Abstract
The mutational status of the immunoglobulin heavy chain variable region defines two clinically distinct forms of chronic lymphocytic leukemia (CLL) known as mutated (M-CLL) and unmutated (UM-CLL). To elucidate the molecular mechanisms underlying the adverse clinical outcome associated with UM-CLL, total proteomes from nine UM-CLL and nine M-CLL samples were analyzed by isobaric tags for relative and absolute quantification (iTRAQ)-based mass spectrometry. Based on the expression of 3521 identified proteins, principal component analysis separated CLL samples into two groups corresponding to immunoglobulin heavy chain variable region 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 underexpressed proteins associated with cytoskeletal remodeling and overexpressed proteins associated with transcriptional and translational activity. Taken together, our findings indicate 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.
Highlights
IntroductionChronic lymphocytic leukemia (CLL) is the most common adult leukemia in Western countries
From the ‡Department of Molecular and Clinical Cancer Medicine, §MRC Centre for Drug Safety Science, Department of Molecular and Clinical Pharmacology, University of Liverpool, Liverpool L69 3GA, UK; ¶Royal Liverpool and Broadgreen University Hospitals NHS Trust, Liverpool L7 8XP, UK
Quantitative Proteomic Analysis Separates Chronic lymphocytic leukemia (CLL) Samples Based on immunoglobulin heavy chain variable region (IGHV) Mutational Status—To generate quantitative information on the whole proteome of CLL cells, mononuclear cells from nine M-CLL and nine UM-CLL samples containing Ͼ90% CLL cells were subjected to isobaric tags for relative and absolute quantification (iTRAQ)-based mass spectrometry (MS) (Table 1)
Summary
Chronic lymphocytic leukemia (CLL) is the most common adult leukemia in Western countries It is characterized by the clonal expansion of antigen-experienced B cells with a distinctive immunophenotype [1, 2]. Many biological variables have been identified in CLL that correlate with clinical outcome Among these variables, the somatic mutational status of the immunoglobulin heavy chain variable region (IGHV) gene expressed by the malignant clone has a unique biological importance as it is the only prognostic biomarker that is fixed at the initiation of clonal expansion, inherited by the entire malignant clone, and stable over time (4 – 6). The abbreviations used are: CLL, chronic lymphocytic leukemia; M-CLL, IGHV-mutated CLL; UM-CLL, IGHV-unmutated CLL; IGHV, immunoglobulin heavy chain variable region; ACN, acetonitrile; MS, mass spectrometry; iTRAQ, isobaric tags for relative and absolute quantification; PANTHER, Protein Analysis Through Evolutionary Relationships; MNDA, myeloid cell nuclear differentiation antigen; PCA, principal component analysis. IGHV status is a strong and independent predictor of outcome in CLL, with M-CLL being associated with a favorable outcome and UM-CLL being associated with early disease progression and shorter survival (4 – 6)
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