Difficulties in dopamine transporter radioligand PET analysis: the example of LBT-999 using [18F] and [11C] labelling: Part II: Metabolism studies

Abstract Introduction LBT-999, ( E )- N -(4-fluorobut-2-enyl)-2β-carbomethoxy-3β-(4'-tolyl)nortropane, has been developed for PET imaging of the dopamine transporter. [ 18 F]LBT-999 PET studies in baboons showed a lower brain uptake than [ 11 C]LBT-999 and a high bone uptake, suggesting the presence of interfering metabolites. Therefore, in vitro and in vivo metabolism of these radiotracers was investigated. Methods Rat and human liver microsomal incubations, baboon plasma and rat brain extracts were analyzed by radio-HPLC and LC-MS-MS. Results In vitro experiments demonstrated the formation by P450s of five polar metabolites. The main routes of LBT-999 metabolism proposed were N -dealkylation, tolyl-hydroxylation and dealkylation plus tolyl-hydroxylation. In vivo in baboons, [ 18 F]LBT-999 was rapidly converted into a [ 18 F]hydroxylated metabolite likely oxidized in plasma into a [ 18 F]carboxylic acid and into unlabeled N -dealkyl-LBT-999. The latter was detected in baboon plasma and in rat brain by LC-MS-MS. The time course of unchanged [ 18 F]LBT-999 decreased rapidly in plasma and was higher than that of [ 11 C]LBT-999 due to the formation of unlabeled N -dealkyl-LBT-999. In rats, striatum-to-cerebellum ratios of [ 18 F]LBT-999, [ 18 F]hydroxylated and [ 18 F]acidic metabolite were 20, 4.2 and 1.65, respectively, suggesting a possible accumulation of the hydroxylated compound in the striatum. Conclusion P450s catalyzed the formation of dealkylated and hydroxylated metabolites of LBT-999. In baboons, an extensive metabolism of [ 18 F]LBT-999, with formation of unlabeled N -dealkyl-LBT-999, [ 18 F]fluorobutenaldehyde (or its oxidation product) and [ 18 F]hydroxy-LBT-999 able to penetrate the brain, prevented an easy and accurate estimation of the input function of the radiotracer. CYP3A4 being the main P450 involved in the metabolism of LBT-999, a similar pathway may occur in humans and confound PET quantification.