Alliance of Glycobiologists for Detection of Cancer

描述（由申请人提供）：该提案旨在通过非侵入性方法检测可治疗阶段的肝细胞癌（HCC）。为此，我们建议量化肝脏分泌的N-糖蛋白（LNP）的位点特异性糖型与肝脏疾病进展为HCC相关。我们和其他人已经表明蛋白质糖基化的变化伴随着HCC的发展。蛋白N-糖型的主要重新分布发生在肝硬化的癌前阶段，但我们最新的研究表明，结合珠蛋白的糖肽的特定糖型仅在HCC中增加。我们证明，这些次要的结合珠蛋白糖型在血清中是可检测的，并且与血清学金标准甲胎蛋白（AFP）相比，具有提高HCC检测的潜力。这支持了我们的假设，即丰富的肝脏分泌的N-糖蛋白的次要位点特异性糖型提供了在循环中可检测到的HCC特异性标志物候选物。我们建议完成HCC特异性N-聚糖的鉴定;我们已经有证据表明，聚糖的特征是多重岩藻糖基化和增加的分支。我们建议通过新优化的质谱方法鉴定携带这些HCC特异性N-聚糖修饰的其他蛋白质的糖肽。这样做是因为触珠蛋白在一部分抗肿瘤药物中下调，并且可能需要其他蛋白质进行最佳检测。我们扩展我们的信息学分析，以分类所有多态性和突变的人类N-糖蛋白与新创建或废除的N-糖基化位点。这些蛋白质变体在人类基因组中过度表达，代表与疾病相关的主要候选者，并且我们预期它们对致癌作用具有直接影响。我们相信，这些变异蛋白质的化学可用资源将刺激糖蛋白组学癌症预防研究。建议的HCC特异性N-糖型的检查是可行的，因为我们已经建立了HCC患者和HCC对照的样本库。该储存库和其他优秀的QC资源对于分离肝脏分泌的N-糖蛋白的HCC特异性次要糖型至关重要;这些糖型在无疾病受试者中不可检测。到研究结束时，我们将通过有针对性的LC-MS选择性反应监测（SRM）方法定量所选位点特异性蛋白糖型。这些方法将以前所未有的准确度定量血清中HCC相关位点特异性糖肽。我们这样做是因为我们相信，特定糖蛋白肽的特定糖型的定量提供了最高的诊断准确性。定义临床适用的癌症标志物对疾病管理和患者健康具有潜在的深远影响。我们的研究有望产生关于蛋白质糖基化对癌症发展的功能性影响的新假设，并刺激一种全新的癌症预防和检测研究。

DESCRIPTION (provided by applicant): This proposal aims to detect hepatocellular carcinoma (HCC) at a treatable stage by non-invasive methods. To this end, we propose to quantify site specific glycoforms of liver secreted N-glycoproteins (LNP) associated with the progression of liver disease to HCC. We and others have shown that changes in protein glycosylation accompany the development of HCC. Major re-distribution of protein N-glycoforms occurs at the premalignant stage of liver cirrhosis but our latest studies show that specific glycoforms of a glycopeptide of haptoglobin increase exclusively in HCC. We demonstrate that these minor glycoforms of haptoglobin are detectable in serum and have the potential to improve detection of HCC compared to the serologic gold standard, alpha fetoprotein (AFP). This supports our hypothesis that minor site specific glycoforms of abundant liver secreted N-glycoproteins provide HCC specific marker candidates detectable in the circulation. We propose to complete identification of the HCC specific N-glycans; we have already evidence that the glycans are characterized by multiple fucosylations and increased branching. We propose to identify glycopeptides of additional proteins carrying these HCC specific N-glycan modifications by newly optimized mass spectrometric methods. This is done because haptoglobin is downregulated in a fraction of cirrhotics and other proteins may be needed for optimal detection. We expand our informatic analysis to classify all polymorphic and mutant human N- glycoproteins with newly created or abolished N-glycosylation sites. These protein variants are over- represented in the human genome, represent prime candidates for association with diseases, and we expect that they have direct impact on carcinogenesis. We believe that a publically available resource of these variant proteins will stimulate glycoproteomic cancer prevention research. The proposed examination of HCC specific N-glycoforms is feasible because we have already created a repository of samples of HCC patients and cirrhotic controls. This repository, and additional outstanding QC resources, is essential for the isolation of the HCC specific minor glycoforms of liver secreted N-glycoproteins; these glycoforms are not detectable in disease free subjects. By the end of the study, we will have quantified the selected site specific protein glycoforms by targeted LC-MS selective reaction monitoring (SRM) methods. These methods will quantify the HCC associated site specific glycopeptides in serum with an unprecedented accuracy. We do this because we believe that quantification of specific glycoforms of specific glycoprotein peptides offers the highest diagnostic accuracy. Defining clinically applicable cancer markers has potentially far-reaching consequences for disease management and patient health. Our study is expected to generate new hypotheses on the functional impact of protein glycosylation on the development of cancer and to stimulate an entirely new line of cancer prevention and detection research.