Endothelial vascular cell adhesion molecule 1 is a marker for high-risk carotid plaques and target for ultrasound molecular imaging

Background: Molecular imaging of carotid plaque vulnerability to atheroembolic events is likely to lead to improvements in selection of patients for carotid endarterectomy (CEA). The aims of this study were to assess the relative value of endothelial inflammatory markers for this application and to develop molecular ultrasound contrast agents for their imaging. Methods: Human CEA specimens were obtained prospectively from asymptomatic (30) and symptomatic (30) patients. Plaques were assessed by semiquantitative immunohistochemistry for vascular cell adhesion molecule 1 (VCAM-1), lectin-like oxidized low-density lipoprotein receptor 1, P-selectin, and von Willebrand factor. Established small peptide ligands to each of these targets were then synthesized and covalently conjugated to the surface of lipid-shelled microbubble ultrasound contrast agents, which were then evaluated in a flow chamber for binding kinetics to activated human aortic endothelial cells under variable shear conditions. Results: Expression of VCAM-1 on the endothelium of CEA specimens from symptomatic patients was 2.4-fold greater than that from asymptomatic patients (P <.01). Expression was not significantly different between groups for P-selectin (P =.43), von Willebrand factor (P =.59), or lectin-like oxidized low-density lipoprotein receptor 1 (P =.99). Although most plaques from asymptomatic patients displayed low VCAM-1 expression, approximately one in five expressed high VCAM-1 similar to plaques from symptomatic patients. In vitro flow chamber experiments demonstrated that VCAM-1-targeted microbubbles bind cells that express VCAM-1, even under high-shear conditions that approximate those found in human carotid arteries, whereas binding efficiency was lower for the other agents. Conclusions: VCAM-1 displays significantly higher expression on high-risk (symptomatic) vs low-risk (asymptomatic) carotid plaques. Ultrasound contrast agents bearing ligands for VCAM-1 can sustain high-shear attachment and may be useful for identifying patients in whom more aggressive treatment is warranted. Clinical Relevance: A noninvasive method for stroke risk assessment in patients with asymptomatic carotid atherosclerosis is long overdue and key to improving treatment. Our data are the first to show that vascular cell adhesion molecule 1 expression is significantly increased in high-risk compared with low-risk carotid plaques in humans (P =.0005), and they demonstrate a novel targeted ultrasound agent that could be used for noninvasive vascular cell adhesion molecule 1 evaluation in patients. Furthermore, these data build on our understanding of carotid plaque biology and define tools for in vivo molecular analysis of atherosclerosis in animal models as well as in humans. This ultrasound technology would be easily translatable to a vascular outpatient setting.