Papers - IKEDA Ryuji
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2-Deoxy-D-ribose inhibits hypoxia-induced apoptosis by suppressing the phosphorylation of p38 MAPK
Ikeda R., Che X., Ushiyama M., Yamaguchi T., Okumura H., Nakajima Y., Takeda Y., Shibayama Y., Furukawa T., Yamamoto M., Haraguchi M., Sumizawa T., Yamada K., Akiyama S.
Biochemical and Biophysical Research Communications 342 ( 1 ) 280 - 285 2006.3
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Biochemical and Biophysical Research Communications
An angiogenic factor, platelet-derived endothelial cell growth factor/thymidine phosphorylase (TP), stimulates the chemotaxis of endothelial cells and confers resistance to apoptosis induced by hypoxia. 2-Deoxy-D-ribose, a degradation product of thymidine generated by TP enzymatic activity, partially prevented hypoxia-induced apoptosis. 2-Deoxy-D-ribose inhibited hypoxia-induced phosphorylation of p38 mitogen-activated protein kinase (MAPK) but not c-jun NH2-terminal kinase/stress-activated protein kinase in human leukemia HL-60 cells. 2-Deoxy-D-ribose also suppressed the levels of Bax attached to mitochondria under hypoxic conditions. SB203580, a specific inhibitor of the p38 MAPK, suppressed the hypoxia-induced apoptosis of HL-60 cells. These findings suggest that one of the molecular bases for resistance to hypoxia-induced apoptosis conferred by 2-deoxy-D-ribose is the inhibition of the p38 signaling pathway. The expression levels of TP are elevated in many malignant solid tumors and thus the 2-deoxy-D-ribose generated by TP in these tumors may play an important role in tumor progression by preventing hypoxia-induced apoptosis. © 2006 Elsevier Inc. All rights reserved.
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Onimaru S., Nakamura K., Kariyazono H., Ikeda R., Ueno T., Fukumoto Y., Yabuki A., Sakata R., Yamada K.
Heart and Vessels 21 ( 2 ) 108 - 115 2006.3
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Heart and Vessels
We evaluated the effects of edaravone, a hydroxyl radical scavenging agent, on the production of tumor necrosis factor-α (TNF-α) in myocardium, and the release of TNF-α and P-selectin from myocardium after ischemia-reperfusion injury in isolated Langendorff-perfused rat hearts. Cardiodynamic function at stable points during perfusion and 5, 15, 30, and 60min after the initiation of reperfusion was evaluated by left ventricular developed pressure, rate of increase in left ventricular pressure and rate of decrease in ventricular pressure, coronary flow, and heart rate. At 60min after the initiation of reperfusion, myocardial infarct size was estimated microscopically using triphenyltetrazolium chloride staining, and expression of TNF-α in myocardium was detected by Western blot and immunohistochemistry. At the same time points as the measurement of cardiodynamic function, TNF-α and the soluble form of P-selectin in coronary effluent were measured by enzyme immunoassay. At all time points during reperfusion, edaravone markedly improved cardiodynamic function and reduced myocardial infarct size in comparison to the control. In myocardium in the control, TNF-α was detected in the endothelial cells and other cells bearing some resemblance to interstitial cells and monocyte cells. Edaravone suppressed this cytokine expression in the corresponding sites. P-selectin as well as TNF-α was found in the coronary effluent of the control, and edaravone significantly decreased soluble P-selectin levels in comparison to the control (P < 0.01). Edaravone might have protective effects on cardiac function through reduction of infarct size via decrease of production of TNF-α in myocardium induced by ischemia-reperfusion injury and through reduction of the release of adhesion molecules such as P-selectin from vascular endothelial cells. © Springer-Verlag Tokyo 2006.
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Hayashi R., Goto Y., Haga A., Kobayashi D., Ikeda R., Yoshida K.
Gene 367 ( 1-2 ) 126 - 134 2006.2
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Gene
Minichromosome maintenance protein (MCM) is composed of six structurally related subunits (MCM2-7) and is essential for eukaryotic DNA replication initiation and early stage of elongation process. Recently human and Xenopus MCM8 was identified as a novel member of MCM protein. Here we characterized MCM8 orthologous genes by using bioinformatics. Human MCM8 showed approximately 90%, 90%, 93%, and 79% total-amino acid identity with mouse, rat, dog, and chicken MCM8, respectively. Human, mouse, rat, dog, and chicken MCM8 gene, consisting of 19, 18, 17, 18, and 18 exons, was mapped to 20p12.3-13, 2F3, 3q36, 24, and 3, respectively. We identified transcription factor E2F-binding motifs in the vicinity of the transcription start site among MCM8 orthologous genes. The mammalian but not chicken E2F-binding motif was accompanied by NF-Y binding motif. MCM8 mRNA was upregulated by E2E1 in human culture cells. Chromatin immunoprecipitation (ChIP) demonstrated the direct association of E2F1 and NF-Y with human MCM8 promoter. The promoter activities of human, rat, and chicken MCM8 were demonstrated to be E2F1-dependent. Analysis of human MCM8 promoter constructs showed that an E2F-binding motif in the vicinity of the transcription initiation site is necessary for the transcriptional activation. We also showed that the transcription of human MCM8 is activated by transcription factors E2F1-4, but not by factors E2F5-8.
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Cepharanthine potently enhances the sensitivity of anticancer agents in K562 cells
Ikeda R., Che X., Yamaguchi T., Ushiyama M., Zheng C., Okumura H., Takeda Y., Shibayama Y., Nakamura K., Jeung H., Furukawa T., Sumizawa T., Haraguchi M., Akiyama S., Yamada K.
Cancer Science 96 ( 6 ) 372 - 376 2005.6
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Cancer Science
A major impediment to cancer treatment is the development of resistance by the tumor. P-glycoprotein (P-gp) and multidrug resistance protein 1 (MRP1) are involved in multidrug resistance. In addition to the extrusion of chemotherapeutic agents through these transporters, it has been reported that there are differences in the intracellular distribution of chemotherapeutic agents between drug resistant cells and sensitive cells. Cepharanthine is a plant alkaloid that effectively reverses resistance to anticancer agents. It has been previously shown that cepharanthine is an effective agent for the reversal of resistance in P-gp-overexpressing cells. Cepharanthine has also been reported to have numerous pharmacological effects besides the inhibition of P-gp. It has also been found that cepharanthine enhanced sensitivity to doxorubicin (ADM) and vincristine (VCR), and enhanced apoptosis induced by ADM and VCR of P-gp negative K562 cells. Cepharanthine changed the distribution of ADM from cytoplasmic vesicles to nucleoplasm in K562 cells by inhibiting the acidification of cytoplasmic organelles. Cepharanthine in combination with ADM should be useful for treating patients with tumors. © Japanese Cancer Association.
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Ikeda R., Yoshida K., Tsukahara S., Sakamoto Y., Tanaka H., Furukawa K., Inoue I.
Journal of Biological Chemistry 280 ( 9 ) 8523 - 8530 2005.3
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Journal of Biological Chemistry
Ossification of the posterior longitudinal ligament of the spine (OPLL) is the leading cause of myelopathy in Japan and is diagnosed by ectopic bone formation in the paravertebral ligament. OPLL is a systemic high bone mass disease with a strong genetic background. To detect genes relevant to the pathogenesis of OPLL, we performed a cDNA microarray analysis of systematic gene expression profiles during the osteoblastic differentiation of ligament cells from OPLL patients (OPLL cells), patients with a disorder called ossification of yellow ligament (OYL), and non-OPLL controls together with human mesenchymal stem cells (hMSCs) after stimulating them with osteogenic differentiation medium (OS). Twenty-four genes were up-regulated during osteoblastic differentiation in OPLL cells. Zinc finger protein 145 (promyelotic leukemia zinc finger or PLZF) was one of the highly expressed genes during osteoblastic differentiation in all the cells examined. We investigated the roles of PLZF in the regulation of osteoblastic differentiation of hMSCs and C2C12 cells. Small interfering RNA-mediated gene silencing of PLZF resulted in a reduction in the expression of osteoblast-specific genes such as the alkaline phosphatase, collagen 1A1 (Col1a1), Runx2/core-binding factor 1 (Cbfa1), and osteocalcin genes, even in the presence of OS in hMSCs. The expression of PLZF was unaffected by the addition of bone morphogenetic protein 2 (BMP-2), and the expression of BMP-2 was not affected by PLZF in hMSCs. In C2C12 cells, overexpression of PLZF increased the expression of Cbfa1 and Col1a1; on the other hand, the overexpression of CBFA1 did not affect the expression of Plzf. These findings indicate that PLZF plays important roles in early osteoblastic differentiation as an upstream regulator of CBFA1 and thereby might participate in promoting the ossification of spinal ligament cells in OPLL patients. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.
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Effects of acute and chronic cyclosporin treatment on digoxin pharmacokinetics
Shibayama Y., Kawachi A., Ikeda R., Motoya T., Yamada K.
Journal of Applied Therapeutic Research 4 ( 4 ) 38 - 45 2004.12
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Journal of Applied Therapeutic Research
Administration of the immunosuppressant cyclosporin A (CsA) increases digoxin (DIG) blood levels through inhibition of P-glycoprotein (P-gp). On the other hand, chronic treatment with CsA has been reported to induce overexpression of P-gp. Therefore, chronic treatment with CsA may change the pharmacokinetic profile of DIG. We report here the experimental results of acute and chronic CsA treatment on the pharmacokinetics of DIG. Rats were treated with CsA once daily for 10 consecutive days. Acute and chronic treatment with CsA was found to increase the area under the blood concentration-time curve for DIG (AUC, acute: 42 ± 3 to 195 ± 30, chronic: 52 ± 2 to 206 ± 30 μg·h/l) and peak blood DIG levels (Cmax, acute: 8.8 ± 0.5 to 23.7 ± 5.4, chronic: 7.2 ± 0.6 to 20.6 ± 2.4 ng/ml). Overexpression of the renal P-gp induced about a 1.4-fold difference between control and chronic treatment with CsA. From the results, the inhibition of P-gp by CsA appears to be a potent determinant on the pharmacokinetic profile of DIG in cases of concomitant administration. Thus, caution is required when using CyA and DIG concomitantly to avoid a potential interaction.
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Shibayama Y., Ikeda R., Motoya T., Yamada K.
Food and Chemical Toxicology 42 ( 6 ) 995 - 1002 2004.6
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Food and Chemical Toxicology
St. John's Wort (Hypericum perforatum, SJW) has been used as a herbal medicine for the treatment of depression in oral doses of 900-1050 mg/day in humans. However, the ingestion of SJW was reported to cause interactions with drugs. In the present study, we examined the effects of SJW treatment on the induction of drug transporters and enzymes in rats. An immunoblot analysis was performed to quantify the expression of the transporters and enzymes. SJW was given at a dose of 400 mg/kg/day, since it was reported that 400 mg/kg/day is antidepressant effective dose in rats. When SJW was administered for 10 days, the amounts of multidrug resistance protein 2 (MRP2), glutathione S-transferase-P (GST-P) and cytochrome P450 1A2 (CYP1A2) in the liver were increased to 304%, 252% and 357% of controls, respectively, although the amounts of P-glycoprotein and multidrug resistance protein 1 were not changed. Under the same conditions, an increase of MRP2 in the kidney was not observed. The increase in the levels of each protein was maximal at 10 days after SJW treatment and lasted for at least 30 consecutive days. These results suggest that SJW induces hepatic MRP2, GST-P and CYP1A2 overexpressions, and thus, it could affect drug metabolism, conjugation and disposition. © 2004 Elsevier Ltd. All rights reserved.
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Inhibition of Metastasis of Tumor Cells Overexpressing Thymidine Phosphorylase by 2-Deoxy-L-Ribose
Nakajima Y., Gotanda T., Uchimiya H., Furukawa T., Haraguchi M., Ikeda R., Sumizawa T., Yoshida H., Akiyama S.
Cancer Research 64 ( 5 ) 1794 - 1801 2004.3
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Cancer Research
Thymidine phosphorylase (TP) catalyzes the reversible conversion of thymidine to thymine, thereby generating 2-deoxy-D-ribose-1-phosphate, which upon dephosphorylation forms 2-deoxy-D-ribose (D-dRib), a degradation product of thymidine. We have previously shown that D-dRib promotes angiogenesis and chemotaxis of endothelial cells and also confers resistance to hypoxia-induced apoptosis in some cancer cell lines. 2-Deoxy-L-ribose (L-dRib), a stereoisomer of D-dRib, can inhibit D-dRib anti-apoptotic effects and suppressed the growth of KB cells overexpressing TP (KB/TP cells) transplanted into nude mice. In this study, we examined the ability of L-dRib to suppress metastasis of KB/TP cells using two different models of metastasis. The antimetastatic effect of L-dRib was first investigated in a liver-metastasis model in nude mice inoculated with KB/TP cells. Oral administration of L-dRib for 28 days at a dose of 20 mg/kg/day significantly reduced the number of metastatic nodules in the liver and suppressed angiogenesis and enhanced apoptosis in KB/TP metastatic nodules. Next, we compared the ability of L-dRib and tegafur alone or in combination to decrease the number of metastatic nodules in organs in the abdominal cavity in nude mice receiving s.c. of KB/TP cells into their backs. L-dRib (20 mg/kg/day) was significantly (P < 0.05) more efficient than tegafur (100 mg/kg/day) in decreasing the number of metastatic nodules in organs in the abdominal cavity. By in vitro invasion assay, L-dRib also reduced the number of invading KB/TP cells. L-dRib anti-invasive activity may be mediated by its ability to suppress the enhancing effect of TP and D-dRib on both mRNA and protein expression of vascular endothelial growth factor and interleukin-8 in cultured KB cells. These findings suggest that L-dRib may be useful in a clinical setting for the suppression of metastasis of tumor cells expressing TP.
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Thymidine phosphorylase inhibits apoptosis induced by anticancer agents
Ikeda R., Furukawa T., Sumizawa T., Haraguchi M., Oiso S., Inoue I., Yamada K., Akiyama S.
Folia Pharmacologica Japonica 122 ( SUPPL. 1 ) 2003.12
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Folia Pharmacologica Japonica
An angiogenic factor, platelet-derived endothelial cell growth factor/thymidine phosphorylase (PD-ECGF/TP), stimulates the chemotaxis of endothelial cells and confers resistance to apoptosis induced by hypoxia. 2-Deoxy-D-ribose, a degradation product of thymidine generated by TP enzymatic activity, partially prevented hypoxia-induced apoptoisis. TP was expressed at higher levels in tumor tissuses compared to the adjacent non-neoplastic tissues in a variety of human carcinomas. High expression of TP is associated with an unfavorable prognosis. To investigate the effect of TP on cisplatin-induced apoptosis, human leukemia Jurkat cells were transfected wild-type or mutant (L148R) TP cDNA. TP inhibits a number of steps in the cisplatin-induced apoptotic pathway, activation of caspase 3, 9 and mitochondrial cytochrome c release. These findings suggest a mechanism by which TP confers the resistance to apoptosis by cisplatin. Moreover, mutant TP that has no enzymatic activity also suppressed cisplatin-induced apoptosis. These findings indicate that TP has cytoprotective functions against cytotoxic agents which are independent of its enzymatic activity.
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Sata T., Ishida K., Motoya T., Nakano R., Honda K., Nakao S., Yamashita K., Iwashita Y., Ikeda R., Yamada K.
Journal of Applied Therapeutic Research 4 ( 2 ) 40 - 45 2003.12
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Journal of Applied Therapeutic Research
The usefulness of drug information leaflets with pictures for elderly patient education to facilitate understanding of their medicines was examined. Sixty inpatients, more than 65 years old, were divided into three groups of 20 patients each. Each group was provided drug information such as trade name, efficacy, administration procedure and cautions for use by one of the following three procedures: i) oral explanation; ii) oral explanation and provision of information written on the package; iii) oral explanation and provision of an information leaflet with pictures. Understanding by patients of their medicines was assessed by an interview with a pharmacist. The patients given drug information leaflets with pictures received significantly (p<0.05) higher understanding scores in all of the information items examined than those in the other groups. After withdrawal of education for 6 months, the patients who had kept the leaflet showed significantly (p<0.05) higher understanding scores than those who had lost it. The score in the latter patient group after the beginning of the second period of education, however, improved faster than that after the first. These results indicate that the use of drug information leaflets with pictures is an effective way to improve the understanding by elderly patients of their medicines.
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Aspirin induces hepatoma-derived cell apoptosis via a hydrogen peroxide-dependent pathway
Tuvdendorj D., Oketani M., Ikeda R., Kohara K., Komorizono Y., Ishibashi K., Munkhtuvshin N., Arima T.
Hepatology Research 26 ( 1 ) 47 - 54 2003.5
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Hepatology Research
Here we studied whether aspirin (ASA) has any influence on viability of human hepatoma-derived SKHep-1 cells and whether hydrogen peroxide (H2O2) has any relation with this effect. ASA inhibited SKHep-1 cell proliferation dose- and time-dependently. Intracellular H2O2increased as early as 15 min after ASA supplementation. Cellular apoptosis correlated with an increase in intracellular H2O2level. Moreover, in the presence of a catalase inhibitor-aminotriazol, ASA showed more apoptotic effect on SKHep-1 cells with increasing intracellular H2O2level. In conclusion, the present results shows that ASA induced SKHep-1 cell apoptosis has a relation with an early increase in intracellular H2O2level and catalase inhibitor synergizes to induce this process. © 2003 Elsevier Science B.V. All rights reserved.
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Thymidine phosphorylase inhibits apoptosis induced by cisplatin
Ikeda R., Furukawa T., Mitsuo R., Noguchi T., Kitazono M., Okumura H., Sumizawa T., Haraguchi M., Che X., Uchimiya H., Nakajima Y., Ren X., Oiso S., Inoue I., Yamada K., Akiyama S.
Biochemical and Biophysical Research Communications 301 ( 2 ) 358 - 363 2003.2
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Biochemical and Biophysical Research Communications
An angiogenic factor, platelet-derived endothelial cell growth factor/thymidine phosphorylase (PD-ECGF/TP), stimulates the chemotaxis of endothelial cells and confers resistance to apoptosis induced by hypoxia. 2-Deoxy-D-ribose, a degradation product of thymidine generated by TP, partially prevents hypoxia-induced apoptosis. TP is expressed at higher levels in tumor tissues compared to the adjacent non-neoplastic tissues in a variety of human carcinomas. High expression of TP is associated with an unfavorable prognosis. To investigate the effect of TP on cisplatin-induced apoptosis, human leukemia Jurkat cells were transfected with wild-type or mutant (L148R) TP cDNA. TP inhibited a number of steps in the cisplatin-induced apoptotic pathway, activation of caspases 3 and 9 and mitochondrial cytochrome c release. These findings suggest a mechanism by which TP confers resistance to apoptosis induced by cisplatin. Moreover, mutant TP that has no enzymatic activity also suppressed cisplatin-induced apoptosis. These findings indicate that TP has cytoprotective functions against cytotoxic agents which are independent of its enzymatic activity. © 2003 Elsevier Science (USA). All rights reserved.
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Che X., Nakajima Y., Sumizawa T., Ikeda R., Ren X., Zheng C., Mukai M., Furukawa T., Haraguchi M., Gao H., Sugimoto Y., Akiyama S.
Cancer Letters 187 ( 1-2 ) 111 - 119 2002.12
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Cancer Letters
A newly synthesized 1,4-benzothiazipine derivate, 4-[3-(4-benzylpiperidin-1-yl) propionyl]-7-methoxy-2,3,4,5-tetrahydro-1, 4-benzothiazepine monohydrochloride (JTV-519) was examined for its ability to reverse P-glycoprotein (P-gp) and multidrug resistance protein 1 (MRP1) mediated multidrug resistance (MDR) in K562/MDR and KB/MRP cells, respectively. JTV-519 at 3μM reversed the resistance of K562/MDR cells to vincristine (VCR), taxol, etoposide (VP16), adriamycin (ADM) and actinomycin D and at 0.5 or 1μM reversed their resistance to STI571. JTV-519 at 10μM enhanced the accumulation of ADM in K562/MDR cells to the level in parental K562 cells and inhibited the efflux of ADM from K562/MDR cells. Photoaffinity labeling of P-gp with3H-azidopine was almost completely inhibited by 500μM JTV-519. JTV-519 at 3μM also partially reversed the resistance of KB/MRP cells to VCR and at 500μM partially inhibited the photoaffinity labeling of MRP1 with125I-II-azidophenyl agosterol A (125I-azidoAG-A). These results suggest that JTV-519 reversed the resistance to the anti-cancer agents in P-gp and MRP1 overexpressing multidrug-resistant cells by directly binding to P-gp and MRP1, and competitively inhibiting transport of the anti-cancer agents. © 2002 Elsevier Science Ireland Ltd. All rights reserved.
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Ren X., Furukawa T., Aoki S., Sumizawa T., Haraguchi M., Nakajima Y., Ikeda R., Kobayashi M., Akiyama S.
Biochemistry 41 ( 48 ) 14132 - 14140 2002.12
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Biochemistry
MRP1 is a 190 kDa membrane glycoprotein that confers multidrug resistance (MDR) to tumor cells. Our recent study demonstrated that GSH is required for the labeling of MRP1932-1531 with a photoanalogue of agosterol A (AG-A) and suggested that GSH interacts with the L0 region of MRP1. In this study, we further characterized the GSH-dependent binding site of azido AG-A on MRP1. Coexpression of the N- and C-terminal halves of MRP1 (residues 1-1222, TM1-16, and 1223-1531, TM17, respectively) in Sf21 insect cells reconstituted a functional drug transporter with a Km for LTC4 (97 nM) similar to that of intact MRP1. In membrane vesicles from those cells, GSH-dependent photolabeling of the MRP1 fragment (1-1222) required the coexpression of the C-terminal MRP1 fragment (1223-1531). An MRP1 fragment extending from residue 1 to 1295 however could be photolabeled by azido AG-A in a GSH-dependent manner. These data indicate that amino acids 1223-1295 of MRP1 are required for AG-A binding to MRP1 in a GSH-dependent manner. However, cross-linking of the photolabel to MRP1 occurs at a more upstream site. An arginine residue at position 1249 of MRP1 was shown to be important for the GSH-dependent binding of AG-A to MRP1. Mutation of this arginine to alanine (R1249A) resulted in a decreased level of GSH-dependent azido AG-A photolabeling of MRP1. Furthermore, this mutant attenuated MRP1 function by decreasing the level of LTC4 substrate transport and impairing resistance to the drug vincristine (VCR). In summary, this study demonstrates that a region of MRP1 (amino acids 1223-1295), which includes TM helix 17, is required for azido AG-A binding to MRP1 in a GSH-dependent manner. A GSH-dependent drug binding site may exist in this region. Furthermore, our findings suggest that the charged amino acid Arg1249 proximal to the C-terminus of TM helix 17 is indispensable for MRP1-substrate interaction and the function of MRP1.
DOI： 10.1021/bi026443s
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Suppression of thymidine phosphorylase-mediated angiogenesis and tumor growth by 2-deoxy-L-ribose
Uchimiya H., Furukawa T., Okamoto M., Nakajima Y., Matsushita S., Ikeda R., Gotanda T., Haraguchi M., Sumizawa T., Ono M., Kuwano M., Kanzaki T., Akiyama S.
Cancer Research 62 ( 10 ) 2834 - 2839 2002.5
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Cancer Research
Thymidine phosphorylase (TP), an enzyme involved in the reversible conversion of thymidine to thymine, is identical to an angiogenic factor, platelet-derived endothelial cell growth factor (PD-ECGF). Both TP and one of the TP-degradation products of thymidine 2-deoxy-D-ribose (dRib) display endothelial cell chemotactic activity in vitro and angiogenic activity in vivo. Recently, we demonstrated that 2-deoxy-L-ribose (IRib) could abolish the inhibitory effect of dRib on hypoxia-induced apoptosis. This suggested that IRib may be a useful inhibitor of dRib and thereby of TP functions. Therefore, we investigated the ability of IRib to inhibit the range of biological activities of TP and dRib. IRib suppressed both dRib-induced endothelial cell migration in a chemotaxis assay and endothelial tube formation induced by dRib in a collagen gel. IRib could also suppress the biological effects of TP in vivo assays of angiogenesis and tumor growth. Thus, in a corneal assay of angiogenesis, IRib inhibited angiogenesis induced by the implantation of recombinant TP. In a dorsal air sac assay of angiogenesis, IRib inhibited angiogenesis induced by the implantation of KB cells overexpressing TP (KB/TP). In a tumor growth assay, IRib treatment considerably decreased the growth rate of KB/TP cells xenografted into nude mice and also resulted in an increase in the proportion of apoptotic cells in KB/TP tumors. These findings demonstrate that TP and dRib play an Important role in angiogenesis and tumor growth, and that these effects can be inhibited by IRib. Thus, IRib is a potentially useful agent for the suppression of TP-dependent anglogenesis and tumor growth.
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Molecular basis for the inhibition of hypoxia-induced apoptosis by 2-deoxy-D-ribose
Ikeda R., Furukawa T., Kitazono M., Ishitsuka K., Okumura H., Tani A., Sumizawa T., Haraguchi M., Komatsu M., Uchimiya H., Ren X., Motoya T., Yamada K., Akiyama S.
Biochemical and Biophysical Research Communications 291 ( 4 ) 806 - 812 2002.3
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Biochemical and Biophysical Research Communications
An angiogenic factor, platelet-derived endothelial cell growth factor/thymidine phosphorylase (PD-ECGF/TP), stimulates the chemotaxis of endothelial cells and confers resistance to apoptosis induced by hypoxia. 2-Deoxy-D-ribose, a degradation product of thymidine generated by TP enzymatic activity partially prevented hypoxia-induced apoptosis. 2-Deoxy-D-ribose inhibits a number of components of the caspase-mediated hypoxia-induced apoptotic pathway. It inhibits hypoxia-induced caspase 3 activation, mitochondrial cytochrome c release, downregulation of Bcl-2 and Bcl-xL, upregulation of hypoxia-inducible factor (HIF)-1α, and loss of mitochondrial transmembrane potential in human leukemia HL-60 cell line. These findings suggest a molecular mechanism by which 2-deoxy-D-ribose confers the resistance to apoptosis. Thus 2-deoxy-D-ribose-modulated suppression of HIF-1α expression could prevent the hypoxia-induced decrease of the anti-apoptotic Bcl-2 and Bcl-xLon the mitochondria. 2-Deoxy-L-ribose and its analogs may enhance apoptosis and suppress the growth of tumors by competitively inhibiting the activities of 2-deoxy-D-ribose and thus these analogs show promise for anti-tumor therapy. © 2002 Elsevier Science Ltd. All rights reserved.
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Ishitsuka K., Ikeda R., Utsunomiya A., Uozumi K., Hanada S., Suzuki S., Takeuchi S., Takatsuka Y., Takeshita T., Ohno N., Arima T.
Leukemia and Lymphoma 43 ( 5 ) 1107 - 1114 2002.1
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Leukemia and Lymphoma
Arsenic trioxide (As2O3) has been reported to induce apoptosis in human T-cell leukemia virus type-I (HTLV-I) infected T-cell lines and fresh adult T-cell leukemia (ATL) cells and to induce G1phase accumulation in HTLV-I infected T-cell lines. The present study aimed to clarify the pathway of AS2O3-induced apoptosis in HTLV-I infected T-cell lines, MT-1 and MT-2, and fresh ATL cells separated from peripheral blood of patients with acute or chronic type ATL. Cells were treated up to 72h at clinically tolerable concentrations of As2O3(1-2 μmol/l) shown to be safe in patients with acute promyelocytic leukemia (APL). Activation of caspases 3, 8, and 9, loss of mitochondrial transmembrane potential and cleavage of poly (adenosine diphosphate-ribose) polymerase (PARP) were observed during As2O3treatment. Furthermore, prior exposure to a broad-spectrum caspase inhibitor blocked AS2O3-induced apoptosis but not G1phase accumulation. While pre-treatment with a CD95 receptor-blocking antibody (Ab) or a TNF-α neutralizing Ab did not show such inhibitions in these cells. In conclusion, As2O3induces apoptosis in HTLV-I infected T-cell lines and fresh ATL cells through CD95 or TNF-α receptor independent caspase activation.
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Mori S., Takao S., Ikeda R., Noma H., Mataki Y., Wang X., Akiyama S., Aikou T.
Biochemical and Biophysical Research Communications 295 ( 2 ) 300 - 305 2002.1
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Biochemical and Biophysical Research Communications
Thymidine phosphorylase (TP) has chemotactic and angiogenic activities resulting from its enzymatic activity in vitro, and it also promotes tumor growth and inhibits apoptosis in vivo. Recently, we have reported that TP plays an important role in Fas-induced apoptosis. Caspase-8 cleavage, subsequent cytochrome c release, and caspase-3 cleavage were prevented in KB cells transfected with a TP cDNA (KB/TP cells). In this study, treatment with thymidine phosphorylase inhibitor (TPI) or thymidine did not affect cell survival of KB/TP cells during Fas-induced apoptosis. Moreover, treatment with thymine or 2-deoxy-D-ribose (degradation products of thymidine generated by TP) also did not affect cell survival of control transfectant (KB/CV) cells during Fas-induced apoptosis. These findings indicate that TP suppresses Fas-induced apoptotic signal transduction independent of its enzymatic activity. © 2002 Elsevier Science Ltd. All rights reserved.
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Role of thymidine phosphorylase in Fas-induced apoptosis.
Mori S., Takao S., Ikeda R., Noma H., Mataki Y., Wang X., Akiyama S., Aiko T.
Human cell : official journal of Human Cell Research Society 14 ( 4 ) 323 - 330 2001.12
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：Human cell : official journal of Human Cell Research Society
Thymidine phosphorylase (TP) has chemotactic and angiogenic activity in vitro, and it promotes tumor growth and inhibits apoptosis in vivo. It plays a key role in the invasiveness and metastasis of TP-expressing solid tumors. KB/TP cells transfected with a TP cDNA have been shown to be resistant to hypoxia-induced apoptosis, suggesting that TP has effects on tumor growth and cell death independent of its effects on angiogenesis. However, the mechanisms of cell death inhibition by TP are unknown. In the present study, we demonstrate that caspase-8 is cleaved in control transfectant KB cells early on during Fas-induced apoptosis. Caspase-8 activation leads to the loss of mitochondrial membrane potential, followed by the release of cytochrome c, the activation of caspase-3, and apoptosis. In contrast, Fas-induced caspase-8 cleavage is inhibited in KB/TP cells, which lead to inhibition of the downstream apoptotic cascade and inhibition of apoptosis. These findings indicate that TP plays an important role in intracellular apoptotic signal transduction in the Fas-induced apoptotic pathway. Therefore, inhibition of TP may suppress the progression of TP-overexpressing solid tumors by inducing apoptosis.
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Reversal of LRP-associated drug resistance in colon carcinoma SW-620 cells
Kitazono M., Okumura H., Ikeda R., Sumizawa T., Furukawa T., Nagayama S., Seto K., Aikou T., Akiyama S.
International Journal of Cancer 91 ( 1 ) 126 - 131 2001.1
Language：Japanese Publishing type：Research paper (scientific journal) Publisher：International Journal of Cancer
Resistance to multiple drugs is mediated by lung resistance-related protein (LRP) as well as P-glycoprotein (P-gp) and multidrug resistance protein (MRP). The levels of expression of LRP mRNA and LRP in a human colon carcinoma cell line, SW-620, were increased by the differentiation-inducing agent, sodium butyrate (NaB). Treatment of SW-620 cells with NaB for 2 weeks conferred resistance to adriamycin (ADM) and VP-16. The resistance was almost completely reversed by PAK-104P, a pyridine analog, but not by cepharanthine. ADM accumulated mainly in the nuclei of SW-620 cells not treated with NaB and in the cytoplasm of SW-620 cells treated with NaB. When the NaB-treated SW-620 cells were incubated with ADM in the presence of PAK-104P, the accumulation of ADM in nuclei was substantially increased. Isolated nuclei from untreated cells accumulated more ADM than nuclei from NaB-treated cells. Efflux of ADM from the nuclei isolated from NaB-treated cells was enhanced. PAK-104P and an antibody against LRP increased the accumulation of ADM in the isolated nuclei from NaB-treated cells, and inhibited the enhanced efflux of ADM from the nuclei. These findings suggest that at leastin part, PAK-104P reverses LRP-mediated drug resistance by inhibiting the efflux of ADM from nuclei. PAK-104P may be useful for reversing MDR in tumors that overexpress LRP. (C) 2001 Wiley-Liss, Inc.
DOI： 10.1002/1097-0215(20010101)91:1<126::AID-IJC1018>3.0.CO;2-8