https://orcid.org/0000-0003-0116-6443
Key Points
We developed of a high-throughput screening assay to identify small-molecule inhibitors of fibrin-mediated clot retraction
Screening 9,710 compounds from drug-repurposing libraries led to the identification of 162 inhibitors with a variety of targets.
Platelet clot retraction, the ultimate phase of platelet thrombus formation, is critical for clot stabilization. It requires functional αIIbβ3 receptors, fibrin, and the integrated actions of the actin-myosin contractile and cytoskeletal systems. Disturbances in clot retraction have been associated with both bleeding and thrombosis. We recently demonstrated that platelets treated with the αIIbβ3 antagonist peptide RGDW, which eliminates fibrinogen-mediated platelet aggregation, are still able to retract clots. We have exploited this observation to developed an unbiased, functional high-throughput assay to identify small molecule inhibitors of fibrin-mediated clot retraction adapted for a 384-well plate format. We tested 9,710 compounds from drug-repurposing libraries (DRL's). These libraries contain compounds that are either FDA-approved or have undergone preclinical/clinical development. We identified 27 compounds from the LOPAC library as inhibitors of clot retraction of which 14 are known inhibitors of platelet function. From the DRL we identified 135 compounds (1.6% hit rate). After extensive curation, these compounds were categorized based on the activity of their reported target. Multiple kinase and phosphodiesterase inhibitors with known antiplatelet effects were identified, along with multiple deubiquitination and receptor inhibitors, as well as compounds that have not previously been reported to have antiplatelet activity. Studies of one of the deubiquitination inhibitors (degrasyn) suggests that its effects are downstream of platelet-fibrinogen interactions and thus may permit the separation of platelet aggregation-mediated events from clot retraction. Additional studies of the identified compounds may lead to novel mechanisms of inhibiting thrombosis.
