Fastlab

[¹⁸F]FB-Tz was synthesised using the GE FASTLab™ automated radiosynthesis platform via reductive amination radiochemistry between precursor 3 and 4-[¹⁸F]fluorobenzaldehyde ([¹⁸F]FBA) (Scheme 1). We described the automated procedure elsewhere;^{21 (link)} in brief, aqueous [¹⁸F]fluoride was dried for the radiosynthesis of [¹⁸F]FBA from the 4-formyl-N,N,N-trimethylanilinium trifluoromethansulfonate precursor and used without further purification. Compound 3 was dissolved in acetonitrile by the addition of triethylamine to quench the TFA-salt, then added to the vessel containing [¹⁸F]FBA. The imine intermediate was formed at 65 °C for 15 min, before the reaction mixture was flowed through a cartridge containing solid-supported cyanoborohydride to reduce the imine to the secondary amine. [¹⁸F]FB-Tz was purified by semi-preparative HPLC (Shimadzu LC20-AT pump attached to a custom-built system, equipped with an Agilent Eclipse XDB-C18, 5μ (250 × 9.4 mm) column. The mobile phase was 20% EtOH/80% sodium phosphate (58 mM, pH 2.4) at a flow rate of 4 mL min⁻¹) and the desired product was diluted in water (30 mL), trapped on a tC18 SPE cartridge, dried under a flow of nitrogen and eluted in EtOH for biological use. [¹⁸F]FB-Tz was characterised by radio-HPLC to determine its identity (by comparison to an authentic reference standard) and purity (Fig. 2B and C).

The previously reported radiosynthesis of ¹⁸F-FPIA was adapted to prepare GMP grade radiopharmaceutical using the GE FASTLab™ automated radiosynthesis platform [11 , 13 ]. A detailed description of the automated radiosynthesis is described in the supplementary information (Supplementary material and methods S1). In brief, the precursor methyl 2,2-dimethyl-3-[(4-methylbenzenesulfonyl)oxy] propanoate [1 (link)] was radiolabelled by displacement of the tosylate group with dry ¹⁸F-fluoride to produce intermediate ¹⁸F-2, the methyl ester of ¹⁸F-FPIA (Scheme S1). Compound ¹⁸F2 was hydrolysed under basic conditions to give ¹⁸F-FPIA which was purified by semi-preparative HPLC using biocompatible solvents (15% EtOH, 85% sodium dihydrogen phosphate buffer, pH 4.5). The fraction containing ¹⁸F-FPIA was diluted in water and passed through a sterile filter into a sterile vial for clinical use. The identity and purity (chemical and radiochemical purity) of the final product were determined by HPLC; other quality control tests were performed, according to European Pharmacopoeia guidelines (Table S1).

[¹⁸F]FRPG and [¹⁸F]FSPG were synthesized from their corresponding naphthalenesulfonyl precursors (Life Molecular Imaging) using a GE FASTlab automated synthesis module using a previously reported method 43 (link). The analysis and identification of [¹⁸F]FRPG and [¹⁸F]FSPG was also conducted as reported previously 43 (link). For a detailed description of precursor and ¹⁹F reference standard syntheses, along with their corresponding spectra, see Supplementary Information: Synthesis and Spectra. The radiochemical purity and molar activity of [¹⁸F]FRPG and [¹⁸F]FSPG were analysed by pre-column derivatization with ortho-phthalaldehyde reagent (OPA). 20 µL [¹⁸F]FRPG or [¹⁸F]FSPG solution was added to 20 µL OPA reagent followed by addition of 80 µL PBS. The mixture was incubated for 5 min at room temperature and then analysed by HPLC. Next, the appropriate ¹⁹F standard was added to the OPA/tracer reaction mixture and left for 15 min before further analysis by HPLC. Co-elution of the ¹⁹F standard OPA-adduct UV peak with the corresponding ¹⁸F OPA-adduct radioactive peak confirmed the identity of both [¹⁸F]FRPG and [¹⁸F]FSPG (Figure S1). HPLC chromatograms showing the different retention times for the [¹⁸F]FRPG-OPA and [¹⁸F]FSPG-OPA adducts with co-elution of their corresponding ¹⁹F reference compounds can be found in the supplemental information (Figure S2).