Development of Molecular Magnetic Resonance Imaging Tools for Risk Stratification of Carotid Atherosclerotic Disease Using Dual-Targeted Microparticles of Iron Oxide

Abstract

Identification of patients with high-risk asymptomatic carotid plaques remains a challenging but crucial step in stroke prevention. Inflammation is the key factor that drives plaque instability. Currently, there is no imaging tool in routine clinical practice to assess the inflammatory status within atherosclerotic plaques. We have developed a molecular magnetic resonance imaging (MRI) tool to quantitatively report the inflammatory activity in atherosclerosis using dual-targeted microparticles of iron oxide (DT-MPIO) against P-selectin and VCAM-1 as a smart MRI probe. A periarterial cuff was used to generate plaques with varying degree of phenotypes, inflammation and risk levels at specific locations along the same single carotid artery in an Apolipoprotein-E-deficient mouse model. Using this platform, we demonstrated that in vivo DT-MPIO-enhanced MRI can (i) target high-risk vulnerable plaques, (ii) differentiate the heterogeneity (i.e. high vs intermediate vs low-risk plaques) within the asymptomatic plaque population and (iii) quantitatively report the inflammatory activity of local plaques in carotid artery. This novel molecular MRI tool may allow characterisation of plaque vulnerability and quantitative reporting of inflammatory status in atherosclerosis. This would permit accurate risk stratification by identifying high-risk asymptomatic individual patients for prophylactic carotid intervention, expediting early stroke prevention and paving the way for personalised management of carotid atherosclerotic disease.

Development of Molecular Magnetic Resonance Imaging Tools for Longitudinal Tracking of Carotid Atherosclerotic Disease Using Fast Imaging with Steady-State Precession

“…The synthesis of contrast agents dual-targeted (DT) and control IgG-MPIOs were reported previously [8]. Briefly, p-toluenesulfonyl surface functionalized MPIO (1 µm diameter; Dynabeads™ MyOne™ Tosylactivated; Invitrogen) were used to conjugate with VCAM-1 antibody (BD Pharmingen™), P-selectin antibody (CD62P; Santa Cruz Biotechnology) and fluorescein cadaverine (Life Technologies).…”

“…All animal experiments were approved by the Institutional Animal Care and Use Committee for Biological Resource Centre at A*STAR, Singapore (IACUC #191,459). ApoE −/− mice (Taconic Biosciences) were used to develop the carotid atherosclerosis model by means of cuff implantation as previously described [7,8,10,11]. The shear-stress modifying cuff induced development of vulnerable, inflamed plaques in the low shear-stress region upstream of cuff placement [7,8,[10][11][12].…”

“…ApoE −/− mice (Taconic Biosciences) were used to develop the carotid atherosclerosis model by means of cuff implantation as previously described [7,8,10,11]. The shear-stress modifying cuff induced development of vulnerable, inflamed plaques in the low shear-stress region upstream of cuff placement [7,8,[10][11][12]. ApoE −/− mice commenced on a high fat diet (HFD) 2 weeks prior to cuff placement.…”

“…Recently, molecular imaging strategies have been developed to look beyond anatomy by assessing the pathophysiological activities within plaques at the molecular level [5]. Molecular MRI has been used to directly report the inflammatory activities within the carotid plaques in mouse models [6][7][8]. Inflammation is not only instrumental in the development of atheromatous plaques, but, more importantly, plays a critical role in plaque destabilisation, converting chronic stable lesions into acute unstable ones with ensuing thromboembolism [9].…”

“…Herein, we have used fluorescentlabelled dual-targeted microparticles of iron oxide (DT-MPIO) against P-selectin and VCAM-1, the key inflammatory biomarkers in plaques. The synthesis and synergistic binding efficacy of DT-MPIO over single-ligand-MPIO have been reported previously [7,8]. Using MR steady-state gradient echo sequence (GRE) developed specifically for molecular imaging of atherosclerosis, this strategy was utilised to (i) identify and characterise vulnerable plaques and (ii) quantitatively track inflammatory status of plaque progression, in the carotid arteries of an apolipoprotein E-deficient (ApoE −/− ) cuff-implanted mouse model [7,8,10].…”