Acute myelomonocytic leukemia with inv(16)(p13q22) complicating Philadelphia chromosome positive chronic myeloid leukemia.

The reciprocal translocation (9;22)(q34;q11) is highly characteristic of chronic myeloid leukemia (CML) and the pericentric inversion inv(16)(p13q22) is almost only found in acute nonlymphocytic leukemia of the myelomonocytic subtype (ANLL M4). Only twice before have an inv(16) and a t(9;22) been found in the same cells, and both times the patients seemed to have de novo ANLL M4. We describe the case of a 21-year-old man who in July 1986 presented with a clinically and hematologically classic chronic phase CML. Treatment with busulfan led to no improvement; instead in September 1986 he developed blast crisis with ANLL M4Eo morphology. He was now cytogenetically examined and the karyotype 45,X,-Y,t(9;22)(q34;q11),inv(16)(p13q22) was found. Southern blot analysis of the bone marrow DNA sampled at this time revealed a standard rearrangement in the 3' end of the M-bcr. Intensive cytostatic treatment caused cytopenia followed by complete hematologic, clinical, and cytogenetic reversal to chronic phase CML, so that in January 1987 the bone marrow karyotype was 46,XY,t(9;22)(q34;q11). Persistent splenomegaly was treated with splenectomy, and a chloroma of the skin was removed by irradiation. In March 1987 he received an allogeneic bone marrow transplant. Since then his only medical problem has been mild graft-versus-host disease; he is well and is working full time as a blacksmith.     

Central diabetes insipidus and inv(3)(q21q26) and monosomy 7 in acute myeloid leukemia

Chromosomal anomalies involving region 3q21q26 have been reported associated with thrombocytosis in various hematological malignancies, such as chronic myeloid leukemia, myelodysplastic syndrome, and acute myeloid leukemia (AML). Recent reports described the association of central diabetes insipidus with AML and chromosomal anomalies involving region 3q21q26. We review the database in our institution and report five cases of inv(3)(q21q26) in consecutive cytogenetic studies of AML and myelodysplastic syndromes from 1992 to 2000, two cases presented as sole abnormalities and three cases were associated with monosomy 7. Only one case was associated with central diabetes insipidus. The literature of 3q21q26 syndrome and central diabetes insipidus in myeloid leukemia is reviewed.     

Translocation 3;21 in Philadelphia chromosome positive chronic myeloid leukemia at diagnosis

We describe a further case of Philadelphia chromosome-positive chronic myeloid leukemia with a t(3;21)(q26.2;q22) present in the chronic phase. Blastic transformation occurred 8 months after presentation. The presence of the t(3;21) may indicate a poor prognosis.     

Nonrandom additional chromosome changes in acute nonlymphocytic leukemia with inv(16)(p13q22)

Five patients with acute nonlymphocytic leukemia and inv(16)(p13q22), all with additional chromosome changes, are reported. Three were diagnosed as having acute myelomonocytic leukemia (FAB-M4), and the other two as having acute monocytic leukemia (FAB-M5b). All five patients had abnormal eosinophils in the bone marrow at diagnosis. Two had a deletion of the long arm of chromosome #7, del(7)(q31), and a trisomy of chromosome #22. These changes have been reported frequently in acute nonlymphocytic leukemia with inv(16), but are extremely rare in leukemias with other specific rearrangements including t(9;22), t(8;21), and t(15;17). Our findings and review of the literature indicate that inv(16) is observed not only in acute myelomonocytic leukemia but also in acute monocytic leukemia, and that del(7q) and +22 are nonrandomly associated with inv(16) as additional abnormalities. No significant differences in the clinical features seem to exist between the patients with only inv(16) and those with inv(16) and additional chromosome changes, except for the lower white blood cell count in the latter group.     

FMS hemizygosity in myeloid dysplasia and acute myeloid leukemia with chromosomal aberration del(5)(q) demonstrated by polymerase chain reaction

Interstitial deletions of the long arm of chromosome 5 del(5)(q), are recurring aberrations in the myelodysplastic syndrome and acute myeloid leukemia. Several genes located in region (5)(q23–34) have been implicated as being of pathogenic importance. In this study seven samples of six patients with myelodysplastic syndrome and acute myeloid leukemia who have the del(5)(q) aberration were analyzed by polymerase chain reaction (PCR) and Southern blot technique. FMS hemizygosity was demonstrated in all patients. PCR analysis from peripheral blood samples confirmed the observations of this aberration found by semiquantitative Southern blot. PCR-based analysis can be used for primary diagnosis in addition to cytogenetic evaluation and for follow-up in patients with del(5)(q) aberration.Abbreviations PCR Polymerase chain reaction - MDS Myelodysplastic syndrome - AML Acute myeloid leukemia - RA Refractory anemia - RAEB Refractory anemia with excess of blasts - RAEBt Refractory anemia with excess of blasts in transformation to acute leukemia     

Detection of single-copy chromosome 17q gain in human neuroblastomas using real-time quantitative polymerase chain reaction.

Regional genomic alterations resulting from single-copy allelic loss or gain have been well characterized in many human cancers and are often of prognostic relevance. Unbalanced gain of 17q material is common in malignant human neuroblastomas and typically results from unbalanced translocations. Unbalanced 17q gain may be an independent predictor of disease outcome, but technical difficulties with quantifying such gain using fluorescent in situ hybridization gives this method limited clinical applicability. We now describe a duplex genomic DNA-based quantitative polymerase chain reaction assay to determine the presence or absence of unbalanced gain of chromosome 17q in primary neuroblastoma specimens. The technique was first refined and validated in a panel of nine human neuroblastoma-derived cell lines by direct comparison with dual-color fluorescent in situ hybridization. Prospective blinded comparison of quantitative polymerase chain reaction and fluorescence in situ hybridization in 40 human neuroblastoma primary tumor samples showed a sensitivity of 96% and 100% specificity for detecting unbalanced 17q gain when a relative 17q copy number ratio of 1.3 was used to define unbalanced gain. Tumors with ratios >1.3 were highly associated with malignant tumor phenotypic features such as metastatic disease (P <.0001) and tumor MYCN amplification (P =.008). These data suggest that quantitative polymerase chain reaction determination of 17q status is feasible and highly specific in primary tumor samples. Sensitivity may be limited because of the inherent complexity of both the chromosomal rearrangements and heterogeneity of some tumor samples. Taken together, quantitative polymerase chain reaction can be used as a high-throughput screening tool for 17q aberrations, but a subset of samples may also require fluorescence in situ hybridization analysis in an attempt to conclusively determine 17q allelic status.     

Pediatric acute myeloid leukemia with t(7;21)(p22;q22)

The t(7;21)(p22;q22) resulting in RUNX1‐USP42 fusion, is a rare but recurrent cytogenetic abnormality associated with acute myeloid leukemia (AML) and myelodysplastic syndromes. The prognostic significance of this translocation has not been well established due to the limited number of patients. Herein, we report three pediatric AML patients with t(7;21)(p22;q22). All three patients presented with pancytopenia or leukopenia at diagnosis, accompanied by abnormal immunophenotypic expression of CD7 and CD56 on leukemic blasts. One patient had t(7;21)(p22;q22) as the sole abnormality, whereas the other two patients had additional numerical and structural aberrations including loss of 5q material. Fluorescence in situ hybridization analysis on interphase cells or sequential examination of metaphases showed the RUNX1 rearrangement and confirmed translocation 7;21. Genomic SNP microarray analysis, performed on DNA extracted from the bone marrow from the patient with isolated t(7;21)(p22;q22), showed a 32.2 Mb copy neutral loss of heterozygosity (cnLOH) within the short arm of chromosome 11. After 2‐4 cycles of chemotherapy, all three patients underwent allogeneic hematopoietic stem cell transplantation (HSCT). One patient died due to complications related to viral reactivation and graft‐versus‐host disease. The other two patients achieved complete remission after HSCT. Our data displayed the accompanying cytogenetic abnormalities including del(5q) and cnLOH of 11p, the frequent pathological features shared with other reported cases, and clinical outcome in pediatric AML patients with t(7;21)(p22;q22). The heterogeneity in AML harboring similar cytogenetic alterations may be attributed to additional uncovered genetic lesions.     

Dysplasia and high proliferation rate are common in acute myeloid leukemia with inv(16)(p13q22)

Acute myeloid leukemia (AML) with inv(16)(p13q22), also known as M4Eo, is a distinct type of AML with a favorable prognosis associated with abnormal bone marrow eosinophils. We reviewed the morphologic findings of archival bone marrow specimens with M4Eo, specifically assessing for dysplasia, and performed immunohistochemical studies to assess the growth fraction using the MIB-1 (Ki-67) antibody. We also assessed the apoptotic rate by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-nick end labeling. All assessable cases had more than 10% dysplastic forms in at least 1 lineage. Seventeen cases had 10% or more dysplastic forms, and 3 cases had more than 50% dysplastic forms in at least 2 lineages. Immunoreactivity for Ki-67 was higher in M4Eo than in other AML types (P = .000). The apoptotic rate in M4Eo was similar to other AML types (P = .724). Our data show that dysplasia is a prominent feature, but not a prognostic indicator, in M4Eo. M4Eo is associated with a significantly higher proliferation rate than other AML types.     

Myelodysplastic syndrome with inv(3)(q21q26.2) or t(3;3)(q21;q26.2) has a high risk for progression to acute myeloid leukemia

Acute myeloid leukemia (AML) with inv(3) (q21q26.2) or t(3;3)(q21;q26.2) is a distinct subtype in the World Health Organization classification. The natural history of myelodysplastic syndrome (MDS) associated with these cytogenetic aberrations is poorly understood. We studied 17 MDS (11 de novo and 6 therapy related) and 3 chronic myelomonocytic leukemia (CMML) cases associated with inv(3) (q21q26.2) or t(3;3)(q21;q26.2). The de novo cases were further classified as refractory cytopenia with multilineage dysplasia (n = 8) and refractory anemia with excess blasts (n = 3). Isolated inv(3)/t(3;3) was identified in 4 cases, whereas -7/7q (n = 13) and -5/5q (n = 6) were common additional aberrations. Nineteen patients died, including 13 in whom the disease progressed to AML after a median of 7 months. Median survival for patients with de novo disease was similar to that for patients with therapy-related MDS (13 vs 17.5 months). MDS or CMML with inv(3)/t(3;3) are aggressive diseases with a high risk of progression to AML.     

Translocation (3;8)(q26;q24): a recurrent chromosomal abnormality in myelodysplastic syndrome and acute myeloid leukemia

We identified a reciprocal translocation between chromosomes 3 and 8, with breakpoints at bands 3q26 and 8q24, in five patients with myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML). The t(3;8)(q26;q24) was the sole cytogenetic aberration in two patients, was associated with trisomy 13 in one patient, and occurred with monosomy 7 in two patients. In three patients, the AML or MDS developed 36, 52, and 57 months following chemotherapy for soft tissue sarcoma, mantle cell lymphoma, and diffuse large B-cell lymphoma, respectively; in these three patients, the neoplasms were considered to be therapy-related. All five patients displayed marked trilineage dysplasia and variable degrees of cytopenias, with marked thrombocytosis noted in one patient and a normal platelet count in another patient. All patients were treated with combination chemotherapy; at writing, four were still alive and one had died during a follow-up period ranging from 1 to 16 months. We conclude that the t(3;8)(q26;q24) is a recurrent translocation associated with therapy-related MDS/AML or de novo AML, and is frequently associated with monosomy 7.     

Rapid prenatal diagnosis of trisomy 21 by real-time quantitative polymerase chain reaction with amplification of small tandem repeats and S100B in chromosome 21

Trisomy 21 (Down syndrome) is the most common congenital anomaly, and it occurs in one out of 700-1000 births. Current techniques such as amniocentesis and chorionic villi sampling (CVS) require lengthy laboratory culture procedures and high costs. This study was undertaken to establish a rapid prenatal diagnosis of trisomy 21 using real-time quantitative polymerase chain reaction (PCR) of fetal DNA from amniotic fluid. Real-time quantitative PCR was performed with DNA templates obtained from 14 normal blood samples, 10 normal amniotic fluid samples, 14 Down syndrome blood samples, and 7 Down syndrome amniotic fluid samples. Primers for D21S167 and S100B of chromosome 21 were used. Primers that direct the amplification of the 165-bp fragment of the insulin-like growth factor (IGF)-1 gene on chromosome 12 using a PCR primer were included to generate an internal standard for quantitation. The relative levels of D21S167 and S100B were 2.6 and 2.4 times higher in the blood of Down syndrome patients than those in the control group. The differences between these two groups were statistically significant (p-values were 0.0012 and 0.0016, respectively). The relative levels of D21S167 and S100B were 2.1 and 2.7 times higher in the amniotic fluid of Down syndrome fetuses than those in the control group. The difference between these two groups was statistically significant (p-values were 0.0379 and 0.0379, respectively). Prenatal diagnosis of trisomy 21 by real-time quantitative PCR using STR (small tandem repeats) amplification of D21S167 and S100B is a useful, accurate and rapid diagnostic method. Furthermore, it may also be useful for prenatal diagnosis with fetal DNA from maternal blood, and for preimplantation genetic diagnosis and prenatal counseling.     

Translocation (3;6)(q21;p21) in acute myeloid leukemia with abnormal thrombopoiesis and basophilia

A case of acute myeloid leukemia (AML) with increased numbers of basophils and abnormal megakaryocytes was shown to have a t(3;6)(q21;p21) in the bone marrow cells. The morphology is described in detail and the case is discussed with reference to t(6;9)(p23;q34) and inv/ins(3)(q21q26) in AML. It is possible that increased numbers of basophils in AML may be associated with a translocation involving 6p21-6p23.     

del(2)(p23) as a sole abnormality in a case of acute myeloid leukemia

We report a case of acute myeloid leukemia (AML) [FAB-M5a] showing a deletion of the short arm of chromosome 2 at band p23 as a sole abnormality in the bone marrow cells. This abnormality deserves to be considered as an established nonrandom entity in AML.     

Detection of 1p and 19q loss in oligodendroglioma by quantitative microsatellite analysis, a real-time quantitative polymerase chain reaction assay

The combined loss of chromosomes 1p and 19q has recently emerged as a genetic predictor of chemosensitivity in anaplastic oligodendrogliomas. Here, we describe a strategy that uses a novel method of real-time quantitative polymerase chain reaction, quantitative microsatellite analysis (QuMA), for the molecular analysis of 1p and 19q loss in oligodendrogliomas and oligoastrocytomas in archival routinely processed paraffin material. QuMA is performed on the ABI 7700 and based on amplifications of microsatellite loci that contain (CA)n repeats where the repeat itself is the target for hybridization by the fluorescently labeled probe. This single probe can therefore be used to determine copy number of microsatellite loci spread throughout the human genome. In genomic DNA prepared from paraffin-embedded brain tumor specimens, QuMA detected combined loss of 1p and 19q in 64% (21 of 32) of oligodendrogliomas and 67% (6 of 9) of oligoastrocytomas. We validate the use of QuMA as a reliable method to detect copy number by showing concordance between QuMA and fluorescence in situ hybridization at 37 of 45 chromosomal arms tested. These results indicate that QuMA is an accurate, high-throughput assay for the detection of copy number at multiple loci; as many as 31 loci of an individual tumor can be analyzed on a 96-well plate in a single 2-hour run. In addition, it has advantages over standard allelic imbalance/loss of heterozygosity assays in that all loci are potentially informative, paired normal tissue is not required, and gain can be distinguished from loss. QuMA may therefore be a powerful molecular tool to expedite the genotypic analysis of human gliomas in a clinical setting for diagnostic/prognostic purposes.     

Identification of a novel fusion gene MLL-MAML2 in secondary acute myelogenous leukemia and myelodysplastic syndrome with inv(11)(q21q23)

We have identified a novel fusion partner of MLL, namely the mastermind like 2 (MAML2 gene), in secondary acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) with inv(11)(q21q23). RT-PCR and sequencing revealed that exon 7 of MLL was fused to exon 2 of MAML2 in the AML and MDS cells. The inv(11)(q21q23) results in the creation of a chimeric RNA encoding a putative fusion protein containing 1,408 amino acids from the NH2-terminal part of MLL and 952 amino acids from the COOH-terminal part of MAML2. The NH2-terminal part of MAML2, a basic domain including a binding site of the intracellular domain of NOTCH, was deleted in MLL-MAML2. MLL-MAML2 in secondary AML/MDS and MECT1-MAML2 in mucoepithelioid carcinoma, benign Wartin's tumor, and clear cell hidradenoma consist of the same COOH-terminal part of MAML2. A luciferase assay revealed that MLL-MAML2 suppressed HES1 promoter activation by the NOTCH1 intracellular domain. MAML2 involving a chimeric gene might contribute to carcinogenesis in multiple neoplasms by the disruption of NOTCH signaling.     

Acute myeloid leukemia (AML) with t(8;21)(q22;q22) relapsing as AML with t(3;21)(q26;q22)

We here report on an 48-year-old male patient with a primary diagnosis of acute myeloid leukemia (AML)-M2 with t(8;21)(q22;q22), who developed complete hematologic and molecular remission after induction chemotherapy. Thirteen months later, he relapsed and showed an AML-M2 with t(3;21)(q26;q22). Retrospectively, polymerase chain reaction (PCR) for AML1-EVI1 and EVI1 overexpression was performed on bone marrow and peripheral blood samples taken at diagnosis and during the first year after the first manifestation of AML to quantify the AML1-EVI1-positive clone. In a bone marrow sample taken 25 days from diagnosis, PCR for AML1-EVI1 was negative, and EVI1 expression, as assessed by quantitative real-time PCR, was within the same range as that of healthy controls. These data suggest that this patient developed a secondary therapy-related AML rather than a relapse.     

Dicentric (19;21)(p13;p13), a novel chromosomal abnormality occurring in a case of Philadelphia chromosome-positive acute lymphoblastic leukemia

We report on a patient with Philadelphia chromosome-positive acute lymphoblastic leukemia, who acquired a novel chromosomal abnormality, a dic(19;21)(p13;p13), during relapse of the disease. The cytogenetic result was confirmed by fluorescence in situ hybridization using alpha-satellite and library probes specific for chromosomes 19 and 21, respectively, as well as a chromosome 19q13.1-specific DNA probe. In our case, the dic(19;21) represents a secondary genetic change and was associated with disease progression and poor prognosis.