Targeted ultrasound contrast agent for molecular imaging of inflammation in high-shear flow

Targeted ultrasound contrast materials (gas‐filled microbubbles carrying ligands to endothelial selectins or integrins) have been investigated as potential molecular imaging agents. Such microbubbles normally exhibit good targeting capability at the slower flow conditions. However, in the conditions of vigorous flow, binding may be limited. Here, we describe a microbubble capable of efficient binding to targets both in slow and fast flow (exceeding 4 dyne/cm2 wall shear stress) using a clustered polymeric form of the fast‐binding selectin ligand sialyl LewisX. Microbubbles were prepared from decafluorobutane gas and stabilized with a monolayer of phosphatidylcholine, PEG stearate and biotin‐PEG‐lipid. Biotinylated PSLex (sialyl LewisX polyacrylamide) or biotinylated anti‐P‐selectin antibody (RB40.34) was attached to microbubbles via a streptavidin bridge. In a parallel plate flow chamber targeted adhesion model, PSLex bubbles demonstrated specific adhesion, retention and slow rolling on P‐selectin‐coated plates. Efficiency of firm targeted adhesion to a P‐selectin surface (140 molecules/µm2) was comparable for antibody‐carrying bubbles and PSLex‐targeted bubbles at 0.68 dyne/cm2 shear stress. At fast flow (4.45 dyne/cm2), PSLex‐targeted bubbles maintained their ability to bind, while antibody‐mediated targeting dropped more than 20‐fold. At lower surface density of P‐selectin (7 molecules/µm2), targeting via PSLex was more efficient than via antibody under all the flow conditions tested. Negative control casein‐coated plates did not retain bubbles in the range of flow conditions studied. To confirm echogenicity, targeted PSLex‐bubbles were visualized on P‐selectin‐coated polystyrene plates by ultrasound imaging with a clinical scanner operated in pulse inversion mode; control plates lacking targeted bubbles did not show significant acoustic backscatter. In vivo, in a murine model of inflammation in the femoral vein setting, targeting efficacy of intravenously administered PSLex‐microbubbles was comparable with targeting mediated by anti‐P‐selectin antibody, and significantly exceeded the accumulation of non‐targeted control bubbles. In the inflamed femoral artery setting, PSLex‐mediated microbubble targeting was superior to antibody‐mediated targeting. Copyright © 2006 John Wiley & Sons, Ltd. Ultrasound contrast microbubbles were decorated with biotinylated polyacrylamide sialyl Lewisx (PSLex) via streptavidin coupling. P‐selectin targeting of PSLex‐bubble