Multiplex ligation-dependent probe amplification for detection of chromosomal abnormalities in myelodysplastic syndrome and acute myeloid leukemia

Abstract We have studied 20 children with therapy-related myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) who were 3 months to 16 years old at diagnosis of their primary neoplasm and 1 to 24 years old at diagnosis of their secondary neoplasm. The median interval from initial treatment for the first malignancy to diagnosis of therapy- related MDS or AML was 46 months (range, 12 to 116 months). Twelve patients had chromosomal abnormalities resulting in loss of material from the long arm of chromosomes 5 and/or 7, three patients had abnormalities of chromosome 11 band q23, one patient had both classes of abnormalities, three patients had other abnormalities, and one patient had a normal karyotype. Ten of 12 patients with chromosome 5 and/or 7 abnormalities had been exposed to an alkylating agent, and two of three patients with 11q23 abnormalities had been exposed to an epipodophyllotoxin. The patient with both classes of abnormalities had been exposed to both types of therapy. We conclude that abnormalities of chromosomes 5 and/or 7 are common in children with therapy-related MDS or AML. The proposed relationships between exposure to alkylating agents and abnormalities of chromosomes 5 and/or 7 and between exposure to epipodophyllotoxins and abnormalities of 11q23 are supported in this pediatric series.

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Mesenchymal stromal cells of myelodysplastic syndrome and acute myeloid leukemia patients have distinct genetic abnormalities compared with leukemic blasts

Blood ◽

10.1182/blood-2011-03-343467 ◽

2011 ◽

Vol 118 (20) ◽

pp. 5583-5592 ◽

Cited By ~ 103

Author(s):

Olga Blau ◽

Claudia Dorothea Baldus ◽

Wolf-Karsten Hofmann ◽

Gundula Thiel ◽

Florian Nolte ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myelodysplastic Syndrome ◽

Mesenchymal Stromal Cells ◽

Chromosomal Aberrations ◽

Stromal Cells ◽

Myeloid Leukemia ◽

Chromosomal Abnormalities ◽

Genetic Alterations ◽

Genetic Abnormalities ◽

Acute Myeloid

Abstract Mesenchymal stromal cells (MSCs) are an essential cell type of the hematopoietic microenvironment. Concerns have been raised about the possibility that MSCs undergo malignant transformation. Several studies, including one from our own group, have shown the presence of cytogenetic abnormalities in MSCs from leukemia patients. The aim of the present study was to compare genetic aberrations in hematopoietic cells (HCs) and MSCs of myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) patients. Cytogenetic aberrations were detected in HCs from 25 of 51 AML patients (49%) and 16 of 43 MDS patients (37%). Mutations of the FLT3 and NPM1 genes were detected in leukemic blasts in 12 (23%) and 8 (16%) AML patients, respectively. Chromosomal aberrations in MSCs were detected in 15 of 94 MDS/AML patients (16%). No chromosomal abnormalities were identified in MSCs of 36 healthy subjects. We demonstrate herein that MSCs have distinct genetic abnormalities compared with leukemic blasts. We also analyzed the main characteristics of patients with MSCs carrying chromosomal aberrations. In view of these data, the genetic alterations in MSCs may constitute a particular mechanism of leukemogenesis.

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Therapy-related myelodysplastic syndrome and acute myeloid leukemia in children: correlation between chromosomal abnormalities and prior therapy

Blood ◽

10.1182/blood.v78.11.2982.bloodjournal78112982 ◽

1991 ◽

Vol 78 (11) ◽

pp. 2982-2988 ◽

Cited By ~ 2

Author(s):

CM Rubin ◽

DC Arthur ◽

WG Woods ◽

BJ Lange ◽

PC Nowell ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myelodysplastic Syndrome ◽

Myeloid Leukemia ◽

Chromosomal Abnormalities ◽

Alkylating Agents ◽

Normal Karyotype ◽

Chromosome 11 ◽

Prior Therapy ◽

11Q23 Abnormalities ◽

Acute Myeloid

We have studied 20 children with therapy-related myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML) who were 3 months to 16 years old at diagnosis of their primary neoplasm and 1 to 24 years old at diagnosis of their secondary neoplasm. The median interval from initial treatment for the first malignancy to diagnosis of therapy- related MDS or AML was 46 months (range, 12 to 116 months). Twelve patients had chromosomal abnormalities resulting in loss of material from the long arm of chromosomes 5 and/or 7, three patients had abnormalities of chromosome 11 band q23, one patient had both classes of abnormalities, three patients had other abnormalities, and one patient had a normal karyotype. Ten of 12 patients with chromosome 5 and/or 7 abnormalities had been exposed to an alkylating agent, and two of three patients with 11q23 abnormalities had been exposed to an epipodophyllotoxin. The patient with both classes of abnormalities had been exposed to both types of therapy. We conclude that abnormalities of chromosomes 5 and/or 7 are common in children with therapy-related MDS or AML. The proposed relationships between exposure to alkylating agents and abnormalities of chromosomes 5 and/or 7 and between exposure to epipodophyllotoxins and abnormalities of 11q23 are supported in this pediatric series.

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Molecular Characterization of Chromosomal Abnormalities In Myelodysplastic Syndrome and Acute Myeloid Leukemia: Validation of An MLPA Protocol and Analysis Method for Use In a Diagnostic Setting

Blood ◽

10.1182/blood.v116.21.4849.4849 ◽

2010 ◽

Vol 116 (21) ◽

pp. 4849-4849

Author(s):

Amber C. Donahue ◽

Adam Abdool ◽

Jay G Wohlgemuth ◽

Chen-Hsiung Yeh

Keyword(s):

Acute Myeloid Leukemia ◽

Data Analysis ◽

Myelodysplastic Syndrome ◽

Copy Number ◽

Myeloid Leukemia ◽

Chromosomal Abnormalities ◽

Analysis Method ◽

Normal Range ◽

Copy Number Changes ◽

Acute Myeloid

Abstract Abstract 4849 Introduction: Current diagnostic screening strategies for copy number variations (CNVs) in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) include fluorescence in situ hybridization (FISH) or karyotyping, both of which are time-consuming, costly, laborious, and lacking in resolution. Multiplex ligation-dependent probe amplification (MLPA) can be used to detect copy number changes in multiple loci simultaneously in a single PCR reaction, and boasts a resolution down to single exons. To adapt MLPA for use in routine clinical diagnostics, we have developed and validated a protocol for automatic data analysis and interpretation of common chromosomal abnormalities in MDS/AML. Patients and Methods: The study used a training set of 45 healthy subjects to establish a normal reference range for each individual probe. Using these ranges we built an automated Excel spreadsheet-based analysis system, which included multiple quality checks, and flagged samples failing these quality controls. Each probe was given a call of “no mutation detected,” “deletion,” or “gain,” based on whether the normalized ratio fell within or outside of the empirically-determined normal range for that probe. We then analyzed over 100 leukemia cases tested by FISH, including both suspected myeloid leukemia samples and suspected chronic lymphocytic leukemia (CLL) samples. Documented chromosomal abnormalities in CLL include 11q-, 17p- (loss of TP53), and trisomy 12, all of which had the potential to be detected by the probes in the MDS MLPA probemix. The greater prevalence of CLL and its associated CNVs provided additional positive controls for the validation of the MDS MLPA probemix and our analysis method. Results: The empirically-determined normal ranges demonstrated that some probes varied widely (3 standard deviation [3SD] normal range of 0.46–1.54), while others were extremely reliable (3SD normal range of 0.84–1.16). The MLPA assay demonstrated excellent overall accuracy (>90%) and specificity (>93%) for both suspected myeloid and CLL samples when compared to FISH. The sensitivity of the MLPA assay is somewhat lower than that of FISH, requiring a probe-dependent 20–40% positivity for a given CNV to be detected. However in several cases, the MDS MLPA assay was able to detect additional lesions too small to be seen by FISH. Conclusions: For MLPA, the total process-to-report time, including data analysis, is 2–3 days, versus the 7–10 days required for FISH analysis. In addition, the MLPA assay is substantially cheaper and considerably less labor-intensive than FISH. Our improved MLPA assay protocol and analysis method provides a clinically robust, multiplexed, high-throughput, high-resolution, and low-cost solution for detection of copy number changes in MDS/AML, and can therefore be used as a first-line screening test in a clinical laboratory. Disclosures: Donahue: Quest Diagnostics Inc.: Employment. Abdool: Quest Diagnostics Inc.: Employment. Wohlgemuth: Quest Diagnostics Inc.: Employment. Yeh: Quest Diagnostics Inc.: Employment.

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