A Sensitive and Automated Diagnostic Assay for Lung Cancer

描述(申请人提供)：一种灵敏的、自动化的肺癌诊断方法肺癌是美国头号癌症杀手。能够在早期、可治愈的阶段识别肺癌将降低死亡率，因此是一个重要的临床目标。为了实现这一目标，我们已经确定了一组可以在痰中检测到的遗传生物标记物，用于肺癌的早期检测(AACR突发新闻)。我们还开发了一种原位微型芯片，包括多个荧光素标记的探针，可以在一次测试中测量遗传生物标记物。然而，探针的荧光信号很弱，褪色速度很快，降低了生物标志物的灵敏度。此外，人工计数多个探针的模糊信号费时费力且变异性高，从而限制了临床应用的前景。最近的研究表明，荧光团与金属的相互作用，即金属增强荧光(MEF)，可以增加荧光信号的强度和光稳定性，并且新兴的自动斑点计数系统可以快速、客观地分析多个探针。R21探索性项目的目标是开发一种原位微型芯片，作为一种可靠的诊断方法，能够可靠地检测痰标本中早期检测肺癌的遗传生物标志物。我们建议1)通过将荧光探针与固定在涂有银纳米颗粒的玻片上的细胞杂交来提高原位微型芯片的灵敏度，2)通过使用自动斑点计数系统分析痰细胞中增强的荧光信号来开发原位微型芯片作为稳健检测，以及3)在我们现有的I期肺癌病例对照队列中利用痰样本确定稳健检测的诊断性能。完成后，原位微型芯片将被改进为一种可靠的、自动化的、非侵入性的肺癌早期检测基因测试。这项研究将为大型前瞻性临床项目(R01)或多中心合作临床研究(R33)奠定基础，旨在进一步评估其可在常规临床实验室迅速广泛接受和应用的全部效用。这项探索性研究将汇集在分子遗传学、纳米技术、细胞病理学、肿瘤学和生物统计学方面具有广泛专业知识的科学家和临床医生的优势互补。

DESCRIPTION (provided by applicant): A Sensitive and Automated Diagnostic Assay for Lung Cancer Lung cancer is the number one cancer killer in the USA. The ability to identify lung cancer at its early, curable stages will reduce the mortality and is thus an important clinical goal. Toward this goal, we have identified a panel of genetic biomarkers that can be detected in sputum for early detection of lung cancer (AACR Breaking News). We have also developed an in situ mini-chip comprising multiple fluorescein-labeled probes that can measure the genetic biomarkers in a single test. However, the fluorescent signals of the probes are weak and fade fast, reducing the sensitivity of the biomarkers. Furthermore, manually counting the blurry signals of the multiple probes is time- consuming and labor intensive with high variability, thus limiting the promising diagnostic approach in clinical setting. The recent studies have shown that fluorophore-metal interaction, namely, metal-enhanced fluorescence (MEF), can increase intensity and photostability of fluorescent signals, and an emerging automated dot counting system permits rapidly and objectively analyze multiple probes. The objective of the R21 exploratory project is to develop the in situ mini-chip as a robust diagnostic assay that can reliably detect the genetic biomarkers for the early detection of lung cancer in sputum samples. We propose to 1) increase the sensitivity of the in situ mini-chip by hybridizing the fluorescent probes to the cells immobilized on the slides coated with silver nanoparticles, 2) develop the in situ mini-chip as robust assay by analyzing the enhanced fluorescent signals in sputum cells using the automated dot-counting system, and 3) determine diagnostic performance of the robust assay for early detection of lung cancer using sputum samples in our existing case-control cohort of patients with stage I lung cancer. At completion, the in situ mini-chip will be refined as a reliable, automated, and noninvasive genetic test for lung cancer early detection. The study will lay the basis for large prospective clinical project (R01) or multicenter cooperative clinical study (R33) designed to further evaluate its full utility that could rapidly and widely be accepted and applied in routine clinical laboratories. This exploratory study will bring together complementary strengths of scientists and clinician with extensive expertise in molecular genetics, nanotechnology, cytopathology, oncology, and biostatistics.