”骨髄腫幹細胞ニッチ”の解析と治療標的分子の同定

2013 Fiscal Year Annual Research Report

• Project/Area Number: 23591404
• Research Institution: Kyoto Pharmaceutical University
• Principal Investigator: 芦原 英司 京都薬科大学, 薬学部, 教授 (70275197)
• Co-Investigator(Kenkyū-buntansha): 平位 秀世 京都大学, 医学(系)研究科(研究院), 助教 (50315933)
• Keywords: 多発性骨髄腫 / 骨髄微小環境 / 低酸素環境 / TGF-β

Research Abstract

多発性骨髄腫（MM）の病態維持に重要なMM幹細胞の骨髄低酸素環境の動態、および治療標的分子の同定のため、以下の検討を行ってきた。まず正所性MM担がんマウスを作製し、MM細胞の骨髄内の動態を解析した。MM細胞は海綿骨周囲の骨内膜下に生着し、低酸素マーカーであるピモニダゾール陽性を示した。このことより、MM細胞は骨髄微小環境下では低酸素状態であることが明らかとなった。低酸素下骨髄微小環境で生存するMM細胞の性状をin vitro系で解析するため、1%低酸素状態下で長期生存可能な低酸素適応MM（HA-MM）細胞株6株（AMO-1、OPM-2、NCIH929、RPMI8226、U266、IM-9）を樹立した。それぞれの細胞増殖を解析したところ、低酸素環境では細胞増殖スピードはそれぞれHA株で有意に遅いことが明らかとなった。Ki67/7-AADを用いた二重染色による細胞周期解析では、親株であるNormoxia下での培養細胞に比して、G0期の細胞集団が有意に増加しており、またSP分画細胞集団も有意に増加していた。さらにOct3/4、Sox2といった幹細胞マーカーのmRNAの発現亢進を認めた。またlimiting dilution法を用いた正所性MM担がんマウス作製にて、HA-MM細胞株では親株と比して、より少ない細胞数ででも生存期間の有意な短縮を認め、これらのことから、HA-MM細胞株はMM幹細胞の性質を有することが明らかとなった。低酸素過去の報告のようなCD138抗原の発現の低下は認めなかった。接着因子としてはCD44の発現亢進を認めた。またTGFβシグナルの下流のSmad2のリン酸化亢進を認め、MMにおいてはTGF-β/Smadシグナル系がMM幹細胞の治療標的である可能性が示唆された。

Research Products

• [Journal Article] Myeloid-Derived Suppressor Cells Play Crucial Roles in the Regulation of Mouse Collagen-Induced Arthritis. (2013)
  • Author(s): 1. Fujii W, Ashihara E, Hirai H, Nagahara H, Kajitani N, Fujioka K, Murakami K, Seno T, Yamamoto A, Ishino H, Kohno M, Maekawa T, Kawahito Y.
  • Journal Title: J Immunol Volume: 191 Pages: 1073-1081
  • DOI: doi: 10.4049/jimmunol.1203535.
  • Peer Reviewed
• [Journal Article] Quantitative Comparison of Endogenous Protein Dynamics between in Live Cell and in vitro by in cell 19F-NMR. (2013)
  • Author(s): 3. Takaoka Y, Kioi Y, Morito A, Ohtani J, Arita K, Ashihara E, Ariyoshi M, Tochio H, Shirakawa M, Hamachi I.
  • Journal Title: ChemCom Volume: 49 Pages: 2801-2803
  • DOI: doi: 10.1039/c3cc39205h.
  • Peer Reviewed
• [Journal Article] Cyclic AMP responsive element binding proteins are involved in 'emergency' granulopoiesis through the upregulation of CCAAT/enhancer binding protein β. (2013)
  • Author(s): 4. Hirai H, Kamio N, Huang G, Matsusue A, Ogino S, Kimura N, Satake S, Ashihara E, Imanishi J, Tenen DG, Maekawa T.
  • Journal Title: PLoS One Volume: 8 Pages: e54862
  • DOI: doi: 10.1371/journal.pone.0054862.
  • Peer Reviewed
• [Journal Article] C/EBPβ promotes BCR-ABL-mediated myeloid expansion and leukemic stem cell exhaustion. (2013)
  • Author(s): 5. Hayashi Y, Hirai H, Kamio N, Yao H, Yoshioka S, Miura Y, Ashihara E, Fujiyama Y, Tenen DG, Maekawa T.
  • Journal Title: Leukemia Volume: 27 Pages: 619-628
  • DOI: doi: 10.1038/leu.2012.258.
  • Peer Reviewed
• [Journal Article] Effectiveness of sulforaphane as a radiosensitizer for murine osteosarcoma cells. (2013)
  • Author(s): 6. Sawai Y, Murata H, Horii M, Koto K, Matsui T, Horie N, Tsuji Y, Ashihara E, Maekawa T, Kubo T, Fushiki S.
  • Journal Title: Oncol Rep Volume: 29 Pages: 941-945
  • DOI: doi: 10.3892/or.2012.2195.
  • Peer Reviewed
• [Journal Article] Therapeutic effects of human mesenchymal and hematopoietic stem cells on rotenone-treated parkinsonian mice. (2013)
  • Author(s): 7. Inden M, Takata K, Nishumura K, Kitamura Y, Ashihara E, Yoshimoto K, Ariga H, Nonmou O, Shimohama S.
  • Journal Title: J Neurosci Res Volume: 91 Pages: 62-72
  • DOI: doi: 10.1002/jnr.23128.
  • Peer Reviewed
• [Journal Article] Zoledronic acid significantly enhances radiation induced apoptosis against human fibrosarcoma cells by inhibiting radioadaptive signaling. (2013)
  • Author(s): 8. Koto K, Murata H, Kimura S, Sawai Y, Horie N, Matsui T, Ryu K, Ashihara E, Maekawa T, Kubo T, Fushiki S.
  • Journal Title: Int J Oncol Volume: 42 Pages: 525-534
  • DOI: doi: 10.3892/ijo.2012.1735.
  • Peer Reviewed
• [Journal Article] 非小細胞性肺がん患者に対する簡易懸濁法を用いたエルロチニブ投与に関する検討． (2013)
  • Author(s): 高田哲也、池見泰明、福土将秀、杉本充弘、石橋直哉、小林政彦、矢野育子、金永学、三嶋理晃、芦原英司、松原和夫.
  • Journal Title: 医療薬学 Volume: 39 Pages: 565-570
  • Peer Reviewed
• [Presentation] Hypoxia-adapted myeloma cells possess stem cell-like character. (2013)
  • Author(s): 1. Ashihara E, Nakagawa Y, Yao H, Yokota A, Miura Y, Takata K, Kitamura Y, Hirai H, Maekawa T.
  • Organizer: The 55th Annual Meeting of American Society of Hematology.
  • Place of Presentation: New Orleans, USA
  • Year and Date: 20131207-20131210
• [Presentation] Direct interaction with bone marrow mesenchymal stromal/stem cells is required for hematopoietic expansion by parathyroid hormone. (2013)
  • Author(s): 2. Yao H, Miura Y, Yoshioka S, Iwasa M, Sato A, Tamura A, Yokota A, Ashihara E, Ichinohe T, Hirai H, Maekawa T.
  • Organizer: The 55th Annual Meeting of American Society of Hematology.
  • Place of Presentation: New Orleans, USA
  • Year and Date: 20131207-20131210
• [Presentation] Expression of C/EBPβ is involved in the exhaustion of hematopoietic stem cells through cell-intrinsic and cell-extrinsic mechanisms. (2013)
  • Author(s): 3. Tamura A, Hirai H, Hayashi Y, Yokota A, Sato A, Yao H, Yoshioka S, Iwasa M, Ashihara E, Miura Y, Maekawa T.
  • Organizer: The 55th Annual Meeting of American Society of Hematology.
  • Place of Presentation: New Orleans, USA
  • Year and Date: 20131207-20131210
• [Presentation] Cytokine-STATs signalings upregulate C/EBPβ in BCR-ABL+ leukemic cells independently from BCR-ABL/JAK-STAT pathway. (2013)
  • Author(s): 4. Yokota A, Hirai H, Hayashi Y, Tamura A, Sato A, Yao H, Yoshioka S, Iwasa M, Ashihara E, Miura Y, Maekawa T.
  • Organizer: The 55th Annual Meeting of American Society of Hematology.
  • Place of Presentation: New Orleans, USA
  • Year and Date: 20131207-20131210
• [Presentation] Microglia-like monocytic cells derived from bone marrow cells phagocytose amyloid-β and facilitate phagocytosis of amyloid-β by resident microglia. (2013)
  • Author(s): 5. Takata K, Toji Y, Kawanishi S, Takada T, Kitamura Y, Ashihara E.
  • Organizer: The 55th Annual Meeting of American Society of Hematology.
  • Place of Presentation: New Orleans, USA
  • Year and Date: 20131207-20131210
• [Presentation] M-CSF刺激による造血幹細胞からのアミロイドβ貪食細胞への分化誘導. (2013)
  • Author(s): 高田和幸、河西翔平、杜氏裕美子、北村佳久、芦原英司
  • Organizer: 第75回日本血液学会学術総会
  • Place of Presentation: 札幌
  • Year and Date: 20131011-20131013
• [Presentation] Glioblastoma-derived exosomes targeting to cancer cells and their lipid characterization. (2013)
  • Author(s): Toda Y,Takata K, Kobayashi K, Hattori Y,Akaji K, Ashihara E.
  • Organizer: 第72回日本癌学会学術総会
  • Place of Presentation: 横浜
  • Year and Date: 20131003-20131005
• [Presentation] Myeloid-Derived Suppressor Cells have regulatory roles in mouse collagen-induced arthritis. (2013)
  • Author(s): 6. Fuji W, Ashihara E, Hirai H, Nagahara H, Fujioka K, Murakami K, Nakamura K, Seno T, Yamamoto A, Ishino H, Kohno M, Maekawa T, Kawahito Y.
  • Organizer: The Annual European Congress of Rheumatology.
  • Place of Presentation: Madrid, Spain.
  • Year and Date: 20130612-20130615
• [Presentation] がん細胞指向性を有するヒト星状膠芽腫細胞由来エキソソームの発見．
  • Author(s): 戸田侑紀、高田和幸、中川由布子、小林数也、服部恭尚、北村佳久、赤路健一、芦原英司．
  • Organizer: 第5回日本RNAi研究会
  • Place of Presentation: 広島
• [Book] 多発性骨髄腫 Updating．第4巻 移植適応骨髄腫の治療．~新規薬剤時代における移植適応患者の治療戦略~ (2013)
  • Author(s): 芦原英司．
  • Total Pages: 203
  • Publisher: 医療ジャーナル社
• [Book] RNA Interference for Oncology: Clinical Prospects Beyond the Hype. In: Emerging Trends in Cell and Gene Therapy. (2013)
  • Author(s): Ashihara E, Maekawa T.
  • Total Pages: 705
  • Publisher: Springer