Protein-Chip Diagnostic Assay Detection for Cancer

描述（由申请人提供）：肿瘤产生循环因子，这些因子有可能用作肿瘤存在的诊断标志物。这些标记物可以是肿瘤类型特异性的、血管生成因子或参与对肿瘤的免疫应答的细胞因子。仅基于一个标记的诊断产生太多的假阳性。所需要的是通过蛋白质组学分析许多标志物（多达100种标志物）的能力，然后在标志物的存在和肿瘤的存在之间建立准确的相关性，从而消除假阳性。我们建议使用超灵敏的多光子检测（MPD）增强免疫测定（IA/MPD）和蛋白质芯片（P-chips/MPD），检测浓度范围从0.01 pg/ml到500 pg/ml的肿瘤标志物。该技术将产生一种更敏感和更特异的诊断方法，这可能使肿瘤的早期检测成为可能。本项目的第一阶段的目标是发展超灵敏的IA/MPD检测技术，用于检测在多种肿瘤类型中表达的血管生成因子成纤维细胞生长因子-1（FGF-1）。然后筛选临床相关的血清样本中的FGF-1，沿着FGF-2、血管内皮生长因子（VEGF）、肝细胞生长因子（HGF）、IL-1 β、IL-6、IL-10、IL-11、IL-12和TNF-α以及肿瘤标志物（如前列腺血清抗原）。目前存在对这些因子中的许多因子的超灵敏IA/MPD测定。临床相关的血清样品将包括癌症治疗前和治疗后的患者以及健康对照受试者。治疗后的癌症患者将随着时间的推移进行监测，以查看标记物水平随时间的变化是否可以预测肿瘤复发。将检查结果以确定标志物水平的特定模式是否与肿瘤的存在相关。第二阶段的目标是开发MPD增强型P芯片。P-芯片/MPD将适用于与肿瘤存在相关的大量标志物，包括PSA、CEA、CA-125和甲胎蛋白、细胞因子和血管生成因子。还将研究其他因子，这些因子被发现由肿瘤表达，但不能被当前相对低灵敏度的诊断检测到。将扩大对临床相关样本的测试。一旦同时测量这些生物标志物的重要性得到确认，我们将开发专用的超灵敏P芯片，用于定量多达100种靶向标记蛋白，检测范围为0.01至500 pg/ml。因此，低成本的癌症诊断和治疗监测测定将变得可用，并且可以提出早期肿瘤形成的潜在机制，因为一些标志物在早期阶段可能仅以较低水平表达。

DESCRIPTION (provided by applicant): Tumors produce circulating factors that have the potential to be used as diagnostic markers for the presence of the tumor. These markers may be tumor type specific, angiogenic factors, or cytokines involved in the immune response to the tumor. The diagnostic based on just one marker yield too many false positives. What is needed is the ability to assay many markers (up to 100 markers) by proteomics and then develop an accurate correlation between the presence of the markers and the presence of the tumor, thus eliminating false positives. We propose use of the supersensitive MultiPhoton Detection (MPD) enhanced to immunoassays (IA/MPD) and protein-chips (P-chips/MPD), to detect tumor markers in concentration ranges from 0.01 pg/ml level to 500 pg/ml. The technology will result in a more sensitive and specific diagnostic method, which may enable the detection of the tumors in an earlier stage of development. The objective of Phase I of this proposal is to develop the supersensitive IA/MPD assay techniques for the angiogenesis factors, Fibroblast Growth Factor-1 (FGF-1) that have been found to be expressed in many tumor types. Then clinically relevant serum samples will be screened for FGF-1, along with FGF-2, Vascular Endothelial Growth Factor (VEGF), Hepatocyte Growth Factor (HGF), IL-lbeta, IL-6, IL-10, IL-11, IL-12, and TNF-alpha, and tumor markers like Prostate Serum Antigen. Ultrasensitive IA/MPD assays currently exist for many of these factors. The clinically relevant serum samples will include pre- and post- cancer treatment patients, and healthy control subjects. Post treatment cancer patients will be monitored over time to see if marker level variations over time can predict tumor recurrence. Results will be examined to determine if a particular pattern of marker levels correlates with the presence of a tumor. The goal of Phase II is the development of the MPD enhanced P-chip. The P-chips/MPD will be adapted to a large set of markers including PSA, CEA, CA-125, and alpha-fetoprotein, cytokines and angiogenesis factors that correlate with the presence of a tumor. Additional factors, which are found expressed by tumors, but can not be detected by current relatively low sensitivity diagnostics will also be studied. Testing on clinically relevant samples will be expanded. Once the importance of concurrent measurement of several of these biomarkers are confirmed, we will develop dedicated supersensitive P-chips for quantifying up to 100 targeted marker proteins with detection ranges from 0.01 to 500 pg/ml. Thus, low cost cancer diagnostics and therapy monitoring assay will become available and a potential mechanism(s) may be suggested for earlier stage tumor formation since some of the markers may only be expressed at lower levels at the early stages.