Cancer is the leading cause of premature deaths in the western world, a circumstance that has spurred the development of new radiopharmaceuticals for cancer diagnosis by positron emission tomography (PET) and for radiotherapy using α- and β-emitting radionuclides (Holland et al., 2012; Zhang et al., 2017). For use in clinical PET imaging, the radiohalogen fluorine-18 (18F) is the preferred radioisotope due to its physical half-life (109.7 min), favorable decay properties, and its ready availability in multi-GBq quantities from medical cyclotrons. A considerable proportion of emerging radiopharmaceuticals are based on small to medium sized peptides, fusion proteins, and endogenous proteins. However, these products are not always obtainable through a direct radiolabeling approach due to their instability at high temperature or the otherwise harsh reaction conditions required to introduce fluorine-18 into their structure. Although labeling of such molecules with fluorine-18 using a chelator conjugated to the targeting moiety with [18F]AlF (aluminum [18F]fluoride) is feasible, this approach can release free [18F]fluoride in vivo (which invariably results in intense bone labelling) and otherwise alter the biodistribution properties of the radiopharmaceutical in an unpredictable fashion (Lütje et al., 2019).
A robust and high yield radiosynthesis amenable to automation is a prerequisite for adopting a PET radiopharmaceutical for widespread clinical or research use (Li et al., 2013). Production of PET radiopharmaceuticals with a multi-step synthetic process can be a challenging task using commercially produced cassette-based modules, mainly due to their lack of modifiability, and the frequent requirement for intermediate or final HPLC purification. 18F-labelled tracers for prostate specific membrane antigen (PSMA) [18F]DCFPyL and [18F]-PSMA-1007 are examples of PET radiopharmaceuticals that formerly required a two-step radiosynthesis from the prosthetic group 6-[18F]fluoronicotinic acid‐2,3,5,6‐tetrafluorophenyl ester ([18F]F‐Py‐TFP) (Chen et al., 2011; Giesel et al., 2017). In the case of [18F]DCFPyL, the first reported radiosynthesis from [18F]F‐Py‐TFP and the PSMA-avid substrate Glu-CO-Lys protected with t-butyl has subsequently been optimized (Chen et al., 2011). Currently, a direct precursor labeling with the trimethylammonium nicotinic acid moiety conjugated to the ε-nitrogen of the lysine residue of Glu-CO-Lys is the favored method for automated multi-dose production of [18F]DCFPyL (Dornan et al., 2018), which has greatly facilitated the use of PSMA PET in clinical routine. However, this introduction of fluoride-18 in organic solvents at a temperature approaching 100°C, is unlikely to provide high labeling efficiencies, or to generalize for larger molecular weight radiopharmaceuticals such as polypeptides and proteins with their bountiful functional groups.
Fairly recently, a simplistic and rapid preparation of [18F]F-Py‐TFP has been reported, wherein the prosthetic group is obtained directly by passing its precursor in a solution of acetonitrile/tert-butanol through a polymeric anion‐exchange cartridge preloaded with [18F]fluoride (Basuli et al., 2016; Olberg, 2017). This procedure yields the bifunctional labeling agent in good yield, with an overall synthesis time of less than ten minutes (EOB), simultaneously avoiding the use of a reaction vessel and a phase transfer catalysts. The on-column radiosynthesis of [18F]F‐Py‐TFP has a very small laboratory footprint and lends itself well for automation and offer radiolabeling of a wide variety of sensitive biomolecules (proteins eg.) (Roy et al., 2019; Lesniak et al., 2019). Expanding on this concept, we report a fully automated, two-step radiosynthesis of [18F]DCFPyL from [18F]F‐Py-TFP and the Glu-CO-Lys pseudo-peptide precursor, which can serve as a blueprint manufacturing process for the preparation of other 18F-labelled radiopharmaceuticals. The approach uses [18F]F‐Py‐TFP produced on column adopted into the commercially available, cassette-based synthesis module FASTlabTM. The automated process implements a hydrophilic interaction liquid chromatography (HILIC) SPE cartridge purification methodology, making final preparative HPLC purification redundant.