A cell-based liquid biopsy approach for early pancreatic cancer detection

 描述(申请人提供)：胰腺癌(PDA)是一种致命的恶性肿瘤，由于大多数患者在诊断时疾病的晚期性质，预后特别差。目前，对于PDA的EARL检测尚无有效的筛查策略。我们实验室的研究表明，在基因工程小鼠模型和胰腺癌前囊性病变患者中，胰腺上皮细胞在疾病的早期阶段被释放到血流中。这些观察为利用这些细胞作为筛选PDA的基础提供了一个独特的机会。该项目的主要目标是开发一种可靠的非侵入性检测方法，可以根据血液中是否存在胰腺来源的循环上皮细胞来检测早期胰腺导管腺癌(PDA)。我们将利用磁性纳米颗粒技术和微流控平台从全血中富集循环上皮细胞(CECs)，然后进行基于RNA的检测，包括RNA荧光原位杂交(RNA FISH)和定量PCR。我们将使用一种血统标记的PDA基因工程小鼠模型来验证这一点，在该模型中，可以很容易地在血液中检测到CEC。此外，我们将利用优化的设备以及其他单细胞分离技术，如DEPArray(Tm)技术或流式细胞仪对CEC进行下一代测序，以便(I)识别新的生物标记物，以便更有效地进行筛查和诊断，以及(Ii)研究胰腺癌进展不同阶段CEC之间的功能差异。最后，我们将在一组已知的PDA患者中实施一项先导性研究，以使用先前AIMS中开发的方法来丰富和检测血液中的CEC。我们预计，这一策略还可以用于其他一些应用，包括评估治疗反应、疾病分期和高危人群的筛查，从而提高胰腺癌的存活率。

 DESCRIPTION (provided by applicant): Pancreatic adenocarcinoma (PDA) is a lethal malignancy with a particularly poor prognosis due to the advanced nature of the disease at the time of diagnosis in most patients. Currently, there are no effective screening strategies for earl detection of PDA. Studies from our lab have demonstrated that pancreatic epithelial cells are shed into the blood stream at early stages of disease in genetically engineered mouse models as well as in patients with pre-malignant cystic lesions of the pancreas. These observations present a unique opportunity to use these cells as the basis for screening for PDA. The primary goal of this project is to develop a robust non-invasive test that can be used to detect early stage pancreatic ductal adenocarcinoma (PDA) based on the presence of circulating epithelial cells of pancreatic origin in blood. We will use magnetic nanoparticle technology and a microfluidic platform to enrich circulating epithelial cells (CECs) from whole blood followed by RNA-based detection, including RNA fluorescent in situ hybridization (RNA FISH) and quantitative PCR. We will validate this using a lineage-labeled genetically engineered mouse model of PDA in which CECs can be easily detected in the blood. Further, we will utilize the optimized device along with additional single cell isolation techniques such as the DEPArray(tm) technology or flow cytometry to perform next-generation sequencing on CECs in order to (i) identify novel biomarkers for more effective screening and diagnosis and (ii) investigate functional differences between CECs at different stages of pancreatic cancer progression. Finally we will implement a pilot study in a cohort of patients with known PDA to enrich and detect CECs in the blood using the assay developed in previous aims. We envision that this strategy could be adapted for a number of other applications including evaluating response to therapy, disease-staging and screening of high-risk populations, thereby leading to improvements in pancreatic cancer survival.