Received for publication November 14, 2005.
Revised January 5, 2006.
Accepted for publication January 5, 2006.

Pharmacological characterization of 4-methoxy-N¹-(4-trans-nitrooxycyclohexyl)-N³-(3-pyridinylmethyl)-1,3-benzenedicarboxamide (2NTX-99), a potential antiatherothrombotic agent with anti-thromboxane and NO-donor activity in platelet and vascular preparations

Abstract

Thromboxane (TXA₂), prostacyclin (PGI₂) and nitric oxide (NO), regulate platelet function and interaction with the vessel wall. Inhibition of TXA₂, implemented synthesis of PGI₂, and supply of exogenous NO, may afford therapeutic benefit. 2NTX-99, a new chemical entity related to picotamide, showed antithromboxane activity and NO-donor properties. 2NTX-99 relaxed rabbit aortic rings precontracted with norepinephrine or U46619 (EC₅₀ 7.9, 17.1 µM, respectively), an effect abolished by 10 µM ODQ. 2NTX-99 inhibited arachidonic acid (AA) induced washed platelet aggregation (EC₅₀ 9.8 µM) and TXB₂ formation (-71% at 10 µM) and its potency increased in the presence of aortic rings (EC₅₀ 1.4 µM). In whole rabbit aorta incubated with homologous platelets, AA caused contraction and TXA₂ formation, reduced by 2NTX-99 (10-40 µM): contraction -28, and -47 %, TXA₂ formation -37 and -75.4 % respectively, with concomitant increase in PGI₂. 2NTX-99 (20-40 µM) inhibited U46619-induced aggregation in rabbit PRP (-74±6.7, -96±2.4% respectively), and inhibited collagen-induced aggregation in human PRP (-48.2±10 and -79.2±6%), while ozagrel was ineffective. In HEK293 cells, transfected with the TP receptor, 2NTX-99 did not compete with the ligand (³H-SQ29,548) nor prevented IP accumulation. After oral administration (50-250 mg/Kg), 2NTX-99 inhibited TXA₂ production in rat clotting blood (-71 and -91 %); at 250 mg/Kg an AUC _{0-16 hours} of 149.5 h·µg/ml and a t_1/2 of 6 h were calculated, with a C_max value of 31.8±8.2 µg/ml. An excellent correlation between plasma concentrations and TXA₂ inhibition occurs. 2NTX-99 controls platelet function and vessel wall interaction by multifactorial mechanisms and possesses therapeutic potential.

Key words: antithrombotic drugs, nitric oxide, platelet aggregation, prostacyclin, thromboxane A2, vasodilation