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1.
FEBS Open Bio ; 8(12): 1977-1991, 2018 Dec.
Article in English | MEDLINE | ID: mdl-30524948

ABSTRACT

Chromosome band 8q24 is the most frequently amplified locus in various types of cancers. MYC has been identified as the primary oncogene at the 8q24 locus, whereas a long noncoding gene, PVT1, which lies adjacent to MYC, has recently emerged as another potential oncogenic regulator at this position. In this study, we established and characterized a novel cell line, AMU-ML2, from a patient with diffuse large B-cell lymphoma (DLBCL), displaying homogeneously staining regions at the 8q24 locus. Fluorescence in situ hybridization clearly detected an elevation in MYC copy numbers corresponding to the homogenously staining region. In addition, a comparative genomic hybridization analysis using high-resolution arrays revealed that the 8q24 amplicon size was 1.4 Mb, containing the entire MYC and PVT1 regions. We also demonstrated a loss of heterozygosity for TP53 at 17p13 in conjunction with a TP53 frameshift mutation. Notably, AMU-ML2 cells exhibited resistance to vincristine, and cell proliferation was markedly inhibited by MYC-shRNA-mediated knockdown. Furthermore, genes involved in cyclin D, mTOR, and Ras signaling were downregulated following MYC knockdown, suggesting that MYC expression was closely associated with tumor cell growth. In conclusion, AMU-ML2 cells are uniquely characterized by homogenously staining regions at the 8q24 locus, thus providing useful insights into the pathogenesis of DLBCL with 8q24 abnormalities.

2.
Int J Hematol ; 102(5): 569-78, 2015 Nov.
Article in English | MEDLINE | ID: mdl-26341959

ABSTRACT

Amylase-producing myeloma exhibits refractoriness to chemotherapy and a dismal prognosis. In this study, we established a human myeloma cell line, 8226/AMY1, in which a lentivirally transfected AMY1 gene was stably expressed and explored its biological characteristics. 8226/AMY1 showed a survival advantage over mock control when treated with dexamethasone, bortezomib, and lenalidomide in vitro partly through inhibition of apoptosis induced by these reagents. In a xenograft murine model, 8226/AMY1 showed rapid tumor growth and reduced sensitivity to bortezomib compared with mock. A microarray gene expression analysis identified TCL1A, which functions as a coactivator of the cell survival kinase Akt, differentially up-regulated in 8226/AMY1. The expression of phosphorylated Akt was increased in the 8226/AMY1 cells following bortezomib treatment, but not in the mock cells. In addition, treatment with perifosine, an inhibitor of Akt phosphorylation, enhanced the anti-myeloma effect of bortezomib in the 8226/AMY1 cells. Our data suggest that amylase-producing myeloma reduced the sensitivity to bortezomib in vitro and in vivo, and the up-regulation of TCL1A may influence the drug susceptibility of 8226/AMY1 via the phosphorylation of Akt. These findings provide clues for developing treatment approaches for not only amylase-producing myeloma, but also relapsed and refractory myelomas.


Subject(s)
Amylases/biosynthesis , Bortezomib/pharmacology , Drug Resistance, Neoplasm/drug effects , Gene Expression Regulation, Enzymologic/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Multiple Myeloma , Animals , Cell Line, Tumor , Humans , Mice , Mice, Inbred NOD , Mice, SCID , Multiple Myeloma/drug therapy , Multiple Myeloma/enzymology , Multiple Myeloma/genetics , Proto-Oncogene Proteins/biosynthesis , Proto-Oncogene Proteins c-akt/metabolism , Xenograft Model Antitumor Assays
3.
Leuk Res ; 37(9): 1132-6, 2013 Sep.
Article in English | MEDLINE | ID: mdl-23806233

ABSTRACT

We examined the effects of diet nutrients on xenotransplanted leukemia cells, THP-1 or NB4. THP-1 tumors showed more growth when fed with high fat diet, while NB4 tumors grew more with high carbohydrate diet. Then, administration of 2-deoxyglucose (a glycolysis inhibitor) showed a significant antitumor effect on both tumors: NB4 tumor showed large necrotic areas, while THP-1 tumor did not, but had augmented expression of enzymes for fatty acid oxidation. 2-Deoxyglucose inhibited the growth of NB4 by cell death because main energy producing pathway (glycolysis) was abolished, while 2-deoxyglucose slowed the growth of THP-1 by shifting energy metabolism to fatty acid ß-oxidation.


Subject(s)
Antimetabolites/pharmacology , Cell Proliferation , Deoxyglucose/pharmacology , Diet , Dietary Supplements , Leukemia, Experimental/drug therapy , Animals , Blotting, Western , Energy Metabolism/drug effects , Female , Glycolysis/drug effects , Humans , Immunoenzyme Techniques , Leukemia, Experimental/metabolism , Mice , Mice, Inbred NOD , Mice, SCID , Neoplasm Transplantation , Transplantation, Heterologous , Tumor Cells, Cultured
4.
Oncol Rep ; 29(5): 2053-7, 2013 May.
Article in English | MEDLINE | ID: mdl-23440281

ABSTRACT

The shift in energy metabolism from oxidative phosphorylation to glycolysis can serve as a target for the inhibition of cancer growth. Here, we examined the metabolic changes induced by 2-deoxyglucose (2-DG), a glycolysis inhibitor, in leukemia cells by metabolome analysis. NB4 cells mainly utilized glucose as an energy source by glycolysis and oxidative phosphorylation in mitochondria, since metabolites in the glycolytic pathway and in the tricarboxylic acid (TCA) cycle were significantly decreased by 2-DG. In THP-1 cells, metabolites in the TCA cycle were not decreased to the same extent by 2-DG as in NB4 cells, which indicates that THP-1 utilizes energy sources other than glucose. TCA cycle metabolites in THP-1 cells may be derived from acetyl-CoA by fatty acid ß-oxidation, which was supported by abundant detection of carnitine and acetylcarnitine in THP-1 cells. 2-DG treatment increased the levels of pentose phosphate pathway (PPP) metabolites and augmented the generation of NADPH by glucose-6-phosphate dehydrogenase. An increase in NADPH and upregulation of glutathione synthetase expression resulted in the increase in the reduced form of glutathione by 2-DG in NB4 cells. We demonstrated that a combination of 2-DG and inhibition of PPP by dehydroepiandrosterone (DHEA) effectively suppressed the growth of NB4 cells. The replenishment of the TCA cycle by fatty acid oxidation by carnitine palmitoyltransferase in THP-1 cells, treated by 2-DG, might be regulated by AMPK, as the combination of 2-DG and inhibition of AMPK by compound C potently suppressed the growth of THP-1 cells. Although 2-DG has been effective in preclinical and clinical studies, this treatment has not been fully explored due to concerns related to potential toxicities such as brain toxicity at high doses. We demonstrated that a combination of 2-DG and DHEA or compound C at a relatively low concentration effectively inhibits the growth of NB4 and THP-1 cells, respectively. These observations may aid in the identification of appropriate combinations of metabolic inhibitors at low concentrations which do not cause toxicities.


Subject(s)
Deoxyglucose/pharmacology , Energy Metabolism/drug effects , Leukemia/metabolism , AMP-Activated Protein Kinases/metabolism , Acetyl Coenzyme A/metabolism , Carnitine O-Palmitoyltransferase/metabolism , Cell Line, Tumor , Citric Acid Cycle/drug effects , Dehydroepiandrosterone/pharmacology , Fatty Acids/metabolism , Glucose/metabolism , Glucosephosphate Dehydrogenase/metabolism , Glutathione/metabolism , Glycolysis/drug effects , Humans , Metabolome/drug effects , Mitochondria/drug effects , Mitochondria/metabolism , NADP/metabolism , Oxidation-Reduction/drug effects , Oxidative Phosphorylation/drug effects , Pentose Phosphate Pathway/drug effects , Pentose Phosphate Pathway/physiology , Pyrazoles/pharmacology , Pyrimidines/pharmacology
5.
Genes Chromosomes Cancer ; 51(1): 42-53, 2012 Jan.
Article in English | MEDLINE | ID: mdl-21965128

ABSTRACT

In this study, we established and analyzed a novel human myeloid leukemia cell line, AMU-AML1, from a patient with acute myeloid leukemia with multilineage dysplasia before the initiation of chemotherapy. AMU-AML1 cells were positive for CD13, CD33, CD117, and HLA-DR by flow cytometry analysis and showed a single chromosomal abnormality, 46, XY, t(12;22)(p13;q11.2), by G-banding and spectral karyotyping. Fluorescent in situ hybridization analysis indicated that the chromosomal breakpoint in band 12p13 was in the sequence from the 5' untranslated region to intron 1 of TEL and that the chromosomal breakpoint in band 22q11 was in the 3' untranslated region of MN1. The chimeric transcript and protein of MN1-TEL could not be detected by reverse-transcriptase polymerase chain reaction or Western blot analysis. However, the MN1 gene was amplified to three copies detected by array comparative genomic hybridization analysis, and the expression levels of the MN1 transcript and protein were high in AMU-AML1 cells when compared with other cell lines with t(12;22)(p13;q11-12). Our data showed that AMU-AML1 cells contain t(12;22)(p13;q11.2) without chimeric fusion of MN1 and TEL. The AMU-AML1 cells gained MN1 copies and had high expression levels of MN1. Thus, the AMU-AML1 cell line is useful for studying the biological consequences of t(12;22)(p13;q11.2) lacking chimeric MN1-TEL.


Subject(s)
Cell Line, Tumor , Chromosomes, Human, Pair 12 , Chromosomes, Human, Pair 22 , Leukemia, Myeloid/genetics , Oncogene Proteins, Fusion/genetics , Transcription Factors/genetics , Translocation, Genetic , Tumor Suppressor Proteins/genetics , Chromosome Banding , Chromosome Breakpoints , Comparative Genomic Hybridization , Gene Expression , Gene Expression Regulation, Leukemic , Gene Order , Humans , Immunophenotyping , In Situ Hybridization, Fluorescence , Leukemia, Myeloid/metabolism , Leukemia, Myeloid/pathology , Male , Middle Aged , Oncogene Proteins, Fusion/metabolism , Spectral Karyotyping , Trans-Activators , Transcription Factors/metabolism , Transcription, Genetic , Tumor Suppressor Proteins/metabolism
6.
Leuk Lymphoma ; 51(11): 2112-9, 2010 Nov.
Article in English | MEDLINE | ID: mdl-20860495

ABSTRACT

For generation of energy, cancer cells utilize glycolysis more vigorously than oxidative phosphorylation in mitochondria (Warburg effect). We examined the energy metabolism of four leukemia cell lines by using glycolysis inhibitor, 2-deoxy-d-glucose (2-DG) and inhibitor of oxidative phosphorylation, oligomycin. NB4 was relatively sensitive to 2-DG (IC(50): 5.75 mM), consumed more glucose and produced more lactate (waste product of glycolysis) than the three other cell lines. Consequently, NB4 was considered as a "glycolytic" leukemia cell line. Dependency on glycolysis in NB4 was confirmed by the fact that glucose (+) FCS (-) medium showed more growth and survival than glucose (-) FCS (+) medium. Alternatively, THP-1, most resistant to 2-DG (IC(50): 16.14 mM), was most sensitive to oligomycin. Thus, THP-1 was recognized to be dependent on oxidative phosphorylation. In THP-1, glucose (-) FCS (+) medium showed more growth and survival than glucose (+) FCS (-) medium. The dependency of THP-1 on FCS was explained, at least partly, by fatty acid oxidation because inhibitor of fatty acid ß-oxidation, etomoxir, augmented the growth suppression of THP-1 by 2-DG. We also examined the mechanisms by which THP-1 was resistant to, and NB4 was sensitive to 2-DG treatment. In THP-1, AMP kinase (AMPK), which is activated when ATP becomes limiting, was rapidly phosphorylated by 2-DG, and expression of Bcl-2 was augmented, which might result in resistance to 2-DG. On the other hand, AMPK phosphorylation and augmentation of Bcl-2 expression by 2-DG were not observed in NB4, which is 2-DG sensitive. These results will facilitate the future leukemia therapy targeting metabolic pathways.


Subject(s)
Energy Metabolism/physiology , Glycolysis/physiology , Leukemia/metabolism , Oxidative Phosphorylation , Antimetabolites/metabolism , Antimetabolites/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Deoxyglucose/metabolism , Deoxyglucose/pharmacology , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/physiology , Energy Metabolism/drug effects , Glucose/metabolism , Glycolysis/drug effects , HL-60 Cells , Humans , Lactic Acid/metabolism , Leukemia/pathology , Oligomycins/pharmacology , Oxidative Phosphorylation/drug effects , Uncoupling Agents/pharmacology
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