骨髄低酸素環境における骨髄腫幹細胞維持分子を標的とした新規治療開発

2013 Fiscal Year Annual Research Report

• Project Area: Integrative Research on Cancer Microenvironment Network
• Project/Area Number: 25112706
• Research Category: Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
• Research Institution: Kyoto University
• Principal Investigator: 前川 平 京都大学, 医学(系)研究科(研究院), 教授 (80229286)
• Project Period (FY): 2013-04-01 – 2015-03-31
• Keywords: 多発性骨髄腫 / 低酸素環境 / 骨髄微小環境 / がん幹細胞

Research Abstract

低酸素環境（1% O2）で長期間（6ヶ月以上）安定して生存可能な多発性骨髄腫細胞株（HA-MM）を樹立し、その性状解析を行った。HA-MM株は幹細胞マーカー（Oct3/4、Sox2、Nanog、等）のmRNAを、Normoxia環境下で生存する親株に比して有意に高発現しており、免疫不全マウスを用いた移植実験において、HA-MM細胞はより少数の細胞での定着を認め、さらに二次移植実験においても親株に比べ生着率が高く、MM幹細胞の性状を有することを明らかにした。当初、MMの治療標的として有効であると報告してきた-カテニンや慢性骨髄性白血病のHA株で治療標的分子の一つとして同定したGlyoxalase 1に注目しシグナル解析を行ってきたが、いずれも有意な差はなく、TGF-β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幹細胞維持に関与することが示唆された。現在TGF-β/Smadシグナル阻害剤にて幹細胞性維機構の破綻をもたらすか、検討を行っている。また骨髄疑似環境の再現の一歩として、ヒト骨髄間質細胞株HS5細胞との共培養系にて検討を行っている。細胞周期解析ではG0期分画の細胞集団は有意に増加するが、幹細胞マーカーの発現亢進は認めなかった。このことより、MM幹細胞の維持には間質細胞との接着より低酸素状態にあることが重要であることが考えられる。

Current Status of Research Progress

3: Progress in research has been slightly delayed.

Reason

仮説を立てたHA-MM細胞株でのWnt経路の活性化ではなくSmad経路の活性化のMM幹細胞維持機構への関与を示唆する結果を得たため、本経路分子をRNA干渉でノックダウンし解析を進めてきたが、導入効率悪くその改善に時間を要している。また低酸素適応MM細胞株の幹細胞性の検討においては、模擬骨髄作製の手技確立に時間を要することにより、若干の進捗の遅れがある。

Strategy for Future Research Activity

RNA干渉による分子機能を制御するためのsiRNAおよびshort-hairpin siRNA導入のためにNucleofection法を検討するとともに、Smad阻害剤による検討を行う。また、模擬骨髄環境の作製に並行し、ヒト骨髄間質細胞株HS5細胞との共培養系による検討を行う。HS5細胞との共培養系は予備実験を既に開始している。

Research Products

• [Journal Article] Two novel susceptibility loci to Takayasu arteritis and synergistic role of the IL12B and HLA-B regions in a Japanese population. (2013)
  • Author(s): Terao C, Yoshifuji H, Kimura A, Matsumura T, Ohmura K, Takahashi M, shimizu M, Kawaguchi T, Chen Z, Naruse TK, Sato-Otubo A, Ebana Y, Maejima Y, Kinoshita H, Murakami K, Kawabata D, Wada Y, Narita I, Tazaki J, Kawaguchi Y, Yamanaka H, Yurugi K, Miura Y, Maekawa T, et al.
  • Journal Title: Am J Human Genet Volume: 93 Pages: 1-9
  • DOI: doi:10.1016/j.ajhg.2013.05.024
  • Peer Reviewed
• [Journal Article] Activity of histone deacetylase inhibitors and an Aurora kinase inhibitor in BCR-ABL expressing leukemic cells. (2013)
  • Author(s): kabe S, Tauchi T, Tanaka Y, Kimura S, Maekawa T, Ohyashiki K.
  • Journal Title: Cancer Cell Int Volume: 13 Pages: 32
  • DOI: doi: 10.1186/1475-2867-13-32.
  • Peer Reviewed
• [Journal Article] Chimerism of bone marrow mesenchymal stem/stromal cells in allogeneic hematopoietic cell transplantation: is it clinically relevant? (2013)
  • Author(s): Miura Y, Yoshioka S, Yao H, Takaori-Kondo A, Maekawa T, Ichinohe T.
  • Journal Title: Chimerism Volume: 4 Pages: 78-83
  • DOI: doi: 10.4161/chim.25609
  • Peer Reviewed
• [Journal Article] Combination of ponatinib with Hedgehog antagonist vismodegib for therapy-resistant BCR-ABL1 positive leukemia. (2013)
  • Author(s): Katagiri S, Tauchi T, Okabe S, Minami Y, Kimura S, Maekawa T, Naoe T, Ohyashiki K.
  • Journal Title: Clin Cancer Res Volume: 19 Pages: 1422-1432
  • DOI: doi:10.1158/1078-0432.CCR-12-1777.
  • Peer Reviewed
• [Journal Article] Zoledronic acid significantly enhances radiation induced apoptosis against human fibrosarcoma cells by inhibiting radioadaptive signaling. (2013)
  • Author(s): Koto K, Murata H, Kimura K, 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] Loss of SMAD4 from colorectal cancer cells promotes CCL15 expression to recruit CCR1+ myeloid cells and facilitate liver metastasis. (2013)
  • Author(s): Itatani Y, Kawada K, Fujishita T, Kakizaki F, Hirai H, Matsumoto T, Iwamoto M, Inamoto S, Hatano E, Hasegawa S, Maekawa T, Uemoto S, Sakai Y, Taketo MM.
  • Journal Title: Gastroenterol Volume: 145 Pages: 1064-1075
  • DOI: doi: 10.1053/j.gastro.2013.07.033
  • Peer Reviewed
• [Journal Article] Myeloid-derived suppressor cells play crucial roles in the regulation of mouse collagen-induced arthritis. (2013)
  • Author(s): Fujii W, Ashihara E, Hirai H, Nagahara H, 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: 10.4049/jimmunol.1203535
  • Peer Reviewed
• [Journal Article] C/EBPβ promotes BCR-ABL-mediated myeloid expansion and leukemic stem cell exhaustion. (2013)
  • Author(s): 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: 10.1038/leu.2012.258
  • 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): 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: 0.1371/journal.pone.0054862
  • Peer Reviewed
• [Journal Article] Effectiveness of sulforaphane as a radiosensitizer for murine osteosarcoma cells. (2013)
  • Author(s): Sawai Y. Murata H. Horii M, Koto K, Matsui T, Horie N, Tsuji Y, Ashihar E, Maekawa T, Kubo T, Fushiki S.
  • Journal Title: Oncol Rep Volume: 29 Pages: 941-945
  • DOI: 10.3892/or.2012.2195
  • Peer Reviewed
• [Journal Article] Association of Takayasu arteritis with HLA-B*67:01 and two amino acids in HLA-B protein. (2013)
  • Author(s): Terao C, Yoshifuji H, Ohmura K, Murakami K, Kawabata D, Yurugi K, Tazaki J, Kinoshita H, Kimura A, Akizuki M, Kawaguchi Y, Yamanaka H, Miura Y, Maekawa T, Saji H, Mimori T.
  • Journal Title: Rheumatology Volume: 52 Pages: 1769-1774
  • DOI: 10.1093/rheumatology/ket241
  • Peer Reviewed
• [Journal Article] Incidence and pattern of hemolytic anemia after minor ABO-mismatched living-donor lobar lung transplantation. (2013)
  • Author(s): Ohsumi A, Chen F, Yurugi K, Maekawa T, Shoji T, Sato M, Aoyama A, Bando T, Date H.
  • Journal Title: Surg Today Volume: 43 Pages: 1250-1253
  • DOI: 10.1007/s00595-012-0422-3
  • Peer Reviewed
• [Presentation] C/EBPβ expressed by bone marrow mesenchymal stromal cells is indispensable for precursor B-cell lymphopoiesis. (2013)
  • Author(s): Yoshioka S, Miura Y, Yao H, Iwasa M, Sato A, Tamura A, Yokota A, Hishita T, Ichinohe T, Hirai H, Maekawa T.
  • Organizer: 55th American Society of Hematology
  • Place of Presentation: New Orleans, LA, USA
  • Year and Date: 20131207-20131210
  • Invited
• [Presentation] Cytokine-STATs signalings upregulate endogenous C/EBPβ in BCR-ABL+ leukemic cells independently from BCR-ABL signaling. (2013)
  • Author(s): Yokota A, Hirai H, Hayashi Y, Tamura A, Sato A, Yao H, Yoshioka S, Iwasa M, Ashihara E, Miura Y, Maekawa T.
  • Organizer: 55th American Society of Hematology
  • Place of Presentation: New Orleans, LA, USA
  • Year and Date: 20131207-20131210
  • Invited
• [Presentation] Cell-intrinsic and -extrinsic involvement of C/EBPβ in the regulation of hematopoietic stem cells. (2013)
  • Author(s): Tamura A, Hirai H, Hayashi Y, Yokota A, Sato A, Yao H, Yoshioka S, Iwasa M, Ashihara E, Miura Y, Maekawa T.
  • Organizer: 55th American Society of Hematology
  • Place of Presentation: New Orleans, LA, USA
  • Year and Date: 20131207-20131210
  • Invited
• [Presentation] Hypoxia-Adapted Myeloma Cells Possess Stem Cell Character. (2013)
  • Author(s): Ashihara E, Nakagawa Y, Yao H, Yokota A, Miura Y, Takata K, Kitamura Y, Hirai H , Maekawa T.
  • Organizer: 55th American Society of Hematology
  • Place of Presentation: New Orleans, LA, USA
  • Year and Date: 20131207-20131210
  • Invited
• [Presentation] Impact of donor-specific anti-HLA and anti-angiotensin-II type 1 receptor antibodies on graft fibrosis after immunosuppression withdrawal in pediatric liver transplantation. (2013)
  • Author(s): Ohe H, Uchida Y, Yoshizawa A, Hirao H, Taniguchi M, Maruya E, Maekawa T, Uemoto S, Terasaki PI.
  • Organizer: The 13th Congress of the Asian Society of Transplantation
  • Place of Presentation: Kyoto, Japan
  • Year and Date: 20130902-20130906
  • Invited
• [Presentation] Myeloid-derived suppressor cells have regulatory roles in mouse collagen-induced arthritis. (2013)
  • Author(s): 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.
  • Organizer: Annual European Congress of Rheumatology 2013.
  • Place of Presentation: Madrid, Spain
  • Year and Date: 20130612-20130615
  • Invited
• [Presentation] Parathyroid hormone enhances expansion through CDH11 in human bone marrow mesenchymal stromal/stem cells. (2013)
  • Author(s): Yao H, Miura Y, Yoshioka S, Hayashi Y, Yokota A, Tamura A, Ichinohe T, Hirai H and Maekawa T.
  • Organizer: AsiaCORD 2013, Scientific Symposium I-1 "New Insights into Stem Cell Biology- Stem Cell Development & Differentiation"
  • Place of Presentation: Kobe, Japan
  • Year and Date: 20130419-20130419
• [Presentation] C/EBPβ in mesenchymal stromal/stem cells regulates early B cell lymphopoiesis. (2013)
  • Author(s): Yoshioka S, Miura Y, Yao H, Hayashi Y, Tamura A, Hishita T, Ichinohe T, Hirai H, Takaori-Kondo A and Maekawa T.
  • Organizer: AsiaCORD 2013, Scientific Symposium I-1 "New Insights into Stem Cell Biology- Stem Cell Development & Differentiation"
  • Place of Presentation: Kobe, Japan
  • Year and Date: 20130419-20130419
  • Invited
• [Book] Emerging Trends in Cell and Gene Therapy. (2013)
  • Author(s): Ashihara E, Maekawa T
  • Total Pages: 705
  • Publisher: Springe, New York Heidelberg Dordrecht London
• [Remarks] 京都大学医学部附属病院輸血細胞治療部 研究年報・業績 2013年
  • URL: http://dtm.kuhp.kyoto-u.ac.jp/report