HA-29: An inhibitor of thromboxane A₂ formation with antagonism of thromboxane A₂/prostaglandin endoperoxide receptor in rabbit platelets

HA-29, 2-[(3-methoxyphenyl)methyl]-pyrano[2,3-c]pyrazol-6(1H)-one, was investigated for its inhibitory mechanism of action in washed rabbit platelets. This compound inhibited the aggregation and ATP release of rabbit platelets induced by arachidonic acid and collagen in a concentration-dependent manner, without affecting those induced by ADP, PAF and thrombin. Prolongation of the incubation time of HA-29 with platelets did not cause further inhibition and the aggregability of the agent-treated platelets could be restored after washing of platelets. The concentration-response curve of u-46619-induced platelet aggregation was shifted to the right by HA-29 in a concentration-dependent manner, but the maximal aggregation was suppressed by HA-29. The pA₂ and PA₁₀ values of HA-29 on U-46619-induced platelet aggregation were 4.26 and 3.58, respectively, with a slope value of -1.4. The U-46619-induced aggregation was markedly disaggregated by HA-29 even it was added 5 min after U-46619. HA-29 inhibited the secondary aggregation and ATP release, but not the primary aggregation of human platelet-rich plasma induced by ADP and epinephrine. Thromboxane B₂ formation caused by arachidonic acid, collagen and thrombin was markedly suppressed by HA-29. HA-29 also inhibited the formation of prostaglandin D₂ caused by arachidonic acid. HA-29 inhibited almost completely the formation of inositol monophosphate caused by U-46619, but not that by collagen or thrombin. HA-29 did not affect U-46619-induced contraction of rat aorta. It is concluded that the antiplatelet effect of HA-29 is due to the inhibition of thromboxane A₂ formation and blockade of thromboxane A₂/prostaglandin endoperoxide receptor.