[11C]-(R)PK11195 tracer kinetics in the brain of glioma patients and a comparison of two referencing approaches

Purpose

Translocator protein (TSPO) is a biomarker of neuroinflammation that can be imaged by PET using [¹¹C]-(R)PK11195. We sought to characterize the [¹¹C]-(R)PK11195 kinetics in gliomas of different histotypes and grades, and to compare two reference tissue input functions (supervised cluster analysis versus cerebellar grey matter) for the estimation of [¹¹C]-(R)PK11195 binding in gliomas and surrounding brain structures.

Methods

Twenty-three glioma patients and ten age-matched controls underwent structural MRI and dynamic [¹¹C]-(R)PK11195 PET scans. Tissue time–activity curves (TACs) were extracted from tumour regions as well as grey matter (GM) and white matter (WM) of the brains. Parametric maps of binding potential (BP_ND) were generated with the simplified reference tissue model using the two input functions, and were compared with each other. TSPO expression was assessed in tumour tissue sections by immunohistochemistry.

Results

Three types of regional kinetics were observed in individual tumour TACs: GM-like kinetics (n?=?6, clearance of the tracer similar to that in cerebellar GM), WM-like kinetics (n?=?8, clearance of the tracer similar to that in cerebral WM) and a form of mixed kinetics (n?=?9, intermediate rate of clearance). Such kinetic patterns differed between low-grade astrocytomas (WM-like kinetics) and oligodendrogliomas (GM-like and mixed kinetics), but were independent of tumour grade. There was good agreement between parametric maps of BP_ND derived from the two input functions in all controls and 10 of 23 glioma patients. In 13 of the 23 patients, BP_ND values derived from the supervised cluster input were systematically smaller than those using the cerebellar input. Immunohistochemistry confirmed that TSPO expression increased with tumour grade.

Conclusion

The three types of [¹¹C]-(R)PK11195 kinetics in gliomas are determined in part by tracer delivery, and indicated that kinetic analysis is a valuable tool in the study of gliomas with the potential for in vivo discrimination between low-grade astrocytomas and oligodendrogliomas. Supervised cluster and cerebellar input functions produced consistent BP_ND estimates in approximately half of the gliomas investigated, but had a systematic difference in the remainder. The cerebellar input is preferred based on theoretical and practical considerations.     

Optimizing an online SPE-HPLC method for analysis of (R)-[11C]1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide [(R)-[11C]PK11195] and its metabolites in humans

(R)-[11C]PK11195 is used as a positron emission tomography tracer for activated microglia in several neurological disorders. Quantification of specific binding requires a metabolite-corrected plasma input function. In this study, a high-performance liquid chromatography (HPLC) procedure with online solid phase extraction was modified for analyzing (R)-[11C]PK11195 plasma samples, yielding total sample recoveries of more than 98%. When applied to human studies, the use of two HPLC systems enabled analysis of up to seven plasma samples under regular conditions. Online radioactivity detection was compared with offline sample measurements of HPLC profiles. Offline measurements provided the most reliable results especially for late plasma samples. In 10 patients, an average decrease of parent compound from 94.6% at 2.5 min to 45.2% at 1 h after administration was observed.     

Microglial activation in perinatal rabbit brain induced by intrauterine inflammation: detection with 11C-(R)-PK11195 and small-animal PET.

Intrauterine infection can lead to a fetal inflammatory response syndrome that has been implicated as one of the causes of perinatal brain injury leading to periventricular leukomalacia (PVL) and cerebral palsy. The presence of activated microglial cells has been noted in autopsy specimens of patients with PVL and in models of neonatal hypoxia and ischemia. Activated microglial cells can cause oligodendrocyte damage and white matter injury by release of inflammatory cytokines and production of excitotoxic metabolites. We hypothesized that exposure to endotoxin in utero leads to microglial activation in the fetal brain that can be monitored in vivo by (11)C-(R)-PK11195 (1-[2-chlorophenyl]-N-methyl-N-[1-methylpropyl]-3-isoquinoline carboxamide)--a positron-emitting ligand that binds peripheral benzodiazepine receptor sites in activated microglia--using small-animal PET. METHODS: Pregnant New Zealand White rabbits underwent laparotomy and were injected with 20 and 30 microg/kg of Escherichia coli lipopolysaccharide along the length of the uterus on day 28 of gestation. The pups were born spontaneously at term (31 d) and were scanned using small-animal PET after intravenous administration of (11)C-(R)-PK11195 and by MRI on postnatal day 1. The standard uptake values (SUVs) of the tracer were calculated for the whole brain at 10-min intervals for 60 min after tracer injection. The pups were euthanized after the scan, and brains were fixed, sectioned, and stained for microglial cells using biotinylated tomato lectin. RESULTS: There was increased brain retention of (11)C-(R)-PK11195--as determined by a significant difference in the slope of the SUV over time--in the endotoxin-treated pups when compared with that of age-matched controls. Immunohistochemical staining showed dose-dependent changes in activated microglia (increased number and morphologic changes) in the periventricular region and hippocampus of the brain of newborn rabbit pups exposed to endotoxin in utero. CONCLUSION: Intrauterine inflammation leads to activation of microglial cells that may be responsible for the development of brain injury and white matter damage in the perinatal period. PET with the tracer (11)C-(R)-PK11195 can be used as a noninvasive, sensitive tool for determining the presence and progress of neuroinflammation due to perinatal insults in newborns.     

Novel reference region model reveals increased microglial and reduced vascular binding of 11C-(R)-PK11195 in patients with Alzheimer's disease.

11C-(R)-PK11195 is a PET radiotracer for the quantification of peripheral benzodiazepine binding sites (PBBSs). The PBBS is a consistent marker of activated microglia, and 11C-(R)-PK11195 has been used to image microglial activity in the diseased brain and in neoplasia. However, the PBBS is also expressed in the brain vasculature (endothelium and smooth muscles), and no evidence, to our knowledge, exists of a change in the vascular PBBS in pathologic brains or of such a change having an effect on the quantification of 11C-(R)-PK11195 binding. To investigate this issue, we have used a modified reference-tissue model (SRTMV) that accounts for tracer vascular activity both in reference and target tissues and applied it for the estimation of binding potential (BP) in a cohort of patients with Alzheimer's disease (AD). METHODS: A total of 10 patients with AD and 10 age-matched healthy subjects who underwent a 11C-(R)-PK11195 scan were considered in the analysis. The time-activity curves of 11 regions of interest were extracted using the Hammersmith maximum probability atlas. BPs were first estimated using the standard simplified reference-tissue model (SRTM) with the reference tissue computed with a supervised selection algorithm. Subsequently, we applied an SRTMV that models PBBS vascular activity using an additional linear term for both target (VbT) and reference (VbR) regions accounting for vascular tracer activity (C(B)), whereas C(B) was extracted directly from the images. VbR was fixed to 5%, and R1, k2, BP, and VbT were estimated. PBBS density in the vasculature was also assessed by immunocytochemistry on a separate cohort of young and elderly controls and 3 AD postmortem brains. RESULTS: The inclusion of a vascular component in the SRTM increased BPs in all subjects, but the amount of the increase was different (about 11.9% in controls and 16.8% in patients with AD). In addition, average VbT values derived using the SRTMV were 4.22% for controls but only 2.87% in patients with AD. Immunochemistry showed reduced PBBS expression in AD due to vascular fibrosis. CONCLUSION: The reduction of VbT in AD can be interpreted as a consequence of 2 independent but concurring phenomena. The vascular fibrosis in the AD brain causes the well-documented decrease of the size of lumens and the reduction of blood volume. At the same time, the fibrotic process determines the loss of vascular PBBS, particularly in smooth muscles, as here documented by immunochemistry. The inclusion of the additional vascular component in the SRTM effectively models these 2 concurrent processes and amplifies the BP in AD more than in controls because of the decrease in tracer binding to the vasculature in the disease cohort.     

Synthesis and preclinical evaluation of [11C]D617, a metabolite of (R)-[11C]verapamil

Objectives(R)-[¹¹C]verapamil is widely used as a positron emission tomography (PET) tracer to evaluate P-glycoprotein (P-gp) functionality at the blood–brain barrier in man. A disadvantage of (R)-[¹¹C]verapamil is the fact that its main metabolite, [¹¹C]D617, also enters the brain. For quantitative analysis of (R)-[¹¹C]verapamil data, it has been assumed that the cerebral kinetics of (R)-[¹¹C]verapamil and [¹¹C]D617 are the same. The aim of the present study was to investigate whether the cerebral kinetics of (R)-[¹¹C]verapamil and [¹¹C]D617 are indeed similar and, if so, whether [¹¹C]D617 itself could serve as an alternative PET tracer for P-gp.Methods[¹¹C]D617 was synthesized and its ex vivo biodistribution was investigated in male rats at four time points following intravenous administration of [¹¹C]D617 (50 MBq) without (n=4) or with (n=4) pretreatment with the P-gp inhibitor tariquidar (15 mg·kg^?1, intraperitoneally). Brain distribution was further assessed using consecutive PET scans (n=8) before and after pretreatment with tariquidar (15 mg·kg^?1, intravenously), as well as metabolite analysis (n=4).ResultsThe precursor for the radiosynthesis of [¹¹C]D617, 5-amino-2-(3,4-dimethoxy-phenyl)-2-isopropyl-pentanitrile (desmethyl D617), was synthesized in 41% overall yield. [¹¹C]D617 was synthesized in 58%–77% decay-corrected yield with a radiochemical purity of ≥99%. The homogeneously distributed cerebral volume of distribution (V_T) of [¹¹C]D617 was 1.1, and this increased 2.4-fold after tariquidar pretreatment.ConclusionV_T of [¹¹C]D617 was comparable to that of (R)-[¹¹C]verapamil, but its increase after tariquidar pretreatment was substantially lower. Hence, (R)-[¹¹C]verapamil and [¹¹C]D617 do not show similar brain kinetics after inhibition of P-gp with tariquidar.     

Peripheral metabolism of (R)-[11C]verapamil in epilepsy patients

Purpose (R)-[¹¹C]verapamil is a new PET tracer for P-glycoprotein-mediated transport at the blood-brain barrier. For kinetic analysis of (R)-[¹¹C]verapamil PET data the measurement of a metabolite-corrected arterial input function is required. The aim of this study was to assess peripheral (R)-[¹¹C]verapamil metabolism in patients with temporal lobe epilepsy and compare these data with previously reported data from healthy volunteers. Methods Arterial blood samples were collected from eight patients undergoing (R)-[¹¹C]verapamil PET and selected samples were analysed for radiolabelled metabolites of (R)-[¹¹C]verapamil by using an assay that measures polar N-demethylation metabolites by solid-phase extraction and lipophilic N-dealkylation metabolites by HPLC. Results Peripheral metabolism of (R)-[¹¹C]verapamil was significantly faster in patients compared to healthy volunteers (AUC of (R)-[¹¹C]verapamil fraction in plasma: 29.4 ± 3.9 min for patients versus 40.8 ± 5.0 min for healthy volunteers; p < 0.0005, Student’s t-test), which resulted in lower (R)-[¹¹C]verapamil plasma concentrations (AUC of (R)-[¹¹C]verapamil concentration, normalised to injected dose per body weight: 25.5 ± 2.1 min for patients and 30.5 ± 5.9 min for healthy volunteers; p = 0.038). Faster metabolism appeared to be mainly due to increased N-demethylation as the polar [¹¹C]metabolite fraction was up to two-fold greater in patients. Conclusions Faster metabolism of (R)-[¹¹C]verapamil in epilepsy patients may be caused by hepatic cytochrome P450 enzyme induction by antiepileptic drugs. Based on these data caution is warranted when using an averaged arterial input function derived from healthy volunteers for the analysis of patient data. Moreover, our data illustrate how antiepileptic drugs may decrease serum levels of concomitant medication, which may eventually lead to a loss of therapeutic efficacy.     

A comparative small-animal PET evaluation of [11C]tariquidar, [11C]elacridar and (R)-[11C]verapamil for detection of P-glycoprotein-expressing murine breast cancer

Purpose

One important mechanism for chemoresistance of tumours is overexpression of the adenosine triphosphate-binding cassette transporter P-glycoprotein (Pgp). Pgp reduces intracellular concentrations of chemotherapeutic drugs. The aim of this study was to compare the suitability of the radiolabelled Pgp inhibitors [¹¹C]tariquidar and [¹¹C]elacridar with the Pgp substrate radiotracer (R)-[¹¹C]verapamil for discriminating tumours expressing low and high levels of Pgp using small-animal PET imaging in a murine breast cancer model.

Methods

Murine mammary carcinoma cells (EMT6) were continuously exposed to doxorubicin to generate a Pgp-overexpressing, doxorubicin-resistant cell line (EMT6AR1.0 cells). Both cell lines were subcutaneously injected into female athymic nude mice. One week after implantation, animals underwent PET scans with [¹¹C]tariquidar (n?=?7), [¹¹C]elacridar (n?=?6) and (R)-[¹¹C]verapamil (n?=?7), before and after administration of unlabelled tariquidar (15?mg/kg). Pgp expression in tumour grafts was evaluated by Western blotting.

Results

[¹¹C]Tariquidar showed significantly higher retention in Pgp-overexpressing EMT6AR1.0 compared with EMT6 tumours: the mean?±?SD areas under the time?Cactivity curves in scan 1 from time 0 to 60?min (AUC_0?C60) were 38.8?±?2.2?min and 25.0?±?5.3?min (p?=?0.016, Wilcoxon matched pairs test). [¹¹C]Elacridar and (R)-[¹¹C]verapamil were not able to discriminate Pgp expression in tumour models. Following administration of unlabelled tariquidar, both EMT6Ar1.0 and EMT6 tumours showed increases in uptake of [¹¹C]tariquidar, [¹¹C]elacridar and (R)-[¹¹C]verapamil.

Conclusion

Among the tested radiotracers, [¹¹C]tariquidar performed best in discriminating tumours expressing high and low levels of Pgp. Therefore [¹¹C]tariquidar merits further investigation as a PET tracer to assess Pgp expression levels in solid tumours.     

Comparative studies of potential cancer biomarkers carbon-11 labeled MMP inhibitors (S)-2-(4'-[11C]methoxybiphenyl-4-sulfonylamino)-3-methylbutyric acid and N-hydroxy-(R)-2-[[(4'-[11C]methoxyphenyl)sulfonyl]benzylamino]-3-methylbutanamide

(S)-2-(4'-[11C]methoxybiphenyl-4-sulfonylamino)-3-methylbutyric acid ([11C]MSMA) and N-hydroxy-(R)-2-[[(4'-[11C]methoxyphenyl)sulfonyl]benzylamino]-3-methylbutanamide ([11C]CGS 25966), carbon-11 labeled matrix metalloproteinase (MMP) inhibitors, have been synthesized for evaluation as new potential positron emission tomography (PET) cancer biomarkers. [11C]MSMA was prepared by appropriate precursor (S)-2-(4'-hydroxybiphenyl-4-sulfonylamino)-3-methylbutyric acid tert-butyl ester, which was synthesized in eight steps from amino acid (L)-valine in 39.4% chemical yield. This precursor was labeled by [11C]methyl triflate through O-[11C]methylation method at the hydroxyl position of biphenol under basic conditions, followed by a quick acid hydrolysis and isolated by solid-phase extraction (SPE) purification to produce pure target compound [11C]MSMA in 35-55% radiochemical yield, based on 11CO2, decay corrected to end of bombardment (EOB), and 20-25 min synthesis time. [11C]CGS 25966 was prepared in our previous work starting from amino acid (D)-valine. The biodistribution of [11C]MSMA and [11C]CGS 25966 were determined at 45 min post iv injection in breast cancer animal models MCF-7's transfected with IL-1alpha implanted athymic mice and MDA-MB-435 implanted athymic mice. The results showed the uptakes of [11C]MSMA and [11C]CGS 25966 in these tumors were 0.95 and 0.42%dose/g in MCF-7's transfected with IL-1alpha implanted mice, 0.98 and 1.53%dose/g in MDA-MB-435 implanted mice, respectively; the ratios of tumor/muscle (T/M) and tumor/blood (T/B) were 1.21 and 1.09 (T/M, MCF-7's), 0.99 and 0.84 (T/B, MCF-7's), 1.38 and 1.27 (T/M, MDA-MB-435), 1.27 and 1.95 (T/B, MDA-MB-435), respectively. The micro-PET images of [11C]MSMA and [11C]CGS 25966 in both breast cancer athymic mice were acquired for 15 min from a MCF-7's transfected with IL-1alpha and/or MDA-MB-435 implanted mouse at 45 min post iv injection of 1 mCi of the tracer using a dedicated high resolution (<3 mm full-width at half-maximum) small FOV (field-of-view) PET imaging system, Indy-PET II scanner, developed in our laboratory, which showed both tumors were invisible with both tracers. The results were compared. From our results, we concluded that both [11C]MSMA and [11C]CGS 25966 might be unsuitable as PET tracers for cancer imaging.     

Human biodistribution and radiation dosimetry of 11C-(R)-PK11195, the prototypic PET ligand to image inflammation

Purpose  

The positron emission tomography (PET) radiotracer ¹¹C-(R)-PK11195 allows the in vivo imaging in humans of the translocator protein 18 kDa (TSPO), previously called peripheral benzodiazepine receptor (PBR), a marker of inflammation. Despite its widespread use, the radiation burden associated with ¹¹C-(R)-PK11195 in humans is not known. To examine this, we performed dynamic whole-body imaging with PET and ¹¹C-(R)-PK11195 in healthy humans.     

PET Measurements of cAMP-Mediated Phosphodiesterase-4 with (R)-[11C]Rolipram

Cyclic adenosine monophosphate (cAMP) is the common second messenger in signal-transduction cascades originating at a number of monoamine receptors involved in neurotransmission, cardiac function and smooth muscle contraction. Altered regulation of cAMP synthesis (at receptors, G-protein subunits or adenylyl cyclase) and breakdown by phosphodiesterase (PDE) enzymes have been implicated in a number of pathologies. The PDE4 inhibitor (R)-rolipram, and the less active (S)- enantiomer, have been labeled with carbon-11 and characterized by in vivo and in vitro experiments for use in the evaluation of altered PDE4 levels in the brain and cardiac tissues. (R)-[11C]Rolipram has been shown to bind selectively to PDE4 over other PDE isozymes, with specific binding reflecting approximately 80 and 40% of the total detected radioactivity in the rat brain and the heart, respectively. Tracer retention in PDE4-rich tissues is increased by cAMP-elevating treatments, as detected by in vivo PET studies and ex vivo biodistribution experiments. In vivo PET imaging studies display strong region-specific signal in the brain and heart, as evaluated in rats, pigs, monkeys and humans. Impaired cAMP-mediated signaling was observed in animal models of aging, obesity, anthracycline-induced cardiotoxicity and myocardial infarction using (R)-[11C]rolipram. Given the critical role of cAMP in multiple hormonal pathways, the good safety profile and well-characterized pharmacokinetics, (R)-[11C]rolipram PET imaging provides a novel tool for serial monitoring of cAMP-mediated signaling at the PDE4 level, yielding insight into pathological progression with potential for directing therapy.     

Serial PET studies of human cerebral malignancy with [1-11C]putrescine and [1-11C]2-deoxy-D-glucose

Serial PET measurements of [1-11C]putrescine ([11C]PUT) uptake and glucose metabolic rate (GMR) using [1-11C]2-deoxy-D-glucose ([11C]2DG) were made on eight human subjects with a radiological and, in most cases, pathological diagnosis of primary or metastatic brain tumor. Blood-to-brain influx constants (Ki) were calculated for [11C]PUT. Tumor uptake of 11C after [11C]PUT injection was unidirectional peaking at 15 min. The mean +/- s.d. Kis for [11C]PUT for tumor and normal brain tissue were 0.78 +/- 0.045 and 0.024 +/- 0.007 ml cc-1 min-1, respectively (average of ratio, 3.11) whereas the ratio of GMR for tumor and normal brain tissue was 1.2 +/- 0.5. The mean Ki for four active, high grade astrocytomas was 0.098 +/- 0.030 in contrast to 0.027 +/- 0.008 ml cc-1 min-1 for two patients with low grade astrocytoma. Active high grade astrocytomas also showed marked CT contrast enhancement and regional glucose hypermetabolism. In one subject with brain metastases, both [11C]PUT and GMR correlated with a declining clinical picture in repeated studies over a 4-mo period. PET studies with [11C]PUT provide a better signal:noise ratio than GMR measurements, are useful for locating small glycolytically hypometabolic tumors and, when used in longitudinal studies in a single subject, appear to provide an index of degree of malignancy.     

Synthesis of racemic [3-11C]phenylalanine and [3-11C]DOPA

The synthesis of racemic [3-¹¹C]phenylalanine and [3-¹¹C]DOPA is reported. The [¹¹C]benzaldehyde and [¹¹C]veratraldehyde prepared in a two-step reaction from the corresponding [¹¹C]acid salt and [¹¹C]alcohol, by means of selective oxidation with tetrabutylammonium hydrogen chromate, were reacted with 2-phenyl-5-oxazolone or 2-(4-chloro)phenyl-5-oxazolone in the presence of a tertiary amine to give the corresponding [α-¹¹C]-4-arylene-2-aryl-5-oxazolones. Ring opening of these olefins, hydrogenation, and removal of protecting groups was carried out in one step using hydroiodic acid/phosphorus, with the production of the racemic [3-¹¹C]amino acids in 8–30% radiochemical yield (starting with ¹¹CO₂) within 52–60 min (including LC separation).     

Enzymatic synthesis of [1-11C]pyruvic acid, L-[1-11C]lactic acid and L-[1-11C]alanine via DL-[1-11C]alanine

L-[1-11C]Lactic acid was prepared enzymatically from [1-11C]pyruvic acid by way of DL-[1-11C]alanine, using remote, semiautomated procedures. The DL isomers of alanine were prepared by a modification of the Bucherer-Strecker reaction from no-carrier-added (NCA) hydrogen [11C]cyanide. The enantiomer mixture was transformed to [1-11C]pyruvic acid by successive elution through columns of (a) immobilized D-amino acid oxidase (D-AAO)/catalase and (b) immobilized L-alanine dehydrogenase (L-AID) or L-amino acid oxidase (L-AAO/catalase). [1-11C]-Pyruvic acid was subsequently converted to L-[1-11C]lactic acid by passage through a L-lactic dehydrogenase (L-LDH) column. L-[1-11C]Alanine and [1-11C]-pyruvic acid were separated chromatographically by way of a cation-exchange column (AG50W-X2, H+ form). Typically the synthesis time was 35-40 min after cyclotron production of hydrogen [11C]cyanide (400 mCi), with radiochemical yields of 25 mCi (25%) for L-[1-11C]lactic acid, 35 mCi (29%) for [1-11C]pyruvic acid, and 20 mCi (20%) for L-[1-11C]alanine. The use of immobilized enzymes eliminates the possibility of protein contamination and assures the production of sterile, pyrogen-free products, allowing for rapid and effective regio- and stereo-specific transformations.     

An automated radiosynthesis of 2-[11C]thymidine using anhydrous [11C]urea derived from [11C]phosgene.

2-[11C]Thymidine has been produced from [11C]methane via [11C]phosgene and [11C]urea. Anhydrous [11C]urea was prepared from [11C]phosgene by reaction with liquid ammonia. This novel approach avoids the problems associated with the synthesis of anhydrous [11C]urea from [11C]cyanide. A fully automated system based on a modular approach and under PLC control has been developed. The system provides 2-[11C]thymidine reliably and reproducibly for clinical PET studies. The radiosynthesis takes 45-50 min from [11C]methane and the average yield was 1.5-3.3 GBq (40-90 mCi). The specific radioactivity was typically in the range 29.6-51.8 GBq mumol-1 (0.8-1.4 Ci mumol-1) at EOS corresponding to 6-12 micrograms of stable thymidine. The radiochemical yield of 2-[11C]thymidine was ca. 14% from [11C]methane.     

Comparison of N-[(11)C]methyl-norchloroepibatidine and N-[(11)C]methyl-2-(2-pyridyl)-7-azabicyclo[2.2.1]heptane with N-[(11)C]methyl-epibatidine in small animal PET studies

Structural variations of the nicotinic acetylcholine receptor radioligand N-[(11)C]methyl-epibatidine were made to form (11)C-labeled N-methyl-norchloroepibatidine (N-methyl-NorchloroEPB) and N-methyl-2-(2-pyridyl)-7-azabicyclo[2.2.1]heptane (N-methyl-2PABH). Radiosyntheses were performed by methylation with high radiochemical purities (>98%) and with specific activities between 140 and 500 GBq/micromol at the end of synthesis. The radiochemical yield (decay-corrected, related to [(11)C]CH(3)I) was between 5 and 10%. Positively and negatively radiolabeled enantiomers were prepared in high optical purity (>98%ee) by labeling of the appropriate optically active substrates, which were obtained via chiral high performance liquid chromatography. For in vivo studies radioligands were administered intravenously in rats. Brain uptake curves were acquired and combined with blocking experiments. Brain uptake of N-[(11)C]methyl-NorchloroEPB was similar to that of N-[(11)C]methyl-EPB whereas N-[(11)C]methyl-2PABH with the modified pyridine ring had a significantly lower uptake.     

Kinetic analysis of 2-[11C]thymidine PET imaging studies: validation studies.

2-[11C]thymidine has been tested as a PET tracer of cellular proliferation. We have previously described a model of thymidine and labeled metabolite kinetics for use in quantifying the flux of thymidine into DNA as a measure of tumor proliferation. We describe here the results of studies to validate some of the model's assumptions and to test the model's ability to predict the time course of tracer incorporation into DNA in tumors. METHODS: Three sets of studies were conducted: (a) The uptake of tracers in proliferative tissues of normal mice was measured early after injection to assess the relative delivery of thymidine and metabolites of thymidine catabolism (thymine and CO2) and calculate relative blood-tissue transfer rates (relative K1s). (b) By using sequential injections of [11C]thymidine and [11C]thymine in normal human volunteers, the kinetics of the first labeled metabolite were measured to determine whether it was trapped in proliferating tissue such as the bone marrow. (c) In a multitumor rat model, 2-[14C]thymidine injection, tumor sampling and quantitative DNA extraction were performed to measure the time course of label uptake into DNA for comparison with model predictions. RESULTS: Studies in mice showed consistent relative delivery of thymidine and metabolites in somatic tissue but, as expected, showed reduced delivery of thymidine and thymine in the normal brain compared to CO2. Thymine studies in volunteers showed only minimal trapping of label in bone marrow in comparison to thymidine. This quantity of trapping could be explained by a small amount of fixation of labeled CO2 in tissue, a process that is included as part of the model. Uptake experiments in rats showed early incorporation of label into DNA, and the model was able to fit the time course of uptake. CONCLUSION: These initial studies support the assumptions of the compartmental model and demonstrate its ability to quantify thymidine flux into DNA by using 2-[11C]thymidine and PET. Results suggest that further work will be necessary to investigate the effects of tumor heterogeneity and to compare PET measures of tumor proliferation to in vitro measures of proliferation and to clinical tumor behavior in patients undergoing therapy.     

Syntheses of racemic [1-11C]-alanine and partially resolved [3-11C]-alanine

Irradiation of N₂(g) with 10 MeV protons gave [¹¹C]-carbon dioxide which was used as such in the synthesis of racemic [1-¹¹C]-alanine. Alternatively it was transformed to [¹¹C]-methyl iodide for the asymmetric synthesis of [3-¹¹C]-alanine, giving the L(+) form in excess, the ratio L(+)/D(−) being 2.9. The syntheses were completed in 20 and 60 min, respectively.