Automated radiosynthesis of [18F]FPEB with clinically viable yields for human use

404 Objectives: A potent and selective mGluR5 inhibitor, [18F]3-fluoro-5-(pyridin-2-ylethynyl)benzonitrile, [18F]FPEB has excellent properties as a central nervous system PET probe. It has been used in numerous clinical studies to evaluate mGluR5 functions in neurological disorders. The production of clinically useful quantities of [18F]FPEB is presently a challenge as the literature procedures demonstrate low radiochemical yielding (1-5% uncorrected) by microwave as well as conventional heating of commercial available precursors (Hamill et al., 2005, Wong et al., 2013, Sullivan, et al., 2013, Wang, et al. 2007). Herein, we present an automated process for the radiosynthesis of [18F]FPEB using a commercially available GE TracerlabTM FXFN module. With our newly validated method, we demonstrate significant reproducible radiochemical yields of [18F]FPEB. Methods: We optimized the reaction conditions using 3-nitro precursor, 1 from ABX with different 18F-eluting reagents and solvents (Table 1). The optimal nucleophilic fluorination of the precursor, 3-nitro-5-(pyridin-2-ylethynyl)benzonitrile, 1 was dissolved in DMSO and treated with azeotropically dried potassium cryptand [18F]fluoride (K[18F]/K222) at 130oC for 5 minutes followed by semi-preparative RP-HPLC purification on a Gemini C6-phenyl 5µ 10x250mm; 32:68 MeCN: 0.2% acetic acid in water; 5mL/min (Scheme 1, Figure 1). The fraction of the semi-preparative mobile phase containing the major radiochemical final product (tR ~23 min) was diluted with 23 mL water contains 2 mL of 8.4% sodium bicarbonate solution, then trapped and released from HLB light SepPak using 1 mL ethanol and 10 mL of 0.9% sodium chloride for injection. Results: Eight different reaction conditions using DMF and DMSO as reaction solvent with either Kryptofix®/potassium carbonate or tetrabutylammonium bicarbonate (TBA HCO3) for F-18 elution were tested and detailed in Table 1. Based on our optimized reaction conditions, three consecutive productions of [18F]FPEB were carried out to validate this radiopharmaceutical for human use. All batches passed quality control according to FDA/USP standards. Therefore, we were able to disclose a new, simple and at the same time high yielding production of radioligand, [18F]FPEB using the commercially available TRACERlab™ FXFN radiosynthesis module (Figure 2). Using the new procedures, demand for this research ligand can be sustainably supported in-house even in larger PET centers by a single production batch. HPLC analysis of formulated product revealed high radiochemical (>98%) and high chemical purity. Formulated [18F]FPEB maintained stability, as measured by radio-HPLC and TLC (Figure 3), and met specifications for visual inspection and pH over a period of 6 hours. Conclusions: [18F]FPEB was validated for human use with a GE TRACERlab™ FXFN radiosynthesis module. The methodology described herein can facilitate multi-center trials and widespread use of this radiopharmaceutical for imaging mGluR5 receptors in vivo. The automated radiosynthesis of [18F]FPEB provides the radiotracer in a form that is ready for injection and in an uncorrected radiochemical yield of 9±2% (relative to starting [18F]fluoride), with a high specific activity of 4600±1200 mCi/μmol at the end of synthesis (63 min; n = 3). Acknowledgements We acknowledge NIH grants: 1R01EB021708 and 1R01NS100164 for generously providing funding for this research. We also thank David F. Lee Jr., Tim Beaudoin, Danielle Vesper and Steven Gallo of the Massachusetts General Hospital for isotope production and technical support.