An azeotropic drying-free approach for copper-mediated radiofluorination without addition of base.

Copper-mediated radiofluorination provides a quick and versatile approach for 18 F-labeling of arenes and heteroarenes. However, this method is known to be base sensitive, which has been a barrier for preparative scale radiosynthesis. In this report, we provide an approach for copper-mediated radiofluorination without azeotropic drying or adding a base. [18 F]Fluoride trapped on a PS-HCO3 Sep-Pak was quantitatively eluted with a solution of 4-dimethylaminopyridinium trifluoromethanesulfonate (DMAP·OTf) in anhydrous N,N-dimethylformamide (DMF). The eluted solution was directly used for copper-mediated radiofluorination. Twelve boronic ester substrates were tested, yielding fluorinated products in 27% to 83% radiochemical yield based on HPLC analysis. This approach was successfully applied to the radiosynthesis of [18 F]flumazenil, a well-known positron emission tomography (PET) tracer for imaging central benzodiazepine receptors, with a radiochemical yield of 47%. This highly efficient protocol significantly augments the powerful copper-mediated radiofluorination approach.

Routine production of [(18)f]flumazenil from iodonium tosylate using a sample pretreatment method: a 2.5-year production report.

PURPOSE: [(18)F]Flumazenil, which has the advantage of a longer half-life than [(11)C]flumazenil, is well known for determining of the central benzodiazepine receptor concentrations. However, [(18)F]flumazenil has not been widely used because fluctuating and relatively low yields render automatic production insufficient for routine and multicenter clinical trials. Here, we describe the results of a 2.5-year production study of [(18)F]flumazenil using an iodonium tosylate precursor, which allowed us to overcome the limitations of low and fluctuating radiochemical yields. PROCEDURES: We developed a clinically applicable production system by modifying a commercial synthesizer for the reliable and reproducible production of [(18)F]flumazenil for routine clinical studies. [(18)F]Flumazenil was prepared at 150 °C for 5 min in the presence of 4-methylphenyl-mazenil iodonium tosylate (4 mg), a radical scavenger (TEMPO, 1 mg), and [(18)F]KF/kryptofix 2.2.2 complex in N,N-dimethylformamide (1 ml). In the purification step, the final mixture was pretreated using different cartridges before performing high-performance liquid chromatography (HPLC) separation. Finally, we measured the radiochemical yield and performed quality-control assays on 94 batches. RESULTS: After carrying out additional purification before HPLC separation using a C18 plus Sep-Pak cartridge, the radiochemical yield of [(18)F]flumazenil increased from 34.4 ± 9.7 % (without the pretreatment, n = 24) to 53.4 ± 9.0 % (n = 94), and the lifetime of the semi-preparative column was five times that of the column without the C18 plus Sep-Pak cartridge. The mean-specific activity of [(18)F]flumazenil was 572 ± 116 GBq/µmol at the end of synthesis, and the radiochemical purity was more than 99 %, as determined by analytical HPLC and radio-TLC. [(18)F]Flumazenil prepared using this method satisfied all quality-control test standards and was highly stable for up to 6 h after preparation. CONCLUSIONS: The results of the 2.5-year production study using an iodonium tosylate precursor indicate that [(18)F]flumazenil has commercial and routine clinical applicability.

Facile aromatic radiofluorination of [18F]flumazenil from diaryliodonium salts with evaluation of their stability and selectivity.

Aromatic radiofluorination of the diaryliodonium tosylate precursor with [(18)F]fluoride ions has been applied successfully to access [(18)F]flumazenil in high radiochemical yields of 67.2 ± 2.7% (decay corrected). The stability and reactivity of the diaryliodonium tosylate precursor plays a key role in increasing the production of (18)F-labelled molecules under the fluorine-18 labelling condition. Various conditions were explored for the preparation of [(18)F]flumazenil from different diaryliodonium tosylate precursors. Optimum incorporation of [(18)F]fluoride ions in the 4-methylphenyl-mazenil iodonium tosylate precursor (5f) was achieved at 150 °C for 5 min by utilizing 4 mg of the precursor, K(2.2.2)/K(2)CO(3) complex, and the radical scavenger in N,N-dimethylformamide. This approach was extended to a viable method for use in automated synthesis with a radiochemical yield of 63.5 ± 3.2% (decay corrected, n = 26) within 60.0 ± 1.1 min. [(18)F]Flumazenil was isolated by preparative HPLC after the reaction was conducted under improved conditions and exhibited sufficient specific activity of 370-450 GBq µmol(-1), with a radiochemical purity of >99%, which will be suitable for human PET studies.

GABAergic dysfunction in essential tremor: an 11C-flumazenil PET study.

UNLABELLED: Essential tremor is the most common movement disorder, but the underlying pathophysiology is not well understood. A primary overactivity of cerebellothalamic output pathways is the most conspicuous finding, as indicated by animal and human studies. It has been argued that this overactivity may be due to impaired central inhibition, and converging evidence points toward a potential role of gamma-aminobutyric acid (GABA) dysfunction in tremor generation. METHODS: Using (11)C-flumazenil and PET, we calculated the distribution volume, an index of availability of benzodiazepine receptor sites of the GABA(A) complex, in a group of 8 patients with bilateral essential tremor, as compared with 11 healthy controls. RESULTS: Significant increases in binding of (11)C-flumazenil at the benzodiazepine receptor site of the GABA(A) receptor in the cerebellum, the ventrolateral thalamus, and the lateral premotor cortex were identified in the essential tremor group. CONCLUSION: Essential tremor is associated with reduced GABAergic function and increased availability of benzodiazepine receptor sites in brain regions implicated specifically in tremor genesis. This finding is thought to reflect overactivity of cerebellothalamic circuits and, hence, lends support to the "GABA hypothesis" of essential tremor.

Improved work-up procedure for the production of [(18)F]flumazenil and first results of its use with a high-resolution research tomograph in human stroke.

INTRODUCTION: The central benzodiazepine receptor (cBZR)-gamma-aminobutyric acid (GABA(A)) receptor complex in the human brain plays an important role in many neurological and psychiatric disorders. (18)F-Labeled flumazenil ([(18)F]FZ) provides a potentially useful tracer to investigate those disorders by means of positron emission tomography (PET). METHODS: [(18)F]Flumazenil was synthesized from its nitro-precursor Ro 15-2344 in DMF at high temperatures between 150 degrees C and 160 degrees C. Other solvents like acetonitrile and dimethylsulfoxide were also investigated as reaction media. A new HPLC method for the final purification of [(18)F]FZ was developed to circumvent some difficulties associated with a previously published procedure sometimes led to a contamination of [(18)F]FZ with Ro 15-2344. The final purification of the radiotracer was achieved using a Waters Symmetry Prep C18 HPLC column with elution with 0.05 M sodium acetate (NaOAc) buffer (pH 5)/THF/MeOH (80:10:10). RESULTS: [(18)F]FZ could be synthesized in reproducible radiochemical yields (RCYs) of 15-20% (decay corrected to EOB) after 80 min overall synthesis time. The synthesized [(18)F]FZ was applied for the first time in a human PET study in a patient with ischemic right middle cerebral artery stroke using the HRRT high-resolution research scanner (Siemens Medical Solution, Knoxville, TN, USA). CONCLUSIONS: [(18)F]FZ is a potentially useful GABA receptor-binding PET ligand. A modified procedure for its preparation in reproducibly high radiochemical yields has been described and the [(18)F]FZ thus produced has been used successfully in a pilot clinical study.

Novel one-pot one-step synthesis of 2'-[(18)F]fluoroflumazenil (FFMZ) for benzodiazepine receptor imaging.

We describe the synthesis of 2'-[(18)F]fluoroflumazenil (FFMZ), which differs from the typically used [(18)F]fluoroethylflumazenil (FEFMZ) for benzodiazepine receptor imaging. For one-pot one-step labeling, the precursors, 2'-tosyloxyflumazenil (TFMZ) and 2'-mesyloxyflumazenil (MFMZ), were synthesized in three steps. The precursors were successfully labeled with no-carrier-added (18)F-fluoride which was activated by repeated azeotropic distillation with Kryptofix 2.2.2./potassium carbonate in MeCN. An automated system for labeling and purification of [(18)F]FFMZ was developed. Labeling efficiency and radiochemical purity of [(18)F]FFMZ after synthesis by the automated system were 68% and 98%, respectively. Specific binding of [(18)F]FFMZ to central benzodiazepine receptor of rats was demonstrated by phosphoimaging.

Synthesis and PET imaging of the benzodiazepine receptor tracer [N-methyl-11C]iomazenil.

The central benzodiazepine receptor tracer [N-methyl-11C]iomazenil (Ro 16-0154) was synthesized by alkylation of the desmethyl precursor noriomazenil with [11C]methyl iodide. The [11C]CH3I (prepared by reduction of [11C]CO2 with LiA1H4 followed by reaction with HI) was reacted with noriomazenil in N,N-dimethylformamide and Bu4N+OH- for 1 min at 80 degrees C and purified by HPLC (C18, 34% CH3CN/H2O 7 mL/min). The product was obtained with synthesis time 35 +/- 5 min (mean +/- SD, n = 7), radiochemical yield (EOB) 36 +/- 16%, radiochemical purity 99 +/- 1%, and specific activity 5100 +/- 2800 mCi/mumol. Absorbed radiation doses were calculated from previously acquired human biodistribution data. The urinary bladder wall received the highest dose (0.099 mGy/MBq) for 4.8 h voiding interval and the effective dose equivalent was 0.015 mSv/MBq. After i.v. injection of [11C]iomazenil in an adult baboon or healthy human volunteer, radioactivity accumulated in the cortex with time-activity curves in agreement with results obtained with [11C]flumazenil PET and [123I]iomazenil SPECT studies. The count rate was sufficient to obtain quantitative images up to 2 h post-injection with a 14 mCi injection. These results suggest that [11C]iomazenil will be a useful agent for measuring benzodiazepine receptors in vivo by positron emission tomography.

Plasma pharmacokinetics and metabolism of the benzodiazepine antagonist [11C] Ro 15-1788 (flumazenil) in baboon and human during positron emission tomography studies.

Flumazenil is a specific antagonist of the central benzodiazepine receptor (CBZR). Labelled with 11C, this compound is the reference radioligand for positron emission tomography (PET) study of the CBZR in humans and primates. The time-course of [11C]-flumazenil radioactivity and its main acid metabolite [11C] Ro 15-3890 were reconstructed from the time-course of total radioactivity in plasma after administration with high or low SRA in primates and humans, applying an extraction procedure validated by TLC. The measured pharmacokinetics of [11C]-flumazenil (T1/2 beta = 45.1 +/- 12.3 min, T1/2 alpha = 1.5 +/- 1.5 min; K = 0.14 +/- 0.14 min-1; Vd area = 44.0 +/- 17.0 l; Clp = 40.0 +/- 8.5 l/h) exhibited a very rapid distribution phase followed by fast elimination, with a large volume of distribution; these results were confirmed by HPLC determinations and agree with previously published data on unlabelled flumazenil. Pharmacokinetics of [11C] Ro 15-3890 acid metabolite show that high drug concentrations in the blood are promptly achieved (kf = 0.13 +/- 0.004 min-1), with a very rapid elimination half-life (T1/2m = 4.47 +/- 1.31 min) comparable to that of [11C]-flumazenil. The percentage metabolization of parent compound to the acid [11C] Ro 15-3890 was constant from the 15th minute and was significantly higher in man compared to the monkey. This percentage was increased by prior eating. The other putative metabolites, i.e. labelled [11C] Ro 15-4965 and unlabelled Ro 15-5528, were never observed at detectable concentrations with TLC and HPLC in rabbit, baboon and human blood samples. This pharmacokinetic study of plasma flumazenil may be useful to implement a dynamic method of CBZR quantification using PET and for analysis of pharmacokinetics in brain tissue.

The utility of 1-[18F]fluoro-3-iodopropane for the synthesis of certain dopamine D-1 and benzodiazepine receptor radioligands.

No-carrier-added (NCA) R(+)-7-chloro-8-hydroxy-3-(3'-[18F]fluoropropyl)-1-phenyl-2,3,4,5- tetrahydro-3-benzazepine (2b) (an analog of dopamine D-1 receptor ligand SCH 23390), ethyl 8-fluoro-5,6-dihydro-5-(3'-fluoropropyl)-6-oxo-4H- imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate (4b) and 3'-[18F]fluoropropyl 8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H- imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate (6b) (analogs of the benzodiazepine RO 15-1788) were synthesized by alkylation of the corresponding nor-compound with NCA 1-[18F]fluoro-3-iodopropane in 10-15% yield (EOB) in approximately 110 min and with a mass of 2-3 nmol. Compound 2 is less potent (approximately 12-14 times) than SCH 23390 in binding to rat striatal membranes in vitro. Compounds 2b, 4b and 6b exhibit no specific anatomical distribution to mouse brain. These results suggest that the substituent at position 3 of SCH 23390, and position 5 and carboxylate group of RO 15-1788 are critical determinants both of affinity and selectivity for receptor binding, and underscores the evaluation necessary when even minor changes (C1 to C3) are made in bioactive compounds.