Contributions
Abstract: PB2076
Type: Publication Only
Background
The dic(1;7)(q10;p10) is a centromeric-centromeric DNA juxtaposition, usually found in Myelodysplastic Syndromes (MDS) and Acute Myeloid Leukemia (AML) arising after chemo/radio-therapy. Due to the absence of genes within the epigenetically defined centromeric region, no specific molecular targets have been identified yet.
Aims
The aim of this study was to characterized biological features of MDS/AML with dic(1;7) by investigating their genetic and epigenetic landscape.
Methods
Three series of 5 cases each were selected for the study: 1) MDS/AML with dic(1;7); 2) t-MDS/AML; 3) non-neoplastic cytopenias. DNA and RNA were extracted from unsorted bone marrow cells. Genomic characterization of dic(1;7) included karyotype, Fluorescent In Situ Hybridization (FISH), Single Nucleotide Polymorphism Array (SNPa), Whole Exome Sequencing (WES), and RNA sequencing. Global epigenetic approach used Enhanced Reduced Representation Bisulfite Sequencing (mERRBS). All amplified libraries were sequenced on Illumina HiSeq2500 using manufacturer’s recommended protocol and were aligned against hg19.
Results
In all dic(1;7) cases breakpoints fell within α-sat DNA at chromosome 1 and 7, resulting in a 1q trisomy and 7q monosomy. SNPa excluded copy neutral loss of heterozygosity or cryptic copy number alterations, confirming dic(1;7) as a cytogenetically primary abnormality. WES did not identify a common mutational background in dic(1;7) series. Two patients harbored 4 somatic mutations never described in MDS/AML (EB1; GGPS1; CCDC8; PSMF1). Gene expression profile (FDR ≤ 0.1, log2FC ≥ I1I) provided us with a specific downregulation signature differentiating dic(1;7) from both controls [4860 differentially expressed genes (DEGs): 889 up and 3971 down] and t-MDS/AML [4317 DEGs: 482 up and 3835 down]. To characterize the biological differences, we analyzed DEGs within functional pathways, which identified downregulation of ATP-binding cassette transporters and upregulation of p53 signaling only in dicentrics (FDR ≤ 0.05). We next investigated the gene dosage effect of 1q trisomy and 7q monosomy. In keeping with 7q monosomy, 95% of DEGs on 7q were downregulated in dic(1;7). Surprisingly, the gene dosage effect of 1q trisomy contributed only partially, with more than 50% of 1q DEGs being downregulated in dicentrics. In exploring the basis for the dic(1;7)-related 1q downregulation, the DNA methylation profile identified hypermethylation at 1q in dicentrics in keeping with downregulation signature despite trisomy. Global epigenome analysis, which capture around 3.2M of CpGs, was able to separate dic(1;7) and t-MDS cases from each other and from controls. Moreover, mERRBS showed a specific dic(1;7) hypermethylation pattern in non-promoter regions, particularly at intronic enhancers. We used PET Module tool to identify putative enhancer-target genes, which hypo-expression gave account of around 30% of the downregulated signature. Furthermore, Hypergeometric Optimization of Motif EnRichment analysis showed that hypermethylated enhancers were specifically enriched for the Krüppel-like factor protein family of transcription factors.
Conclusion
In conclusion our results first showed that the cytogenetically unbalanced dic(1;7) in MDS/AML is the hallmark of a specific gene expression profile originated by epigenetic events, mostly enhancer hypermethylation. Biological features of dic(1;7), particularly its rare occurrence in complex karyotypes and the presence of p53 up-regulation, are separating MDS/AML with dic(1;7) from t-MDS/AML.
Session topic: 9. Myelodysplastic syndromes – Biology & Translational Research
Keyword(s): Chromosomal abnormality, DNA methylation, Gene expression profile, Myelodysplasia
Abstract: PB2076
Type: Publication Only
Background
The dic(1;7)(q10;p10) is a centromeric-centromeric DNA juxtaposition, usually found in Myelodysplastic Syndromes (MDS) and Acute Myeloid Leukemia (AML) arising after chemo/radio-therapy. Due to the absence of genes within the epigenetically defined centromeric region, no specific molecular targets have been identified yet.
Aims
The aim of this study was to characterized biological features of MDS/AML with dic(1;7) by investigating their genetic and epigenetic landscape.
Methods
Three series of 5 cases each were selected for the study: 1) MDS/AML with dic(1;7); 2) t-MDS/AML; 3) non-neoplastic cytopenias. DNA and RNA were extracted from unsorted bone marrow cells. Genomic characterization of dic(1;7) included karyotype, Fluorescent In Situ Hybridization (FISH), Single Nucleotide Polymorphism Array (SNPa), Whole Exome Sequencing (WES), and RNA sequencing. Global epigenetic approach used Enhanced Reduced Representation Bisulfite Sequencing (mERRBS). All amplified libraries were sequenced on Illumina HiSeq2500 using manufacturer’s recommended protocol and were aligned against hg19.
Results
In all dic(1;7) cases breakpoints fell within α-sat DNA at chromosome 1 and 7, resulting in a 1q trisomy and 7q monosomy. SNPa excluded copy neutral loss of heterozygosity or cryptic copy number alterations, confirming dic(1;7) as a cytogenetically primary abnormality. WES did not identify a common mutational background in dic(1;7) series. Two patients harbored 4 somatic mutations never described in MDS/AML (EB1; GGPS1; CCDC8; PSMF1). Gene expression profile (FDR ≤ 0.1, log2FC ≥ I1I) provided us with a specific downregulation signature differentiating dic(1;7) from both controls [4860 differentially expressed genes (DEGs): 889 up and 3971 down] and t-MDS/AML [4317 DEGs: 482 up and 3835 down]. To characterize the biological differences, we analyzed DEGs within functional pathways, which identified downregulation of ATP-binding cassette transporters and upregulation of p53 signaling only in dicentrics (FDR ≤ 0.05). We next investigated the gene dosage effect of 1q trisomy and 7q monosomy. In keeping with 7q monosomy, 95% of DEGs on 7q were downregulated in dic(1;7). Surprisingly, the gene dosage effect of 1q trisomy contributed only partially, with more than 50% of 1q DEGs being downregulated in dicentrics. In exploring the basis for the dic(1;7)-related 1q downregulation, the DNA methylation profile identified hypermethylation at 1q in dicentrics in keeping with downregulation signature despite trisomy. Global epigenome analysis, which capture around 3.2M of CpGs, was able to separate dic(1;7) and t-MDS cases from each other and from controls. Moreover, mERRBS showed a specific dic(1;7) hypermethylation pattern in non-promoter regions, particularly at intronic enhancers. We used PET Module tool to identify putative enhancer-target genes, which hypo-expression gave account of around 30% of the downregulated signature. Furthermore, Hypergeometric Optimization of Motif EnRichment analysis showed that hypermethylated enhancers were specifically enriched for the Krüppel-like factor protein family of transcription factors.
Conclusion
In conclusion our results first showed that the cytogenetically unbalanced dic(1;7) in MDS/AML is the hallmark of a specific gene expression profile originated by epigenetic events, mostly enhancer hypermethylation. Biological features of dic(1;7), particularly its rare occurrence in complex karyotypes and the presence of p53 up-regulation, are separating MDS/AML with dic(1;7) from t-MDS/AML.
Session topic: 9. Myelodysplastic syndromes – Biology & Translational Research
Keyword(s): Chromosomal abnormality, DNA methylation, Gene expression profile, Myelodysplasia