11 research outputs found
Frequent deletions of JARID2 in leukemic transformation of chronic myeloid malignancies
Chronic myeloproliferative neoplasms (MPN) and myelodysplastic syndromes (MDS) have an inherent tendency to progress to acute myeloid leukemia (AML). Using high-resolution SNP microarrays, we studied a total of 517 MPN and MDS patients in different disease stages, including 77 AML cases with previous history of MPN (N = 46) or MDS (N = 31). Frequent chromosomal deletions of variable sizes were detected, allowing the mapping of putative tumor suppressor genes involved in the leukemic transformation process. We detected frequent deletions on the short arm of chromosome 6 (del6p). The common deleted region on 6p mapped to a 1.1-Mb region and contained only the JARID2 genemember of the polycomb repressive complex 2 (PRC2). When we compared the frequency of del6p between chronic and leukemic phase, we observed a strong association of del6p with leukemic transformation (P = 0.0033). Subsequently, analysis of deletion profiles of other PRC2 members revealed frequent losses of genes such as EZH2, AEBP2, and SUZ12; however, the deletions targeting these genes were large. We also identified two patients with homozygous losses of JARID2 and AEBP2. We observed frequent codeletion of AEBP2 and ETV6, and similarly, SUZ12 and NF1. Using next generation exome sequencing of 40 patients, we identified only one somatic mutation in the PRC2 complex member SUZ12. As the frequency of point mutations in PRC2 members was found to be low, deletions were the main type of lesions targeting PRC2 complex members. Our study suggests an essential role of the PRC2 complex in the leukemic transformation of chronic myeloid disorders. Am. J. Hematol. 2012
Complex Patterns of Chromosome 11 Aberrations in Myeloid Malignancies Target CBL, MLL, DDB1 and LMO2
Exome sequencing of primary tumors identifies complex somatic mutation patterns. Assignment of relevance of individual somatic mutations is difficult and poses the next challenge for interpretation of next generation sequencing data. Here we present an approach how exome sequencing in combination with SNP microarray data may identify targets of chromosomal aberrations in myeloid malignancies. The rationale of this approach is that hotspots of chromosomal aberrations might also harbor point mutations in the target genes of deletions, gains or uniparental disomies (UPDs). Chromosome 11 is a frequent target of lesions in myeloid malignancies. Therefore, we studied chromosome 11 in a total of 813 samples from 773 individual patients with different myeloid malignancies by SNP microarrays and complemented the data with exome sequencing in selected cases exhibiting chromosome 11 defects. We found gains, losses and UPDs of chromosome 11 in 52 of the 813 samples (6.4%). Chromosome 11q UPDs frequently associated with mutations of CBL. In one patient the 11qUPD amplified somatic mutations in both CBL and the DNA repair gene DDB1. A duplication within MLL exon 3 was detected in another patient with 11qUPD. We identified several common deleted regions (CDR) on chromosome 11. One of the CDRs associated with de novo acute myeloid leukemia (P=0.013). One patient with a deletion at the LMO2 locus harbored an additional point mutation on the other allele indicating that LMO2 might be a tumor suppressor frequently targeted by 11p deletions. Our chromosome-centered analysis indicates that chromosome 11 contains a number of tumor suppressor genes and that the role of this chromosome in myeloid malignancies is more complex than previously recognized
Clinical significance of genetic aberrations in secondary acute myeloid leukemia
The study aimed to identify genetic lesions associated with secondary acute myeloid leukemia (sAML) in comparison with AML arising de novo (dnAML) and assess their impact on patients' overall survival (OS). High-resolution genotyping and loss of heterozygosity mapping was performed on DNA samples from 86 sAML and 117 dnAML patients, using Affymetrix Genome-Wide Human SNP 6.0 arrays. Genes TP53, RUNX1, CBL, IDH1/2, NRAS, NPM1, and FLT3 were analyzed for mutations in all patients. We identified 36 recurrent cytogenetic aberrations (more than five events). Mutations in TP53, 9pUPD, and del7q (targeting CUX1 locus) were significantly associated with sAML, while NPM1 and FLT3 mutations associated with dnAML. Patients with sAML carrying TP53 mutations demonstrated lower 1-year OS rate than those with wild-type TP53 (14.3% +/- 9.4% vs. 35.4% +/- 7.2%; P = 0.002), while complex karyotype, del7q (CUX1) and del7p (IKZF1) showed no significant effect on OS. Multivariate analysis confirmed that mutant TP53 was the only independent adverse prognostic factor for OS in sAML (hazard ratio 2.67; 95% CI: 1.335.37; P = 0.006). Patients with dnAML and complex karyotype carried sAML-associated defects (TP53 defects in 54.5%, deletions targeting FOXP1 and ETV6 loci in 45.4% of the cases). We identified several co-occurring lesions associated with either sAML or dnAML diagnosis. Our data suggest that distinct genetic lesions drive leukemogenesis in sAML. High karyotype complexity of sAML patients does not influence OS. Somatic mutations in TP53 are the only independent adverse prognostic factor in sAML. Patients with dnAML and complex karyotype show genetic features associated with sAML and myeloproliferative neoplasms. Am. J. Hematol., 2012
Complex Patterns of Chromosome 11 Aberrations in Myeloid Malignancies Target <i>CBL, MLL, DDB1</i> and <i>LMO2</i>
<div><p>Exome sequencing of primary tumors identifies complex somatic mutation patterns. Assignment of relevance of individual somatic mutations is difficult and poses the next challenge for interpretation of next generation sequencing data. Here we present an approach how exome sequencing in combination with SNP microarray data may identify targets of chromosomal aberrations in myeloid malignancies. The rationale of this approach is that hotspots of chromosomal aberrations might also harbor point mutations in the target genes of deletions, gains or uniparental disomies (UPDs). Chromosome 11 is a frequent target of lesions in myeloid malignancies. Therefore, we studied chromosome 11 in a total of 813 samples from 773 individual patients with different myeloid malignancies by SNP microarrays and complemented the data with exome sequencing in selected cases exhibiting chromosome 11 defects. We found gains, losses and UPDs of chromosome 11 in 52 of the 813 samples (6.4%). Chromosome 11q UPDs frequently associated with mutations of <i>CBL</i>. In one patient the 11qUPD amplified somatic mutations in both <i>CBL</i> and the DNA repair gene <i>DDB1</i>. A duplication within <i>MLL</i> exon 3 was detected in another patient with 11qUPD. We identified several common deleted regions (CDR) on chromosome 11. One of the CDRs associated with <i>de novo</i> acute myeloid leukemia (P=0.013). One patient with a deletion at the <i>LMO2</i> locus harbored an additional point mutation on the other allele indicating that <i>LMO2</i> might be a tumor suppressor frequently targeted by 11p deletions. Our chromosome-centered analysis indicates that chromosome 11 contains a number of tumor suppressor genes and that the role of this chromosome in myeloid malignancies is more complex than previously recognized.</p> </div
Summary of chromosome 11 aberrations.
<p>Large chromosomal aberrations are indicated with colored bars around the ideogram of chromosome 11. Green – gains; red – deletions; blue – uniparental disomies. The position of the bars relative to the chromosome ideogram indicates the position and size of the aberration. For the two patients of whom two samples were analyzed (UPN 23 and UPN 42 – see Table S1) recurrent aberrations are depicted only once. The positions of <i>CBL, MLL, EED, SF1</i>, <i>DDB1</i> and <i>LMO2</i> are indicated by vertical lines. Mutations in these genes are depicted by orange circles along these lines. Common deleted regions are indicated at the bottom of Figure 2 listing the genes they cover. The ideogram depicts G-banding pattern at ~850-band resolution level.</p
Mutations detected in <i>DDB1, MLL</i> and <i>LMO2</i>.
<p><b>A</b>: Sample 36 harbored an 11q UPD as indicated by the blue bar below the chromosome 11 ideogram. We found two somatic mutations in <i>DDB1</i> and <i>CBL</i>. As can be seen in the Sanger sequencing traces, both mutations are homozygous due to amplification by the UPD. <b>B</b>: In sample 50 a tandem duplication in <i>MLL</i> exon 3 was detected. The top graph shows whole exome coverage data across <i>MLL</i> exon 3. The data is plotted as the log<sub>2</sub> ratio of the normalized exome sequencing coverage in the patient sample divided by the median normalized coverage of 8 independent control samples at each genomic position (X-axis). The position of the duplication is indicated by the red bar. Sanger sequencing confirmed an in-frame tandem duplication of 171 amino acids as shown at the bottom. <b>C</b>: A common deleted region on chromosome 11p targets <i>LMO2</i>. All deletions in the analyzed cohort that span the <i>LMO2</i> locus are depicted next to the chromosome 11 ideogram. Red bars indicate deletions, green bars indicate gains. In sample 42, which harbored a deletion spanning the LMO2 locus, we also detected a point mutation in <i>LMO2</i>. The middle section shows a signal intensity plot measuring copy number from Affymetrix microarrays. The plot depicts signal intensity (log<sub>2</sub> scale) differences between the patient and a healthy control pool for each probe (as implemented in the Affymetrix Genotyping Console software). The deletion in sample 42 can be seen as the deviation from 0 for all probes in the deleted genomic region (X-axis). The point mutation in <i>LMO2</i> as identified by Sanger sequencing is depicted at the bottom of panel C. <b>A</b>, <b>B</b> and <b>C</b>: Depicted are the genomic (letters) as well as the respective amino acid (box chains) sequences. Numbers above the boxes indicate amino acid positions in the proteins. Amino acids substituted in the patient samples are indicated by red boxes. The red circle indicates a splice site mutation. Reference and mutant sequences are shown. The arrows indicate the site of mutations below the Sanger sequencing traces.</p
Mutational patterns in <i>CBL</i>.
<p><b>A</b>: Sample 45 had a 6 bp tandem duplication in CBL leading to the insertion of the amino acids valine (V) and aspartic acid (D) after position 390. <b>B</b>: One sample identified in a cohort screen for mutations in <i>CBL</i> exons 8 and 9 carried two mutations, one in exon 8 (W408C) and a second one in intron 8 at the splice acceptor site (G to A). PCR subcloning and analysis of colony DNA revealed that the two mutations are on different alleles. Depicted are two representative colonies. Colony 43 has the mutation in exon 8 but not in intron 8 whereas colony 17 shows the opposite case. <b>A</b>, <b>B</b>: Depicted are the genomic (letters) as well as the respective amino acid (box chains) sequences. Numbers indicate amino acid positions in the Cbl protein. Amino acids, which are substituted due to mutations are in red boxes. The splice site alteration is a red circle. Black arrows indicate the positions of the mutations below the Sanger sequencing traces. <b>C</b>: Overview of <i>CBL</i> mutagenesis in MPN. Different genetic mechanisms are involved in increasing mutant gene dosage of <i>CBL</i>. Each panel shows schematically the two parental copies of chromosomes 11 (blue and yellow) and the <i>CBL</i> gene (white rectangles). Mutations are indicated with asterisks. From left to right: heterozygous mutation in <i>CBL</i>; uniparental disomy introduces homozygous <i>CBL</i> mutations; gain of a part of chromosome 11q leads to a duplication of the <i>CBL</i> mutation while one wild type allele is still present; compound heterozygosity established by two different mutations on the different alleles of the <i>CBL</i> gene in one cell. In addition, the loss of a part of chromosome 11q deleting one <i>CBL</i> allele and leaving the other allele unaffected (wild type <i>CBL</i>) is likely to introduce phenotypes due to haploinsufficiency.</p
Cohort distribution.
<p><b>A</b>: Distribution of the 813 samples analyzed by Affymetrix microarrays according to diagnosis. <b>B</b>: Fraction of samples that harbor chromosome 11 aberrations (black bars) for each disease entity in percent. The P-value indicates an association of chromosome 11 aberrations with disease progression in MPN. MPN, myeloproliferative neoplasm; CML, chronic myeloid leukemia; AML, acute myeloid leukemia; MDS, myelodysplastic syndrome.</p