. Using this method, 4-(dimethylphenylsilyl)nicotine was prepared and oxidized to afford (S)-4-hydroxynicotine.     

Synthesis of [N-methyl-3H]-7-chloro-1,3-dihydro-5-(2-fluorophenyl)-1-methyl-2 H-1,4-benzodiazepin-2- one (3H-fludiazepam) by phase transfer catalysis

[N-methyl-3H]-7-Chloro-1,3-dihydro-5-(2-fluorophenyl)-1-methyl-2H-1,4- benzodiazepin-2- one ( [3H]fludiazepam) was prepared by phase transfer catalytic N-methylation of the corresponding nor-derivative with [3H]methyl iodide [407 GBq/mmol (11 Ci/mmol)] in a radiochemical yield of 34%. The specific activity was 271.6 GBq/mmol (7.34 Ci/mmol).     

A novel strategy for the preparation of the injectable PET/CT radiopharmaceutical (-)-[11C]-(1R,2S)-meta-hydroxyephedrine ((-)-[11C]HED

Positron emission tomography (PET) had been applied in clinical early diagnosis of various tumors and other diseases. The methylated synthetic conditions of (-)-[¹¹C]-(1R,2S)-meta-hydroxyephedrine ((-)-[¹¹C]HED), considered as one of the most important radiopharmaceuticals for PET, were optimized through single factor and orthogonal design methods. Here, we reported an improved purification protocol. The radiochemical yields of the final product were over 45% (decay-corrected and based on [¹¹C]methyl iodide) (n?=?50). The radiochemical purities and chemical purities were over 99% (n?=?50) and 97% (n?=?50), respectively. The automatic radiosynthesis procedure of (-)-[¹¹C]HED with relatively high radiochemical yield was convenient and reliable.

     

N-[11C]methyl-1-(1,3-benzodioxol-5-yl)-2-butanamine (MBDB): synthesis, quality control and biodistribution

N-[¹¹C]methyl-1-(1,3-benzodioxol-5-yl)-2-butanamine ([¹¹C]MBDB) 3 was prepared by methylation of the demethyl precursor BDB with [¹¹C]CHI. The radiosynthesis was optimized with regard to temperature, reaction time and amount of precursor, best results (i.e., 84% radiochemical yield, based on [¹¹C]CH₃I activity) were obtained using 3 mg BDB at a reaction temperature of 130 °C in 8 minutes. With respect to a facilitated workup routine, productions were performed with 0.6 mg BDB at 110 °C for 10 minutes, yielding more than 50% of 3. The radiochemical purity of the final tracer solution was >98%, the specific activity was determined to be 300 GBq/mol (8000 Ci/mmol). Biodistribution, studies in rats showed two major metabolic pathways as indicated by an increasing liver uptake (9.1% ID/organ at 5 minutes to 21% ID/organ at 30 minutes) and a high urine activity (up to 16% ID/g). In brain tracer uptake was more than 1%, with a brain to blood ratio of almost 12 resulting from a very rapid blood clearance of 3.     

Radiosynthesis of N-11C-meisoindigotin as a novel PET agent for imaging of cyclin-dependent kinases and GSK-3β

Meisoindigotin has been demonstrated as a new type of cancer chemotherapeutic agent. N-11C-Meisoindigotin was synthesized by N-11C-methylation of the isoindigotin precursor with 11C-labelled methyl triflate. The decay corrected radiochemical yields were 15−25 % ,and the specific radioactivity was 1.0−1.2 Ci/?mol at the end of synthesis. The cellular uptake of [N-11C]meisoindigotin was evaluated in four different lung cancer cell lines. Our results showed that the A549, GLC-82, 95D cell lines exhibited higher uptake than 95C cell line after incubation for 60 min. N-11C-Meisoindigotin was a promising candidate for further development as a novel PET radiotracer for imaging of cyclin-dependent kinases (CDKs) and GSK-3β.     

Preliminary Assessment of the Anti-inflammatory Activity of New Structural Honokiol Analogs with a 4′-O-(2-Fluoroethyl) Moiety and the Potential of Their 18F-Labeled Derivatives for Neuroinflammation Imaging

Neolignans honokiol and 4′-O-methylhonokiol (MH) and their derivatives have pronounced anti-inflammatory activity, as evidenced by numerous pharmacological studies. Literature data suggested that cyclooxygenase type 2 (COX-2) may be a target for these compounds in vitro and in vivo. Recent studies of [¹¹C]MPbP (4′-[¹¹C]methoxy-5-propyl-1,1′-biphenyl-2-ol) biodistribution in LPS (lipopolysaccharide)-treated rats have confirmed the high potential of MH derivatives for imaging neuroinflammation. Here, we report the synthesis of four structural analogs of honokiol, of which 4′-(2-fluoroethoxy)-2-hydroxy-5-propyl-1, 1′-biphenyl (F-IV) was selected for labeling with fluorine-18 (T_1/2 = 109.8 min) due to its high anti-inflammatory activity confirmed by enzyme immunoassays (EIA) and neuromorphological studies. The high inhibitory potency of F-IV to COX-2 and its moderate lipophilicity and chemical stability are favorable factors for the preliminary evaluation of the radioligand [¹⁸F]F-IV in a rodent model of neuroinflammation. [¹⁸F]F-IV was prepared with good radiochemical yield and high molar activity and radiochemical purity by ¹⁸F-fluoroethylation of the precursor with Boc-protecting group (15) with [¹⁸F]2-fluoro-1-bromoethane ([¹⁸F]FEB). Ex vivo biodistribution studies revealed a small to moderate increase in radioligand uptake in the brain and peripheral organs of LPS-induced rats compared to control animals. Pretreatment with celecoxib resulted in significant blocking of radioactivity uptake in the brain (pons and medulla), heart, lungs, and kidneys, indicating that [¹⁸F]F-IV is likely to specifically bind to COX-2 in a rat model of neuroinflammation. However, in comparison with [¹¹C]MPbP, the new radioligand showed decreased brain uptake in LPS rats and high retention in the blood pool, which apparently could be explained by its high plasma protein binding. We believe that the structure of [¹⁸F]F-IV can be optimized by replacing the substituents in the biphenyl core to eliminate these disadvantages and develop new radioligands for imaging activated microglia.     

Synthesis of nicotine derivatives via reductive disilylation of (S)-nicotine

[reaction: see text] A variety of novel nicotine derivatives were prepared via reductive disilylation of (S)-nicotine. Treatment of nicotine with lithium powder and chlorotrimethylsilane afforded 1,4-bis(trimethylsilyl)-1,4-dihydronicotine in high yield. Addition of various carbonyl electrophiles and a catalytic amount of TBAF provided either C-5 substituted nicotines or 1,4-dihydronicotine derivatives.     

[11C]-labeling of some caffeine derivatives for mapping adenosine A2a receptors by PET technique

[¹¹C]-labeled form of ten A_2a adenosine receptor specific 8-styryl-7-methyl-xanthine derivatives ([¹¹C]-caffeines) were synthesised by N-methylation of the corresponding 8-styryl-xanthine derivatives using [¹¹C]-methyl iodide in optimized reaction conditions. The results show that the [¹¹C]-methylations take place with excellent radiochemical yields (35–93%), and can be utilised easily in online preparations. These labeled ligands may facilitate the positron emission tomographic (PET) investigation of adenosine A_2a receptors.     

Radical-mediated carboxylation of alkyl iodides with [11C]carbon monoxide in solvent mixtures

[reaction: see text] [carboxyl-(11)C]Carboxylic acids were prepared from alkyl iodides via photoinitiated radical reactions using 10(-)(8) mol of [(11)C]carbon monoxide in binary and ternary homogeneous solvent mixtures. Short- (isobutyric), medium-, and long-chain saturated fatty acids (heptadecanoic) were labeled with isolated decay-corrected radiochemical yields ranging from 55% to 70% in 5-7-min reactions. The conversion of [(11)C]carbon monoxide to products reached 80-90%. To obtain good yields in the reactions performed in water-acetonitrile and water-THF mixtures, the addition of tetrabutylammonium hydroxide or potassium hydroxide was essential. The carboxylation was efficient for primary and secondary alkyl iodides. The carboxylation of tertiary iodides was feasible for 1-iodoadamantane but not for tert-butyl iodide. The dependence of the radiochemical yields on reaction time, photoirradiation conditions, and organic and inorganic additives was studied. The method provides a one-step route to [carboxyl-(11)C]carboxylic acids; traditional methods, in contrast, would require several steps. For example, using the devised reaction conditions, 3.19 GBq of purified [1-(11)C]1,10-decanedicarboxylic acid (specific radioactivity 188 GBq/mumol) was obtained within 35 min of the end of 10 muAh bombardment. (1-(13)C)4-Phenylbutyric acid was synthesized using ((13)C)carbon monoxide for identifying the labeling position with (1)H and (13)C NMR.     

The synthesis of a new cardiac sympathetic nerve imaging agent N-[11C]CH3-dopamine and biodistribution study

In this study, we synthesized and characterized N-[¹¹C]methyl-dopamine ([¹¹C]MDA) for cardiac sympathetic nerve imaging. [¹¹C]MDA was synthesized by direct N-methylation of dopamine with [¹¹C]methyl iodide and purified by semi-preparation reverse high pressure liquid chromatography (HPLC). The total synthesis time was 45 min including HPLC purification. The radiochemical yields of [¹¹C]MDA was 20 ± 3 %, without decay correction. The radiochemical purity was >98 % and the specific activity was about 50 GBq/mmol. The biological properties of [¹¹C]MDA were evaluated by biodistribution study in normal mice. PET imaging was performed in healthy Chinese mini-swines. Biodistribution study showed that [¹¹C]MDA had high myocardium uptake. PET/CT imaging showed [¹¹C]MDA had clear and symmetrical myocardium uptake, which was blocked obviously by injecting imipramine hydrochloride. [¹¹C]MDA would be a promising candidate of radiotracer for cardiac sympathetic nervous system imaging.     

Six-step synthesis of (S)-brevicolline from (S)-nicotine

[reaction: see text] A six-step synthesis of (S)-brevicolline from (S)-nicotine is reported. Regioselective trisubstitution of the pyridine ring of nicotine, followed by successive Suzuki cross-coupling and Buchwald amination reactions, afforded the enantiopure beta-carboline alkaloid, brevicolline.     

(11)C] Carbon monoxide in selenium-mediated synthesis of (11)C-carbamoyl compounds

Using either amines, amino alcohols, or alcohols in selenium-mediated synthesis with [(11)C]carbon monoxide, 3 ureas, 6 carbamates, and 1 carbonate were labeled. Tetrabutylammonium fluoride ((TBA)F) was discovered to form a soluble and reactive complex with selenium and drastically increase the radiochemical yields. Of the selected carbamoyl compounds, one was a receptor ligand, one was an enzyme inhibitor, and one was a muscular relaxant pharmaceutical. The (11)C-target compounds were obtained in radiochemical yields ranging from low to almost quantitative and with specific radioactivity up to 1300 GBq/micromol. The radiochemical purity of the final products exceeded 98%. In one case, the corresponding (13)C-substituted compound was produced to verify the position of the (11)C-label. In a typical experiment starting with 16.4 GBq [(11)C]carbon monoxide, 7.0 GBq of LC-purified 5-phenyl-1,3-oxazolidin-[2-(11)C]-2-one was obtained within 20 min from start of the carbonylation reaction (84% decay-corrected radiochemical yield). The presented approach is an interesting alternative to the use of [(11)C]phosgene in labeling chemistry.     

Synthesis of [125I]iodoclorgyline, a selective monoamine oxidase A inhibitor, and its biodistribution in mice.

A new radioiodinated monoamine oxidase A (MAO-A) specific inhibitor, [125I]iodoclorgyline, was synthesized from its tin precursor by iododestannylation reaction using sodium [125I]iodide and hydrogen peroxide with high yield and site specificity. The product possessed a high radiochemical purity as well as high specific activity. The method can be readily applicable for labeling with 123I, a very suitable radioisotope for in vivo imaging with single photon emission computer tomography (SPECT). Biodistribution studies of the [125I]iodoclorgyline in mice showed high initial uptake in the brain, and brain radioactivity reached a constant level at 60 min after intravenous injection. The results suggested that [125I]iodoclorgyline might have potential as a radiopharmaceutical for MAO-A studies in the brain with SPECT.     

Synthesis of [11C]/(13C)amines via carbonylation followed by reductive amination

Twelve 11C-labelled amines were prepared via 11C-carbonylation followed by reductive amination. The 11C-carbonylation was performed in the presence of tetrakis(triphenylphosphine)palladium using aryl iodides or aryl triflates, [11C]carbon monoxide and phenyl-/methylboronic acid. The [11C]ketones formed in this step were then transformed directly into amines by reductive amination using different amines in the presence of TiCl4 and NaBH3CN. The 11C-labelled amines were obtained with decay-corrected radiochemical yields in the range 2-78%. The radiochemical purity of the isolated products exceeded 98%. (13C)Benzhydryl-phenyl-amine was synthesised and analysed by NMR spectroscopy for confirmation of the labelling position. Specific radioactivity was determined for the same compound. The reference compounds were prepared by reductive amination of ketones using conventional reaction conditions and three of the compounds were novel. The presented approach is a new method for the synthesis of [11C]/(13C)amines.     

Efficient synthesis of [¹¹C]ramelteon as a positron emission tomography probe for imaging melatonin receptors involved in circadian rhythms

Ramelteon (TAK-375) is a novel melatonin receptor agonist that is used for clinical treatment of insomnia. The present report describes radiolabeling of ramelteon with the short-lived positron-emitter 11C (T(1/2)=20.4 min) by 2 methods. One method was [11C]methylation of an acetoamide precursor and the other was [11C]acylation of the corresponding amine precursor. First, [11C]methylation method showed the low reproducibility together with the production of many kinds of side products from which the [11C-methyl]Ramelteon was separated with chemical purity of <28% and radiochemical purity of >98%. Whereas, the [11C]acylation method showed high efficiency and reproducibility with a good radiochemical yield (22-43%, decay corrected), high chemical and radiochemical purities (>99% each), and high specific activity (43-162 GBq/μmol) (n=5) after HPLC purification. [11C]Ramelteon is a potential positron emission tomography (PET) probe for imaging the melatonin receptor.     

Microfluidic reactions using [11C]carbon monoxide solutions for the synthesis of a positron emission tomography radiotracer

Microfluidic technology has been used to perform [(11)C]carbonylation reactions using solutions containing [(11)C]CO in the form of the complex, copper(i)tris(3,5-dimethylpyrazolyl)borate-[(11)C]carbonyl (Cu(Tp*)[(11)C]CO). The synthesis of the model compound [(11)C]N-benzylbenzamide and the known tracer molecule [(11)C]trans-N-[5-(2-flurophenyl)-2-pyrimidinyl]-3-oxospiro[5-azaisobenzofurane-1(3H),1'-cyclohexane]-4'-carboxamide ([(11)C]MK-0233), a ligand for the neuropeptide Y Y5 receptor, have been performed using this technique. Following semi-preparative HPLC purification and reformulation, 1262 ± 113 MBq of [(11)C]MK-0233 was produced at the end of the synthesis with a specific activity of 100 ± 30 GBq μmol(-1) and a >99% radiochemical purity. This corresponds to a decay corrected radiochemical yield of 7.2 ± 0.7%. Using a 3 mL vial as the reaction vessel, and following semi-preparative HPLC purification and reformulation, 1255 ± 392 MBq of [(11)C]MK-0233 was produced at the end of the synthesis with a specific activity of 100 ± 15 GBq μmol(-1) and a >99% radiochemical purity. This corresponds to a decay corrected radiochemical yield of 7.1 ± 2.2%.     

Synthesis of O-(2-[~(18)F]fluoroethyl)-L-tyrosine and its biological evaluation in B16 melanoma-bearing mice as PET tracer for tumor imaging

O-(2-[18F]fluoroethyl) -L-tyrosine([18F]FET) ,a fluorine-18 labeled analogue of tyrosine,has been syn-thesized and biologically evaluated in tumor-bearing mice. The whole synthesis procedure is com-pleted within 50 min. The radiochemical yield is about 40%(no decay corrected) and radiochemical purity more than 97% after simplified solid phase extraction. [18F]FET shows rapid,high uptake and long retention in the tumor as well as low uptake in the brain. The ratios of tumor-to-muscle(T/M) and tumor-to-blood(T/B) of [18F]FET are similar to those of [18F]FDG,but the ratios of tumor-to-brain(T/Br) are 2-3 times higher than that of [18F]FDG. Autoradiography of [18F]FET demonstrates a remarkable accumulation in melanoma with high contrast. It appears to be a probable competitive candidate for melanoma imaging with PET.