MR and Fluorescent Imaging of Low-Density Lipoprotein Receptors 1 Hui Li, Brian D. Gray, Ian Corbin, Corinna Lebherz, Hoon Choi, Sissel Lund-Katz, James M. Wilson, Jerry D. Glickson, Rong Zhou Rationale and Objectives. Over-expression of low-density lipoprotein receptors (LDLRs) occurs in many types of malignan- cies and is related to the requirement for lipids for rapid proliferation of the tumors. On the other hand, LDLRs that are unable to bind LDL are found on hepatocytes of patients with familial hypercholesterolemia (FH), a genetic disease that leads to pre- mature atherosclerosis and death. The highly selective binding of LDL to LDLR makes these particles ideal carriers of therapeu- tic and diagnostic contrast agents into the targeted cells. The objectives of this paper are to examine whether a prototype con- trast agent (PTIR267) with dual detection properties is suitable for labeling of LDL particles for in vivo detection of LDLR by magnetic resonance imaging (MRI) and for in vitro monitoring of cellular localization by confocal fluorescence microscopy. Materials and Methods. PTIR267 is a lipophilic GdDTPA derivative conjugated to a fluorescent dye. The conjugated dye molecule makes the probe sufficiently water soluble to allow labeling of LDL by a brief incubation of LDL with PTIR267 dis- solved in PBS at 37°C (mole ratio LDL: PTIR267 = 0.09:1). The molar relaxivity of PTIR267 in saline is 26 mM -1 s -1 . Spe- cific LDLR-mediated uptake of PTIR267-labeled LDL was demonstrated in vitro by confocal fluorescence imaging of B16 mel- anoma cells using confocal fluorescence imaging. In vivo uptake of PTIR267-labeled LDL by a subcutaneously implanted B16 melanoma in mice leads to 30% decrease in longitudinal relaxation time (T 1 ) in the tumor. In vivo uptake of PTIR267-labeled LDL leads to 70% decrease in T 1 in a normal C57BL/6 mouse liver; however, in the liver of LDL receptor gene knockout (LDLr-/-) mice with C57BL/6 background, only 12% decrease in T 1 is observed. Conclusions. The dual fluorescence and MR imaging properties of PTIR267, combined with the ease of LDL labeling, suggest that it will be a useful tool for optimization of LDLR-targeted cancer diagnosis or therapy and for monitoring the efficacy of gene therapy of FH. Key Words. Low-density lipoprotein; low-density lipoprotein receptor; MR imaging; gadolinium; familial hypercholester- olemia; cancer; B16 melanoma; PTIR267; confocal fluorescent microscopy. © AUR, 2004 Low-density lipoprotein (LDL), the major carrier of cho- lesterol in the blood, binds exclusively to the LDL recep- tor (LDLR) and consists of a polar phospholipid mono- layer surrounding a neutral lipid core of cholesterol esters and triglycerides. A large apolipoprotein, apoB-100 (534 kD; 4,536 amino acids), adsorbed on the surface of the phospholipid monolayer, solubilizes the lipid core and contains a receptor binding site. The binding site contains a sequence of cationic arginine and lysine residues that bind stereospecifically complementary to anionic sites on the surface of the receptor. After binding to the receptor, LDL is internalized and incorporated into endosomes, in which the lipoproteins are degraded into constituent Acad Radiol 2004; 11:1251–1259 1 From the Departments of Radiology (H.L., I.C., J.D.G., R.Z.), Institute of Human Gene Therapy (C.L.), Material Science (H.C.) University of Pennsyl- vania, Philadelphia, PA, PTI Research Inc., Exton, PA (B.D.G), Children’s Hospital of Philadelphia, Philadelphia, PA 19104 (S.L.K.) Received May 20, 2004; revision requested July 22; revision received August 9; revision ac- cepted August 12. This research was supported by American Heart Associ- ation (RZ) and the National Cancer Institute grants R24CA83105 (JDG), P20-CA86255-01 (JDG), an SBIR grant from the National Heart, Lung and Blood Institute, R43 HL62042-01(BDG), and a contract from the National Cancer Institute, BAA N01-CM-97065-32 (Britton Chance, JDG). Address correspondence to: RZ. e-mail: zhou@rad.upenn.edu © AUR, 2004 doi:10.1016/j.acra.2004.08.007 1251