Molecularly targeted optical contrast agents have the potential to enable surgeons to visualize specific molecular markers that can help improve surgical precision and thus outcomes. Fluorescently quenched substrates can be used to highlight tumor lesions by targeting proteases that are highly abundant in the tumor microenvironment. However, the majority of these and other molecularly targeted optical contrast agents are labeled with reporter dyes that are not ideally matched to the properties of clinical camera systems, which are typically optimized for detection of indocyanine-green (ICG). While a wide range of near-infrared (NIR) dyes are suitable for use with highly sensitive and highly tunable research-focused small animal imaging systems, most have not been evaluated for use with commonly used clinical imaging systems. Here we report the optimization of a small molecule fluorescently quenched protease substrate probe 6QC-ICG, which uses the indocyanine green (ICG) dye as its optical reporter. We evaluated dosing and kinetic parameters of this molecule in tumor-bearing mice and observed optimal tumor over background signals in as little as 90 min with a dose of 2.3 mg/kg. Importantly, the fluorescence intensity of the probe signal in tumors did not linearly scale with dose, suggesting the importance of detailed dosing studies. Furthermore, when imaged using the FDA approved da Vinci Si surgical system with Firefly detection, signals were significantly higher for the ICG probe compared to a corresponding probe containing a dye with similar quantum yield but with a slightly shifted excitation and emission profile. The increased signal intensity generated by the optimal dye and dose of the ICG labeled probe enabled detection of small, flat lesions that were less than 5 mm in diameter. Therefore, 6QC-ICG is a highly sensitive probe that performs optimally with clinical imaging systems and has great potential for applications in optical surgical navigation.

中文翻译：

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用于光学手术导航的蛋白酶激活探针的优化

分子靶向的光学对比剂具有使外科医生可视化特定分子标记物的潜力，这些分子标记物可帮助提高手术精度并从而改善结果。通过靶向在肿瘤微环境中高度丰富的蛋白酶，荧光淬灭的底物可用于突出肿瘤损伤。但是，大多数这些和其他分子靶向的光学对比剂都用报告染料标记，这些染料与临床相机系统的特性并不理想匹配，通常针对检测吲哚菁绿（ICG）进行了优化。尽管各种各样的近红外（NIR）染料都适合与高度敏感和高度可调的以研究为重点的小型动物成像系统一起使用，但大多数尚未经过评估，可以与常用的临床成像系统一起使用。在这里，我们报告优化的小分子荧光淬灭蛋白酶底物探针6QC-ICG，它使用吲哚菁绿（ICG）染料作为其光学报告分子。我们评估了荷瘤小鼠中该分子的剂量和动力学参数，并在短短90分钟内以2.3 mg / kg的剂量观察到最佳的肿瘤信号超过背景信号。重要的是，肿瘤中探针信号的荧光强度并未随剂量线性变化，这表明进行详细剂量研究的重要性。此外，当使用FDA批准进行成像时 我们评估了荷瘤小鼠中该分子的剂量和动力学参数，并在短短90分钟内以2.3 mg / kg的剂量观察到最佳的肿瘤信号超过背景信号。重要的是，肿瘤中探针信号的荧光强度并未随剂量线性变化，这表明进行详细剂量研究的重要性。此外，当使用FDA批准进行成像时 我们评估了荷瘤小鼠中该分子的剂量和动力学参数，并在短短90分钟内以2.3 mg / kg的剂量观察到最佳的肿瘤信号超过背景信号。重要的是，肿瘤中探针信号的荧光强度并未随剂量线性变化，这表明进行详细剂量研究的重要性。此外，当使用FDA批准进行成像时在具有萤火虫检测功能的da Vinci Si外科手术系统中，ICG探针的信号明显高于相应的探针，该探针所含的染料具有相似的量子产率，但激发和发射曲线略有偏移。由最佳染料和ICG标记探针的最佳剂量产生的信号强度增加，使得能够检测直径小于5 mm的小而扁平的病变。因此，6QC-ICG是一种高度灵敏的探针，在临床成像系统中表现最佳，并且在光学手术导航中具有巨大的应用潜力。