Improved Molecular Imaging Contrast Agent for Detection of Human Thrombus Patrick M. Winter, 1,2 * Shelton D. Caruthers, 1,3 Xin Yu, 1,2 Sheng-Kwei Song, 4 Junjie Chen, 2 Brad Miller, 5 Jeff W.M. Bulte, 5,6 J. David Robertson, 7 Patrick J. Gaffney, 8 Samuel A. Wickline, 1 and Gregory M. Lanza 1 Molecular imaging of microthrombus within fissures of unsta- ble atherosclerotic plaques requires sensitive detection with a thrombus-specific agent. Effective molecular imaging has been previously demonstrated with fibrin-targeted Gd-DTPA-bis- oleate (BOA) nanoparticles. In this study, the relaxivity of an improved fibrin-targeted paramagnetic formulation, Gd-DTPA- phosphatidylethanolamine (PE), was compared with Gd-DTPA- BOA at 0.05-4.7 T. Ion- and particle-based r 1 relaxivities (1.5 T) for Gd-DTPA-PE (33.7 (s*mM) -1 and 2.48  10 6 (s*mM) -1 , respec- tively) were about twofold higher than for Gd-DTPA-BOA, per- haps due to faster water exchange with surface gadolinium. Gd-DTPA-PE nanoparticles bound to thrombus surfaces via anti-fibrin antibodies (1H10) induced 72%  5% higher change in R 1 values at 1.5 T (R 1  0.77  0.02 1/s) relative to Gd-DTPA- BOA (R 1  0.45  0.02 1/s). These studies demonstrate marked improvement in a fibrin-specific molecular imaging agent that might allow sensitive, early detection of vascular microthrombi, the antecedent to stroke and heart attack. Magn Reson Med 50:411– 416, 2003. © 2003 Wiley-Liss, Inc. Key words: contrast agent; nanoparticle; molecular imaging; vulnerable plaque; fibrin The acute formation of thrombus on ruptured atheroscle- rotic plaques is well recognized as the source of unstable angina, myocardial infarction, transient ischemic attacks, and stroke (1). Although myriad medical advances in the detection and treatment of advanced carotid and coronary artery disease have emerged, early detection of the most common source of thromboembolism—rupturing athero- sclerotic plaques in arteries with modest 40-60% stenosis (2)—remains diagnostically elusive with the use of routine angiography or duplex ultrasound techniques. A variety of approaches have been proposed to improve detection of vulnerable plaques, including intravascular ultrasound elastography (3), radionuclide imaging (4), and thermography (5). Magnetic resonance imaging (MRI) also is emerging as a particularly sensitive modality to nonin- vasively visualize thromboses within the carotid artery (6). However, the proximate cause of heart attacks and strokes—rupture of the vulnerable plaque— cannot be re- liably detected with any nondestructive imaging modality. Recent studies reveal that vulnerable plaque rupture and microthrombus formation precedes acute myocardial in- farction by days to months (7), providing a window of opportunity to intercede and prevent serious sequelae. Sensitive molecular imaging and detection of micro- thrombi along the intimal surface of vulnerable plaques will require a high-avidity, target-specific probe with ro- bust signal amplification compatible with a sensitive, high-resolution imaging modality. Until recently, the sig- nal amplification required to detect and visualize impor- tant molecular or cellular moieties present in nano- and picomolar concentrations in vivo was obtainable only with nuclear imaging modalities. However, more recently, mo- lecular imaging with magnetic resonance has shown prom- ise (8,9). A new approach entails the use of a fibrin-specific para- magnetic molecular imaging agent to improve detection and quantification of these occult microthrombi (9). The contrast agent is a ligand-directed, lipid-encapsulated liq- uid perfluorocarbon nanoparticle (250 nm nominal di- ameter) that has high target avidity and a prolonged sys- temic halflife, and can carry high gadolinium-DTPA pay- loads (50,000 Gd 3+ atoms per particle) for high detection sensitivity. The present report describes methods for enhancing the detection of thrombus with the use of a more favorable lipophilic anchor to attach the gadolinium chelate onto the nanoparticle surface. The influence of this improved sur- face chelate on T 1 -weighted contrast relaxivity is assessed both in solution and when targeted to thrombus via anti- fibrin antibodies. Variable field relaxometry at different temperatures is employed to characterize the relaxation differences among species of molecular imaging agents. This new generation of fibrin-specific nanoparticles re- veals the importance of the gadolinium chelate for optimi- zation of MR molecular imaging agents. 1 Cardiovascular MR Laboratories, Department of Medicine, Cardiovascular Division, Barnes-Jewish Hospital, Washington University School of Medicine, St. Louis, Missouri. 2 Department of Biomedical Engineering, Washington University, St. Louis, Missouri. 3 Philips Medical Systems, Best, The Netherlands. 4 Department of Chemistry, Washington University School of Medicine, St. Louis, Missouri. 5 Laboratory of Diagnostic Radiology Research, National Institutes of Health, Bethesda, Maryland. 6 Department of Radiology, Johns Hopkins University School of Medicine, Baltimore, Maryland. 7 Analytical Chemistry Group, University of Missouri Research Reactor, Co- lumbia, Missouri. 8 Department of Surgery, St. Thomas’s Hospital, London, UK. Presented in part at the 9th Annual Meeting of ISMRM, 2001, Glasgow, Scotland. Grant sponsor: NIH; Grant numbers: CO07121; HL59865; Grant sponsor: American Heart Association; Grant number: 0235125N; Grant sponsor: Washington University Small Animal Imaging Resource (WUSAIR), funded in part by the NCI Small Animal Imaging Research Program (SAIRP); Grant number: R24 CA83060; Grant sponsor: Philips Medical Systems. *Correspondence to: Dr. Patrick M. Winter, Department of Cardiology, Cam- pus Box 8086, Washington University Medical School, 660 S. Euclid Ave., St. Louis, MO 63110. E-mail: Patrick@cvu.wustl.edu Received 12 July 2002; revised 26 March 2003; accepted 30 March 2003. DOI 10.1002/mrm.10532 Published online in Wiley InterScience (www.interscience.wiley.com). Magnetic Resonance in Medicine 50:411– 416 (2003) © 2003 Wiley-Liss, Inc. 411