Abstract 13226: Dual Microbubble Targeting Improves in vitro Capture Efficiency of Ultrasound Micro¬Bubbles Targeted to Selectins and Activated Platelets Although not Providing Additional Signal Effect for in vivo Imaging of Arterial Thrombosis

Abstract

Purpose. Platelets can be found on the surface of inflamed and ruptured plaques. Hence, membrane proteins of activated platelets are of special interest regarding molecular imaging of atherosclerosis. Capture and firm adhesion under high shear stress are important for the success of ultrasound molecular imaging in atherosclerosis. Therefore, we developed and studied a targeted microbubble (MB) with two ligands bound on the surface: an antibody against ligand-induced binding sites of the activated GP IIb/IIIa receptor (LIBS-IgG), and the fast binding selectin ligand sialyl Lewis^a (sLe^a).

Methods. LIBS and sLe^a were coupled to MB using a biotin-streptavidin system, either individually (MB_LIBS and MB_sLe), or dually (MB_dual). Control MB (MBCtr) bearing a scrambled sLea and an isotype control antibody were also prepared. MB adhesion on platelets was assessed in vitro in a flow chamber at different shear rates. In-vivo, a non-occlusive carotid artery thrombosis was induced by ferric chloride in a mouse model, and MB adhesion was assessed by ultrasound.

Results. In the flow chamber, all targeted MB showed significantly increased adhesion to platelets compared to MBCtr. Whereas MB_sLe were slowly rolling on the platelet surface, MB_LIBS and MB_dual showed firm adhesion. MB_dual showed a higher capture efficiency than single targeted MB_LIBS and MB_sLe (p<0.05). On in vivo ultrasound imaging, MB_sLe and MB_dual showed a significantly higher acoustic intensity than MBCtr (p<0.05), however, there was no increase in mean signal intensity for MB_dual over MB_sLe (p>0.05). The degree of carotid stenosis on histology correlated well with the ultrasound acoustic intensity for MB_sLe and MB_dual (p<0.05).

Conclusions. Dual targeting of microbubbles using a combination of fast binding polymers and specific antibodies is a promising strategy to obtain firm adhesion on activated platelets under high shear flow conditions in vitro. Noninvasive imaging of activated platelets in arteries with high shear stress is feasible. However, the advantages of the dual targeting strategy that were apparent in the in vitro setting are not readily translatable into in vivo models.