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Despite their limitations, unfractionated heparin (UFH) and bivalirudin remain standard-of-care parenteral anticoagulants for percutaneous coronary intervention (PCI). We discovered novel direct thrombin inhibitors (DTIs) from tick salivary transcriptomes and optimised their pharmacologic activity. The most potent, ultravariegin, inhibits thrombin with a Ki of 4.0 pM, 445-fold better than bivalirudin. Unexpectedly, despite their greater antithrombotic effect, variegin/ultravariegin demonstrated less bleeding, achieving a 3-to-7-fold wider therapeutic index in rodent thrombosis and bleeding models. When used in combination with aspirin and ticagrelor in a porcine model, variegin/ultravariegin reduced stent thrombosis compared with antiplatelet therapy alone but achieved a 5-to-7-fold lower bleeding time than UFH/bivalirudin. Moreover, two antibodies screened from a naïve human antibody library effectively reversed the anticoagulant activity of ultravariegin, demonstrating proof-of-principle for antidote reversal. Variegin and ultravariegin are promising translational candidates for next-generation DTIs that may reduce peri-PCI bleeding in the presence of antiplatelet therapy., Bleeding complications limits the use of effective antithrombotics therapeutics. Here, the authors developed next-generation direct thrombin inhibitors with low bleeding risks as safe peri-percutaneous coronary intervention anticoagulants when used in combination with antiplatelets.

We recently discovered that Mfsd2b, which is the S1P exporter found in blood cells. Here, we report that Mfsd2b is critical for the release of all S1P species in both resting and activated platelets. We show that resting platelets store S1P in the cytoplasm. After activation, this S1P pool is delivered to the plasma membrane, where Mfsd2b is predominantly localized for export. Employing knockout mice of Mfsd2b, we reveal that platelets contribute a minor amount of plasma S1P. Nevertheless, Mfsd2b deletion in whole body or platelets impairs platelet morphology and functions. In particular, Mfsd2b knockout mice show significantly reduced thrombus formation. We show that loss of Mfsd2b affects intrinsic platelet functions as part of remarkable sphingolipid accumulation. These findings indicate that accumulation of sphingolipids including S1P by deletion of Mfsd2b strongly impairs platelet functions, which suggests that the transporter may be a target for the prevention of thrombotic disorders., The mechanisms by which platelets release sphingosine-1-phosphate (S1P) is not well characterized. Here the authors show that Mfsd2b is required for S1P release from both resting and activated platelets and that deletion of Mfsd2b impairs thrombotic functions of platelets.

Background We have isolated, variegin, a unique direct thrombin inhibitor (DTI) from tropical bont tick Amblyomma variegatum. Variegin inhibits thrombin active site and exosite-1 with an inhibitory constant of 0.3 nM (9-fold better than bivalirudin). It is also >5 orders of magnitude more selective for thrombin than other blood coagulation serine proteases. Variegin has a plasma half-life of 50 minutes (compared with bivalirudin 25 minutes and heparin ∼ hours). Purpose We aimed to develop variegin into a parenteral anticoagulant for percutaneous coronary intervention (PCI) and tested variegin in several pre-clinical models. Methods In rats, variegin was tested for efficacy (anticoagulation intensity) in a FeCl3-induced carotid artery thrombosis model while safety (bleeding risk) was tested in a tail incision model that recapitulated the time-frame of PCI (∼1 hour) in humans (time-response model). In pigs, an extracorporeal circuit with modified Badimon chambers containing coronary stents was used to assess efficacy, while bleeding risk was evaluated through needle-induced injury on a superficial ear vein, with or without concurrent administration of aspirin and ticagrelor (DAPT). Unfractionated heparin (UFH) and bivalirudin at dosages recommended for PCI were used as references. Ex vivo clotting analyses including thrombin generation test, rotational thromboelastometry, activated partial thromboplastin time and clot waveform analysis were performed in human blood spiked with DAPT and the three anticoagulants. Results In the rat time-response model, a single variegin bolus conferred better antithrombotic effect than a continuous infusion of bivalirudin and more rapid recovery of haemostasis than a single bolus of heparin. In the porcine ex vivo model, without DAPT, UFH, bivalirudin and 1 mg/kg variegin reduced stent thrombus by 35% (P Figure 1 Conclusion In the presence of aspirin and ticagrelor, a low dose of variegin, a novel direct thrombin inhibitor, achieved superior antithrombotic effect with significantly lower bleeding risk than heparin or bivalirudin in pre-clinical PCI models.