Quantum Dots for NIR Fluorescence Imaging of Tumor Angiogenesis

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DESCRIPTION (provided by applicant): Our long-term objective is to develop appropriate probes for multimodality molecular imaging of tumor angiogenesis and metastasis. Intermediate objective of this application is to develop cyclic RGD peptide conjugated quantum dots (QDs) for near-infrared (NIR) fluorescence imaging of av¿3 integrin expression in vivo. Cell adhesion molecule av¿3 integrin plays a key role in angiogenesis related diseases including cancer, atherosclerosis, rheumatoid arthritis, restenosis and diabetic retinopathy. Inhibition of av¿3 integrin activity by mAbs, cyclic RGD peptide antagonists, and peptidomimetics has been shown to induce endothelial apoptosis, to inhibit angiogenesis, and to increase endothelial monolayer permeability. Suitably labeled RGD peptides and antibodies have also been developed by us and others for MRI, ultrasound, NIR fluorescence, and radionuclide (PET and SPECT) imaging of integrin expression in vivo. Recently we demonstrated for the first time that RGD peptide conjugated QDs are able to target the extracellular segment of integrin av¿3 in an xenograft model. However, the enthusiasm for further studies of these nanoconjugates is mitigated by the unfavorable in vivo kinetics and cadmium-based cytotoxicity of the traditional quantum dots. We propose here to develop the next generation of biocompatible QDs for NIR fluorescence imaging. These particles are ultra small with thin coating material and are not made of cadmium chalcogenide materials. This type of QDs are suitable for animal imaging studies and eventually human use. First, we will develop a small library of non-Cd based QDs and conjugate the new QDs with RGD peptide. The performance of the newly developed non-Cd QDs will be compared with traditional Cd-based QDs. Efforts will also be spent to reduce non-specific binding. Second, we will evaluate the integrin targeting efficacy of QD-RGD in vivo. We will also directly compare biologically modified QDs with traditional molecular imaging probes by labeling RGD peptide with fluorescent dyes. In order to fully characterize the biodistribution and pharmacokinetics of the QD-RGD conjugates, we will radiolabel the newly developed QD-RGD conjugates with Cu (t1/2 = 12.7 h) and 64 124 I (t1/2 = 4.2 d) for combined NIR fluorescence and PET imaging. Finally, both Cd and non-Cd based QD-RGD conjugates will be subjected to acute and chronic toxicity studies. We expect that the newly developed non-Cd QDs with little or no toxicity will be amenable for clinical translation. The success of this study with RGD peptide modification for integrin av¿3 can be extended to QD multiplexing to provide the real-time information about cell surface markers (indicating malignancy, tumor type, and potentially influencing therapeutic procedure). Such information will be crucial for fluorescence-guided surgery by sensitive, specific, and real-time intraoperative visualization of molecular features of normal and diseased processes. Fluorescent semiconductor nanocrystals (a.k.a. quantum dots) have evolved over the last two decades from pure electronic materials science to biological applications. Various coating techniques have been applied to make QDs biocompatible (water-soluble and biologically stable). Suitably conjugated QDs (through antibody, proteins, peptides or oligonucleotides) are most commonly used in cell biology applications, including DNA array technology, immunofluorescence assays. However, preparation of QDs for molecular imaging has, so far, been severely under-developed. In addition, cadmium based cytotoxicity further hampered the development of such nanoconstructs for in vivo applications. In this application we thrive to develop non- cadmium based ultra small QDs for tumor angiogenesis imaging. We will develop and characterize a library of QD-RGD conjugates, followed by in vitro and in vivo screening. The QDs with suitable imaging quality will be subjected to acute and chronic toxicity studies. The success of this approach will allow clinical translation of QD based probe for fluorescence imaging.