Regulating Cellular Metabolism and Energy Flow to Target Thrombosis

Project summary Platelets and leukocytes play a crucial role in thrombotic cardiovascular disease (CVD) pathophysiology. While conventional antiplatelet therapies effectively reduce mortality and nonfatal events in over 50% of high-risk patients, they also increase the risk of bleeding, a significant concern in modern CVD treatment. Given that the interplay between platelets and leukocytes contributes to thrombotic vascular occlusion, an ideal therapeutic approach would be the one that inhibits thrombo-inflammatory responses without increasing bleeding risk. We propose to test an innovative concept that metabolic reprogramming through cytosolic malic enzyme 1 (ME1) inhibition in platelets and leukocytes will inhibit their hyperactivation and decrease susceptibility to arterial thrombosis while minimal bleeding risk. The premise is that ME1, a nicotinamide adenine dinucleotide phosphate (NADP+)-dependent enzyme, plays a crucial role in energy metabolism by facilitating the conversion of malate to pyruvate. This process links glycolysis to the tricarboxylic acid cycle, with the resulting pyruvate transported to the mitochondria. The NADP(H) generated is utilized by NADP(H)-oxidases, leading to increased reactive oxygen species that drive platelet and leukocyte activation. Pilot studies have demonstrated that reducing ME1 activity in human platelets or genetically ablating ME1 in mouse platelets inhibits agonist- induced platelet aggregation, total ATP production, and NADP(H) generation. Platelet-specific ME1-deficient mice exhibited reduced susceptibility to arterial thrombosis without compromising hemostasis. Additionally, inhibition of ME1 in human neutrophils or deficiency of ME1 in mouse neutrophils inhibited neutrophil extracellular trap formation. To pursue this research, we will utilize human samples, novel cell-specific mutant mice, experimental thrombosis models, and a range of molecular and immunological assays alongside metabolomics. Our specific aims include 1) assessing the regulatory role of ME1 in platelet and leukocyte function, 2) evaluating the regulatory role of ME1 in in vivo thrombosis and hemostasis models, and 3) determining if pharmacological inhibition of ME1 activity can reduce platelet hyperactivation and thrombosis in models with preexisting comorbidities. Our team possesses extensive expertise, enhancing this project's feasibility and potential success. The proposed research is significant because existing antiplatelet drugs carry bleeding risks and simultaneously do not target both platelets and leukocytes, which play a crucial role in triggering a thrombotic event. Reprogramming metabolic and energy pathways in platelets and leukocytes through a single target to inhibit arterial thrombosis represents a novel strategy that could be investigated for efficacy and safety in future studies.