ono-11120 and picotamide

1. Picotamide was shown to inhibit platelet binding of thromboxane A2 (TxA2)-mimetics and to cause a reduction of TxA2 platelet receptors after in vivo administration. The present study aimed to investigate directly [3H]-picotamide binding to human platelets and in particular the relationship between binding kinetics and antiaggregating properties. 2. [3H]-picotamide time-dependently bound to a single class of platelet TxA2 receptors with a KD of 325 nmol l-1 at equilibrium. The binding was displaceable by TxA2 analogues U46619 and ONO11120 (Ki 19 and 28 nmol l-1 respectively) but not by prostacyclin (PGI2), prostaglandin E2 (PGE2) and TxB2. Antiaggregating activity and TxA2 formation inhibition paralleled with binding kinetics. 3. By prolonging the incubation time from 30 to 120 min, picotamide showed a progressively increasing non-displaceable binding, whereas specific displaceable binding decreased in comparison to the values reached at 30 min. Non displaceable binding was specific, temperature-dependent saturable and followed a Michaelis-Menten kinetic (Vmaxapp = 130 fmol per 10(8) platelets h-1, KMapp = 330 nmol l-1). Picotamide progressively underwent a specific stable interaction with its platelet receptor. 4. In conclusion, after an initial reversible binding, a progressive stabilization of picotamide binding takes place resulting in a progressively more stable interaction with platelets.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adult; Binding, Competitive; Blood Platelets; Collagen; Humans; In Vitro Techniques; Kinetics; Phthalic Acids; Platelet Aggregation; Platelet Aggregation Inhibitors; Prostaglandin Endoperoxides, Synthetic; Receptors, Thromboxane; Thromboxane A2

On the basis of indirect pharmacological evidence, picotamide, a methoxy derivative of 4-hydroxy-isophthalic acid (N,N'bis(3-picolyl)-4-methoxy-isophthalamide) has been postulated to inhibit platelet aggregation by competitively interfering with the thromboxane A2 (TxA2) platelet receptor. In the present study the interaction between picotamide and TxA2 receptors on human platelets was investigated by a direct radioligand assay method with [125I]PTA-OH and [3H]U46619 as labelled radioligands. The ONO11120 and U46619 inhibitory constants (Ki) for [125I]PTA-OH binding were 19 +/- 4 and 17 +/- 3 nM, respectively. Picotamide displaced [125I]PTA-OH binding with a Ki of 1472 +/- 321 nM. The Ki for ONO 11120 and U46619 on [3H]U46619 binding were 42 +/- 12 and 16 +/- 5 nM, respectively, whereas the Ki for picotamide was 1648 +/- 431 nM. These data provide evidence that picotamide can directly inhibit the TxA2 platelet receptor.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Adult; Binding, Competitive; Blood Platelets; Female; Humans; In Vitro Techniques; Kinetics; Male; Middle Aged; Phthalic Acids; Platelet Aggregation Inhibitors; Prostaglandin Endoperoxides, Synthetic; Radioligand Assay; Receptors, Prostaglandin; Receptors, Thromboxane; Thermodynamics; Thromboxane A2