Blocking P2Y14R Emerges to Combat Venous Thromboembolism via Neutrophil-Plaque Interaction

Background

Venous thromboembolism (VTE), encompassing deep vein thrombosis (DVT) and pulmonary embolism, remains a leading cause of cardiovascular mortality globally. Despite advances in anticoagulant therapies, significant gaps persist in managing thrombo-inflammatory complications. Neutrophil extracellular traps (NETs) – web-like DNA structures decorated with histones and antimicrobial proteins – have emerged as critical mediators of pathological thrombosis by promoting platelet aggregation and endothelial activation.

The P2Y14 receptor (P2Y14R), a G-protein-coupled receptor activated by extracellular UDP-glucose, is highly expressed on neutrophils and implicated in inflammatory responses. However, its precise role in VTE pathogenesis and the potential for therapeutic targeting remained unexplored until this landmark study.

Study Design

This multidisciplinary investigation employed a translational approach combining clinical observations with mechanistic experiments:

• Clinical cohort: Analyzed neutrophil P2Y14R expression in venous blood from VTE patients
• Animal models: Utilized DVT and disseminated intravascular coagulation models in wild-type and neutrophil-specific P2Y14R knockout mice
• Mechanistic studies: Integrated transcriptomics, phosphoproteomics, and immunofluorescence to dissect signaling pathways
• Drug discovery: Screened a repurposing drug library via high-throughput Glide docking for P2Y14R antagonists

Key Findings

The study yielded several groundbreaking discoveries:

Clinical Correlations

Neutrophils from VTE patients demonstrated significant upregulation of P2Y14R compared to controls, establishing a clinical association between receptor expression and thrombotic risk.

Genetic Evidence

Neutrophil-specific P2Y14R deficiency in mice resulted in:

• 57% reduction in venous thrombus weight
• 73% decrease in NET formation markers (citrullinated histone H3)
• Improved microvascular perfusion in disseminated intravascular coagulation

Mechanistic Insights

P2Y14R deletion activated a novel PKA/AKAP13/RhoA signaling axis:

1. Enhanced PKA-mediated phosphorylation of AKAP13 scaffolding protein
2. Subsequent inhibition of RhoA GTPase activation
3. Impaired cytoskeletal rearrangements required for neutrophil-platelet aggregation
4. Reduced NETosis and thrombo-inflammation

Therapeutic Discovery

The study identified proglumide, an existing cholecystokinin receptor antagonist, as a potent P2Y14R inhibitor with:

• Superior binding affinity (-9.3 kcal/mol) in docking studies
• Confirmed antagonism in calcium flux assays
• In vivo efficacy matching genetic P2Y14R deletion

Expert Commentary

“This work elegantly bridges molecular discovery with therapeutic potential,” notes Dr. Lisa Baumann Kreuziger (Medical College of Wisconsin), a thrombosis specialist unaffiliated with the study. “By elucidating how P2Y14R orchestrates NETosis through RhoA cytoskeletal dynamics, it offers a precision medicine approach for VTE patients with neutrophil-driven pathology.”

The researchers acknowledge limitations including the need for human pharmacokinetic studies with proglumide and investigation of potential off-target effects in chronic P2Y14R inhibition.

Conclusion

This study establishes P2Y14R as a master regulator of thrombo-inflammatory NET formation and validates pharmacological P2Y14R blockade as a viable strategy for VTE intervention. The rapid repurposing potential of proglumide could accelerate clinical translation, offering hope for patients refractory to current anticoagulants.

Funding and Registration

Supported by National Natural Science Foundation of China grants. Preclinical data available upon request; human trials in planning phase (NCT pending).