Non-invasive Differentiation of Benign Lesions from Aggressive Pancreatic Cancer

描述（由申请人提供）：目前，我们无法区分胰腺良性病变和侵袭性肿瘤，除非进行手术切除和随后的组织学评价。这项提案的长期目标是开发一种正电子发射断层扫描（PET）显像剂用于临床，以改善胰腺肿瘤的危险分层。该R21项目的目标是通过证明使用我们的蛋白酶活性探针与光学和放射性核素成像技术在动物模型中成功成像浸润前和浸润性肿瘤来证明可行性。最近的研究表明组织蛋白酶样蛋白酶活性与胰腺上皮内肿瘤（PanIN）分级相关。这些数据提高了这种活性也可用于区分良性病变和侵袭性癌症的可能性。我们开发了一种新技术，可以灵敏地、非侵入性地检测蛋白酶活性。初步研究表明，非侵入性检测，区分和分级胰腺疾病，包括急性和慢性胰腺炎，导管内乳头状粘液性肿瘤（IPMN），PanIN和胰腺导管腺癌（PDAC）的承诺。这些数据支持追求以下目的以测试我们的假设，即我们可以使用蛋白水解来激活膜结合成像剂，用于胰腺癌的准确和定量的非侵入性成像，以提供关于肿瘤状态的功能和动态信息。通过使用光学成像来优化探针和放射性核素成像，我们打算展示最终开发用于临床使用的PET成像剂的原理证明。目标1.胰腺癌检测中先导药物的优化与筛选。将对探针进行优化，以提高选择性和信噪比，并验证它们在各种癌细胞系中的作用。将通过小鼠和人胰腺癌细胞系组中的荧光成像测定对探针进行排序。目标2.在胰腺癌转基因模型中使用胰腺癌进展的非侵入性成像优化探针。将在小鼠中评价来自目标1的最佳探针。我们将使用胰腺癌的基因工程小鼠模型，其中胰腺定向表达的组成型活性Kras驱动PanIN形成。用光学成像和切片组织中检测到的探针积聚将与组织学分级相关，并通过免疫荧光定向至细胞特异性标志物。激光捕获显微切割将用于收获高蛋白水解活性区域并确定MS中存在哪些蛋白酶。这些数据将直接将特定蛋白酶活性与体内胰腺癌及其前体病变联系起来。根据光学成像的结果，将用放射性核素标记少量选定的探针，并进行PET成像。将进行药效学和生物分布研究，以证明可用于临床。这些目标将确定是否可以使用细胞周围蛋白水解来识别胰腺疾病，以及是否可以非侵入性地区分不同的病理。

DESCRIPTION (provided by applicant): At present, we cannot differentiate benign lesions of the pancreas from aggressive tumors without surgical resection and subsequent histological evaluation. The long-term goal of this proposal is to develop a positron emission tomography (PET) imaging agent for clinical use to improve risk stratification among pancreatic neoplasms. The goal of this R21 project is to establish proof of feasibility by demonstrating successful imaging of pre-invasive and invasive tumors in an animal model using our protease activity probes with optical and radionuclide imaging techniques. Recent work has shown cathepsin-like protease activity correlates with pancreatic intraepithelial neoplasm (PanIN) grade. These data raise the possibility that such activity could also be used to differentiate benign lesions from aggressive cancer. We have developed new technology that sensitively and non-invasively detects protease activities. Preliminary studies have shown promise for non-invasively detecting, differentiating, and grading pancreatic diseases including acute and chronic pancreatitis, intraductal papillary mucinous neoplasm (IPMN), PanIN, and pancreatic ductal adenocarcinoma (PDAC). These data support pursuing the following aims to test our hypothesis that we can use proteolysis to activate a membrane binding imaging agent for accurate and quantitative non-invasive imaging of pancreatic cancer to provide functional and dynamic information about tumor status. By using optical imaging to optimize the probe and radionuclide imaging we intend to show proof of principle for ultimately developing a PET imaging agent for clinical use. Aim 1. Optimize and Validate Lead Agents for the Detection of Pancreatic Cancer. The probes will be optimized to increase selectivity and signal to noise and to verify that they work in a variety of cancer cell lines. Probes will be ranked by fluorescence imaging assays in panels of mouse and human pancreatic cancer cell lines. Aim 2. Optimize Probes using Non-Invasive Imaging of Pancreatic Cancer Progression in a Transgenic Model of Pancreatic Cancer. The best probes from Aim 1 will be evaluated in mice. We will use a genetically-engineered mouse model of pancreatic cancer where pancreas-directed expression of constitutively active Kras drives PanIN formation. Probe accumulation, as detected with optical imaging and in sectioned tissue will be correlated to histological grade and oriented by immunofluorescence to cell specific markers. Laser capture microdissection will be used to harvest regions of high proteolytic activity and determine which protease(s) are present with MS. These data will directly link specific protease activities to pancreatic cancer and their precursor lesions in vivo. Based on results from optical imaging, a small number of selected probes will be labeled with radionuclides and PET imaging will be conducted. Pharmacodynamics and biodistribution studies will be carried out to demonstrate translatability to the clinic. These aims will determine whether pericellular proteolysis can be used to identify pancreatic disease and whether different pathologies can be distinguished non-invasively.