Comparative Clinical Study Of Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia And Lymphoid Blast Crisis Of Chronic Myelogenous Leukemia

Background and ObjectiveThe Philadelphia (Ph) chromosome is the reciprocal translocation between the bcr (breakpoint cluster region) gene on the chromosome22and the abl (Abelson tyrosine kinase) gene on the chromosome9, which leads the formation of bcr/abl fusion gene, and is closely related to the development of chronic myeloid leukemia (CML). It is important to monitor the bcr/abl fusion gene transcriptions levels, because it can predict the relapse of leukemia and is a reliable factor for the therapeutic response of leukemia. The Ph chromosome is the specific chromosomal change in CML and plays a key role in the development of CML. About70%of CML cases end in myeloid blast crisis, and20%-30%cases end in lymphoid blast crisis. The Ph chromosome is also present in the blast cells of CML. With the advances of cytogenetics, the Ph chromosome is also found in approximately20%～40%of adults cases with acute lymphoblastic leukemia (ALL), and is considered as one of the worst prognostic factors in these patients. To explore the clinical features of patients with Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ALL) and chronic myeloid leukemia in lymphoid blast crisis (CML LBC), we retrospectively analyzed the clinical data of21cases with Ph+ALL and31cases with CML LBC. Materials and methods1Patients:The research enrolled21cases of Ph+ALL and31cases of CML LBC, which were treated in the department of Hematology of the First Affiliated Hospital of ZhengZhou University from July,2005to July,2010, and the follow-up time over12months. The diagnosis of leukemia was based on the marrow cell morphology, immunophenotypes and cytogenetic analysis, according to the diagnostic criteria. All the patients were treated with chemotherapy.2Methods2.1Reverse Transcription-Polymerase chain Reaction (QT-PCR):Bone marrow mononuclear cells were separated from heparinized bone marrow. RNA was extracted from cells by Trizl. The gene copies of bcr/abl were assayed by nest QT-PCR.2.2FISH:About300to500interphase cells were analyzed. Interphase FISH was used to detect bcr/abl gene. Probes for bcr and abl gene sequence probes were purchased from Jin Beijia Company, Beijing. The fluorescence hybridization signal of interphase cells were visualized with red fluorescence signal for bcr gene, green fluorescent signals for abl gene, and yellow fluorescent signal for bcr/abl fusion gene. There were two red and two green fluorescent signal in normal cells. One red one green and one yellow signal were considered as positive cells.2.3Immunophenotype:Indirect immunofluorescence flow cytometry was used to detect immunophenotype of leukemic cells. Monoclonal antibodies for lymphoid lineage included T-cell lineage (CD2, CD3, CD4, CD7, CD8) and B-cell lineage (CD10, CD19, CD20, CD22, cCD79a). Monoclonal antibodies for myeloid lineage included CD13, CD14, CD15, CD117, and CD33. Monoclonal antibodies for progenitor cell were HLA-DR, CD34. Total10000cells were collected by FACsort and analyzed by a software Cell-Quest. Cells were grouped by gating on CD45expression and side scatter.2.4. Morphological analysis:Bone marrow aspiration slides were stained by GIEMSA staining and histochemistry staining and cells were differentially counted. The subtypes of leukemia were based on FAB classification.2.5. Chromosome karyotype:At least20metaphase cells of each sample were analyzed using G-banding technique. Chromosome karyotype was defined according to the International Naming System of Human Chromosomes (ISCN). At least two cells with the same structural chromosome rearrangement were confirmed as cloning abnormality.2.6. Remission induction:All the patients received the standard chemotherapy of VDCLP regimen (V:Vincristine, D:Daunorubicin, C:Cyclophosphamide, L: L-asparaginase, P:Prednisone) for their induction treatments.2.7. Consolidation therapy:Patients achieved complete remission were treated using sequential chemotherapy of VDCLP, CODP, CHOP, EA, MAE, and VMCP regimens (O:Vincristine, M:Mitoxantrone, A:Cytarabine, E:Etoposide). Some patients were treated with the addition of imatinib or nilotinib, who could afford the agents.3. Statistics analysis:The data were analyzed using software SPSS15.0. Numeric data were expressed as mean±standard deviation (x±s). The difference between two groups was compared by student t or t’test. The probability of survival and correlation was analyzed by Kaplan-Meier. The standard of significant level was α=0.05, when P<0.05had a statistical significance.Results1. Clinical data:The patients with Ph+ALL had a mean age of43.33±13.68years old (ranged form17to64years old), with13males and8females. The patients with CML LBC had a mean age of40.39±16.46years old (ranged form14to75years old), with19males and12females. There was no statistical significance between the two groups(P>0.05). The incidence of hepatosplenomegaly of the CML LBC patients was80.65%, and the Ph+ALL patients was14.28%, there was statistical significance between the two groups (P<0.05).31cases of CML LBC patients all have gone through chronic phase. And there were9cases occurred with hepatomegaly and11cases occurred with splenomegaly before the lymphoid blast crisis.2. Laboratory data:The white blood cell count was more than10×109/L in8cases (38.10%) of Ph+ALL, and in23cases (74.19%) of CML LBC. There was statistical significance between the two groups (P<0.05).3. Flow cytometry:the immunophenotype were detected in21cases of Ph+ALL, among which16cases (76.19%) expressed B-lymphoid lineage marker CD10, CD19, CD79a, and HLA-DR; One cases (4.76%) expressed B-lymphoid lineage marker with T-lymphoid lineage marker (CD2, CD3, CD7); Four cases (19.05%) expressed B-lymphoid lineage marker with myeloid lineage marker (CD13,CD33). The immunophenotype of31cases of CML LBC were analyzed with20cases (64.52%) only expressing B-lymphoid lineage marker (CD10, CD19, HLA-DR, CD34);10cases (32.26%) expressing B-lymphoid lineage marker with myeloid lineage marker (CD13, CD33); and1cases (3.23%) only expressing T-lymphoid lineage marker. There was no statistical significance between the two groups (P>0.05).4. Karyotype analysis:Classical Ph chromosomes were detected in19out of21cases of Ph+ALL, with one cases carried hyperdiploid abnormalities and Ph chromosomes, and the another case carried Ph chromosomes and-9chromosome abnormalities. Classical Ph chromosomes were detected in29out of31cases of CML LBC, with one case carried4q+abnormalities and Ph chromosomes, and the another case carried Ph chromosomes and45XY abnormalities. The Ph chromosomes disappeared when the Ph+ALL patients achieved complete remission. But the Ph chromosomes were continuous present when the CML LBC patients achieved complete remission. The count of Ph chromosome cell was as high as80%.5. Detection of bcr/abl:the bcr/abl fusion gene copy number of pre-treatment was70.12±20.33in21cases of Ph+ALL, which was74.26±22.05in31cases of CML LBC. Bcr/abl was detected in9cases out of21cases of Ph+ALL who has achieved complete remission, and the gene copy number was0.32±0.15. There were12cases of CML LBC achieved hematological response with Bcr/abl copies at49.54±16.34.There were5cases out of31cases of CML LBC has used TKIs, and3cases out of the5cases who has returned to chronic phase received the detection of bcr/abl, and the gene copy number was68.74±13.09.All of the CML LBC patients expressed with p210, and all of the Ph+ALL patients expressed with p190.6. Clinical response:the complete remission rate in Ph+ALL (69.10%) was higher than those in CML LBC (48.39%)(P<0.05). The median survival of Ph+ALL was10.76±6.91months, and these in CML LBC was7.06±6.03months, there was statistical significance between the two groups (P<0.05).7. Side effects:Eighteen out of21cases (85.17%) of Ph+ALL patients and27out of31cases (87.10%) of CML LBC experienced varied degree of complications after chemotherapy, such as mouth ulcers, bleeding, headache, dizziness, nausea, vomiting, abdominal pain, diarrhea, etc. There was no statistical significance between the two groups (P>0.05).Conclusion1. There are differences of clinical manifestations between the Ph+ALL and CML LBC.2. There is no Ph chromosomes detected in bone marrow cells when the Ph+ALL patients achieved complete remission. The Ph chromosomes are continuously detected in bone marrow cells when the CML LBC patients achieved complete remission.