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With the goal of improving intraoperative cancer visualization, we have developed AVB-620, a novel intravenously administered, in vivo fluorescent peptide dye conjugate that highlights malignant tissue and is optimized for human use. Matrix metalloproteinases (MMPs) hydrolyze AVB-620 triggering tissue retention and a ratiometric fluorescence color change which is visualized using camera systems capable of imaging fluorescence and white light simultaneously. AVB-620 imaging visualizes primary tumors and demonstrated high in vivo diagnostic sensitivity and specificity (both &gt;95%) for identifying breast cancer metastases to lymph nodes in two immunocompetent syngeneic mouse models. It is well tolerated and single-dose toxicology studies in rats determined a no-observed-adverse-effect-level (NOAEL) at &gt;110-fold above the imaging and estimated human dose. Protease specificity and hydrolysis kinetics were characterized and compared using recombinant MMPs. To understand the human translation potential, an in vitro diagnostic study was conducted to evaluate the ability of AVB-620 to differentiate human breast cancer tumor from healthy adjacent tissue. Patient tumor tissue and healthy adjacent breast tissue were homogenized, incubated with AVB-620, and fluorogenic responses were compared. Tumor tissue had 2-3 fold faster hydrolysis than matched healthy breast tissue; generating an assay sensitivity of 96% and specificity of 88%. AVB-620 has excellent sensitivity and specificity for identifying breast cancer in mouse and human tissue. Significant changes were made in the design of AVB-620 relative to previous ratiometric protease-activated agents. AVB-620 has pharmaceutical properties, fluorescence ratio dynamic range, usable diagnostic time window, a scalable synthesis, and a safety profile that have enabled it to advance into clinical evaluation in breast cancer patients.

Sensitive in vivo Visualization of Breast Cancer Using Ratiometric Protease-activatable Fluorescent Imaging Agent, AVB-620