Details of a Researcher - Ohtani, Hisakazu

Papers - Ohtani, Hisakazu

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Kinetic analysis of P-glycoprotein-mediated transport by using normal human placental brush-border membrane vesicles

Ushigome F., Koyabu N., Satoh S., Tsukimori K., Nakano H., Nakamura T., Uchiumi T., Kuwano M., Ohtani H., Sawada Y.

Pharmaceutical Research (Pharmaceutical Research) 20 ( 1 ) 38 - 44 2003.01

ISSN 07248741

Purpose. P-Glycoprotein (Pgp) plays an important role in drug disposition and excretion in various tissues such as the brain, intestine, and kidney. Moreover, we have demonstrated that Pgp is expressed on the brush-border membranes of trophoblast cells in the placenta and restricts drug transfer from the maternal circulation to the fetus. However, the transport kinetics of physiologically expressed Pgp has scarcely been investigated. Methods. In this study, we assessed the functional kinetics of transport mediated by Pgp that is physiologically expressed in normal tissue by using human placental brush-border membrane vesicles (BBMVs). Digoxin and vinblastine were used as typical substrates of Pgp. Results. The uptakes of [3H] digoxin and [3H] vinblastine into BBMVs were significantly increased in the presence of an ATP-regenerating system. The ATP-dependent uptakes of [3H]digoxin and [3H] vinblastine into BBMVs exhibited saturable kinetics. The Michaelis constants (Kt values) were 2.65 ± 1.80 μM and 21.9 ± 3.37 μM, respectively. In the presence of a Pgp inhibitor such as verapamil, cyclosporine A, or progesterone, the ATP-dependent uptakes of [3H]digoxin and [3H] vinblastine into BBMVs were significantly reduced. Anti-Pgp monoclonal antibody C219 completely inhibited the uptake of [3H] digoxin. Conclusions. The transport kinetics of [3H]digoxin and [3H]vinblastine by physiologically expressed Pgp were successfully evaluated by using BBMVs prepared from normal human placenta. The present method enabled us to evaluate the function of physiologically expressed Pgp and is superior to the use of cultured transfectants in terms of the yield of vesicles. The present method may also be applicable to investigating the influence of various factors such as the genotype of the MDR1 gene or various pathophysiologic states of neonates on the function of Pgp.

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Agranulocytosis possibly caused by ranitidine in a patient with renal failure

Takami N., Yamamoto Y., Matsuo H., Ohtani H., Sawada Y.

International Journal of Clinical Pharmacology and Therapeutics (International Journal of Clinical Pharmacology and Therapeutics) 40 ( 11 ) 520 - 523 2002.11

ISSN 09461965

A 70-year-old Japanese woman with renal dysfunction under hemodialysis presented with vomiting and chill with fever. Over the previous 24 weeks she had been taking 75 mg of ranitidine after hemodialysis. Other medications taken were prednisolone, furosemide, α-calcidol, amlodipine and calcium carbonate. Before starting ranitidine, she had been treated with famotidine for about 2 years without complication. Hematological inspection on admission revealed agranulocytosis with WBC of 400/mm3. Ranitidine was discontinued and granulocyte colony-stimulating factor (G-CSF) was started. On Day 3, laboratory data showed slight improvement of cytopenia with WBC Of 1,000/mm3. On Day 6, her hemogram showed marked improvement with WBC of 11,700/mm3 and G-CSF was discontinued. She was discharged on Day 10. Several cases describing ranitidine-induced cytopenia are associated with the use of ranitidine at a dose of 150 mg/day or higher, and adverse reactions were found within 2 - 35 days after beginning ranitidine treatment. In the case described here, however, the adverse reaction occurred after a longer treatment period with ranitidine at a lower dose. In conclusion, ranitidine should be administered with great caution to patients with severe renal dysfunction.

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Nightmares and panic disorder associated with carvedilol overdose

Maebara C., Ohtani H., Sugahara H., Mine K., Kubo C., Sawada Y.

Annals of Pharmacotherapy (Annals of Pharmacotherapy) 36 ( 11 ) 1736 - 1740 2002.11

ISSN 10600280

OBJECTIVE: To report a case of nightmares and sleep disorder associated with improper use of carvedilol, an α/β-blocker, and to model the time course of receptor occupancy in this patient. CASE SUMMARY: A 41-year-old man with panic disorder had been treated with alprazolam 1.2 mg/d (3 times daily), carvedilol 10 mg/d (once in the morning), and etizolam 0.5 mg (for anxiety attack). Although the physical and psychological symptoms gradually improved, he reported nightmares and panic attacks. An interview revealed that he had been taking carvedilol 5 mg twice a day after lunch and dinner on his own initiative, in addition to the prescribed dosage. The patient was asked to take carvedilol 10 mg only after breakfast, as had been advised. Consequently, the sleep disorder and nightmares disappeared. METHODS: We calculated the time courses of β2-adrenoceptor binding occupancy in the central nervous system after oral administration of carvedilol with the ordinary and improper regimens by using pharmacokinetic/pharmacodynamic parameters obtained from the literature. RESULTS: Compared with the ordinary dose of carvedilol 10 mg once a day, the improper regimen (10 mg after breakfast followed by 5 mg after lunch and dinner) increases the β2-adrenoceptor binding occupancy at night (2300) to as high as the mean β2-adrenoceptor binding occupancy after an ordinary dose of propranolol. CONCLUSIONS: The sleep disorder and nightmares experienced by this patient had been induced by elevation of central β2-adrenoceptor binding occupancy at night as the result of improper use of carvedilol.

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非循環系薬物による致死性心電学的副作用の定量的予測法の確立

大谷壽一

上原記念生命科学財団研究報告集 ((公財)上原記念生命科学財団) 16 30 - 32 2002.11

薬物による催不整脈作用をin vitroの実験から定量的に予測するために,動物実験におけるin vivo-in vitro相関の解析と,ヒトカリウムチャンネル発現系におけるカリウム電流阻害作用と臨床的報告に基づく副作用リスクとの解析を試みた.モルモットを用い,in vivo実験系として心電図の薬物動態・動力学的解析を,in vitro実験系としては摘出心臓還流標本における心電図測定を行い,両者の関係を解析した.in vivo実験ではキニジン(30mg/kg/h),ソタロール(5mg/kg/h)のいずれによっても心電図QTc間隔は最大30-40msec程度延長した.in vitro実験でもQTcはいずれの薬剤でも延長し,灌流液濃度とQTc延長との関係は直線関係が認められた.いずれの薬剤でもin vivoではin vitroよりもQTc延長の強度が強いことが明らかとなった.従って,in vitro実験結果をin vivoに外挿する場合には,両者の濃度的乖離を十分に考慮する必要があると考えられた

Functional characterization of mouse cation transporter mOCT2 compared with mOCT1

Kakehi M., Koyabu N., Nakamura T., Uchiumi T., Kuwano M., Ohtani H., Sawada Y.

Biochemical and Biophysical Research Communications (Biochemical and Biophysical Research Communications) 296 ( 3 ) 644 - 650 2002.10

ISSN 0006291X

We characterized the function of mouse organic cation transporter OCT2 (TC 2.A.19.1.5) in comparison with that of OCT1 (TC 2.A.19.1.1). Uptake of [3H]1-methyl-4-phenylpyridinium ([3H]MPP+) by Xenopus laevis oocytes injected with mOCT1 (Slc22a1) or mOCT2 (Slc22a2) cRNA was attenuated by an increase of extracellular K+ concentration and under acidic extracellular conditions. The uptakes of [3H]MPP+ via mOCT1 and mOCT2 were saturable, with similar Michaelis constants (Kt) of 10 and 24 μM, respectively. mOCT2 also mediated the uptake of [14C]tetraethylammonium with a Kt value of 36 μM, which is similar to that of mOCT1. Quinine, tetraethylammonium, cimetidine, procainamide, choline, and N′-methylnicotinamide inhibited the uptake of [3H]MPP+ via mOCT1, as well as via mOCT2, and the inhibitory potencies for mOCT1 were comparable to but slightly higher than those for mOCT2. Thus, although the transport properties of mOCT2 are similar to those of mOCT1 in respect to the membrane-potential dependency, pH-sensitivity, and affinities for MPP+ and tetraethylammonium, several organic cations had weaker inhibitory effects on [3H]MPP+ uptake by mOCT2 than by mOCT1. © 2002 Elsevier Science (USA). All rights reserved.

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Transport and uptake of nateglinide in Caco-2 cells and its inhibitory effect on human monocarboxylate transporter MCT1

Okamura A., Emoto A., Koyabu N., Ohtani H., Sawada Y.

British Journal of Pharmacology (British Journal of Pharmacology) 137 ( 3 ) 391 - 399 2002.10

ISSN 00071188

1 Nateglinide, a novel oral hypoglycemic agent, rapidly reaches the maximum serum concentration after oral administration, suggesting that it is rapidly absorbed in the gastrointestinal tract. The aim of this work is to clarify the intestinal absorption mechanism of nateglinide by means of in vitro studies. 2 We examined the transcellular transport and the apical uptake of [ 14C]nateglinide in a human colon carcinoma cell line (Caco-2). We also examined whether nateglinide is transported via monocarboxylate transport-1 (MCT1) by means of an uptake study using MCT1-expressing Xenopus laevis oocytes. 3 In Caco-2 cells, the transcellular transport of [ 14C]nateglinide from the apical to basolateral side was greater than that in the opposite direction. The uptake of [ 14C]nateglinide from the apical side was concentration-dependent, H +-dependent, and Na +-independent. Kinetic analysis revealed that the Kt and Jmax values of the initial uptake rate of [ 14C]nateglinide were 448 μM and 43.2 nmol mg protein -1 5 min -1, respectively. Various monocarboxylates, including salicylic acid and valproic acid, and glibenclamide significantly inhibited the uptake of [ 14C]nateglinide. 4 The uptake study using MCT1-expressing oocytes showed that nateglinide inhibits the MCT1-mediated uptake of [ 14C]L-lactic acid, though nateglinide itself is not transported by MCT1. 5 Taken together, these results suggest that the uptake of nateglinide from the apical membranes of Caco-2 cells is, at least in part, mediated by a proton-dependent transport system(s) distinct from MCT1.

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Characteristics of L-lactic acid transport in basal membrane vesicles of human placental syncytiotrophoblast

Inuyama M., Ushigome F., Emoto A., Koyabu N., Satoh S., Tsukimori K., Nakano H., Ohtani H., Sawada Y.

American Journal of Physiology - Cell Physiology (American Journal of Physiology - Cell Physiology) 283 ( 3 52-3 ) C822 - 30 2002.09

ISSN 03636143

The characteristics of L-lactic acid transport across the trophoblast basal membrane were investigated and compared with those across the brush-border membrane by using membrane vesicles isolated from human placenta. The uptake of L-[14C]lactic acid into basal membrane vesicles was Na+ independent, and an uphill transport was observed in the presence of a pH gradient ([H+]out > [H+]in). L-[14C]lactic acid uptake exhibited saturation kinetics with a Km value of 5.89 ± 0.68 mM in the presence of a pH gradient. p-Chloromercuribenzenesulfonate and α-cyano-4-hydroxycinnamate inhibited the initial uptake, whereas phloretin or 4,4′-diisothiocyanostilbene-2,2′-disulfonate did not. Mono- and dicarboxylic acids suppressed the initial uptake. In conclusion, L-lactic acid transport in the basal membrane is H+ dependent and Na+ independent, as is also the case for the brush-border membrane transport, and its characteristics resemble those of monocarboxylic acid transporters. However, there were several differences in the effects of inhibitors between basal and brush-border membrane vesicles, suggesting that the transporter(s) involved in L-lactic acid transport in the basal membrane of placental trophoblast may differ from those in the brush-border membrane.

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Contribution of P-glycoprotein to efflux of ramosetron, a 5-HT<inf>3</inf> receptor antagonist, across the blood-brain barrier

Yamamoto C., Murakami H., Koyabu N., Takanaga H., Matsuo H., Uchiumi T., Kuwano M., Naito M., Tsuruo T., Ohtani H., Sawada Y.

Journal of Pharmacy and Pharmacology (Journal of Pharmacy and Pharmacology) 54 ( 8 ) 1055 - 1063 2002.08

ISSN 00223573

In-situ rat and mouse brain perfusion data indicated that the brain distribution of ramosetron (R-ramosetron), a 5-hydroxytryptamine3 (5-HT3) receptor antagonist, was extremely low compared with that expected from its lipophilicity. We hypothesized the involvement of an efflux system(s) and investigated the contribution of P-glycoprotein to efflux transport of ramosetron across the blood-brain barrier by means of an in-vitro uptake study in cell lines that over-express P-glycoprotein. We examined the contribution of mdr1a, mdr1b and MDR1 P-glycoprotein by using LV500 cells, MBEC4 cells and LL-GA5-COL300 cells, which over-express mdr1a P-glycoprotein, mdr1b P-glycoprotein and MDR1 P-glycoprotein, respectively. The uptake of [14C]ramosetron by LV500 cells and LLC-GA5-COL300 cells was significantly lower than that by the respective parental cells. Next, we studied the effects of P-glycoprotein inhibitors, verapamil and ciclosporin, on uptake of [14C] ramosetron by these cell lines. The uptake of [14C]ramosetron by LV500 cells and LLC-GA5-COL300 cells was significantly increased in the presence of verapamil or ciclosporin, while verapamil did not affect the uptake of [14C]ramosetron by MBEC4 cells. These results indicate that the efflux of [14C]ramosetron is partly mediated by mdr1a P-glycoprotein, but not by mdr1b P-glycoprotein, and that there is a difference in substrate specificity between mdr1a P-glycoprotein and mdr1b P-glycoprotein. Further, [14C]ramosetron was confirmed to be effluxed by human MDR1 P-glycoprotein. We conclude that the limited distribution of ramosetron to the brain is due, at least in part, to efflux mediated by the P-glycoprotein at the blood-brain barrier.

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Role of human MDR1 gene polymorphism in bioavailability and interaction of digoxin, a substrate of P-glycoprotein

Kurata Y., Ieiri I., Kimura M., Morita T., Irie S., Urae A., Ohdo S., Ohtani H., Sawada Y., Higuchi S., Otsubo K.

Clinical Pharmacology and Therapeutics (Clinical Pharmacology and Therapeutics) 72 ( 2 ) 209 - 219 2002.08

ISSN 00099236

Objective: Our objective was to quantitate the contribution of the genetic polymorphism of the human MDR1 gene to the bioavailability and interaction profiles of digoxin, a substrate of P-glycoprotein. Methods: The pharmacokinetics of digoxin was studied in 15 healthy volunteers, who were divided into 3 groups (n = 5 each) on the basis of genotyping for the MDR1 gene, in a 4-dose study after single doses of digoxin alone (0.5 mg orally and intravenously) and coadministered with clarithromycin (400 mg orally for 8 days). The dose of digoxin was reduced during the clarithromycin phase (0.25 mg orally and intravenously). Results: The bioavailability of digoxin in G/G2677C/C3435, G/T2677C/T3435, and T/T2677T/T3435 subjects were 67.6% ± 4.3%, 80.9% ± 8.9%, and 87.1% ± 8.4%, respectively, and the difference between G/G2677C/C3435 and T/T2677T/T3435 subjects was statistically significant (P < .05). The MDR1 variants were also associated with differences in disposition kinetics of digoxin, with the renal clearance being almost 32% lower in T/T2677T/T3435 subjects (1.9 ± 0.1 mL/min per kilogram) than G/G2677C/C3435 subjects (2.8 ± 0.3 mL/min per kilogram), and G/T2677C/T3435 subjects having an intermediate value (2.1 ± 0.6 mL/min per kilogram). Coadministration of clarithromycin did not consistently affect digoxin clearance or renal clearance. However, a significant increase in digoxin bioavailability was observed in G/G2677C/C3435 subjects (67.6% ± 4.3% versus 85.4% ± 6.1%; P < .05) but not in the other 2 genotype groups. Conclusion: The allelic variants in the human MDR1 gene are likely to be associated with altered absorption and/or disposition profiles of digoxin and P-glycoprotein-mediated drug interaction.

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Transluminal gene transfer into brain capillary endothelial cells in vivo with HVJ-liposomes

Jiang C., Matsuo H., Koyabu N., Ohtani H., Fujimoto H., Yonemitsu Y., Kaneda Y., Naito M., Tsuruo T., Sawada Y.

Journal of Drug Targeting (Journal of Drug Targeting) 10 ( 4 ) 345 - 352 2002.07

ISSN 1061186X

Bioactive proteins or peptides cannot be effectively delivered into brain capillary endothelial cells (BCECs) or brain parenchyma. In this study, we selectively transferred Escherichia coli β-galactosidase gene (β-gal) as a model gene into BCECs by using the hemagglutination virus of Japan (HVJ)-liposomes. HVJ-liposomes encapsulating a β-gal plasmid were used to transfect MBEC4 cells in vitro, and were administrated via the internal carotid artery to rat in vivo. Success of the procedure was confirmed by the detection of 116 kDa β-gal protein in transfected MBEC4 cells and in brain capillaries isolated from transfected rats, by Western blot analysis and histological staining. The enzymatic activities of β-galactosidase were 5- to 10-fold and 20-fold higher than when β-gal-containing liposomes without fusogenic activity (uncoated liposomes) or plasmid alone were employed in vitro and in vivo, respectively. Thus, HVJ-liposomes were demonstrated to be a useful vector to transfer a foreign gene into the brain capillary endothelium in vivo via the transluminal route.

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Distribution of domperidone into the rat brain is increased by brain ischaemia or treatment with the P-glycoprotein inhibitor verapamil

Dan Y., Murakami H., Koyabu N., Ohtani H., Sawada Y.

Journal of Pharmacy and Pharmacology (Journal of Pharmacy and Pharmacology) 54 ( 5 ) 729 - 733 2002.05

ISSN 00223573

Domperidone (DOM), a peripheral dopamine D2 receptor antagonist, is a substrate of P-glycoprotein (P-gp). Therefore, the transport of DOM into the brain may be restricted by P-gp function at the blood-brain barrier, and when the function of P-gp is impaired by ATP depletion under conditions of brain ischaemia (e.g. cerebral thrombosis), side-effects may be induced as a result of increased distribution of DOM into the brain. In this study, we investigated the effects of brain ischaemia and verapamil, a P-gp inhibitor, on the permeability coefficient-surface area product (PS values) of DOM across the blood-brain barrier by using an in-situ rat brain perfusion technique. The PS values of DOM were increased 3.4- and 3.6-fold after brain ischaemia for 10 and 20 min, respectively. Furthermore, co-administration of verapamil significantly increased the PS values of DOM by 42.6- and 43.3-fold in the normal and ischaemia groups, respectively. Brain vascular volume was not affected by ischaemia or verapamil. These results show that the transport of DOM into the brain is restricted by P-gp and that the brain distribution of DOM can be increased by cerebral ischaemia or co-administration of a P-gp inhibitor.

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Inhibition of P-glycoprotein by flavonoid derivatives in adriamycin-resistant human myelogenous leukemia (K562/ADM) cells

Ikegawa T., Ohtani H., Koyabu N., Juichi M., Iwase Y., Ito C., Furukawa H., Naito M., Tsuruo T., Sawada Y.

Cancer Letters (Cancer Letters) 177 ( 1 ) 89 - 93 2002.03

ISSN 03043835

We investigated the effects of natural flavones, quercetin and morin, and their pentamethyl, pentaethyl, pentapropyl, pentabutyl and pentaallyl ethers, on the function of P-glycoprotein (P-gp) assessed by an increase in the uptake of [3H]vincristine by human myelogenous leukemia (K562) cells and adriamycin-resistant human myelogenous leukemia (K562/ADM) cells. Pentamethyl, pentaethyl, pentapropyl and pentaallyl ethers of morin and quercetin (20 μM) all increased the uptake of [3H]vincristine by K562/ADM cells, while quercetin, morin and their pentabutyl ethers had no effect. Pentamethylquercetin, pentaallylquercetin and pentaethylmorin remarkably increased the uptake of [3H]vincristine by K562/ADM cells by 10.6, 10.8 and 14.4-fold, respectively. These inhibitory potencies for P-gp were more potent than typical P-gp inhibitors, cyclosporine A and verapamil. Taking into consideration that these flavonoid derivatives possess antitumor promoter activity, they may become candidates of effective multidrug resistance-reversing agents in cancer chemotherapy. © 2002 Elsevier Science Ireland Ltd. All rights reserved.

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Proton gradient-dependent transport of valproic acid in human placental brush-border membrane vesicles

Nakamura H., Ushigome F., Koyabu N., Satoh S., Tsukimori K., Nakano H., Ohtani H., Sawada Y.

Pharmaceutical Research (Pharmaceutical Research) 19 ( 2 ) 154 - 161 2002.03

ISSN 07248741

Purpose. To investigate the transport mechanism of valproic acid across the human placenta, we used human placental brush-border membrane vesicles and compared them with that of lactic acid. Methods. Transport of [3H]valproic acid and [14C]lactic acid was measured by using human placental brush-border membrane vesicles. Results. The uptakes of [3H]valproic acid and [14C]lactic acid into brush-border membrane vesicles were greatly stimulated at acidic extravesicular pH. The uptakes of [3H]valproic acid and [14C]lactic acid were inhibited by various fatty acids, p-chloromercuribenzene sulfonate, α-cyano-4-hydroxycinnamate, and FCCP. A kinetic analysis showed that it was saturable, with Michaelis constants (Kt) of 1.04 ± 0.41 mM and 1.71 ± 0.33 mM for [3H]valproic acid and [14C]lactic acid, respectively. Furthermore, lactic acid competitively inhibited [3H]valproic acid uptake and vice versa. Conclusion. These results suggest that the transport of valproic acid across the microvillous membrane of human placenta is mediated by a proton-linked transport system that also transports lactic acid. However, some inhibitors differentially inhibited the uptakes of [3H]valproic acid and [14C]lactic acid, suggesting that other transport systems may also contribute to the elevated fetal blood concentration of valproic acid in gravida.

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Ejects of Single and Repeated Treatment with Itraconazole on the Pharmacokinetics of Midazolam in Rats

Sawada Y., Takedomi S., Matsuo H., Ohtani H., Yamano K., Iga T.

Drug Metabolism and Pharmacokinetics (Drug Metabolism and Pharmacokinetics) 17 ( 4 ) 275 - 283 2002

ISSN 13474367

To estimate the influence of repeated administration of drug metabolism inhibitors on the extent of drug interaction, we investigated the ejects of single intravenous or repeated oral administration of itraconazole on the pharmacokinetics of midazolam in rats. In the single administration study, the plasma concentration of itraconazole was maintained by intravenous infusion, and midazolam was administered into the portal vein to investigate its kinetics. In the repeated administration study, the kinetics of midazolam was investigated after seven-day oral treatment with itraconazole. The in vitro metabolism of midazolam and the contents of cytochrome P450 were investigated using liver microsomes from the itraconazole-treated rats. © 2002, The Japanese Society for the Study of Xenobiotics. All rights reserved.

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Effects of grapefruit juice and orange juice on the intestinal efflux of P-glycoprotein substrates

Tian R., Koyabu N., Takanaga H., Matsuo H., Ohtani H., Sawada Y.

Pharmaceutical Research (Pharmaceutical Research) 19 ( 6 ) 802 - 809 2002

ISSN 07248741

Purpose. The aim of this study is to investigate the effects of 50% ethyl acetate extracts of grapefruit juice (GFJ) and orange juice (OJ) on the transport activity of P-glycoprotein (P-gp) in the rat small intestine. Methods. The efflux of P-gp substrates from rat everted sac in the absence or presence of verapamil, GFJ, OJ or erythromycin was measured. Rhodamine123, fexofenadine and saquinavir were used as P-gp substrates. P-gp expression levels in the rat jejunum and ileum were determined by Western blot analysis. Results. The efflux of rhodamine123 from the everted sac increased from the apex of the jejunum to the low ileum and the expression of P-gp in the ileum was 2.31-fold higher than that in the jejunum. Verapamil and the 50% GFJ and OJ extracts inhibited the efflux from the intestine of all three drugs tested. Erythromycin decreased the efflux of rhodamine123 and fexofenadine, but did not affect the efflux of saquinavir in the intestine. Conclusions. GFJ and OJ extracts inhibited the efflux of P-gp substrates from the small intestine. Therefore, they may enhance the oral bioavailability of P-gp substrates by increasing absorption in the small intestine.

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Polymorphism of MDR1 Gene in Healthy Japanese Subjects: A Novel SNP with an Amino Acid Substitution (Glu108Lys)

Honda T., Dan Y., Koyabu N., Ohtani H., Sawada Y., Ieiri I., Otsubo K., Higuchi S.

Drug Metabolism and Pharmacokinetics (Drug Metabolism and Pharmacokinetics) 17 ( 5 ) 479 - 481 2002

ISSN 13474367

We discovered a novel single nucleotide polymorphism (SNP) at position 325 (G325A) in exon 5 of the multidrug-resistance 1 (MDR1) gene in a study of 37 healthy Japanese subjects. Details are as follows. SNP, 020614Honda001; GENE NAME, human P-glycoprotein (MDR1); ACCESSION NUMBER, M29427; LENGTH, 25 bases; 5' -ATGAATCTGGAGG/AAAGACATGACCA-3'. This SNP is expected to cause an amino acid substitution (Glu108Lys). In this study, one homozygote and one heterozygote for G325A were identified. © 2002, The Japanese Society for the Study of Xenobiotics. All rights reserved.

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H<sup>+</sup>-linked transport of salicylic acid, an NSAID, in the human trophoblast cell line BeWo

Emoto A., Ushigome F., Koyabu N., Kajiya H., Okabe K., Satoh S., Tsukimori K., Nakano H., Ohtani H., Sawada Y.

American Journal of Physiology - Cell Physiology (American Journal of Physiology - Cell Physiology) 282 ( 5 51-5 ) C1064 - 75 2002

ISSN 03636143

We investigated the transport of salicylic acid and L-lactic acid across the placenta using the human trophoblast cell line BeWo. We performed uptake experiments and measured the change in intracellular pH (pHi). The uptakes of [14C] salicylic acid and L-[14C] lactic acid were temperature- and extracellular pH-dependent and saturable at higher concentrations. Both uptakes were also reduced by FCCP, nigericin, and NaN3. Various nonsteroidal anti-inflammatory drugs (NSAIDs) strongly inhibited the uptake of L-[14C] lactic acid. Salicylic acid and ibuprofen noncompetitively inhibited the uptake of L-[14C] lactic acid. α-Cyano-4-hydroxycinnamate (CHC), a monocarboxylate transporter inhibitor, suppressed the uptake of L-[14C] lactic acid but not that of [14C] salicylic acid. CHC also suppressed the decrease of pHi induced by L-lactic acid but had little effect on that induced by salicylic acid or diclofenac. These results suggest that NSAIDs are potent inhibitors of lactate transporters, although they are transported mainly by a transport system distinct from that for L-lactic acid.

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Transcellular transport of genistein, a soybean-derived Isoflavone, across human colon carcinoma cell line

Oitate M., Nakaki R., Koyabu N., Takanaga H., Matsuo H., Ohtani H., Sawada Y.

Biopharmaceutics and Drug Disposition (Biopharmaceutics and Drug Disposition) 22 ( 1 ) 23 - 29 2001.12

ISSN 01422782

Genistein, a soybean-derived isoflavone, is thought to have an anticarcinogenic action, but little is known about the cellular mechanisms of its intestinal absorption. This study was designed to investigate the absorption mechanisms of genistein using human colon carcinoma cell line, Caco-2 cells. The apical-to-basolateral transcellular transport of genistein across a Caco-2 cell monolayer was significantly greater than that in the opposite direction. An uptake experiment revealed that cellular uptake of genistein by Caco-2 cells was concentrative. The transcellular transport of genistein was saturable and temperature-dependent, and was inhibited by other flavonoids such as rutin, quercetin, (+)-catechin and (-)-epicatechin. These results suggest that genistein is transported across Caco-2 cells by a carrier-mediated system, located on the apical membrane. Copyright © 2001 John Wiley & Sons, Ltd.

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Quantitative relationship between myocardial concentration of tacrolimus and QT prolongation in guinea pigs: Pharmacokinetic/pharmacodynamic model incorporating a site of adverse effect

Minematsu T., Ohtani H., Yamada Y., Sawada Y., Sato H., Iga T.

Journal of Pharmacokinetics and Pharmacodynamics (Journal of Pharmacokinetics and Pharmacodynamics) 28 ( 6 ) 533 - 554 2001.12

ISSN 1567567X

Clinical cases have been reported of tacrolimus (FK506)-induced QT prolongation. We have previously demonstrated sustained QT prolongation by FK506 in guinea pigs. Herein, we aimed to conduct a pharmacokinetic/pharmacodynamic (PK/PD) analysis of FK506, using a model involving the myocardial compartment. The pharmacokinetics of FK506 and its effects on QTc intervals were investigated in guinea pigs. In the pharmacokinetic study, whole blood and ventricular FK506 concentrations were analyzed, using a 4-compartment model during and after intravenous infusion of FK506 (0.01 or 0.1 mg/hr/kg). Subsequently, the concentration-response relationship between ventricular FK506 concentration and change in QTc interval was analyzed, using the maximal effect (Emax) model. Pharmacokinetic profiles of FK506 showed a delayed distribution of FK506 into the ventricle. Furthermore, the observed QT prolongation paralleled the ventricular FK506 concentrations, with no lag-time between the two. The Emax model successfully described the relationship between changes in QTc interval and ventricular FK506 concentrations. In conclusion, the PK/PD model where the myocardial drug concentration of FK506 was linked with its adverse effect could describe, for the first time, the anti-clockwise hysteresis observed in the relationship between blood FK506 concentration and QT prolongation. Such a hysteresis pattern for QT prolongation might be caused, therefore, mainly by the delayed disposition of FK506 to ventricular myocytes.

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Pharmacokinetic and pharmacodynamic analysis of the antihypercalcemic effect of incadronate disodium in rats

Ohno Y., Yamada Y., Usui T., Takahashi K., Tsuchiya F., Ohtani H., Sato H., Sawada Y., Iga T.

Biological and Pharmaceutical Bulletin (Biological and Pharmaceutical Bulletin) 24 ( 11 ) 1290 - 1293 2001.11

ISSN 09186158

Incadronate concentrates into the bone as a target organ after intravenous administration of incadronate disodium. Mature osteoclasts has take up incadronate from the bone surface and convert it from an active to an inactive form. As a result, incadronate decreases the plasma calcium concentration by suppressing bone resorption. In this study, the pharmacokinetic and pharmacodynamic (PK/PD) analysis model for ascertaining the anti-hypercalcemic effects of incadronate disodium was developed in rats. Data on both the concentration of incadronate in bone and that of free calcium in blood after intravenous administration from our previous study were used for analysis. To estimate the concentration in the surface layer of bone, data on the concentration of incadronate in bone after single intravenous administration were analyzed based on the PK model considering three-compartments. The estimated concentrations in the surface layer in bone were applied to the PD model as an input function. The PD model was developed to analyze the changes in the plasma calcium concentration after a single intravenous administration considering an irreversible inhibition of osteoclast activity. The obtained fitted curves were in good agreement with the observed data. The model could explain the long duration of the antihypercalcemic effect of incadronate disodium and should be useful for planning rational dose regimens for effective antihypercalcemic therapy.

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