NGR imaging of angiogenesis in myocardial infarction /ischemia models

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DESCRIPTION (provided by applicant): Recent studies from our laboratory have demonstrated that angiogenic vasculature in the heart can be targeted and imaged using a synthetic peptide sequence consisting of asparagine-glycine-arginine (NGR). The vascular address for this homing sequence is a cell surface aminopeptidase, aminopeptidase N (APN) also known as CD13. In cardiac angiogenesis we have established, that CD13/APN is strongly up-regulated on endothelial cells in the neo-vasculature, and that it cannot be detected on quiescent vessels outside the angiogenic area. Furthermore, we have shown, using conjugates of NGR with different fluorescent molecules and with biotin, that cardiac neo-vessels can be targeted and imaged selectively with these conjugates. In other studies, we have shown that on endothelial cells CD13/APN is up-regulated in response to hypoxia, angiogenic growth factors (HIF-la, bFGF and VEGF). Based on these studies we propose to utilize this novel vascular address and the versatile NGR homing sequence to study the role played by the angiogenic vasculature that is identified by targeted imaging in the setting of myocardial infarction (MI), and in hind limb ischemia (HLI). In mouse MI and HLI models we will co-register NGR fluorescence intensity and tomography with 3D images of neo-vessels by micro-CT, 2 photon microscopy and serial histological sections together with indices of tissue perfusion, oxygenation and left ventricular (L.V.) function. Molecular characterization of neo-vessels that bind NGR, will be performed by using laser dissection microscopy to isolate endothelial cells that bind fluorescently labeled NGR and endothelial cells that do not bind NGR that will then be subjected to microarray expression analysis. Clinical applicability of NGR imaging will be tested with NGR-iron particles in a porcine model of myocardial ischemia and bio-distribution, pharmacokinetics and internalization will be studied in mice and by confocal and electron microscopy of live endothelial cells. (End of Abstract)