CLL iTRAQ1 - Total proteome analysis identifies migration defects as a major pathogenetic factor in immunoglobulin heavy chain variable region (IGHV)-unmutated chronic lymphocytic leukemia

The mutational status of the immunoglobulin heavy chain variable region (IGHV) defines two clinically distinct forms of chronic lymphocytic leukemia (CLL) known as mutated (M-CLL) and un-mutated (UM-CLL). To elucidate the molecular mechanisms underlying the adverse clinical outcome associated with UM-CLL, total proteomes from 9 UM-CLL and 9 M-CLL samples were analysed by isobaric tags for relative and absolute quantification (iTRAQ)-based mass spectrometry. Unsupervised clustering, based on the expression of 3521 identified proteins, separated CLL samples into two groups corresponding to IGHV mutational status. Computational analysis showed that 43 cell migration/adhesion pathways were significantly enriched by 39 differentially expressed proteins, 35 of which were expressed at significantly lower levels in UM-CLL samples. Furthermore, UM-CLL cells under-expressed proteins associated with cytoskeletal remodelling and over-expressed proteins associated with transcriptional and translational activity. Taken together, our findings indicated that UM-CLL cells are less migratory and more adhesive than M-CLL cells, resulting in their retention in lymph nodes where they are exposed to proliferative stimuli. In keeping with this hypothesis, analysis of an extended cohort of 120 CLL patients revealed a strong and specific association between UM-CLL and lymphadenopathy. Our study illustrates the potential of total proteome analysis to elucidate pathogenetic mechanisms in cancer.

免疫球蛋白重链可变区（immunoglobulin heavy chain variable region, IGHV）的突变状态定义了两种临床特征迥异的慢性淋巴细胞白血病（chronic lymphocytic leukemia, CLL）亚型，即突变型（M-CLL）与未突变型（UM-CLL）。为阐明与UM-CLL相关的不良临床结局背后的分子机制，本研究采用基于同位素相对与绝对定量标签（isobaric tags for relative and absolute quantification, iTRAQ）的质谱分析法，对9例UM-CLL与9例M-CLL样本的全蛋白质组进行了分析。基于3521个已鉴定蛋白质的表达水平进行无监督聚类，可将CLL样本按照IGHV突变状态划分为两个组别。生物信息学分析显示，39个差异表达蛋白显著富集于43条细胞迁移/黏附通路，其中35个蛋白在UM-CLL样本中的表达水平显著下调。此外，UM-CLL细胞中与细胞骨架重塑相关的蛋白表达下调，而与转录及翻译活性相关的蛋白表达上调。综合以上研究结果，本研究表明UM-CLL细胞的迁移能力弱于M-CLL细胞，而黏附能力更强，这使得肿瘤细胞滞留于淋巴结内并受到增殖信号的刺激。与这一假说相符的是，对包含120例CLL患者的扩展队列进行分析后发现，UM-CLL与淋巴结病存在显著且特异性的关联。本研究证实了全蛋白质组分析在阐明癌症发病机制方面的应用潜力。