Uniparental disomy may be associated with microsatellite instability in acute myeloid leukemia (AML) with a normal karyotype

Abstract A large proportion of acute myeloid leukemia (AML) exhibits a normal karyotype in which the underlying pathomechanisms still have to be determined. Novel techniques like arrayCGH or single nucleotide polymorphism (SNP) chip analysis allow the identification and characterization of molecular rearrangements at the sub-megabase level. Recently, the application of genome-wide SNP array technology revealed frequent uniparental disomy (UPD) in approximately 20% of AML suggesting that UPD represents a nonrandom event in leukemogenesis. Uniparental disomy is acquired by somatic recombination and therefore not accessible by conventional cytogenetic methods or arrayCGH. In this study we analyzed DNA from AML patients with normal karyotype for the presence of LOH. SNP analysis was performed on the Mapping 100k GeneChip (Affymetrix, Santa Clara, CA). DNA was extracted from paired samples of 56 de novo AML patients with normal karyotype at diagnosis and in complete remission, respectively. Signal intensity data were analyzed by the GCOS GeneChip analysis software and statistical analysis of SNP call data was performed by the dChipSNP software. In addition, standard mutation screening of the genes encoding NPM1, FLT3, CEBPA, MLL and NRAS was performed in all cases. Using the 100k SNP array, a mean SNP call rate of 98.2% was reached, resulting in > 110,000 SNP genotype calls per sample. Signal intensity data analysis revealed submicroscopic chromosomal deletions resulting in hemizygosity in three patients. Patient 1 had a single 2 Mb deletion in chromosomal band 3p14.1, patient 2 had two small deletions affecting chromosome 12q23 and 12p13, the latter encompassing the ETV6 locus, and patient 3 had two small deletions within the long arm of chromosome 8. Besides these small chromosomal regions of copy number alterations, we found 4 large stretches of somatically acquired homozygosity without numeric alterations, affecting chromosome 6 (6p21 to 6 pter and 6q26 to 6 qter), chromosome 11 (11p12 to 11pter) and chromosome 13 (13q11 to 13qter). Noteworthy, in the case with uniparental disomy of chromosome 13, we could detect a homozygous FLT3-ITD mutation, supporting the findings that acquired isodisomy for chromosome 13 is common in AML, and associated with FLT3-ITD mutations (Griffiths et al., Leukemia, 2005). In summary, high resolution SNP assay technology in AML patients with normal karyotype allowed the identification of distinct chromosomal regions affected by UPD, supporting the postulated nonrandom mechanism of acquired mitotic recombination events in AML. Besides known chromosomal regions known to be affected by genomic aberrations in AML, we found additional submicroscopic chromosomal aberrations in cases with normal karyotype. Analysis of larger patient series will allow the identification of novel regions of interest harboring genes that might be involved in the pathogenesis of AML.

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A Genome-Wide Map of Acquired Uniparental Disomy in Acute Myeloid Leukemia.

Blood ◽

10.1182/blood.v110.11.996.996 ◽

2007 ◽

Vol 110 (11) ◽

pp. 996-996 ◽

Cited By ~ 1

Author(s):

Manu Gupta ◽

Manoj Raghavan ◽

Rosemary E. Gale ◽

Claude Chelala ◽

Christopher Allen ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Copy Number ◽

Myeloid Leukemia ◽

Uniparental Disomy ◽

Normal Karyotype ◽

Intermediate Risk ◽

Snp Arrays ◽

Risk Patients ◽

Control Set ◽

Acute Myeloid

Abstract The recent discovery of acquired uniparental disomy (aUPD) in acute myeloid leukemia (AML) has been linked to homozygosity for mutations in certain genes (Raghavan et al, Cancer Res. 2005, Fitzgibbon et al, Cancer Res. 2005). Although this phenomenon, which is undetectable by conventional cytogenetics, has been confirmed in subsequent small-scale studies, its extent and frequency remains uncertain. To determine the frequency and distribution of aUPD, DNA samples from 455 young adult AML patients entered in the UK Medical Research Council AML10 trial were analyzed using Mapping 10K 2.0 single nucleotide polymorphism (SNP) arrays (Affymetrix Inc.). Genomic DNA from blood samples of ten non-leukemic individuals was used as control to estimate the copy number values (control set I). We defined aUPD as 50 consecutive homozygous markers but allowed 2 heterozygous calls to accommodate contaminating normal tissue. Using this criterion a false positive rate of 3.3% was calculated from an available data of 90 independent controls (control set I). Overall, 120 regions of UPD were observed in 79 AML cases (17%), 87% of which involved at least one breakpoint, i.e. resulted from mitotic recombination, and 13% were whole chromosome aUPDs arising from chromosomal non-disjunction. They were the sole aberration, as detected by SNP arrays, in 61 samples (13%), and 84% of these had only a single region of aUPD. There was a non-random distribution across chromosomes; 13q (n=18 cases), 11p (n=8) and 11q (n=9) were the most frequently affected. Other chromosomes with regions of recurrent aUPD were 2p (n=7), 2q (n=6), 1p (n=5), 19q (n=4), 17q12–q21.2 (n=4), 21q (n=4), 9p (n=3), Xq (n=3), 6p (n=2), and 17p (n=2). Acquired UPDs were observed across all cytogenetic risk groups: in 25% of adverse risk patients, 11% of favorable risk, 19% of normal karyotype and 10% of the remaining intermediate risk patients. Samples with aUPD13q (5% of samples) belonged exclusively to the intermediate risk group. Chromosome 13 was the only chromosome to show recurrent whole chromosome aUPD. Fifteen samples with aUPD13q covered the region of the FLT3 gene at 13q12.2; all 15 had a FLT3-internal tandem duplication (ITD) and all cases with a high FLT3-ITD mutant level > 50% of total had 13q aUPD. Gains and losses were observed in 12% and 14% of the samples respectively. As expected, gains on chromosome 8 and losses on chromosomes 5 and 7 were common, confirming the general utility of this approach. No homozygous losses were observed. Comparison of arrays with cytogenetic analysis showed that additional information (aUPDs and/or copy number changes) was obtained in 23% of cases with a normal karyotype and 38% of cases without available cytogenetics. This study highlights the importance of aUPD in the development of AML and pinpoints regions that may contain novel mutational targets.

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Genome-Wide Micro-Deletions and Amplifications Acquired at Relapse in Acute Myeloid Leukemia.

Blood ◽

10.1182/blood.v114.22.166.166 ◽

2009 ◽

Vol 114 (22) ◽

pp. 166-166 ◽

Cited By ~ 2

Author(s):

Manoj Raghavan ◽

Manu Gupta ◽

Tracy Chaplin ◽

Sabah Khalid ◽

T. Andrew Lister ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Copy Number ◽

Myeloid Leukemia ◽

Conflicts Of Interest ◽

Snp Array ◽

Uniparental Disomy ◽

Normal Karyotype ◽

P Value ◽

Copy Number Changes ◽

Acute Myeloid

Abstract Abstract 166 Recurrence of acute myeloid leukemia AML has a poor prognosis with only 20% of adults surviving to 5 years. Therefore it is of importance to identify molecular changes that explain the pathogenesis of relapsed AML. Previous studies had not identified consistently acquired cytogenetic changes at relapse. Recently, acquired uniparental disomy due to mitotic recombination was described in 40% of relapsed AML (Raghavan et al 2008). Most of the events lead to homozygosity for FLT3 mutations. This study aimed to discover if there are further genetic abnormalities acquired at disease recurrence that cannot be identified by conventional cytogenetics, i.e. microdeletions or gains. Twenty-one presentation and relapse paired AML patient blood and marrow samples were stored with consent at St Bartholomew's Hospital, London. Eleven patient samples had a normal karyotype at diagnosis, two had favourable prognosis cytogenetics (inv(16) and t(8;21)) and others had varying numerical cytogenetic abnormalities and rearrangements associated with an intermediate prognosis. DNA from the samples was analysed by array based high-resolution single nucleotide polymorphism (SNP) genotyping (Affymetrix Human SNP array 6.0). Data was analysed using Partek Genome Browser (Partek, MO). In all cases, the leukemia infiltrate of the marrow or blood was greater than 60% and most cases were greater than 90% allowing accurate identification of DNA copy number changes. Abnormalities of a size that would be identified by cytogenetics were disregarded. Using segmentation analysis using a p-value less than 0.001, over 400 microdeletions and gains were detected that were acquired at relapse in the 21 pairs. Each of the copy number changes was less than 2 megabases in size. One AML sample with a normal karyotype at diagnosis and trisomy 8 and add(9)(q34) at relapse had not acquired any microdeletions or gains. In contrast, in other samples as many as 69 microdeletions/gains were detected. There was no correlation between increased complexity of the karyotype of the leukemia and the number of microdeletions/gains. Several of the acquired microdeletions/gains were in regions containing genes known to be involved in AML, including a deletion of 234Kb at 13q12.2 involving FLT3 and CDX2, and an acquired deletion at 21p11.2 of 150Kb involving exons encoding the runt domain of RUNX1. Another copy number gain was detected at the MLL locus, suggestive of partial tandem duplication. Other detected locations are in Table 1.Table 1Location by cytobandCopy number changeSize / KbP valueGene13q12.2Deletion23410−33FLT3, CDX221q22.12Deletion15010−13RUNX111q23.3Gain5.10.0099MLL11p15.4Gain830.00001NUP9817q21.31Deletion8.00.0007BRCA1The results indicate that recurrent AML may be associated with the deletion or gain of several genes involved in leukaemogenesis. Many other locations are involved throughout the genome, suggesting at least some of these are also involved in the clonal evolution of the leukaemia at recurrence. Further studies should identify novel genes from these regions involved in the pathogenesis of AML. Disclosures: No relevant conflicts of interest to declare.

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FLT3 Internal Tandem Duplication in Acute Myeloid Leukemia with Normal Karyotype

The Korean Journal of Hematology ◽

10.5045/kjh.2007.42.3.250 ◽

2007 ◽

Vol 42 (3) ◽

pp. 250 ◽

Cited By ~ 1

Author(s):

Sang-Ho Kim ◽

Yeo-Kyeoung Kim ◽

Il-Kwon Lee ◽

Deog-Yeon Jo ◽

Jong-Ho Won ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Tandem Duplication ◽

Normal Karyotype ◽

Internal Tandem Duplication ◽

Flt3 Internal Tandem Duplication ◽

Acute Myeloid

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Mutational profiling of acute myeloid leukemia with normal cytogenetics in Brazilian patients: the value of next-generation sequencing for genomic classification

Journal of Investigative Medicine ◽

10.1136/jim-2017-000566 ◽

2017 ◽

Vol 65 (8) ◽

pp. 1155-1158 ◽

Cited By ~ 6

Author(s):

Thiago Rodrigo de Noronha ◽

Miguel Mitne-Neto ◽

Maria de Lourdes Chauffaille

Keyword(s):

Acute Myeloid Leukemia ◽

Next Generation Sequencing ◽

Myeloid Leukemia ◽

Regulation Of Gene Expression ◽

Uniparental Disomy ◽

Driver Mutations ◽

Next Generation ◽

Normal Cytogenetics ◽

Generation Sequencing ◽

Acute Myeloid

Karyotype (KT) aberrations are important prognostic factors for acute myeloid leukemia (AML); however, around 50% of cases present normal results. Single nucleotide polymorphism array can detect chromosomal gains, losses or uniparental disomy that are invisible to KT, thus improving patients’ risk assessment. However, when both tests are normal, important driver mutations can be detected by the use of next-generation sequencing (NGS). Fourteen adult patients with AML with normal cytogenetics were investigated by NGS for 19 AML-related genes. Every patient presented at least one mutation:DNMT3Ain nine patients;IDH2in six;IDH1in three;NRASandNPM1in two; andTET2,ASXL1,PTPN11, andRUNX1in one patient. No mutations were found inFLT3,KIT,JAK2,CEBPA,GATA2,TP53,BRAF,CBL,KRAS,andWT1genes. Twelve patients (86%) had at least one mutation in genes related with DNA methylation (DNMT3A,IDH1,IDH2,andTET2), which is involved in regulation of gene expression and genomic stability. All patients could be reclassified based on genomic status and nine had their prognosis modified. In summary, NGS offers insights into the molecular pathogenesis and biology of cytogenetically normal AML in Brazilian patients, indicating that the prognosis could be further stratified by different mutation combinations. This study shows a different frequency of mutations in Brazilian population that should be confirmed.

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Normal karyotype acute myeloid leukemia with the CD7+ CD15+ CD34+ HLA-DR + immunophenotype is a clinically distinct entity with a favorable outcome

Annals of Hematology ◽

10.1007/s00277-014-2013-4 ◽

2014 ◽

Vol 93 (6) ◽

pp. 957-963 ◽

Cited By ~ 10

Author(s):

Noriyoshi Iriyama ◽

Norio Asou ◽

Yasushi Miyazaki ◽

Shunichiro Yamaguchi ◽

Shinya Sato ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Normal Karyotype ◽

Favorable Outcome ◽

Distinct Entity ◽

Hla Dr ◽

Acute Myeloid

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FLT3 mutations have no prognostic impact in elderly patients with acute myeloid leukemia and normal karyotype

American Journal of Hematology ◽

10.1002/ajh.21458 ◽

2009 ◽

Vol 84 (8) ◽

pp. 532-534 ◽

Cited By ~ 11

Author(s):

Felicetto Ferrara ◽

Clelia Criscuolo ◽

Cira Riccardi ◽

Tiziana Izzo ◽

Mariangela Pedata ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Elderly Patients ◽

Myeloid Leukemia ◽

Normal Karyotype ◽

Prognostic Impact ◽

Flt3 Mutations ◽

Acute Myeloid

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BCOR Mutations in Acute Myeloid Leukemia: Clonal Evolution and Prognosis

Blood ◽

10.1182/blood-2020-137127 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 4-4

Author(s):

Ashley Zhang ◽

Yuntao Liu ◽

Shuning Wei ◽

Benfa Gong ◽

Chunlin Zhou ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Risk Stratification ◽

Myeloid Leukemia ◽

Statistical Difference ◽

Tp53 Mutation ◽

Clonal Evolution ◽

Normal Karyotype ◽

Abnormal Karyotype ◽

Acute Myeloid

Background BCOR gene is a transcription repressor that may influence normal hematopoiesis and is associated with poor prognosis in acute myeloid leukemia (AML) with normal karyotype. However, due to the rare mutation frequency in AML (3.8%-5%), clinical characteristics and prognosis of AML patients with BCOR mutation including abnormal karyotype are still unknown. In addition, the clonal evolution of AML patients with BCOR mutation has not been fully investigated. Methods By means of next generation of sequencing, we performed sequencing of 114 genes related to hematological diseases including BCOR on 509 newly diagnosed AML patients (except for acute promyelocytic leukemia) from March 2017 to April 2019. The 2017 European Leukemia Net (ELN) genetic risk stratification was used to evaluate prognosis. Overall survival (OS) was defined as the time from diagnosis to death or last follow-up. Relapse-free survival (RFS) was measured from remission to relapse or death. Clonal evolution was investigated through analyzing bone marrow samples at diagnosis, complete remission (CR) and relapse from the same patient. Result Among 509 AML patients, we found BCOR mutations in 23 patients (4.5%). BCOR mutations were enriched in patients with mutations of RUNX1 (p = 0.008) and BCORL1 (p = 0.0003). Patients with BCOR mutation were more at adverse ELN risk category compared to patients without BCOR mutation (p = 0.007). Besides, there was a larger proportion of patients with normal karyotype in BCOR mutation group but it had not reached statistical difference (62.5% vs 45.5%, p = 0.064). The abnormal karyotype in patients with BCOR mutations included trisomy 8, t(9;11), inv(3), -7 and complex karyotype.There were no significant differences in age, sex, white blood cell count, hemoglobin or platelet count between the two groups. More patients died during induction (13.0% vs 3.5%, p = 0.56) and fewer patients achieved CR after 2 cycles of chemotherapy when patients had BCOR mutations (69.6% vs 82.5%, p = 0.115) but the difference had not reached statistical difference . Patients with BCOR mutations had inferior 2-year OS (52.1% vs 70.7%, p = 0.0094) and 2-year RFS (29.8% vs 61.1%, p = 0.0090). After adjustment for ELN risk stratification, BCOR mutation was still remain a poor prognostic factor. However, the adverse prognostic impact of BCOR mutation is overcome by hematopoietic stem cell transplantation (HSCT), in which there was no difference between BCOR mutation group and wild type group (p = 0.474) (Figure 1). Through analysis of paired bone marrow sample at diagnosis, remission and relapse, we revealed the clonal evolution that BCOR mutation was only detected at diagnosis sample as a subclone and diminished at CR and relapse while TP53 mutation was only detected at relapse with a variant allele frequency (VAF) of 25.5%. We also found BCOR mutation at another patient's diagnosis and relapse sample while TP53 mutation was detected at relapse with VAF of 11.8%. Conclusion BCOR is associated with RUNX1 mutation and higher ELN risk. AML patients with BCOR mutation including normal and abnormal karyotype conferred a worse impact on OS that can be overcome by HSCT. BCOR mutation is a subclone at diagnosis or relapse in some patients, in which TP53 mutation clone occurred at relapse. Disclosures No relevant conflicts of interest to declare.

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