Biphenotypic acute leukemia with Ph1 chromosome, M-BCR-, myeloperoxidase+, and CALLA+]

A 63 year-old woman was referred to our hospital because of fever and increased number of blasts in the bone marrow. On physical examination she had slight hepatomegaly but no splenomegaly. Laboratory tests disclosed a hemoglobin level of 8.5 g/dl; a WBC count of 13,200/microliter with 26% blasts; a platelet count of 51,000/microliter. A bone marrow aspirate was normocellular with 74% blasts and 37% blasts were stained positive for myeloperoxidase. Cell surface markers for HLA-DR, CD10, CD19, CD13, CD33 were positive. Karyotype analysis revealed 46, XX, t (9q+; 22q-) and 45XX, -7, t (9q+; 22q-). Southern analysis showed rearrangement of immunoglobulin heavy chain but not T cell receptor beta gene. Rearrangements in M-BCR were not detected with 5' or 3' bcr probes. After 2 courses of chemotherapy, blasts decreased to 7% with recovery of normal elements and 11 out of 20 metaphases of the bone marrow cells were normal karyotype. These findings suggest that this case was de novo Ph1 positive acute leukemia which demonstrated both lymphoid and myeloid features.     

An unclassifiable case of hypoplastic leukemia in old age treated successfully with vincristine and prednisolone

An 80-year-old male was admitted because of dizziness and palpitation. Laboratory investigation revealed pancytopenia. A bone marrow aspirate showed a markedly hypocellular marrow with 41.6% blast cells. Peroxidase activity was negative and PAS reaction was block positive in the blast cells. Surface markers of these cells were positive for HLA-DR antigen and partially positive for CD13 (MY7). Other markers, such as T, B or myeloid antigens were all negative. These blast cells were classified as L1 according to the FAB system but suggested essentially unclassifiable in cell differentiation. The patient was treated successfully with vincristine and prednisolone and induced into complete remission although repeated marrow examination findings revealed hypocellular. As for the classification of hypoplastic leukemia, lymphoid or primitive "stem cell" leukemia also should be considered as other categories of acute leukemias and be treated according to each case.     

Philadelphia-positive acute mixed leukemia with monosomy 7

A 26-year-old male was admitted to our hospital because of fever and leukocytosis. On admission, a white blood cell count was 28,300/microliters with 46.5% blast cells and 16.0% atypical monocytoid cells, a hemoglobin level 13.7 g/dl, and a platelet count 15.0 X 10(4)/microliters. Bone marrow contained 58.8% of peroxidase-negative blast cells. He was diagnosed as acute lymphoblastic leukemia (ALL L2) according to the FAB classification. Chromosome analysis revealed the marrow cells to contain 45, XY, -7, t(9; 22) (q34; q11). On surface marker analysis, the leukemic cells were positive for both lymphoid (CD10) and myeloid markers (CD13). Two color flow-cytometric analysis showed two distinct populations with CD10 and CD1 3, respectively. Rearrangements of both immunoglobulin heavy chain and T cell receptor beta-chain were observed. The "breakpoint cluster region" on chromosome 22 was not rearranged. On the basis of these findings, we thought this case being acute mixed leukemia. He was refractory to AdVP therapy and BHAC-DMP therapy. He is now under treatment with A-Triple-V therapy.     

Spontaneous Epstein-Barr virus transformed B-cell line sharing the identical immunoglobulin gene rearrangement with acute myeloid leukemia

Mononuclear cells from a 44-year-old patient with acute myeloid leukemia (AML) gave rise to a spontaneous permanent cell line cultured in suspension. The cell line was shown to be positive for Epstein-Barr virus nuclear antigen (EBNA). As expected, its composite phenotype was of B-cell type with B-cell antigens (CD 20, CD 21) and with monoclonal surface IgM of kappa type, but without detectable IgM secretion. Surprisingly, identical monoclonal rearrangements of the immunoglobulin heavy chain (JH) sequences could be demonstrated in the uncultured bone marrow AML cells and in the cell line that also had kappa light chain gene rearrangement. This is the first case to our knowledge of an EBNA positive B-cell line with identical monoclonal Ig heavy chain rearrangement as detected in myeloblastic leukemia cells.     

Treatment of a Child With Myeloid/NK Cell Precursor Acute Leukemia With L-Asparaginase and Unrelated Cord Blood Transplantation

A 2-year-old Japanese boy who presented with multiple cervical, axillary, and inguinal lymphadenopathy was diagnosed by immunocytochemical analysis as having myeloid/natural killer (NK) cell precursor acute leukemia. Leukemic blasts in the bone marrow were positive for CD56 (NK marker), CD7 (T-cell marker), CD33 (myeloid marker), CD34, and HLA-DR. Tumor cells in a lymph node were also positive for CD2, cytoplasmic CD3 (T-cell marker), CD7, CD33, CD34, and CD56, but negative for peroxidase staining and other T-cell, NK, and myeloid markers. Southern blot analysis showed no rearrangement bands for T-cell receptor delta and immunoglobulin heavy chain. Chromosomal analysis revealed 46,XY,inv(7)(p21q21). Neither chemotherapy for acute lymphoblastic leukemia nor that for acute myeloid leukemia induced remission in this patient. However, complete remission was achieved by the administration of L-asparaginase (6,000 U/m2 for 5 days). Because the disease was considered refractory to standard chemotherapy, cord blood transplantation was performed from an HLA 1-locus mis-matched unrelated donor. The conditioning regimen consisted of total body irradiation, cytarabine, and cyclophosphamide, and cyclosporine and short-term methotrexate were employed for graft-versus-host disease (GVHD) prophylaxis. Hematological reconstitution was rapid, and only grade I acute GVHD was observed. The patient has been in remission for more than 24 months after transplantation. Our findings indicate that combination therapy with L-asparaginase and allogeneic stem cell transplantation may be useful for the treatment of myeloid/NK cell precursor acute leukemia.     

Basophilic differentiation of leukemic cells in a patient with acute leukemia carrying minor bcr/abl chimeric mRNA

A 77-year-old woman was referred to our hospital because of leukocytosis and leukoblastosis in September 1999. She was healthy except for hypertension, and no abnormal findings in the peripheral blood had been observed up to December 1998. Physical examination revealed neither hepatosplenomegaly nor superficial lymphadenopathy. A bone marrow film showed massive proliferation of blast cells (87.8%), some of which contained coarse basophilic granules (38.6%). The cells were negative for peroxidase and esterase (alpha-naphtyl butyrate and ASD-chloroacetate) staining, but the granules showed metachromasia upon toluidine blue staining. As immunophenotypic analysis of the cells showed double positive for CD13/CD19 but negativity for CD33, this case did not meet the diagnostic criteria for biphenotypic acute leukemia. Chromosome and gene analysis showed positivity for the Ph1 chromosome with minor bcr/abl chimeric mRNA. A homogenate of the peripheral mononuclear cells demonstrated a high concentration of histamine. Electron microscopy analysis confirmed that some of the blast cells contained dense granules, which closely resembled "immature basophil granules" morphologically. These results suggested that the blast cells showed basophilic differentiation. As the clinical course and peripheral blood findings were different from blastic crisis of chronic myelogenous leukemia (CML) and CML with minor bcr/abl chimeric mRNA, the present case was diagnosed as "multiphenotypic acute leukemia", a type of acute basophilic leukemia classified by Duchayne.     

Acute myelomonocytic leukemia (M4) with CD19 antigen expression, eosinophilia and basophilia in bone marrow

A 12-year old boy was admitted to Saitama Children's Medical Center because of fever and epistaxis. He had leukocytosis (WBC 40,800/microliters, blast 75%), anemia, thrombocytopenia and high levels of serum LDH, lysozyme, Vitamin B12, and plasma histamine. Bone marrow aspiration revealed hypercellular marrow with 31.2% blasts, 15.2% eosinophils, and 14.2% basophils. Blasts had Auer rods and were positive for peroxidase and negative for alpha-naphthyl butyrate esterase and PAS stainings. Ia, CD13 (My7), and CD19 (B4) antigens were expressed on his leukemic cells. Chromosomal study showed 46, XY, t(7;8) (q35;q22), del(9) (q13q22). Southern blot analysis using immunoglobulin constant region (C) probes revealed germline patterns of C mu, C kappa, C lambda, and breakpoint cluster region. A diagnosis of acute myelomonocytic leukemia (AMMoL, M4) was made. He attained a complete remission with daunorubicin and cytarabine, and 6 months later he received bone marrow transplantation from HLA-identical sister. This case had the common breakpoint 8q22 with ANLL with t(8;21) (q22;q22), and was unique AMMoL with proliferation of eosinophils and basophils in bone marrow.     

Acute Myelogenous Leukemia Concurrent With Untreated Chronic Lymphocytic Leukemia

We report a case of acute myelogenous leukemia (AML) concurrent with untreated chronic lymphocytic leukemia (CLL). An 84-year-old Japanese man was admitted to the Chihaya Hospital with persistent high-grade fever. Morphologic observation of peripheral blood and bone marrow smears revealed a proliferation of blasts and lymphocytosis with small and mature phenotypes. Immunophenotyping of the blast cells revealed CD13+, CD33+, CD34+, and HLA-DR+, and that of the lymphocytes revealed CD5+, CD19+, CD20+, and lambda+ on the cell surface. The peripheral lymphocytes revealed an IgH gene rearrangement. Chromosome analysis of 20 metaphase cells from bone marrow showed numerous abnormalities, containing +8,+11,+21. The patient's disease was diagnosed as AML with trilineage dysplasia concurrent with CLL. The simultaneous occurrence of AML and CLL is extremely rare but should not be overlooked as a possible underlying cause of lymphocyte abnormalities in AML patients.     

Acute leukemias associated with the 4;11 chromosome translocation have rearranged immunoglobulin heavy chain genes

Studies of acute leukemia with the 4;11 translocation have yielded conflicting results regarding the lineage of the cell of origin in this disease. To investigate this issue further, we have examined the state of immunoglobulin genes in tumor cells from two affected patients, immunophenotyped their leukemic cells using a number of monoclonal antibody reagents with specificities for lymphoid or myelomonocytic antigens, and examined the malignant cells by electron microscopy. DNA was extracted from leukemic bone marrow cells and hybridized with radiolabeled DNA fragment probes specific for the constant region of immunoglobulin heavy chain and kappa and lambda light chain genes. Autoradiographs revealed rearrangement of both allelic heavy chain genes, but a germline configuration of light chain genes in both cases. Surface marker analysis showed that blasts from both patients expressed HLA-DR and the myeloid antigens Leu-M1, 1C2, 2D1, and 4B3, but lacked common acute lymphocytic leukemia antigen or T antigens. Furthermore, they did not have sheep erythrocyte receptors nor did they express surface or cytoplasmic immunoglobulin or B cell precursor determinants. Electron microscopy analysis showed that blast cells from patient 1 exhibited numerous monoribosomes, polyribosomes, and isolated strands of rough endoplasmic reticulum in their cytoplasm. These ultrastructural features are characteristic for both common acute lymphocytic leukemia and pre-B-ALL cells, but not for T-ALL or acute myelogenous leukemia cells. Peroxidase was undetectable in cells from both patients. Our study suggests that this disorder represents a unique subtype of leukemia. The cell of origin may be an early B cell progenitor that shares certain surface antigens with myeloid cells or a stem cell with the potential for both lymphoid and myelomonocytic differentiation.     

Rearrangement of immunoglobulin heavy chain genes in T cell acute lymphoblastic leukemia

We studied the arrangement of the immunoglobulin heavy chain genes by Southern blot analysis of DNA freshly obtained from marrow blast cells of 14 children with T cell acute lymphoblastic leukemia (T-ALL) using probes to the C mu and JH gene segments: At least one of the C mu-gene alleles was rearranged in three cases. In two of these, one C mu gene had the germ-line configuration and one was rearranged, whereas both alleles were rearranged in the third case. In one case, a rearranged heavy chain gene hybridized to the C mu-region probe, but not to the JH probe, indicating that the entire JH region had been deleted. These results demonstrate that immunoglobulin heavy chain gene rearrangements are not restricted to B lineage lymphoproliferative diseases in humans.     

髓系／NK前体细胞急性白血病1例并文献复习

目的:提高对髓系/NK前体细胞急性白血病的认识.方法:报道1例髓系/NK前体细胞急性白血病资料,并复习文献,总结该病临床及实验室检查特点.结果:患者以腹部症状起病,肝脾显著肿大.流式细胞仪免疫表型分析HLA-DR( )、CD11c( )、CD11b( )、CD13( )、CD33( )、CD38( )、CD56( )、MPO( ).经两疗程CHOP方案化疗,骨髓象部分缓解,但是髓外浸润加重,后改用VDP方案,最终因出现感染、DIC、多器官功能衰竭而死亡.结论:急性NK细胞白血病具有独特的临床、组织病理学和免疫表型特征,对现有化疗不敏感,预后恶劣.     

Rituximab can be useful as treatment for minimal residual disease in bcr-abl-positive acute lymphoblastic leukemia

We report the results of administering CD20 monoclonal antibody (MoAb) in a 32-year-old man with bcr-abl-positive acute lymphoblastic leukemia. Morphological complete remission was achieved after two lines of chemotherapy with persistence of blast cells (2%) in flow cytometric analysis of marrow cells. Since no HLA-matched donor for allogeneic bone marrow transplantation (BMT) was found, anti-CD20 MoAb therapy was administered for in vivo marrow purging, prior to autologous peripheral blood stem cell (PBSC) harvest and transplantation. After MoAb therapy <0.1% of blast cells were observed and the molecular abnormality (bcr-abl gene rearrangement) disappeared.     

Successful allogeneic bone marrow transplantation in a case of adult precursor B-lymphoblastic lymphoma

A 42-year-old woman was admitted to the National Cancer Center Hospital in Tokyo, Japan, because of lumbago and bilateral leg pain. Clinical examination revealed a retroperitoneal bulky tumor, a breast tumor, and lymphoblasts in bone marrow. She did not have surface lymph node swelling or systemic symptoms such as weight loss and night sweats. Lymphoblasts in bone marrow were positive for CD10 and CD19 but negative for CD20, surface immunoglobulin, and T-cell antigens. Needle biopsy of a retroperitoneal mass revealed diffuse infiltration of lymphoblastic tumor cells. Because the cells were immunoreactive for CD79a, CD10, and terminal deoxynucleotidyl transferase, the patient was diagnosed as having precursor B-lymphoblastic lymphoma (which is rare in adults) with bone marrow involvement. The patient achieved complete remission by an induction therapy for acute lymphoblastic leukemia, underwent allogeneic bone marrow transplantation, and has remained in complete remission for more than 3 years.     

Establishment and characterization of acute B-cell lymphocytic leukemia cell line showing (8;14) and (14;18) chromosome translocation

Cell line KHM-2B expressing two oncogene products, c-myc and bcl-2, was established from a patient with acute lymphocytic leukemia with an 8;14 and 14;18 chromosome translocation. Surface marker studies of the cell line showed that the cells were positive for HLA-DR, CALLA (CD10), B1 (CD20) and B4 (CD19), but negative for T11 (CD2). The fresh cells from peripheral blood of the patient had no surface immunoglobulins, whereas KHM-2B cells were positive for mu.lambda type surface immunoglobulin. A cytogenetic analysis of the cell line revealed two translocations, t (8;14) (q24;q32) and t(14;18)(q32;q21). Rearrangement of the c-myc and bcl-2 genes was detected by Southern blot analysis of the KHM-2B DNA. Northern blot analysis revealed production of c-myc and bcl-2 mRNAs. These results indicated that two oncogenes were activated by two translocations to immunoglobulin genes.     

Diagnosis of stem cell leukemias in view of phenotypic and genotypic analysis

To identify the biological characteristics of so called stem cell leukemia (SCL), of which leukemic blast cells should be derived from pluripotent stem cells, immunophenotypical and genotypical analysis and response to several hematopoietic cytokines were studied in 272 cases with acute de novo leukemia. In 132 cases with acute myelogenous leukemia (AML), some cases of CD19+ and/or CD7+ AML were considered as SCL. In cases with myeloperoxidase negative acute lymphoblastic leukemia (ALL), cases of CD7 + CD1 - CD3 - CD4 - CD8 - My-Ag (myeloid antigens) +ALL, considered as those of T-precursor ALLs, and cases of HLA-DR + CD19 + CD20 - My-Ag + ALL, considered as those of B-precursor ALLs, were though to be SCL. We did not think the cases of ALL with dual genotype to be SCL, since dual genotype could not be considered as sings of ability to differentiate to multilineage but as products of the process of active V-DJ rearrangements of Ig heavy chain gene.     

Simultaneous establishment of myeloid and B-lymphoid cell lines with identical chromosome abnormalities from Philadelphia chromosome-positive chronic myelogenous leukaemia

Two continuously growing cell lines, designated YOS-M and YOS-B, were established simultaneously from a patient with Philadelphia (Ph1) chromosome-positive chronic myelogenous leukaemia (CML) in myeloid blast crisis. Both YOS-M and YOS-B had the Ph1 chromosome and identical additional chromosome abnormalities, which were not detected in the chronic phase. Cytochemical analysis showed that YOS-M was significantly positive for peroxidase, whereas YOS-B was entirely negative. YOS-M expressed myeloid-associated antigens (CD14, CD33) as well as CD4, CD25 and CD34. The surface phenotype of YOS-M was identical to that of the leukaemic blasts found in the patient. On the other hand, YOS-B expressed mature B-cell markers, CD19, CD20, CD21 and surface immunoglobulin, but not myeloid-associated antigens. These two cell lines showed an identical rearrangement pattern of the break point cluster region on chromosome 22, but rearrangement of the immunoglobulin heavy chain gene was detected only in YOS-B. These findings provide definite evidence that CML cells still have the capability to differentiate and mature along different haematopoietic cell lineages even after blast crisis.     

CD56-positive acute lymphoblastic leukemia

We report a patient with lung cancer who developed CD56-positive acute lymphoblastic leukemia. He was referred to our hospital for thrombocytopenia. Atypical cells were found in the blood and the bone marrow. These cells were immunophenotypically positive for CD3epsilon, CD56, and terminal deoxynucleotidyl transferase, and negative for surface CD3, CD4, CD19, CD33, and myeloperoxidase. A small proportion of leukemic cells express CD13. There were no rearrangements of T-cell receptor (TCR)-beta, TCR-gamma, or immunoglobulin heavy chain. No Epstein-Barr virus was detected. Systemic examination did not detect any tumors other than pulmonary adenocarcinoma, and the patient was diagnosed as having acute natural killer (NK) cell leukemia. Chemotherapy was effective, and he achieved complete remission. The course of the disease was complicated by a lung abscess, and the patient died 3 months after the diagnosis. We considered that the diagnosis was blastic NK cell lymphoma/leukemia subtype. However, it actually was myeloid/NK cell precursor leukemia subtype that weakly expressed CD13.     

Acute myeloid leukemia with mediastinal myeloid sarcoma refractory to acute myeloid leukemia therapy but responsive to l-asparaginase

We report the case of a 14-year-old female with acute myeloid leukemia (AML) and myeloid sarcomas (MS) in the anterior mediastinum and around numerous bones. Laboratory tests showed a white blood cell count of 4.0?×?10(9)/l with 7.0?% blasts. Computed tomography revealed a mediastinal mass and pleural effusion; pleural effusion cytology was negative for malignant cells. In addition, disseminated intravascular coagulation (DIC) was present. Following DIC therapy, thoracoscopic and bone marrow biopsies were performed. Immunostaining and surface marker analysis revealed that the blast cells were positive for cytoplasmic myeloperoxidase, CD4, CD7, CD33, CD44, CD117, and HLA-DR, but negative for CD34 and CD56. Karyotype was normal. MS associated with AML was diagnosed. Multidrug chemotherapy for AML was completely ineffective, and MS continued to progress. Immunohistochemistry revealed that the blasts were negative for asparagine synthetase (AS); therefore, chemotherapy including L: -asparaginase was initiated. After the first administration of L: -asparaginase, the patient's condition improved; however, she subsequently developed tumor lysis syndrome and sepsis, which eventually led to death. Aggressive MS in childhood is rare and refractory to existing AML chemotherapy. Chemotherapy including L: -asparaginase may prove to be effective in such cases, especially those in which blast cells show negative AS expression.     

Intracellular expression of CD61 precedes surface expression

 We report a case of acute myeloid leukemia which we have classified as acute megakaryoblastic leukemia because of intracytoplasmic expression of CD61. Light microscopy demonstrated agranular blasts which, by cytochemical staining, were negative for myeloperoxidase. Using flow cytometry, the blast cells were seen to be positive for HLA-DR, CD7, CD13, CD33, and CD34, thus revealing their myeloid origin. Immunophenotyping on fixed blood smears additionally showed positive reaction with the CD61 antibody, demonstrating the megakaryoblastic differentiation of the blasts. After permeabilization of the cell membrane, the intracytoplasmic CD61 expression was confirmed by flow cytometry. Cytogenetic analysis disclosed a del(7)(q21–22). Our findings suggest that cytoplasmic expression of CD61 may precede the cell-surface expression of this antigen and should therefore be investigated in all cases of acute leukemias with undifferentiated morphology and negative cytochemistry to set apart early FAB-M7 from FAB-M0. Received: February 8, 1999 / Accepted: May 5, 1999     

Acute leukemia with the phenotype of a natural killer/T cell bipotential precursor

 An acute leukemia with an unusual immunophenotype developed in a 17-year-old girl. At the initial presentation, extramedullary involvement was not evident, but with advancing disease, massive splenomegaly and an osteolytic rib tumor developed. The disease was aggressive and refractory to intensive chemotherapeutic regimens for myeloid and lymphoid malignancies, and the patient died 3 months after the initial presentation. The leukemic cells were of irregular shape and variable size; they had deeply indented or bi-lobed nuclei and relatively fine, azurophilic granules in their cytoplasm. They were positive for acid phosphatase and β-glucuronidase in granular staining, but they were negative for myeloperoxidase. The leukemic cells had a unique immunophenotype: it was positive for T-cell antigens (CD1a, CD2, cytoplasmic CD3, CD4), myeloid antigens (CD13 and CD33), NK-cell antigen (CD56), CD19 and CD30. DNA analysis revealed no gene rearrangement in the T-cell receptor β, γ and δ, or immunoglobulin heavy chain genes. The leukemic cells of our patient are thought to have arisen from the transformation of a putative precursor cell common to both the T- and NK-cell lineage in the bone marrow. The current literature on precursor NK-cell malignancy is reviewed, and its clinicopathological feature is discussed. Received: 29 April 1998 / Accepted: 28 August 1998