title: Difficulties in dopamine transporter radioligand PET analysis: the example of LBT-999 using [18F] and [11C] labelling: part II: Metabolism studies.
abstract: BACKGROUND
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: CEA, I2BM, Service Hospitalier Frédéric Joliot, F-91406 Orsay, France. marie-anne.schollhorn@shfj.cea.fr
authors.forename: Marie-Anne
authors.initials: MA
authors.lastname: Peyronneau
authors.forename: Wadad
authors.initials: W
authors.lastname: Saba
authors.forename: Frédéric
authors.initials: F
authors.lastname: Dollé
authors.forename: Sébastien
authors.initials: S
authors.lastname: Goutal
authors.forename: Christine
authors.initials: C
authors.lastname: Coulon
authors.forename: Michel
authors.initials: M
authors.lastname: Bottlaender
authors.forename: Héric
authors.initials: H
authors.lastname: Valette
mesh_terms: D000818:Animals; D001921:Brain / Q000000981:diagnostic imaging / Q000378:metabolism; D002250:Carbon Radioisotopes; D002853:Chromatography, Liquid; D003042:Cocaine / Q000031:analogs & derivatives* / Q000097:blood / Q000493:pharmacokinetics; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D050483:Dopamine Plasma Membrane Transport Proteins / Q000378:metabolism; D005462:Fluorine Radioisotopes; D006801:Humans; D008297:Male; D008862:Microsomes, Liver / Q000000981:diagnostic imaging / Q000378:metabolism; D015394:Molecular Structure; D010215:Papio; D049268:Positron-Emission Tomography / Q000379:methods; D011869:Radioligand Assay; D019275:Radiopharmaceuticals / Q000097:blood / Q000493:pharmacokinetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D053719:Tandem Mass Spectrometry
chemical_list: C508666:8-((E)-4-fluoro-but-2-enyl)-3beta-p-tolyl-8-aza-bicyclo(3.2.1)octane-2beta-carboxylic acid methyl ester; D002250:Carbon Radioisotopes; D065607:Cytochrome P-450 Enzyme Inhibitors; D018076:DNA, Complementary; D050483:Dopamine Plasma Membrane Transport Proteins; D005462:Fluorine Radioisotopes; D019275:Radiopharmaceuticals; D003577:Cytochrome P-450 Enzyme System; D003042:Cocaine