New complex t(2;11;17)(p21;q23;q11), a variant form of t(2;11), associated with del(5)(q23q32) in myelodysplastic syndrome-derived acute myeloblastic leukemia

The t(2;11)(p21;q23) is a rare recurrent aberration observed in myelodysplastic syndrome (MDS) and acute myeloblastic leukemia (AML). It has been suggested that t(2;11) is specifically associated with a deletion of the long arm of chromosome 5 (5q). A 63-year-old man was initially diagnosed as AML with del(5)(q23q32) as a sole abnormality. At relapse, t(2;11;17)(p21;q23;q11) in association with del(5q) appeared in 14 of 20 cells by G-banding. Spectral karyotyping confirmed three derivative chromosomes, der(11)t(2;11), der(17)t(11;17), and der(2)t(2;17). Fluorescence in situ hybridization analysis with a probe for MLL demonstrated that the breakpoint at 11q23 was telomeric to the MLL gene. Nine of 10 reported cases with t(2;11) and del(5q) had MDS including 5q- syndrome and four of them evolved to AML, as observed in the present case. Our results indicated that t(2;11;17) was a secondary genetic change, which appeared during disease progression after del(5q) was observed. Furthermore, considering another reported case, the MLL gene seems to be not involved in the pathogenesis of MDS/AML with t(2;11) and del(5q).     

Isochromosome of a deleted 20q: a rare but recurrent chromosome abnormality in myelodysplastic syndromes

Interstitial deletion of the long arm of chromosome 20, as the sole abnormality, is commonly observed in myeloid malignancies, including myeloproliferative disorder, myelodysplastic syndrome, and acute myeloid leukemia. The breakpoints of the deletion are typically located in the region 20q11.2 approximately q13.3, although smaller deletions within this region have also been reported. We present here 4 patients with myelodysplastic syndrome with an isochromosome of the deleted long arm of chromosome 20: ider(20)(q10)del(20)(q11q13). Fluorescence in situ hybridization studies were performed on the bone marrow samples from these patients to prove the identity of this unusual chromosome abnormality.     

Distinct sequences on 11q13.5 and 11q23-24 are frequently coamplified with MLL in complexly organized 11q amplicons in AML/MDS patients

Amplification within chromosome arm 11q involving the mixed-lineage leukemia gene (MLL) locus is a rare but recurrent aberration in acute myeloid leukemia and myelodysplastic syndrome (AML/MDS). We and others have observed that 11q amplifications in most AML/MDS cases have not been restricted to the chromosomal region surrounding the MLL gene. Therefore, we implemented a strategy to characterize comprehensively 11q amplicons in a series of 13 AML/MDS patients with MLL amplification. Analysis of 4 of the 13 cases by restriction landmark genomic scanning in combination with virtual genome scan and by matrix-based comparative genomic hybridization demonstrated that the 11q amplicon in these four cases consisted of at least three discontinuous sequences derived from different regions of the long arm of chromosome 11. We defined a maximally 700-kb sequence around the MLL gene that was amplified in all cases. Apart from the core MLL amplicon, we detected two additional 11q regions that were coamplified. Using fluorescence in situ hybridization (FISH) analysis, we demonstrated that sequences in 11q13.5 and 11q23-24 were amplified in 8 of 13 and 10 of 12 AML/MDS cases, respectively. Both regions harbor a number of potentially oncogenic genes. In all 13 cases, either one or both of these regions were coamplified with the MLL amplicon. Thus, we demonstrated that 11q amplicons in AML/MDS patients display a complex organization and have provided evidence for coamplification of two additional regions on the long arm of chromosome 11 that may harbor candidate target genes.     

Molecular definition of chromosome arm 5q deletion end points and detection of hidden aberrations in patients with myelodysplastic syndromes and isolated del(5q) using oligonucleotide array CGH

Isolated deletions of the long arm of chromosome 5, del(5q), are observed in 10% of myelodysplastic syndromes (MDS) and are associated with a more favorable prognosis, although the clinical course varies considerably. If one or more additional chromosomal aberrations are present, this correlates with a significantly shorter overall survival. To assess the frequency of hidden abnormalities in cases with an isolated cytogenetic del(5q), we have performed a genome wide high resolution 44 K 60mer oligonucleotide array comparative genomic hybridization (aCGH) study using DNA from bone marrow cells of 12 MDS and one AML patient. In one case a single additional hidden 5.6 Mb deletion of 13q14 and in another case multiple larger aberrations involving many chromosomes were found. Fluorescence in situ hybridization demonstrated that aberrations present in 35% of the bone marrow cells can be detected by aCGH. Furthermore with oligonucleotide aCGH the deletion end points in 5q were mapped precisely, revealing a cluster of proximal breakpoints in band q14.3 (n = 8) and a distal cluster between bands q33.2 and q34 (n = 11). This study shows the high resolution of oligonucleotide CGH arrays for precisely mapping genomic alterations and for refinement of deletion end points. In addition, the high sensitivity of this method enables the study of whole bone marrow cells from MDS patients, a disease with a low blast count.     

A der(13)t(7;13)(p13;q14) with monoallelic loss of RB1 and D13S319 in myelodysplastic syndrome

Deletions or translocations of chromosome band 13q14, the locus of the retinoblastoma gene (RB1), have been observed in a variety of hematological malignancies including myelodysplastic syndrome (MDS). We describe here a novel unbalanced translocation der(13)t(7;13)(p13;q14) involving 13q14 in a patient with MDS. A 66-year-old woman was diagnosed as having MDS, refractory anemia with excess of blasts (RAEB-1) because of 7.4% blasts and trilineage dysplasia in the bone marrow cells. G-banding and spectral karyotyping analyses showed complex karyotypes as follows: 46,XX,der(6)t(6;7)(q11;?),der(7)del(7)(?p13)t(6;7)(q?;q11)t(6;13)(q?;q?),der(13)t(7;13)(p13;q14). Fluorescence in situ hybridization (FISH) analyses demonstrated that one allele of the RB1 gene and the microsatellite locus D13S319, located at 13q14 and telomeric to the RB1 gene, was deleted. Considering other reported cases, our results indicate that submicroscopic deletions accompanying 13q14 translocations are recurrent cytogenetic aberrations in MDS. The RB1 gene or another tumor suppressor gene in the vicinity of D13S319, or both, may be involved in the pathogenesis of MDS with 13q14 translocations by monoallelic deletion.     

A comparison of two contrasting recurrent isochromosomes 20 found in myelodysplastic syndromes suggests that retention of proximal 20q is a significant factor in myeloid malignancies

We compare two different isochromosomes of chromosome 20 in myelodysplastic syndromes (MDS): an isochromosome of the short arm of chromosome 20, idic(20)(q11), and an isochromosome of the long arm of a deleted chromosome 20, ider(20)(q10)del(20)(q11.2). The isochromosomes are of contrasting morphology, because opposite arms are duplicated, but they both show loss of the critical region at 20q12, as well as retention and duplication of the centromere and proximal long arm (20q11). We speculate that a region of proximal 20q is preferentially retained during deletions of the critical region in MDS and acute myeloid leukemia.     

21q− in primary thrombocythemia

In 11 of 18 patients with primary thrombocythemia (PT), karyotype analysis of marrow cells after Giemsa chromosome banding revealed a previously unrecognized specific abnormality, consisting in a deletion of the long arm of a chromosome #21 (21q?), del 21(q21). This deletion was observed in 11.1 to 47.6% of marrow cell metaphases. In 8 patients, 21q? cells were detected prior to any therapy. In 3 other patients, 21q? cells were found during relapse after busulfan therapy. In one patient, a translocation of the deleted material on the long arm of a #11, t(11;21)(q25;q21), was observed.     

Novel chromosome 16 abnormality--der(16)del(16) (q13)t(16;21)(p11.2;q22)--associated with acute myeloid leukemia.

Inversion of chromosome 16 is a common feature of acute myeloid leukemia (AML) M4, while t(16;21), although also associated with AML, appears to be a separate entity. We present a patient with myelodysplastic syndrome (MDS) who transformed to AML-M1. The karyotype was normal at diagnosis; at 15 months, hematological evidence of transformation was present, and repeat cytogenetics showed a novel rearrangement of one chromosome 16. Two breaks had occurred; one in the short arm at 16p11, with translocation of the segment distal to this onto chromosome 21q, and the other in the long arm at 16q22 with subsequent deletion of the segment from 16q22-->qter. Fluorescence in situ hybridization (FISH) confirmed the abnormalities detected by cytogenetics and excluded involvement of the AML1 gene on 21q22. While the 16q22 breakpoint was at the usual site for the inv(16), the 16p11 was not. The patient is more characteristic of t(16;21) than inv(16), and adds to the spectrum of chromosome 16 abnormalities in AML.     

Shwachman syndrome as mutator phenotype responsible for myeloid dysplasia/neoplasia through karyotype instability and chromosomes 7 and 20 anomalies

An investigation of 14 patients with Shwachman syndrome (SS), using standard and molecular cytogenetic methods and molecular genetic techniques, showed that (1) the i(7)(q10) is not, or not always, an isochromosome but may arise from a more complex mechanism, retaining part of the short arm; (2) the i(7)(q10) has no preferential parental origin; (3) clonal chromosome changes, such as chromosome 7 anomalies and del(20)(q11), may be present in the bone marrow (BM) for a long time without progressing to myelodysplastic syndrome (MDS)/acute myeloid leukemia (AML); (4) the del(20)(q11) involves the minimal region of deletion typical of MDS/AML; (5) the rate of chromosome breaks is not significantly higher than in controls, from which it is concluded that SS should not be considered a breakage syndrome; (6) a specific kind of karyotype instability is present in SS, with chromosome changes possibly found in single cells or small clones, often affecting chromosomes 7 and 20, in the BM. Hence, we have confirmed our previous hypothesis that the SS mutation itself implies a mutator effect that is responsible for MDS/AML through these specific chromosome anomalies. This conclusion supports the practice of including cytogenetic monitoring in the follow-up of SS patients.     

FISH studies identify the i(20q-) anomaly as a der(20)del(20)(q11q13)idic(20)(p11)

Fluorescence in situ hybridization (FISH) analyses were performed on six of seven patients who had been reported in 2004 to have an i(20q-) anomaly expressed as ider(20)(q10)del(20)(q11q13). The i(20q-) was investigated with a series of probes: a centromere-specific probe for chromosome 20, two paint probes for 20p and 20q, and a panel of locus-specific probes prepared from BAC/PAC clones mapped to 20p. The results showed that: (1) i(20q-) was a dicentric chromosome; (2) both of its arms comprised a deleted 20q and a small part of 20p near the centromere of chromosome 20; and (3) the breakpoints and reunion sites of i(20q-) differed, residing in the region 20p11.21-20p11.22 delineated by BAC/PAC clones RP11-96L6 and RP13-401N8. Thus, i(20q-) could be more precisely described as a der(20)del(20)(q11q13)idic(20)(p11).     

Deletion of chromosome 15 represents a rare but recurrent chromosomal abnormality in myelocytic malignancies

We report on three cases with myelocytic malignancies cytogenetically characterized by a deletion of chromosome 15 occurring as the sole cytogenetic aberration. The deletions were defined as del(15) (q12q21) (two cases) and del(15)(q11q21) (one case). Cytogenetic analysis was supplemented by fluorescence in situ hybridization (FISH) using a chromosome 15 specific whole chromosome painting probe and probes hybridizing to the UBE3A gene on 15q11~q13, the PML gene on 15q22, and the telomeric region of 15q. Hereby, an interstitial deletion of 15q including UBE3A, but not PML and the telomeric region of 15q could be demonstrated. Two of our patients were diagnosed as acute myelocytic leukemia (AML) with bone marrow dysplasia classified as AML-M6 and AML-M4, respectively, according to the French-American-British classification; the third patient suffered from a chronic myelomonocytic leukemia (CMMoL). In two cases, the aberration was found at the time of primary diagnosis, whereas the third case showed the del(15) only during relapse of leukemia. Both cases with acute leukemia did not adequately respond to intensive chemotherapeutic treatment and died 13 and 11 months, respectively, after primary diagnosis. Our findings and the data of five previously published cases with an isolated del(15) indicate that: 1) del(15) represents a rare but recurrent abnormality in myelocytic hemopathies; 2) in our cases, del(15) was interstitial and included the region 15q11~q13/UBE3A, but not 15q22/PML and the telomeric region of 15q as shown by FISH; 3) del(15) occurs frequently in disorders with myelodysplastic or myeloproliferative features and may therefore affect early hematopoietic progenitor cells; and 4) del(15) may occur during disease progression and is often associated with an unfavorable prognosis.     

Interstitial deletion of the long arm of chromosome 11 determined by fluorescencein situ hybridization

Summary An interstitial deletion, del(11)(q14q22), found in a female infant was examined by fluorescencein situ hybridization with cosmid DNA markers mapped on the long arm of chromosome 11. Three cosmids mapped on 11q14.1-11q22.1 region were not hybridized to the del(11) chromosome, while all the other DNA markers mapped on 11cen-11q14.1 and 11q23.1-11 qter region gave hybridization signals on the del(11) chromosome. Cytogenetic analysis after R-banding confirmed an apparent deletion of 11q14-q22, but containing a small R-negative band, a part of 11q22.3 and/or 11q14.1, in the middle part of del(11) chromosome. The karyotype thus was determined to be 46, XX, del(11)(q14.1q22.3).     

del(2)(p23): a consistent recurrent abnormality in acute myelogenous leukemia.

A case of acute myelogenous leukemia (AML-M2) with an unusual chromosomal finding is presented. In addition to the most frequently observed translocation in this neoplasia, involving the long arms of chromosomes 8 and 21, there was a partial deletion of the short arm of chromosome 2 band (p23), i.e., 46,XX,del(2)(p23),t(8;21)(q22;q22). Deletion of the short arm of chromosome 2 has been described in association with other chromosome abnormalities in two other cases of AML and as the sole abnormality in three cases of AML, indicating that this abnormality is nonrandom and may be associated with leukemic transformation of hematopoietic cells. Therefore, we propose that the del(2)(p23),t(8;21)(q22;q22) abnormality be accorded status III and possibly considered a subset of AML (M2).     

Dual-probe fluorescence in situ hybridization assay for detecting deletions associated with VCFS/DiGeorge syndrome I and DiGeorge syndrome II loci

Over 90% of patients with DiGeorge syndrome (DGS) or velocardiofacial syndrome (VCFS) have a microdeletion at 22q11.2. Given that these deletions are difficult to visualize at the light microscopic level, fluorescence in situ hybridization (FISH) has been instrumental in the diagnosis of this disorder. Deletions on the short arm of chromosome 10 are also associated with a DGS-like phenotype. Since deletions at 22q11.2 and at 10p13p14 result in similar findings, we have developed a dual-probe FISH assay for screening samples referred for DGS or VCFS in the clinical laboratory. This assay includes two test probes for the loci, DGSI at 22q11.2 and DGSII at 10p13p14, and centromeric probes for chromosomes 10 and 22. Of 412 patients tested, 54 were found to be deleted for the DGSI locus on chromosome 22 (13%), and a single patient was found deleted for the DGSII locus on chromosome 10 (0. 24%). The patient with the 10p deletion had facial features consistent with VCFS, plus sensorineural hearing loss, and renal anomalies. Cytogenetic analysis showed a large deletion of 10p [46, XX,del(10)(p12.2p14)] and FISH using a 10p telomere region-specific probe confirmed the interstitial nature of the deletion. Analysis for the DGSI and the DGSII loci suggests that the deletion of the DGSII locus on chromosome 10 may be 50 times less frequent than the deletion of DGSI on chromosome 22. The incidence of deletions at 22q11.2 has been estimated to be 1 in 4000 newborns; therefore, the deletion at 10p13p14 may be estimated to occur in 1 in 200,000 live births.     

The paradox of 20q11.21 amplification in a subset of cases of myeloid malignancy with chromosome 20 deletion

Deletion of the long arm of one chromosome 20 (del(20q)) is a well‐recognized abnormality in acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) and is presumed to cause loss of a tumor suppressor gene at 20q12. In a previously published series of MDS and AML cases, which had lost this region via unbalanced translocation, around 40% of cases were shown to have additional copies of the chromosome 20 abnormalities, with resulting gain or amplification of the retained parts of chromosome 20, most often 20q11.2. We have used FISH and array comparative genomic hybridization, to define further the retained and amplified regions. We now report targeted amplification of 20q11.21 in four of the 22 cases selected for further study and in one new case. The shortest amplified region of 250 kb in a series of five patients with three to ten copies of the 20q11.21 region contained the complete HCK, TM9SF4, PLAGL2, and POFUT1 genes. By RT‐PCR we have shown that there is correlation between amplification and increased expression of these four genes in most cases. Localized and high level amplification of the common 250 kb region is evidence for activation of an oncogene in this region in these MDS and AML cases. Cases with 20q11.21 amplification tended to have a high proportion of erythroblasts in the marrow, with two cases diagnosed as erythroleukemia (AML‐M6). Chromosome sub‐band 20q11.21 amplification may therefore prove to be a marker of a specific subset of AML/MDS with a significant erythroid component. © 2010 Wiley‐Liss, Inc.     

Molecular cytogenetics localizes two new breakpoints on 11q23.3 and 21q11.2 in myelodysplastic syndrome with t(11;21) translocation.

Translocation t(11;21)(q24;q11.2) is a rare but recurrent chromosomal abnormality associated with myelodysplastic syndrome (MDS) that until now has not been characterized at the molecular level. We report here results of a molecular cytogenetic analysis of this translocation in a patient with refractory anemia. Using FISH with a panel of 11q and 21q cosmid/YAC probes, we localized the chromosome 11 breakpoint at q23.3 in a region flanked by CP-921G9 and CP-939H3 YACs, distal to the HRX/MLL locus frequently involved in acute leukemias. The chromosome 21 breakpoint was mapped in a 800-kb fragment inserted into the CP-145E3 YAC at 21q11.2, proximal to the AML1 gene. It is noteworthy that in all four cases with a t(11;21) reported until now, a second der(11)t(11;21) and loss of normal chromosome 11 could be observed either at diagnosis or during the course of the disease. Since in our case heteromorphism was detected by FISH on the centromeric region of the two der(11), the second der(11) chromosome could be the result of a mitotic recombination that had occurred on the long arm of chromosome 11, rather than of duplication of the original der(11). Constancy of secondary karyotypic changes resulting in an extra copy of the putative chimeric gene at der(11), loss of 11 qter sequences, and partial trisomy 21 suggest that neoplastic progression of MDS cases with a t(11;21) may be driven by the same mechanism(s).     

Delineation of multiple deleted regions in 7q in myeloid disorders.

Loss of chromosome material due to deletions of the long arm of chromosome 7, del(7q), is a consistent finding in all types of myeloid disorders, invariably associated with a poor prognosis. Two different segments, 7q22 and 7q32-q33, have been implicated as critical regions of gene loss associated with these disorders. In the present study, we used fluorescence in situ hybridization (FISH) to characterize the 7q22 breakpoint of an apparently balanced t(7;7)(p13;q22) in an acute myeloid leukemia patient. FISH analysis on bone marrow metaphases from this patient revealed that the sequence corresponding to a series of three ordered cosmids from 7q22 was deleted from one of the der(7) chromosomes. These cosmids contain the human homologue of the Drosophila homeobox gene cut (CUTL1) and span a region of approximately 150 kb. Although the proximal boundary of the deleted segment could not be exactly defined, we estimate the size of this deletion to be approximately 500 kb. Subsequently, we carried out FISH studies using the CUTL1 cosmids on a further 16 patients with deletions of 7q and myeloid disorders. The sequence corresponding to at least two of the cosmids was deleted from the del(7q) in 11 out of 14 cases with a proximal breakpoint within 7q22. Further detailed FISH mapping in this series of 17 patients has identified two other nonoverlapping commonly deleted segments at 7q31-q32 and 7q33, respectively. These data confirm and refine other studies, implying that several different genes on 7q may be involved in the pathogenesis of myeloid diseases. Genes Chromosomes Cancer 25:384-392, 1999.