Generation of an Anticoagulant Aptamer that Targets Factor V/Va and Disrupts the FVa-Membrane Interaction in Normal and COVID-19 Patient Samples. Soule et al

Coagulation cofactors profoundly regulate hemostasis and are appealing targets for anticoagulants. However, targeting such proteins has been challenging because they lack an active site. To address this, we isolate an RNA aptamer termed T18.3 that binds to both Factor V and Va (FV/FVa) with nanomolar affinity and demonstrates clinically relevant anticoagulant activity in both plasma and whole blood. The aptamer also shows synergy with low molecular weight heparin, and delivers potent anticoagulation in plasma collected from COVID-19 patients. Moreover, the aptamer's anticoagulant activity can be rapidly and efficiently reversed using protamine sulfate, which potentially allows fine-tuning of aptamer’s activity post-administration. We further show that the aptamer achieves its anticoagulant activity by abrogating FV/FVa interaction with phospholipid membranes. Our success in generating an anticoagulant aptamer targeting FV/Va demonstrates the feasibility of using cofactor-binding aptamers as therapeutic protein-inhibitors and reveals a unconventional working mechanism of aptamer by interrupting protein-membrane interactions.

凝血辅因子 (coagulation cofactors) 可显著调控机体止血 (hemostasis) 过程，是抗凝药物的极具吸引力的研发靶点。然而，由于此类蛋白缺乏活性位点，靶向它们的研发颇具挑战。为解决这一难题，我们分离得到一种命名为T18.3的RNA适配体 (RNA aptamer)，该适配体以纳摩尔级亲和力结合凝血因子V及其活化形式 (FV/FVa)，并在血浆与全血中均展现出具有临床相关性的抗凝活性。
该适配体还可与低分子量肝素 (low molecular weight heparin) 产生协同效应，并在新冠患者采集的血浆中呈现出强效抗凝作用。此外，使用硫酸鱼精蛋白 (protamine sulfate) 可快速且高效地逆转该适配体的抗凝活性，这为给药后精准调控适配体活性提供了潜在可能。
我们进一步证实，该适配体通过阻断FV/FVa与磷脂膜 (phospholipid membranes) 的相互作用来实现其抗凝活性。
本研究成功开发出靶向FV/FVa的抗凝适配体，这证明了利用结合辅因子的适配体作为治疗性蛋白抑制剂的可行性，同时揭示了适配体通过阻断蛋白-膜相互作用发挥功能的非常规作用机制。