Targeted Imaging Mass Spectroscopy for Biomarker Detection in Human Tissues

描述（由申请人提供）：生物标志物在患者组织活检中测量，以改进诊断，并指导治疗。对病人来说，这意味着接受最有可能起作用的治疗或药物，同时避免替代疗法的费用和毒副作用。在病理学中，一个长期存在的问题是，组织中最准确的生物标志物检测方法需要提取和破坏组织。由于显微诊断仍然是诊断的金标准，因此需要在完整组织中测量生物标志物的方法。现有的方法依赖于人眼检查和对指示生物标志物的色素量的定性评分。生物标记通常是一次执行一个。这导致评分的可变性，并限制了在小组织活检中可以进行的测试量。我们的项目是用自动靶向成像质谱法取代人眼检测，它可以同时测量每个组织活检上的许多生物标志物。我们使用“条形码”可切割标签的探针（抗体），以识别完整组织切片中的生物标志物目标。条形码标签就像人们在杂货店里找到的商品上的条形码。我们的机器检测器相当于杂货店收银台的扫描仪。商店的条形码几乎是无限可变的，但就像指纹一样，每个条形码唯一地标识产品。通过现有的化学方法，我们可以创建许多不同的质量标签条形码，以便在每个组织切片上进行许多不同的测试。激光扫描组织，打破标签从他们的探针，使他们飞离组织到一个自动探测器。激光的位置是精确知道的，所以每次条形码标签飞离组织时，它与病理学家使用的标准图像有关。长期目标是以更低的成本和更准确地测量组织生物标志物。第一阶段假设：使用HER-2作为模型，我们假设TIMS产生的组织生物标志物结果与乳腺癌中HER-2的IHC检测结果相当。具体目标1：设计和实现新一代的光可切割（三酰基）分子偶联抗体。具体目标2：开发多路TIMS协议。第二阶段计划：检查每个抗体上有多少个标签；确定结合和未结合三基标签的稳定性和保质期；确定蛋白质，DNA和RNA是否可以在多重分析中同时靶向；检查激光是否可以缩小范围，将其分辨率提高到单个细胞；确定激光诱导的质量标签释放是激光强度的线性函数还是非线性函数商业应用：我们将寻求药物开发商作为客户。如果用于早期发现，TIMS很可能也将用于后续的临床试验，以识别和招募肿瘤中含有适合药物靶点的患者。这将推动该方法进入更广泛的诊断市场。

DESCRIPTION (provided by applicant): Biomarkers are measured in patient tissue biopsies to refine diagnosis, and to guide therapy. For a patient, this means receiving the therapy or drug most likely to work while avoiding the expense and toxic side effects of alternatives. In pathology, a long-standing problem is that the most accurate methods for biomarker detection in tissues require extraction and disruption of the tissue. Because microscopic diagnosis remains the gold standard for diagnosis, methods to measure biomarkers in intact tissue are needed. Existing methods depend upon human eye inspection and qualitative scoring of the amount of a pigment indicating a biomarker. Biomarkers are typically performed one at a time. This causes variability in scoring, and limits the amount of testing that can be done on small tissue biopsies. Our project is to replace human eye detection with automated targeted imaging mass spectroscopy which allows simultaneous measurement of many biomarkers on each tissue biopsy. We use "bar coded" cleavable tags for probes (antibodies) that identify biomarker targets in intact tissue sections. The bar coded tags are like the bar code one finds on items at the grocery store. Our machine detector is the counterpart of the scanner at the grocery store checkout line. The store bar codes are almost infinitely variable but like a fingerprint, each uniquely identifies the product. Through existing chemical methods we can create many different mass tag bar codes so that many different tests can be performed on each tissue section. A laser scans the tissue, breaking the tags from their probes so that they fly off the tissue to an automatic detector. The location of the laser is precisely known so that each time a bar coded tag flies off the tissue, it is related to the standard picture used by the pathologist. The long term goal is to measure tissue biomarkers at lower cost and more accurately. PHASE I HYPOTHESIS: Using HER-2 as a model, we hypothesize that TIMS produces tissue biomarker results that are comparable to IHC detection of HER-2 in breast cancer. Specific Aim 1: Design and implement a new generation of photo cleavable (trityl) molecules conjugated to antibodies. Specific Aim 2: Development of a multiplexed TIMS protocol. PLANS FOR PHASE II: Examine how many tags there are on each antibody; Determine stability and shelf life of bound and unbound trityl tags; Determine whether proteins, DNA, and RNA can be simultaneously targeted in a multiplexed assay; Examine whether the laser can be narrowed, refining its resolution to individual cells; Determine whether laser induced release of mass tags is a linear or non-linear function of laser intensity COMMERCIAL APPLICATION: We will seek drug developers as customers. If used in early phase discovery, it is likely that TIMS would also be used prospectively in subsequent clinical trials to identify and enroll patients whose tumors contain targets appropriate for the drug. This would pull the method along into the wider diagnostics marketplace. PUBLIC HEALTH RELEVANCE: Diseases are complex and differ from patient to patient, and our ability to see these differences is improving. Patients should receive the therapy or drug most likely to benefit them while avoiding the expense and toxic side effects of ineffective alternatives. More effective measurement of markers of disease in human tissue biopsies can more accurately align patients with the treatment most likely to benefit them.