Serum Glyco-Markers of Early Hepatocellular Carcinoma Using a Mass Spec Approach

摘要：肝细胞癌（HCC）是世界范围内第四大常见的癌症相关死亡原因 在美国的发病率正在上升。在美国，超过80%的HCC患者有潜在的肝硬化， 专业协会指南建议对所有肝硬化患者进行监测，以促进早期发现。 不幸的是，只有20-30%的患者在早期阶段被发现时，潜在的治愈性治疗， 可能我们的建议，基于我们强有力的初步数据评估糖蛋白质组学概况，解决了 明确未满足的对准确的早期HCC检测生物标志物的需求。蛋白质中糖基化的独特变化， 涉及聚糖基团的结构变化，已被证明是早期糖尿病的重要血清生物标志物。 HCC检测。重要的是，即使是涉及次要结构的细微变化也可以非常具体地区分 肝硬化与早期HCC。在这项研究中，我们将使用质谱法，这已经证明， 是一种高度灵敏和准确的方法来检测和定量监测聚糖结构的变化。我们 已发表的质量分析发现工作，串联质谱测量，和新的生物信息学 用于聚糖和糖肽分析的工具表明，这些结构上的微小变化可以作为有希望的 早期HCC检测生物标志物。在目标1中，这些方法将被证明用于糖肽筛选， 血清，使用新型质谱分析，使用基于分步的平行反应监测（PRM）分析， HCD片段化方法，最终可以用于区分早期HCC和肝硬化。的 PRM方法是一种靶向检测方法，可以区分聚糖结构的微小变化，并且可以进行多重检测 以同时监测大量标记。此外，使用与PRM策略相结合的分离， 我们将能够区分聚糖结构中岩藻糖基化和唾液酸化的不同异构形式 其可能含有重要的疾病相关标记。在目标2中，本文开发的方法将被优化 然后应用于300个早期HCC和肝硬化样本的大型确认集，用于20个靶点， 结合珠蛋白的糖肽以及发现和评价的其他几种潜在的糖肽标记物 在我们之前的工作中。在目标3中，我们将在盲态II期生物标志物研究中验证生物标志物的性能 使用800-1000名早期HCC和肝硬化患者的大集合，在性别、种族/民族方面不同， 和肝病病因学。通过这些目标，我们将确定一组糖肽， 用于早期HCC检测的准确生物标志物。

Abstract: Hepatocellular carcinoma (HCC) is the fourth most common cause of cancer-related death worldwide and is rising in incidence in the US. Over 80% of patients in the US with HCC have underlying cirrhosis, and professional societies guidelines recommend surveillance in all patients with cirrhosis to facilitate early detection. Unfortunately, only 20-30% of patients are detected at an early stage when potentially curative treatments are possible. Our proposal, based on our strong preliminary data evaluating glycoproteomic profiles, addresses the clear unmet need for accurate early HCC detection biomarkers. Unique changes in glycosylation in proteins, which involve structural changes in glycan groups, have been shown to be important serum biomarkers for early HCC detection. Importantly, even subtle changes involving minor structures can be very specific in differentiating cirrhosis versus early stage HCC. In this study, we will use a mass spectrometry approach, which has proven to be a highly sensitive and accurate way to detect and quantitatively monitor glycan structural changes. Our published mass analysis discovery work, tandem mass spectrometry measurements, and novel bioinformatic tools for glycan and glycopeptide analysis demonstrate these minor changes in structure can act as promising early HCC detection biomarkers. In aim 1 these methods will be demonstrated for glycopeptide screening from serum using novel mass spec assays using parallel reaction monitoring (PRM) analysis based on a stepped HCD fragmentation method which can ultimately be used to distinguish early stage HCC from cirrhosis. The PRM method is a targeted assay which can distinguish small changes in glycan structure and can be multiplexed to monitor large numbers of markers simultaneously. In addition, using separations coupled to the PRM strategy, we will be able to distinguish different isomeric forms of fucosylation and sialylation in glycan structures which may contain important disease related markers. In aim 2 the methods developed herein will be optimized and then applied to a large confirmation set of 300 early stage HCC and cirrhosis samples for 20 target glycopeptides from Haptoglobin and to several other potential glycopeptide markers discovered and evaluated in our prior work. In aim 3 we will then validate the biomarker performance in a blinded phase II biomarker study using a large set of 800-1000 early stage HCC and cirrhosis patients, diverse with regard to sex, race/ethnicity, and liver disease etiology. Through these aims, we will identify a panel of glycopeptides that can serve as highly accurate biomarkers for early HCC detection.