Sechenov Institute of Evolutionary Physiology and Biochemistry of the Russian Academy of Sciences, Saint Petersburg, Russia (V.S., N.R., S.G.); Saint Petersburg State University, Saint Petersburg, Russia (N.A.A., A.P., A.K.); and Department of Internal Medicine, Division of Cardiology, University of Texas Houston Medical School, Houston, Texas (I.S., E.M.).
Sechenov Institute of Evolutionary Physiology and Biochemistry of the Russian Academy of Sciences, Saint Petersburg, Russia (V.S., N.R., S.G.); Saint Petersburg State University, Saint Petersburg, Russia (N.A.A., A.P., A.K.); and Department of Internal Medicine, Division of Cardiology, University of Texas Houston Medical School, Houston, Texas (I.S., E.M.)
J Pharmacol Exp Ther. 2022 May;381(2):164-175. doi: 10.1124/jpet.121.000973. Epub 2022 Feb 23.
12-lipoxigenase (12-LOX) is implicated in regulation of platelet activation processes and can be a new promising target for antiplatelet therapy. However, investigations of 12-LOX were restricted by the lack of specific and potent 12-LOX inhibitors and by controversial data concerning the role of 12-LOX metabolites in platelet functions. A novel specific 12-LOX inhibitor ML355 was shown to inhibit platelet aggregation without adverse side effects on hemostasis; however, the molecular mechanisms of its action on platelets are poorly understood. Here, we showed that ML355 inhibited platelet activation induced by thrombin or thromboxane A, but not by collagen-related peptide. ML355 blocked protein kinase B, phosphoinositide 3-kinase, and extracellular signal-regulated kinase, but not p38 kinase, spleen tyrosine kinase (Syk), or phospholipase C2 phosphorylation in activated platelets. The main inhibitory effect of low doses of ML355 (1-20 μM) on thrombin activated platelets was mediated by the decrease in reactive oxygen species level, whereas high doses of ML355 (50 μM) caused cyclic adenosine monophosphate activation. ML355 did not affect the activity of nitric oxide-dependent soluble guanylyl cyclase, nor did it affect the relaxation of preconstricted aortic rings in mice. ML355 itself did not affect platelet viability, but at 50 μM dose blocked caspase-dependent apoptosis induced by B-cell lymphoma II inhibitor ABT-737. SIGNIFICANCE STATEMENT: The current paper provides novel and original data concerning molecular mechanisms of 12-LOX inhibitor ML355 action on platelets. These data reveal antiplatelet and protective effects of ML355 on platelets and may be of importance for both antiplatelet and anticancer therapy.
12-脂氧合酶（12-LOX）参与调节血小板激活过程，可能成为抗血小板治疗的新靶点。然而，由于缺乏特异性和有效的 12-LOX 抑制剂，以及关于 12-LOX 代谢物在血小板功能中的作用的争议性数据，对 12-LOX 的研究受到限制。一种新型特异性 12-LOX 抑制剂 ML355 已被证明可抑制血小板聚集，且对止血无不良副作用；然而，其在血小板上的作用机制尚不清楚。在这里，我们发现 ML355 抑制由凝血酶或血栓烷 A 诱导的血小板激活，但不抑制由胶原相关肽诱导的血小板激活。ML355 阻断蛋白激酶 B、磷酸肌醇 3-激酶和细胞外信号调节激酶，但不阻断 p38 激酶、脾酪氨酸激酶（Syk）或磷脂酶 C2 在激活的血小板中的磷酸化。低剂量 ML355（1-20 μM）对凝血酶激活的血小板的主要抑制作用是通过降低活性氧水平介导的，而高剂量 ML355（50 μM）导致环磷酸腺苷激活。ML355 不影响一氧化氮依赖性可溶性鸟苷酸环化酶的活性，也不影响预先收缩的小鼠主动脉环的舒张。ML355 本身不影响血小板的活力，但在 50 μM 剂量下可阻断 B 细胞淋巴瘤 II 抑制剂 ABT-737 诱导的半胱天冬酶依赖性细胞凋亡。意义声明：本论文提供了关于 12-LOX 抑制剂 ML355 作用于血小板的分子机制的新的原始数据。这些数据揭示了 ML355 对血小板的抗血小板和保护作用，可能对抗血小板和抗癌治疗都很重要。
