Identification of Ovarian Cancer Plasma Biomarkers

描述（由申请人提供）：上皮性卵巢癌（EOC）是美国最致命的妇科癌症，迫切需要新的血液生物标志物。CA 125是FDA批准的EOC最常用的生物标志物，但由于灵敏度和特异性不足，未被批准用于早期诊断。此外，尽管CA 125通常用于在诊断后辅助疾病的临床管理，但它不能用于其中CA 125在诊断时没有显著升高的大部分EOC患者，其包括透明细胞和粘液亚型。我们的长期目标是确定新的EOC血液和肿瘤组织生物标志物，并进行实验室规模的验证，以确定哪些生物标志物可以改善这种疾病的早期诊断和/或临床管理。我们的总体工作假设是，最特异的EOC血液生物标志物将是卵巢肿瘤脱落到血液中的蛋白质，并且在无癌症个体的血液中处于非常低的丰度水平。在过去的一年里，我们实现了最初提案的所有目标。具体来说，我们使用一种新的深入蛋白质组分析方法鉴定了29种低丰度的新型生物标志物，以鉴定含有人类肿瘤的异种移植小鼠血清或血浆中的人类蛋白质。然后，使用无标记多路复用多反应监测（MRM）质谱（MS）测定，与非癌症对照相比，这些蛋白质在EOC患者的血液中升高。我们下一个资助期的具体目标是在几个较大的患者队列中对这12种生物标志物进行实验室规模的验证，以确定用于早期诊断的最佳生物标志物。 EOC和改善这种疾病的临床管理。我们还将对这些生物标志物区分主要EOC亚型的能力进行初步评估，因为这些是遗传上不同的疾病。为了确定某些生物标志物是否可以补充CA 125，我们将对透明细胞和粘液性卵巢肿瘤的分泌组进行重点评价。我们还将单独评估我们的生物标志物在我们新的较大患者队列中表现出低CA 125水平的患者血浆样本子集中的预测价值。将在5年内追求的两个目标是：1）为我们的12种新型EOC生物标志物开发组织和高通量临床前血浆测定，以及2）检测来自多个独立患者队列的血浆或血清中我们的12种新型EOC生物标志物水平的灵敏度和特异性。将使用新型ImmunoMRM方法定量患者和对照血浆或血清中的生物标志物，该方法使用稳定同位素标记的完整蛋白作为内标物，通过市售抗体多重下拉靶向完整蛋白。预计我们的12种生物标志物的子集被证明对EOC的早期诊断和/或临床管理有用，未来将在CLIA认证的实验室中进行更大规模的多中心临床前测试。

DESCRIPTION (provided by applicant): There is an urgent need for new blood biomarkers for epithelial ovarian cancer (EOC), which is the most lethal gynecological cancer in the United States. CA125 is the most commonly used FDA approved biomarker for EOC, but it is not approved for early diagnosis due to inadequate sensitivity and specificity. Furthermore, although CA125 is commonly used to assist in clinical management of the disease after diagnosis, it cannot be used for the substantial portion of EOC patients where CA125 is not substantially elevated at time of diagnosis, which includes the clear cell and mucinous subtypes. Our long-term goal is to identify novel EOC blood-based and tumor tissue biomarkers and conduct laboratory-scale validation to determine which biomarkers can improve early diagnosis and/or clinical management of this disease. Our overall working hypothesis is that the most specific EOC blood biomarkers will be proteins shed by the ovarian tumor into the blood and that are at very low abundance levels in the blood of cancer-free individuals. During the past grant year, we achieved all goals of the original proposal. Specifically, we identified 29 low-abundance, novel biomarkers using a novel in-depth proteome analysis method to identify human proteins in serum or plasma of xenografted mice containing human tumors. These proteins were then shown to be elevated in blood from EOC patients compared with non-cancer controls using a label-free multiplexed multiple reaction monitoring (MRM) mass spectrometry (MS) assay. We have now narrowed this list to 12 high priority biomarkers using biological and functional criteria Our specific goal for the next grant period is to conduct laboratory-scale validation of these 12 biomarkers in several larger patient cohorts to identify the best biomarkers for early diagnosis of EOC and for improved clinical management of this disease. We will also conduct preliminary evaluations of the capacities of these biomarkers to distinguish among major EOC subtypes, as these are genetically different diseases. To determine whether some biomarkers could complement CA125, we will conduct a focused evaluation of secretomes of clear cell and mucinous ovarian tumors. We will also separately evaluate the predictive value of our biomarkers in the subset of patient plasma samples that exhibit low CA125 levels in our new larger patient cohorts. The two aims that will be pursued over a 5- year period are: 1) Develop tissue and high-throughput preclinical plasma assays for our 12 novel EOC biomarkers, and 2) Validate sensitivity and specificity of our 12 novel EOC biomarker levels in plasma or serum from multiple independent patient cohorts. The biomarkers will be quantified in patient and control plasma or serum using a novel ImmunoMRM method that multiplexes pull-down of targeted intact proteins by commercially-available antibodies using stable isotope-labeled intact proteins as internal standards. It is expected that the subset of our 12 biomarkers that are shown to be useful for early diagnosis and/or clinical management of EOC will advance to larger-scale multi-center, preclinical testing in CLIA certified laboratories in the future.