Kinetic study of benzyl [1-14C]acetate as a potential probe for astrocytic energy metabolism in the rat brain: Comparison with benzyl [2-14C]acetate.

Brain uptake of [(14)C]acetate has been reported to be a useful marker of astrocytic energy metabolism. In addition to uptake values, the rate of radiolabeled acetate washout from the brain appears to reflect CO2 exhaustion and oxygen consumption in astrocytes. We measured the time-radioactivity curves of benzyl [1-(14)C]acetate ([1-(14)C]BA), a lipophilic probe of [1-(14)C]acetate, and compared it with that of benzyl [2-(14)C]acetate ([2-(14)C]BA) in rat brains. The highest brain uptake was observed immediately after injecting either [1-(14)C]BA or [2-(14)C]BA, and both subsequently disappeared from the brain in a single-exponential manner. Estimated [1-(14)C]BA washout rates in the cerebral cortex and cerebellum were higher than those of [2-(14)C]BA. These results suggested that [1-(14)C]BA could be a useful probe for estimating the astrocytic oxidative metabolism. The [1-(14)C]BA washout rate in the cerebral cortex of immature rats was lower than that of mature rats. An autoradiographic study showed that the washout rates of [1-(14)C]BA from the rat brains of a lithium-pilocarpine-induced status epilepticus model were not significantly different from the values in control rat brains except for the medial septal nucleus. These results implied that the enhancement of amino acid turnover rate rather than astrocytic oxidative metabolism was increased in status epilepticus.

Improvement of brain uptake for in vivo PET imaging of astrocytic oxidative metabolism using benzyl [1-(11)C]acetate.

Brain uptake of acetate is insufficient for obtaining a quantitative image of astrocytic oxidative metabolism. To improve the brain uptake of [1-(11)C]acetate, we synthesized benzyl [1-(11)C]acetate ([1-(11)C]BA) and conducted a positron emission tomography (PET) study assessing astrocytic oxidative metabolism. The brain uptake of [1-(11)C]BA was markedly higher compared with [1-(11)C]acetate, and disappeared with a half-life of 20 min in all regions studied. The brain uptake of [1-(11)C]BA was significantly decreased by fluorocitrate. The results indicate that [1-(11)C]BA could be a useful PET probe for assessing astrocytic oxidative metabolism.

Synthesis and evaluation of PET probes for the imaging of I2 imidazoline receptors in peripheral tissues.

INTRODUCTION: To explore the possible use of positron emission tomography (PET) probes for imaging of I(2)-imidazoline receptors (I(2)Rs) in peripheral tissues, we labeled two new I(2)R ligands, 2-[2-(o-tolyl)vinyl]-4,5-dihydro-1H-imidazole (K(i) for I(2)Rs, 3.7 nM) and 2-[2-(o-tolyl)ethyl]-4,5-dihydro-1H-imidazole (K(i) for I(2)Rs, 1.7 nM) with (11)C ([(11)C]metrazoline and [(11)C]TEIMD), respectively, and evaluated these ligands and the recently developed I(2)R ligand 2-[3-fluoro-[4-(11)C]tolyl]-4,5-dihydro-1H-imidazole ([(11)C]FTIMD) by in vivo studies. METHODS: [(11)C]Metrazoline and [(11)C]TEIMD were prepared by a palladium-promoted cross-coupling reaction of the tributylstannyl precursor and [(11)C]methyl iodide. Their biodistribution in mice was investigated by tissue dissection. In addition, PET scans and metabolite analysis were performed. RESULTS: [(11)C]Metrazoline and [(11)C]TEIMD were successfully synthesized with a suitable radioactivity for injection. In the liver and pancreas expressing I(2)Rs, coinjection with the high-affinity I(2)R ligand, BU224, induced a reduction in the radioactivity level at 30 min after injection of [(11)C]metrazoline and [(11)C]FTIMD. However, the radioactivity level after injection of [(11)C]TEIMD was unchanged. In the PET study, coinjection with BU224 induced a decrease in the radioactivity level in the liver and pancreas after more than 15 min of injection of [(11)C]metrazoline and [(11)C]FTIMD as compared with the results obtained for controls. In metabolite analysis, coinjection with BU224 induced a significant reduction in the percentage of unchanged [(11)C]metrazoline at 30 min after injection as compared with that in the control, although no significant difference was observed in the percentage of unchanged [(11)C]FTIMD. CONCLUSION: [(11)C]Metrazoline may be a more useful PET probe than [(11)C]FTIMD for imaging of I(2)Rs in peripheral tissues.

Evaluation of the P-glycoprotein- and breast cancer resistance protein-mediated brain penetration of 11C-labeled topotecan using small-animal positron emission tomography.

INTRODUCTION: Topotecan (TPT) is a camptothecin derivative and is an anticancer drug working as a topoisomerase-I-specific inhibitor. But TPT cannot penetrate through the blood-brain barrier. In this study, we synthesized a new positron emission tomography (PET) probe, [(11)C]TPT, to evaluate the P-glycoprotein (Pgp)- and breast cancer resistance protein (BCRP)-mediated brain penetration of [(11)C]TPT using small-animal PET. METHODS: [(11)C]TPT was synthesized by the reaction of a desmethyl precursor with [(11)C]CH(3)I. In vitro study using [(11)C]TPT was carried out in MES-SA and doxorubicin-resistant MES-SA/Dx5 cells in the presence or absence of elacridar, a specific inhibitor for Pgp and BCRP. The biodistribution of [(11)C]TPT was determined using small-animal PET and the dissection method in mice. RESULTS: The transport of [(11)C]TPT to the extracellular side was determined in MES-SA/Dx5 cells exhibiting the expressions of Pgp and BCRP at high levels. This transport was inhibited by coincubation with elacridar. In Mdr1a/b(-/-)Bcrp1(-/-) mice, PET results indicated that the brain uptake of [(11)C]TPT was about two times higher than that in wild-type mice. Similarly, the brain penetration of [(11)C]TPT in wild-type mice was increased by treatment with elacridar. The radioactivity in the brain of elacridar-treated mice was maintained at a certain level after the injection of [(11)C]TPT, although the radioactivity in the blood decreased with time. CONCLUSIONS: We demonstrated the increase of brain penetration of [(11)C]TPT by deficiency and inhibition of Pgp and BCRP functions using small-animal PET in mice.

Synthesis and evaluation of novel carbon-11 labeled oxopurine analogues for positron emission tomography imaging of translocator protein (18 kDa) in peripheral organs.

To develop a PET ligand for imaging TSPO in peripheral organs, we designed three novel oxopurine analogues [(11)C]3a-c (LogD: 1.81-2.17) by introducing a pyridine ring in place of a benzene ring in the lead compound [(11)C]2 (LogD: 3.48). The desmethyl precursors 10 for radiosynthesis were synthesized by reacting glycine 7 with picolylamines 4, followed by hydrolysis and by Curtius rearrangement with diphenylphosphoryl azide. Methylation of 10a-c with methyl iodide produced unlabeled compounds 3a-c. The radiosynthesis of [(11)C]3a-c was performed by reacting 10a-c with [(11)C]methyl iodide. Compounds 3a-c displayed high or moderate in vitro binding affinities (K(i): 5-40 nM) for TSPO. PET with [(11)C]3a-c in rats showed high uptake in the lung, heart, and kidney, which are organs with high TSPO expression. Metabolite analysis with [(11)C]3a showed that radioactivity in these organs mainly corresponded with unchanged [(11)C]3a. PET with [(11)C]3a using a rat model of lung inflammation showed a significant signal in the lipopolysaccharide-treated lung.

Evaluation of chemotherapy response in VX2 rabbit lung cancer with 18F-labeled C2A domain of synaptotagmin I.

UNLABELLED: The C2A domain of synaptotagmin I can target apoptotic cells by binding to exposed anionic phospholipids. The goal of this study was to synthesize and develop (18)F-labeled C2A-glutathione-S-transferase (GST) as a molecular imaging probe for the detection of apoptosis and to assess the response of paclitaxel chemotherapy in VX2 rabbit lung cancer. METHODS: (18)F-C2A-GST was prepared by labeling C2A-GST with N-succinimidyl 4-(18)F-fluorobenzoate ((18)F-SFB). (18)F-C2A-GST was confirmed by high-performance liquid chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. The binding of (18)F-C2A-GST toward apoptosis was validated in vitro using camptothecin-induced Jurkat cells. Biodistribution of (18)F-C2A-GST was determined in mice by a dissection method and small-animal PET. Single-dose paclitaxel was used to induce apoptosis in rabbits bearing VX2 tumors (n = 6), and 2 VX2 rabbits without treatment served as control. (18)F-C2A-GST PET was performed before and at 72 h after therapy, and (18)F-FDG PET/CT was also performed before treatment. To confirm the presence of apoptosis, tumor tissue was analyzed and activated caspase-3 was measured. RESULTS: (18)F-C2A-GST was obtained with more than 95% radiochemical purity and was stable for 4 h after formulation. (18)F-C2A-GST bound apoptotic cells specifically. Biodistribution in mice showed that (18)F-C2A-GST mainly excreted from the kidneys and rapidly cleared from blood and nonspecific organs. High focal uptake of (18)F-C2A-GST in the tumor area was determined after therapy, whereas no significant uptake before therapy was found in the tumor with (18)F-FDG-avid foci. The maximum standardized uptake value after therapy was 0.47 ± 0.28, significantly higher than that in the control (0.009 ± 0.001; P < 0.001). The apoptotic index was 79.81% ± 8.73% in the therapy group, significantly higher than that in the control (5.03% ± 0.81%; P < 0.001). Activated caspase-3 after paclitaxel treatment increased to 69.55% ± 16.27% and was significantly higher than that in the control (12.26% ± 5.39%; P < 0.001). CONCLUSION: (18)F-C2A-GST was easily synthesized by conjugation with (18)F-SFB and manifested a favorable biodistribution. Our results demonstrated the feasibility of (18)F-C2A-GST for the early detection of apoptosis after chemotherapy in a VX2 lung cancer model that could imitate the human lung cancer initiation, development, and progress.

Determination of radioactivity in infant, juvenile and adult rat brains after injection of anti-influenza drug [¹¹C]oseltamivir using PET and autoradiography.

Oseltamivir phosphate (Tamiflu(®)) is an orally active anti-influenza drug, which is hydrolyzed to its metabolite Ro 64-0802 inhibiting the influenza virus with potent activity. The abnormal behavior of young influenza patients associated with the use of oseltamivir has developed to a social problem in countries where Tamiflu is often prescribed. It is important to determine the amount of oseltamivir in the brain and to elucidate the relationship between its presence and neuropsychiatric side effects. The aim of this study was to determine the radioactivity in the infant, juvenile and adult rat brains after injection of [(11)C]oseltamivir into the rats using PET and autoradiography. After injection of this radioligand, the highest radioactivity was found in the infant brain and the radioactivity level decreased with age. Ex vivo autoradiography on the infant brain displayed a relatively higher radioactivity in the cerebellum than that in the cerebrum. Pretreatment with cyclosporin A (an inhibitor for P-glycoprotein) increased the brain radioactivity. These results give helpful insights into elucidating why the neuropsychiatric side effects of oseltamivir occur in young patients.

Visualization of early infarction in rat brain after ischemia using a translocator protein (18 kDa) PET ligand [11C]DAC with ultra-high specific activity.

The aim of this study was to visualize early infarction in the rat brain after ischemia using a translocator protein (TSPO) (18 kDa) PET ligand [(11)C]DAC with ultra-high specific activity (SA) of 3670-4450 GBq/µmol. An infarction model of rat brain was prepared by ischemic surgery and evaluated 2 days after ischemia using small-animal PET and in vitro autoradiography. Early infarction with a small increase of TSPO expression in the brain was visualized using PET with high SA [(11)C]DAC (average 4060 GBq/µmol), but was not distinguished clearly with usually reported SA [(11)C]DAC (37 GBq/µmol). Infarction in the rat brain 4 days after ischemia was visualized using high and usually reported SAs [(11)C]DAC. Displacement experiments with unlabeled TSPO-selective AC-5216 or PK11195 diminished the difference in radioactivity between ipsilateral and contralateral sides, confirming that the increased uptake on the infracted brain was specific to TSPO. In vitro autoradiography with high SA [(11)C]DAC showed that the TSPO expression increased on early infarction in the rat brain. High SA [(11)C]DAC is a useful and sensitive biomarker for the visualization of early infarction and the characterization of TSPO expression which was slightly elevated in the infarcted brain using PET.

Evaluation of limiting brain penetration related to P-glycoprotein and breast cancer resistance protein using [(11)C]GF120918 by PET in mice.

PURPOSE: GF120918 has a high inhibitory effect on P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). We developed [(11)C]GF120918 as a positron emission tomography (PET) probe to assess if dual modulation of P-gp and BCRP is useful to evaluate brain penetration. PROCEDURES: PET studies using [(11)C]GF120918 were conducted on P-gp and/or Bcrp knockout mice as well as wild-type mice. RESULTS: In PET studies, the AUC(brain) ([0-60 min]) and K (1) value in P-gp/Bcrp knockout mice were nine- and 26-fold higher than that in wild-type mice, respectively. These results suggest that brain penetration of [(11)C]GF120918 is related to modulation of P-gp and BCRP and is limited by two transporters working together. CONCLUSIONS: PET using [(11)C]GF120918 may be useful for evaluating the function of P-gp and BCRP. PET using P-gp/Bcrp knockout mice may be an effective method to understand the overall contributions the functions of P-gp and BCRP.

Synthesis and in vivo evaluation of ¹8F-fluoroethyl GF120918 and XR9576 as positron emission tomography probes for assessing the function of drug efflux transporters.

The purpose of this study was to synthesize two new positron emission tomography (PET) probes, N-(4-(2-(1,2,3,4-tetrahydro-6,7-dimethoxy-2-isoquinolinyl)ethyl)phenyl)-9,10-dihydro-5-[¹8F]fluoroethoxy-9-oxo-4-acridine carboxamide ([¹8F]3) and quinoline-3-carboxylic acid [2-(4-{2-[7-(2-[¹8F]fluoroethoxy)-6-methoxy-3,4-dihydro-1H-isoquinolin-2-yl]ethyl}phenylcarbamoyl)-4,5-dimethoxyphenyl]amide ([¹8F]4), and to evaluate the potential of these PET probes for assessing the function of two major drug efflux transporters, P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). [¹8F]3 and [¹8F]4 were synthesized by ¹8F-alkylation of each O-desmethyl precursor with [¹8F]2-fluoroethyl bromide for injection as PET probes. In vitro accumulation assay showed that treatment with P-gp/BCRP inhibitors (1 and 2) enhanced the intracellular accumulation capacity of P-gp- and BCRP-overexpressing MES-SA/Dx5 cells. In PET studies, the uptake (AUC(brain[0-)60 (min])) of [¹8F]3 and [¹8F]4 in wild-type mice co-injected with 1 were approximately sevenfold higher than that in wild-type mice, and the uptake of [¹8F]3 and [¹8F]4 in P-gp/Bcrp knockout mice were eight- to ninefold higher than that in wild-type mice. The increased uptake of [¹8F]3 and [¹8F]4 was similar to that of parent compounds ([¹¹C]1 and [¹¹C]2) previously described, indicating that radioactivity levels in the brain after injection of [¹8F]3 and [¹8F]4 are related to the function of drug efflux transporters. Also, these results suggest that the structural difference between parent compounds ([¹¹C]1 and [¹¹C]2) and fluoroethyl analogs ([¹8F]3 and [¹8F]4) do not obviously affect the potency against drug efflux transporters. In metabolite analysis of mice, the unchanged form in the brain and plasma at 60 min after co-injection of [¹8F]4 plus 1 were higher (95% for brain; 81% for plasma) than that after co-injection of [¹8F]3 plus 1. [¹8F]4 is a promising PET probe to assess the function of drug efflux transporters.

Synthesis and evaluation of 6-[1-(2-[(18)F]fluoro-3-pyridyl)-5-methyl-1H-1,2,3-triazol-4-yl]quinoline for positron emission tomography imaging of the metabotropic glutamate receptor type 1 in brain.

The purpose of this study was to synthesize 6-[1-(2-[(18)F]fluoro-3-pyridyl)-5-methyl-1H-1,2,3-triazol-4-yl]quinoline ([(18)F]FPTQ, [(18)F]7a) and to evaluate its potential as a positron emission tomography ligand for imaging metabotropic glutamate receptor type 1 (mGluR1) in the rat brain. Compound [(18)F]7a was synthesized by [(18)F]fluorination of 6-[1-(2-bromo-3-pyridyl)-5-methyl-1H-1,2,3-triazol-4-yl]quinoline (7b) with potassium [(18)F]fluoride. At the end of synthesis, 1280-1830MBq (n=8) of [(18)F]7a was obtained with >98% radiochemical purity and 118-237GBq/µmol specific activity using 3300-4000MBq of [(18)F]F(-). In vitro autoradiography showed that [(18)F]7a had high specific binding with mGluR1 in the rat brain. Biodistribution study using a dissection method and small-animal PET showed that [(18)F]7a had high uptake in the rat brain. The uptake of radioactivity in the cerebellum was reduced by unlabeled 7a and mGluR1-selective ligand JNJ-16259685 (2), indicating that [(18)F]7a had in vivo specific binding with mGluR1. Because of a low amount of radiolabeled metabolite present in the brain, [(18)F]7a may have a limiting potential for the in vivo imaging of mGluR1 by PET.

PET study on mice bearing human colon adenocarcinoma cells using [11C]GF120918, a dual radioligand for P-glycoprotein and breast cancer resistance protein.

OBJECTIVE: To evaluate the functions of P-glycoprotein (Pgp) and breast cancer resistance protein (BCRP) in human colon adenocarcinoma (Caco-2), we carried out an in-vitro study and a small animal positron emission tomography (PET) study using [C]GF120918 (elacridar). METHODS: [C]GF120918 was synthesized by reacting the desmethyl precursor with [C]CH3I. An in-vitro study using [C]GF120918 was carried out in Caco-2 and Madin-Darby canine kidney cells in the presence or absence of a transporter inhibitor (cyclosporine A and unlabeled GF120918). The biodistribution of radioactivity after the injection of [C]GF120918 was determined in Caco-2-bearing mice using a small animal PET scanner. RESULTS: In Caco-2 cells expressing Pgp and BCRP, coincubation with unlabeled GF120918 caused an approximately two-fold increase in [C]GF120918 uptake compared with that of the control ([C]GF120918 only). In Caco-2-bearing mice, PET results indicated that [C]GF120918 uptake in the tumor was low, but was significantly increased by treatment with unlabeled GF120918. In metabolite analysis, the radioactive component in the tumor almost corresponded to intact [C]GF120918. CONCLUSION: A PET study combining the administration of [C]GF120918 with unlabeled GF120918 may be a useful tool for evaluating the functions of Pgp and BCRP in tumors.

In vivo imaging and quantitative analysis of TSPO in rat peripheral tissues using small-animal PET with [18F]FEDAC.

INTRODUCTION: The translocator protein (18 kDa) (TSPO) is widely expressed in peripheral tissues, including the heart, lung, and kidney. Our laboratory developed N-benzyl-N-methyl-2-[7,8-dihydro-7-(2-[(18)F]fluoroethyl)-8-oxo-2-phenyl-9H-purin-9-yl]acetamide ([(18)F]FEDAC) as a TSPO positron emission tomography (PET) ligand. Here, using small-animal PET with [(18)F]FEDAC, we performed TSPO imaging and quantitative analysis of TSPO binding in rat peripheral tissues. METHODS: The in vivo distribution and kinetics of [(18)F]FEDAC were measured in rat peripheral tissues (heart, lung and kidney). Using the in vivo pseudo-equilibrium method, TSPO binding parameters [TSPO density (B(max)), dissociation constant (K(D))] and receptor occupancy were estimated in these peripheral tissues. RESULTS: [(18)F]FEDAC was highly distributed in the lung, heart and kidney, and these TSPO-enriched tissues could be clearly visualized. The kinetics of this radioligand in these tissues was rapid, which is suitable for the determination of in vivo TSPO binding parameters and receptor occupancy. The B(max) value of TSPO in the heart, lung, and kidney was 393, 141, and 158 pmol/ml, respectively. The K(D) value of the radioligand in the heart, lung, and kidney was 119, 36 and 123 nM, respectively. By pretreatment with 5 mg/kg Ro 5-4864 (a TSPO ligand), about 90% of binding sites for TSPO in the heart and lung were occupied. In the kidney, the binding sites were completely occupied by 5 mg/kg Ro 5-4864. CONCLUSIONS: [(18)F]FEDAC is a suitable PET ligand for TSPO imaging and quantitative analysis of TSPO binding in rat peripheral tissues. The utilization of [(18)F]FEDAC-PET and the pseudo-equilibrium method can contribute to the study of the TSPO function and evaluate the in vivo binding parameters and receptor occupancy of TSPO therapeutic compounds.

18F-FEAC and 18F-FEDAC: PET of the monkey brain and imaging of translocator protein (18 kDa) in the infarcted rat brain.

UNLABELLED: We evaluated two (18)F-labeled PET ligands, N-benzyl-N-ethyl-2-[7,8-dihydro-7-(2-(18)F-fluoroethyl)-8-oxo-2-phenyl-9H-purin-9-yl]acetamide ((18)F-FEAC) and N-benzyl-N-methyl-2-[7,8-dihydro-7-(2-(18)F-fluoroethyl)-8-oxo-2-phenyl-9H-purin-9-yl]acetamide ((18)F-FEDAC), by investigating their kinetics in the monkey brain and by performing in vitro and in vivo imaging of translocator protein (18 kDa) (TSPO) in the infarcted rat brain. METHODS: Dissection was used to determine the distribution of (18)F-FEAC and (18)F-FEDAC in mice, whereas PET was used for a monkey. With each (18)F-ligand, in vitro autoradiography and small-animal PET were performed on infarcted rat brains. RESULTS: (18)F-FEAC and (18)F-FEDAC had a high uptake of radioactivity in the heart, lung, and other TSPO-rich organs of mice. In vitro autoradiography showed that the binding of each (18)F-ligand significantly increased on the ipsilateral side of rat brains, compared with the contralateral side. In a small-animal PET study, PET summation images showed the contrast of radioactivity between ipsilateral and contralateral sides. Pretreatment with TSPO ligands N-benzyl-N-ethyl-2-(7-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide (AC-5216) or (R)-N-methyl-N-(1-methylpropyl)-1-(2-chlorophenyl)isoquinoline-3-carboxamide (PK11195) diminished the difference in uptake between the 2 sides. The PET study showed that each (18)F-ligand had uptake and distribution patterns in the monkey brain similar to those of (11)C-AC-5216. After injection into the monkey during PET, the uptake of each (18)F-ligand in the brain decreased over time whereas (11)C-AC-5216 did not. In the brain homogenate of mice, the percentage of the fraction corresponding to intact (18)F-FEAC and (18)F-FEDAC was 68% and 75% at 30 min after injection. In monkey plasma, each (18)F-ligand was scarcely metabolized until the end of the PET scan. CONCLUSION: (18)F-FEAC and (18)F-FEDAC produced in vitro and in vivo signals allowing visualization of the increase in TSPO expression in the infarcted rat brain. The kinetics of both (18)F-ligands in the monkey brain and tolerance for in vivo metabolism suggested their usefulness for imaging studies of TSPO in primates.

Radiosynthesis and evaluation of [11C]YM-202074 as a PET ligand for imaging the metabotropic glutamate receptor type 1.

INTRODUCTION: Developing positron emission tomography (PET) ligands for imaging metabotropic glutamate receptor type 1 (mGluR1) is important for studying its role in the central nervous system. N-cyclohexyl-6-{[N-(2-methoxyethyl)-N-methylamino]methyl}-N-methylthiazolo[3,2-a]benzimidazole-2-carboxamide (YM-202074) exhibited high binding affinity for mGluR1 (K(i)=4.8 nM), and selectivity over other mGluRs in vitro. The purpose of this study was to label YM-202074 with carbon-11 and to evaluate in vitro and in vivo characteristics of [(11)C]YM-202074 as a PET ligand for mGluR1 in rodents. METHODS: [(11)C]YM-202074 was synthesized by N-[(11)C]methylation of its desmethyl precursor with [(11)C]methyl iodide. The in vitro and in vivo brain regional distributions were determined in rats using autoradiography and PET, respectively. RESULTS: [(11)C]YM-202074 (262-630 MBq, n=5) was obtained with radiochemical purity of >98% and specific activity of 27-52 GBq/mumol at the end of synthesis, starting from [(11)C]CO(2) of 19.3-21.5 GBq. In vitro autoradiographic results showed that the high specific binding of [(11)C]YM-202074 for mGluR1 was presented in the cerebellum, thalamus and hippocampus, which are known as mGluR1-rich regions. In ex vivo autoradiography and PET studies, the radioligand was specifically distributed in the cerebellum, although the uptake was low. Furthermore, the regional distribution was fairly uniform in the whole brain by pretreatment with JNJ16259685 (a mGluR1 antagonist). However, radiometabolite(s) was detected in the brain. CONCLUSIONS: From these results, especially considering the low brain uptake and the influx of radiometabolite(s) into brain, [(11)C]YM-202074 may not be a useful PET ligand for in vivo imaging of mGluR1 in the brain.

Imaging of I2-imidazoline receptors by small-animal PET using 2-(3-fluoro-[4-11C]tolyl)-4,5-dihydro-1H-imidazole ([11C]FTIMD).

INTRODUCTION: Imidazoline receptors (IRs) have been established as distinct receptors, and have been categorized into at least two subtypes (I(1)R and I(2)R). I(2)Rs are associated with depression, Alzheimer's disease, Huntington's disease and Parkinson's disease. A few positron emission tomography (PET) probes for I(2)Rs have been synthesized, but a selective PET probe has not been evaluated for the imaging of I(2)Rs by PET. We labeled a selective I(2)R ligand 2-(3-fluoro-4-tolyl)-4,5-dihydro-1H-imidazole (FTIMD) with (11)C and performed the first imaging of I(2)Rs by PET using 2-(3-fluoro-[4-(11)C]tolyl)-4,5-dihydro-1H-imidazole ([(11)C]FTIMD). METHODS: [(11)C]FTIMD was prepared by a palladium-promoted cross-coupling reaction of the tributylstannyl precursor and [(11)C]methyl iodide in the presence of tris(dibenzylideneacetone)dipalladium(0) and tri(o-tol)phosphine. Biodistribution was investigated in rats by tissue dissection. [(11)C]FTIMD metabolites were measured in brain tissues and plasma. Dynamic PET scans were acquired in rats, and the kinetic parameters estimated. RESULTS: [(11)C]FTIMD was successfully synthesized with a suitable radioactivity for the injection. Co-injection with 0.1 mg/kg of cold FTIMD and BU224 induced a significant reduction in the brain-to-blood ratio 15 and 30 min after the injection. In metabolite analysis, unchanged [(11)C]FTIMD in the brain was high (98%) 30 min after the injection. In PET studies, high radioactivity levels were observed in regions with a high density of I(2)R. The radioactivity levels and V(T) values in the brain regions were prominently reduced by 1.0 mg/kg of BU224 pretreatment as compared with control. CONCLUSION: [(11)C]FTIMD showed specific binding to I(2)Rs in rat brains with a high density of I(2)R.

Synthesis and evaluation of [11C]XR9576 to assess the function of drug efflux transporters using PET.

OBJECTIVE: XR9576 (tariquidar) is an anthranilic acid derivative and potent P-glycoprotein (P-gp) inhibitor. XR9576 has undergone phase I and II studies as combined chemotherapy against cancer. XR9576 has been developed as a useful therapeutic agent but not as a PET probe. We therefore developed [(11)C]XR9576 as a PET probe and assessed whether PET studies using [(11)C]XR9576 are a promising approach to assess P-gp function primarily. METHODS: We synthesized [(11)C]XR9576 by methylation of 7-O-desmethyl XR9576 with [(11)C]methyl iodide. In in vivo tissue distribution, the effects of co-injection with XR9576 on the uptake of [(11)C]XR9576 in mice were investigated. PET studies using [(11)C]XR9576 were performed in P-gp and/or Bcrp knockout mice as well as in wild-type mice. Metabolites of [(11)C]XR9576 were measured in the brain and plasma of mice. RESULTS: [(11)C]XR9576 was successfully synthesized with suitable radioactivity for injection as well as appropriate radiochemical purity and stability. In in vivo tissue distribution, the brain uptake of [(11)C]XR9576 significantly increased about tenfold of control on co-injection with >10 mg/kg of XR9576. In PET studies, the AUC(brain) ([0-60 min]) in P-gp and P-gp/Bcrp knockout mice was 2- and 11-fold higher than that in wild-type mice. [(11)C]XR9576 showed a high metabolic stability (>90% unchanged form) in the brain and plasma of mice 30 min after injection. These results suggest that a tracer amount of [(11)C]XR9576 behave as the P-gp and Bcrp substrate, and the increased brain uptake or AUC(brain) of [(11)C]XR9576 correlates with P-gp and Bcrp functions. CONCLUSIONS: PET studies using [(11)C]XR9576 may be a promising approach for evaluating deficiency of the function of drug efflux transporters targeting intracranial diseases and tumors.

Radiosynthesis of novel carbon-11-labeled triaryl ligands for cannabinoid-type 2 receptor.

Two novel triaryl ligands 2 and 5 with potent in vitro binding affinities for the cannabinoid subtype-2 (CB2) receptor were labeled with a positron-emitting radioactive nuclide (11)C. Radioligands [(11)C]2, [(11)C]5, and their analogs [(11)C]3 and [(11)C]4 were synthesized by O-[(11)C]methylation of their corresponding phenol precursors with [(11)C]CH(3)I. [(11)C]2-5 had relatively high uptakes (>1.2% injected dose/g tissue) in mouse brains.

[11C]Gefitinib ([11c]Iressa): radiosynthesis, in vitro uptake, and in vivo imaging of intact murine fibrosarcoma.

OBJECTIVE: Gefitinib (N-(3-chloro-4-fluorophenyl)-7-methoxy-6-[3-(morpholin-4-yl)propoxy]quinazolin-4-amine, Iressa) is an approved anticancer drug. In this study, we labeled gefitinib with carbon-11 and evaluated [(11)C]gefitinib to explore its specific binding in intact fibrosarcoma (NFSa)-bearing mice. METHODS: [(11)C]Gefitinib was synthesized by the reaction of desmethyl precursor (1) with [(11)C]CH(3)I. In vitro uptake of [(11)C]gefitinib into NFSa, human-A431 epidermoid carcinoma, and Jurkat T cells was determined. Positron emission tomography (PET) imaging using [(11)C]gefitinib was performed for NFSa-bearing mice. RESULTS: [(11)C]Gefitinib accumulated into NFSa cells with 2.1 uptake ratio (UR)/mg protein in cells. Addition of nonradioactive gefitinib decreased uptake in a concentration-dependent manner. [(11)C]Gefitinib also had high uptake (2.6 UR/mg protein) into epidermal growth factor receptor/tyrosine kinase (EGFR/TK)-rich A431 cells but low uptake (0.2 UR/mg protein) into EGFR/TK-poor Jurkat cells. In vivo distribution study on NFSa-bearing mice by the dissection method revealed that [(11)C]gefitinib specifically accumulated into the tumor. The ratio of radioactivity in tumors to that in blood and muscle as two comparative regions increased from 0.4 to 6.0 and from 0.6 to 5.0 during this experiment (0-60 min), respectively. PET for NFSa-bearing mice produced a clear tumor image, although high radioactivity was distributed throughout the body. Treatment with nonradioactive gefitinib (100 mg/kg) decreased uptake in the tumor. In vivo metabolite analysis demonstrated that [(11)C]gefitinib was stable in the tumor, liver, kidney, and blood. CONCLUSION: These results demonstrated the promising potential of [(11)C]gefitinib to serve as a PET ligand for in vivo imaging of NFSa-bearing mice.