Glioblastoma is one of the most malignant primary brain cancer. Despite surgical resection with modern technology followed by chemo-radiation therapy with temozolomide, resistance to the treatment and recurrence is common due to its aggressive and infiltrating nature of the tumor with high proliferation index. The median survival time of the patients with glioblastomas is less than 15 months. Till now there has been no report of molecular target specific for glioblastomas. Early diagnosis and development of molecular target specific for glioblastomas are essential for longer survival of the patients with glioblastomas. Development of biomarkers specific for glioblastomas is most important for early diagnosis, estimation of the prognosis, and molecular target therapy of glioblastomas. To that end, in this study, we have conducted a comprehensive proteome study using primary cells and tissues from patients with glioblastoma. In the discovery stage, we have identified 7429 glioblastoma-specific proteins, where 476 proteins were quantitated using Tandem Mass Tag (TMT) method; 228 and 248 proteins showed up and down-regulated pattern, respectively. In the validation stage (20 selected target proteins), we developed quantitative targeted method (MRM: Multiple reaction monitoring) using stable isotope standards (SIS) peptide. In this study, five proteins (CCT3, PCMT1, TKT, TOMM34, UBA1) showed the significantly different protein levels (t-test: p value ≤ 0.05, AUC ≥ 0.7) between control and cancer groups and the result of multiplex assay using logistic regression showed the 5-marker panel showed better sensitivity (0.80 and 0.90), specificity (0.92 and 1.00), error rate (10 and 2%), and AUC value (0.94 and 0.98) than the best single marker (TOMM34) in primary cells and tissues, respectively. Although we acknowledge that the model requires further validation in a large sample size, the 5 protein marker panel can be used as baseline data for the discovery of novel biomarkers of the glioblastoma. For the discovery of multi-diagnostic biomarker, we have conducted a comprehensive proteome study using primary cells from patients with glioblastoma. In this study, 7429 glioblastoma-specific proteins were identified and then 20 selected target proteins were verified using MRM method. Finally, five proteins (CCT3, PCMT1, TKT, TOMM34, UBA1) showed the significantly different protein levels (t-test: p value ≤ 0.05, AUC ≥ 0.7) between control and cancer groups and the result of multiplex assay using logistic regression showed the 5-marker panel showed better sensitivity (0.80 and 0.90), specificity (0.92 and 1.00), error rate (10 and 2%), and AUC value (0.94 and 0.98) than the best single marker (TOMM34) in primary cells and tissues, respectively.

中文翻译：

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使用靶向定量方法的胶质细胞蛋白质组作为潜在的多重诊断生物标志物

胶质母细胞瘤是最恶性的原发性脑癌之一。尽管采用现代技术进行手术切除，然后进行替莫唑胺化放疗治疗，但由于肿瘤具有高增殖指数的侵袭性和浸润性，对治疗的抵抗和复发仍然很常见。胶质母细胞瘤患者的中位生存时间不到15个月。到目前为止，还没有针对胶质母细胞瘤的特异性分子靶点的报道。早期诊断和开发胶质母细胞瘤特异性分子靶点对于延长胶质母细胞瘤患者的生存至关重要。开发胶质母细胞瘤特异性生物标志物对于胶质母细胞瘤的早期诊断、预后评估和分子靶向治疗至关重要。为此，在本研究中，我们利用胶质母细胞瘤患者的原代细胞和组织进行了全面的蛋白质组研究。在发现阶段，我们已经鉴定了7429个胶质母细胞瘤特异性蛋白，其中476个蛋白使用串联质量标签（TMT）方法进行了定量；228 和 248 个蛋白质分别显示上调和下调模式。在验证阶段（20 个选定的目标蛋白），我们开发了使用稳定同位素标准品 (SIS) 肽的定量靶向方法（MRM：多反应监测）。在本研究中，五种蛋白质（CCT3、PCMT1、TKT、TOMM34、UBA1）在对照组和癌症组之间显示出显着不同的蛋白质水平（t 检验：p 值≤ 0.05，AUC ≥ 0.7），并使用逻辑回归进行多重测定的结果回归显示，与原发性最佳单一标记物 (TOMM34) 相比，5 标记物组合显示出更好的敏感性（0.80 和 0.90）、特异性（0.92 和 1.00）、错误率（10 和 2%）以及 AUC 值（0.94 和 0.98）。分别是细胞和组织。尽管我们承认该模型需要在大样本量中进一步验证，但 5 种蛋白质标记物组合可以用作发现胶质母细胞瘤新型生物标记物的基线数据。为了发现多重诊断生物标志物，我们使用胶质母细胞瘤患者的原代细胞进行了全面的蛋白质组研究。在本研究中，鉴定了 7429 个胶质母细胞瘤特异性蛋白，然后使用 MRM 方法验证了 20 个选定的靶蛋白。最后，五种蛋白质（CCT3、PCMT1、TKT、TOMM34、UBA1）在对照组和癌症组之间显示出显着不同的蛋白质水平（t 检验：p 值 ≤ 0.05，AUC ≥ 0.7），并且使用逻辑回归的多重测定结果显示在原代细胞和组织，分别。