title: Synthesis and evaluation in rats of the dopamine D2/3 receptor agonist 18F-AMC20 as a potential radioligand for PET.
abstract: Dopamine D(2/3) receptor (D(2/3)R) agonist PET tracers are better suited for the imaging of synaptic dopaminergic neurotransmission than D(2/3)R antagonists and may also offer the opportunity to study in vivo the high-affinity state of D(2/3)R (D(2/3)RHigh). With the aim to develop (18)F-labeled D2/3R agonists suitable for widespread clinical application, we report here on the synthesis and in vitro and in vivo evaluation of a D(2/3)R agonist ligand from the aminomethyl chromane (AMC) class-(R)-2-[(4-(18)F-fluorobenzylamino)methyl]chroman-7-ol ((18)F- AMC20: ).
  
  
  METHODS
  In vitro affinities of AMC20: toward dopaminergic receptor subtypes were measured in membrane homogenates prepared from HEK293 cells expressing human dopamine receptors. Agonism of AMC20: was assessed in the arrestin recruitment assay in Chinese hamster ovary-K(1) cells expressing the long isoform of D(2)R (D(2)RLong). D(2/3)R-specific binding of (18)F- AMC20: was evaluated in brain slices of Sprague-Dawley rats by in vitro autoradiography and in living rats by in vivo small-animal PET imaging and ex vivo autoradiography. PET data were analyzed with 1- and 2-tissue compartmental models, the simplified reference tissue model, and Logan graphical analysis. Specificity of binding was tested by blocking D(2/3)R with raclopride (coincubation with 10 μM in vitro, administration of 1.0 mg/kg in vivo).
  
  
  RESULTS
  In membrane homogenates, AMC20: demonstrated picomolar affinity at D(2)RHigh (mean inhibition constant [K(i)] = 85 pM) and excellent selectivity against the low-affinity state of D(2)R (D(2)RLow) (mean K(i) = 84 nM, 988-fold selectivity) and D(1)-like receptors (mean K(i) = 5,062 nM at D1R). The efficacy of AMC20: was 90% of that of dopamine in the arrestin recruitment assay. Up to 70% of total binding of (18)F- AMC20: in the D2/3R-rich striatum in rat brain slices was D(2/3)R-specific; in living rats, the uptake ratio between the striatum and the D(2/3)R-poor cerebellum reached 2.0-2.5, depending on the measurement method. Radiometabolites of (18)F- AMC20: did not enter the brain. Striatal binding potential of (18)F- AMC20: varied between 0.49 and 0.59 depending on the estimation method. Pretreatment with 1 mg of raclopride per kilogram reduced the apparent specific binding of (18)F- AMC20: in the striatum.
  
  
  CONCLUSIONS
  (18)F- AMC20: shows specific binding to D(2/3)R in the striatum of living rats. Further optimization of the chemical structure of (18)F- AMC20: can lead to (18)F-labeled D(2/3) agonist PET tracers more suitable for in vivo clinical application.
authors.affiliation: Department of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Shalgunov
authors.affiliation: Department of Nuclear Medicine, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Jan-Peter
authors.initials: JP
authors.lastname: van Wieringen
authors.affiliation: SyMO-Chem BV, Eindhoven, The Netherlands; and.
authors.forename: Henk M
authors.initials: HM
authors.lastname: Janssen
authors.affiliation: SyMO-Chem BV, Eindhoven, The Netherlands; and.
authors.forename: P Michel
authors.initials: PM
authors.lastname: Fransen
authors.affiliation: Department of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.
authors.forename: Rudi A J O
authors.initials: RA
authors.lastname: Dierckx
authors.affiliation: Department of Pharmacology, Johannes Gutenberg University, Mainz, Germany.
authors.forename: Martin C
authors.initials: MC
authors.lastname: Michel
authors.affiliation: Department of Nuclear Medicine, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: Jan
authors.initials: J
authors.lastname: Booij
authors.affiliation: Department of Nuclear Medicine and Molecular Imaging, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands p.h.elsinga@umcg.nl.
authors.forename: Philip H
authors.initials: PH
authors.lastname: Elsinga
mesh_terms: D000818:Animals; D001578:Benzopyrans / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D001596:Benzylamines / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D001692:Biological Transport / Q000187:drug effects; D001921:Brain / Q000000981:diagnostic imaging / Q000187:drug effects / Q000378:metabolism; D016466:CHO Cells; D060326:Chemistry Techniques, Synthetic; D002839:Chromans / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D018491:Dopamine Agonists / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D008024:Ligands; D008297:Male; D049268:Positron-Emission Tomography / Q000379:methods*; D020891:Raclopride / Q000494:pharmacology; D011854:Radiochemistry; D051381:Rats; D017207:Rats, Sprague-Dawley; D017448:Receptors, Dopamine D2 / Q000819:agonists*; D050637:Receptors, Dopamine D3 / Q000819:agonists*; D013237:Stereoisomerism
chemical_list: C000595795:2-((4-fluorobenzylamino)methyl)chroman-7-ol; D001578:Benzopyrans; D001596:Benzylamines; D002839:Chromans; D018491:Dopamine Agonists; D008024:Ligands; D017448:Receptors, Dopamine D2; D050637:Receptors, Dopamine D3; D020891:Raclopride
keywords: 18F; PET; agonist tracer; dopamine receptor