title: In vitro binding characteristics of [3H]AZ11637326, a novel alpha7-selective neuronal nicotinic receptor agonist radioligand.
abstract: AZ11637326 (5'-(2-fluoro[3,4,5(-3)H3]phenyl)-spiro[1-azabicyclo [2.2.2]octane-3,2'(3'H)-furo[2,3-b]pyridine) is a potent partial agonist at the human alpha7 neuronal nicotinic receptor with sub-nanomolar affinity for the human and rat alpha7 [(125)I]alpha-bungarotoxin binding sites. In a search for novel agonist radioligands and imaging ligands for the alpha7 nicotinic receptor, [(3)H]AZ11637326 was synthesized and its in vitro membrane binding properties were characterized. [(3)H]AZ11637326 bound to halpha7-HEK membranes with high specificity (>95%), high affinity (230 pM) and a B(max) of 460 fmol/mg. The rank order of affinity of nicotinic standards determined with [(3)H]AZ11637326 strongly correlated with those determined using the classical alpha7 antagonist [(125)I]alpha-bungarotoxin, indicating that [(3)H]AZ11637326 bound to halpha7-HEK membranes with an alpha7 nicotinic-like pharmacology. The K(i) values for the standards were on average 2.3-fold lower affinity than determined using the prototypical alpha7 nicotinic antagonist [(125)I]alpha-bungarotoxin. Because [(3)H]AZ11637326 specific binding is rapid and reversible, the K(i) values determined using this ligand are more accurate estimates of affinity than those determined using the kinetically sluggish [(125)I]alpha-bungarotoxin. [(3)H]AZ11637326 also bound to a high affinity (510 pM), nicotine-sensitive site on rat hippocampal membranes with an average B(max) of 55 fmol/mg. With rat hippocampal membranes, the nicotine-sensitive fraction of total binding was sub-optimal for a radioligand (~50%), yet the potencies and rank order of the K(i) values for standards were consistent with an alpha7 nicotinic pharmacology. Overall, these studies indicate that [(3)H]AZ11637326 is a useful new in vitro probe of the alpha7 nicotinic receptor agonist site and support its potential utility for in vivo receptor occupancy studies.
authors.affiliation: CNS Discovery, AstraZeneca, Wilmington, Delaware 19850-5437, USA. john.gordon@astrazeneca.com
authors.forename: John C
authors.initials: JC
authors.lastname: Gordon
authors.forename: Eifion
authors.initials: E
authors.lastname: Phillips
authors.forename: David A
authors.initials: DA
authors.lastname: Gurley
authors.forename: J Richard
authors.initials: JR
authors.lastname: Heys
authors.forename: Lois Ann
authors.initials: LA
authors.lastname: Lazor
authors.forename: Herbert G
authors.initials: HG
authors.lastname: Barthlow
authors.forename: Mike A
authors.initials: MA
authors.lastname: Mallamaci
authors.forename: Rich A
authors.initials: RA
authors.lastname: Keith
mesh_terms: D000818:Animals; D053961:Azabicyclo Compounds / Q000378:metabolism / Q000494:pharmacology*; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D001952:Bridged-Ring Compounds / Q000378:metabolism / Q000494:pharmacology; D002038:Bungarotoxins / Q000378:metabolism / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D008297:Male; D009474:Neurons / Q000378:metabolism; D018722:Nicotinic Agonists / Q000378:metabolism / Q000494:pharmacology*; D018733:Nicotinic Antagonists / Q000378:metabolism / Q000494:pharmacology; D011869:Radioligand Assay; D051381:Rats; D017207:Rats, Sprague-Dawley; D011978:Receptors, Nicotinic / Q000378:metabolism*; D013141:Spiro Compounds / Q000378:metabolism / Q000494:pharmacology*; D064569:alpha7 Nicotinic Acetylcholine Receptor
chemical_list: C555067:5'-(2-fluorophenyl)-spiro(1-azabicyclo(2.2.2)octane-3,2'(3'H)-furo(2,3-b)pyridine); C408128:AR-R 17779; D053961:Azabicyclo Compounds; D001952:Bridged-Ring Compounds; D002038:Bungarotoxins; C579167:Chrna7 protein, human; C579165:Chrna7 protein, rat; D008024:Ligands; D018722:Nicotinic Agonists; D018733:Nicotinic Antagonists; D011978:Receptors, Nicotinic; D013141:Spiro Compounds; D064569:alpha7 Nicotinic Acetylcholine Receptor