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Objective: To report a rare complex karyotypic abnormalities including ins (15;17),t(2;17;20) and trisomy 8 in a patient with acute promyelocytic leukemia (APL)., Methods: Chromosomes were prepared after 24 h culture of bone marrow cells. R-banding technique was used to analyze the karyotype. Multiplex fluorescence in situ hybridization (M-FISH), chromosome painting using whole chromosome parint (WCP) 2, 15, 17 and 20 and interphase-FISH (I-FISH) using PML-RARa dual-colour dual-fusion translocation probe were performed to ascertain the essence and origin of the abnormal chromosomes detected by conventional karyotypic analysis., Results: Karyotypic analysis revealed a karyotype of 47, XY, 2q-, + 8, 17q+ , 20p+ . M-FISH analysis showed a karyotype of 47, XY, t(2;17;20) (q24;q21;p13), + 8, which was confirmed by chromosome painting. PML-RARa fusion gene which lied in the derivative chromosome 15 was detected by I-FISH suggesting a cryptic insertion (15;17)(q22;q21.1q21.3)., Conclusion: FISH is a reliable method for characterization of cryptic ins (15;17) and other complex translocations. It should be used in all suspected APL patients lacking t(15;17) by conventional karyotypic analysis.

Objective: To investigate the characteristics of the abnormalities of chromosome 17 in myeloid malignancies with complex chromosomal abnormalities (CCAs)., Methods: Abnormalities of chromosome 17 were analyzed in 73 patients with myeloid malignancies with CCAs showed by R banding and conventional karyotyping, including 21 acute myeloid leukemia (AML), 36 chronic myeloid leukemia (CML) and 16 myelodysplastic syndrome (MDS). All CCAs were further analyzed by multiplex fluorescence in situ hybridization (M-FISH)., Results: Among the 73 myeloid malignancies with CCAs, chromosome 17 was the most frequently involved chromosome. It was found in 46.5% (34/73) of all cases, including 12 AML, 13 CML in blast crisis (BC) and 9 MDS. However, it was not found in the 9 CML cases in chronic phase (CP). The majority of changes were structural rearrangements which were identified in 43.8%(32/73)of all cases, among them the frequency was 52.4% (11/21), 33.3% (12/36) and 56.3% (9/16) in AML, CML and MDS, respectively. Numerical abnormalities were detected in 15.1% (11/73) cases, all were monosomy 17, and the frequency was 25.0% (3/12), 38.5% (5/13) and 33.3% (3/9) in AML, CML and MDS, respectively. Both numerical and structural abnormalities of chromosome 17 were found in 9 cases. Unbalanced translocations involving chromosome 17 were much more frequent than balanced ones. In the 3 groups, 16, 15 and 8 unbalanced translocations were found respectively. Only two kind of balanced translocations including t(15;17) in AML and t(15;17;22) in CML were found. All chromosomes were involved except chromosomes 5, 6 and 22 as partner chromosomes, the most common one was chromosome 15 (8.2%), followed by chromosome 2 (5.4%). Five of the 6 cases with translocation of chromosomes 15 and 17 were acute promyelocytic leukemia, the other case was CML-BC., Conclusion: Abnormalities of chromosome 17 were the most frequently involved chromosomal aberrations in myeloid malignancies, and structural rearrangements were more common. All the numerical abnormalities were monosomy 17, unbalanced translocations were much more frequent than balanced ones.

Background and Objectives: Human leukemia cell lines are of great value in leukemia research. Thus far 36 leukemia cell lines carrying the 11q23 translocation and MLL rearrangements, including two cell lines with t(6;11)(q27;q23) and an MLL-AF6 fusion gene have been described. We have established a new monocytic cell line with t(6;11), designated SHI-1, and herein describe its biological characteristics., Design and Methods: Mononuclear cells isolated from the bone marrow of a patient with acute monocytic leukemia (AML-M5b) at relapse were inoculated and passaged by liquid culture. The biological features of the cell line were characterized by morphological assays, flow cytometry, cytogenetic analysis, reverse transcription polymerase chain reaction (RT-PCR), direct sequencing, fluorescence in situ hybridization (FISH), clonogenic culture, quantitative fluorescent PCR, zymography, short tandem repeating sequences-PCR (STR-PCR), multiplex-FISH (M-FISH), and tumorigenic capacity in nude mice., Results: The SHI-1 cell line has been maintained in continuous culture without any external cytokines for three years. The morphology and immunoprofile of the cells show typical features of monocytic lineage. Karyotypic analysis demonstrated a t(6;11)(q27;q23) translocation accompanied by a deletion of 17p, which are the same abnormalities as were seen in the leukemia cells of this patient in relapse. The MLL-AF6 fusion transcript and the loss of one p53 allele were proven by chromosome painting, FISH and RT-PCR analysis in both SHI-1 cells and the primary leukemia cells. A point mutation of ATC-->ACC at codon 195 of exon 6 in another p53 allele was found by direct sequencing of DNA in SHI-1 cells as well as in the primary leukemia cells. Neither Epstein-Barr virus nor mycoplasma was detected in SHI-1 cells. Tumor masses were found in all sixteen mice 9-19 days after subcutaneous injection of SHI-1 cells. DNA fingerprinting confirmed the authenticity of the cell line., Interpretation and Conclusions: SHI-1 is a new monocytic leukemia cell line with the t(6;11) translocation, p53 gene alterations, and high tumorigenicity in nude mice. It could be a valuable tool in the study of leukemogenesis.

Objective: To investigate the clinical and biological characteristics of childhood acute myeloid leukemia(AML)with 8;21 translocation., Methods: A retrospective analysis including clinical information, cell morphology, chromosome, immunophenotype and molecular biology was performed on 41 cases of childhood t(8;21)AML. The control group included 19 cases of AML without t(8;21) translocation detected during the same period., Results: The 41 cases of t(8;21)AML accounted for 68.3% of 60 continuous childhood AML patients. Among them, classical t(8;21) translocation was seen in 29 cases; variant t(8;21) translocation, simple 8q-, near-tetraploidy characterized by the duplication of t(8;21) translocation each came into view in 2 cases; and cryptic t(8;21) translocation was seen in 6 cases. Thirty seven cases (80.4%) belonged to M2 subtype of AML. Most of them had the morphological changes such as the leukemia cells' indent nucleus with a light stain region of perinucleus, basophilic cytoplasm, differentiation with maturation, megaloblastoid changes and nuclear-cytoplasm imbalance; the high expression of CD13 antigen; and the AML1/ETO fusion transcript in 23 cases examined by reverse transcription-polymerase chain reaction (RT-PCR) assay, including 6 cases with normal karyotype. The difference in complete remission rate between t(8;21) positive patients group and t(8;21) negative patients group was not significant in statistics (82.4% vs 75%, P>0.05). However the difference in recurring rate of the leukemia was statistically significant (10.7% vs 41.7%, P<0.05)., Conclusion: t(8;21)AML is the most frequent type of childhood AML. It is predominantly associated with M2 subtype of AML and has unique morphological, immunological prognostic features .

Objective: To report five cases of acute myeloid leukemia (AML) with t(16;21)(p11;q22) translocation and the result of chromosome painting analysis on one of them., Methods: Chromosome specimens were prepared by short-term culture of bone marrow cells. Karyotype analysis was made by R-banding technique. Chromosome painting was performed using whole chromosome probes 16 and 21 in 1 case., Results: Karyotype analysis showed identical translocation t(16;21)(p11;q22) in all five cases, accounting for 0.3% of 1448 cases of acute myeoid leukemia examined in the past fifteen years. Moreover, chromosome painting distinctly demonstrated t(16;21) in one of them. Leukemia blasts did not show hemophagocytosis in all of them., Conclusion: t(16;21) translocation is a rare and recurring chromosome rearrangement. It represents a specific type of AML. Chromosome painting technique is a more reliable means for detecting it, compared with the conventional karyotype analysis.

Objective: To evaluate the value of a panel fluorescence in situ hybridization (FISH) in the detection of common chromosome abnormalities in myelodysplastic syndrome (MDS)., Methods: Twenty cases of MDS patients, whose karyotypes were unknown by the FISH examiner beforehand, were analyzed with a panel FISH using YAC248F5 (5q31), YAC938G5 (7q32), CEP8 and YAC 912C3 (20q12) probes to detect the frequently occurring chromosome abnormalities (-5/5q, -/7q-, +8, 20q-) in MDS. Then the results were compared to those of conventional cytogenetics (CC)., Results: Among 20 cases, 13 cases were found to carry common chromosome abnormalities by panel FISH (trisomy 8, five cases; -5/5q-, one case; 20q-, five cases; 5q- accompanying 20q-, one case; complex abnormalities, one case). However, on CC examination, only five cases were found to have common chromosomal abnormalities (20q-, four cases; 5q- accompanying 20q-, one case). In addition, trisomy 21, marker chromosome and complex abnormalities comprising -5, -7 and marker chromosomes were seen in one case each, the rest were normal., Conclusion: Panel FISH is a useful tool of molecular cytogenetics in the detection of common chromosome abnormalities in MDS.

Objective: To study the clinical and cytogenetic characteristics of four patients with myelodysplastic syndrome (MDS) and one with acute myeloid leukemia experiencing t (1;7)., Methods: Five patients seen in our hospital from 1992 to 2001 were diagnosed as MDS and acute myelocytic leukemia (AML) according to the French-American-British (FAB) criteria. Chromosomes were prepared using the direct method as well as 24-hour unstimulated cultures of fresh heparinized bone marrow for each subject, while R-banding was used to analyze karyotypes. Dual-color fluorescence in situ hybridization (FISH) using SpectrumRed and SpectrumGreen directly labeled chromosome 1-specific alpha-satellite DNA probe (red) and chromosome 7- specific alpha-satellite DNA probe (green) was performed for three cases., Results: Of the five patients, three had 1;7 translocation due to a long history of exposure to benzene. In three cases, dual-color FISH resulted in three red signals and two green ones, in which one red signal adjoining one green signal in 27.6%, 84% and 18.5% metaphases, respectively., Conclusions: Exposure to benzene may be the cause for Chinese MDS and AML patients with t (1;7) translocation. The result of dual-color FISH convincingly confirmed that the centromere of the derivative chromosome 7p/1q resulting from 1;7 translocation was made up of centromeres from both chromosomes 1 and 7.

Objective: To study the origin of aberrant clones with chromosome abnormalities in MDS., Methods: Antibodies recognizing myeloid cells (CD33), B cells (CD19) and T cells (CD2) were used to sort bone marrow cells of ten MDS patients with chromosome abnormalities (trisomy 8 five cases, trisomy 8 accompanying trisomy 9 one case, trisomy 8 accompanying 7p- one case, 20q- two cases, t (1; 7) one case) by Immuno-Magnetic Bead technique, then the positive cells for CD33, CD19 or CD2 were detected respectively by interphase FISH using the centromeric probes for chromosomes 1, 7and 8, and the locus specific probe for 20q12. 200 to 300 cells were scored for each detection., Results: FISH showed that trisomy 8, t (1; 7) and 20q- all affected CD33 positive cells, but not CD2 positive cells in these ten MDS patients. Only in one MDS patient, 20q- affected the CD19 positive cells., Conclusion: Our data suggest that the chromosomal aberrations in MDS are mostly restricted to myeloid cell lineages, but B lymphocytes are involved in minor MDS implying that abnormal clone originated from a multipotent hematopoietic progenitor.

Objective: To explore the morphological, immunophenotypical and cytogenetic (MIC) characteristics of chronic myeloid leukemia in blast crisis (CML-BC)., Methods: Marrow cells from 31 CML-BC patients were studied by bone marrow smears with morphological and cytochemical stain analysis. Immunophenotypic analysis was performed by micro blood method and flow cytometry. Cytogenetic analysis was performed using bone marrow cells prepared directly and or after 24 hours culture. RHG banding was used for karyotypic analysis., Results: 23 patients (74.2%) were found to have myeloid blast crisis. 5 cases (16.1%) had a B-lymphoid immunophenotype and 4 of them expressed myeloid associated antigens at the same time. Another 3 cases were classified as acute undifferentiated leukemia (AUL 1 patient) and acute myeloid -lymphoid leukemia (AMLL 2 patients) with B and myeloid markers expression. A significant proportion (67.7%) of blast crisis cases were CD(34) positive. CD(34) was expressed in 80.0% of lymphoid and 65.2% of myeloid BC. Analysis of the relationship between CD(7) and CD(34) expression in blasts of myeloid crisis showed their dual expression in 8/23 (34.8%) of the cases. Cytogenetic analysis indicated that 14/27 (51.9%) of the patients had developed secondary cytogenetic abnormalities in addition to the Ph chromosome. These newly emerging abnormalities included a trisomy 8 in 3 patients, a double Ph chromosome in 3 patients, an iso chromosome 17q in 2 patients, loss of Y chromosome in 1 patent and other complicated translocation in 5 patients., Conclusion: Blast crisis of CML is a kind of disease of stem cell. The differentiation of blast cell is blocked in the early stage. The prognosis of such patients is poor. MIC showed its important values in diagnosis, judgement of prognosis and determination of therapeutic protocol of CML-BC.

Objective: To verify the value of sister chromatid differentiation (SCD) analysis of bone marrow cells in establishing the diagnosis and predicting prognosis of myelodysplastic syndrome (MDS)., Methods: S-bromodeoxyuridine-SCD and karyotyping studies were performed in 50 cases with clinically suspected MDS., Results: Prolonged cell cycle time (CT) was found in 33 of the 50 cases (66.0%). In 13 cases with acute myeloid leukemia (AML), all had prolonged CT. However, only 2 (10.5%) of the 19 cases with nonmalignant hematological disorders had prolonged CT. There were significant differences of CT between the cases with MDS and those with AML, aplastic anemia (AA) and benign hypercellular anemias as well as healthy individuals. However, there were no significant differences between healthy individuals and cases with AA and, with benign hepercellular anemia. CT gradually prolonged during the evolution of MDS. 24 of the 50 patients (48.0%) were found to have clonal chromosomal abnormalities. In seven cases not compatible with MDS according to FAB criteria, six were found to have clonal chromosomal abnormalities. The mortality rates were 11.8 and 50.0% in the groups of normal SCD and of prolonged SCD, respectively (P < 0.05). The rates of leukemia transformation in both the groups were 5.9% and 16.7% (P > 0.05). The median survival of both the groups was 12 months and 8 months (P = 0.051)., Conclusion: SCD analysis is valuable for establishing the diagnosis and predicting the prognosis of MDS.

Objective: To study the myelodysplastic syndrome(MDS) with 1;7 translocation in five cases and to determine further the constitution and origin of centromere of the derivative chromosome resulting from 1;7 translocation., Methods: Bone marrow chromosome preparation of five cases was made using direct method or short- term culture. Karyotypic analysis was carried out by R-banding technique. Dual-color fluorescence in situ hybridization(FISH) using Spectrum Red and Spectrum Green directly labeled chromosome 1-specific a-satellite DNA probe(red) and chromosome 7-specific a-satellite DNA probe(green) was performed in three patients of them., Results: All of the five cases had 1;7 translocation. The centromere of the derivative chromosome 7p/1q was constituted with red and green signals in three of them., Conclusion: The result of dual-color FISH confirms that the centromere of the derivative chromosome resulting from 1;7 translocation originated from both centromeres of chromosome 1 and chromosome 7.

Objective: To evaluate the association between isolated trisomy 11 and the clinical, hematological, immunological, prognostic aspects in hematological malignancies., Methods: Bone marrow cell cytogenetic analysis was performed by direct method and/or 24 h culture method. RHG banding was used for karyotype analysis. Immunophenotype analysis was carried out by flow cytometry. Ten patients with acute myeloid leukemia (AML) were treated with HA regimen chemotherapy and followed up., Results: The isolated trisomy 11 was found in 11 of 1 * ! 763 hematological malignancies cases (0.6%). The diagnoses included 10 AML (6 M(2), 2 M(5), 1 M(1), 1 M(4)), and 1 myelodysplastic syndromes. Ten of them have no hepatosplenomegaly. The immunophenotypical analysis of leukemia cells showed positive for CD(13), CD(33) and CD(34) in 5 cases. Follow-up data were available in 10 cases. The complete remission rate was 40% with a median survival of 10 months., Conclusion: The isolated trisomy 11 was mainly seen in AML, especially in M(2) subtype. Their prognosis was poor.

Objective: To study the clinical, biological features and prognosis of acute biphenotypic leukemia (BAL) in the adults., Methods: Bone marrow specimens of 63 BAL patients were evaluated to prove the diagnosis and the classification by morphologic, cytochemical, immunologic and cytogenetic (MIC) examinations. These patients were treated with protocols suitable for acute myeloid leukemia (AML), or acute lymphoblastic leukemia (ALL), or both., Results: No significant difference in clinical features was observed between BAL, AML or ALL. Morphologically, the subtypes of M(5), M(1) and M(2) were predominant in AML, as L(2) and L(1) were in ALL. Immunologically, coexpression of myeloid and B lineage associated antigens was predominant and CD(34) was hyperexpressed in BAL, which suggested that BAL might originate from malignant transformation of earlier hematopoietic cells. Cytogenetically, Ph chromosome was observed in 25.5% (13/51) of BAL patients. Prognostically, both the treatment response and the overall survival of BAL patients were poor., Conclusion: Patients with BAL have unique clinical, biological and prognostic features.

Objective: To evaluate the four techniques for clonal analysis in the early diagnosis of myelodysplastic syndromes (MDS)., Methods: Four techniques for clonal analysis were performed in bone marrow samples from fifty patients with suspected MDS: (1) Conventional cytogenetics (CC) for clonal chromosomal abnormalities; (2) BrdU-sister chromatid differentiation (BrdU-SCD) for cell cycle analysis; (3) Fluorescence in situ hybridization (FISH) for trisomy 8; (4) PCR-SSCP for N-ras mutation., Results: The diagnosis of forty-five patients was compatible with FAB criteria of MDS, the other five patients didn't fully meet the FAB criteria. They had either only one lineage dyspoiesis or no any obvious dysplastic features and two of them were diagnosed as suspicious refractory anemia (RA), one as anemia with hypercellular bone marrow and two as chronic aplastic anemia. The results of the four techniques performed in them showed that four patients had clonal karyotype abnormalities, two had prolonged cell cycle, three had trisomy 8 of different proportions, and one had N-ras mutation. Thus, they were all diagnosed as RA., Conclusion: The untypical MDS patients can be diagnosed early by examination with combining several clonal analysis techniques.

Objective: To report two myelodysplatic syndromes (MDS) patients with t(3; 5) (q25; q34)., Methods: Chromosome specimens were prepared by short-term culture of bone marrow cells. Karyotype analysis was performed by R banding technique, chromosome painting (fluorescence in situ hybridization, FISH) by using whole chromosome 3 and 5 probes in case 1., Results: The clinical and hematological findings were compatible with diagnosis of MDS. Karyotype analysis showed that both patients had identical t(3; 5) (q25; q34) translocation. A reciprocal translocation between chromosomes 3q and 5q was proved by FISH in one patient., Conclusions: t(3; 5) translocation is a rare chromosome abnormality specifically associated with MDS and frequently displays trilineage dysplasia. Chromosome painting technique is a reliable tool for detecting this translocation.

Objective: To present a special case with the karyotype and molecular marker of acute myeloid leukemia (AML)-M2 who was induced to complete remission by all-trans retinoic acid (ATRA) alone., Methods: A recently hospitalized young female patient with acute leukemia was initially diagnosed as M3 subtype based on morphological French-American-British (FAB) classification. Karyotype analysis using standard G and R banding techniques and RT-PCR were applied to further define the diagnosis. After primarily cultured bone marrow cells from the iliac aspiration were tested for in vitro induced differentiation, the patient was treated with oral all-trans retinoic acid alone, 60 mg per day until complete remission was achieved. Peripheral blood and bone marrow changes were monitored over the whole treatment course., Results: The characteristic chromosomal aberration for M3, the t(15;17) reciprocal translocation, was not found while a t(8;21) translocation was verified. Furthermore, an amplified product of the AML-1/ETO fusion gene instead of the PML/RAR alpha fusion gene was detected by RT-PCR and the diagnosis was corrected from M3 to M2. Primary cultured bone marrow cells can be fully induced to terminal differentiation after 4 days exposure to ATRA. A hematological complete remission was achieved after 40 days treatment with ATRA as a single therapeutic agent, suggesting an alternative pathway mediating ATRA-induced myeloid differentiation., Conclusion: A leukemia patient with a subtype other than M3, such as M2 in this case, may also be induced to complete remission by the mechanism of ATRA-induced terminal differentiation. This implies that there may be a pathway other than PML/RAR alpha fusion gene product which mediates ATRA-induced myeloid maturation in leukemia cells.

Objective: To evaluate fluorescence in situ hybridization (FISH) in the detection of inv (16) (p13; q22)., Methods: Spectrum red labeled yeast artificial chromosome (YAC) clone 854E2 which spans the breakpoint cluster region in MYH11 in band 16p13 and single color interphase FISH were used to detect inv (16) in 26 cases of acute myelomonocytic leukemias (AML-M(4)), and the results were compared with that of conventional cytogenetic analysis., Results: R banding karyotyping test revealed no inv (16) in 25 cases, one AML M(4Eo) case showed inv (16) by G banding. Nine cases including all three M(4Eo) had inv (16) by FISH analysis, among whom the characteristic fluorescence signal pattern of the inv (16) was seen in 13.3% to 32.1% (median, 21.3%) of the tested cells., Conclusion: YAC 854E2 and interphase FISH provide a powerful technique in the detection of inv (16) (p13q22).

To investigate the value of metaphase-fluorescence in situ hybridization (M-FISH) in the diagnosis of acute promyelocytic leukemia (APL) and the detection of its minimal residual disease (MRD), 10 cases of untreated acute myeloid leukemia (AML) (de novo APL 5 cases, relapsed APL 3 cases, AML-M(1) and AML-M(5) one case each) diagnosis by cell morphology at presentation and 10 cases of APL after complete remission (CR) were studied by M-FISH using a whole chromosome 17 painting probe labeled by digoxigenin and the results were compared with that of conventional cytogenetic examination and reverse transcription-polymerase chain reaction (RT-PCR). Among 10 untreated AML cases, 7 had positive M-FISH results, of whom 4 had t (15;17) translocation, 3 had normal karyotype. Six of them had PML/RARalpha fusion transcript except one, in whom RT-PCR did not be performed; 3 had negative M-FISH results, of whom one had del (2q) x 2 abnormalities, who was RT-PCR-positive for PML/RARalpha fusion transcript; one had complex karyotype abnormalities, whose RT-PCR was negative for PML/RARalpha fusion transcript; one had t (9;22) translocation, whose RT-PCR was negative for PML/RARalpha fusion transcript, but positive for BCR/ABL fusion transcript. Thus the diagnosis of AML-M(3) was revised as AML-M(2) for the latter two cases. 10 APL cases after CR had normal karyotype, but 12/15 M-FISH assays detected 1 - 5 t (15;17) positive cells in 9 of them. This finding is compatible with the results of RT-PCR assays. Leukemia relapse was seen in one case, and two positive M-FISH results were appeared in the 2 assays at a 13 months' interval. This study suggests that M-FISH had important practical value in the diagnosis of APL and the detection of MRD, and that it is less sensitive than RT-PCR, however, it seems to be more potential for prediction of the relapse of leukemia due to its capacity of detecting quantitatively the chromosome translocation in proliferative cells.