Comparative Tissue Proteomics of Microdissected Specimens Reveals Novel Candidate Biomarkers of Bladder Cancer
Comparative Tissue Proteomics of Microdissected Specimens Reveals Novel Candidate Biomarkers of Bladder Cancer
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DOI：
10.1074/mcp.m115.051524
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发表时间：
2015-09-01
影响因子：
7
通讯作者：
Chen, Yi-Ting
中科院分区：
文献类型：
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作者：
Chen, Chien-Lun;Chung, Ting;Chen, Yi-Ting
全世界诊断出超过380，000例膀胱癌新病例，每年约有150，200例死亡。为了发现膀胱癌的潜在生物标志物，我们采用了一种结合激光显微切割、用于相对和绝对定量标记的同量异位素标签以及液相色谱-串联质谱（LC-MS/MS）分析的策略来分析新鲜冷冻膀胱肿瘤标本中的蛋白质组变化。从四对手术切除的原发性膀胱癌肿瘤和邻近的非肿瘤组织中提取细胞蛋白，用于两批同量异位素标签的相对和绝对定量实验，共鉴定了3220种蛋白质。大卫（注释、可视化和综合发现数据库）对失调蛋白的分析显示，三个最重要的生物过程是细胞外基质组织、细胞外结构组织和氧化还原。膀胱癌组织中蛋白质的表达上调，反映了膀胱癌组织对有机物、金属离子和无机物的反应。选择7个差异表达蛋白作为潜在的膀胱癌生物标志物进行进一步验证。免疫组化分析显示，与非癌膀胱上皮细胞相比，肿瘤细胞中的SLC 3A 2、STMN 1和TAGLN 2三种蛋白的水平显著升高，并且表明TAGLN 2可以是用于诊断（AUC = 0.999）和评估膀胱癌患者的淋巴结转移的有用的肿瘤组织标志物。ELISA结果显示，与对照组相比，膀胱癌亚组中STMN 1和TAGLN 2的尿液水平显著增加。与年龄匹配的疝尿液样本相比，膀胱癌样本中的尿液TAGLN 2显示出最大的倍数变化（7.13倍），曲线下面积值为0.70（p < 0.001，n = 205）。总体而言，TAGLN 2在个体膀胱癌组织和尿液样本中显示出最显著的过表达，因此代表了用于膀胱癌非侵入性筛查的潜在生物标志物。我们的研究结果突出了膀胱组织蛋白质组的价值，为将来尿路上皮癌潜在生物标志物的验证研究提供了有价值的信息。
More than 380,000 new cases of bladder cancer are diagnosed worldwide, accounting for similar to 150,200 deaths each year. To discover potential biomarkers of bladder cancer, we employed a strategy combining laser microdissection, isobaric tags for relative and absolute quantitation labeling, and liquid chromatography-tandem MS (LC-MS/MS) analysis to profile proteomic changes in fresh-frozen bladder tumor specimens. Cellular proteins from four pairs of surgically resected primary bladder cancer tumor and adjacent nontumorous tissue were extracted for use in two batches of isobaric tags for relative and absolute quantitation experiments, which identified a total of 3220 proteins. A DAVID (database for annotation, visualization and integrated discovery) analysis of dysregulated proteins revealed that the three top-ranking biological processes were extracellular matrix organization, extracellular structure organization, and oxidation-reduction. Biological processes including response to organic substances, response to metal ions, and response to inorganic substances were highlighted by up-expressed proteins in bladder cancer. Seven differentially expressed proteins were selected as potential bladder cancer biomarkers for further verification. Immunohistochemical analyses showed significantly elevated levels of three proteins-SLC3A2, STMN1, and TAGLN2-in tumor cells compared with noncancerous bladder epithelial cells, and suggested that TAGLN2 could be a useful tumor tissue marker for diagnosis (AUC = 0.999) and evaluating lymph node metastasis in bladder cancer patients. ELISA results revealed significantly increased urinary levels of both STMN1 and TAGLN2 in bladder cancer subgroups compared with control groups. In comparisons with age-matched hernia urine specimens, urinary TAGLN2 in bladder cancer samples showed the largest fold change (7.13-fold), with an area-under-the-curve value of 0.70 (p < 0.001, n = 205). Overall, TAGLN2 showed the most significant overexpression in individual bladder cancer tissues and urine specimens, and thus represents a potential biomarker for noninvasive screening for bladder cancer. Our findings highlight the value of bladder tissue proteome in providing valuable information for future validation studies of potential biomarkers in urothelial carcinoma.