Metabolic Stability of the Demyelination Positron Emission Tomography Tracer [18F]3-Fluoro-4-Aminopyridine and Identification of Its MetabolitesS

[18F]3-fluoro-4-aminopyridine ([18F]3F4AP) is a positron emission tomography (PET) tracer for imaging demyelination based on the multiple sclerosis drug 4-aminopyridine (4AP, dalfampridine). This radiotracer was found to be stable in rodents and nonhuman pri-mates imaged under isoflurane anesthesia. However, recent find-ings indicate that its stability is greatly decreased in awake humans and mice. Since both 4AP and isoflurane are metabolized primarily by cytochrome P450 enzymes, particularly cytochrome P450 family 2 subfamily E member 1 (CYP2E1), we postulated that this enzyme may be responsible for the metabolism of 3F4AP. Here, we investi-gated the metabolism of [18F]3F4AP by CYP2E1 and identified its metabolites. We also investigated whether deuteration, a common approach to increase the stability of drugs, could improve its stabil-ity. Our results demonstrate that CYP2E1 readily metabolizes 3F4AP and its deuterated analogs and that the primary metabolites are 5-hydroxy-3F4AP and 3F4AP N-oxide. Although deuteration did not decrease the rate of the CYP2E1-mediated oxidation, our findings explain the diminished in vivo stability of 3F4AP compared with 4AP and further our understanding of when deuteration may improve the metabolic stability of drugs and PET ligands