Comparison of L-[1-11C]methionine and L-methyl-[11C]methionine for measuring in vivo protein synthesis rates with PET

To evaluate the feasibility of using either L-[1-11C]-methionine or L-[methyl-11C]methionine for measuring protein synthesis rates by positron emission tomography (PET) in normal and neoplastic tissues, distribution and metabolic studies with 14C- and 11C-labeled methionines were carried out in rats bearing Walker 256 carcinosarcoma. The tissue distributions of the two 14C-labeled methionines were similar except for liver tissue. Similar distribution patterns were observed in vivo by PET using 11C-labeled methionines. The highest 14C incorporation rate into the protein-bound fraction was found in the liver followed by tumor, brain, and pancreas. The incorporation rates in liver and pancreas were different for the two methionines. By chloroform-methanol fractionation of these four tissues, in liver significantly different amounts of 14C were observed in macromolecules. Also in brain tissue slight differences were found. By HPLC analyses of the protein-free fractions of plasma, tumor, and brain tissue at 60 min after injection, for both methionines several 14C-labeled metabolites in different amounts, were detected. About half of the 14C-labeled material in the protein-free fraction was found to be methionine. In these three tissues the amount of nonprotein metabolites and [14C]bicarbonate amount ranged from 10% to 17% and 12% to 15% for L-[1-14C]methionine and L-[methyl-14C]methionine, respectively. From these results it can be concluded that the minor metabolic pathways have to be investigated in order to quantitatively model the protein synthesis by PET.     

Effect of tracer metabolism on PET measurement of [11C]pyrilamine binding to histamine H1 receptors

The present study was carried out to investigate the time course of [11C]pyrilamine metabolism and the degree of entry of metabolites into the brain. PET studies were performed in seven healthy volunteers and arterial plasma concentrations of [11C]pyrilamine and its labeled metabolites were determined. After intravenous injection, [11C]pyrilamine metabolized gradually in the human body, with less than 10% of plasma activity being original radioligand at 60 min. Tracer metabolism markedly affected the input function and the calculated impulse response function of the brain. Rat experiments demonstrated that although metabolites of [11C]pyrilamine might enter the brain, they were not retained for prolonged periods of time. At 30-90 min after injection of [11C]pyrilamine, less than 1% of the radioactivity in the brain was originating from metabolites of [11C]pyrilamine. Based on the rat data, the contribution of 11C-labeled metabolites to total [11C]pyrilamine radioactivity in the human brain was estimated and found to be negligible. These results suggest that the metabolites of [11C]pyrilamine do not accumulate within the cerebral extravascular space and that there is minimal metabolism of [11C]pyrilamine by brain tissue itself. Therefore, [11C]pyrilamine metabolites can be neglected in kinetic analysis, using either a compartmental or a noncompartmental model, of the [11C]pyrilamine binding to histamine H1 receptors.     

Quantitative evaluation of L-[methyl-C-11] methionine uptake in tumor using positron emission tomography

Tumor uptake of L-[methyl-11C]methionine ([11C]Met) was assessed in six patients with brain tumors and three patients with lung cancer using positron emission tomography (PET). In arterial plasma samples taken at 5, 15, 30, and 60 min after injection, a fraction of [11C]Met was measured using high performance liquid chromatography in individual patients. Employing curve fitting, a history of [11C]Met activity was obtained as an input function. By means of sequential PET scannings and graphic analysis, uptake rate and distribution volume of [11C]Met in tumor tissue were calculated. In two studies irreversible uptake into the tumors was not seen when total plasma 11C activity was used as the input; however when [11C]Met plasma activity was used, definite irreversible uptake was seen, indicating tumor viability. In other studies, up to 24% underestimation of uptake rate was found. The present results demonstrated the importance of measuring [11C]Met in plasma for quantitative assessment of in vivo amino acid metabolism in tumors.     

Contribution of labeled carbon dioxide to PET imaging of carbon-11-labeled compounds.

11CO2 is one of the major metabolites of many [11C]-labeled radiopharmaceuticals, including glucose, thymidine, acetate, amino acids, and fatty acids. Our data contradict the notion that the contribution of labeled CO2 to PET images can be disregarded because of its rapid elimination through the lungs. We have measured the retention and excretion of 11CO2 in dogs after the intravenous injection of labeled CO2/HCO3-, which had been equilibrated ex vivo with blood. Only 58% of the label was exhaled as CO2 over the first 60 min after injection, with the rest retained in the body. The injection of [11C]thymidine labeled in the ring-2 position or [11C]acetate labeled in the carboxylate position resulted in the production of large amounts of labeled CO2 with the exhalation of about 47% and 23%, respectively, of the injected label over 60 min. At 10 min after injection of either [11C]thymidine and [11C] acetate, approximately 60% to 70% of total blood activity was in labeled CO2 or bicarbonate. On the other hand, the use of [1-11C]glucose only resulted in exhalation of 5% of the injected dose and CO2/HCO3- made up less than 10% of blood activity at 10 min. Our results indicate that retention and distribution of labeled CO2 needs to be considered when interpreting PET data obtained from 11C-labeled compounds.     

Evaluation of [11C]GB67, a novel radioligand for imaging myocardial alpha 1-adrenoceptors with positron emission tomography

Dysfunction of the sympathetic nervous system underlies a number of myocardial disorders. Positron emission tomography (PET) offers a way of assessing receptor function non-invasively in humans, but there are no PET radioligands for assessing myocardial alpha-adrenoceptors. GB67, a structural and pharmacological analogue of the alpha 1-adrenoceptor antagonist prazosin, was labelled with positron-emitting carbon-11 (t1/2 = 20.4 min) by 11C-methylation of N-desmethylamido-GB67 (GB99). [11C]GB67 was injected intravenously into conscious rats. Serial arterial blood samples were taken. Rats were killed and tissues removed to determine radioactivity. The percentages of unchanged [11C]GB67 and its radioactive metabolites in plasma and tissues were assessed by HPLC. Plasma clearance of radioactivity was rapid. Myocardial uptake was maximal at 1-2 min and decreased slowly during 60 min. Predosing with adrenoceptor antagonists demonstrated selectivity for myocardial alpha 1-adrenoceptors. GB67 and prazosin blocked uptake of radioactivity; the non-selective antagonist, phentolamine, partially blocked uptake; the alpha 2-adrenoceptor antagonist, RX 821002, only blocked uptake at high dose and the beta-adrenoceptor antagonist, CGP 12177, had no effect. Additionally, injection of prazosin at 20 min after radioligand displaced radioactivity. In vivo competition curves obtained by injecting [11C]GB67 with varying amounts of either unlabelled GB67 or its precursor GB99 were fitted to a competitive binding model to provide estimates of the maximum number of binding sites (Bmax) and half saturation doses (K) for myocardium. Assuming a tissue protein content of 10%, the values of Bmax [approximately 13 pmol.(g tissue)-1[ were similar to those ]50-170 fmol.(mg protein)-1] reported for myocardial alpha 1-adrenoceptors assessed in vitro. Both GB67 and its precursor GB99 had high affinity for alpha 1-adrenoceptors [KGB67 = 1.5 nmol.(kg body weight)-1, KGB99 = 4.8 nmol.(kg body weight)-1]. HPLC demonstrated four radioactive metabolites in plasma. [11C]GB67 was 80% of the radioactivity at 5 min and 50% at 45 min. No radioactive metabolites were detected in myocardium up to 60 min after injection. [11C]GB67 was assessed in two male human volunteers. PET demonstrated high myocardial uptake. The profile of radioactive metabolites in plasma was comparable to that in the rat, although metabolism was slower in humans. Thus, [11C]GB67 is a promising radioligand for assessing alpha 1-adrenoceptors in human myocardium with PET.     

Synthesis and evaluation of [18F]1-amino-3-fluorocyclobutane-1-carboxylic acid to image brain tumors.

We have developed a new tumor-avid amino acid, 1-amino-3-fluorocyclobutane-1-carboxylic acid (FACBC), labeled with 18F for nuclear medicine imaging. METHODS: [18F]FACBC was prepared with high specific activity (no carrier added [NCA]) and was evaluated for its potential in tumor localization. A comparative study was performed for [18F]FACBC and [18F]2-fluorodeoxyglucose (FDG) in which the uptake of each agent in 9L gliosarcoma (implanted intracerebrally in Fisher 344 rats) was measured. In addition, the first human PET study of [18F]FACBC was performed on a patient with residual glioblastoma multiforme. Quantitative brain images of the patient were obtained by using a Siemens 921 47-slice PET imaging system. RESULTS: In the rat brain, the initial level of radioactivity accumulation after injection of [18F]FACBC was low (0.11 percentage injected dose per gram [%ID/g]) at 5 min and increased slightly to 0.26 %ID/g at 60 min. The tumor uptake exhibited a maximum at 60 min (1.72 %ID/g), resulting in a tumor-to-brain ratio increase of 5.58 at 5 min to 6.61 at 60 min. In the patient, the uptake of [18F]FACBC in the tumor exhibited a maximum concentration of 146 nCi/mL at 35 min after injection. The uptake of radioactivity in the normal brain tissue was low, 21 nCi/mL at 15 min after injection, and gradually increased to 29 nCi/mL at 60 min after injection. The ratio of tumor to normal tissue was 6 at 20 min after injection. The [18F]FACBC PET scan showed intense uptake in the left frontal region of the brain. CONCLUSION: The amino acid FACBC can be radiofluorinated for clinical use. [18F]FACBC is a potential PET tracer for tumor imaging.     

18F]Fluoroethylflumazenil: a novel tracer for PET imaging of human benzodiazepine receptors.

5-(2'-[18F]Fluoroethyl)flumazenil ([18F]FEF) is a fluorine-18 labelled positron emission tomography (PET) tracer for central benzodiazepine receptors. Compared with the established [11C]flumazenil, it has the advantage of the longer half-life of the fluorine-18 label. After optimisation of its synthesis and determination of its in vitro receptor affinities, we performed first PET studies in humans. PET studies in seven healthy human volunteers were performed on a Siemens ECAT EXACT whole-body scanner after injection of 100-280 MBq [L8F]FEF. In two subjects, a second PET scan was conducted after pretreatment with unlabelled flumazenil (1 mg or 2.5 mg i.v., 3 min before tracer injection). A third subject was studied both with [18F]FEF and with [11C]flumazenil. Brain radioactivity was measured for 60-90 min p.i. and analysed with a region of interest-oriented approach and on a voxelwise basis with spectral analysis. Plasma radioactivity was determined from arterial blood samples and metabolites were determined by high-performance liquid chromatography. In human brain, maximum radioactivity accumulation was observed 4 +/- 2 min p.i., with a fast clearance kinetics resulting in 50% and 20% of maximal activities at about 10 and 30 min, respectively. [18F]FEF uptake followed the known central benzodiazepine receptor distribution in the human brain (occipital cortex >temporal cortex >cerebellum >thalamus >pons). Pretreatment with unlabelled flumazenil resulted in reduced tracer uptake in all brain areas except for receptor-free reference regions like the pons. Parametric images of distribution volume and binding potential generated on a voxelwise basis revealed two- to three-fold lower in vivo receptor binding of [18F]FEF compared with [11C]flumazenil, while relative uptake of [18F]FEF was higher in the cerebellum, most likely owing to its relatively higher affinity for benzodiazepine receptors containing the alpha6 subunit. Metabolism of [18F]FEF was very rapid. Polar metabolites represented about 50%-60% of total plasma radioactivity at 5 min and 80%-90% at 20 min p.i. Although [11C]flumazenil has some advantages over [18F]FEF (higher affinity, slower metabolism, slower kinetics), our results indicate that [18F]FEF is a suitable PET ligand for quantitative assessment of central benzodiazepine receptors, which can be used independently of an on-site cyclotron.     

Different brain radioactivity curves in a PET study with [11C]beta-CIT labelled in two different positions.

The cocaine congener 2beta-carbomethoxy-3beta-(4'-iodophenyl)tropane (beta-CIT) has a chemical structure that enables labelling with carbon-11 either by N-methylation or by O-methylation. The regional brain uptake of [N-methyl-11C]beta-CIT and [O-methyl-11C]beta-CIT was compared in cynomolgus monkeys using positron emission tomography (PET). The striatal uptake of radioactivity after intravenous injection of [O-methyl-11C]beta-CIT reached a plateau at 30-40 min, whereas the uptake of [N-methyl-11C]beta-CIT increased continuously during the time of the PET measurement. Two of the putative labelled metabolites, [N-methyl-11C]beta-CIT-acid and [O-methyl-11C]nor-beta-CIT were prepared and examined with PET to investigate if they may enter the brain and thus add to the radioactivity uptake obtained with [11C]beta-CIT. Less than 0.4% of injected [N-methyl-11C]beta-CIT-acid entered the brain whereas 5-6% of [O-methyl-11C]nor-beta-CIT entered the brain and accumulated in the striatum and in the thalamus. The fraction of [O-methyl-11C]nor-beta-CIT obtained in plasma after intravenous injection of [O-methyl-11C]nor-beta-CIT, however, never exceeded 3%. Consequently, the formation of [N-methyl-11C]beta-CIT-acid and [O-methyl-11C]nor-beta-CIT cannot be the explanation for the different time-activity curves in the monkey brain demonstrated with [11C]beta-CIT labelled in two different positions. An unidentified labelled lipophilic metabolite, detected in monkey plasma after injection of [O-methyl-11C]beta-CIT, remains as the only possible explanation for the differences between [N-methyl-11C]beta-CIT and [O-methyl-11C]beta-CIT.     

In vivo imaging of insulin receptors by PET: preclinical evaluation of iodine-125 and iodine-124 labelled human insulin.

[A(14)-*I]iodoinsulin was prepared for studies to assess the suitability of labeled iodoinsulin for positron emission tomography (PET). Iodine-125 was used to establish the methods and for preliminary studies in rats. Further studies and PET scanning in rats were carried out using iodine-124. Tissue and plasma radioactivity was measured as the uptake index (UI = [cpm x (g tissue)(-1)]/[cpm injected x (g body weight)(-1)]) at 1 to 40 min after intravenous injection of either [A(14)-(125)I]iodoinsulin or [A(14)-(124)I]iodoinsulin. For both radiotracers, initial clearance of radioactivity from plasma was rapid (T(1/2) approximately 1 min), reaching a plateau (UI = 2.8) at approximately 5 min which was maintained for 35 min. Tissue biodistributions of the two radiotracers were comparable; at 10 min after injection, UI for myocardium was 2.4, liver, 4.0, pancreas, 5.4, brain, 0.17, kidney, 22, lung, 2.3, muscle, 0.54 and fat, 0.28. Predosing rats with unlabelled insulin reduced the UI for myocardium (0.95), liver (1.8), pancreas (1.2) and brain (0.08), increased that for kidney (61) but had no effect on that for lung (2.5), muscle (0.50) or fat (0.34). Analysis of radioactivity in plasma demonstrated a decrease of [(125)I]iodoinsulin associated with the appearance of labeled metabolites; the percentage of plasma radioactivity due to [(125)I]iodoinsulin was 40% at 5 min and 10% at 10 min. The heart, liver and kidneys were visualized using [(124)I]iodoinsulin with PET.     

Evaluation of (R)-[11C]verapamil as PET tracer of P-glycoprotein function in the blood-brain barrier: kinetics and metabolism in the rat

There is evidence that P-glycoprotein (P-gp) in the blood-brain barrier (BBB) may be involved in the aetiology of neurological disorders. For quantification of P-gp function in vivo, (R)-[11C]verapamil can be used as a positron emission tomography (PET) tracer, provided that a mathematical model describing kinetics of uptake and clearance of verapamil is available. To develop and validate such a model, the kinetic profile and metabolism of (R)-[11C]verapamil have to be known. The aim of this study was to investigate the presence of labeled metabolites of [11C]verapamil in the plasma and (brain) tissue of Wistar rats. For this purpose, extraction and high-performance liquid chromatography (HPLC) methods were developed. The radioactive metabolites of (R)-[11C]verapamil in the liver were N-dealkylated compounds, O-demethylated compounds and a polar fraction formed from N-demethylation products of (R)-[11C]verapamil. Apart from this [11C] polar fraction, other radioactive metabolites of [11C]verapamil were not detected in the brain tissue. Thirty minutes after injection, unmetabolized (R)-[11C]verapamil accounted for 47% of radioactivity in the plasma and 69% in the brain. Sixty minutes after injection, unmetabolized (R)-[11C] verapamil was 27% and 48% in the plasma and the brain, respectively.     

Metabolite analysis of [11C]flumazenil in human plasma: Assessment as the standardized value for quantitative PET studies

Analysis of carbon-11 labeled metabolites in plasma was carried out during positron emission tomography (PET) studies with a central benzodiazepine receptor ligand [¹¹C]flumazenil ([¹¹C]FMZ) in 24 human subjects (14–76 y.o.) including five normal volunteers and 19 patients with neurological disorders. Arterial plasma samples were obtained at 3, 5, 10, 15, 20, 30 and 60 min after i.v. injection of the tracer, and were analyzed by high-performance liquid chromatography. The rate of plasma [¹¹C]FMZ degradation was associated with a large individual variation, but no significant difference was found in the degradation of [¹¹C]FMZ either between male and female, young and old, or between normal subjects and patient groups. When the mean fraction of unchanged [¹¹C]FMZ at each time point was used instead of individually measured metabolite data for the arterial input function, as much as a 30% error occurred in the distribution volume of the [¹¹C]FMZ binding in the brain. These results indicate that the mean percentage of unchanged [¹¹C]FMZ fraction in subjects cannot be used as the standardized value, and that the analysis of metabolites in plasma is necessary to determine the exact arterial input function for quantitative PET measurement.     

In vivo imaging of adenosine A2A receptors in rat and primate brain using [11C]SCH442416

Purpose The aim of this study was to evaluate the suitability of [¹¹C]SCH442416 for the in vivo imaging of adenosine A_2A receptors.Methods In rats and Macaca nemestrina, we evaluated the time course of the cerebral distribution of [¹¹C]SCH442416. Furthermore, in rats we investigated the rate of metabolic degradation, the inhibitory effects of different drugs acting on adenosine or dopamine receptors and the modification induced by the intrastriatal administration of quinolinic acid (QA).Results The rate of metabolic degradation of [¹¹C]SCH442416 in rats was slow; 60 min after tracer injection, more than 40% of total plasma activity was due to unmetabolised [¹¹C]SCH442416. At the time of maximum uptake, radioactive metabolites represented only 6% of total extractable activity in the cerebellum and less than 1% in the striatum. In the striatum, the region with the highest expression of A_2A receptors, the in vivo uptake of [¹¹C]SCH442416 was significantly reduced only by drugs acting on A_2A receptors or by QA, a neurotoxin that selectively reduces the number of intrastriatal GABAergic neurons. Position emission tomography (PET) studies in monkeys indicated that the tracer rapidly accumulates in brain, reaching maximum uptake between 5 and 10 min. Twenty minutes after the injection, radioactivity concentration in the striatum was two times that in the cerebellum.Conclusion The specificity of binding, the rank order of regional distribution in the brain of rats and M. nemestrina, the good signal to noise ratios and the low amount of radioactive metabolites in brain and periphery indicate that [¹¹C]SCH442416 is a promising tracer for the in vivo imaging of A_2A adenosine receptors using PET.     

[18F]Fluoroethylflumazenil: a novel tracer for PET imaging of human benzodiazepine receptors

5-(2'-[¹⁸F]Fluoroethyl)flumazenil ([¹⁸F]FEF) is a fluorine-18 labelled positron emission tomography (PET) tracer for central benzodiazepine receptors. Compared with the established [¹¹C]flumazenil, it has the advantage of the longer half-life of the fluorine-18 label. After optimisation of its synthesis and determination of its in vitro receptor affinities, we performed first PET studies in humans. PET studies in seven healthy human volunteers were performed on a Siemens ECAT EXACT whole-body scanner after injection of 100-280 MBq [¹⁸F]FEF. In two subjects, a second PET scan was conducted after pretreatment with unlabelled flumazenil (1 mg or 2.5 mg i.v., 3 min before tracer injection). A third subject was studied both with [¹⁸F]FEF and with [¹¹C]flumazenil. Brain radioactivity was measured for 60-90 min p.i. and analysed with a region of interest-oriented approach and on a voxelwise basis with spectral analysis. Plasma radioactivity was determined from arterial blood samples and metabolites were determined by high-performance liquid chromatography. In human brain, maximum radioactivity accumulation was observed 4DŽ min p.i., with a fast clearance kinetics resulting in 50% and 20% of maximal activities at about 10 and 30 min, respectively. [¹⁸F]FEF uptake followed the known central benzodiazepine receptor distribution in the human brain (occipital cortex >temporal cortex >cerebellum >thalamus >pons). Pretreatment with unlabelled flumazenil resulted in reduced tracer uptake in all brain areas except for receptor-free reference regions like the pons. Parametric images of distribution volume and binding potential generated on a voxelwise basis revealed two- to three-fold lower in vivo receptor binding of [¹⁸F]FEF compared with [¹¹C]flumazenil, while relative uptake of [¹⁸F]FEF was higher in the cerebellum, most likely owing to its relatively higher affinity for benzodiazepine receptors containing the Ő subunit. Metabolism of [¹⁸F]FEF was very rapid. Polar metabolites represented about 50%-60% of total plasma radioactivity at 5 min and 80%-90% at 20 min p.i. Although [¹¹C]flumazenil has some advantages over [¹⁸F]FEF (higher affinity, slower metabolism, slower kinetics), our results indicate that [¹⁸F]FEF is a suitable PET ligand for quantitative assessment of central benzodiazepine receptors, which can be used independently of an on-site cyclotron.     

Metabolic fate of L-[methyl-11C]methionine in human plasma

The metabolites of L-[methyl-¹¹C]methionine in the plasma of 8 patients with tumor were measured for 60 min after injection. In the plasma, after a rapid clearance, the total radioactivity remained constant, and protein-bound radioactivity increased rapidly. Non protein metabolites detected by HPLC as at least two components besides methionine, increased with time. Significant individual variations for the metabolism were observed. AT 60 min after injection, 36.5% (range: 16%–72%) and 45.3% (range: 13%–74%) of the ¹¹C was measured as methionine and labeled proteins, respectively.     

In vivo distribution and identification of 11C-activity after injection of [methyl-11C]thymidine in Wistar rats.

[Methyl-11C]thymidine and PET offer an in vivo, noninvasive quantitative approach for studying nucleoside uptake in cells on the condition the fraction of [methyl-11C]thymidine (in deoxyribonucleic acid [DNA] or as DNA precursors) versus the total accumulated activity is known. METHODS: In a group of normal (n = 6) and a group of tumor-bearing (n = 3) Wistar rats, the biodistribution of 11C-activity was studied dynamically. In a second group of rats (n = 6), the animals were killed at 20 min postinjection and the organs and tissues of interest (liver, heart, brain, duodenum and tumor) were measured for activity and then homogenized. 11C-activity in each fraction (cell debris, protein/ DNA-fraction, lipids and supernatant) was measured. The supernatant was analyzed by high-performance liquid chromatography (HPLC)-radiochromatography for identification of different 11C-labeled compounds. RESULTS: After venous injection, most of the 11C-activity was rapidly trapped in the liver and in fast-dividing tissue (e.g., duodenum); minor activity was located in the bladder, kidneys, heart and brain. HPLC separation showed that the 11C-activity of the liver tissue consisted of metabolites only. For the duodenum and tumor, at least 55% of the 11C-activity was precipitated in the protein/DNA-fraction and about 60% as DNA precursors (thymidine, 2'-deoxythymidine 5'-monophosphate and 2'-deoxythymidine 5'- triphosphate ) in the supernatant. CONCLUSION: Despite the in vivo metabolism, major 11C-activity in rapidly dividing tissue consists of [methyl-11C]thymidine incorporated in the DNA. Catabolism takes place mainly in the liver where the degradation products are stored. PET quantification data using [methyl-11C] thymidine can give information about thymidine incorporation in DNA and cell proliferation of tumors.     

Evaluation of [O-methyl-11C]RS-15385-197 as a positron emission tomography radioligand for central alpha2-adrenoceptors

Carbon-11 labelled RS-15385-197 and its ethylsulphonyl analogue, RS-79948-197, were evaluated in rats as potential radioligands to image central alpha2-adrenoceptors in vivo. The biodistributions of both compounds were comparable with that obtained in an earlier study using tritiated RS-79948-197 and were consistent with the known localisation of alpha2-adrenoceptors. The maximal signals (total to non-specific binding) were, however, reduced, in the order [11C]RS-79948-197 < [11C]RS-15385-197 < [3H]RS-79948-197, primarily due to the difference in radiolabel position (O-methyl for carbon- 11 compared with S-ethyl for tritium). This resulted in the in-growth of radiolabelled metabolites in plasma, which, in turn, contributed to the non-specific component of brain radioactivity. Nonetheless, the signal ratio of approximately 5 for a receptor-dense tissue compared with the receptor-sparse cerebellum, at 90-120 min after radioligand injection, encouraged the development of [O-methyl-11C]RS-15385-197 for human positron emission tomography (PET). Unfortunately, in two human PET scans (each of 90 min), brain extraction of the radioligand was minimal, with volumes of distribution more than an order of magnitude lower than that measured in rats. Following intravenous injection, radioactivity was retained in plasma and metabolism of the radiolabelled compound was very low. Retrospective measurements of in vitro plasma protein binding and in vivo brain uptake index (BUI) in rats demonstrated a higher protein binding of the radioligand in human compared with rat plasma and a lower BUI in the presence of human plasma. It is feasible that a higher affinity of RS-15385-197 for human plasma protein compared with receptor limited the transport of the radioligand. Although one of the PET scans showed a slight heterogeneity in biodistribution of radioactivity which was consistent with the known localisation of alpha2-adrenoceptors in human brain, it was concluded that [O-methyl-11C]RS-15385-197 showed little promise for routine quantification of alpha2-adrenoceptors in man.     

Preparation and evaluation of ethyl [18F]fluoroacetate as a proradiotracer of [18F]fluoroacetate for the measurement of glial metabolism by PET

IntroductionChanges in glial metabolism in brain ischemia, Alzheimer's disease, depression, schizophrenia, epilepsy and manganese neurotoxicity have been reported in recent studies. Therefore, it is very important to measure glial metabolism in vivo for the elucidation and diagnosis of these diseases. Radiolabeled acetate is a good candidate for this purpose, but acetate has little uptake in the brain due to its low lipophilicity. We have designed a new proradiotracer, ethyl [¹⁸F]fluoroacetate ([¹⁸F]EFA), which is [¹⁸F]fluoroacetate ([¹⁸F]FA) esterified with ethanol, to increase the lipophilicity of fluoroacetate (FA), allowing the measurement of glial metabolism.MethodsThe synthesis of [¹⁸F]EFA was achieved using ethyl O-mesyl-glycolate as precursor. The blood–brain barrier permeability of ethyl [1-¹⁴C]fluoroacetate ([¹⁴C]EFA) was estimated by a brain uptake index (BUI) method. Hydrolysis of [¹⁴C]EFA in the brain was calculated by the fraction of radioactivity in lipophilic and water fractions of homogenized brain. Using the plasma of five animal species, the stability of [¹⁴C]EFA was measured. Biodistribution studies of [¹⁸F]EFA in ddY mice were carried out and compared with [¹⁸F]FA. Positron emission tomography (PET) scanning using common marmosets was performed for 90 min postadministration. At 60 min postinjection of [¹⁸F]EFA, metabolite studies were performed. Organs were dissected from the marmosets, and extracted metabolites were analyzed with a thin-layer chromatography method.ResultsThe synthesis of [¹⁸F]EFA was accomplished in a short time (29 min) and with a reproducible radiochemical yield of 28.6±3.6% (decay corrected) and a high radiochemical purity of more than 95%. In the brain permeability study, the BUI of [¹⁴C]EFA was 3.8 times higher than that of sodium [1-¹⁴C]fluoroacetate. [¹⁴C]EFA was hydrolyzed rapidly in rat brains. In stability studies using the plasma of five animal species, [¹⁴C]EFA was stable only in primate plasma. Biodistribution studies in mice showed that the uptake of [¹⁸F]EFA in selected organs was higher than that of [¹⁸F]FA. From nonprimate PET studies, [¹⁸F]EFA was initially taken into the brain after injection. Metabolites related to the tricarboxylic acid (TCA) cycle were detected in common marmoset brain.Conclusion[¹⁸F]EFA rapidly enters the brain and is then converted into TCA cycle metabolites in the brains of common marmosets. [¹⁸F]EFA shows promise as a proradiotracer for the measurement of glial metabolism.     

A radiometabolite study of the serotonin transporter PET radioligand [11C]MADAM

Introduction¹¹C]MADAM is a radioligand suitable for PET studies of the serotonin transporter (SERT). Metabolite analysis in human and non-human plasma samples using HPLC separation has shown that [¹¹C]MADAM was rapidly metabolized. A possible metabolic pathway is the S-oxidation which could lead to SOMADAM and SO₂MADAM.In vitro evaluation of these two potential metabolites has shown that SOMADAM exhibited a good affinity for SERT and a good selectivity for SERT over NET and DAT.MethodsComparative PET imaging studies in non-human primate brain with [¹¹C]MADAM and [¹¹C]SOMADAM were carried out, and plasma samples were analyzed using reverse phase HPLC. We have explored the metabolism of [¹¹C]MADAM in rat brain with a view to understand its possible interference for brain imaging with PET.ResultsPET imaging studies in non-human primate brain using [¹¹C]SOMADAM indicated that this tracer does not bind with high amounts to brain regions known to be rich in SERT. The fraction of [¹¹C]SOMADAM in non-human primate plasma was approximately 5% at 4 min and 1% at 15 min after [¹¹C]MADAM injection. HPLC analysis of brain sample after [¹¹C]MADAM injection to rats demonstrated that [¹¹C]SOMADAM was not detected in the brain.Conclusions¹¹C]SOMADAM is not superior over [¹¹C]MADAM as a SERT PET radioligand. Nevertheless, [¹¹C]SOMADAM has been identified as a minor labeled metabolite of [¹¹C]MADAM measured in monkey plasma. [¹¹C]SOMADAM was not detected in rat brain.     

Pharmacokinetics of [18F]FETNIM: a potential marker for PET.

18F-labeled fluoroerythronitroimidazole (FETNIM) has been suggested as a marker of tumor hypoxia for use with PET. Our goal was to evaluate the pharmacokinetic properties of [18F]FETNIM in rats and analyze metabolites in human, dog, and rat plasma and urine. Metabolites in liver and tumor homogenates from tumor-bearing rats, as well as the biodistribution of the tracer, were also studied. METHODS: Radio-thin-layer chromatography and digital autoradiography were used to distinguish metabolites from the parent drug in urine and plasma from 8 patients, 3 dogs, and 18 rats, as well as in liver and tumor homogenates from Sprague-Dawley rats bearing 7,12-dimethylbenzanthracene-induced rat mammary carcinoma. Biodistribution of [18F]FETNIM was also studied in rats at 15, 30, 60, 120, and 240 min after tracer injection. RESULTS: Most of the radioactivity in plasma and urine was the unchanged tracer, whereas rat liver homogenates contained almost only metabolites of [18F]FETNIM. None of the species studied showed binding of tracer to plasma proteins. A large variation-3%-70%-in the radioactivity represented by unchanged [18F]FETNIM was found in rat tumor. A negative correlation was found between the percentage of radioactivity represented by unchanged [18F]FETNIM in tumor tissue and tumor uptake (percentage injected dose per gram of tissue) at later times. The highest radioactivity was seen in urine and kidney; the lowest uptake was in fat, cerebellum, and bone matrix. In contrast to matrix, bone marrow had high uptake of 18F. The tumor-to-blood ratio reached a maximum of 1.80 +/- 0.64 at 2 h. CONCLUSION: We conclude that [18F]FETNIM shows low peripheral metabolism, little defluorination, and possible metabolic trapping in hypoxic tumor tissue. These suggest a potential use for this tracer in PET studies on hypoxia of cancer patients.     

N-[11C-Methyl]chlorphentermine and N,N-[11C-dimethyl]chlorphentermine as brain blood-flow agents for positron emission tomography

N-[11C-methyl]chlorphentermine ([11C]NMCP) and N,N-[11C-dimethyl]chlorphentermine ([11C]NDMCP) were prepared from chlorphentermine and 11CH3I in DMF and evaluated in rats as brain blood-flow agents for positron emission tomography (PET). Tissue distribution of [11C]NMCP showed that brain uptake was 2.70 +/- 0.40% of injected dose per organ at 5 min with no change in radioactivity concentration up to 30 min after i.v. injection. Approximately 80% of the initial brain uptake remained at 60 min. On the other hand, initial brain uptake of [11C] NDMCP (3.66 +/- 0.31 and 3.63 +/- 0.88% injected dose per organ at 5 and 15 min, respectively) was greater than that of [11C]NMCP. The brain activity however, rapidly decreased to 2.38 +/- 0.17 and 1.82 +/- 0.32% at 30 and 60 min, respectively. Because of its longer retention in the brain compared with [11C]NDMCP, [11C]NMCP would be a potential brain blood-flow agent for quantitative PET studies.