Glycan Biomarkers of Prostate Cancer in Prostatic Fluids

描述（由申请人提供）：前列腺特异性抗原（PSA）和前列腺酸性磷酸酶（PAP）是由前列腺上皮细胞分泌的糖蛋白，作为前列腺癌的血清生物标志物具有长期的临床应用史。这两种蛋白质和许多其他前列腺衍生的蛋白质在精浆中的浓度显著更高，并且在直肠指诊前列腺按摩后尿液中的前列腺分泌物（EPS）也显著更高。质谱仪器和技术的新进展推动了对表征与癌症相关的蛋白质上聚糖结构改变的重新重视。先前发表的关于少数样本的研究表明，PSA和PAP上不同N-连接聚糖结构的存在可以区分良性前列腺疾病和前列腺癌。我们对来自大量精浆的PSA和PAP聚糖的初步数据以及对单个样本点的比较证实了这一点。提出了一种组合的多重质谱和基于ELISA的方法来表征前列腺液中源自PSA和PAP的N-连接聚糖。这种方法专门测试了以下假设：随着前列腺癌在疾病严重程度上的进展，多种聚糖结构变化是可检测的，并且在分泌的前列腺蛋白如PSA和PAP上持续发生。我们的方法代表了前列腺癌生物标志物策略从传统的基于血清或组织的来源的范式转变。它还强调了基于与癌症表型相关的表面和分泌糖蛋白的碳水化合物表达的充分描述的变化的诊断测定的发展。为了实现这些目标，我们组建了一个协同合作团队，该团队拥有前列腺癌转化研究和蛋白质组学方面的专业知识，并结合聚糖分析专家。必要的临床样本、仪器和聚糖分析工作流程可用于实现以下特定目标。1)识别近端前列腺液中PAP和PSA上的N-连接聚糖生物标志物，反映前列腺癌病理学和疾病状态。2)直肠指检后大规模EPS尿液中前列腺疾病特异性PAP和PSA糖型的检测和预验证将开发特异性唾液酸和岩藻糖靶向凝集素ELISA。3)使用iTRAQ比较在EPS样本队列中识别其他候选前列腺疾病糖蛋白生物标志物，包括疾病状态间聚糖变化的综合特征。我们期望PSA和PAP上聚糖的表征将确定改善前列腺癌检测和危险分层的分子标记物。公共卫生相关性：目前PSA血清检测用于前列腺癌早期检测和治疗的优势和局限性已得到充分证明，很明显，这种疾病需要新的诊断生物标志物。我们建议表征前列腺分泌的液体中蛋白质聚糖组分的癌症特异性变化。这些分子标志物的鉴定将改善前列腺癌的检测和活检和前列腺切除术前的风险分层。

DESCRIPTION (provided by applicant): Prostate-specific antigen (PSA) and prostatic acid phosphatase (PAP) are glycoproteins secreted by prostate epithelial cells, and have a long clinical history of use as serum biomarkers of prostate cancers. These two proteins and many other prostatic-derived proteins are present at significantly higher concentrations in seminal plasma and expressed prostatic secretions (EPS) in urine following a digital rectal exam prostate massage. New advances in mass spectrometry instrumentation and techniques have fueled a renewed emphasis on characterizing alterations in glycan structures on proteins associated with cancers. Previous published studies on a handful of samples have suggested that the presence of different N-linked glycan structures on PSA and PAP could distinguish benign prostate diseases from prostate cancer. Our preliminary data on PSA and PAP glycans derived from large seminal plasma cohorts and comparisons of individual sample spots has confirmed this. A combination of multiple mass spectrometry and ELISA- based approaches to characterizing N-linked glycans derived from PSA and PAP in prostatic fluids are proposed. This approach specifically tests the hypothesis that multiple glycan structural changes are detectable and consistently occur on secreted prostatic proteins like PSA and PAP as prostate cancers progress in disease severity. Our approach represents a paradigm shift for prostate cancer biomarker strategies away from the traditional serum or tissue based sources. It also emphasizes the development of diagnostic assays based on the well described changes in carbohydrate expression of surface and secreted glycoproteins associated with the cancer phenotype. To accomplish these goals, we have assembled a synergistic collaborative team with expertise in prostate cancer translational research and proteomics combined with glycan analysis specialists. The requisite clinical samples, instrumentation and glycan analysis workflows are available to accomplish the following specific aims. 1) Identify N-linked glycan biomarkers on PAP and PSA in proximal prostatic fluids reflective of prostate cancer pathology and disease state. 2) The detection and pre-validation of prostate-disease specific PAP and PSA glycoforms in a large-scale cohort of EPS urines obtained after digital rectal exams. Specific sialic acid and fucose targeted lectin ELISAs will be developed. 3) Identify additional candidate prostatic disease glycoprotein biomarkers in the EPS sample cohort using iTRAQ comparisons, including a comprehensive profile of the glycan changes across disease states. We expect that characterization of the glycans on PSA and PAP will identify molecular markers that improve prostate cancer detection and risk stratification. PUBLIC HEALTH RELEVANCE: The current strengths and limitations of the PSA serum test for early detection and treatment of prostate cancers are well documented, and it is clear new diagnostic biomarkers are needed for this disease. We propose to characterize cancer specific changes in the glycan components of proteins in fluids secreted by the prostate. Identification of these molecular markers will improve prostate cancer detection and risk stratification prior to biopsy and prostatectomy procedures.