OC 77.1 - Inhibition of the Class II PI 3-kinase, PI3KC2a, prevents thrombosis in hyperlipidaemic mice

Background: Inhibition of platelet activation is the basis of standard-of-care therapy for the prevention of arterial thrombosis. A major limitation of current anti-platelet agents is inadequate efficacy in high-risk populations, such as those with hyperlipidaemia. The class II PI3-kinase, PI3KC2α, is an intracellular signaling enzyme that has been shown to be important for platelet function. Targeting PI3KC2α in mice results in striking anti-thrombotic effects with preserved haemostatic function, making PI3KC2α a promising candidate to pursue improved anti-thrombotic therapy. However, whether these effects are preserved in the setting of hyperlipidaemia remains unknown.

Aims: To examine whether genetic deficiency or pharmacological inhibition of PI3KC2α provides anti-thrombotic effects in blood taken from hyperlipidaemic mice.

Methods: Thrombosis was evaluated using an ex vivo microfluidic whole-blood continuous perfusion assay. Whole blood taken from hyperlipidaemic ApoE-/- mice was flowed at arterial shear rates over type I collagen fibers and thrombus volume measured in real-time via confocal microscopy for five minutes. The effect of genetic deficiency of PI3KC2α was examined by using blood from mice with combined deficiency of ApoE-/- and PI3KC2α, while a recently-developed PI3KC2α inhibitor, MIPS-21335 (10 µM), pretreated in whole blood from ApoE-/- mice to evaluate the effect of acute pharmacological inhibition of PI3KC2α.

Results: Hyperlipidaemia had the anticipated pro-thrombotic effect, with thrombus volume 1.5-fold greater in blood from ApoE-/- mice than in blood from wild-type mice (p=0.009). This increased thrombosis in blood from ApoE-/- mice was entirely prevented in blood from mice with a combined deficiency of ApoE and PI3KC2α (p=0.007). Acute inhibition of PI3KC2α with MIPS-21335 also reduced thrombosis in blood from ApoE-/- mice, albeit to a lesser extent (p=0.066).

Conclusion(s): These findings demonstrate that the anti-thrombotic effect of PI3KC2α-deficiency or -inhibition observed in normolipidaemia is largely retained in the face of hyperlipidaemia and suggests PI3KC2α is a promising target for anti-thrombotic therapy in this high-risk population.