title: SK&F 96365, a novel inhibitor of receptor-mediated calcium entry.
abstract: A novel inhibitor of receptor-mediated calcium entry (RMCE) is described. SK&F 96365 (1-(beta-[3-(4-methoxy-phenyl)propoxy]-4-methoxyphenethyl)-1H- imidazole hydrochloride) is structurally distinct from the known 'calcium antagonists' and shows selectivity in blocking RMCE compared with receptor-mediated internal Ca2+ release. Human platelets, neutrophils and endothelial cells were loaded with the fluorescent Ca2(+)-indicator dyes quin2 or fura-2, in order to measure Ca2+ or Mn2+ entry through RMCE as well as Ca2+ release from internal stores. The IC50 (concn. producing 50% inhibition) for inhibition of RMCE by SK&F 96365 in platelets stimulated with ADP or thrombin was 8.5 microM or 11.7 microM respectively; these concentrations of SK&F 96365 did not affect internal Ca2+ release. Similar effects of SK&F 96365 were observed in suspensions of neutrophils and in single endothelial cells. SK&F 96365 also inhibited agonist-stimulated Mn2+ entry in platelets and neutrophils. The effects of SK&F 96365 were independent of cell type and of agonist, as would be expected for a compound that modulates post-receptor events. Voltage-gated Ca2+ entry in fura-2-loaded GH3 (pituitary) cells and rabbit ear-artery smooth-muscle cells held under voltage-clamp was also inhibited by SK&F 96365; however, the ATP-gated Ca2(+)-permeable channel of rabbit ear-artery smooth-muscle cells was unaffected by SK&F 96365. Thus SK&F 96365 (unlike the 'organic Ca2+ antagonists') shows no selectivity between voltage-gated Ca2+ entry and RMCE, although the lack of effect on ATP-gated channels indicates that it discriminates between different types of RMCE. The effects of SK&F 96365 on functional responses of cells thought to be dependent on Ca2+ entry via RMCE were also studied. Under conditions where platelet aggregation is dependent on stimulated Ca2+ entry via RMCE, the response was blocked by SK&F 96365 with an IC50 of 15.9 microM, which is similar to the IC50 of 8-12 microM observed for inhibition of RMCE. Adhesion and chemotaxis of neutrophils were also inhibited by SK&F 96365. SK&F 96365 is a useful tool to distinguish RMCE from internal Ca2+ release, and to probe the role of RMCE in mediating functional responses of cells. However, SK&F 96365 is not as potent (IC50 around 10 microM) or selective (also inhibits voltage-gated Ca2+ entry) as would be desirable, so caution must be exercised when using this compound.
authors.affiliation: SmithKline Beecham Pharmaceuticals, Frythe, Welwyn, Herts, U.K.
authors.forename: J E
authors.initials: JE
authors.lastname: Merritt
authors.forename: W P
authors.initials: WP
authors.lastname: Armstrong
authors.forename: C D
authors.initials: CD
authors.lastname: Benham
authors.forename: T J
authors.initials: TJ
authors.lastname: Hallam
authors.forename: R
authors.initials: R
authors.lastname: Jacob
authors.forename: A
authors.initials: A
authors.lastname: Jaxa-Chamiec
authors.forename: B K
authors.initials: BK
authors.lastname: Leigh
authors.forename: S A
authors.initials: SA
authors.lastname: McCarthy
authors.forename: K E
authors.initials: KE
authors.lastname: Moores
authors.forename: T J
authors.initials: TJ
authors.lastname: Rink
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000634:Aminoquinolines; D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D015220:Calcium Channels / Q000378:metabolism*; D002448:Cell Adhesion / Q000187:drug effects; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism; D005456:Fluorescent Dyes; D016257:Fura-2; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology*; D015640:Ion Channel Gating / Q000187:drug effects; D008345:Manganese / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D011817:Rabbits; D014471:Umbilical Veins
chemical_list: D000634:Aminoquinolines; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D005456:Fluorescent Dyes; D007093:Imidazoles; D010975:Platelet Aggregation Inhibitors; D008345:Manganese; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; C035739:Quin2; D002118:Calcium; D016257:Fura-2