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Gene Expression and DNA Methylation in Acute Lymphoblastic Leukemia
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Molecular Medicine.
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Pediatric acute lymphoblastic leukemia (ALL) is the most common malignancy in children, which results from the malignant transformation of progenitor cells in the bone marrow into leukemic cells. The precise mechanisms for this transformation are not well defined, however recent studies suggest that aberrant regulation of gene expression or DNA methylation may play an important role. Hence, the aim of this thesis was to use novel methods to investigate genome-wide gene expression and DNA methylation patterns in a large collection of primary ALL cells from pediatric patients. With these studies, we aimed to increase the understanding of factors that regulate gene expression and DNA methylation in ALL.

In the first study of the thesis we found that data obtained from genome-wide digital gene expression analysis enabled excellent cytogenetic subtype-specific classification of ALL cells and revealed new features of gene expression within the disease, such as prevalent antisense transcription and alternative polyadenylation. In the second study we used technology developed for large-scale single nucleotide polymorphism (SNP) genotyping for quantitative analysis of allele-specific gene expression (ASE), revealing widespread ASE in ALL cells. Analysis of DNA methylation in promoter regions of the genes displaying ASE using DNA-microarrays revealed frequent regulation of gene expression by DNA methylation. In the third study, using the same DNA methylation array, we identified differences in the DNA methylation patterns in ALL cells at diagnosis compared to healthy mononuclear cells from the bone marrow of the same children at remission. In the fourth study we measured the DNA methylation of >450,000 CpG sites across the genome in a large collection of ALL samples and non-leukemic control cells. We found that ALL cells displayed highly divergent DNA methylation patterns depending on their cytogenetic subtype and widespread regions of differential methylation were enriched for repressive histone marks. DNA methylation levels at distinct regions in the genome were substantially increased at relapse compared to matched cells from diagnosis.

Collectively, the results presented in this thesis provide new insights into the patterns of gene expression and epigenetic changes in ALL and further increase our understanding of the development and progression of the disease, which will hopefully lead to better treatment options in the future.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2012. , 52 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 799
Keyword [en]
Digital gene expression, Allele-specific gene expression, DNA methylation, Epigenetics, Acute lymphoblastic leukemia
National Category
Pediatrics Hematology Cancer and Oncology
Research subject
Molecular Medicine
Identifiers
URN: urn:nbn:se:uu:diva-179680ISBN: 978-91-554-8442-2 (print)OAI: oai:DiVA.org:uu-179680DiVA: diva2:545964
Public defence
2012-10-05, Rudbecksalen, Rudbeck Laboratoriet, Uppsala, 13:00 (English)
Opponent
Supervisors
Available from: 2012-09-13 Created: 2012-08-21 Last updated: 2013-01-22Bibliographically approved
List of papers
1. Digital gene expression profiling of primary acute lymphoblastic leukemia cells
Open this publication in new window or tab >>Digital gene expression profiling of primary acute lymphoblastic leukemia cells
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2012 (English)In: Leukemia, ISSN 0887-6924, E-ISSN 1476-5551, Vol. 26, no 6, 1218-1227 p.Article in journal (Refereed) Published
Abstract [en]

We determined the genome-wide digital gene expression (DGE) profiles of primary acute lymphoblastic leukemia (ALL) cells from 21 patients taking advantage of 'second-generation' sequencing technology. Patients included in this study represent four cytogenetically distinct subtypes of B-cell precursor (BCP) ALL and T-cell lineage ALL (T-ALL). The robustness of DGE combined with supervised classification by nearest shrunken centroids (NSC) was validated experimentally and by comparison with published expression data for large sets of ALL samples. Genes that were differentially expressed between BCP ALL subtypes were enriched to distinct signaling pathways with dic(9;20) enriched to TP53 signaling, t(9;22) to interferon signaling, as well as high hyperdiploidy and t(12;21) to apoptosis signaling. We also observed antisense tags expressed from the non-coding strand of ∼50% of annotated genes, many of which were expressed in a subtype-specific pattern. Antisense tags from 17 gene regions unambiguously discriminated between the BCP ALL and T-ALL subtypes, and antisense tags from 76 gene regions discriminated between the 4 BCP subtypes. We observed a significant overlap of gene regions with alternative polyadenylation and antisense transcription (P<1 × 10(-15)). Our study using DGE profiling provided new insights into the RNA expression patterns in ALL cells.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-168670 (URN)10.1038/leu.2011.358 (DOI)000305081000009 ()22173241 (PubMedID)
Available from: 2012-02-14 Created: 2012-02-14 Last updated: 2017-12-07Bibliographically approved
2. Allele-specific gene expression patterns in primary leukemic cells reveal regulation of gene expression by CpG site methylation
Open this publication in new window or tab >>Allele-specific gene expression patterns in primary leukemic cells reveal regulation of gene expression by CpG site methylation
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2009 (English)In: Genome Research, ISSN 1088-9051, E-ISSN 1549-5469, Vol. 19, no 1, 1-11 p.Article in journal (Refereed) Published
Abstract [en]

To identify genes that are regulated by cis-acting functional elements in acute lymphoblastic leukemia (ALL) we determined the allele-specific expression (ASE) levels of 2, 529 genes by genotyping a genome-wide panel of single nucleotide polymorphisms in RNA and DNA from bone marrow and blood samples of 197 children with ALL. Using a reproducible, quantitative genotyping method and stringent criteria for scoring ASE, we found that 16% of the analyzed genes display ASE in multiple ALL cell samples. For most of the genes, the level of ASE varied largely between the samples, from 1.4-fold overexpression of one allele to apparent monoallelic expression. For genes exhibiting ASE, 55% displayed bidirectional ASE in which overexpression of either of the two SNP alleles occurred. For bidirectional ASE we also observed overall higher levels of ASE and correlation with the methylation level of these sites. Our results demonstrate that CpG site methylation is one of the factors that regulates gene expression in ALL cells.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-98392 (URN)10.1101/gr.083931.108 (DOI)000262200000001 ()18997001 (PubMedID)
Available from: 2009-02-20 Created: 2009-02-20 Last updated: 2017-12-13Bibliographically approved
3. DNA Methylation Analysis of Bone Marrow Cells at Diagnosis of Acute Lymphoblastic Leukemia and at Remission
Open this publication in new window or tab >>DNA Methylation Analysis of Bone Marrow Cells at Diagnosis of Acute Lymphoblastic Leukemia and at Remission
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2012 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 4, e34513- p.Article in journal (Refereed) Published
Abstract [en]

To detect genes with CpG sites that display methylation patterns that are characteristic of acute lymphoblastic leukemia (ALL) cells, we compared the methylation patterns of cells taken at diagnosis from 20 patients with pediatric ALL to the methylation patterns in mononuclear cells from bone marrow of the same patients during remission and in non-leukemic control cells from bone marrow or blood. Using a custom-designed assay, we measured the methylation levels of 1,320 CpG sites in regulatory regions of 413 genes that were analyzed because they display allele-specific gene expression (ASE) in ALL cells. The rationale for our selection of CpG sites was that ASE could be the result of allele-specific methylation in the promoter regions of the genes. We found that the ALL cells had methylation profiles that allowed distinction between ALL cells and control cells. Using stringent criteria for calling differential methylation, we identified 28 CpG sites in 24 genes with recurrent differences in their methylation levels between ALL cells and control cells. Twenty of the differentially methylated genes were hypermethylated in the ALL cells, and as many as nine of them (AMICA1, CPNE7, CR1, DBC1, EYA4, LGALS8, RYR3, UQCRFS1, WDR35) have functions in cell signaling and/or apoptosis. The methylation levels of a subset of the genes were consistent with an inverse relationship with the mRNA expression levels in a large number of ALL cells from published data sets, supporting a potential biological effect of the methylation signatures and their application for diagnostic purposes.

National Category
Hematology Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-178755 (URN)10.1371/journal.pone.0034513 (DOI)000305012700038 ()22493696 (PubMedID)
Available from: 2012-08-01 Created: 2012-08-01 Last updated: 2017-12-07Bibliographically approved
4. DNA hypermethylation signatures for genetic subtypes of pediatric ALL at diagnosis and relapse
Open this publication in new window or tab >>DNA hypermethylation signatures for genetic subtypes of pediatric ALL at diagnosis and relapse
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(English)Manuscript (preprint) (Other academic)
National Category
Hematology Pediatrics Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-179542 (URN)
Available from: 2012-08-19 Created: 2012-08-19 Last updated: 2013-01-22

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