Allen F. Brooks

Late-stage [18F]fluorination: new solutions to old problems

The last 2-3 years have seen numerous relationships develop between organometallic chemists, fluorine chemists and PET Centers around the world. These collaborations have led to the development of many new strategies for the late-stage introduction of fluorine-18 into complex bioactive molecules. In this perspective we highlight recent developments and key milestones since 2011.

Copper-Catalyzed [18F]Fluorination of (Mesityl)(aryl)iodonium Salts

A practical, rapid, and highly regioselective Cu-catalyzed radiofluorination of (mesityl)(aryl)iodonium salts is described. This protocol utilizes [18F]KF to access 18F-labeled electron-rich, -neutral, and -deficient aryl fluorides under a single set of mild conditions. This methodology is applied to the synthesis of protected versions of two important radiotracers: 4-[18F]fluorophenylalanine and 6-[18F]fluoroDOPA.

Synthesis of [18F]Arenes via the Copper-Mediated [18F]Fluorination of Boronic Acids

A copper-mediated radiofluorination of aryl- and vinylboronic acids with K18F is described. This method exhibits high functional group tolerance and is effective for the radiofluorination of a range of electron-deficient, -neutral, and -rich aryl-, heteroaryl-, and vinylboronic acids. This method has been applied to the synthesis of [18F]FPEB, a PET radiotracer for quantifying metabotropic glutamate 5 receptors.

Evaluation of [11C]N-Methyl Lansoprazole as a Radiopharmaceutical for PET Imaging of Tau Neurofibrillary Tangles

[(11)C]N-Methyl lansoprazole ([(11)C]NML, 3) was synthesized and evaluated as a radiopharmaceutical for quantifying tau neurofibrillary tangle (NFT) burden using positron emission tomography (PET) imaging. [(11)C]NML was synthesized from commercially available lansoprazole in 4.6% radiochemical yield (noncorrected RCY, based upon [(11)C]MeI), 99% radiochemical purity, and 16095 Ci/mmol specific activity (n = 5). Log P was determined to be 2.18. A lack of brain uptake in rodent microPET imaging revealed [(11)C]NML to be a substrate for the rodent permeability-glycoprotein 1 (PGP) transporter, but this could be overcome by pretreating with cyclosporin A to block the PGP. Contrastingly, [(11)C]NML was not found to be a substrate for the primate PGP, and microPET imaging in rhesus revealed [(11)C]NML uptake in the healthy primate brain of ∼1600 nCi/cc maximum at 3 min followed by rapid egress to 500 nCi/cc. Comparative autoradiography between wild-type rats and transgenic rats expressing human tau (hTau +/+) revealed 12% higher uptake of [(11)C]NML in the cortex of brains expressing human tau. Further autoradiography with tau positive brain samples from progressive supranuclear palsy (PSP) patients revealed colocalization of [(11)C]NML with tau NFTs identified using modified Bielschowsky staining. Finally, saturation binding experiments with heparin-induced tau confirmed K d and Bmax values of [(11)C]NML as 700 pM and 0.214 fmol/μg, respectively.

High affinity radiopharmaceuticals based upon lansoprazole for PET imaging of aggregated tau in Alzheimer's disease and progressive supranuclear palsy: synthesis, preclinical evaluation, and lead selection.

Abnormally aggregated tau is the hallmark pathology of tauopathy neurodegenerative disorders and is a target for development of both diagnostic tools and therapeutic strategies across the tauopathy disease spectrum. Development of carbon-11- or fluorine-18-labeled radiotracers with appropriate affinity and specificity for tau would allow noninvasive quantification of tau burden using positron emission tomography (PET) imaging. We have synthesized [(18)F]lansoprazole, [(11)C]N-methyl lansoprazole, and [(18)F]N-methyl lansoprazole and identified them as high affinity radiotracers for tau with low to subnanomolar binding affinities. Herein, we report radiosyntheses and extensive preclinical evaluation with the aim of selecting a lead radiotracer for translation into human PET imaging trials. We demonstrate that [(18)F]N-methyl lansoprazole, on account of the favorable half-life of fluorine-18 and its rapid brain entry in nonhuman primates, favorable kinetics, low white matter binding, and selectivity for binding to tau over amyloid, is the lead compound for progression into clinical trials.

Copper-Mediated Radiofluorination of Arylstannanes with [18F]KF

A copper-mediated nucleophilic radiofluorination of aryl- and vinylstannanes with [18F]KF is described. This method is fast, uses commercially available reagents, and is compatible with both electron-rich and electron-deficient arene substrates. This method has been applied to the manual synthesis of a variety of clinically relevant radiotracers including protected [18F]F-phenylalanine and [18F]F-DOPA. In addition, an automated synthesis of [18F]MPPF is demonstrated that delivers a clinically validated dose of 200 ± 20 mCi with a high specific activity of 2400 ± 900 Ci/mmol.

Evolution of eukaryal tRNA-guanine transglycosylase: insight gained from the heterocyclic substrate recognition by the wild-type and mutant human and Escherichia coli tRNA-guanine transglycosylases

The enzyme tRNA-guanine transglycosylase (TGT) is involved in the queuosine modification of tRNAs in eukarya and eubacteria and in the archaeosine modification of tRNAs in archaea. However, the different classes of TGTs utilize different heterocyclic substrates (and tRNA in the case of archaea). Based on the X-ray structural analyses, an earlier study [Stengl et al. (2005) Mechanism and substrate specificity of tRNA-guanine transglycosylases (TGTs): tRNA-modifying enzymes from the three different kingdoms of life share a common catalytic mechanism. Chembiochem, 6, 1926–1939] has made a compelling case for the divergent evolution of the eubacterial and archaeal TGTs. The X-ray structure of the eukaryal class of TGTs is not known. We performed sequence homology and phylogenetic analyses, and carried out enzyme kinetics studies with the wild-type and mutant TGTs from Escherichia coli and human using various heterocyclic substrates that we synthesized. Observations with the Cys145Val (E. coli) and the corresponding Val161Cys (human) TGTs are consistent with the idea that the Cys145 evolved in eubacterial TGTs to recognize preQ1 but not queuine, whereas the eukaryal equivalent, Val161, evolved for increased recognition of queuine and a concomitantly decreased recognition of preQ1. Both the phylogenetic and kinetic analyses support the conclusion that all TGTs have divergently evolved to specifically recognize their cognate heterocyclic substrates.

Cu-Mediated C-H 18F-Fluorination of Electron-Rich (Hetero)arenes

This communication describes a method for the nucleophilic radiofluorination of electron-rich arenes. The reaction involves the initial C(sp2)–H functionalization of an electron-rich arene with MesI(OH)OTs to form a (mesityl)(aryl)iodonium salt. This salt is then used in situ in a Cu-mediated radiofluorination with [18F]KF. This approach leverages the stability and availability of electron-rich arene starting materials to enable mild late-stage radiofluorination of toluene, anisole, aniline, pyrrole, and thiophene derivatives. The radiofluorination has been automated to access a 41 mCi dose of an 18F-labeled nimesulide derivative in high (2800 ± 700 Ci/mmol) specific activity.

Synthesis of Diverse 11C-Labeled PET Radiotracers via Direct Incorporation of [11C]CO2

Three new positron emission tomography (PET) radiotracers of interest to our functional neuroimaging and translational oncology programs have been prepared through new developments in [(11)C]CO2 fixation chemistry. [(11)C]QZ (glutaminyl cyclase) was prepared via a tandem trapping of [(11)C]CO2/intramolecular cyclization; [(11)C]tideglusib (glycogen synthase kinase-3) was synthesized through a tandem trapping of [(11)C]CO2 followed by an intermolecular cycloaddition between a [(11)C]isocyanate and an isothiocyanate to form the 1,2,4-thiadiazolidine-3,5-dione core; [(11)C]ibrutinib (Brutons tyrosine kinase) was synthesized through a HATU peptide coupling of an amino precursor with [(11)C]acrylic acid (generated from [(11)C]CO2 fixation with vinylmagnesium bromide). All radiochemical syntheses are fully automated on commercial radiochemical synthesis modules and provide radiotracers in 1-5% radiochemical yield (noncorrected, based upon [(11)C]CO2). All three radiotracers have advanced to rodent imaging studies and preliminary PET imaging results are also reported.

A fully-automated one-pot synthesis of [18F]fluoromethylcholine with reduced dimethylaminoethanol contamination via [18F]fluoromethyl tosylate.

INTRODUCTION A novel one-pot method for preparing [(18)F]fluoromethylcholine ([(18)F]FCH) via in situ generation of [(18)F]fluoromethyl tosylate ([(18)F]FCH2OTs), and subsequent [(18)F]fluoromethylation of dimethylaminoethanol (DMAE), has been developed. METHODS [(18)F]FCH was prepared using a GE TRACERlab FXFN, although the method should be readily adaptable to any other fluorine-(18) synthesis module. Initially ditosylmethane was fluorinated to generate [(18)F]FCH2OTs. DMAE was then added and the reaction was heated at 120 °C for 10 min to generate [(18)F]FCH. After this time, reaction solvent was evaporated, and the crude reaction mixture was purified by solid-phase extraction using C(18)-Plus and CM-Light Sep-Pak cartridges to provide [(18)F]FCH formulated in USP saline. The formulated product was passed through a 0.22 µm filter into a sterile dose vial, and submitted for quality control testing. Total synthesis time was 1.25 h from end-of-bombardment. RESULTS Typical non-decay-corrected yields of [(18)F]FCH prepared using this method were 91 mCi (7% non-decay corrected based upon ~1.3 Ci [(18)F]fluoride), and doses passed all other quality control (QC) tests. CONCLUSION A one-pot liquid-phase synthesis of [(18)F]FCH has been developed. Doses contain extremely low levels of residual DMAE (31.6 µg/10 mL dose or ~3 ppm) and passed all other requisite QC testing, confirming their suitability for use in clinical imaging studies.