ABC蛋白スーパーファミリーによる異物排出機構の解析

• Project/Area Number: 10217202
• Research Category: Grant-in-Aid for Scientific Research on Priority Areas
• Allocation Type: Single-year Grants
• Review Section: Biological Sciences
• Research Institution: The University of Tokyo
• Principal Investigator: 鈴木 洋史 東京大学, 大学院・薬学系研究科, 助教授 (80206523)
• Co-Investigator(Kenkyū-buntansha): 伊藤 晃成 千葉大学, 大学院・薬学研究院, 助手 (30323405) ; 加藤 将夫 東京大学, 大学院・薬学系研究科, 助手 (30251440)
• Project Period (FY): 1998 – 2002
• Project Status: Completed (Fiscal Year 2002)
• Budget Amount: ¥45,200,000 (Direct Cost: ¥45,200,000); Fiscal Year 2002: ¥8,600,000 (Direct Cost: ¥8,600,000); Fiscal Year 2001: ¥9,000,000 (Direct Cost: ¥9,000,000); Fiscal Year 2000: ¥9,200,000 (Direct Cost: ¥9,200,000); Fiscal Year 1999: ¥9,200,000 (Direct Cost: ¥9,200,000); Fiscal Year 1998: ¥9,200,000 (Direct Cost: ¥9,200,000)
• Keywords: 胆汁排泄 / 経細胞輸送 / ベクトル輸送 / ABCトランスポーター / BCRP / 硫酸抱合体 / MRP2 / MRP3 / 有機アニオン / 胆汁酸 / 極性輸送 / BSEP / trans-inhibition / 肝潅流 / 有機アニオン輸送 / 胆汁酸輸送 / 血液脳関門 / 小腸上皮細胞 / 多剤耐性 / MRP1 / MRP5 / GS-Xポンプ

Research Abstract

我々はすでに異物解毒に関与するMRP(ABCC)ファミリーについて検討を加え、これらが細胞内で生成されたグルクロン酸抱合体・グルタチオン抱合体の排出に関与することを示してきた。今年度は特にMRP2および3の機能についても検討を加えたが、現在まで未解明であった硫酸抱合体の排出機構についても検討を加えた。我々はbreast cancer resistance protein(BCRP/ABCG2)が正常細胞のapical側膜に発現することに着目し、この蛋白質の機能について検討を加えた。その結果、BCRPが生体内因化合物estrone-3-sulfateやdehydroepiandrosterone sulfateを基質とすることを見出した。これはABCトランスポーターが硫酸抱合体を基質とすることを示した始めての例である。さらに、estrone-3-sulfateに対する阻害実験を行ったところ、一連の硫酸抱合体は低いIC_<50>値により阻害をかけたのに対し、グルクロン酸抱合体やグルタチオン抱合体はより高いIC_<50>値を示した。さらに、4-methylumbelliferone sulfateやE3040 sulfateはBCRPにより顕著に取り込まれた。一方、4-methylumbelliferone glucuronideやE3040 glucuronideはBCRPにより有意に取り込まれるものの、取り込みの絶対値は硫酸抱合体に比べて顕著に低いものであった。これらの結果は、BCRPが硫酸抱合体の排出に関与することを示している。MRPがグルクロン酸抱合体・グルタチオン抱合体を基質とすることから、これらのトランスポーターは互いに相補的に機能することにより、異物解毒に関与することが示唆された。

Research Products

• [Publications] Akita H et al.: "Sinusoidal Efflux of Taurocholate Correlates with the Hepatic Expression Level of Mrp3"Biochem Biophys Res Commun. 299. 681-687 (2002)
• [Publications] Suzuki H, Sugiyama Y: "Single nucleotide polymorphisms in multidrug resistance associated protein 2 (MRP2/ABCC2) : Its impact on drug disposition"Adv Drug Deliv Rev. 54. 1311-1331 (2002)
• [Publications] Xiong H et al.: "Mechanisms of impaired biliary excretion of acetaminophen glucuronide after acute phenobarbital treatment or phenobarbital pretreatment"Drug Metab Dispos. 30. 962-969 (2002)
• [Publications] Ji B et al.: "Multidrug resistance-associated protein 2 (MRP2) plays an important role in the biliary excretion of glutathione conjugates of 4-hydroxynonenal"Free Radic Biol Med. 33. 370-378 (2002)
• [Publications] Sasaki, M., Suzuki, H., Ito, K., Abe, T., Sugiyama, Y.: "Transcellular Transport of Organic Anions Across a Double-transfected Madin-Darby Canine Kidney II Cell Monolayer Expressing Both Human Organic Anion-transporting Polypeptido (OATP2/SLC21A6) and Multidrug Resistance-associated Protein 2 (MRP2/ABCC2)"J. Biol. Chem.. (in press). (2002)
• [Publications] Itoda, M., Saito, Y., Soyama, A., Saeki, M., Murayama, N., Ishida, S., Sai, K., Nagano, M., Suzuki, H., Sugiyama, Y., Ozawa, S., Sawada: "J. Polymorphisms in the ABCC2 (cMOAT/MRP2) gene found in 72 established cell lines derived from Japanese individuals-An association between single nucleotide polymorphisms in the 5'-untranslated region and exon 28"Drug Metab. Dispos.. (in press). (2002)
• [Publications] Akita, H., Suzuki, H., Hirohashi, T., Takikawa, T., Sugiyama, Y.: "Transport activity of human MRP3 expressed in Sf9 cells : Comparative studies with rat MRP3"Pharm. Res.. 19. 34-41 (2002)
• [Publications] Ito, K., Suzuki, H., Sugiyama, Y.: "Single amino acid substitution of rat MRP2 results in acquired transport activity for taurocholate"Am. J. Physiol.. 281. G1034-G1043 (2001)
• [Publications] Akita, H., Suzuki, H., Sugiyama, Y.: "Sinusoidal efflux of taurocholate is enhanced in Mrp2-deficient rat liver"Pharm. Res.. 18. 1119-1125 (2001)
• [Publications] T.Hirohashi: "ATP-dependent transport of bile salts by rat multidrug resistance-associated protein 3 (MRP3)."J.Biol.Chem.. 275. 2905-2910 (2000)
• [Publications] H.Akita: "Characterization of bile acid transport mediated by multidrug resistance associated protein 2 ahd bile salt export pump."Biochim.Biophys.Acta. 1511. 7-16 (2001)
• [Publications] H.Suzuki: "Transport of drugs across the hepatic sinusoidal membrane : Sinusoidal drug influx and efflux in the liver."Semin.Liv.Dis.. 20. 251-263 (2000)
• [Publications] H.Suzuki: "Hepatocellular extrusion of bile acids by ABC transporters."In "Biology of bile acids in health and disease" ed. by van Berge Henegouwen GP, Keppler D, Leuschner U, Paumgartner G and Stiehl A (Kluwer), Kluwer Academic Publishers. (2001)
• [Publications] M. Homma: "High affinity efflux transport for glutathione conjugates on the luminal membrane of a mouse brain capillary endothelia cell line (MBEC4)"J. Pharmacol. Exp. Ther.. 288. 198-203 (1999)
• [Publications] X..-Y. Chu: "Active efflux of CPT-11 and its metabolites in human KB-derived cell lines"J. Pharmacol. Exp. Ther.. 288. 735-741 (1999)
• [Publications] T. Hirohashi: "Characterization of the transport properties of cloned rat multidrug resistance associated protein 3 (MRP3)"J. Biol. Chem.. 274. 15181-15185 (1999)
• [Publications] J. Nishino: "Transepithelial transport of organic anions across the choroid plexus : Possible involvement of organic anion transporter and multidrug resistance associated protein"J. Pharmacol. Exp. Ther.. 290. 289-294 (1999)
• [Publications] T. Hirohashi: "The function and expression of multidrug resistance associated protein (MRP) family in human colon adenocarcinoma cells (Caco-2)"J. Pharmacol. Exp. ther.. 292. 265-270 (2000)
• [Publications] T. Hirohashi: "ATP-dependent transport of bile salts by rat multidrug resistance-associated protein 3 (MRP3)"J. Biol. Chem.. 275. 2905-2910 (2000)
• [Publications] H.Suzuki: "Transporters for bile acids and organic anions"In "Membrane transporters as drug targets" edited by G.Amidon and W.Sadee, Plenum Publishing Corp., New York. 54 (1999)
• [Publications] H.Suzuki: "Excretion of GSSG and glutathione conjugates mediated by MRP1 and cMOAT/MRP2." Semin.Liv.Dis.18. 359-376 (1998)
• [Publications] X.-Y.Chu: "Active efflux of CPT-11 and its metabolites in human KB-derived cell lines." J.Pharmacol.Exp.Ther.(in press).
• [Publications] K.Ueda: "Different substrate specificity among GS-X pump family members : Comparison between MRP and novel transporter expressed on a cisplatin resistant cell (KCP-4)." Jpn.J.Cancer Res.(in press).
• [Publications] H.Suzuki: "Transporters for bile acids and organic anions. In “Menbrane transporters as drug targets" edited by G.Amidon and W.Sadee" Plenum Publishing Corp., New York.,