First Selective 12-LOX Inhibitor, ML355, Impairs Thrombus Formation and Vessel Occlusion In Vivo With Minimal Effects on Hemostasis

Abstract

Objective Adequate platelet reactivity is required for maintain hemostasis. However, excessive platelet reactivity can also lead to the formation of occlusive thrombi. 12-lipoxygenase (12-LOX), an oxygenase highly expressed in the platelet, has been demonstrated to regulate platelet function and thrombosis ex vivo, supporting a key role for 12-LOX in the regulation of in vivo thrombosis. However, the ability to pharmacologically target 12-LOX in vivo has not been established to date. Here, we studied the effect of the first highly selective 12-LOX inhibitor, ML355, on in vivo thrombosis and hemostasis. Approach and Results ML355 dose-dependently inhibited human platelet aggregation and 12-LOX oxylipin production, as confirmed by mass spectrometry. Interestingly, the antiplatelet effects of ML355 were reversed following exposure to high concentrations of thrombin in vitro. Ex vivo flow chamber assays confirmed that human platelet adhesion and thrombus formation at arterial shear over collagen were attenuated in whole blood treated with ML355 comparable to aspirin. Oral administration of ML355 in mice showed reasonable plasma drug levels by pharmacokinetic assessment. ML355 treatment impaired thrombus growth and vessel occlusion in FeCl3-induced mesenteric and laser-induced cremaster arteriole thrombosis models in mice. Importantly, hemostatic plug formation and bleeding following treatment with ML355 was minimal in mice in response to laser ablation on the saphenous vein or in a cremaster microvasculature laser-induced rupture model. Conclusions Our data strongly supports 12-LOX as a key determinant of platelet reactivity in vivo and inhibition of platelet 12-LOX with ML355 may represent a new class of antiplatelet therapy.

Activation of metabolite receptor GPR91 promotes platelet aggregation and transcellular biosynthesis of leukotriene C4

“…Low‐dose ASA targets COX and is widely used for this purpose, while thromboxane receptor antagonists have been developed without making it to clinical settings. 12‐LOX and 12‐HETE have been implicated in platelet functions and thrombus formation and recently, ML355, a potent and selective 12‐LOX inhibitor, was suggested to impair platelet reactivity and clotting . In fact, as ASA could only partially inhibit succinate potentiated platelet aggregation (Figure B), the effectiveness of ML355 in the current study supports targeting 12‐LOX as a potential new antiplatelet therapy, especially in patients with ASA resistance or intolerance.…”

“…12-LOX and 12-HETE have been implicated in platelet functions and thrombus formation 24 and recently, ML355, a potent and selective 12-LOX inhibitor, was suggested to impair platelet reactivity and clotting. 25 In fact, as ASA could only partially inhibit succinate potentiated platelet aggregation 12 (Figure 3B), the effectiveness of ML355 in inflammation, especially in aspirin-exacerbated respiratory disease. 28 In fact, this notion can be extended to atherosclerosis and associated cardiovascular diseases.…”

Novel aspects of antiplatelet therapy in cardiovascular disease

“…However, early switching from prasugrel or ticagrelor to clopidogrel based on the results of platelet function testing may become an alternative option to reduce bleeding risk while maintaining adequate platelet inhibition following ACS, though more study is needed . Finally, new antiplatelet agents have yielded promising results in preclinical trials and may in the future become meaningful additions to the current pharmacological armamentarium in cardiovascular disease …”

“…They found a dose‐dependent decrease of human platelet aggregation by ML355, an effect that was reversed after exposure to high concentrations of thrombin in vitro. Moreover, oral administration of ML355 in mice reduced thrombus formation and vessel occlusion in FeCl 3 ‐induced mesenteric and laser‐induced cremaster arteriole thrombosis models with only minimal effects on hemostasis …”

“…Recently, Adili et al. studied the impact of the selective 12‐LOX inhibitor ML355 on thrombosis and hemostasis (Table ) . They found a dose‐dependent decrease of human platelet aggregation by ML355, an effect that was reversed after exposure to high concentrations of thrombin in vitro.…”