Simon M. Ametamey

Targeting of renal carcinoma with 67/64Cu-labeled anti-L1-CAM antibody chCE7: selection of copper ligands and PET imaging.

In order to optimize radiocopper labeling of anti-L1-CAM antibody chCE7, five bifunctional copper chelators were synthesized and characterized (CPTA-N-hydoxysuccinimide, DO3A-L-p-isothiocyanato-phenylalanine, DOTA-PA-L-p-isocyanato-phenylalanine, DOTA-glycyl-L-p-isocyanato-phenylalanine and DOTA-triglycyl-L-p-isocyanato-phenylalanine). Substitution with more than 11 chelators per antibody molecule was found to influence immunoreactivity and biodistributions of (67)Cu-MAb chCE7 significantly. CPTA-labeled antibody achieved the best tumor to normal tissue ratios when biodistributions of the different (67)Cu-chCE7 conjugates were assessed in tumor-bearing mice. High resolution PET imaging with (64)Cu-CPTA-labeled MAb chCE7 showed uptake in lymph nodes and heterogeneous distribution in tumor xenografts.

Association of 82 Rubidium and 11C-methionine uptake in brain tumors measured by positron emission tomography

SummaryPositron emission tomography (PET) studies have indicated that alteration of active transport contributes to increased net amino acid accumulation into human brain tumors. We compared the uptake of 11C-methionine (MET) and the K+ analog 82Rubidium (RUB) in 30 patients suffering from various brain tumors using PET. MET and RUB accumulated rapidly in tumor tissue and remained on average at a stable level thereafter from which normalized uptake values were calculated (tissue radioactivity over injected radioactivity × body weight (NU)). K1 (RUB) and K1, k2, 0 (MET) were also estimated using non-linear rate constant fitting in 17/30 patients. NU and K1 values were significantly correlated for MET (Spearman Rank p < 0.005) and RUB (p < 0.001). NU and K1 values for MET and RUB were higher in meningiomas compared to gliomas and were significantly correlated for the whole spectrum of tumors (p < 0.001). When meningiomas were excluded, the correlation was maintained. K3 values for MET (metabolic rate) in tumors were in the range of normal brain. No correlation between RUB and MET was found for normal brain. With increasing RUB uptake, the ratio of NU MET over NU RUB approached the value of 1.0. These results suggest that apart from active transport, also passive diffusion across the blood-brain barrier (BBB) may account for MET uptake from blood into tumor tissue. This probably limits the use of MET in the differential diagnosis of brain lesions where BBB disruption is present.

NanoPET imaging of [18F]fluoromisonidazole uptake in experimental mouse tumours

PurposeThe purpose of this study was to assess the potential and utility of ultra-high-resolution hypoxia imaging in various murine tumour models using the established hypoxia PET tracer [18F]fluoromisonidazole ([18F]FMISO).Methods[18F]FMISO PET imaging was performed with the dedicated small-animal PET scanner NanoPET (Oxford Positron Systems) and ten different human tumour xenografts in nude mice as well as B16 melanoma tumours in syngeneic Balb/c mice. For comparison, [18F]fluorodeoxyglucose ([18F]FDG) PET scans were also performed in the mice bearing human tumour xenografts.ResultsIn 10 out of 11 experimental tumour models, [18F]FMISO PET imaging allowed clear-cut visualisation of the tumours. Inter- and intratumoural heterogeneity of tracer uptake was evident. In addition to average TMRR (tumour-to-muscle retention ratio including all voxels in a volume of interest (VOI)), the parameters TMRR75% and TMRR5 (tumour-to-muscle retention ratio including voxels of 75% or more of the maximum radioactivity in a VOI and the five hottest pixels, respectively) also served as measures for quantifying the heterogeneous [18F]FMISO uptake in the tumours. The variability observed in [18F]FMISO uptake was related neither to tumour size nor to the injected mass of the radiotracer. The pattern of normoxic and hypoxic regions within the human tumour xenografts, however, correlated with glucose metabolism as revealed by comparison of [18F]FDG and [18F]FMISO images.ConclusionThis study demonstrates the feasibility and utility of [18F]FMISO for imaging murine tumour models using NanoPET.

Electrophysiological study, biodistribution in mice, and preliminary PET evaluation in a rhesus monkey of 1-amino-3-[18F]fluoromethyl-5-methyl-adamantane (18F-MEM): a potential radioligand for mapping the NMDA-receptor complex.

The effect of the fluorinated memantine derivative and NMDA receptor antagonist, 1-amino-3-fluoromethyl-5-methyl-adamantane (19F-MEM), at the NMDA receptor ion channel was studied by patch clamp recording. The results showed that 19F-MEM is a moderate NMDA receptor channel blocker. A procedure for the routine preparation of the 18F-labelled analog 18F-MEM has been developed using a two-step reaction sequence. This involves the no-carrier-added nucleophilic radiofluorination of 1-[N-(tert-butyloxy)carbamoyl]-3-(toluenesulfonyloxy)methyl- 5-methyl-adamantane and the subsequent cleavage of the BOC-protecting group using aqueous HCI. The 18F-MEM was obtained in 22 +/- 7% radiochemical yield (decay-corrected to EOB) in a total synthesis time including HPLC purification of 90 min. A biodistribution study after i.v. injection of 18F-MEM in mice showed a fast clearance of radioactivity from blood and relatively high initial uptake in the kidney and in the lung, which gradually decreased with time. The brain uptake was high (up to 3.6% ID/g, 60 min postinjection) with increasing brain-blood ratios: 2.40, 5.10, 6.33, and 9.27 at 5, 30, 60, and 120 min, respectively. The regional accumulation of the radioactivity in the mouse brain was consistent with the known distribution of the PCP recognition site. Preliminary PET evaluation of the radiotracer in a rhesus monkey demonstrated good uptake and prolonged retention in the brain, with a plateau from 35 min onwards p.i. in the NMDA receptor-rich regions (frontal cortex, striata, and temporal cortex). Delineation of the hippocampus, a region known to contain a high density of NMDA receptors, was not possible owing to the resolution of the PET tomograph. The regional brain uptake of 18F-MEM was changed by memantine and by a pharmacological dose of (+)-MK-801, indicating competition for the same binding sites. In a preliminary experiment, haloperidol, a dopamine D2 and sigma receptor antagonist, decreased the binding of 18F-MEM from the brain regions examined, suggesting that binding was also occurring to the sigma recognition sites.

Synthesis and preliminary in vitro evaluation of a new memantine derivative 1-amino-3-[18F]fluoromethyl-5-methyl-adamantane: A potential ligand for mapping the N-methyl-D-aspartate receptor complex

The new memantine derivative 1-amino-3-[18F]fluoromethyl-5-methyl-adamantane (18F-MEM) was prepared in a two-step reaction sequence for evaluation as a PET tracer. This involves the no-carrier-added nucleophilic radiofluorination of 1-[N-(tert-butyloxy)-carbamoyl]-3-(toluenesulfonyloxy)methyl-5-methyl-adamantane (3) with K18F/Kryptofix 2.2.2 in DMSO and the subsequent deprotection of the resulting 18F-BOC-MEM by addition of aqueous HCl. 18F-MEM was obtained after purification by reversed phase HPLC in 22±7 % radiochemical yield (decay corrected to EOB) with a radiochemical purity > 99% and a total synthesis time of 100 min. 18F-MEM is stable up to 6 h in aqueous solution at room temperature and revealed appropriate lipophilicity for good diffusion through the blood-brain-barrier. In vitro studies with the non-radioactive analog, 1-amino-3-fluoromethyl-5-methyl-adamantane (19F-MEM) indicated that this compound binds selectively to the phencyclidine (PCP) binding site within the NMDA receptor complex.

Efficient synthesis of enantiomerically pure thioester precursors of [11c]McN-5652 from racemic McN-5652

An improved synthesis of the anantiomerically pure thioester precursors of [11C](+)-McN-5652 ([11C](+)−1]), and [11C](−)-McN5652 ([11C](−)−1]) starting from racemic McN-5652 ((±)−1) is described. The synthetic method includes the resolution of (±)−1 by repeated crystallization of the (+)- and (−)-di-p-toluoyltartrates yielding (+)-McN-5652 ((+)−1) and (−)-McN-5652 ((−)-1), each with >98% enantiomeric purity. S-Demethylation of (±)−1, (+)−1, and (−)-1, respectively was achieved by treatment with sodium amide at low temperatures (−78°C) followed by conversion of the intermediate thiols 2 with acetyl chloride to give the corresponding thioester precursors (±)−3, (+)−3 or (−)−3. This demethylation method almost completely suppressed the isomerization of the pharmacologically active trans diastereomer into the inactive cis form. Chiral HPLC analyses confirmed that the S-demethylation proceeded without any racemization. 11C-Labelling of (+)−3 or (−)−3, yields enantiomerically pure [11C](+)-McN-5652 or [11C](−)-McN5652, each in 22% radiochemical yield (decay-corrected, related to [11C]CO2) and a specific radioactivity of 74 GBq/μmol (2 Ci/μmol) at the end of synthesis (EOS) Copyright

Radiosynthesis of [11C]brofaromine, a potential tracer for imaging monoamine oxidase A

Brofaromine (4-5(-methoxy-7-bromobenzofuranyl)-2-piperidine-HCl) is a potent and selective inhibitor of monoamine oxidase (MAO) A. Two methods for its synthesis and a preliminary positron emission tomography (PET) evaluation in monkey brain are described. The first method, at low carrier concentration of CO2, consisted of direct O-methylation of (4-(5-hydroxy-7-bromobenzofuranyl)-2-piperidine). The total radiochemical yield achieved ranged from 30 to 50% (from end of bombardment [EOB] and decay corrected) with an overall synthesis time of 45 min. The second approach, with high carrier amounts of CO2 arising from inherent target problems, was accomplished in a three-step route involving protection of secondary amino functionality, O-methylation and deprotection. The total radiochemical yield was 10% (from EOB and decay corrected) with a total synthesis time of 70 min. For both methods methylation was achieved using the classical methylating agent [11C]CH3I, and radiochemical purity was higher than 98%. PET evaluation of the radioligand in a Rhesus monkey showed a high uptake of radioactivity in the brain. Using the irreversible MAO-A inhibitor clorgyline and reversible MAO-A inhibitors moclobemide and brofaromine, three blockade experiments were designed to determine the extent of specific binding of [11C]brofaromine to MAO-A. No apparent decrease in accumulation of radioactivity in the monkey brain was observed when compared to a baseline scan.

In Vivo properties of N-(2-aminoethyl)-5-halogeno-2-pyridinecarboxamide 18F- and 123I-labelled reversible inhibitors of monoamine oxidase B

The reversible and highly selective monoamine oxidase B (MAO-B) inhibitor Ro 19-6327, a picolinic acid derivative, was selected for the development of new radiopharmaceuticals, whereby in place of Cl either 123I or 18F was introduced. The respective labelling procedures have been described earlier. In this study, some metabolic properties were investigated. Blood and urine samples were analysed, and halogenated picolinylglycine, a more hydrophilic compound, was identified as the main metabolite. This shows that the amine is oxidised to the respective carboxylate, but the intermediate imine or aldehyde that was proposed earlier could not be detected. First experiments with single photon emission tomography and positron emission tomography (PET) showed that the iodo compound can be used to investigate MAO-B in vivo while the fluoro compound is accumulated in the brain to such a low degree that no PET studies can be performed. We conclude that the main reason for the poor uptake of the fluoro compound is its lower lipophilicity as compared to the iodo compound and, to a lesser degree, its metabolism, which is similar for both compounds.

123I-MSP and F[11C]MSP: New selective 5-HT2A receptor radiopharmaceuticals for in vivo studies of neuronal 5-HT2 serotonin receptors. Synthesis, in vitro binding study with unlabelled analogues and preliminary in vivo evaluation in mice

In vitro binding study on bovine brain membranes using [3H]SCH23390, [3H]spiperone, [3H]prazosin and [3H]RP62203 as radioligands (for D1, D2, alpha1 and 5-HT2A receptors respectively) indicate that the new butyrophenones 8-[3-(4-fluorobenzoyl)propyl]-1-methyl-1,3,8-triazaspiro[4,5]de can-4-one (FMSP) and 8-[3-(4-iodobenzoyl)propyl]-1-methyl-1,3,8-triazaspiro[4,5]deca n-4-one (IMSP) exhibit a significantly higher selectivity for the 5-HT2A over D1, D2 and alpha1 receptors. Consequently, the radiolabelled analogues F[11C]MSP and 123I-MSP were prepared in attempt to obtain potential radiopharmaceuticals for in vivo imaging of neuronal 5-HT2A receptors with positron emission tomography (PET) and single photon emission tomography (SPET). F[11C]MSP was synthesized by reaction of [11C]CH3I with 8-[3-(4-fluorobenzoyl)propyl]-1,3,8-triazaspiro[4,5]decan-4- one (DMSP) in 12 +/- 3% radiochemical yield, whereas 123I-MSP was obtained in 82 +/- 8% radiochemical yield by a no-carrier-added Cu(I)-assisted [123I]iododebromination of 8-[3-(4-bromo-benzoyl)propyl]-1-methyl-1,3,8-triazaspiro[4,5]de can-4-ene (BrMSP). In vivo pharmacokinetic and brain binding characterization of 123I-MSP assessed in mice following intravenous injection, showed a fast clearance of 123I-MSP from blood and relatively high initial uptakes in the liver, kidneys and in the lung. Significant uptake and long retention were observed in the brain (up to 1.64% i.d., 60 min p.i.), with a regional accumulation of radioactivity consistent with the reported 5-HT2A receptors distribution in the brain. Frontal cortex to cerebellum ratio of 3.5 was calculated at 60 min p.i. Furthermore, the initial brain uptake was significantly reduced after pretreatment of the animals with ritanserin, a selective 5-HT2 antagonist, and by preinjection of the non-radiolabelled analog IMSP, thus indicating the specificity of the brain uptake. These data suggest that 123I-MSP may be a promising compound for studying the serotoninergic 5-HT2 receptors with SPET. Due to the low specific activity of F[11C]MSP currently obtained by the [11C]methylation reaction, systematic in vivo investigation of F[11C]MSP are as yet not feasable.

11C-radiolabeling of hallucinogenic psilocin, a potential radioligand for studying the role of serotonin receptors in psychotic symptom formation

The desmethyl compound, 4-hydroxy-N-methyltryptamine (4), was synthesized via a four-step reaction sequence starting from 4-benzyloxyindole (1). Psilocin (4-hydroxy-N,N-dimethyltryptamine), an indole hallucinogen, was labeled by N-monomethylation of the side chain using the classical methylating agent [ 11 C]CH 3 I in 20±5% (decay corrected from [ 11 C]CH 3 I) radiochemical yield. Specific activities obtained ranged from 900-2300Ci/mmol at EOS (end of synthesis). The radiosynthesis, semi-preparative HPLC and formulation were completed in an average time of 45 minutes.