Automation of Copper-Mediated 18F-Fluorination of Aryl Pinacol Boronates Using 4-Dimethylaminopyridinium Triflate.

Currently, the copper-mediated radiofluorination of aryl pinacol boronates (arylBPin) using the commercially available, air-stable Cu(OTf)2Py4 catalyst is one of the most efficient synthesis approaches, greatly facilitating access to a range of radiotracers, including drug-like molecules with nonactivated aryl scaffolds. Further adjustment of this methodology, in particular, the [18F]fluoride recovery step for the routine preparation of radiotracers, has been the focus of recent research. In our recent study, an organic solution of 4-dimethylaminopyridinium trifluoromethanesulfonate (DMAPOTf) was found to be an efficient PTC for eluting radionuclides retained on the weak anion exchange cartridge, Oasis WAX 1cc, employing the inverse sorption-elution protocol. Notably, the following Cu-mediated radiofluorination of arylBPin precursors in the presence of the Cu(OTf)2(Py)4 catalyst can be performed with high efficiency in the same solvent, bypassing not only the conventional azeotropic drying procedure but any solvent replacement. In the current study, we aimed to translate this methodology, originally developed for remote-controlled operation with manual interventions, into the automated synthesis module on the TRACERlab automation platform. The adjustment of the reagent amounts and solvents allowed for high efficiency in the radiofluorination of a series of model arylBPin substrates on the TRACERlab FXFE Pro synthesis module, which was adapted for nucleophilic radiofluorinations. The practical applicability of the developed radiofluorination approach with DMAPOTf elution was demonstrated in the automated synthesis of 6-L-[18F]FDOPA. The radiotracer was obtained with an activity yield (AY; isolated, not decay-corrected) of 5.2 ± 0.5% (n = 3), with a synthesis time of ca. 70 min on the TRACERlab FX N Pro automation platform. The obtained AY was comparable with one reported by others (6 ± 1%) using the same boronate precursor, while a slightly higher AY of 6-L-[18F]FDOPA (14.5 ± 0.5%) was achieved in our previous work using commercially available Bu4NOTf as the PTC.

One-Pot Radiosynthesis of [18F]Anle138b-5-(3-Bromophenyl)-3-(6-[18F]fluorobenzo[d][1,3]dioxol-5-yl)-1H-pyrazole-A Potential PET Radiotracer Targeting α-Synuclein Aggregates.

Availability of PET imaging radiotracers targeting α-synuclein aggregates is important for early diagnosis of Parkinson's disease and related α-synucleinopathies, as well as for the development of new therapeutics. Derived from a pyrazole backbone, 11C-labelled derivatives of anle138b (3-(1,3-benzodioxol-5-yl)-5-(3-bromophenyl)-1H-pyrazole)-an inhibitor of α-synuclein and prion protein oligomerization-are currently in active development as the candidates for PET imaging α-syn aggregates. This work outlines the synthesis of a radiotracer based on the original structure of anle138b, labelled with fluorine-18 isotope, eminently suitable for PET imaging due to half-life and decay energy characteristics (97% ß+ decay, 109.7 min half-life, and 635 keV positron energy). A three-step radiosynthesis was developed starting from 6-[18F]fluoropiperonal (6-[18F]FP) that was prepared using (piperonyl)(phenyl)iodonium bromide as a labelling precursor. The obtained 6-[18F]FP was used directly in the condensation reaction with tosylhydrazide followed by 1,3-cycloaddition of the intermediate with 3'-bromophenylacetylene eliminating any midway without any intermediate purifications. This one-pot approach allowed the complete synthesis of [18F]anle138b within 105 min with RCY of 15 ± 3% (n = 3) and Am in the range of 32-78 GBq/µmol. The [18F]fluoride processing and synthesis were performed in a custom-built semi-automated module, but the method can be implemented in all the modern automated platforms. While there is definitely space for further optimization, the procedure developed is well suited for preclinical studies of this novel radiotracer in animal models and/or cell cultures.

Automated Optimized Synthesis of [18F]FLT Using Non-Basic Phase-Transfer Catalyst with Reduced Precursor Amount.

3'-deoxy-3'-[18F]fluorothymidine ([18F]FLT) is a positron emission tomography (PET) tracer useful for tumor proliferation assessment for a number of cancers, particularly in the cases of brain, lung, and breast tumors. At present [18F], FLT is commonly prepared by means of the nucleophilic radiofluorination of 3-N-Boc-5'-O-DMT-3'-O-nosyl thymidine precursor in the presence of a phase-transfer catalyst, followed by an acidic hydrolysis. To achieve high radiochemical yield, relatively large amounts of precursor (20−40 mg) are commonly used, leading to difficulties during purification steps, especially if a solid-phase extraction (SPE) approach is attempted. The present study describes an efficient method for [18F]FLT synthesis, employing tetrabutyl ammonium tosylate as a non-basic phase-transfer catalyst, with a greatly reduced amount of precursor employed. With a reduction of the precursor amount contributing to lower amounts of synthesis by-products in the reaction mixture, an SPE purification procedure using only two commercially available cartridges­OASIS HLB 6cc and Sep-Pak Alumina N Plus Light­has been developed for use on the GE TRACERlab FX N Pro synthesis module. [18F]FLT was obtained in radiochemical yield of 16 ± 2% (decay-corrected) and radiochemical purity >99% with synthesis time not exceeding 55 min. The product was formulated in 16 mL of normal saline with 5% ethanol (v/v). The amounts of chemical impurities and residual solvents were within the limits established by European Pharmacopoeia. The procedure described compares favorably with previously reported methods due to simplified automation, cheaper and more accessible consumables, and a significant reduction in the consumption of an expensive precursor.

Production of 6-l-[18F]Fluoro-m-tyrosine in an Automated Synthesis Module for 11C-Labeling.

6-l-[18F]Fluoro-m-tyrosine (6-l-[18F]FMT) represents a valuable alternative to 6-l-[18F]FDOPA which is conventionally used for the diagnosis and staging of Parkinson's disease. However, clinical applications of 6-l-[18F]FMT have been limited by the paucity of practical production methods for its automated production. Herein we describe the practical preparation of 6-l-[18F]FMT using alcohol-enhanced Cu-mediated radiofluorination of Bpin-substituted chiral Ni(II) complex in the presence of non-basic Bu4ONTf using a volatile iPrOH/MeCN mixture as reaction solvent. A simple and fast radiolabeling procedure afforded the tracer in 20.0 ± 3.0% activity yield within 70 min. The developed method was directly implemented onto a modified TracerLab FX C Pro platform originally designed for 11C-labeling. This method enables an uncomplicated switch between 11C- and 18F-labeling. The simplicity of the developed procedure enables its easy adaptation to other commercially available remote-controlled synthesis units and paves the way for a widespread application of 6-l-[18F]FMT in the clinic.

Rapid 18F-labeling via Pd-catalyzed S-arylation in aqueous medium.

We report radiolabeling of thiol-containing substrates via Pd-catalyzed S-arylation with 2-[18F]fluoro-5-iodopyridine, which is readily accessible using the "minimalist" radiofluorination method. The practicality of the procedure was confirmed by preparation of a novel PSMA-specific PET-tracer as well as labeling of glutathione, Aß oligomer-binding RD2 peptide, bovine serum albumin and PSMA I&S.

Nucleophilic Synthesis of 6-l-[18F]FDOPA. Is Copper-Mediated Radiofluorination the Answer?

Positron emission tomography employing 6-l-[18F]fluoro-3,4-dihydroxyphenylalanine (6-l-[18F]FDOPA) is currently a highly relevant clinical tool for detection of gliomas, neuroendocrine tumors and evaluation of Parkinson's disease progression. Yet, the deficiencies of electrophilic synthesis of 6-l-[18F]FDOPA hold back its wider use. To fulfill growing clinical demands for this radiotracer, novel synthetic strategies via direct nucleophilic 18F-radiloabeling starting from multi-Curie amounts of [18F]fluoride, have been recently introduced. In particular, Cu-mediated radiofluorination of arylpinacol boronates and arylstannanes show significant promise for introduction into clinical practice. In this short review these current developments will be discussed with a focus on their applicability to automation.

Alcohol-Supported Cu-Mediated 18F-Fluorination of Iodonium Salts under "Minimalist" Conditions.

In the era of personalized precision medicine, positron emission tomography (PET) and related hybrid methods like PET/CT and PET/MRI gain recognition as indispensable tools of clinical diagnostics. A broader implementation of these imaging modalities in clinical routine is closely dependent on the increased availability of established and emerging PET-tracers, which in turn could be accessible by the development of simple, reliable, and efficient radiolabeling procedures. A further requirement is a cGMP production of imaging probes in automated synthesis modules. Herein, a novel protocol for the efficient preparation of 18F-labeled aromatics via Cu-mediated radiofluorination of (aryl)(mesityl)iodonium salts without the need of evaporation steps is described. Labeled aromatics were prepared in high radiochemical yields simply by heating of iodonium [18F]fluorides with the Cu-mediator in methanolic DMF. The iodonium [18F]fluorides were prepared by direct elution of 18F- from an anion exchange resin with solutions of the corresponding precursors in MeOH/DMF. The practicality of the novel method was confirmed by the racemization-free production of radiolabeled fluorophenylalanines, including hitherto unknown 3-[18F]FPhe, in 22-69% isolated radiochemical yields as well as its direct implementation into a remote-controlled synthesis unit.

Synthesis and biological evaluation of 2-(3,4-dimethoxyphenyl)-6-(2-[18F]fluoroethoxy)benzothiazole ([18F]FEDBT) for PET imaging of breast cancer.

Given the ever-present demand for improved PET radiotracer in oncology imaging, we have synthesized 2-(3,4-dimethoxyphenyl)-6-(2-[18F]fluoroethoxy)benzothiazole ([18F]FEDBT), a fluorine-18-containing fluoroethylated benzothiazole to explore its utility as a PET imaging tracer. [18F]FEDBT was prepared via kryptofix-mediated nucleophilic substitution of the tosyl group precursor. Fractionated ethanol-based solid-phase (SPE cartridge-based) purification afforded [18F]FEDBT in 60% radiochemical yield (EOB), with radiochemical purity in excess of 98% and the specific activity was 35GBq/µmol. The radiotracer displayed clearly higher cellular uptake ratio in various breast cancer cell lines MCF7, MDA-MB-468 and MDA-MB-231. However, both biodistribution and microPET studies have showed an higher abdominal accumulation of [18F]FEDMBT and the tumor/muscle ratio of 1.8 was observed in the MDA-MB-231 xenograft tumors mice model. Further the lipophilic improvement is needed for the reducement of hepatobilliary accumulation and to promote the tumor uptake for PET imaging of breast cancer.

4-[18F]fluoroglutamic acid (BAY 85-8050), a new amino acid radiotracer for PET imaging of tumors: synthesis and in vitro characterization.

There is a high demand for tumor specific PET tracers in oncology imaging. Besides glucose, certain amino acids also serve as energy sources and anabolic precursors for tumors. Therefore, (18)F-labeled amino acids are interesting probes for tumor specific PET imaging. As glutamine and glutamate play a key role in the adapted intermediary metabolism of tumors, the radiosynthesis of 4-[(18)F]fluoro l-glutamic acid (BAY 85-8050) as a new specific PET tracer was established. Cell-uptake studies revealed specific tumor cell accumulation.

[18F]flumazenil binding to central benzodiazepine receptor studies by PET--quantitative analysis and comparisons with [11C]flumazenil.

[(11)C]flumazenil is the reference radioligand for Positron Emission Tomography (PET) studies of central benzodiazepine (BZ) receptors. Fluorine is available in the flumazenil molecule and [(18)F]flumazenil has recently been prepared. The aim of the present PET-study in 8 male subjects was to examine the binding of [(18)F]flumazenil in the human brain by direct comparison with [(11)C]flumazenil. Each subject participated in two 93-minute PET-measurements with [(11)C]flumazenil and [(18)F]flumazenil, respectively. Data were analyzed using compartment models with metabolite-corrected arterial plasma input and reference tissue models using the pons as reference region. There was no evident difference between the kinetic behaviors of the two ligands. Overall, the noise in the time activity curves for [(18)F]flumazenil was lower at late time points, and the variance of the kinetic parameters was lower than for [(11)C]flumazenil. In BZ receptor rich regions, such as the neocortex, the 3-compartment model was statistically favored, whereas the 2-compartment model was favored in the pons. Binding potential values obtained by the reference tissue models were in good agreement with those obtained by the kinetic analysis. There was no support for the presence of specific binding in the pons. In conclusion, the binding and the kinetic behavior of [(11)C]flumazenil and [(18)F]flumazenil were similar. The present analysis supports the use of pons as reference region in simplified protocols without arterial blood sampling. [(18)F]flumazenil should thus be an excellent choice for applied studies at centers not having a cyclotron.

A fully automated one-pot synthesis of [carbonyl-11C]WAY-100635 for clinical PET applications.

A fully automated synthesis of the important 5HT(1A) receptor radioligand, [carbonyl-(11)C]WAY-100635 (I), was developed based on the optimized one-pot "wet" synthesis procedure. A modern automated apparatus was constructed from commercially available components and operated via LabView software. In average, (906+/-525)MBq (n=94) of (I) was obtained from 40 min bombardment at 50 microA beam current within 50 min synthesis time. The specific radioactivity (SA) at the time of injection was (50.5+/-29.3)GBq/mumol (n=94).

No carrier added synthesis of O-(2'-[18F]fluoroethyl)-L-tyrosine via a novel type of chiral enantiomerically pure precursor, NiII complex of a (S)-tyrosine Schiff base.

O-(2'-[(18)F]fluoroethyl)-l-tyrosine ([(18)F]FET) has gained much attention as a promising amino acid radiotracer for tumor imaging with positron emission tomography (PET) due to favorable imaging characteristics and relatively long half-life of (18)F (110min) allowing remote-site application. Here we present a novel type of chiral enantiomerically pure labeling precursor for [(18)F]FET, based on NiII complex of a Schiff's base of (S)-[N-2-(N'-benzylprolyl)amino]benzophenone (BPB) with alkylated (S)-tyrosine, Ni-(S)-BPB-(S)-Tyr-OCH2CH2X (X=OTs (3a), OMs (3b) and OTf (3c)). A series of compounds 3a-c was synthesized in three steps from commercially available reagents. Non-radioactive FET as a reference was prepared from 3a in a form of (S)-isomer and (R,S) racemic mixture. Radiosynthesis comprised two steps: (1) n.c.a. nucleophilic fluorination of 3a-c (4.5-5.0mg) in the presence of either Kryptofix 2.2.2.or tetrabutylammonium carbonate (TBAC) in MeCN at 80 degrees C for 5min, followed by (2) removal of protective groups by treating with 0.5M HCl (120 degrees C, 5min). The major advantages of this procedure are retention of enantiomeric purity during the (18)F-introduction step and easy simultaneous deprotection of amino and carboxy moieties in 3a-c. Radiochemically pure [(18)F]FET was isolated by semi-preparative HPLC (C18 mu-Bondapak, Waters) eluent aq 0.01M CH(3)COONH(4), pH 4/C(2)H(5)OH 90/10 (v/v). Overall synthesis time operated by Anatech RB 86 laboratory robot was 55min. In a series of compounds 3a-c, tosyl derivative 3a provided highest radiochemical yield (40-45%, corrected for radioactive decay). Enantiomeric purity was 94-95% and 96-97%, correspondingly, for Kryptofix and TBAC assisted fluorinations. The suggested procedure involved minimal number of synthesis steps and suits perfectly for automation in the modern synthesis modules for PET radiopharmaceuticals. Preliminary biodistribution study in experimental model of turpentine-induced aseptic abscess and Glioma35 rat's tumor (homografts) in Wistar rats has demonstrated the enhanced uptake of radiotracer in the tumor area with minimal accumulation in the inflamed tissues.

Preparation of highly specific radioactivity [18F]flumazenil and its evaluation in cynomolgus monkey by positron emission tomography.

A straightforward method for the preparation of no-carrier-added (n.c.a.) [18F]flumazenil via standard nucleophilic radiofluorination of the corresponding nitro-analog Ro 15-2344 has been developed. The labeling was performed by employing the K18F/kryptofix complex in DMF at 160 degrees C for 30 min and equimolar ratio [K/K2.2.2]+18F-/precursor. Under these conditions, an 18F incorporation rate into flumazenil was in the range of 55-60%. The final product was isolated by HPLC purification within a total synthesis time of 75 min and a radiochemical yield of about 30% (EOB). Human post-mortem whole-hemisphere autoradiography of brain sections demonstrated selective uptake of the radioligand in the areas of high density of the central benzodiazepine receptors (BZR). PET studies in a cynomolgus monkey and metabolite studies by HPLC demonstrated similar results by [18F]flumazenil as for [11C]flumazenil. In blocking experiments, almost all radioactivity was inhibited by the addition of unlabeled flumazenil. [18F]Flumazenil is a suitable radioligand for PET assessment of the BZR.