Exploring the Metabolism of (+)-[¹⁸F]Flubatine in vitro and in vivo: LC-MS/MS aided Identification of Radiometabolites in a Clinical PET Study

Both (+)-[18F]flubatine and its enantiomer (-)-[18F]flubatine are radioligands for the neuroimaging of a4ß2 nicotinic acetylcholine receptors (nAChRs) by positron emission tomography (PET). Within a clinical study in patients with early Alzheimer’s disease, (+)-[18F]flubatine ((+)-[18F]1) was examined regarding its metabolic fate, in particular by identification of degradation products detected in plasma and urine. The investigations included an in vivo study of (+)-flubatine ((+)-1) in pig and structural elucidation of formed metabolites by LC-MS/MS. Incubations of (+)-1 and (+)-[18F]1 with human liver microsomes were performed to generate in vitro metabolites as well as radiometabolites, which enabled an assignment of their structures by comparison of LC-MS/MS and radio-HPLC data. Plasma and urine samples taken after administration of (+)-[18F]1 into human were examined by radio-HPLC and, on the basis of results obtained in vitro and in vivo, formed radiometabolites were identified.
In pig, (+)-1 was monohydroxylated at different sites of the azabicyclic ring system of the molecule. Additionally, one intermediate metabolite underwent glucuronidation, as also demonstrated in vitro. In human, 95.9 ? 1.9% (N = 10) of unchanged tracer remained in plasma, 30 min after injection. However, despite the low metabolic degradation, both radiometabolites formed could be characterized as i.) a product of C-hydroxylation at the azabicyclic ring system and ii.) a glucuronide conjugate of previously N8-hydroxylated (+)-[18F]1.