Makoto Takei

Sensitive measurement of positron emitters eluted from HPLC

For sensitive analysis of the radioactive-metabolite in human PET, a radio-HPLC system coupled to a newly designed positron detector was constructed. The detector had the advantages of low noise level (1.7 +/- 1.0 cpm) and high sensitivity (32 +/- 1%) due to coincidence counting and large BGO crystals. Furthermore, the detector was easy to move, since a pair of the BGO housings coupled to photomultipliers was effectively arranged in parallel and a HPLC cell with different volume could be inserted between the BGO housing. This radio-HPLC system was useful for analyzing samples with low radioactivity. It was applied to the measurement of [11C]FLB457 in plasma, having high affinity and high selectivity with dopamine D2 receptors. Extremely low radioactivity of [11C]FLB457 (2500 dpm) could be analyzed by using the radio-HPLC system. The performance of this detector was compared with those of commercially available systems that had been used as sensitive detectors for HPLC.

Radiosynthesis, Photoisomerization, Biodistribution, and Metabolite Analysis of 11C-PBB3 as a Clinically Useful PET Probe for Imaging of Tau Pathology

2-((1E,3E)-4-(6-(11C-methylamino)pyridin-3-yl)buta-1,3-dienyl)benzo[d]thiazol-6-ol (11C-PBB3) is a clinically useful PET probe that we developed for in vivo imaging of tau pathology in the human brain. To ensure the availability of this probe among multiple PET facilities, in the present study we established protocols for the radiosynthesis and quality control of 11C-PBB3 and for the characterization of its photoisomerization, biodistribution, and metabolism. Methods: 11C-PBB3 was synthesized by reaction of the tert-butyldimethylsilyl desmethyl precursor (1) with 11C-methyl iodide using potassium hydroxide as a base, followed by deprotection. Photoisomerization of 11C-PBB3 under fluorescent light was determined. The biodistribution and metabolite analysis of 11C-PBB3 was determined in mice using the dissection method. Results: 11C-PBB3 was synthesized with 15.4% ± 2.8% radiochemical yield (decay-corrected, n = 50) based on the cyclotron-produced 11C-CO2 and showed an averaged synthesis time of 35 min from the end of bombardment. The radiochemical purity and specific activity of 11C-PBB3 were 98.0% ± 2.3% and 180.2 ± 44.3 GBq/μmol, respectively, at the end of synthesis (n = 50). 11C-PBB3 showed rapid photoisomerization, and its radiochemical purity decreased to approximately 50% at 10 min after exposure to fluorescent light. After the fluorescent light was switched off, 11C-PBB3 retained more than 95% radiochemical purity over 60 min. A suitable brain uptake (1.92% injected dose/g tissue) of radioactivity was observed at 1 min after the probe injection, which was followed by rapid washout from the brain tissue. More than 70% of total radioactivity in the mouse brain homogenate at 5 min after injection represented the unchanged 11C-PBB3, despite its rapid metabolism in the plasma. Conclusion: 11C-PBB3 was produced with sufficient radioactivity and high quality, demonstrating its clinical utility. The present results of radiosynthesis, photoisomerization, biodistribution, and metabolite analysis could be helpful for the reliable production and application of 11C-PBB3 in diverse PET facilities.

Evaluation of Limiting Brain Penetration Related to P-glycoprotein and Breast Cancer Resistance Protein Using [11C]GF120918 by PET in Mice

PurposeGF120918 has a high inhibitory effect on P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). We developed [11C]GF120918 as a positron emission tomography (PET) probe to assess if dual modulation of P-gp and BCRP is useful to evaluate brain penetration.ProceduresPET studies using [11C]GF120918 were conducted on P-gp and/or Bcrp knockout mice as well as wild-type mice.ResultsIn PET studies, the AUCbrain[0–60 min] and K1 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 [11C]GF120918 is related to modulation of P-gp and BCRP and is limited by two transporters working together.ConclusionsPET using [11C]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.

Noninvasive and quantitative assessment of the function of multidrug resistance-associated protein 1 in the living brain

Multidrug resistance-associated protein 1 (MRP1) acts as a defense mechanism by pumping xenobiotics and endogenous metabolites out of the brain. The currently available techniques for studying brain-to-blood efflux have significant limitations related to either their invasiveness or the qualitative assessment. Here, we describe an in vivo method, which overcomes these limitations for assessing MRP1 function, using positron emission tomography (PET) and a PET probe. 6-Bromo-7-[11C]methylpurine was designed to readily enter the brain after intravenous administration and to be efficiently converted to its glutathione conjugate (MRP1 substrate) in situ. Dynamic PET scan provided the brain time—activity curve after injection of 6-bromo-7-[11C]methylpurine into mice. The efflux rate of the substrate was kinetically estimated to be 1.4 h−1 with high precision. Moreover, knockout of Mrp1 gene caused approximately a 90% reduction of the efflux rate, compared with wild-type mice. In conclusion, our method allows noninvasive and quantitative assessment for MRP1 function in the living brain.

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.

Production possibility of 60,61,62Cu radioisotopes by alpha induced reactions on cobalt for PET studies

Abstract Excitation functions were measured by the stacked-foil technique for 59 Co ( α , n ) 62 Cu , 59 Co ( α ,2 n ) 61 Cu and 59 Co ( α ,3 n ) 60 Cu nuclear reactions up to 60 MeV. The excitation functions were compared with the published data. The optimum energy range for the production of 61 Cu and 62 Cu was found to be 39 → 18 and 18.5 → 6 MeV, respectively. The calculated thick target yield of 61 Cu in this energy range was 21.0 mCi / μA (supposing one half-life activation time); and 16.2 mCi / μA (supposing three half-life activation time) for 62 Cu . The level of 60 Cu and 62 Cu impurities at 61 Cu production decreases to around 1% after a 1 h cooling time. The practical yield in this case is 17.2 mCi / μA . For production of 62 Cu the contamination level of 61 Cu increases continuously after EOB, but remains below 1% if the cooling time is less than 0.5 h ( 1.9 mCi / μA at 0.5 h after EOB). Unfortunately, in the case of 60 Cu production, the contamination level of 61 Cu and 62 Cu at EOB was found to be 18.4% and 47.9%, respectively, of the produced 60 Cu activity ( 6.4 mCi / μA , after 60 min irradiation time, in the energy interval 60 → 44 MeV ).

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, Alzheimers disease, Huntingtons disease and Parkinsons 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.

Identification of a major radiometabolite of [11C]PBB3.

INTRODUCTION [(11)C]PBB3 is a clinically used positron emission tomography (PET) probe for in vivo imaging of tau pathology in the brain. Our previous study showed that [(11)C]PBB3 was rapidly decomposed to a polar radiometabolite in the plasma of mice. For the pharmacokinetic evaluation of [(11)C]PBB3 it is important to elucidate the characteristics of radiometabolites. In this study, we identified the chemical structure of a major radiometabolite of [(11)C]PBB3 and proposed the metabolic pathway of [(11)C]PBB3. METHODS Carrier-added [(11)C]PBB3 was injected into a mouse for in vivo metabolite analysis. The chemical structure of a major radiometabolite was identified using LC-MS. Mouse and human liver microsomes and liver S9 samples were incubated with [(11)C]PBB3 in vitro. In silico prediction software was used to assist in the determination of the metabolite and metabolic pathway of [(11)C]PBB3. RESULTS In vivo analysis showed that the molecular weight of a major radiometabolite of [(11)C]PBB3, which was called as [(11)C]M2, was m/z 390 [M+H(+)]. In vitro analysis assisted by in silico prediction showed that [(11)C]M2, which was not generated by cytochrome P450 enzymes (CYPs), was generated by sulfated conjugation mediated by a sulfotransferase. CONCLUSION The major radiometabolite, [(11)C]M2, was identified as a sulfated conjugate of [(11)C]PBB3. [(11)C]PBB3 was metabolized mainly by a sulfotransferase and subsidiarily by CYPs.

Imaging of peripheral-type benzodiazepine receptor in tumor: in vitro binding and in vivo biodistribution of N-benzyl-N-[11C]methyl-2-(7-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide

INTRODUCTION The aim of this study was to evaluate N-benzyl-N-[(11)C]methyl-2-(7-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide ([(11)C]DAC) as a novel peripheral-type benzodiazepine receptor (PBR) ligand for tumor imaging. METHODS [(11)C]DAC was synthesized by the reaction of a desmethyl precursor with [(11)C]CH(3)I. In vitro uptake of [(11)C]DAC was examined in PBR-expressing C6 glioma and intact murine fibrosarcoma (NFSa) cells. In vivo distribution of [(11)C]DAC was determined using NFSa-bearing mice and small-animal positron emission tomography (PET). RESULTS [(11)C]DAC showed specific binding to PBR in C6 glioma cells, a standard cell line with high PBR expression. Specific binding of [(11)C]DAC was also confirmed in NFSa cells, a target tumor cell line in this study. Results of PET experiments using NFSa-bearing mice, showed that [(11)C]DAC was taken up specifically into the tumor, and pretreatment with PK11195 abolished the uptake. CONCLUSIONS [(11)C]DAC was taken up into PBR-expressing NFSa. [(11)C]DAC is a promising PET ligand that can be used for imaging PBR in tumor-bearing mice.

Alpha beam monitoring via natCu + alpha processes in the energy range from 40 to 60 MeV

Abstract Excitation functions were measured using the stacked-foil method for alpha particle-induced nuclear reactions on natural copper leading to the formation of 61Cu, 66Ga and 67Ga in the energy range from 16 to 60 MeV. Extending the recommended database of the nat Cu ( α , x ) 66 Ga and nat Cu ( α , x ) 67 Ga processes with our new values, now the above processes can be used for alpha beam energy and intensity monitoring up to 60 MeV. On the basis of the present study it was found that the nat Cu ( α , x ) 61 Cu process is also a useful candidate for monitor purposes in the 40–60 MeV energy region.