Bouchal Pavel, Dvořáková Monika, Roumeliotis Theodoros, Bortlíček Zbyněk, Ihnatová Ivana, Procházková Iva, Ho Jenny T C, Maryáš Josef, Imrichová Hana, Budinská Eva, Vyzula Rostislav, Garbis Spiros D, Vojtěšek Bořivoj, Nenutil Rudolf
From the ‡Masaryk Memorial Cancer Institute, Regional Centre for Applied Molecular Oncology, Brno, Czech Republic; §Masaryk University, Faculty of Science, Department of Biochemistry, Brno, Czech Republic;
¶Proteomics Mass Spectrometry, The Wellcome Trust Sanger Institute, Cambridge CB10 1SA, UK;
Mol Cell Proteomics. 2015 Jul;14(7):1814-30. doi: 10.1074/mcp.M114.041335. Epub 2015 Apr 22.
Current prognostic factors are insufficient for precise risk-discrimination in breast cancer patients with low grade breast tumors, which, in disagreement with theoretical prognosis, occasionally form early lymph node metastasis. To identify markers for this group of patients, we employed iTRAQ-2DLC-MS/MS proteomics to 24 lymph node positive and 24 lymph node negative grade 1 luminal A primary breast tumors. Another group of 48 high-grade tumors (luminal B, triple negative, Her-2 subtypes) was also analyzed to investigate marker specificity for grade 1 luminal A tumors. From the total of 4405 proteins identified (FDR < 5%), the top 65 differentially expressed together with 30 previously identified and control markers were analyzed also at transcript level. Increased levels of carboxypeptidase B1 (CPB1), PDZ and LIM domain protein 2 (PDLIM2), and ring finger protein 25 (RNF25) were associated specifically with lymph node positive grade 1 tumors, whereas stathmin 1 (STMN1) and thymosin beta 10 (TMSB10) associated with aggressive tumor phenotype also in high grade tumors at both protein and transcript level. For CPB1, these differences were also observed by immunohistochemical analysis on tissue microarrays. Up-regulation of putative biomarkers in lymph node positive (versus negative) luminal A tumors was validated by gene expression analysis of an independent published data set (n = 343) for CPB1 (p = 0.00155), PDLIM2 (p = 0.02027) and RELA (p = 0.00015). Moreover, statistically significant connections with patient survival were identified in another public data set (n = 1678). Our findings indicate unique pro-metastatic mechanisms in grade 1 tumors that can include up-regulation of CPB1, activation of NF-κB pathway and changes in cell survival and cytoskeleton. These putative biomarkers have potential to identify the specific minor subpopulation of breast cancer patients with low grade tumors who are at higher than expected risk of recurrence and who would benefit from more intensive follow-up and may require more personalized therapy.
目前的预后因素不足以对低级别乳腺肿瘤的乳腺癌患者进行精确的风险区分，这类肿瘤偶尔会发生早期淋巴结转移，这与理论预后情况不符。为了识别这类患者的标志物，我们对24例淋巴结阳性和24例淋巴结阴性的1级管腔A型原发性乳腺肿瘤采用了iTRAQ-2DLC-MS/MS蛋白质组学技术。另外一组48例高级别肿瘤（管腔B型、三阴性、Her-2亚型）也进行了分析，以研究1级管腔A型肿瘤标志物的特异性。在总共鉴定出的4405种蛋白质（FDR<5%）中，对前65种差异表达的蛋白质以及30种先前鉴定的蛋白质和对照标志物也进行了转录水平分析。羧肽酶B1（CPB1）、PDZ和LIM结构域蛋白2（PDLIM2）以及环指蛋白25（RNF25）水平升高与淋巴结阳性的1级肿瘤特异性相关，而在高级别肿瘤中，无论在蛋白质水平还是转录水平，微管蛋白1（STMN1）和胸腺素β10（TMSB10）都与侵袭性肿瘤表型相关。对于CPB1，在组织微阵列上通过免疫组织化学分析也观察到了这些差异。通过对一个独立的已发表数据集（n = 343）进行基因表达分析，验证了淋巴结阳性（与阴性相比）管腔A型肿瘤中假定生物标志物的上调情况，其中CPB1（p = 0.00155）、PDLIM2（p = 0.02027）和RELA（p = 0.00015）。此外，在另一个公共数据集（n = 1678）中发现了与患者生存具有统计学意义的关联。我们的研究结果表明，1级肿瘤中存在独特的促转移机制，可能包括CPB1上调、NF-κB通路激活以及细胞存活和细胞骨架的变化。这些假定的生物标志物有可能识别出肿瘤级别较低但复发风险高于预期的特定小群体乳腺癌患者，这些患者将受益于更密切的随访，可能需要更个性化的治疗。
