Epigenetic signature of ionizing radiation in therapy-related AML patients.

Therapy-related acute myeloid leukaemia (t-AML) is a late side effect of previous chemotherapy (ct-AML) and/or radiotherapy (rt-AML) or immunosuppressive treatment. t-AMLs, which account for ∼10-20 % of all AML cases, are extremely aggressive and have a poor prognosis compared to de novo AML. Our hypothesis is that exposure to radiation causes genome-wide epigenetic changes in rt-AML. An epigenome-wide association study was undertaken, measuring over 850K methylation sites across the genome from fifteen donors (five healthy, five de novo, and five t-AMLs). The study predominantly focussed on 94K sites that lie in CpG-rich gene promoter regions. Genome-wide hypomethylation was discovered in AML, primarily in intergenic regions. Additionally, genes specific to AML were identified with promoter hypermethylation. A two-step validation was conducted, both internally, using pyrosequencing to measure methylation levels in specific regions across fifteen primary samples, and externally, with an additional eight AML samples. We demonstrated that the MEST and GATA5 gene promoters, which were previously identified as tumour suppressors, were noticeably hypermethylated in rt-AML, as opposed to other subtypes of AML and control samples. These may indicate the epigenetic involvement in the development of rt-AML at the molecular level and could serve as potential targets for drug therapy in rt-AML.

Region-Specific Methylation Profiling in Acute Myeloid Leukemia.

Alteration of DNA methylation level in cancer diseases leads to deregulation of gene expression-silencing of tumor suppressor genes and enhancing of protooncogenes. There are several tools devoted to the problem of identification of CpG sites' demethylation but majority of them focuses on single site level and does not allow for quantification of region methylation changes. The aim was to create an adaptive algorithm supporting detection of differentially methylated CpG sites and genomic regions specific for acute myeloid leukemia. Knowledge on AML methylation fingerprint helps in better understanding the epigenetics of leukemogenesis. Proposed algorithm is data driven and does not use predefined quantification thresholds. Gaussian mixture modeling supports classification of CpG sites to several levels of demethylation. p value integration allows for translation from single site demethylation to the demethylation of gene promoter and body regions. Methylation profiles of healthy controls and AML patients were examined (GEO:GSE63409). The differences in whole genome methylation profiles were observed. The methylation profile differs significantly among genomic regions. The lowest methylation level was observed for promoter regions, while sites from intergenic regions were by average higher methylated. The observed number of AML related down methylated sites has not substantially exceeded the expected number by chance. Intergenic regions were characterized by the highest percentage of AML up methylated sites. Methylation enhancement/diminution is the most frequent for intergenic region while methylation compensation (positive or negative) is specific for promoter regions. Functional analysis performed for AML down methylated or extreme high up methylated genes showed strong connection to the leukemic processes.