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.

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.

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.

Evaluation of N-benzyl-N-[11C]methyl-2- (7-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide ([11C]DAC) as a novel translocator protein (18 kDa) radioligand in kainic acid-lesioned rat.

The aim of this study was to evaluate N-benzyl-N-[11C]methyl-2-(7-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide ([11C]DAC) as a new translocator protein (18 kDa) [TSPO, formerly known as the peripheral-type benzodiazepine receptor (PBR)] positron emission tomography (PET) ligand in normal mice and unilateral kainic acid (KA)-lesioned rats. DAC is a derivative of AC-5216, which is a potent and selective PET ligand for the clinical investigation of TSPO. The binding affinity and selectivity of DAC for TSPO were similar to those of AC-5216, and DAC was less lipophilic than AC-5216. The distribution pattern of [11C]DAC was in agreement with TSPO distribution in rodents. No radioactive metabolite of [11C]DAC was found in the mouse brain, although it was metabolized rapidly in mouse plasma. Using small-animal PET, we examined the in vivo binding of [11C]DAC for TSPO in KA-lesioned rats. [11C]DAC and [11C]AC-5216 exhibited similar brain uptake in the lesioned and nonlesioned striatum, respectively. The binding of [11C]DAC to TSPO was increased significantly in the lesioned striatum, and [(11)C]DAC showed good contrast between the lesioned and nonlesioned striatum (the maximum ratio was about threefold). In displacement experiments, the uptake of [11C]DAC in the lesioned striatum was eventually blocked using an excess of either unlabeled DAC or PK11195 injected. [11C]DAC had high in vivo specific binding to TSPO in the injured rat brain. Therefore, [11C]DAC is a useful PET ligand for TSPO imaging, and its specific binding to TSPO is suitable as a new biomarker for brain injury.

Biodistribution and metabolism of the anti-influenza drug [11C]oseltamivir and its active metabolite [11C]Ro 64-0802 in mice.

INTRODUCTION: Oseltamivir phosphate (Tamiflu) is an orally active anti-influenza drug, which is hydrolyzed by esterase to its carboxylate metabolite Ro 64-0802 with potent activity to inhibit the influenza virus. The abnormal behavior and death associated with the use of oseltamivir have developed into a major problem in Japan where Tamiflu is often prescribed for seasonal influenza. It is critical to determine the amount of oseltamivir and Ro 64-0802 in the human brain and to elucidate the relationship between their amounts and neuropsychiatric side effects. The aim of this study was to evaluate [(11)C]oseltamivir and [(11)C]Ro 64-0802 in mice as promising positron emission tomography (PET) ligands for measuring their amounts in living brains. METHODS: Whole-body biodistribution of [(11)C]oseltamivir and [(11)C]Ro 64-0802 was determined in mice using the dissection method and micro-PET. In vitro and in vivo metabolite assay was performed in the plasma and brain of mice. RESULTS: Between 1 and 60 min after injection of [(11)C]oseltamivir and [(11)C]Ro 64-0802, 0.20-0.06% and 0.39-0.03% ID/g were detected in the mouse brains, respectively (dissection method). Radioactivity concentrations in the living brains between 0 and 90 min after injection were measured at standardized uptake values of 0.25-0.05 for [(11)C]oseltamivir and 0.38-0.02 for [(11)C]Ro 64-0802 (micro-PET). In vivo metabolite assay demonstrated the presence of [(11)C]oseltamivir and [(11)C]Ro 64-0802 in the brains after [(11)C]oseltamivir injection. CONCLUSION: This study determined the distribution and metabolism of [(11)C]oseltamivir and [(11)C]Ro 64-0802 in mice. PET could be used to measure their amounts in the living brain and to elucidate the relationship between the amounts in the brain and the side effects of Tamiflu in the central nervous system.

[18F]FEAC and [18F]FEDAC: Two novel positron emission tomography ligands for peripheral-type benzodiazepine receptor in the brain.

[(18)F]FEAC ([(18)F]4a) and [(18)F]FEDAC ([(18)F]4b) were developed as two novel positron emission tomography (PET) ligands for peripheral-type benzodiazepine receptor (PBR). [(18)F]4a and [(18)F]4b were synthesized by fluoroethylation of precursors 8a and 8b with [(18)F]FCH(2)CH(2)Br ([(18)F]9), respectively. Small-animal PET scan for a neuroinflammatory rat model showed that the two radioligands had high uptakes of radioactivity in the kainic acid-infused striatum, a brain region where PBR density was increased.

In vitro binding of [(11)C]raclopride with ultrahigh specific activity in rat brain determined by homogenate assay and autoradiography.

OBJECTIVE: The aim of this study was to characterize the in vitro binding of [(11)C]raclopride with ultrahigh specific activity (SA) in the striatum and cerebral cortex of rat brain. METHODS: [(11)C]Raclopride, a dopamine D(2) receptor ligand, with an ultrahigh SA of 4880+/-2360 GBq/micromol (132+/-64 Ci/micromol, n=25) was synthesized. In vitro binding experiment was performed using brain homogenate assay and autoradiography (ARG). RESULTS: In vitro homogenate assay demonstrated that high SA [(11)C]raclopride (2520-6340 GBq/micromol; 68-171 Ci/micromol) had two-affinity (high and low) binding sites in the striatum and cerebral cortex of rat brain. In the striatum, K(d,high) and B(max,high) values were 0.005+/-0.002 nM and 0.19+/-0.04 fmol/mg tissue, respectively, while K(d,low) and B(max,low) values were 2.2+/-1.0 nM and 35.8+/-16.4 fmol/mg tissue, respectively. In the cerebral cortex, K(d,high) and B(max,high) values were 0.061+/-0.087 nM and 0.2+/-0.2 fmol/mg tissue, respectively, while K(d,low) and B(max,low) values were 2.5+/-3.2 nM and 5.5+/-4.8 fmol/mg tissue, respectively. On the other hand, only one binding site was found in the striatum and no binding site was identified in the cerebral cortex using low SA [(11)C]raclopride (44 GBq/micromol; 1.2 Ci/micromol). In vitro ARG for the rat brain using high SA [(11)C]raclopride (6212 GBq/micromol; 168 Ci/micromol) gave a coronal image of the striatum and cerebral cortex with a higher signal/noise ratio than using low SA [(11)C]raclopride (40 GBq/micromol; 1.1 Ci/micromol). CONCLUSION: Using ultrahigh SA [(11)C]raclopride for the in vitro homogenate assay, we succeeded in detecting two-affinity binding sites of [(11)C]raclopride, not only in the striatum but also in the cerebral cortex of rat brain.

Radiosyntheses of two positron emission tomography probes: [11C]Oseltamivir and its active metabolite [11C]Ro 64-0802.

Oseltamivir phosphate (Tamiflu, 1.H(3)PO(4) is an orally active anti-influenza drug, which is hydrolyzed by esterase to its carboxylate metabolite Ro 64-0802 (2) with potent activity inhibiting neuraminidase. In this study, for the first time, we synthesized carbon-11-labeled oseltamivir ([(11)C]1) and Ro 64-0802 ([(11)C]2) as two novel positron emission tomography probes and demonstrated that [(11)C]1 had twofold higher radioactivity concentration in the mouse brains than [(11)C]2.

11C-AC-5216: a novel PET ligand for peripheral benzodiazepine receptors in the primate brain.

UNLABELLED: Developing a PET ligand for imaging of the peripheral benzodiazepine receptor (PBR; Translocator Protein [18 kDa] TSPO) is of great importance for studying its role in glial cells in the injured brain and in neurodegenerative disorders, such as Alzheimer's disease. The aim of this study was to synthesize and evaluate N-benzyl-N-ethyl-2-(7-(11)C-methyl-8-oxo-2-phenyl-7,8-dihydro-9H-purin-9-yl)acetamide ((11)C-AC-5216) as a PET ligand for imaging PBR in the primate brain. METHODS: AC-5216 and its desmethyl precursor (compound 1) were synthesized starting from commercially available compounds. The radiosynthesis of (11)C-AC-5216 was performed through the reaction of compound 1 with (11)C-CH(3)I in the presence of NaH. The in vivo brain regional distribution was determined in mice (dissection) and a monkey (PET). RESULTS: (11)C-AC-5216 (800-1,230 MBq; n = 25) was obtained with a radiochemical purity of 98% and a specific activity of 85-130 GBq/mumol at the end of synthesis. After injection of (11)C-AC-5216 into mice, a high accumulation of radioactivity was found in the lungs, heart, adrenal glands, and other PBR-rich organs. In the mouse brain, high radioactivity was observed in the olfactory bulb and cerebellum. Radioactivity in these regions was inhibited by nonradioactive AC-5216 or PK11195 but was not decreased by central benzodiazepine receptor-selective flumazenil and Ro15-4513. A PET study of the monkey brain determined that (11)C-AC-5216 had a relatively high uptake in the occipital cortex, a rich PBR-dense area in the primate brain. Pretreatment with nonradioactive AC-5216 and PK11195 reduced the radioactivity of (11)C-AC-5216 in the occipital cortex significantly, suggesting its high specific binding with PBR in the brain. Metabolite analysis demonstrated that (11)C-AC-5216 was stable in vivo in the mouse brain, although it was metabolized in the plasma of mice and the monkey. CONCLUSION: (11)C-AC-5216 is a promising PET ligand for imaging PBR in rodent and primate brains.

In vitro and ex vivo autoradiography studies on peripheral-type benzodiazepine receptor binding using [11C]AC-5216 in normal and kainic acid-lesioned rats.

AC-5216 was reported as a novel ligand for peripheral-type benzodiazepine receptor (PBR) with a different chemical structure from DAA1106 analogues. This ligand had potent affinity for PBR and selectivity for PBR over other neurotransmitters. We have previously labeled AC-5216 using positron-emitter (11)C. The aim of this study was to evaluate [(11)C]AC-5216 in a rat brain model with neuroinflammation using an autoradiography (ARG) technique. In vitro ARG of normal rat brain showed that [(11)C]AC-5216 accumulated highly in the olfactory bulb, choroid plexus and cerebellum. The distribution pattern agreed with the localization of PBR in the rodent brain. Infusion of kainic acid (KA: 1, 2.5 and 5 nmol) into the rat striatum resulted in neuroinflammation. In vitro and ex vivo ARG revealed that the radioactivity level of [(11)C]AC-5216 was increased significantly in the KA-lesioned striatum compared to the non-lesioned striatum. Increasing the amount of KA infused into the striatum augmented radioactivity in the striatum as well as the cerebral cortex and hippocampus of the lesioned side. Treatment with a large amount of non-radioactive AC-5216 or PK11195 inhibited the binding of [(11)C]AC-5216 and diminished the difference of radioactivity levels between the lesion and non-lesioned sides. These results demonstrated that [(11)C]AC-5216 had high specific binding to PBR in the KA-lesioned rat brain. Thus, [(11)C]AC-5216 is a promising PET ligand for imaging PBR in a brain with neuroinflammation.

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.

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.

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 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 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.

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.