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Fms-like tyrosine kinase 3 (FLT3) is highly expressed in mixed-lineage leukemia (MLL) gene - rearranged acute lymphoblastic leukemia ( MLL + ALL) with a dismal prognosis. We previously reported that FLT3 ligand (FL) stimulation induced cell cycle arrest in MLL + ALL cells leading to resistance against anti-leukemic agents. Given that FL stimulation enhanced transforming growth factor (TGF)β1 mRNA levels in MLL + ALL cells, we extensively examined the effect of TGFβ1 on the cell cycle progression and chemosensitivity in MLL + ALL cells, and found that TGFβ1 stimulation induced MLL + ALL cells into cell cycle arrest resistant to arabinosyl cytosine; its effect was markedly enhanced in synergy with FL. Thus, it is likely that TGFβ1 and FL, both abundantly produced by bone marrow stromal cells, function in a coordinated manner to render MLL + ALL cells chemoresistant, which should lead to the development of minimal residual disease (MRD) resulting in relapse. The use of inhibitors against FLT3 and TGFβ1 may become a useful strategy for eradicating MRD in MLL + ALL. [ABSTRACT FROM AUTHOR]

Allogeneic stem cell transplantation (allo-SCT) is a potentially curative therapy for chronic myeloid leukemia and Philadelphia chromosome-positive (Ph(+)) acute lymphoblastic leukemia, and the graft-vs-leukemia (GVL) effect can eradicate residual leukemia after allo-SCT. Ph(+) leukemia cells frequently express death-inducing receptors (DR4 and DR5) for tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), which is one of the cytotoxic ligands expressed on cytotoxic T cells and natural killer cells mediating the GVL effect. Here we demonstrate that imatinib specifically downregulated DR4 and DR5 expression in cell lines and clinical samples of Ph(+) leukemia. Second-generation tyrosine kinase inhibitors (dasatinib and nilotinib) and short hairpin RNA against bcr-abl also downregulated DR4 and DR5 expression in Ph(+) leukemia cells, and transfection of bcr-abl into a Ph(−) leukemia cell line induced DR4 and DR5 expression, which was abrogated by imatinib treatment. Accordingly, Ph(+) leukemia cells that had been pretreated with imatinib showed resistance to the pro-apoptotic activity of recombinant human soluble TRAIL. These observations demonstrate that BCR-ABL is critically involved in the leukemia-specific expression of DR4 and DR5 and in the susceptibility of Ph(+) leukemia to TRAIL-mediated anti-leukemic activity, providing new insight into the mechanisms of the tumor-specific cytotoxic activities of TRAIL. [ABSTRACT FROM AUTHOR]

t(17;19)-acute lymphoblastic leukemia (ALL) shows extremely poor prognosis. E2A-HLF derived from t(17;19) blocks apoptosis induced by the intrinsic mitochondrial pathway and has a central role in leukemogenesis and chemoresistance. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is expressed on cytotoxic T cells and natural killer cells and binds with death receptors (DR4/DR5), inducing apoptosis by dual activation of intrinsic and extrinsic pathways, and TRAIL mediates the graft-versus-leukemia (GVL) effect after allogeneic stem cell transplantation (allo-SCT). We found that cell lines and patients' samples of t(17;19)-ALL expressed death receptors for TRAIL, and recombinant soluble TRAIL immediately induced apoptosis into t(17;19)-ALL cell lines. E2A-HLF induced gene expression of DR4/DR5, which was dependent on the DNA-binding and transactivation activities of E2A-HLF through the 5′ upstream region of the start site at least in the DR4 gene. Introduction of E2A-HLF into non-t(17;19)-ALL cell line upregulated DR4 and DR5 expression, and sensitized to proapoptotic activity of recombinant soluble TRAIL. Finally, a newly diagnosed t(17;19)-ALL patient underwent allo-SCT immediately after induction of first complete remission, and the patient has survived without relapse for over 3-1/2 years after allo-SCT. These findings suggest that E2A-HLF sensitizes t(17;19)-ALL to the GVL effect by upregulating death receptors for TRAIL. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented related to acute lymphoblastic leukemia (ALL).

The Fcγ receptor IIIb (FcγRIIIb), a receptor for the Fcγ region of IgG, is specifically expressed on neutrophils. It has two allelic polymorphisms, NA1 and NA2, which are highly immunogenic and act as targets in alloimmune or autoimmune neutropenia. Thus, neutrophil antigens (NA) compatibility of donor/recipient pairs might be expected to affect the engraftment of neutrophils after allogeneic SCT (allo-SCT). Here, the impact of NA compatibility of 17 patients and their donors undergoing allo-SCT with a myeloablative regimen was determined. Leukocyte depletion filters were used for all transfusions before and post-SCT; most patients received G-CSF after transplant. Major mismatches for NA1 and NA2 were present in 1 and 7 patient/donor pairs, respectively. These eight patients receiving NA major-mismatched allo-SCT were compared with nine patients who received NA compatible allo-SCT. Engraftment of neutrophils and the incidence of post-engraftment neutropenia were found to be identical in the two groups. Despite the limitations in statistical power because of the small number of patients analyzed, these observations suggest that the major mismatching for NA2 antigen has little impact on the engraftment of neutrophils after myeloablative allo-SCT, at least in patients transfused using leukocyte depletion filters and receiving G-CSF after transplantation.Bone Marrow Transplantation (2009) 43, 229–235; doi:10.1038/bmt.2008.311; published online 22 September 2008 [ABSTRACT FROM AUTHOR]

Malignant cells generally acquire some immune escape mechanisms for clonal expansion. Immune escape mechanisms also contribute to the failure of graft-versus-leukemia (GVL) effect after allogeneic hematopoietic stem cell transplantation (allo-SCT). Infant leukemias with mixed-lineage leukemia (MLL) rearrangement have a remarkably short latency, and GVL effect after allo-SCT has not been clearly evidenced in these leukemias. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)- and FasL-mediated cytotoxic pathways play important roles in cytotoxic T-lymphocyte- and natural killer cell-mediated antitumor immunity and optimal GVL activity. We investigated the in vitro sensitivity of MLL-rearranged acute lymphoblastic leukemia (ALL) and acute myeloblastic leukemia (AML) cells to TRAIL- and FasL-mediated cytotoxicity. Most of cell lines and primary leukemia cells were highly resistant to TRAIL primarily owing to low cell-surface expression of death receptors in ALL and simultaneous expression of decoy receptors in AML. Nearly half of cell lines and majority of primary leukemia cells showed low sensitivity to FasL. These results suggest that resistance to death-inducing ligands, particularly to TRAIL, could be one of the mechanisms for a rapid clonal expansion and a poor sensitivity to the GVL effect in infant leukemias with MLL rearrangement. [ABSTRACT FROM AUTHOR]

The peroxisome proliferator-activated receptor γ (PPAR γ), a member of the nuclear receptor superfamily, is expressed at highest levels in adipose tissue and functions as a central regulator in the process of adipocyte differentiation. In the present study, we showed that human leukemic cell lines, not only myeloid but also lymphoid, express PPAR γ and its activation by natural ligand (15-deoxy-δ[sup 12,14]-prostaglandin J2) and synthetic ligand (troglitazone) profoundly inhibited their proliferation by induction of apoptosis preferentially in the serum-free culture. We pursued its mechanism using the representative cell lines, and found that induction of apoptosis was accompanied by caspase-3 activation and specifically blocked by its inhibitor. While status of several apoptosisrelated molecules remained unchanged, the c-Myc expression was markedly down-regulated within 24 h after troglitazone treatment. The c-myc mRNA levels were dramatically reduced at 1 h and became undetectable at 12 h after troglitazone treatment, which proved to be accompanied by complete blockade of the Tcf-4 activity in the electrophoretic mobility shift assay. We succeeded in establishing HL-60 cell lines growing well in the presence of troglitazone in the long-term serum-free culture. They showed neither induction of apoptosis nor down-regulation of the c-Myc expression via blockade of the Tcf-4 activity after troglitazone treatment. This is the first identification of the linkage between PPAR γ-mediated apoptosis and down-regulation of the c-myc gene expression. [ABSTRACT FROM AUTHOR]

The Janus kinase (JAK) family is one of intracellular protein tyrosine kinases (PTKs) present in hematopoietic and lymphoid cells and has been shown to play a crucial role in a variety of biological responses. It was reported that a human B-precursor leukemic cell line was potently inhibited in its proliferation by one of synthetic PTK inhibitors (tyrphostins), AG490, via anti-JAK2 activity. However, no extensive studies about it have been performed. In the present study, we tested 16 human lymphoid leukemic cell lines (B-precursor, 12; T cell, four) for their sensitivity to AG490 using 3H-thymidine incorporation and colony formation assays, and found that B-precursor cell lines with 11q23 translocation or Philadelphia chromosome (Ph1) whose JAK2 proved to be constitutively phosphorylated were predominantly sensitive to AG490 at a concentration that has few inhibitory effect on normal hematopoiesis. We first revealed the association of JAK2 with BCR-ABL in Ph1-positive cell lines and with Bruton's tyrosine kinase (BTK) in cell lines with 11q23 translocation by coimmunoprecipitation experiments. Of interest, AG490 markedly down-regulated phosphorylation of JAK2, but rather transiently up-regulated phosphorylation of BCR-ABL and BTK, suggesting direct implication of AG490 in the process of the JAK2 dephosphorylation. These results indicate that AG490 exerts a potent inhibitory activity to B-precursor leukemia with specific chromosomal abnormalities, and a therapeutic approach using AG490 is expected. [ABSTRACT FROM AUTHOR]

Reports on the study conducted on seven pairs of primary sample and cell line from childhood B-precursor acute lymphoblastic leukemia without nonrandom translocations. Translocations that were available to be used in the study; Diagnosis conducted on the patients; Abnormalities that were revealed in the genes of the patients.

Thrombopoietin (TPO) is a hematopoietic growth factor which plays a central role in normal megakaryocytopoiesis and thrombopoiesis. Although the interaction between TPO and its receptor c-Mpl encoded by the c-mpl gene is now known to be implicated in the proliferation and/or differentiation of abnormal myeloid cells and normal hematopoietic stem cells, little is known about a role of the TPO/c-Mpl system in lymphoid leukemia cells. In the present study, we first examined the expression of c-mpl/c-Mpl in 23 human lymphoid leukemic cell lines (T-lineage 4, B-lineage 19) using three distinct methods. The c-mpl mRNA was detectable in as many as 20 cell lines (T-lineage 3, B-lineage 17) by reverse transcriptase-polymerase chain reaction, but its translated product, c-Mpl, was demonstrable by Western blot only in B-lineage cell lines. Flow cytometric analysis revealed the surface c-Mpl expression in 13 of 17 B-lineage cell lines, but its higher expression (>40%) was restricted in nine B-precursor cell lines, eight of which had 11q23 translocation or Philadelphia chromosome (Ph1). We also demonstrated that two of eight cell lines with 11q23 translocation or Ph1 exhibited a significant proliferative response to TPO in the 3H-thymidine uptake and colony-forming assays. Triggering of these cell lines by TPO transiently up-regulated tyrosine phosphorylation of JAK-2 and Shc, indicating that their receptor is functional. Primary leukemia cells separated from patients with B-precursor acute lymphoblastic leukemia with Ph1 or 11q23 translocation also showed the surface c-Mpl expression and a significant responsiveness to TPO. These results suggest that the TPO/c-Mpl interaction may play a physiological role in the growth regulation of B-precursor leukemia cells particularly with specific chromosomal abnormalities. [ABSTRACT FROM AUTHOR]

Granulocyte colony-stimulating factor (G-CSF) has been shown to support the growth of multipotential hematopoietic stem cells in addition to the cells of neutrophilic lineage. Philadelphia chromosome (Ph1)-positive leukemia has its origin in the hematopoietic stem cell. In the present study, we demonstrated that the proliferation of leukemic cells from chronic myeloid leukemia in blast crisis (CML-BC) and Ph1-positive acute lymphoblastic leukemia (ALL) cases is frequently stimulated with G-CSF in vitro. We next studied a total of 12 leukemic cell lines established from CML-BC (n= 6) and Ph1-positive acute leukemia (n= 6): four 'myeloid', five 'biphenotypic', and three 'lymphoid' types. All cell lines expressed G-CSF receptor (G-CSFR) in flow cytometric analysis, but their proliferative response to G-CSF in 3H-thymidine incorporation assay varied. The 'biphenotypic' cell lines expressed G-CSFR at higher levels and showed the most pronounced response to G-CSF. The 'lymphoid' cell lines showed intermediate G-CSFR expression with the modest response to G-CSF. Unexpectedly, 'myeloid' cell lines showed lower G-CSFR expression and lower G-CSF response compared with 'biphenotypic' cell lines. In three of four 'myeloid' cell lines, proliferation was partially inhibited by an addition of anti-G-CSF neutralizing monoclonal antibody into culture medium. Further, the % inhibition of 3H-thymidine uptake of cell lines positively correlated with the amount of their intracellular G-CSF measured by enzyme immunoassay, suggesting an autocrine growth mechanism via the G-CSF/G-CSFR interaction. These results suggest that G-CSF play an important role in the growth regulation of leukemia cells from Ph1-positive acute leukemia and CML-BC. [ABSTRACT FROM AUTHOR]

The p16 gene encoding a specific inhibitor of cyclin-dependent kinases 4 and 6 has been reported to be inactivated at a variety of rates in malignant tumors. We studied frequency and mechanism of inactivation of the p16 gene in various types of childhood acute lymphoblastic leukemia (ALL) using 36 leukemic cell lines established from children (B precursor-ALL, 28; B-ALL/Burkitt's lymphoma, 3; and T-ALL, 5). On Southern blot, homozygous deletions or hemizygous deletions with rearrangement were detected in 14 cell lines. The expression of p16 protein was not observed on Western blot in 18 of 22 cell lines with intact p16 gene, but induced in 16 cell lines after treatment with the demethylating agent, indicating the silencing of the p16 gene by hypermethylation. Of note, the p16 gene was inactivated by hypermethylation of the 5' CpG island in nine of nine cell lines with 11q23 translocation, but was restored with the treatment of the demethylating agent. Partial methylation of the p16 gene was also demonstrated in three of eight primary leukemia samples with this translocation, suggesting that the p16 gene inactivation by hypermethylation might play a role in the leukemogenesis and disease progression of ALL with 11q23 translocation. [ABSTRACT FROM AUTHOR]

We previously reported a novel monoclonal antibody KOR-SA3544 which predominantly reacted with a surface antigen (sSA3544) expressed on Philadelphia chromosome (Ph1)-positive acute lymphoblastic leukemia (ALL). In the present study, we demonstrate that the antibody specifically recognized nonspecific cross-reacting antigen (NCA)-50/90 (CD66c), one of the carcinoembryonic antigen (CEA)-related glycoproteins encoded by a member of the CEA gene family. In addition, we show that the SA3544 antigen (NCA-50/90) was invariably expressed in cytoplasm of all of the human leukemic cell lines examined (sSA3544-positive B-lymphoid two, sSA3544-negative T or B-lymphoid and non-lymphoid 24) regardless of the presence or absence of surface expression of this antigen. Immunoelectromicroscopic examination revealed that the cytoplasmic antigen was mainly present in granules in sSA3544-positive leukemia cells, whereas it was diffusely present in cytosol in sSA3544-negative leukemia cells. Thus, among members of the CEA family, NCA-50/90 was first demonstrated to be expressed not only on the surface of some leukemia cells, but also in cytoplasm of various types of leukemia cells. [ABSTRACT FROM AUTHOR]

We report a 20-month-old boy with acute lymphoblastic leukemia with the 11q23 translocation whose blasts markedly increased in peripheral blood after intravenous granulocyte colony-stimulating factor (G-CSF) administration, but disappeared after stopping G-CSF. The in vitro study showed that the leukemic cells separated from this patient expressed G-CSF receptor (G-CSFR) and an addition of G-CSF stimulated their proliferation by 3H-thymidine incorporation assay (stimulation index, 4.9). To clarify whether or not leukemic cells with 11q23 translocations generally express G-CSFR and show proliferative response to G-CSF, we performed the similar in vitro experiments using eight leukemic cell lines with 11q23 translocations. We found that all cell lines examined expressed G-CSFR (20-98%) and proliferation of seven leukemic cell lines was significantly enhanced in response to G-CSF (stimulation index >1.5 in five cell lines), suggesting a possible participation of the G-CSF/G-CSFR interaction in the process of growth regulation of leukemic cells with 11q23 translocations. [ABSTRACT FROM AUTHOR]