Kalathottukaren Manu Thomas, Abraham Libin, Kapopara Piyushkumar R, Lai Benjamin F L, Shenoi Rajesh A, Rosell Federico I, Conway Edward M, Pryzdial Edward L G, Morrissey James H, Haynes Charles A, Kizhakkedathu Jayachandran N
Centre for Blood Research.
Department of Pathology and Laboratory Medicine.
Blood. 2017 Mar 9;129(10):1368-1379. doi: 10.1182/blood-2016-10-747915. Epub 2016 Dec 29.
Anticoagulant therapy-associated bleeding and pathological thrombosis pose serious risks to hospitalized patients. Both complications could be mitigated by developing new therapeutics that safely neutralize anticoagulant activity and inhibit activators of the intrinsic blood clotting pathway, such as polyphosphate (polyP) and extracellular nucleic acids. The latter strategy could reduce the use of anticoagulants, potentially decreasing bleeding events. However, previously described cationic inhibitors of polyP and extracellular nucleic acids exhibit both nonspecific binding and adverse effects on blood clotting that limit their use. Indeed, the polycation used to counteract heparin-associated bleeding in surgical settings, protamine, exhibits adverse effects. To address these clinical shortcomings, we developed a synthetic polycation, Universal Heparin Reversal Agent (UHRA), which is nontoxic and can neutralize the anticoagulant activity of heparins and the prothrombotic activity of polyP. Sharply contrasting protamine, we show that UHRA does not interact with fibrinogen, affect fibrin polymerization during clot formation, or abrogate plasma clotting. Using scanning electron microscopy, confocal microscopy, and clot lysis assays, we confirm that UHRA does not incorporate into clots, and that clots are stable with normal fibrin morphology. Conversely, protamine binds to the fibrin clot, which could explain how protamine instigates clot lysis and increases bleeding after surgery. Finally, studies in mice reveal that UHRA reverses heparin anticoagulant activity without the lung injury seen with protamine. The data presented here illustrate that UHRA could be safely used as an antidote during adverse therapeutic modulation of hemostasis.
抗凝治疗相关的出血和病理性血栓形成对住院患者构成严重风险。通过开发能够安全中和抗凝活性并抑制内源性血液凝固途径激活剂（如多聚磷酸盐（polyP）和细胞外核酸）的新型疗法，可以减轻这两种并发症。后一种策略可以减少抗凝剂的使用，潜在地减少出血事件。然而，先前描述的多聚磷酸盐和细胞外核酸的阳离子抑制剂表现出非特异性结合以及对血液凝固的不利影响，这限制了它们的使用。事实上，用于抵消手术中肝素相关出血的聚阳离子鱼精蛋白就有不良影响。为了解决这些临床缺陷，我们开发了一种合成聚阳离子——通用肝素逆转剂（UHRA），它无毒，能够中和肝素的抗凝活性以及多聚磷酸盐的促血栓形成活性。与鱼精蛋白形成鲜明对比的是，我们发现UHRA不与纤维蛋白原相互作用，不影响凝血过程中纤维蛋白的聚合，也不会消除血浆凝血。通过扫描电子显微镜、共聚焦显微镜和凝块溶解试验，我们证实UHRA不会掺入凝块中，并且凝块具有正常的纤维蛋白形态，是稳定的。相反，鱼精蛋白会与纤维蛋白凝块结合，这可以解释鱼精蛋白如何引发凝块溶解并增加术后出血。最后，在小鼠身上进行的研究表明，UHRA能够逆转肝素的抗凝活性，且不会出现鱼精蛋白导致的肺损伤。本文所展示的数据表明，UHRA可以安全地用作止血不良治疗调节过程中的解毒剂。
