title: Negative and positive allosteric modulators of the P2X(7) receptor.
abstract: OBJECTIVE
  Antagonist effects at the P2X(7) receptor are complex with many behaving in a non-competitive manner. In this study, the effects of N-[2-({2-[(2-hydroxyethyl)amino]ethyl}amino)-5-quinolinyl]-2-tricyclo[3.3.1.1(3,7)]dec-1-ylacetamide (compound-17) and N (2)-(3,4-difluorophenyl)-N (1)-[2-methyl-5-(1-piperazinylmethyl)phenyl]glycinamide dihydrochloride (GW791343) on P2X(7) receptors were examined and their mechanism of action explored.
  
  
  METHODS
  Antagonist effects were studied by measuring agonist-stimulated ethidium accumulation in cells expressing human or rat recombinant P2X(7) receptors and in radioligand binding studies.
  
  
  RESULTS
  Compound-17 and GW791343 were non-competitive inhibitors of human P2X(7) receptors. Receptor protection studies using decavanadate and pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS) showed that neither compound-17 nor GW791343 competitively interacted at the ATP binding site and so were probably negative allosteric modulators of the P2X(7) receptor. GW791343 prevented the slowly reversible blockade of the human P2X(7) receptor produced by compound-17 and inhibited [(3)H]-compound-17 binding to the P2X(7) receptor suggesting they may bind to similar or interacting sites. At rat P2X(7) receptors, compound-17 was a negative allosteric modulator but the predominant effect of GW791343 was to increase agonist responses. Antagonist interaction and radioligand binding studies revealed that GW791343 did not interact at the ATP binding site but did interact with the compound-17 binding site suggesting that GW791343 is a positive allosteric modulator of the rat P2X(7) receptor.
  
  
  CONCLUSIONS
  Compound-17 was a negative allosteric modulator of human and rat P2X(7) receptors. GW791343 was a negative allosteric modulator of the human P2X(7) receptor but at the rat P2X(7) receptor its predominant effect was positive allosteric modulation. These compounds should provide valuable tools for mechanistic studies on P2X(7) receptors.
authors: Neurology Centre of Excellence for Drug Discovery, GlaxoSmithKline Research & Development Limited, Harlow, Essex, UK. anton.d.michel@gsk.com
authors: A D
authors: AD
authors: Michel
authors: L J
authors: LJ
authors: Chambers
authors: D S
authors: DS
authors: Walter
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D000494:Allosteric Regulation; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004996:Ethidium / Q000378:metabolism; D005998:Glycine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D015394:Molecular Structure; D010879:Piperazines / Q000378:metabolism / Q000494:pharmacology*; D011487:Protein Conformation; D011732:Pyridoxal Phosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D011804:Quinolines / Q000378:metabolism / Q000494:pharmacology*; D011869:Radioligand Assay; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D058486:Receptors, Purinergic P2X7; D011994:Recombinant Proteins / Q000378:metabolism; D013045:Species Specificity; D013997:Time Factors; D014162:Transfection; D014638:Vanadates / Q000378:metabolism / Q000494:pharmacology
chemical_list: C528279:GW791343; C528278:N-(2-((2-((2-hydroxyethyl)amino)ethyl)amino)-5-quinolinyl)-2-tricyclo(3.3.1.1(3,7))dec-1-ylacetamide; C546468:P2RX7 protein, human; C546470:P2rx7 protein, rat; D010879:Piperazines; D011804:Quinolines; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D011994:Recombinant Proteins; C077792:pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid; D014638:Vanadates; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; D011732:Pyridoxal Phosphate; C063302:KN 62; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000255:Adenosine Triphosphate; D004996:Ethidium; D005998:Glycine