title: Inhibition of platelet functions and thrombosis through selective or nonselective inhibition of the platelet P2 receptors with increasing doses of NF449 [4,4',4'',4'''-(carbonylbis(imino-5,1,3-benzenetriylbis-(carbonylimino)))tetrakis-benzene-1,3-disulfonic acid octasodium salt].
abstract: Our aim was to determine whether the newly described P2X1 antagonist NF449 [4,4',4'',4'''-(carbonylbis(imino-5,1,3-benzenetriylbis(carbonylimino)))tetrakis-benzene-1,3-disulfonic acid octasodium salt] could selectively antagonize the platelet P2X1 receptor and how it affected platelet function. NF449 inhibited alpha,beta-methyleneadenosine 5'-triphosphate-induced shape change (IC50 = 83 +/- 13 nM; n = 3) and calcium influx (pA2 = 7.2 +/- 0.1; n = 3) (pIC50 = 6.95) in washed human platelets treated with apyrase to prevent desensitization of the P2X1 receptor. NF449 also antagonized the calcium rise mediated by the P2Y1 receptor, but with lower potency (IC50 = 5.8 +/- 2.2 microM; n = 3). In contrast, it was a very weak antagonist of the P2Y12-mediated inhibition of adenylyl cyclase activity. Selective blockade of the P2X1 receptor with NF449 led to reduced collagen-induced aggregation, confirming a role of this receptor in platelet activation induced by collagen. Intravenous injection of 10 mg/kg NF449 into mice resulted in selective inhibition of the P2X1 receptor and decreased intravascular platelet aggregation in a model of systemic thromboembolism (35 +/- 4 versus 51 +/- 3%) (P = 0.0061; n = 10) but without prolongation of the bleeding time (106 +/- 16 versus 78 +/- 7 s; n = 10) (N.S.; P = 0.1209). At a higher dose (50 mg/kg), NF449 inhibited the three platelet P2 receptors. This led to a further reduction in platelet consumption compared with mice injected with saline (13 +/- 4 versus 42 +/- 3%) (P = 0.0002; n = 5). NF449 also reduced dose-dependently the size of thrombi formed after laser-induced injury of mesenteric arterioles. Overall, our results indicate that NF449 constitutes a new tool to investigate the functions of the P2X1 receptor and could be a starting compound in the search for new antithrombotic drugs targeting the platelet P2 receptors.
authors.affiliation: INSERM U. 311, Etablissement Français du Sang-Alsace, 10, rue Spielmann, Strasbourg Cédex, France.
authors.forename: Béatrice
authors.initials: B
authors.lastname: Hechler
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Magnenat
authors.forename: Maddalena L
authors.initials: ML
authors.lastname: Zighetti
authors.forename: Matthias U
authors.initials: MU
authors.lastname: Kassack
authors.forename: Heiko
authors.initials: H
authors.lastname: Ullmann
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Cazenave
authors.forename: Richard
authors.initials: R
authors.lastname: Evans
authors.forename: Marco
authors.initials: M
authors.lastname: Cattaneo
authors.forename: Christian
authors.initials: C
authors.lastname: Gachet
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001557:Benzenesulfonates / Q000008:administration & dosage / Q000494:pharmacology*; D001760:Bleeding Time; D001792:Blood Platelets / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism; D003094:Collagen / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004837:Epinephrine / Q000494:pharmacology; D005343:Fibrinolytic Agents; D006801:Humans; D066298:In Vitro Techniques; D007275:Injections, Intravenous; D051379:Mice; D008855:Microscopy, Electron, Scanning; D009865:Oocytes / Q000378:metabolism; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D010979:Platelet Function Tests; D018048:Receptors, Purinergic P2 / Q000187:drug effects*; D058469:Receptors, Purinergic P2X; D013923:Thromboembolism / Q000517:prevention & control; D013927:Thrombosis / Q000188:drug therapy; D014982:Xenopus laevis
chemical_list: C110258:4,4,',4'',4'''-(carbonylbis(imino-5,1,3-benzenetriylbis(carbonylimino)))tetrakis(benzene-1,3-disulfonate); D001557:Benzenesulfonates; D005343:Fibrinolytic Agents; D010975:Platelet Aggregation Inhibitors; D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D003094:Collagen; D000262:Adenylyl Cyclases; C002630:alpha,beta-methyleneadenosine 5'-triphosphate; D002118:Calcium; D004837:Epinephrine