OC 58.4 - Aptamer Inhibitor of Von Willebrand Factor Improves Vascular Patency in a Microfluidic Model of Ischemic Stroke by Prevention of Platelet-Mediated Reocclusion

Background: Von Willebrand factor (VWF) is mechanistically critical in arterial thrombosis. Previous studies have shown aptamer BB-031, a VWF-A1 inhibitor, induces thrombolysis via a mechanism yet to be elucidated.

Aims: This study utilizes a microfluidic model of arterial thrombosis to evaluate BB-031 against the current therapeutic standard for ischemic stroke.

Methods: Heparinized blood (paired n=10/group) incubated with fluorescent anti-CD41 was perfused via gravity through a collagen-coated, stenosed, Y-chamber to simulate arterial occlusion. 0, 3, or 6hrs post-occlusion, blood labelled with disparate anti-CD41 and dosed with vehicle or treatment was reperfused at a pressure head mimicking the middle cerebral artery for 2hrs. Reperfusion was achieved without disturbing the thrombus. Thrombus and patency were monitored via real-time imaging, immunofluorescent z-stacks, and outflow mass. Combined patency data were analyzed via one-way analysis of variance and Tukey’s test at 95% CI. A separate subset of experiments(n=3) analyzed platelet and VWF incorporation in clots fixed 3hrs post-occlusion via paired t-test.

Results: Thrombi substantially remodeled post-occlusion(Fig1A-B). At all timepoints, BB-031 increased mean outflow mass(Fig1C). In brief, 6hrs post-occlusion, BB-031 increased outflow 1.354±1.433g compared to 0.368±0.538g with vehicle(mean±SD). Inclusive of all timepoints, BB-031 maximum outflow mass was significantly higher compared to controls and other treatment groups(Fig1D). Z-stacked renderings pre- and post-reperfusion(Fig1E) show volumetric decreases in total thrombus size with BB-031, especially at 6hrs post-occlusion(Fig1F). While all treatment conditions showed an average decrease in original thrombus volume(Fig1G), BB-031 showed minimal deposition of newly reperfused platelets(Fig1H). Dynamic platelet and VWF incorporation post thrombus formation is illustrated in confocal images of fixed clots, which was reduced with BB-031 treatment(Fig2A-B).

Conclusion(s): This study establishes an in vitro high-throughput microfluidic platform studying arterial occlusion, clot retraction, drug delivery, and recanalization. BB-031 could be a safe and more efficacious therapeutic alternative to current standards of care, especially after prolonged periods of stroke onset, by reducing re-occlusive platelet deposition.