Details of a Researcher - ITAGAKI Shiro

Papers - ITAGAKI Shiro

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In vitro and in vivo antioxidant properties of chlorogenic acid and caffeic acid

Yuki Sato, Shirou Itagaki, Toshimitsu Kurokawa, Jiro Ogura, Masaki Kobayashi, Takeshi Hirano, Mitsuru Sugawara, Ken Iseki

INTERNATIONAL JOURNAL OF PHARMACEUTICS ( ELSEVIER SCIENCE BV ) 403 ( 1-2 ) 136 - 138 2011.01 [Refereed]

Dietary polyphenols are thought to be beneficial for human health as antioxidants. Coffee beans contain a common polyphenol, chlorogenic acid. Chlorogenic acid is the ester of caffeic acid and quinic acid. Although these polyphenols have received much attention, there is little evidence indicating a relationship between the effect and the rate of absorption. In this study, we focused on the beneficial effects of chlorogenic acid and caffeic acid, a major metabolite of chlorogenic acid. We carried out in vitro and in vivo experiments. In the in vitro study, caffeic acid had stronger antioxidant activity than that of chlorogenic acid. The uptake of chlorogenic acid by Caco-2 cells was much less than that of caffeic acid. The physiological importance of an orally administered compound depends on its availability for intestinal absorption and subsequent interaction with target tissues. We then used an intestinal ischemia-reperfusion model to evaluate antioxidant activities in vivo. We found that both chlorogenic acid and caffeic acid had effects on intestinal ischemia-reperfusion injury. Since caffeic acid has a stronger antioxidant activity than that of chlorogenic acid and chlorogenic acid is hydrolyzed into caffeic acid in the intestine, it is possible that caffeic acid plays a major role in the protective effect of chlorogenic acid against ischemia-reperfusion injury. (C) 2010 Elsevier B.V. All rights reserved.

DOI PubMed

AMP-activated protein kinase regulates the expression of monocarboxylate transporter 4 in skeletal muscle

Ayako Furugen, Masaki Kobayashi, Katsuya Narumi, Meguho Watanabe, Sho Otake, Shirou Itagaki, Ken Iseki

LIFE SCIENCES ( PERGAMON-ELSEVIER SCIENCE LTD ) 88 ( 3-4 ) 163 - 168 2011.01 [Refereed]

Aims: The aim of this study was to determine the effect of 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), an AMP-activated protein kinase (AMPK) activator, on monocarboxylate transporter 4 (MCT4) expression in rat skeletal muscle and a prototypic embryonal rhabdomyosarcoma cell line (RD cells). Main methods: We examined the alteration in Glucose transporter 4 (GLUT4) and MCT4 mRNA levels by quantitative real-time PCR. Alteration in GLUT4 and MCT4 protein levels was examined by Western blotting. Key findings: In an in vivo study. AICAR increased MCT4 mRNA and protein levels in a fiber-type specific manner. In an in vitro study, AICAR increased MCT4 mRNA and protein levels. Moreover, AICAR-induced MCT4 expression was blocked by Compound C, an AMPK inhibitor. Significance: In this study, we found that AMPK activation induced expression of MCT4 in RD cells and rat skeletal muscle in a fiber-type specific manner. These results indicate the possible involvement of an AMPK-mediated pathway associated with MCT4 expression in skeletal muscle. (C) 2010 Elsevier Inc. All rights reserved.

DOI PubMed CiNii

有機アニオントランスポータOATP2B1の臨床的有用性

板垣史郎

医薬品相互作用研究 35 ( 1 ) 1 - 10 2011 [Refereed] [Invited]

Regulation of Monocarboxylate Transporter 1 in Skeletal Muscle Cells by Intracellular Signaling Pathways

Katsuya Narumi, Ayako Furugen, Masaki Kobayashi, Sho Otake, Shirou Itagaki, Ken Iseki

BIOLOGICAL & PHARMACEUTICAL BULLETIN ( PHARMACEUTICAL SOC JAPAN ) 33 ( 9 ) 1568 - 1573 2010.09 [Refereed]

Skeletal muscle is the major producer of lactic acid in the body, but its oxidative fibers also use lactic acid as a respiratory fuel. Monocarboxylate transporter (MCT) 1 has been suggested to play a major role in influx of L-lactic acid for oxidation. The regulation mechanism of MCT1 was characterized utilizing rhabdomyosarcoma cells as an in vitro skeletal muscle model. The uptake of L-lactic acid via MCT1 was studied in the presence of various intracellular regulatory pathways, including pathways mediated by protein kinases A, C and G (PKA, PKC and PKG), protein tyrosine kinase (PTK), and Ca2+/calmodulin modulators. The results showed that PKG-, PTK-, and Ca2+/calmodulin-mediated regulatory pathways play no role in the regulation of L-lactic acid uptake, but a role for PKC- and PKA-mediated pathways was apparent. Uptake of L-lactic acid appeared to be stimulated by phorbol 12-myristate 13-acetate (PMA, a PKC activator) via an increase in V-max of transport processes with no alteration in K-m. In parallel, PMA treatment also resulted in an increase in the level of MCT1 expression. On the other hand, exposure to 8-Br-cAMP, a cAMP analog, and to forskolin, an adenylyl cyclase activator, resulted in a significant decrease in L-lactic acid uptake. Additionally, 8-Br-cAMP reduced V-max but not K-m values. Parallel to the decrease in V-max of L-lactic acid uptake, the level of MCT1 expression was decreased in response to incubation with 8-Br-cAMP. These results indicate the possible involvement of a PKC- and PKA-mediated pathway associated with expression of MCT1 and lactate transport.

DOI CiNii

放射線治療に伴う口腔粘膜障害に対する予防・軽減効果が期待される物質の探索

板垣史郎, 中田千絵, 平野剛, 鷹野瑠美, 笠師久美子, 菅原満, 高橋夏子, 小林正紀, 井関健

医療薬学 ( (一社)日本医療薬学会 ) 36 ( 9 ) 696 - 702 2010.09 [Refereed]

放射線台療に伴う口腔粘膜障害に対する予防・軽減効果が期待される物質を探索した。16Gyの放射線照射を行った後、5日間培養したHO-1-N-1細胞を用いて、種々の化合物の障害抑制効果について検討した。抗酸化作用を有するN-アセチルシステイン(NAC)およびカテキン類に着目し、その障害抑制効果について検討した。高濃度のNACの添加により、放射線照射による細胞生存率の低下が有意に抑制された。カテキン類の添加により、放射線照射による細胞生存率の低下は有意に抑制された。NACおよびカテキン類の添加により、放射線照射による蛍光強度の増大は抑制された。HO-1-N-1細胞に放射線を照射したところ、アポトーシスに特徴的なDNAの断片化が観察され、アポトーシスが誘導されていることが示された。一方、NACおよびカテキン類の添加により、DNAの断片化は顕著に抑制された。

DOI

Pharmacokinetics of Oral and Intravenous Administration of Digoxin after Intestinal Ischemia-Reperfusion

Jiro Ogura, Hajime Maruyama, Masaki Kobayashi, Shirou Itagaki, Ken Iseki

BIOLOGICAL & PHARMACEUTICAL BULLETIN ( PHARMACEUTICAL SOC JAPAN ) 33 ( 5 ) 922 - 925 2010.05 [Refereed]

Intestinal ischemia-reperfusion (I/R) causes gut dysfunction characterized by decreased basement membrane integrity and decreased barrier function. Indeed, it has been reported that the absorption of several drugs is altered after intestinal I/R. Intestinal I/R also promotes multi-organ failure (MOF). The liver and kidney can be affected by MOF after intestinal I/R. However, little is known about the alteration of pharmacokinetics after intravenous administration in intestinal I/R injury. In the present study, we investigated pharmacokinetics of digoxin after oral administration and intravenous administration in intestinal I/R injury. Plasma digoxin concentration in I/R rats after oral administration was not significantly altered at any time compared with that in sham-operated rats. Plasma digoxin concentration in rats reperfused for 1 h after intravenous administration was significantly higher than that in sham-operated rats. Plasma digoxin concentrations in rats reperfused for 6 and 24 h were the same as those in sham-operated rats. The area under the concentration time curve after intravenous administraion (AUC(i.s.)) and total clearance (CLtot) in rats reperfused for 1 h was 1.89- and 0.57-fold higher than that in sham-operated rats. However, elimination rate (k(e)) and half-life (t(1/2)) in rats reperfused for 1 h were not altered. Distribution volume (V-d) in rats reperfused for 1 h was decreased than that in sham-operated rats, but there was not statistical difference. These results suggest that intestinal I/R affected the V-d of digoxin, and plasma concentration of digoxin was increased. The present study suggests that understanding pharmacokinetics of drug after intravenous administration in intestinal I/R injury is important to provide valuable information for safe drug therapy for intestinal I/R patients.

DOI

Grapefruit juice enhance the uptake of coenzyme Q10 in the human intestinal cell-line Caco-2

Shirou Itagaki, Akiko Ochiai, Masaki Kobayashi, Mitsuru Sugawara, Takeshi Hirano, Ken Iseki

FOOD CHEMISTRY ( ELSEVIER SCI LTD ) 120 ( 2 ) 552 - 555 2010.05 [Refereed]

Coenzyme Q10 (CoQ10) is very widely consumed by humans as a food supplement. However, CoQ10 is taken up from the intestine into the circulation at a low rate. The absorption of compounds from the gastrointestinal tract is one of the important determinants for oral bioavailability. Secretory transport limits the Oral bioavailability of compounds. It has been reported that efflux transport of CoQ10 is mediated by P-glycoprotein (P-gp) in Caco-2 cells. We tried to improve intestinal absorption of CoQ10 by modulating P-gp. Since grapefruit juice (GFJ) is reported to inhibit P-gp function, we investigated the effect of GFJ on the transport of CoQ10 by Caco-2 cells. In the presence of GFJ, the basal-to-apical transport of CoQ10 was decreased and the uptake of CoQ10 was increased. These findings suggest that the combined administration of CoQ10 and GFJ could enhance CoQ10 absorption. (C) 2009 Elsevier Ltd. All rights reserved.

DOI CiNii

In vitro実験系による肺がん化学療法レジメンの抗腫瘍効果の評価

斎藤由起子, 平野剛, 沖洋充, 笠師久美子, 菅原満, 小林正紀, 高橋夏子, 板垣史郎, 井関健

医療薬学 ( (一社)日本医療薬学会 ) 36 ( 4 ) 220 - 226 2010.04 [Refereed]

In vitro実験系による肺がん化学療法レジメンの抗腫瘍効果について検討した。肺がんのモデルとしてヒト肺扁平上皮がん由来A549細胞を使用した。CDDP+CPT-11(IP)療法において、CDDPは長期間にわたって抗腫瘍効果を示すことが示唆された。SN-38はday2からday5にかけて細胞生存率の回復を認め、CPT-11の1回日の投与により死滅したがん細胞が回復してきたところを再投与によりさらに攻撃するというレジメンの投与スケジュールに相関した。CDDP+VP-16(EP)療法において、各々の単剤のときと比較して抗腫瘍効果が増大することが示された。

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Diclofenac-induced stimulation of SMCT1 (SLC5A8) in a heterologous expression system: A RPE specific phenomenon

Sudha Ananth, Lina Zhuang, Elangovan Gopal, Shiro Itagaki, Babu Ellappan, Sylvia B. Smith, Vadivel Ganapathy, Pamela Martin

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS ( ACADEMIC PRESS INC ELSEVIER SCIENCE ) 394 ( 1 ) 75 - 80 2010.03 [Refereed]

SMCT1 is a Na(+)-coupled monocarboxylate transporter expressed in a variety of tissues including kidney, thyroid, small intestine, colon, brain, and retina. We found recently that several non-steroidal anti-inflammatory drugs (NSAIDs) inhibit the activity of SMCT1. Here we evaluated the effect of diclofenac, also a NSAID, on SMCT1. SMCT1 cDNA was expressed heterologously in the human retinal pigment epithelial cell lines HRPE and ARPE-19, the human mammary epithelial cell line MCF7, and in Xenopus laevis oocytes. Transport was monitored by substrate uptake and substrate-induced currents. Na(+)-dependent uptake/current was considered as SMCT1 activity. The effect of diclofenac was evaluated for specificity, dose-response, and influence on transport kinetics. To study the specificity of the diclofenac effect, we evaluated the influence of this NSAID on the activity of several other cloned transporters in mammalian cells under identical conditions. In contrast to several NSAIDs that inhibited SMCT1, diclofenac stimulated SMCT1 when expressed in HRPE and ARPE-19 cells. The stimulation was marked, ranging from 2- to 5-fold depending on the concentration of diclofenac. The stimulation was associated with an increase in the maximal velocity of the transport system as well as with an increase in substrate affinity. The observed effect on SMCT1 was selective because the activity of several other cloned transporters, when expressed in HRPE cells and studied under identical conditions, was not affected by diclofenac. Interestingly, the stimulatory effect on SMCT1 observed in HRPE and ARPE-19 cells was not evident in MCF7 cells nor in the X. laevis expression system, indicating that SMCT1 was not the direct target for diclofenac. The RPE-specific effect suggests that the target of diclofenac that mediates the stimulatory effect is expressed in RPE cells but not in MCF7 cells or in X. laevis oocytes. Since SMCT1 is a concentrative transporter for metabolically important compounds such as pyruvate, lactate, beta-hydroxybutyrate, and nicotinate, the stimulation of its activity by diclofenac in RPE cells has biological and clinical significance. (C) 2010 Elsevier Inc. All rights reserved.

DOI PubMed

Protective effects of quercetin-3-rhamnoglucoside (rutin) on ischemia-reperfusion injury in rat small intestine

Shirou Itagaki, Setsu Oikawa, Jiro Ogura, Masaki Kobayashi, Takeshi Hirano, Ken Iseki

FOOD CHEMISTRY ( ELSEVIER SCI LTD ) 118 ( 2 ) 426 - 429 2010.01 [Refereed]

Quercetin-3-rhamnoglucoside (rutin) has a wide spectrum of biochemical and pharmacological activities. Orally administered compounds are absorbed in the intestine. Although absorption of rutin from the jejunum of the rat was good, binding of rutin to the intestinal wall components may limit its absorption from the small intestine. The physiological importance of an orally administered compound depends on its interaction with target tissues. Since there is limited information on the importance of rutin in vivo, we focused on the protective effect of rutin on intestinal injury. The intestinal mucosa is extremely sensitive to reactive oxygen species. We used a rat mesenteric ischemia-reperfusion (I/R) injury model as a model of oxidative injury and investigated the antioxidant activities of rutin in vivo. We found that rutin, which has combined antioxidant activity from radical-scavenging, xanthine oxidase inhibition and chain-breaking effects, exhibits a protective effect on I/R injury in the rat small intestine. (C) 2009 Elsevier Ltd. All rights reserved.

DOI

In vitro Evaluation of Anticancer Effects of FOLFIRI and mFOLFOX 6 Chemotherapy Regimens and Their Effects on EGFR Expression

sei mei, sei mei, sei mei, sei mei, sei mei, sei mei, sei mei

Iryo Yakugaku (Japanese Journal of Pharmaceutical Health Care and Sciences) ( Japanese Society of Pharmaceutical Health Care and Sciences ) 36 ( 12 ) 855 - 862 2010 [Refereed]

Recently,the number of patients with colorectal cancer has been increasing.Many patients are not aware of any symptoms until the disease is advanced.It is important to select effective regimens for colorectal cancer therapy and combination therapy with a molecular-targeted agent has recently been used for this purpose.However,the effects of anticancer drugs on epidermal growth factor receptor (EGFR),a molecular-targeted protein,have not been clarified.We therefore evaluated the effects of folinic acid/5-fluorouracil/irinotecan (FOLFIRI) and folinic acid/5-fluorouracil/oxaliplatin (mFOLFOX citation=6)treatments,using the 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2 H-tetrazolium bromide (MTT) assay,as well as the effects of these treatments on EGFR expression in LoVo cells.
The results suggested that 5-fluorouracil and irinotecan have a stronger antiproliferative effect than oxaliplatin.The reduction in viability for LoVo cells treated with FOLFIRI was greater than that for Lovo cells treated with mFOLFOX 6 on day 22.Moreover,EGFR mRNA and protein levels were increased more by mFOLFOX 6 than by FOLFIRI.These results suggest that FOLFIRI therapy may be more effective than mFOLFOX 6 therapy against LoVo cells in consideration of EGFR expression.

DOI CiNii

Expression and Role of SNAT3 in the Placenta

C. Yoshioka, S. Yasuda, F. Kimura, M. Kobayashi, S. Itagaki, T. Hirano, K. Iseki

PLACENTA ( W B SAUNDERS CO LTD ) 30 ( 12 ) 1071 - 1077 2009.12 [Refereed]

Glutamine is the most versatile amino acid and its plasma concentration is the highest of all amino acid. Many transporters are therefore involved in glutamine uptake or efflux. Glutamine is actively released from the placenta into fetal circulation. In this study, we examined the alteration of transporters that transport glutamine into fetal circulation as gestation progresses. High expression levels of system A and y(+)L were found in the rat placenta in the late period of pregnancy and the expression levels of these transporters increased as gestation progressed (p < 0.05). On the other hand, the expression of SNAT3, the system N transporter, was detected in the early period of pregnancy and its expression level decreased as gestation progressed (p < 0.05). SNAT3 was also found to be expressed in isolated human primary cytotrophoblast cells and its expression level was decreased by their differentiation into syncytiotrophoblast cells (p < 0.05). Since this regulation is closely related to glutamine synthetase expression, SNAT3 may play a key role in providing glutamine corresponding to glutamine synthetase function in the early period of gestation. This is the first report on the expression of SNAT3 in the placenta in the early stage of pregnancy. (C) 2009 Elsevier Ltd. All rights reserved.

DOI

Liver X receptor regulates expression of MRP2 but not that of MDR1 and BCRP in the liver

Ikumi Chisaki, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki

BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES ( ELSEVIER SCIENCE BV ) 1788 ( 11 ) 2396 - 2403 2009.11 [Refereed]

Liver X receptors (LXRs) belong to the nuclear hormone receptor superfamily. Multidrug resistance-associated protein 2 (MRP2), multidrug resistance 1 (MDR1) and breast cancer resistance protein (BCRP) play an important role in the efflux of a broad range of endogenous and xenobiotic compounds from hepatocytes. Since the effects of LXR activation on there transporters have been obscure, we investigated the effects of LXR agonists, TO901317 and 25-hydroxycholesterol, on MRP2, MDR1, BCRP expression in HepG2 cells and the rat liver. in an in vitro study, TO901317 increased ABCA1, an LXR target gene, and MRP2 mRNA and protein levels. On the other hand, TO901317 had little effect on MDR1 and BCRP mRNA levels. In an in vivo study, Abca1 and Mrp2 mRNA and protein levels were increased by TO901317. but TO901317 had no effect on Mdr1a and Bcrp mRNA levels in the rat liver. Moreover, TO901317-induced MRP2 mRNA expression was blocked by LXR alpha knockdown. In this study, we demonstrated that LXR activation induced expression of MRP2 but not that of MDR1 and BCRP in hepatocytes. The results suggest that agonists for LXR activate transcription of the MRP2 gene in order to promote excretion of endogenous and xenobiotic compounds from hepatocytes into bile. (C) 2009 Elsevier B.V. All rights reserved.

DOI

Liver X receptor regulates expression of MRP2 but not that of MDR1 and BCRP in the liver.

Chisaki I, Kobayashi M, Itagaki S, Hirano T, Iseki K

Biochimica et biophysica acta ( 11 ) 1788 ( 11 ) 2396 - 2403 2009.11 [Refereed]

Liver X receptors (LXRs) belong to the nuclear hormone receptor superfamily. Multidrug resistance-associated protein 2 (MRP2), multidrug resistance 1 (MDR1) and breast cancer resistance protein (BCRP) play an important role in the efflux of a broad range of endogenous and xenobiotic compounds from hepatocytes. Since the effects of LXR activation on there transporters have been obscure, we investigated the effects of LXR agonists, TO901317 and 25-hydroxycholesterol, on MRP2, MDR1, BCRP expression in HepG2 cells and the rat liver. in an in vitro study, TO901317 increased ABCA1, an LXR target gene, and MRP2 mRNA and protein levels. On the other hand, TO901317 had little effect on MDR1 and BCRP mRNA levels. In an in vivo study, Abca1 and Mrp2 mRNA and protein levels were increased by TO901317. but TO901317 had no effect on Mdr1a and Bcrp mRNA levels in the rat liver. Moreover, TO901317-induced MRP2 mRNA expression was blocked by LXRα knockdown. In this study, we demonstrated that LXR activation induced expression of MRP2 but not that of MDR1 and BCRP in hepatocytes. The results suggest that agonists for LXR activate transcription of the MRP2 gene in order to promote excretion of endogenous and xenobiotic compounds from hepatocytes into bile.

DOI CiNii

Liver X receptor regulates expression of MRP2 but not that of MDR1 and BCRP in the liver.

Chisaki I, Kobayashi M, Itagaki S, Hirano T, Iseki K

Biochimica et biophysica acta ( Elsevier B.V. ) 1788 ( 11 ) 2396 - 2403 2009.11 [Refereed]

Liver X receptors (LXRs) belong to the nuclear hormone receptor superfamily. Multidrug resistance-associated protein 2 (MRP2), multidrug resistance 1 (MDR1) and breast cancer resistance protein (BCRP) play an important role in the efflux of a broad range of endogenous and xenobiotic compounds from hepatocytes. Since the effects of LXR activation on there transporters have been obscure, we investigated the effects of LXR agonists, TO901317 and 25-hydroxycholesterol, on MRP2, MDR1, BCRP expression in HepG2 cells and the rat liver. in an in vitro study, TO901317 increased ABCA1, an LXR target gene, and MRP2 mRNA and protein levels. On the other hand, TO901317 had little effect on MDR1 and BCRP mRNA levels. In an in vivo study, Abca1 and Mrp2 mRNA and protein levels were increased by TO901317. but TO901317 had no effect on Mdr1a and Bcrp mRNA levels in the rat liver. Moreover, TO901317-induced MRP2 mRNA expression was blocked by LXRα knockdown. In this study, we demonstrated that LXR activation induced expression of MRP2 but not that of MDR1 and BCRP in hepatocytes. The results suggest that agonists for LXR activate transcription of the MRP2 gene in order to promote excretion of endogenous and xenobiotic compounds from hepatocytes into bile.

DOI PubMed CiNii

Intestinal Absorption and Secretion Mechanism of Carboxylate Drugs

Shirou Itagaki

YAKUGAKU ZASSHI-JOURNAL OF THE PHARMACEUTICAL SOCIETY OF JAPAN ( PHARMACEUTICAL SOC JAPAN ) 129 ( 11 ) 1341 - 1349 2009.11 [Refereed] [Invited]

Oral drug delivery is generally the most desirable means of administration, mainly because of patient. acceptance, convenience in administration Intestinal absorption mechanisms of anionic drugs have been mainly explained by the passive diffusion of nonionized compounds. However, several studies have suggested the involvement of specific transporters in intestinal absorption of weak acids including monocarboxylates. (-)-N-(trans-4-Isopropylcyclohexanecarbonyl)-D-phenylalanine (nateglinide) is a oral hypoglycemic agent possessing a carboxyl group and a peptide-type bond in its structure. Although nateglinide quickly reaches the maximal serum concentration after oral administration, nateglinide itself is not transported by PepT1 or MCT1. We demonstrated that nateglinide transport occurs via a single system that is H+ dependent but is distinct from PepT1 or MCT1. In clinical, patients usually take many kinds of drugs at the same dine. Thus, drug-drug interactions involving transporters can often directly affect the therapeutic safety and efficacy of many drugs However, there have been few studies on food-drug interactions involving transporters. Dietary polyphenols have been widely assumed to be beneficial to human health. Polyphenols are commercially prepared and used as functional foods. We reported that ferulic acid, which is widely used as a functional food, affects the transport of clinical agents. The major dose-limiting toxicity after administration of trinotecan hydrochloride. 7-ethyl-10-(4-[1-piperidino]-1-piperidino)-carbonyloxycamptothecin (CPT-11) is severe diarrhea. We have found that a specific transport system mediates the uptake of active metabolite 7-ethyl-10-hydroxycamptothecin (SN-38) across the apical membrane in Caco-2 cells. Baicalin and sulfobromophthatlcin inhibit this transporter. Inhibition of this transporter would be a useful means for reducing late-onset diarrhea.

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Transport by SLC5A8 With Subsequent Inhibition of Histone Deacetylase 1 (HDAC1) and HDAC3 Underlies the Antitumor Activity of 3-bromopyruvate

Muthusamy Thangaraju, Senthil K. Karunakaran, Shiro Itagaki, Elangovan Gopal, Selvakumar Elangovan, Puttur D. Prasad, Vadivel Ganapathy

CANCER ( JOHN WILEY & SONS INC ) 115 ( 20 ) 4655 - 4666 2009.10 [Refereed]

BACKGROUND: 3-Bromopyruvate is an alkylating agent with antitumor activity. It is currently believed that blockade of adenosine triphosphate production from glycolysis and mitochondria is the primary mechanism responsible for this antitumor effect. The current studies uncovered a new and novel mechanism for the antitumor activity of 3-bromopyruvate. METHODS: The transport of 3-bromopyruvate by sodium-coupled monocarboxylate transporter SMCT1 (SLC5A8), a tumor suppressor and a sodium (Na(+))-coupled, electrogenic transporter for short-chain monocarboxylates, was studied using a mammalian cell expression and the Xenopus laevis oocyte expression systems. The effect of 3-bromopyruvate on histone deacetylases (HDACs) was monitored using the lysate of the human breast cancer cell line MCF7 and human recombinant HDAC isoforms as the enzyme sources. Cell viability was monitored by fluorescence-activated cell-sorting analysis and colony-formation assay. The acetylation status of histone H4 was evaluated by Western blot analysis. RESULTS: 3-Bromopyruvate is a transportable substrate for SLC5A8, and that transport process is Na(+)-coupled and electrogenic. MCF7 cells did not express SLC5A8 and were not affected by 3-bromopyruvate. However, when transfected with SLC5A8 or treated with inhibitors of DNA methylation, these cells underwent apoptosis in the presence of 3-bromopyruvate. This cell death was associated with the inhibition of HDAC1/HDAC3. Studies with different isoforms of human recombinant HDACs identified HDAC1 and HDAC3 as the targets for 3-bromopyruvate. CONCLUSIONS: 3-Bromopyruvate was transported into cells actively through the tumor suppressor SLC5A8, and the process was energized by an electrochemical Na(+) gradient. Ectopic expression of the transporter in MCF7 cells led to apoptosis, and the mechanism involved the inhibition of HDAC1/HDAC3. Cancer 2009;115:4655-66. (C) 2009 American Cancer Society.

DOI PubMed

Response of the ABCG2 promoter in T47D cells and BeWo cells to sex hormone treatment

Satoru Yasuda, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki

MOLECULAR BIOLOGY REPORTS ( SPRINGER ) 36 ( 7 ) 1889 - 1896 2009.09 [Refereed]

The aim of this study was to elucidate the effects of sex hormones on activity of the ABCG2 promoter in different cell lines. T47D cells and BeWo cells were used as models for ABCG2-expressing cell lines, and luciferase assays using ABCG2 promoter-luciferase constructs were performed. It was shown that progesterone increased the response of the ABCG2 promoter in T47D cells but not in BeWo cells. On the other hand, estradiol had no effect on response of the ABCG2 promoter in either cell line. However, response of the ABCG2 promoter was enhanced by overexpression of ER alpha in both T47D cells and BeWo cells. T47D cells had higher sensitivity to ER alpha than did BeWo cells. Furthermore, it was shown that the inductive effect of progesterone on the ABCG2 promoter was inhibited by addition of RU486 or mithramycin A. Therefore, it was thought that the ABCG2 promoter responded to stimulation of the progesterone receptor (PR)-Sp1 pathway in T47D cells. Furthermore, progesterone suppressed the response of the ABCG2 promoter by changing the expression levels of PR-A and PR-B in BeWo cells. These findings suggested that there are differences between cell lines in the regulation mechanism of ABCG2 expression by sex hormone treatment.

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Food-Drug Interaction Involving the Ferulic Acid/H plus Transporter

Shirou Itagaki, Yoko Kobayashi, Yoshitaka Saito, Masaki Kobayashi, Takeshi Hirano, Ken Iseki

DRUG METABOLISM REVIEWS ( TAYLOR & FRANCIS INC ) 41 60 - 60 2009.08 [Refereed]

Contribution of organic anion transporting polypeptide OATP2B1 to amiodarone accumulation in lung epithelial cells

Satoru Seki, Masaki Kobayashi, Shirou Itagaki, Takeshi Hirano, Ken Iseki

BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES ( ELSEVIER SCIENCE BV ) 1788 ( 5 ) 911 - 917 2009.05 [Refereed]

The accumulation mechanisms of amiodarone (AMD) involving transporters in lung alveolar epithelial type 11 cells were studied. The uptake of AMD was examined using human alveolar epithelia I-derived cell line A549 as a model. AMD was transported by the carrier-mediated system, and the apparent K-m and V-max values were 66.8 +/- 30.3 mu M and 49.7 +/- 9.7 nmol/mg protein/5 min, respectively. The uptake of AMD by A549 cells was Na+-independent and was inhibited by substrates of human organic anion transporting polypeptide (OATP). The inhibition profiles were similar to the inhibitory effects of several compounds on OATP2B1 -mediated E-3-S transport, and RT-PCR analysis showed mRNA expression of OATP2B1 and 1B3 in A549 cells. SiRNAs targeted to the OATP2B1 gene decreased the OATP2B1 mRNA expression level in A549 cells up to about 50% and reduced the uptake of AMD up to about 40%. These results indicate that AMD uptake mediated by carriers, including OATP2B1, might lead to accumulation of AMD in the lung and AMD-induced pulmonary toxicity (AIPT). (C) 2009 Elsevier B.V. All rights reserved.

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