Thrombolysis Resistance in Acute Ischemic Stroke - PROJECT SUMMARY This project aims to improve outcomes for patients with large vessel occlusion (LVO) acute ischemic stroke (AIS), a leading cause of global morbidity and mortality. Although mechanical thrombectomy is increasingly adopted, intravenous thrombolysis (IVT) with tissue plasminogen activator (tPA) remains the most accessible therapy. However, IVT often fails to achieve successful recanalization in most eligible patients, which reflects clot resistance to tPA therapy. Recent studies highlight the structural and cellular heterogeneity of AIS clot, but how clot metabolic changes contribute to tPA resistance remains poorly defined. Our preliminary studies indicate that hypoxia core develops within clot, driving anerobic glycolysis with lactate accumulation, ionic imbalance, and pH reduction. This acidic microenvironment impairs tPA activity, which requires a neutral pH for optimal function. In addition, we detected bacteria and their metabolites within AIS clots and showed presence of viral genome in clots. These microbial signals can promote neutrophil extracellular traps (NETs), intensify hypoxia, and increase plasminogen activator inhibitor-1 (PAI-1) activity, all of which can collectivity reduce fibrinolysis. To address these challenges, we will evaluate nanoparticle-based, fibrin-targeted tPA delivery. Specifically, this project will 1) define metabolic and hypoxic mechanisms underlying thrombolysis resistance in human AIS clots using multi-omics, histology, and advanced pH imaging, 2) characterize microbial contributions to resistance, assessing bacterial and viral presence, metabolites, and their effects on clot structure, NET formation, and PAI-1 activity, and 3) develop and test fibrin-targeted, pH-responsive nanoparticles designed to deliver and stabilize tPA within acidic thrombus cores, thereby restoring its local activity and improving lysis efficiency. By integrating molecular, microbial, and engineering approaches, this research will elucidate the mechanisms driving tPA resistance and introduce a targeted strategy to overcome it. The findings are expected to enhance recanalization rates, minimize complications, and transform intravenous thrombolysis from a uniform protocol into a precision-guided therapy for AIS.
