Protein disulfide isomerase inhibitors constitute a new class of antithrombotic agents

Abstract

Thrombosis, or blood clot formation, and its sequelae remain a leading cause of morbidity and mortality, and recurrent thrombosis is common despite current optimal therapy. Protein disulfide isomerase (PDI) is an oxidoreductase that has recently been shown to participate in thrombus formation. While currently available antithrombotic agents inhibit either platelet aggregation or fibrin generation, inhibition of secreted PDI blocks the earliest stages of thrombus formation, suppressing both pathways. Here, we explored extracellular PDI as an alternative target of antithrombotic therapy. A high-throughput screen identified quercetin-3-rutinoside as an inhibitor of PDI reductase activity in vitro. Inhibition of PDI was selective, as quercetin-3-rutinoside failed to inhibit the reductase activity of several other thiol isomerases found in the vasculature. Cellular assays showed that quercetin-3-rutinoside inhibited aggregation of human and mouse platelets and endothelial cell-mediated fibrin generation in human endothelial cells. Using intravital microscopy in mice, we demonstrated that quercetin-3-rutinoside blocks thrombus formation in vivo by inhibiting PDI. Infusion of recombinant PDI reversed the antithrombotic effect of quercetin-3-rutinoside. Thus, PDI is a viable target for small molecule inhibition of thrombus formation, and its inhibition may prove to be a useful adjunct in refractory thrombotic diseases that are not controlled with conventional antithrombotic agents.

Inhibition of thiol isomerase activity diminishes endothelial activation of plasminogen, but not of protein C

“…In our experiments, plasminogen activation and fibrin lysis were inhibited by RL90 antibody, which is known to cross react with ERp57 [10], but also (to a lesser degree) by rutin, which is a more specific inhibitor of PDI [38]. Addition of exogenous PDI produced a statistically significant increase in the rate of plasminogen activation.…”

Rutin- potent natural thrombolytic agent

“…Thus, rutin binds to and reversibly inhibits PDI but shows only minimal activity towards other extracellular thiol isomerases present in the vasculature. Evaluation of the effect of flavonol ingestion on cardio-vascular events demonstrated protection from myocardial infarction and stroke with increased intake (Jasuja et al, 2012).…”

“…Thus, PDI inhibition is a viable target for small molecule inhibition of thrombus formation and its inhibition can prove to be a useful adjunct in refractoty thrombotic diseases that are not controlled with conventional anti-thrombotic agents (Jasuja et al, 2012).…”

“…Plasma levels of rutin decrease rapidly after either intra-venous or oral administration (Jasuja et al, 2012). Ingested rutin is hydrolyzed to quercetin in the intestine and further changed to other conjugated metabolites of quercetin (Gee et al, 2000).…”

“…Thrombosis or blood clot formation and its consequences remain a leading cause of morbidity and mortality, and recurrent thrombosis is common despite current optimal therapy (Jasuja et al, 2012). Clots in arteries are platelet rich where as in veins they are fibrin rich.…”