Dual orthogonal fluorescent protease sensors for image guided surgery

项目摘要 虽然有越来越多的有用的化疗和放射性方法可用于癌症 在治疗中，主要的治疗策略仍然是手术切除肿瘤。的最大挑战 对于外科医生来说，1）精确和快速地定位原发性病变，2）清楚地定义边缘 肿瘤和正常组织之间的关系。手术后复发的失败通常是由于 不完全清除所有癌细胞。最近在使用光学传感器方面取得了许多进展， 用于荧光成像引导手术的造影剂（图1A和1B）。然而，FDA唯一批准的光学 试剂是主要用于灌注成像的非靶向染料， 肿瘤组织中的摄取。许多分子靶向光学造影剂最近已经发展到 临床试验此外，还有许多利用酶活性的光学探针，例如 在肿瘤微环境中被激活的蛋白酶。我们开发了光学“智能探头”， 只有当被发现在活化的巨噬细胞内的一类蛋白酶作用时， 肿瘤微环境虽然这些探针已经被证明是真实的有价值的代理 尽管使用FDA批准的达芬奇芬奇手术系统进行手术期间的肿瘤成像， 某些组织中高背景和取决于局部探针的有限的信号动态范围 浓度的为了解决这些局限性，我们概述了一个范式转移类光学设计 需要多个肿瘤特异性酶签名用于激活的探针，以及FRET成像 提高探针灵敏度和选择性的策略。我们将把这些探测器和摄像机连接起来， 图像处理系统，这将进一步使其最终的临床翻译。该提案是建立在 Bogyo实验室和 Intuitive Surgical Inc这些初步结果验证了使用蛋白酶探针成像的关键概念 使用现有的FDA批准的手术系统在手术过程中的肿瘤边缘，并作为坚实的基础 对于旨在增强现有光学成像探头在外科手术中的实用性的拟议研究， 工作流

Project Summary While there are an increasing number of useful chemotherapy and radiological methods available for cancer therapy, the predominant treatment strategy remains surgical resection of the tumor. The greatest challenges for the surgeon are 1) precisely and rapidly locating the primary lesion and 2) clearly defining the margins between tumor and normal tissues. Failure in the form of recurrence after surgery usually results from incomplete removal of all cancer cells. There have been a number of recent advances in the use of optical contrast agents for fluorescence imaging guided surgery (FIGS). However, the only FDA approved optical agents are non-targeted dyes that are mainly used for perfusion imaging and in some cases have increased uptake in tumor tissues. A number of molecularly targeted optical contrast agents have recently advanced to clinical trials. In addition, there are a number of optical probes that exploit the activity of enzymes such as proteases that are activated in the tumor microenvironment. We have developed optical ‘smart probes’ that produce a signal only when acted upon by a class of proteases found in activated macrophages within the tumor microenvironment. While these probes have already been proven to be valuable agents for real time tumor imaging during surgery using the FDA approved da Vinci surgical system, they still suffer from relatively high background in some tissues and limited dynamic range of signal that depends on local probe concentrations. To address these limitations, we outline the design of a paradigm-shifting class of optical probes that require multiple tumor-specific enzyme signatures for activation, as well as a FRET imaging strategy to enhance both sensitivity and selectivity of probes. We will couple these probes with camera and image processing systems that will further enable their eventual clinical translation. This proposal is built around a large body of preliminary data resulting from an active collaboration between the Bogyo Lab and Intuitive Surgical Inc. These initial results have validated the key concept of using protease probes to image tumor margins during surgery using existing FDA approved surgical systems and serve as a strong foundation for the proposed studies aimed at enhancing the utility of existing optical imaging probes in the surgical workflow.