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J-M Luo,
H Yoshida,
S Komura,
N Ohishi, L Pan,
K Shigeno,
I Hanamura,
K Miura,
S Iida,
R Ueda,
T Naoe,
Y Akao,
R Ohno,
K Ohnishi
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ABSTRACT: The SH2 domain-containing inositol 5'-phosphatase (SHIP) is crucial in hematopoietic development. To evaluate the possible tumor suppressor role of the SHIP gene in myeloid leukemogenesis, we examined primary leukemia cells from 30 acute myeloid leukemia (AML) patients, together with eight myeloid leukemia cell lines. A somatic mutation at codon 684, replacing Val with Glu, was detected in one patient, lying within the signature motif 2, which is the phosphatase active site. The results of an in vitro inositol 5'-phosphatase assay revealed that the mutation reduced catalytic activity of SHIP. Leukemia cells with the mutation showed enhanced Akt phosphorylation following IL-3 stimulation. K562 cells transfected with the mutated SHIP-V684E cDNA showed a growth advantage even at lower serum concentrations and resistance to apoptosis induced by serum deprivation and exposure to etoposide. These results suggest a possible role of the mutated SHIP gene in the development of acute leukemia and chemotherapy resistance through the deregulation of the phosphatidylinositol-3,4,5-triphosphate (PI(3,4,5)P3)/Akt signaling pathway. This is the first report of a mutation in the SHIP gene in any given human cancer, and indicates the need for more attention to be paid to this gene with respect to cancer pathogenesis.
Leukemia 02/2003; 17(1):1-8. · 9.56 Impact Factor
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L Pan,
K Ohnishi,
W J Zhang,
H Yoshida,
L Maksumova,
F Muratkhodjaev,
K Shigeno,
S Nakamura,
J M Luo,
H L Hao,
S Fujisawa,
K Naito,
K Shinjo,
A Takeshita,
R Ohno
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ABSTRACT: Interleukin-12 (IL-12) has potent antitumor activities. We examined whether IL-12 enhanced the cytotoxicity of peripheral blood mononuclear cells (PBMNC) and decreased leukemia cells in 30 patients with leukemia or myelodysplastic syndromes (MDS): 12 acute myeloid leukemia (AML) (five in complete remission (CR) and seven in non-CR); six chronic myeloid leukemia (CML); and 12 MDS (three refractory anemia (RA), eight RA with excess of blasts and one chronic myelomonocytic leukemia). PBMNC from patients and five healthy volunteers were cultured at 5 x 10(5)/ml parallel with or without 100 units/ml of IL-12 for 3 days. Cytotoxicity of PBMNC against K562 cells was assessed by flow cytometry. To quantify the amount of leukemia cells, WT1 mRNA was measured by competitive reverse transcription polymerase chain reaction (RT-PCR), since WT1 mRNA is considered as a marker of minimal residual disease (MRD) in leukemia or MDS. The cytotoxicity of non-IL-12-treated PBMNC of 30 patients was 13.4+/-9.3% at the effector to target (E:T) ratio of 20:1, and significantly lower than that of normal subjects (25.7+/-8.4%). The cytotoxicity increased to 30.6+/-17.9% in the IL-12-treated PBMNC. WT1 mRNA in PBMNC of five healthy volunteers was less than 10(3) copies/microg of total RNA. Following the 3-day IL-12 treatment, mean WT1 mRNA of PBMNC was reduced from 10(4.8) to 10(4.2) copies/microg of total RNA in six CML patients, from 10(5.4) to 10(4.8) copies/microg in 12 MDS patients and from 10(5.0) to 10(4.2) copies/microg in five AML patients in CR, but not reduced in five of seven AML in non-CR. These results showed that IL-12 significantly enhanced PBMNC cytotoxicity and decreased the quantity of leukemia cells in PBMNC of most patients with MDS, CML and AML in CR. IL-12 might be of considerable benefit in the elimination of MRD in patients with hematological malignancies.
Leukemia 10/2000; 14(9):1634-41. · 9.56 Impact Factor
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ABSTRACT: Lovastatin, a competitive inhibitor of HMG-CoA reductase, reportedly inhibits proliferation and induces apoptosis of tumor cells with MDR-1 coded P-glycoprotein (Pgp) expression. In this study we investigated the sensitivity to lovastatin of eight myeloid leukemia cell lines: K562, NOMO-1, NB4 and its retinoic acid (RA) resistant subline NB4/RA, and their multidrug-resistant (MDR) sublines: K562/ADR, NOMO-1/ADR, NB4/MDR and NB4/RA/MDR. MTT and apoptosis assays revealed that K562/ADR, NOMO-1/ADR and NB4/RA/MDR were more sensitive to lovastatin than their parental cell lines, while NB4/MDR showed the same level of sensitivity as parental NB4 cells, which already were very sensitive to lovastatin. Significant elevation of transcript levels of HMG-CoA reductase was observed by semiquantitative RT-PCR analysis in more than three lovastatin-sensitive MDR sublines, but not in NB4/MDR compared with the parental cell lines. HMG-CoA reductase mRNA levels were up-regulated more than two-fold by the exposure to lovastatin in all of the parental non-Pgp-expressing cell lines. In NB4/MDR, HMG-CoA reductase mRNA level was elevated to a similar extent as in parental NB4, whereas in three other MDR sublines which showed preferential sensitivity to lovastatin, their HMG-CoA reductase mRNA levels were not significantly elevated after 24- and 48-h treatment with lovastatin. These results indicate a connection between drug resistance and regulation of the mevalonate pathway, and further strengthen the clinical possibility that drug resistant leukemias would be susceptible to treatment with lovastatin.
Leukemia 09/2000; 14(8):1444-50. · 9.56 Impact Factor
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W J Zhang,
K Ohnishi,
H Yoshida, L Pan,
L Maksumova,
F Muratkhodjaev,
J M Luo,
K Shigeno,
S Fujisawa,
K Naito,
S Nakamura,
K Shinjo,
A Takeshita,
R Ohno
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ABSTRACT: Spicamycin is a potent inducer of differentiation of human myeloid leukemia cells (HL-60) and murine myeloid leukemia cells (M1). One of the spicamycin derivatives, KRN5500, shows a broad spectrum of antitumor activity against human tumor xenografts in nude mice. In this study, we first investigated the differentiation efficacy of spicamycin and KRN5500 in HL-60 and acute promyelocytic leukemia cell line, NB4, and found that low concentrations of both compounds induced differentiation to a small extent in both cell lines, but markedly induced apoptosis in NB4 cells. Further investigation in a myeloid leukemia cell line, NKM-1, a lymphoma cell line, Daudi, and a multiple myeloma cell line, NOP-1, showed that high concentrations of both compounds also induced apoptosis in these cells. The 50% inhibitory concentration (IC(50)) determined by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that myeloid cells were more sensitive to both compounds than lymphoid cells, and spicamycin was more potent than KRN5500. Western blot analysis of Bcl-2, Bcl-xL and Bax expression and immunofluorescence analysis of promyelocytic leukemia (PML) protein indicated that apoptosis induced by spicamycin and KRN5500 was associated with down-regulation of Bcl-2 expression and modulation of PML protein. Thus, spicamycin and KRN5500 may be useful for the treatment of myeloid and lymphoid neoplasms.
Japanese journal of cancer research: Gann 07/2000; 91(6):604-11.
