PB1218 - Aptamer Inhibitor of Von Willebrand Factor Induces Thrombolysis and Improves Vascular Patency Upon Immediate and Delayed Treatment in a Microfluidic Model of Stroke

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

Aims: This study aims to utilize a microfluidic model of arterial occlusion and prolonged retraction to evaluate BB-031 thrombolysis compared to current therapeutic alternatives.

Methods: Heparinized whole blood from healthy volunteers (n=6-10/group) was incubated with fluorescent anti-CD41 and perfused via gravity through a collagen-coated, stenosed Y-channel microfluidic channel to simulate complete arterial occlusion. Thrombosis was imaged in realtime. After 0, 3, or 6hrs post-occlusion, the Y-system allowed for reperfusion immediately upstream with matched donor blood containing BB-031, alteplase (ALT), tenecteplase (TEN), or vehicle. Thrombolysis and patency were monitored for 2hrs via outflow mass measurement and z-stacked immunofluorescent imaging. Mixed-effects analysis was conducted on patency data.

Results: After 6hrs of retraction, thrombi substantially remodeled. Fluorescence intensity-derived profiles of occlusive clots demonstrate even distribution post-occlusion shifting to a central peak after 6hrs (Fig1). Allowing no retraction (0hrs), 1692nM BB-031 increased outflow by 0.250±0.629g compared to 0.031±0.065g with vehicle, 0.074±0.061g with 0.7nM ALT, and 0.101±0.150g with 0.7nM TEN (Fig2). After 3hrs retraction, BB-031 outflow was 0.951±1.279g compared to 0.301±0.380g with vehicle, 0.412±0.681g with ALT, and 0.569±0.714g with TEN. After 6hrs retraction, outflow increased by 1.354±1.433g with BB-031, 0.368±0.538g with vehicle, 0.158±0.195g with ALT, and 0.520±0.539g with TEN (retraction time: p=0.0147) (treatment: p=0.0490). Representative z-stacked renderings pre- and post-reperfusion from 0hr retraction clots show a volumetric decrease in thrombus size of 39% with BB-031 compared to 25% and 18% reduction with vehicle and ALT, respectively, in a donor-matched dataset.

Conclusion(s): This study establishes an in vitro high-throughput microfluidic platform to study arterial occlusion, clot retraction, drug delivery, and thrombolysis. BB-031 improves patency in this model compared to current standards of care, which is enhanced 3hrs and 6hrs post-occlusion, potentially due to VWF-dependent lysis reducing thrombus volume.