Yoshinobu Asano

Effect of (-)-epigallocatechin gallate on leukemic blast cells from patients with acute myeloblastic leukemia.

Information on the anti-carcinogenic effect of EGCG, the main constituent of the polyphenols present in Japanese green tea leaves, has recently been accumulating. In this report, we evaluate the effect of EGCG on leukemic blast cells from AML patients. The results showed that EGCG inhibited the proliferation of AML cells in all cases examined. Since AML cells might proliferate by autocrine or paracrine growth mechanisms, we also examined the effect of EGCG on the production of GM-CSF from AML cells. Although EGCG did not directly inhibit the production of GM-CSF, it did inhibit the effect of TNF-alpha or TPA, both of which stimulated AML cells to produce GM-CSF. On the other hand, the modulation of receptors for growth factors might play a role in the proliferation or carcinogenesis of AML cells. We also found that EGCG inhibited the modulation of c-kit, a receptor for stem cell factor, on leukemic cells. These findings suggested that EGCG might be available as a new therapeutic tool for AML patients.

Cyclosporin A therapy for patients with myelodysplastic syndrome: multicenter pilot studies in Japan

We examined the efficacy of cyclosporin A (CsA) in 50 patients with myelodysplastic syndrome (MDS) consisting from 47 of RA, 1 of RARS, and 2 of RAEB. These patients showed various marrow cell types including hypo-, normo-, and hypercellularity. Patients belonged to the following International Prognostic Scoring System (IPSS) risk groups: 4 of low, 41 of intermediate-1, and 5 of intermediate-2. The median CsA dose was 4.58mg/kg, and treatment responses were classified according to the International Working Group (IWG) criteria. Hematological improvement (HI) was observed in 30 (60%) patients, and all of them were belonged to RA. In the patients with RARS or RAEB, no efficacy was observed. Four (8%) of the responders achieved partial remission (PR) with granulocytes > or = 1500microl(-1), Hb>11g/dl and platelets > or = 100,000microl(-1). Higher response rate (53%) was shown in erythroid lineage (HI-E) compared to platelet (HI-P, 36%) or neutrophil lineage (HI-N, 35%). When we analyzed the correlation between the response to CsA therapy and the karyotype or HLA type, there were significantly more responders with good karyotype or DRB1*1501 than with intermediate/poor karyotypes or with other HLA types. These results indicate the usefulness of CsA therapy for MDS patients with any marrow cellularity, especially for erythroid lineage and patients with good karyotype or DRB1*1501.

Early viral complications following CD34-selected autologous peripheral blood stem cell transplantation for non-Hodgkin's lymphoma

A patient with non‐Hodgkins lymphoma who received a CD34‐selected autologous peripheral blood stem cell transplant (PBSCT) developed cytomegalovirus retinitis, adenovirus‐associated haemorrhagic cystitis (HC) and fatal herpes simplex virus pneumonia. Depletion of mature T cells from the graft and a persistent decrease in CD4+ lymphocytes following PBSCT may have predisposed this patient to such viral infections. Infusion of cryopreserved autologous PBSC (containing mature T cells) was effective for adenovirus‐associated HC. Immunosuppression and resultant viral infections may affect patients receiving CD34‐selected autologous transplantation.

Immunosuppressive therapy for patients with refractory anemia.

Abstract. Trials of immunosuppressive therapy have been reported in some case reports of hypoplastic myelodysplastic syndrome (MDS). In this study, we gave immunosuppressive therapies to eight patients with normo- or hyperplastic MDS of refractory anemia subtype without karyotypic abnormalities and analyzed the HLA-DRB1 type or the presence of paroxysmal nocturnal hemoglobinuria (PNH) neutrophils in these patients. Cyclosporin A (CyA) therapy was effective for improving cytopenia in four of the eight MDS patients. While the side effects of CyA were mostly mild and transient, one patient demonstrated karyotypic abnormality following CyA therapy and accelerated to refractory anemia with an excess of blasts. Additional antithymocyte globulin (ATG) therapy was effective in one of three nonresponders to CyA therapy. One patient died due to leukemic transformation after ATG therapy. When we analyzed the correlation between the response to CyA therapy and the HLA-DRB1 type, there were more responders with DRB1*1501 (three of four patients) than without (one of four patients), but a statistically significant difference was not evident between the two groups. In addition, the presence of PNH neutrophils was not correlated with the response to CyA and/or ATG therapy. These results indicate the usefulness of immunosuppressive therapies even for normo- or hyperplastic MDS patients. Further trials using more patients with a long follow-up period would be worthwhile in order to clarify the possibility of disease progression and in order to predict the response of patients.

Phase II study of cladribine (2-chlorodeoxyadenosine) in relapsed or refractory adult T-cell leukemia-lymphoma

Adult T-cell leukemia-lymphoma (ATL) is a retrovirus-associated T-cell malignancy with an extremely poor prognosis; the median survival time of ATL patients with the acute or lymphoma type is less than 1 year with various combination chemotherapies. Cladribine (2-chlorodeoxyadenosine; 2-CdA), a purine analog resistant to degradation by adenosine deaminase, has shown definitive clinical activity against various lymphoid malignancies, including hairy cell leukemia, indolent lymphoma, and cutaneous T-cell lymphoma. An in vitro study showed the sensitivity of T-lymphoblastoid cell lines to cladribine, and a preceding Japanese phase I study of cladribine showed that 1 refractory patient with ATL achieved an objective response. To evaluate the therapeutic efficacy of cladribine in treating ATL, we conducted a multicenter phase II study. The plan was to administer cladribine to 30 ATL patients as 0.09 mg/kg per day by 7-day continuous intravenous infusion every 4 weeks for up to 3 courses. Before the planned interim analysis, 16 patients with relapsed or refractory ATL were enrolled, 15 of whom were eligible. Only 1 of the 15 eligible patients showed an objective response (overall response rate, 7%; 90% confidence interval, 0% to 28%), and 11 patients (73%) showed progressive disease, mostly during the first course of treatment. Because the upper limit of the 90% confidence interval of the overall response rate did not reach 30% in the interim analysis, the Independent Monitoring Committee advised us to discontinue patient enrollment. In conclusion, cladribine is not worthy of further investigation for the treatment of ATL.

Human granulocyte colony‐stimulating factor receptors in acute myelogenous leukemia

Abstract: Human granulocyte colony‐stimulating factor (G‐CSF) receptors on human acute leukemia cells were investigated using human G‐CSF iodolabeled by the lactoperoxidase method. Among various human leukemic cell lines, only cells of myelogenous lineage including HL‐60, THP‐1 and U937 had one type of high‐affinity receptor for G‐CSF, as shown by Scatchard analysis. Fresh leukemia cells from 19 patients with acute myelogenous leukemia (AML) were then studied. Specific receptors for G‐CSF were demonstrated on blast cells in all 19 cases, the mean number of G‐CSF receptors per AML cell ranging from 95 to 1436. G‐CSF receptors on AML cells appeared to be a single affinity type, although some variations were observed. The mean number of G‐CSF receptors on leukemic cells from patients with either FAB M3 or FAB M2 was greater than that of cells from patients with Ml (p < 0.01, p < 0.10, respectively). Moreover, the mean number of receptors for G‐CSF on CD13‐and CD34‐positive AML cells was higher than that on CD13‐negative and CD34‐positive AML cells (p < 0.01), and the mean number of G‐CSF receptors on CD7‐positive AML cells was lower than that for CD7‐negative AML cells (p < 0.10). Since the FAB classification and surface phenotypes reflect maturation stages, our findings indicate that the distribution of G‐CSF receptors, even on AML cells, may be related to the maturation process.

Fas/Fas ligand and hematopoietic progenitor cells.

Fas antigen is a receptor that crosslinks with a ligand or antibody initiating a signal transduction cascade that leads to apoptosis. During normal hematopoiesis, Fas antigen is not expressed on CD34+ cells, including premature hematopoietic progenitor cells. Functional Fas antigen expression is induced by several hematopoietic regulators. These changes may appear not only in the process of differentiation of hematopoietic progenitor cells, but also as a negative feedback mechanism that controls chaotic proliferation of these cells. These findings suggest that the Fas/Fas ligand system is closely related to the maintenance of homeostasis during the process of normal hematopoiesis. Furthermore, increased Fas antigen expression is observed on CD34+ cells from patients with aplastic anemia, suggesting that it might cause bone marrow suppression. The use of Fas-mediated apoptosis of malignant cells as a tool for eliminating hematologic malignancies is promising. Increased Fas ligand expression is observed on natural killer lymphoma cells and may be associated with the pathogenesis of failure of several organs. The Fas/Fas ligand system plays an important role in the physiologic and pathologic processes of hematopoiesis. The development of treatments using this system are forthcoming.

Acquired Pelger-Huët anomaly in association with concomitant tacrolimus and fluconazole therapy following allogeneic bone marrow transplantation

A 38-year-old Japanese woman with severe aplastic anemia received an allogeneic bone marrow transplant from her serologically HLA-identical father. Cyclosporine and methotrexate were administered to prevent graft-versus-host disease (GVHD). However, grade III acute GVHD developed on day 44, which was successfully treated with methylprednisolone and tacrolimus. Fluconazole therapy was started for oral candidiasis on day 112, but she complained of headache soon after. In addition to glycosuria and increased serum creatinine levels, Pelger–Huët anomaly of granulocytes was found in her blood, which disappeared after discontinuation of tacrolimus. Transient occurrence of Pelger–Huët cells may be associated with tacrolimus toxicity due to drug interaction with fluconazole. Bone Marrow Transplantation (2000) 26, 1255–1257.

Analysis of the granulocyte colony-stimulating factor receptor gene structure using PCR-SSCP in myeloid leukemia and myelodysplastic syndrome

Abstract: The membrane‐proximal cytoplasmic region of the granulocyte colony‐stimulating factor receptor (G–CSFR) is known to be essential for the proliferation signal, with a more distal region being required for the differentiation signal. Such a separation of functional domains raises the possibility that mutations occurring at these regions may contribute to cell proliferation in the absence of differentiation, this being the most important characteristic in acute leukemia cells. Therefore, we analysed the structural abnormalities at the transmembrane and cytoplasmic region of G–CSFR in a significant number of patients with various myeloid malignancies. When we examined the genomic DNA of G–CSFR obtained from 41 patients with acute myelogenous leukemia (AML), 18 with chronic myelogenous leukemia (CML), 7 with myelodysplastic syndrome (MDS), 2 with chronic myelomonocytic leukemia and 1 with chronic neutrophilic leukemia, we found a polymorphism in 3 patients, but no significant pathogenic mutations in any patients. The screening for this polymorphism in 100 hematologically normal controls revealed that it may be useful as a linkage marker for population and family studies, because the heterozygosity index is at a high level (0.055). While there have been several reports discussing the leukemogenic potential of mutations in the cytokine/hematopoietin receptor superfamily, genetic alterations in the transmembrane and cytoplasmic region of G–CSFR do not seem to play a pathogenic role in leukemia.

Effect of Interleukin 10 on the Hematopoietic Progenitor Cells from Patients with Aplastic Anemia

The overproduction of cytokines with inhibitory effects on hematopoiesis is considered to play a role in the pathogenesis of aplastic anemia. While interleukin 10 (IL‐10) is a cytokine production inhibitory factor, the possibility of immunosuppressive therapy using IL‐10 for aplastic anemia has not been explored. In this study, therefore, we examined the effect of IL‐10 on progenitor cells obtained from seven patients with severe aplastic anemia. Our study indicated that IL‐10 dramatically enhanced the erythroid colony formation in a dose‐dependent manner in two of the seven cases examined. When we examined the concentration of cytokines in the culture supernatants of unstimulated bone marrow cells, the spontaneous production of interferon‐γ (IFN‐γ) was observed in one of these two cases, and this production was completely inhibited by addition of IL‐10. These findings suggested that IL‐10 enhanced the erythroid colony formation by inhibiting the pathological production of IFN‐γ in this case. This study provides an experimental support for the clinical application of IL‐10 in some patients with aplastic anemia.