title: Characterization of a selective and potent antagonist of human P2X(7) receptors, AZ11645373.
abstract: OBJECTIVE
  The ATP-gated P2X(7) receptor has been shown to play a role in several inflammatory processes, making it an attractive target for anti-inflammatory drug discovery. We have recently identified a novel set of cyclic imide compounds that inhibited P2X(7) receptor-mediated dye uptake in human macrophage THP-1 cells. In this study the actions and selectivity of one of these compounds, AZ11645373, were characterized.
  
  
  METHODS
  We measured membrane currents, calcium influx, and YOPRO-1 uptake from HEK cells expressing individual P2X receptors, and YOPRO1 uptake and interleukin-1beta release from THP-1 cells in response to ATP and the ATP analogue benzoylbenzoyl ATP (BzATP).
  
  
  RESULTS
  AZ11645373 up to 10 microM, had no agonist or antagonist actions on membrane currents due to P2X receptor activation at human P2X(1), rat P2X(2), human P2X(3), rat P2X(2/3), human P2X(4), or human P2X(5) receptors expressed in HEK cells. AZ11645373 inhibited human P2X(7) receptor responses in HEK cells in a non-surmountable manner with K (B) values ranging from 5 - 20 nM, with mean values not significantly different between assays. K (B) values were not altered by removing extracellular calcium and magnesium. ATP-evoked IL-1beta release from lipopolysaccharide-activated THP-1 cells was inhibited by AZ11645373, IC(50) = 90 nM. AZ11645373 was > 500-fold less effective at inhibiting rat P2X(7) receptor-mediated currents with less than 50% inhibition occurring at 10 microM.
  
  
  CONCLUSIONS
  AZ11645373 is a highly selective and potent antagonist at human but not rat P2X(7) receptors and will have much practical value in studies of human cells.
authors.affiliation: Department of Biomedical Science, University of Sheffield, Sheffield, UK.
authors.forename: L
authors.initials: L
authors.lastname: Stokes
authors.forename: L-H
authors.initials: LH
authors.lastname: Jiang
authors.forename: L
authors.initials: L
authors.lastname: Alcaraz
authors.forename: J
authors.initials: J
authors.lastname: Bent
authors.forename: K
authors.initials: K
authors.lastname: Bowers
authors.forename: M
authors.initials: M
authors.lastname: Fagura
authors.forename: M
authors.initials: M
authors.lastname: Furber
authors.forename: M
authors.initials: M
authors.lastname: Mortimore
authors.forename: M
authors.initials: M
authors.lastname: Lawson
authors.forename: J
authors.initials: J
authors.lastname: Theaker
authors.forename: C
authors.initials: C
authors.lastname: Laurent
authors.forename: M
authors.initials: M
authors.lastname: Braddock
authors.forename: A
authors.initials: A
authors.lastname: Surprenant
mesh_terms: D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000814:Aniline Compounds; D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D001583:Benzoxazoles; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D005456:Fluorescent Dyes; D006801:Humans; D007094:Imides / Q000494:pharmacology*; D053583:Interleukin-1beta / Q000378:metabolism; D015640:Ion Channel Gating / Q000187:drug effects; D008070:Lipopolysaccharides / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D058919:Purinergic P2 Receptor Antagonists; D011806:Quinolinium Compounds; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000378:metabolism; D058486:Receptors, Purinergic P2X7; D013045:Species Specificity; D013844:Thiazoles / Q000737:chemistry / Q000494:pharmacology*; D014162:Transfection; D014966:Xanthenes
chemical_list: C516674:AZ 11645373; D000814:Aniline Compounds; D000893:Anti-Inflammatory Agents; D001583:Benzoxazoles; C409648:Fluo 4; D005456:Fluorescent Dyes; D007094:Imides; D053583:Interleukin-1beta; D008070:Lipopolysaccharides; C546468:P2RX7 protein, human; C546470:P2rx7 protein, rat; D058919:Purinergic P2 Receptor Antagonists; D011806:Quinolinium Compounds; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D013844:Thiazoles; D014966:Xanthenes; C089813:YO-PRO 1; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate