Cattaneo Marco, Lecchi Anna, Ohno Michihiro, Joshi Bhalchandra V, Besada Pedro, Tchilibon Susanna, Lombardi Rossana, Bischofberger Norbert, Harden T Kendall, Jacobson Kenneth A
Hematology and Thrombosis Unit, Ospedale San Paolo, DMCO-University of Milan, Milan, Italy.
Biochem Pharmacol. 2004 Nov 15;68(10):1995-2002. doi: 10.1016/j.bcp.2004.06.026.
Activation of the P2Y(1) nucleotide receptor in platelets by ADP causes changes in shape and aggregation, mediated by activation of phospholipase C (PLC). Recently, MRS2500(2-iodo-N(6)-methyl-(N)-methanocarba-2'-deoxyadenosine-3',5'-bisphosphate) was introduced as a highly potent and selective antagonist for this receptor. We have studied the actions of MRS2500 in human platelets and compared these effects with the effects of two acyclic nucleotide analogues, a bisphosphate MRS2298 and a bisphosphonate derivative MRS2496, which act as P2Y(1) receptor antagonists, although less potently than MRS2500. Improved synthetic methods for MRS2500 and MRS2496 were devised. The bisphosphonate is predicted to be more stable in general in biological systems than phosphate antagonists due to the non-hydrolyzable CP bond. MRS2500 inhibited the ADP-induced aggregation of human platelets with an IC(50) value of 0.95 nM. MRS2298 and MRS2496 also both inhibited the ADP-induced aggregation of human platelets with IC(50) values of 62.8 nM and 1.5 microM, respectively. A similar order of potency was observed for the three antagonists in binding to the recombinant human P2Y(1) receptor and in inhibition of ADP-induced shape change and ADP-induced rise in intracellular Ca(2+). No substantial antagonism of the pathway linked to the inhibition of cyclic AMP was observed for the nucleotide derivatives, indicating no interaction of these three P2Y(1) receptor antagonists with the proaggregatory P2Y(12) receptor, which is also activated by ADP. Thus, all three of the bisphosphate derivatives are highly selective antagonists of the platelet P2Y(1) receptor, and MRS2500 is the most potent such antagonist yet reported.
血小板中的P2Y(1)核苷酸受体被二磷酸腺苷(ADP)激活后,会通过磷脂酶C(PLC)的激活引发形状变化和聚集。最近,MRS2500(2-碘-N(6)-甲基-(N)-甲碳环-2'-脱氧腺苷-3',5'-双磷酸)作为该受体的一种高效且选择性的拮抗剂被引入。我们研究了MRS2500在人血小板中的作用,并将这些效应与两种无环核苷酸类似物的效应进行了比较,这两种类似物分别是双磷酸酯MRS2298和双膦酸盐衍生物MRS2496,它们作为P2Y(1)受体拮抗剂,但其效力不如MRS2500。我们设计了改进的MRS2500和MRS2496合成方法。由于不可水解的C-P键,预计双膦酸盐在生物系统中总体上比磷酸盐拮抗剂更稳定。MRS2500抑制人血小板的ADP诱导聚集,IC(50)值为0.95 nM。MRS2298和MRS2496也都抑制人血小板的ADP诱导聚集,IC(50)值分别为62.8 nM和1.5 μM。在与重组人P2Y(1)受体结合以及抑制ADP诱导的形状变化和ADP诱导的细胞内Ca(2+)升高方面,观察到这三种拮抗剂具有相似的效力顺序。对于核苷酸衍生物,未观察到与环磷酸腺苷抑制相关途径的实质性拮抗作用,这表明这三种P2Y(1)受体拮抗剂与同样被ADP激活的促聚集P2Y(12)受体没有相互作用。因此,所有这三种双磷酸酯衍生物都是血小板P2Y(1)受体的高度选择性拮抗剂,而MRS2500是迄今报道的此类最有效的拮抗剂。