Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Investigating epigenetic effects of activation-induced deaminase in chronic lymphocytic leukemia

ABSTRACT: Activation induced deaminase (AID) has two distinct and well defined roles, both relying on its deoxycytidine (dC) deaminating function: one as a DNA mutator and another in DNA demethylation. In chronic lymphocytic leukemia (CLL), AID was previously shown to be an independent negative prognostic factor. While there is substantial impact on DNA mutations, effects of AID on gene expression by promoter demethylation of disease related target genes in leukemia has not been addressed. To shed light on this question, we aimed at determining genome wide methylation changes as well as gene expression changes in response to AID expression in CLL. Although we found minor differences in individual methylation variable positions following AID expression, we could not find recurrent methylation changes of specific target sites or changes in global methylation.

ORGANISM(S): Homo sapiens

SUBMITTER: Maria Schubert

PROVIDER: E-MTAB-7431 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Chronic lymphocytic leukaemia (CLL) is renowned for its variable clinical course and response to therapy, suggesting a role for precision medicine. However, the molecular basis of CLL variability remains incompletely understood. Recent developments in data independent acquisition (DIA) -MS technologies, such as SWATH (Sequential Windowed Acquisition of all THeoretical fragments), provide an opportunity to study the pathophysiology of CLL at the proteome level. This report describes the sample replication and data handling requirements for SWATH-MS analysis of clinical samples. A CLL-specific spectral library compromising over 1,500,000 spectra with digital information for over 150,000 peptides has been generated to enable the quantification of up to 7736 proteins. To assess the reproducibility of the assay, SWATH-MS analysis was performed on 6 cryopreserved CLL patient samples, incorporating biological (IGHV mutational status), sample preparation and MS technical replicates. Quantitative information was obtained for 5169 proteins (<1% FDR) across 54 SWATH-MS acquisitions. These analyses showed that SWATH-MS is a highly reproducible technique. However, replicate sample preparations on different days was identified as the main source of variation. To overcome the effect of variation between sample preparation batches, different computational approaches for batch correction were tested. All approaches successfully removed technical variability whilst retaining proteomic differences between clinical subgroups. Functional enrichment analysis of proteins associated to IGHV mutational status highlighted the importance of metabolic remodelling in the biology of CLL and overlapped significantly with previous studies based on gene expression profiling. Finally, an approach to perform statistical power analysis in proteomics studies was developed. This approach could be used to drive the design of future CLL SWATH-MS studies. These fundamental requirements for DIA approaches should be widely applicable to other clinical proteomics studies.

Project description:Prospective series of 136 clinical monoclonal B lymphocytosis (cMBL) and 216 chronic lymphocytic leukemia (CLL) Rai 0 patients, were investigated in this study. While the distribution of CD38 and ZAP-70 positivity was similar, IGHV-mutated cases were more frequent among cMBL (P = 0.005). A Cox multivariate analysis on the whole patient cohort showed that cMBL condition was predictive of longer PFS, while CD38 expression and IGHV-unmutated status and CD38 expression correlated significantly with a shorter PFS in cMBL and Rai0-CLL, respectively. Trisomy 12, 11q- and 17p- abnormalities were scanty and of no predictive value in both conditions. Notably, gene and miRNA expression profiling showed no significant differences between cMBL and Rai0-CLL. Furthermore, similar gene and miRNA expression signatures were found in cMBL and Rai0-CLL according to the IGHV gene mutational status: that is, unmutated cases had different signatures from mutated cases, irrespectively of the cMBL or CLL condition. Overall, our study based on a prospective series of patients indicates that no major biological differences exist in cMBL compared to Rai0-CLL, suggesting that this two entities mainly differ for the initial size of the monoclonal cell population which may reflect in the longer time for clonal expansion. This series of microarray experiments contains the gene expression profiles of purified B-cell chronic lymphocytic leukemia (B-CLL) cells obtained from 160 patients (Binet stage A) including 45 cMBL and 115 Rai0-CLL. Peripheral blood mononuclear cells from CLL patients were isolated by Ficoll-Hypaque (Seromed, Biochrom KG, Berlin, Germany) density-gradient centrifugation. For gene and miRNA expression profiling experiments CLL cells were enriched by negative selection with the EasySep-Human B cell enrichment kit without CD43 depletion (Stem Cell Technologies) using the fully automated protocol of immunomagnetic cell separation with RoboSepTM (Stem Cell Technologies).

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Investigating epigenetic effects of activation-induced deaminase in chronic lymphocytic leukemia

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