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YM-254890, which was isolated from the culture broth of Chromobacterium sp., inhibits ADP-induced platelet aggregation and has antithrombotic and thrombolytic effects. YM-254890 blocks Gαq/11-coupled ADP receptor P2Y1-mediated Ca2+ mobilization. Here we report that YM-254890 is a selective Gαq/11 inhibitor. YM-254890 blocked Ca2+ mobilization mediated by several Gαq/11coupled receptors but not by Gαi- or Gα15-coupled receptor, indicating that phospholipase Cfi activation and subsequent signaling molecules are not the target of YM-254890. YM-254890 completely prevented the serum response factor (SRF)-mediated gene transcription induced by GαqR183C, which is constitutively active in a receptor-dependent manner because of its reduced kcat of GTP hydrolysis. Conversely, YM-254890 had only a modest effect on the SRF-mediated gene transcription by GαqQ209L, which is GTPase-deficient (activated) Gαg. These suggested that the acting point of YM-254890 is receptor-Gαg interaction or the subsequent guanine nucleotide exchange step. The fact that YM-254890 (i) inhibited the SRF-mediated gene transcription by Gαqi5, which interacts with Gαi-coupled receptor and possesses the effector function of Gαq, and (ii) had no effect on the Kd value of high affinity [³H]2MeSADP binding to P2Y1, which reflects the agonist-receptor-Gα ternary complex, suggested that receptor-Gαq/11 interaction is not the target of YM-254890. On the other hand, specific [35S]GTPγS binding to Gαq/11 stimulated by the M1 muscarinic acetylcholine receptor and P2Y1 were inhibited by YM-254890. These data indicate that YM-254890 blocks the exchange of GDP for GTP in Gαq/11 activation. This novel Gαq/11selective inhibitor is a promising and powerful tool for studying Gαq/11 protein activation, Gαq/11-coupled receptor signaling, and Gαq/11-mediated biological events. [ABSTRACT FROM AUTHOR]

The structure elucidation and biological activity of novel YM-254890 (1) analogues and semi-synthetic derivatives are described. Three natural analogues, YM-254891 (2), YM-254892 (3), and YM-280193 (4), were isolated from the fermentation broth of Chromobacterium sp. QS3666, and two hydrogenated derivatives, YM-385780 (5) and YM-385781 (6), were synthesized from YM-254890. Their structures were determined by one- and two-dimensional NMR studies and mass spectrometry. Among these compounds, two natural analogues 2–3 which possessed acyl groups at β-HyLeu-1 and one derivative 6 whose conformation was similar to that of 1 showed comparable Gαq/11 inhibitory activity to that of 1. This indicates that the acyl β-HyLeu residue plays an important role in activity and also that the α,β-unsaturated carbonyl group of the N-MeDha residue is not critical to activity. The other hydrogenated derivative 5 had significantly less activity, which could be attributed to conformational differences. [Copyright &y& Elsevier]

1 The effects of YM-254890, a specific Galpha(q/11) inhibitor, on platelet functions, thrombus formation under high-shear rate condition and femoral artery thrombosis in cynomolgus monkeys were investigated. 2 YM-254890 concentration dependently inhibited ADP-induced intracellular Ca(2+) elevation, with an IC(50) value of 0.92+/-0.28 microM. 3 P-selectin expression induced by ADP or thrombin receptor agonist peptide (TRAP) was strongly inhibited by YM-254890, with IC(50) values of 0.51+/-0.02 and 0.16+/-0.08 microM, respectively. 4 YM-254890 had no effect on the binding of fibrinogen to purified GPIIb/IIIa, but strongly inhibited binding to TRAP-stimulated washed platelets. 5 YM-254890 completely inhibited platelet shape change induced by ADP, but not that induced by collagen, TRAP, arachidonic acid, U46619 or A23187. 6 YM-254890 attenuated ADP-, collagen-, TRAP-, arachidonic acid- and U46619-induced platelet aggregation with IC(50) values of <1 microM, whereas it had no effect on phorbol 12-myristate 13-acetate-, ristocetin-, thapsigargin- or A23187-induced platelet aggregation. 7 High-shear stress-induced platelet aggregation and platelet-rich thrombus formation on a collagen surface under high-shear flow conditions were concentration dependently inhibited by YM-254890. 8 The antithrombotic effect of YM-254890 was evaluated in a model of cyclic flow reductions in the femoral artery of cynomolgus monkeys. The intravenous bolus injection of YM-254890 dose dependently inhibited recurrent thrombosis without affecting systemic blood pressure or prolonging template bleeding time. 9 YM-254890 is a useful tool for investigating Galpha(q/11)-coupled receptor signaling and the physiological roles of Galpha(q/11).

We examined the antithrombotic and thrombolytic effects of the G(q/11) inhibitor YM-254890 in an electrically-induced carotid artery thrombosis model in rats. YM-254890 dose-dependently inhibited ex vivo ADP-induced platelet aggregation after i.v. bolus injection. In the thrombosis study, YM-254890 dosedependently prolonged time to occlusion at doses of 3 and 10 g/kg i.v. and decreased occlusion rate at 10 g/kg i.v. In the thrombolysis study, YM-254890 at 30 micro g/kg i.v. shortened the time to reperfusion and prevented reocclusion after thrombolysis with a modified tissue-type plasminogen activator. YM-254890, at 10 micro g/kg and more, significantly improved carotid patency status after thrombolysis. However, at 30 micro g/kg and more, YM-254890 decreased systemic blood pressure. These results suggest that YM-254890 may be effective for treating G(q)-mediated diseases, and that YM-254890 is a useful tool for investigating the biological roles of G(q/11).