ｉＰＳ細胞由来の造血幹細胞を利用した再生不良性貧血自己抗原の同定

• Project/Area Number: 14F04419
• Research Category: Grant-in-Aid for JSPS Fellows
• Allocation Type: Single-year Grants
• Section: 外国
• Research Field: Hematology
• Research Institution: Kanazawa University
• Principal Investigator: 中尾 眞二 金沢大学, 医学系, 教授 (70217660)
• Co-Investigator(Kenkyū-buntansha): ESPINOZA CALDERO JORGE 金沢大学, 医学系, 外国人特別研究員 ; ESPINOZA CALDERON JORGE LUIS 金沢大学, 医学系, 外国人特別研究員 ; ESPINOZA CALDERON Jorge Luis 金沢大学, 医学系, 外国人特別研究員
• Project Period (FY): 2014-04-25 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥2,300,000 (Direct Cost: ¥2,300,000); Fiscal Year 2016: ¥600,000 (Direct Cost: ¥600,000); Fiscal Year 2015: ¥1,100,000 (Direct Cost: ¥1,100,000); Fiscal Year 2014: ¥600,000 (Direct Cost: ¥600,000)
• Keywords: 再生不良性貧血 / 細胞傷害性T細胞 / iPS細胞 / CD34陽性細胞 / 6pLOH陽性細胞 / T細胞レセプター / HLA-Bアレル / 6pLOH / 造血前駆細胞 / 自己抗原 / HLA-B*40:02 / 6pUPD / 造血幹細胞

Outline of Annual Research Achievements

平成27年度までの研究により、シクロスポリン（CsA）依存性の1例（KANA1）と、CsAで改善した1例（KANA6）から、6pLOH(+)、6pLOH(-)（野生型）、HLA-Bアレル（B4002またはB5401）単独欠失の3種類のiPS細胞クローンを樹立することに成功した。これらのiPS細胞を種々の造血因子の存在下で3週間培養することにより、約70%のCD34陽性細胞を含む造血前駆細胞集団が誘導された。KANA1由来の野生型CD34 陽性細胞で患者自身のCD8陽性T細胞を刺激することにより、野生型CD34陽性細胞のみを選択的に傷害する細胞傷害性T細胞（CTL）株が誘導された。このCTL株からCTLクローンの樹立を試みたが、増殖が不良で、維持することが困難であったため、CTL株のT細胞レセプター（TCR）レパトアを解析し、頻度の高いTCRβ鎖陽性T細胞からTCRα鎖β鎖のcDNAを作製した。このうち1種類のTCRモチーフは、初発時に保存されていた骨髄T細胞でもっとも頻度の高いCD8陽性PD-1陽性T細胞中のTCRモチーフと一致していた。このｃDNAを用いてJurkat細胞のトランスフェクタントを作製し、野生型CD34陽性細胞に対する特異性を現在検討している。
一方、KANA6については、未輸血・無治療状態の骨髄が保存されていたため、PD-1陽性CD137陽性T細胞分画から単一細胞ソーティングを行い、頻度の高い4種類のTCRモチーフcDNAを用いて、KANA1と同様にJurkat細胞にトランスフェクトした。作製された4種類のトランスフェクタントは、野生型CD34陽性細胞には反応したが、B5401欠失CD34陽性細胞には反応しなかった。このため、現在CD34陽性細胞からcDNAライブラリーを作製し、HLA-B*54:01をトランスフェクトしたCOS細胞への導入を準備している。

Research Progress Status

28年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

28年度が最終年度であるため、記入しない。

Research Products

• [Journal Article] Microbe-Induced Inflammatory Signals Triggering Acquired Bone Marrow Failure Syndromes. (2017)
  • Author(s): Espinoza JL, Kotecha R, Nakao S
  • Journal Title: Front Immunol Volume: 8 Pages: 186-186
  • DOI: 10.3389/fimmu.2017.00186
  • Peer Reviewed
• [Journal Article] The simultaneous inhibition of the mTOR and MAPK pathways with Gnetin-C induces apoptosis in acute myeloid leukemia. (2017)
  • Author(s): Espinoza JL, Elbadry MI, Taniwaki M, Harada K, Trung LQ, Nakagawa N, Takami A, Ishiyama K, Yamauchi T, Takenaka K, Nakao S
  • Journal Title: Cancer Lett Volume: 400 Pages: 127-136
  • DOI: 10.1016/j.canlet.2017.04.027
  • Peer Reviewed / Open Access
• [Journal Article] Induced pluripotent stem cell technology: A window for studying the pathogenesis of acquired aplastic anemia and possible applications. (2017)
  • Author(s): Elbadry MI, Espinoza JL, Nakao S
  • Journal Title: Exp Hematol Volume: 49 Pages: 9-18
  • DOI: 10.1016/j.exphem.2016.12.011
  • Peer Reviewed / Open Access
• [Journal Article] Epstein-Barr virus-associated leukemic lymphoma after allogeneic stem cell transplantation. (2016)
  • Author(s): Takamatsu H, Araki R, Nishimura R, Yachie A, Espinoza JL, Okumura H, Yoshida T, Kuzushima K, Nakao S
  • Journal Title: J Clin Virol Volume: 80 Pages: 82-86
  • DOI: 10.1016/j.jcv.2016.04.020
  • NAID: 120005771106
  • Peer Reviewed
• [Journal Article] A functional polymorphism in the NKG2D gene modulates NK-cell cytotoxicity and is associated with susceptibility to Human Papilloma Virus-related cancers. (2016)
  • Author(s): Espinoza JL, Nguyen VH, Ichimura H, Pham TT, Nguyen CH, Pham TV, Elbadry MI, Yoshioka K, Tanaka J, Trung LQ, Takami A, Nakao S
  • Journal Title: Sci Rep Volume: 6 Issue: 1 Pages: 39231-39231
  • DOI: 10.1038/srep39231
  • Peer Reviewed / Open Access
• [Journal Article] An altered gut microbiota may trigger autoimmune-mediated acquired bone marrow failure syndromes. (2016)
  • Author(s): Espinoza JL, Elbadry MI, Nakao S
  • Journal Title: Clin Immunol Volume: 171 Pages: 62-64
  • DOI: 10.1016/j.clim.2016.08.008
  • Peer Reviewed
• [Journal Article] Cyclosporine restores hematopoietic function by compensating for decreased Tregs in patients with pure red cell aplasia and acquired aplastic anemia. (2016)
  • Author(s): Dao AT, Yamazaki H, Takamatsu H, Sugimori C, Katagiri T, Maruyama H, Zaimoku Y, Maruyama K, Ly TQ, Espinoza L, Nakao S
  • Journal Title: Ann Hematol Volume: 95 Issue: 5 Pages: 771-781
  • DOI: 10.1007/s00277-016-2629-7
  • Peer Reviewed
• [Journal Article] Induction of HLA-B*40:02-restricted T cells possessing cytotoxic and suppressive functions against haematopoietic progenitor cells from a patient with severe aplastic anaemia. (2015)
  • Author(s): Inaguma Y, Akatsuka Y, Hosokawa K, Maruyama H, Okamoto A, Katagiri T, Shiraishi K, Murayama Y, Tsuzuki-Iba S, Mizutani Y, Nishii C, Yamamoto N, Demachi-Okamura A, Kuzushima K, Ogawa S, Emi N, Nakao S.
  • Journal Title: Br J Haematol. Volume: in press Issue: 1 Pages: 131-134
  • DOI: 10.1111/bjh.13464
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Somatic Mutations and Clonal Hematopoiesis in Aplastic Anemia. (2015)
  • Author(s): Yoshizato T, Dumitriu B, Hosokawa K, Makishima H, Yoshida K, Townsley D, Sato-Otsubo A, Sato Y, Liu D, Suzuki H, Wu CO, Shiraishi Y, Clemente MJ, Kataoka K, Shiozawa Y, Okuno Y, Chiba K, Tanaka H, Nagata Y, Katagiri T, Kon A, Sanada M, Scheinberg P, Miyano S, Maciejewski JP, Nakao S, Young NS, Ogawa S.
  • Journal Title: N Engl J Med. Volume: 373 Issue: 1 Pages: 35-47
  • DOI: 10.1056/nejmoa1414799
  • NAID: 120005623244
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Paroxysmal nocturnal hemoglobinuria induced by the occurrence of BCR-ABL in a PIGA mutant hematopoietic progenitor cell. (2015)
  • Author(s): Tominaga R, Katagiri T, Kataoka K, Wee RK, Maeda A, Gomyo H, Mizuno I, Murayama T, Ogawa S, Nakao S.
  • Journal Title: Leukemia. Volume: - Issue: 5 Pages: 1208-1210
  • DOI: 10.1038/leu.2015.268
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Somatic mutations and clonal hematopoiesis in aplastic anemia. (2015)
  • Author(s): Yoshizato T, Dumitriu B, Hosokawa K, Makishima H, Yoshida K, Townsley D, Sato-Otsubo A, Sato Y, Liu D, Suzuki H, Wu C, Shiraishi Y, Clemente MJ, Kataoka K, Shiozawa Y, Okuno Y, Chiba K, Tanaka H, Nagata Y, Katagiri T, Kon A, Sanada M, Scheinberg P, Miyano S, Maciejewski JP, Nakao S, Young NS, Ogawa S.
  • Journal Title: N Eng J Med Volume: -
  • NAID: 120005623244
  • Peer Reviewed
• [Journal Article] Induction of HLA-B*40:02-restricted T cells possessing cytotoxic and suppressive functions against hematopoietic progenitor cells from a patient with severe aplastic anemia. (2015)
  • Author(s): Inaguma Y, Akatsuka Y, Hosokawa K, Maruyama H, Okamoto A, Katagiri T, Shiraishi K, Murayama Y, Tsuzuki-Iba S, Mizutani Y, Nishii C, Yamamoto N, Demachi-Okamura A, Kuzushima K, Ogawa S, Emi N, Nakao S.
  • Journal Title: Br J Haematol Volume: -
  • Peer Reviewed
• [Journal Article] Imatinib for bronchiolitis obliterans after allogeneic hematopoietic stem cell transplantation. (2015)
  • Author(s): Watanabe S, Waseda Y, Kimura H, Takato H, Ohata K, Kondo Y, Kasahara K, Nakao S.
  • Journal Title: Bone Marrow Transplant Volume: -
  • Peer Reviewed
• [Journal Article] Persistent molecular remission of refractory acute myeloid leukemia with inv(16)(p13.1q22) in an elderly patient induced by cytarabine ocfosfate hydrate. (2015)
  • Author(s): Arahata M, Shimizu Y, Asakura H, Nakao S.
  • Journal Title: J Hematol Oncol Volume: - Issue: 1 Pages: 5-5
  • DOI: 10.1186/s13045-014-0100-6
  • Peer Reviewed / Open Access
• [Journal Article] Paraptosis cell death induction by the thiamine analog benfotiamine in leukemia cells. (2015)
  • Author(s): Sugimori N, Espinoza JL, Trung LQ, Takami A, Kondo Y, An DT, Sasaki M, Wakayama T, Nakao S.
  • Journal Title: PloS One Volume: - Issue: 4 Pages: e0120709-e0120709
  • DOI: 10.1371/journal.pone.0120709
  • Peer Reviewed / Open Access
• [Journal Article] Increased glycosylphosphatidylinositol-anchored protein-deficient granulocytes define a benign subset of bone marrow failures in patients with trisomy 8. (2014)
  • Author(s): Hosokawa K, Sugimori N, Katagiri T, Sasaki Y, Saito C, Seiki Y, Mochizuki K, Yamazaki H, Takami A, Nakao S.
  • Journal Title: Eur J Haemat Volume: 18 Issue: 3 Pages: 230-238
  • DOI: 10.1111/ejh.12484
  • NAID: 120005661927
  • Peer Reviewed / Open Access
• [Journal Article] Relative incidences and outcomes of Clostridium difficile infection following transplantation of unrelated cord blood, unrelated bone marrow, and related peripheral blood in adult patients: a single institute study. (2014)
  • Author(s): Hosokawa K, Takami A, Tsuji M, Araoka H, Ishiwata K, Takagi S, Yamamoto H, Asano-Mori Y, Matsuno N, Uchida N, Masuoka K, Wake A, Makino S, Yoneyama A, Nakao S, Taniguchi S.
  • Journal Title: Transp Infectious Dis Volume: 16 Issue: 3 Pages: 412-420
  • DOI: 10.1111/tid.12224
  • Peer Reviewed / Open Access
• [Journal Article] Successful hyperbaric oxygen therapy for refractory BK virus-associated hemorrhagic cystitis after cord blood transplantation. (2014)
  • Author(s): Hosokawa K, Yamazaki H, Nakamura T, Yoroidaka T, Imi T, Shima Y, Ohata K, Takamatsu H, Kotani T, Kondo Y, Takami A, Nakao S.
  • Journal Title: Transp Infectious Dis Volume: 16 Issue: 5 Pages: 843-846
  • DOI: 10.1111/tid.12266
  • Peer Reviewed / Open Access
• [Presentation] Generation of Ips Cell-Derived Hematopoietic Progenitor Cells from Patients with Acquired Aplastic Anemia Harboring Copy Number Neutral Loss of Heterozygosity of the Short Arm of Chromosome 6 (2015)
  • Author(s): Luis Espinoza, Yoshinori Yoshida, Kazuhisa Chonabayashi, Takamasa Katagiri,Yoshitaka Zaimoku, and Shinji Nakao.
  • Organizer: The American Society of Hematology 57th Annual Meeting
  • Place of Presentation: Orange County Convention Center
  • Year and Date: 2015-12-05
  • Int'l Joint Research