A Lectin Glycoarray Approach for Markers of Pancreatic Cancer

描述（由申请人提供）：胰腺癌是最致命的人类恶性肿瘤之一，早期发现将大大提高患者的生存率。糖微阵列方法将用于在人血浆中寻找胰腺癌的早期检测标记物。我们将利用凝集素亲和柱对完整的n -连接血浆糖蛋白进行多维液相分离。完整糖蛋白的液体分离将首先涉及无孔色谱，然后是液体毛细管等电聚焦以进一步分离蛋白质糖型。每个液体部分将被标记在硝化纤维素包被的显微镜载玻片上，以产生天然糖蛋白微阵列，然后将被各种荧光标记的凝集素询问，以探测每个微阵列点是否存在特定的聚糖部分。将分析来自癌症患者、正常患者和炎性病变患者的血浆样本，以寻找揭示癌症进展过程中发生的特定聚糖结构变化的模式。将对来自癌症和正常患者以及相关炎症病变（慢性胰腺炎）患者的30个样本的分析测试集进行初步分析，以确定这些潜在的标志物。新的软件将用于分析这些阵列和结果测试集。利用凝集素提取/LC ESI/MS/MS和QIT-TOF （MALDI MSn）质谱法鉴定与癌症或炎症相关的糖蛋白，并鉴定其结构部分，以检查可能作为癌症标志物的结构的详细变化。虽然许多标记糖蛋白本身在血浆中含量相对较高，但它们可能是基于糖结构变化的癌症特异性。这项研究在公共卫生方面具有重要意义，因为它可能为寻找血浆中的疾病标志物提供了一种方法。这些标志物可能与疾病的分期有关，因此可能对个性化治疗很重要，这是临床医学的一个重要问题。

DESCRIPTION (provided by applicant): Pancreatic cancer is one of the most deadly forms of human malignancy for which early detection would greatly facilitate increased patient survival. A glyco-microarray approach will be used to search for early detection markers of pancreatic cancer in human plasma. We will use multi-dimensional liquid-phase fractionation of intact N-linked plasma glycoproteins isolated by lectin affinity columns. The liquid-based fractionation of intact glycoproteins will initially involve nonporous chromatography, followed by liquid capillary isoelectric focusing to further separate protein glycoforms. Each liquid fraction will be spotted on nitrocellulose- coated microscope slides to produce a natural glycoprotein microarray which will then be interrogated by various fluorescently-labeled lectins to probe each microarray spot for the presence of specific glycan moieties. Plasma samples from cancer and normal patients and patients with inflammatory lesions will be analyzed to search for patterns that reveal specific glycan structural changes that occur during cancer progression. Initial analysis will be performed on an analytical test set of 30 samples each from cancer and normal patients and from patients with related inflammatory lesions (chronic pancreatitis) to identify such potential markers. Novel software will be used to analyze these arrays and the resulting test set. Glycoproteins that are associated with cancer versus normal or inflammation will be identified and the structural moieties identified using lectin extraction/LC ESI/MS/MS and QIT-TOF (MALDI MSn) mass spectrometry to examine the detailed changes in structure that may serve as markers of cancer. While many of the marker glycoproteins themselves are of relatively high abundance in plasma, they may be cancer-specific based upon changes in glycan structure. This research has important implications in public health in that it may provide a methodology for searching for markers of disease in plasma. These markers may be related to the stage of the disease and, thus, may become important for personalized treatment, an important issue in clinical medicine.