レトロウイルス遺伝子導入を用いた抗癌剤耐性・感受性機構の研究

• Project/Area Number: 13218137
• Research Category: Grant-in-Aid for Scientific Research on Priority Areas
• Allocation Type: Single-year Grants
• Review Section: Biological Sciences
• Research Institution: Kyoritsu University of Pharmacy (2004); Japanese Foundation For Cancer Research (2001-2003)
• Principal Investigator: 杉本 芳一 共立薬科大学, 薬学部, 教授 (10179161)
• Project Period (FY): 2001 – 2004
• Project Status: Completed (Fiscal Year 2004)
• Budget Amount: ¥36,500,000 (Direct Cost: ¥36,500,000); Fiscal Year 2004: ¥9,500,000 (Direct Cost: ¥9,500,000); Fiscal Year 2003: ¥9,500,000 (Direct Cost: ¥9,500,000); Fiscal Year 2002: ¥9,000,000 (Direct Cost: ¥9,000,000); Fiscal Year 2001: ¥8,500,000 (Direct Cost: ¥8,500,000)
• Keywords: ABC transporter / ABCG2 / BCRP / estrogen / flavonoid / gefitinib / irinotecan / SNP / 癌化学療法 / 抗癌剤耐性 / ABCトランスポーター / ステロイドホルモン / レトロウイルス

Research Abstract

BCRP(Breast cancer resistance protein、ABCG2)はirinotecan、mitoxantrone、topotecanなどの抗癌剤の耐性に関与するABC輸送体である。我々は、BCRPがS-S結合を介したホモ2量体として存在することを明らかにし、このホモ2量体化の阻害によりBCRP機能をdominant-negativeに阻害できることを示した。また、BCRPのR482変異体発現細胞の多くがmitoxantroneとdoxorubicinに対して野生型BCRP発現細胞より高い耐性を示すことを明らかにした。次いで我々は、estrone、estradiolがBCRPによる抗癌剤耐性を克服することを明らかにした。この知見を元にBCRPによる基質輸送を解析し、BCRPはestrogen本体ではなくその硫酸抱合体であるestrone sulfateをATP依存的に輸送することを示した。さらに種々の化合物の輸送を調べ、BCRPが硫酸基を持つsteroidに選択性の高いtransporterであることを明らかにした。また、合成estrogen剤のdiethylstilbestrol、新規tamoxifen誘導体TAG-139、phytoestrogenであるgenisteinやnaringeninなどのestrogen関連化合物がBCRPの基質あるいは阻害剤となることを示した。しかし化合物の抗BCRP作用と抗estrogen作用は相関しなかった。また、EGFRのtyrosine kinase阻害剤であるgefitinibがBCRPの基質となることを示した。BCRP遺伝子のSNPの検索を行い、exon部分にBCRPタンパクの発現を消失させるBCRP-C376T(Q126stop)と発現を低下させるBCRP-C421A(Q141K)の2つのSNPを同定した。この2つのSNPが抗癌剤投与時の副作用と関係する可能性を示唆する複数の症例を同定した。estrogenがBCRP mRNAの発現低下を介さずにBCRPタンパクの発現を低下させることを見出した。阻害剤、SNP、タンパク発現の抑制など、種々の機序による生体内でのBCRPの活性の低下は、irinotecan、gefitinibなどを用いたがん化学療法における効果と副作用の発現に大きな影響を与えると考えられる。

Research Products

• [Journal Article] Antimitotic activity and reversal of breast cancer resistance protein-mediated drug resistance by stilbenoids from Bletilla striata. (2005)
  • Author(s): Morita H.
  • Journal Title: Bioorganic & Medicinal Chemistry Letters 15 Pages: 1051-1054
• [Journal Article] Estrogen-mediated Post-transcriptional Down-regulation of Breast Cancer Resistance Protein/ABCG2. (2005)
  • Author(s): Imai, Y.
  • Journal Title: Cancer Res. 65 Pages: 596-604
• [Journal Article] Induction of cellular resistance to nucleoside reverse transcriptase inhibitors by the wild-type breast cancer resistance protein. (2004)
  • Author(s): Wang, X.
  • Journal Title: Biochem.Pharmacol. 68 Pages: 1363-1370
• [Journal Article] Gefitinib reverses breast cancer resistance protein-mediated drug resistance. (2004)
  • Author(s): Yanase, K.
  • Journal Title: Mol Cancer Ther. 3 Pages: 1119-1125
• [Journal Article] Phytoestrogens/flavonoids reverse breast cancer resistance protein/ABCG2-mediated multidrug resistance. (2004)
  • Author(s): Imai, Y.
  • Journal Title: Cancer Res. 64 Pages: 4346-4352
• [Publications] Miwa, M.: "Single amino acid substitutions in the transmembrane domains of breast cancer resistance protein (BCRP) alter cross resistance patterns in the transfectants."Int.J.Cancer. 107. 757-767 (2003)
• [Publications] Imai, Y.: "Breast cancer resistance protein exports sulfated estrogens but not free estrogens."Mol.Pharmacol.. 64. 610-618 (2003)
• [Publications] Chen, Z.: "Transport of methotrexate, methotrexate polyglutamates and 17beta-estradiol 17-(beta-D-glucuronide) by ABCG2 (BCRP/MXR):Effects of acquired mutations at R482 on methotrexate transport."Cancer Res.. 63. 4048-4054 (2003)
• [Publications] Suzuki, M.: "ABCG2 transports sulfated conjugates of steroids and xenobiotics."J.Biol.Chem.. 278. 22644-22649 (2003)
• [Publications] Mukai, M.: "Reversal of the resistance to STI571 in human chronic myelogenous leukemia K562 cells."Cancer Science. 94. 557-563 (2003)
• [Publications] Sngimoto, Y.: "Drug-selected co-expression of P-glycoprotein and gp91 in vivo from an MDR1-bicistronic retrovirus vector Ha-MDR-IRES-gp91"J.Gene Med.. (in press).
• [Publications] Hong, Y.: "Construction of a high efficiency retroviral vector for gene therapy of Hunter's syndrome"J.Gene Med.. 5. 18-29 (2003)
• [Publications] Sugimoto, Y.: "Reversal of breast cancer resistance protein-mediated drug resistance by estrogen antagonists and agonists"Mol.Cancer Ther.. 2. 105-112 (2003)
• [Publications] Wang, X.: "Breast cancer resistance protein (BCRP) induces resistance to HIV-nucleoside reverse transcriptase inhibitors"Mol.Pharmacol.. 63. 65-72 (2003)
• [Publications] Che, X.F.: "Reversal of P-glycoprotein mediated multidrug resistance by a newly synthesized 1,4-benzothiazipine derivative, JTV-519"Cancer Lett.. 187. 111-119 (2002)
• [Publications] Horie, K.: "SUMO-1 conjugation to intact DNA topoisomerase I amplifies cleavable complex formation induced by camptothecin"Oncogene. 21. 7913-7922 (2002)
• [Publications] Imai, Y.: "C421A polymorphism in the human breast cancer resistant protein gene is associated with low expression of Q141K protein and low-level drug resistance"Mol.Cancer Ther.. 1. 611-616 (2002)
• [Publications] Imai, Y.: "Estrone and 17β-estradiol reverse breast cancer resistance protein-mediated multidrug resistance"Jpn.J.Cancer Res.. 93. 231-235 (2002)
• [Publications] Kage, K.: "Dominant-negative inhibition of breast cancer resistance protein as drug efflux pump through the inhibition of S-S dependent homodimerization"Int.J.Cancer. 97. 626-630 (2002)
• [Publications] Kage, K.: "Dominant-negative inhibition of breast cancer resistance protein as drug efflux pump through the inhibition of S-S dependent homodimerization"Int. J. Cancer. 97. 626-630 (2002)
• [Publications] Imai, Y.: "Estrone and 17β-estradiol reverse breast cancer resistance protein-mediated multidrug resistance"Jpn. J. Cancer Res.. (in press).
• [Publications] Sugimoto, Y.: "Bicistronic retrovirus vectors encoding drug-resistant genes. In: R.A.Aubin (ed.), Transgene Delivery and Expression in Mammalian Cells (a volume of Methods in Molecular Biology)"Humana Press (in press).