蛋白酶激活受体-4拮抗剂的研究进展

蛋白酶激活受体-4(PAR-4)是一个由300个氨基酸经过7次跨膜形成的G蛋白偶联受体,它可通过与凝血酶的结合诱导血小板聚集,参与凝血过程。因此,PAR-4受体可以作为抗血小板药物的新靶点。近年来研究者发现了3种不同结构类型的小分子PAR-4受体拮抗剂,分别含有吲唑、吲哚和咪唑并噻二唑结构。对这几种拮抗剂的结构、药理活性等方面的研究进展进行综述。     

抗血小板药物研究进展及新靶点药物的发现

抗血小板药物是预防和治疗心肌梗死和缺血性脑卒中等动脉血栓疾病的主要药物,目前应用的该类药物主要包括阿司匹林、P2Y₁₂受体拮抗剂、糖蛋白IIb/IIIa（GPIIb/IIIa）受体拮抗剂和蛋白酶激活受体（PAR1）拮抗剂等。尽管已有抗血小板药物能够在一定程度上降低动脉血栓疾病的发生,但出血副作用不容忽视。随着对血小板相关机制研究的不断深入,越来越多抗血小板药物新靶点被发现,包括蛋白酶激活受体（PARs）、血小板糖蛋白VI（GPVI）、磷脂酰肌醇激酶（PI3Kβ）和蛋白二硫键异构酶（PDI）等。综述了已上市的抗血小板药物、目前临床应用存在的问题以及以新靶点研发的研究状况。     

FDA批准抗血小板药Zontivity用于心梗和卒中高危患者

2014年5月美国FDA批准Zontivity（vorapaxar）用于心肌梗死、卒中或外周动脉疾病患者,以减少心梗、卒中、心血管相关的死亡和冠脉血管重建的风险。PAR-1是一种可被凝血酶激活的受体,而凝血酶是一种有效的血小板激活剂。Zontivity是一种首创的蛋白酶激活受体1拮抗剂,可以抑制凝血酶诱导的血小板激活,抑制血小板聚集。     

凝血酶受体拮抗剂在抗血栓方面的研究进展

20世纪90年代初克隆和发现的凝血酶受体(PAR-1)为研制抗血栓药物提供了一个新靶点,引起学术界和制药业的广泛关注。由于凝血酶受体的特殊活化机制,其自身相连的活化氮端与活化中心近在咫尺,只有结合力很高的小分子化合物才能有效地拮抗凝血酶受体。因此,多年来,只有少数化合物被发现具有较好的凝血酶受体拮抗活性,其中vorapaxar和atopaxar进入了临床试验。vorapaxar在Ⅲ期临床试验发现有显著疗效,但也有出血不良反应,尤其不适用于有中风史的患者。最近,vorapaxar与PAR-1结合的晶体结构已经发表。这些结果为设计和研制新一代凝血酶受体拮抗剂指出了优化的方向,提供了分子水平的结构信息。本文从药物化学角度综述近年来凝血酶受体拮抗剂的研究进展和现状,重点描述vorapaxar、atopaxar以及相关化合物和最新发表的PAR-1拮抗剂。     

抗血栓单克隆抗体药物研究进展

血栓性疾病是严重危害人类健康的疾病之一 ,在血栓形成过程中血小板发挥着至关重要的作用 ,因此 ,抗血小板药物已经作为治疗血栓性疾病的有效手段在临床上得到了广泛应用。最常用的抗血小板药物是阿司匹林 ,它通过抑制血小板花生四烯酸的代谢 ,阻断血栓烷A2的产生 ,而具有抗血小板聚集功能的作用。抵克力得 (ticlid ,TP)是血小板二磷酸腺苷 (ADP)受体拮抗剂 ,它能阻断ADP诱导的血小板聚集、活化。然而 ,血小板还可以通过胶原受体、凝血酶受体以及非花生四烯酸代谢途径黏附、活化和聚集而形成血栓。寻找新的抗血栓药物已经是医药研究热点…     

应用于抗血栓治疗的新凝血酶受体抑制剂SCH-530348

喜巴辛衍生物SCH-530348是蛋白酶激活受体1(PAR-1)的抑制剂,PAR-1是人血小板凝血酶最主要的一种受体。SCH-530348是第一种能抑制凝血酶诱导的血小板聚集而不影响凝血酶对胶原酶原活化能力的化合物。文中综述了SCH-530348的临床前和Ⅰ～Ⅲ期临床试验结果。这些研究表明SCH-530348具有降低缺血性事件发生风险的几率,同时不会明显增加机体出血的风险,临床上可应用于急性冠脉综合征的治疗和缺血性心血管事件的预防。     

凝血酶蛋白酶激活受体-1作为转移性黑色素瘤治疗潜在靶点的研究进展

黑色素瘤由于转移性强,成为皮肤癌中死亡率最高的恶性肿瘤之一,目前没有有效的治疗方法。凝血酶蛋白酶激活受体-1(PAR-1)在黑色素瘤的发病过程中起到重要作用,PAR-1通过激活肿瘤细胞黏附和侵袭以及新生血管因子生成促进黑色素瘤转移。PAR-1有望成为治疗转移性黑色素瘤药物新靶点。     

缺血性心脏病的抗血小板治疗

冠状动脉粥样硬化斑块的破裂或损伤,常发生于急性冠脉综合征或经皮冠脉介入（PCI）术后患者,可诱发血小板的激活和血栓的形成,从而导致心肌梗死或心源性猝死。血小板表面糖蛋白Ⅱb／Ⅲa受体的激活是激活血小板及最终形成血栓的最后共同途径。由于血小板在斑块破裂后血栓形成的发病机制中起着关键作用,故应用抗血小板药物,如阿司匹林、噻氯吡啶类药物或糖蛋白Ⅱb／Ⅲa受体拮抗剂,可以有效地减少急性冠脉综合征或PCI术后患者的心血管事件的发生率。不同的抗血小板药物,由于起效时间、作用方式、效能以及花费不同,故在不同疾病状态下,需要选用或联用不同的药物以达到安全、充分有效的抗血小板作用。     

新的抗血栓药物的研究进展

血小板活化在血栓性疾病中起关键作用，阿司匹林一直是预防血检性疾病的主要药物。血栓素合成酶抑制剂和血栓素受体拮抗剂等是新的有效的抗血小板药物。     

血栓烷A2受体信号传导及相关抗血小板药物研究进展

摘 要血栓烷A2受体广泛存在于人体不同的组织器官,属于G蛋白偶联受体超家族中的一种七重跨膜蛋白,在血小板膜上也有分布。血栓烷A2受体激活导致一系列连锁反应也是活化血小板的重要途径之一,进而促进止血和血栓形成。目前,抗血小板药物的研究主要集中在血小板活化通路中的受体和信号分子水平,血栓烷A2受体相关的抗血小板药物也受到广泛关注。了解血小板内该受体下的信号转导,对于探讨相关药物作用机制、开发新型或中药有效成分类抗血小板药物具有重要意义。     

Thrombin receptor antagonists; recent advances in PAR-1 antagonist development

The receptor for the serine protease thrombin, the protease-activated receptor-1 (PAR-1), has been recently characterized. Its key roles in thrombin-stimulated human platelet activation, vascular endothelial and smooth muscle proliferation, inflammatory responses and neurodegeneration suggest receptor involvement in various disorders such as arterial thrombosis, atherosclerosis, restenosis, inflammation and myocardial infarction. It has been established that thrombin elicits the majority of its effects via PAR-1. PAR-1 has a novel mechanism of activation. The receptor, a member of the seven-transmembrane domain receptor family, is cleaved by thrombin at a specific site on the N-terminal extension, and a newly exposed N-terminus acts as a tethered ligand to activate the receptor itself. The need for development of a PAR-1 antagonist that may be valuable as a therapeutic agent has been recognized. An intriguing challenge is the necessity of the antagonist to compete with an intramolecular ligand while showing no intrinsic activity. The lead compounds were found to be synthetic peptides containing N-terminal hexapeptide or pentapeptide (Ser-Phe-Leu-Leu-Arg-Asn, Ser-Phe-Leu-Leu-Arg) or modified sequences (TRAPs; thrombin receptor-activating peptides), which exhibit full PAR-1 agonist activity. Selective PAR-1 antagonists have already been synthesized. Though their potency is still not enough to justify therapeutic use, it is clear that future progress will bring a novel class of drugs-thrombin receptor antagonists. The emphasis of this review, therefore, will be placed on advances in the discovery of potent and selective PAR-1 antagonists.     

G-protein coupled receptor antagonists-1: protease activated receptor-1 (PAR-1) antagonists as novel cardiovascular therapeutic agents

Inhibition of thrombin receptor (PAR-1) is a promising therapeutic approach for the treatment of various cardiovascular disorders such as unstable angina, acute myocardial infarction, stroke, and restenosis. Since a PAR-1 antagonist is specific for the cellulalr actions of thrombin, and does not interfere with fibrin generation, it is expected to have less bleeding liability than the currently available treatments. Several peptide and non-peptide PAR-1 antagonists with potent inhibition of platelet aggregation have been reported. Antithrombotic effect of a PAR-1 antibody has been demonstrated in a baboon thrombosis model and the antirestenosis property of a PAR-1 antagonist has been demonstrated in a rat model.     

Discovery of an orally available PAR-1 antagonist as a novel antiplatelet agent

Antiplatelet therapy is a key treatment in atherothrombotic disease and platelet is activated via multiple pathways. Current agents do not interfere with all pathways including the protease-activated receptor-1 (PAR-1) pathway stimulated by thrombin. New antiplatelet agents targeting PAR-1 are aimed to reduce thrombosis ideally without increasing bleeding risk. This article provides a review of the new class of agents, PAR-1 antagonists.     

Development of agonists/antagonists for protease-activated receptors (PARs) and the possible therapeutic application to gastrointestinal diseases

Protease-activated receptors (PARs), a family of G-protein-coupled seven-transmembrane-domain receptors, are activated by proteolytic unmasking of the N-terminal cryptic tethered ligand by certain serine proteases. Among four PAR family members cloned to date, PAR-1, PAR-2, and PAR-4 can also be activated through a non-enzymatic mechanism, which is achieved by direct binding of exogenously applied synthetic peptides based on the tethered ligand sequence, known as PARs-activating peptides, to the body of the receptor. Various peptide mimetics have been synthesized as agonists for PARs with improved potency, selectivity, and stability. Some peptide mimetics and/or nonpeptide compounds have also been developed as antagonists for PAR-1 and PAR-4. PARs are widely distributed in the mammalian body, especially throughout the alimentary systems, and play various roles in physiological/pathophysiological conditions, i.e., modulation of salivary, gastric, or pancreatic glandular exocrine secretion, gastrointestinal smooth muscle motility, gastric mucosal cytoprotection, suppression/facilitation of visceral pain and inflammation, etc. Thus PARs are now considered novel therapeutic targets, and development of selective agonists and/or antagonists for PARs might provide a novel strategy for the treatment of various diseases that are resistant to current therapeutics.     

Discovery of potent orally active thrombin receptor (protease activated receptor 1) antagonists as novel antithrombotic agents

Structurally novel thrombin receptor (protease activated receptor 1, PAR-1) antagonists based on the natural product himbacine are described. The prototypical PAR-1 antagonist 55 showed a Ki of 2.7 nM in the binding assay, making it the most potent PAR-1 antagonist reported. 55 was highly active in several functional assays, showed excellent oral bioavailability in rat and monkey models, and showed complete inhibition of agonist-induced ex vivo platelet aggregation in cynomolgus monkeys after oral administration.     

Review article: proteinase-activated receptors - novel signals for gastrointestinal pathophysiology

Proteinase-activated receptors (PARs) have the common property of being activated by the proteolytic cleavage of their extracellular N-terminal domain. The new NH2-terminus acts as a 'tethered ligand' binding and activating the receptor itself. Four members of this family have been cloned, three of which are activated by thrombin (PAR-1, PAR-3 and PAR-4) while the fourth (PAR-2) is activated by trypsin or mast cell tryptase. In physiological or pathophysiological conditions, the gastrointestinal tract is exposed more than other tissues to proteinases (digestive enzymes, proteinases from pathogens or proteinases from inflammatory cells) that can activate PARs. Since PARs are highly expressed throughout the gastrointestinal tract, the study of the role of PARs in these tissues appears to be particularly important. It has already been shown that PAR-2 activation induces calcium mobilization and eicosanoid production in enterocytes as well as changes in ion transport in jejunal tissue segments. PAR-2 activation also causes calcium mobilization and stimulates amylase release from pancreatic acini. Moreover, both PAR-1 and PAR-2 activation can alter the gastrointestinal motility. In inflammatory or allergic conditions, the proteinases that constitute the major agonists for PARs (thrombin, trypsin and mast cell tryptase) are usually released. The activation of PARs by these proteinases might contribute to the gastrointestinal disorders associated with these pathologies. A complete understanding of the role of PARs in the gastrointestinal tract will require the development of selective receptor antagonists that are not yet available. Nonetheless, the use of PAR agonists has already highlighted new potential functions for proteinases in the gastrointestinal tract, thus the control of PAR activation might represent a promising therapeutic target.     

Proteinase-activated receptor-2 mediates hyperresponsiveness in isolated guinea pig bronchi

The mast cell serine protease tryptase has been implicated as a critical mediator of airway hyperresponsiveness in vitro and in vivo. We have previously demonstrated that tryptase promotes hyperresponsiveness in isolated guinea pig bronchi. In this study, we have investigated the potential role of tryptase-mediated activation of proteinase-activated receptor-2 (PAR-2) in promoting airway hyperresponsiveness. Ex vivo exposure of guinea pig bronchi to the PAR-2 agonists H(2)N-Ser-Leu-Ile-Gly-Arg-Leu-CONH(2) (SLIGRL) and t-cinnamoyl-H(2)N-Leu-Ile-Gly-Arg-Leu-O-CONH(2) (t-c-LIGRLO) (0.1-10 microM) induced a concentration-dependent increase of contractile response to histamine. Treatment with 10 microM SLIGRL or t-c LIGRLO for 45 min increased subsequent responsiveness to histamine (0.3mM) by 54+/-3% and 69+/-5%, respectively (P<0.05 vs. control). In contrast, the PAR-1 agonist peptide H(2)N-Ser-Phe-Leu-Leu-Arg-Asn-CONH(2) (SFLLRN) did not promote significant changes in the airway. Effects of the peptides were observed following at least a 30-min preincubation with the tissue. Coincubation with indomethacin or removal of epithelial cells is required for PAR-2-mediated hyperreactivity. The inactive analogue H(2)N-Leu-Ser-Ile-Gly-Arg-Leu-CONH(2) (LISGRL; 10 microM) failed to promote hyperresponsiveness. Neuropeptide antagonists blocked the effect of the PAR-2 agonists. Selective antagonists of NK1 (L-703,606), NK2 (L-659,877), and CGRP (alphaCGRP 8-37) provided additive inhibition of PAR-2-mediated hyperreactivity. Pretreatment of bronchi with capsaicin (0.8 microM) also prevented the effects of SLIGRL. These results demonstrate the potential involvement of tryptase-mediated activation of PAR-2 in promoting airway hyperresponsiveness. These results further demonstrate that the PAR-2-mediated response involves a neurogenic mechanism involving neuropeptide release.     

Proteinase-activated receptors

Proteinase-activated receptors are a recently described, novel family of seven-transmembrane G-protein-coupled receptors. Rather then being stimulated through ligand receptor occupancy, activation is initiated by cleavage of the N terminus of the receptor by a serine protease resulting in the generation of a new tethered ligand that interacts with the receptor within extracellular loop-2. To date, four proteinase-activated receptors (PARs) have been identified, with distinct N-terminal cleavage sites and tethered ligand pharmacology. In addition to the progress in the generation of PAR-1 antagonists, we describe the role of thrombin in such processes as wound healing and the evidence implicating PAR-1 in vascular disorders and cancer. We also identify advances in the understanding of PAR-1-mediated intracellular signaling and receptor desensitization. The cellular functions, signaling events, and desensitization processes involved in PAR-2 activation are also assessed. However, other major aspects of PAR-2 are highlighted, in particular the ability of several serine protease enzymes, in addition to trypsin, to function as activators of PAR-2. The likely physiological and pathophysiological roles for PAR-2 in skin, intestine, blood vessels, and the peripheral nervous system are considered in the context of PAR-2 activation by multiple serine proteases. The recent discovery of PAR-3 and PAR-4 as additional thrombin-sensitive PARs further highlights the complexity in assessing the effects of thrombin in several different systems, an issue that remains to be fully addressed. These discoveries have also highlighted possible PAR-PAR interactions at both functional and molecular levels. The future identification of other PARs and their modes of activation are an important future direction for this expanding field of study.     

Therapeutic potential of protease-activated receptor-1 antagonists

The serine protease thrombin (EC 3.4.21.5) is central to the maintenance of haemostatic balance through its coagulant, anticoagulant and platelet activating properties. In addition, this enzyme affects numerous cellular responses in a wide variety of cells, such as cell proliferation, cytokine and growth factor release, lipid metabolism and tissue remodelling. A family of G-protein-coupled protease-activated receptors (PARs) mediates these cellular actions of thrombin. While thrombin can activate three of the four PAR family members, PAR-1 represents the primary thrombin-responsive receptor in human cells. The expression of PAR-1 in platelets, the vasculature and myocardium, in cells within atherosclerotic plaque and tissues after vascular injury, indicates that this receptor plays an important role during the response to tissue injury and associated inflammatory processes. With the development of PAR-deficient mice and small-molecule antagonists, it is now clear that intervening in processes mediated by PAR-1 presents a new approach to treating a variety of disorders dependent on thrombin generation, including thrombosis and restenosis. The full potential of PAR-1 antagonists has yet to be realised, but the promise of novel therapeutics that modulate receptor function rather than thrombin's proteolytic activity, provides an alternative and, perhaps, more desirable means to dampen the pathological effects of thrombin.