Application of difluoromethylene phosphonic acids to development of biologically active nucleotides

1)根据9-（5′，5′-二氟-5′-磷戊基）鸟嘌呤（DFPP-G）与小牛脾脏PNP二元配合物的x射线晶体学数据，设计9-（5′，5′-二氟-5′-磷戊基）-9-去氮鸟嘌呤核苷磷酸化酶（PNP）多底物类似抑制剂。ddfpp - dg及其类似化合物通过9-二氮杂-9-碘鸟嘌呤衍生物与ω-炔基二氟亚甲基膦酸盐作为关键反应的Sonogashira偶联反应获得的连接物长度进行调节。DFPP-DG是一种非常有效的PNP抑制剂，对小牛脾脏和人红细胞PNP的表观抑制常数（在1 mM磷酸盐存在下）分别为4.4 nM和8.1 nM。其中一种类似物，具有较长链连接膦酸盐和9-去氮鸟嘌呤的homo- dppg - dg，对人酶的抑制作用更强，明显抑制常数为53 nM（在1mM磷酸盐存在下）。2)天然核苷的二磷酸衍生物，包括腺苷3',5'-二磷酸（A3P5P），据报道可作为P2Y1受体的竞争性拮抗剂或部分激动剂。选择性P2Y1受体拮抗剂被认为具有抗血栓作用的潜力，而选择性受体激动剂可能具有抗高血压或抗糖尿病作用的潜力。因此，合成了多种修饰的核苷酸类似物，以讨论其与P2Y1受体拮抗剂和激动剂的构效关系。其中，MRS2179据报道对P2Y1受体具有选择性拮抗剂活性。MRS2179对体外血小板聚集的影响较强，但持续时间较短。为了开发一种耐磷酸酶的P2Y1受体拮抗剂，我们研究了P2Y1受体配体的修饰核苷酸类似物的合成途径，其中MRS2216的一个或两个磷酸基被二氟甲基膦基取代。

1) 9-(5',5'-Difluoro-5'-phosphonopentyl)-9-deazaguanine (DFPP-DG) was designed as a multi-substrate analogue inhibitor against purine nucleoside phosphorylase (PNP) on the basis of X-ray crystallographic data obtained for a binary complex of 9-(5',5'-difluoro-5'-phosphonopentyl)guanine (DFPP-G) with calf spleen PNP. DFPP-DG and its analogous compounds were adjusted by length of the linker achieved by the Sonogashira coupling reaction between a 9-deaza-9-iodoguanine derivative and ω-alkynyldifluoromethylene phosphonates as a key reaction. DFPP-DG is a very potent PNP inhibitor with apparent inhibition constants (in the presence of 1 mM phosphate) of 4.4 nM and 8.1 nM vs calf spleen and human erythrocyte PNPs, respectively. One of its analogues, homo-DFPP-DG, with longer chain linking phosphonate and 9-deazaguanine is even more potent vs human enzyme, with an apparent inhibition constant of 53 nM (in the presence of 1mM phosphate).2) The bisphosphate derivatives of naturally occurring nucleosides including adenosine 3',5'-bisphosphate (A3P5P) are reported to act as competitive antagonists or partial agonists of P2Y1 receptors. A selective P2Y1 receptor antagonist is believed to have potential as an antithrombotic agent, while a selective receptor agonist may have potential as an antihypertensive or antidiabetic agent. Therefore, a variety of modified nucleotide analogues were synthesized to discuss the structure-activity relationships to P2Y1 receptor antagonists and agonists. Among them, MRS2179 is reported to show selective antagonist activities against P2Y1 receptor. However, the effect of MRS2179 on ex vivo platelet aggregation was strong but of short duration. To develop a phosphatase-resistant P2Y1 receptor antagonist, we examined to synthetic routes to modified nucleotide analogues for P2Y1 receptor ligands, in which one or two of phosphate groups for MRS2216 are replaced by a difluoromethylenephosphonyl group.