title: Identification and pharmacological characterization of [125I]L-750,667, a novel radioligand for the dopamine D4 receptor.
abstract: We identified a novel azaindole derivative, L-750,667, that has high affinity (Ki = 0.51 nM) and >2000-fold selectivity for D4 dopamine receptors compared with its activity at D2 and D3 dopamine receptors. L-750,667 had little affinity for rat D1/D5 dopamine receptors, sigma binding sites, or 5-hydroxytryptamine1A or 5-hydroxytryptamine2 receptors. In functional studies, L-750,667 exhibited high affinity antagonist activity at D4 receptors, reversing dopamine (1 microM)-induced inhibition of cAMP accumulation in human embryonic kidney (HEK) cells expressing the human D4 receptor (hD4 HEK) with an EC50 value of 80 nM. The radioiodinated form of L-750,667 bound specifically to the human dopamine D4 receptor expressed in HEK cells and saturation analysis revealed a single high affinity binding site for [125I]L-750,667 (Kd = 0.16 +/- 0.06 nM). The maximum number of binding sites (Bmax) estimated using [125I]L-750,667 in hD4 HEK cells was 251 +/- 71 fmol/mg, which correlated well with the Bmax value determined using [3H]spiperone (227 +/- 83 fmol/mg) in the same membrane preparations. The pharmacological profile of [125I]L-750,667 binding to hD4 HEK cells was evaluated using known dopamine receptor agonists and antagonists. The rank order of potencies for dopamine receptor agonists was dopamine > quinpirole > 6,7-aminodihydroxytetralin > 5,6-aminodihydroxytetralin. Dopamine receptor antagonists also showed high affinity, with a rank order of haloperidol > chlorpromazine > domperidone > (+)-butaclamol > (-)-sulpiride = (+)-sulpiride > (+)-SCH23390 > (-)-butaclamol. [125I]L-750,667, bound to D4 receptors in a stereoselective manner with (+)-butaclamol showing higher activity than its respective enantiomer (-)-butaclamol. These results show that [125I]L-750,667 is a novel, highly selective radioligand for dopamine D4 receptors and may be used to investigate the dopamine D4 receptor population in the central nervous system.
authors.affiliation: Department of Biochemistry and Molecular Biology, Neuroscience Research Centre, Merck Sharp and Dohme Research Laboratories, Harlow, Essex, UK.
authors.forename: S
authors.initials: S
authors.lastname: Patel
authors.forename: S
authors.initials: S
authors.lastname: Patel
authors.forename: R
authors.initials: R
authors.lastname: Marwood
authors.forename: F
authors.initials: F
authors.lastname: Emms
authors.forename: D
authors.initials: D
authors.lastname: Marston
authors.forename: P D
authors.initials: PD
authors.lastname: Leeson
authors.forename: N R
authors.initials: NR
authors.lastname: Curtis
authors.forename: J J
authors.initials: JJ
authors.lastname: Kulagowski
authors.forename: S B
authors.initials: SB
authors.lastname: Freedman
mesh_terms: D000818:Animals; D002460:Cell Line; D018492:Dopamine Antagonists / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007457:Iodine Radioisotopes; D007700:Kinetics; D011725:Pyridines / Q000378:metabolism / Q000494:pharmacology*; D011758:Pyrroles / Q000378:metabolism / Q000494:pharmacology*; D011869:Radioligand Assay; D051381:Rats; D017448:Receptors, Dopamine D2 / Q000187:drug effects* / Q000378:metabolism; D050638:Receptors, Dopamine D4
chemical_list: C493599:DRD4 protein, human; D018492:Dopamine Antagonists; C493601:Drd4 protein, rat; D007457:Iodine Radioisotopes; C103081:L 750667; D011725:Pyridines; D011758:Pyrroles; D017448:Receptors, Dopamine D2; D050638:Receptors, Dopamine D4