Establishment of Gene Engineered Organoids for Functional Analysis of Human Colorectal Driver Gene Mutations

• Project/Area Number: 26293177
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Partial Multi-year Fund
• Section: 一般
• Research Field: Gastroenterology
• Research Institution: Keio University
• Principal Investigator: Sato Toshiro 慶應義塾大学, 医学部, 准教授 (70365245)
• Project Period (FY): 2014-04-01 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥16,510,000 (Direct Cost: ¥12,700,000、Indirect Cost: ¥3,810,000); Fiscal Year 2016: ¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000); Fiscal Year 2015: ¥5,330,000 (Direct Cost: ¥4,100,000、Indirect Cost: ¥1,230,000); Fiscal Year 2014: ¥5,980,000 (Direct Cost: ¥4,600,000、Indirect Cost: ¥1,380,000)
• Keywords: 大腸癌 / 浸潤 / 転移 / オルガノイド / 幹細胞 / 下部消化管学 / 大腸がん

Outline of Final Research Achievements

Recent advance in sequence technology has enabled comprehensive analyses of genetic mutations in human colorectal cancer. In this study, we combined organoid culture technology and CRISPR-Cas9 technology to establish gene engineered organoids for analysis of genetic mutations in human colorectal cancers. The engineered organoids were xenotransplanted into immune-deficient mice and pathologically examined for their invasive and metastatic capacity. The functional analysis platform for driver genetic mutations are instrumental and holds a promise for applying to drug screening. 

Research Products

• [Journal Article] Visualization and Targeting of LGR5+ Human Colon Cancer Stem Cells. (2017)
  • Author(s): Shimokawa M, Ohta Y, Nishikori S, Matano M, Takano A, Fujii M, Date S, Sugimoto S, Kanai T, Sato T.
  • Journal Title: Nature Volume: 印刷中
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Cell competition with normal epithelial cells promotes apical extrusion of transformed cells through metabolic?changes. (2017)
  • Author(s): Shunsuke Kon, et al.
  • Journal Title: Nat Cell Biol Volume: 印刷中 Pages: 530-41
  • DOI: 10.1038/ncb3509
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] IL-22BP dictates characteristics of Peyer's patch follicle-associated epithelium for antigen uptake (2017)
  • Author(s): Jinnohara T, Kanaya T, Hase K, Sakakibara S, Kato T, Tachibana N, Sasaki T, Hashimoto Y, Sato T, Watarai H, Kunisawa J, Shibata N, Williams I, Kiyono H, and Ohno H
  • Journal Title: J Exp Med Volume: 印刷中
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Expansion of adult human pancreatic tissue yields organoids harbouring progenitor cells with endocrine differentiation potential (2017)
  • Author(s): Loomans CJM, Giuliani NW, Balak J, Ringnalda F,van Gurp L, Huch M, Boj SF, Sato T, Kester L, de Sousa Lopes SMC, Roost MS, Bonner-Weir S, Engelse MA, Rabelink TJ, Heimberg H, Vries RGJ, van Oudenaarden A, Carlotti F, Clevers H, de Koning EJP
  • Journal Title: Science Translational Medicine Volume: 印刷中
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] A Single Species of Clostridium Subcluster XIVa Decreased in Ulcerative Colitis Patients (2016)
  • Author(s): Takeshita K, Mizuno S, Mikami Y, Sujino T, Saigusa K, Matsuoka K, Naganuma M, Sato T, Takada T, Tsuji H, Kushiro A, Nomoto K, Kanai T
  • Journal Title: Inflamm Bowel Dis Volume: 22 Pages: 2802-2810
  • DOI: 10.1097/mib.0000000000000972
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Tissue-specific mutation accumulation in human adult stem cells during life. (2016)
  • Author(s): Blokzijl F,de Ligt J,Jager M,Sasselli V,Roerink S,Sasaki N,Huch M,Boymans S,Kuijk E,Prins P, Nijman I,Martincorena I,Mokry M,Wiegerinck CL,Middendorp S,Sato T,Schwank G, Nieuwenhuis EES,Verstegen MMA,van der Laan LJW,de Jonge J,IJzermans JNM, Vries RG,van de Wetering M,Stratton MR,Clevers H,Cuppen E,van Boxtel R
  • Journal Title: nature Volume: 238 Pages: 260-264
  • DOI: 10.1038/nature19768
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Zinc transporter SLC39A7/ZIP7 promotes intestinal epithelial self-renewal by resolving ER stress. (2016)
  • Author(s): Ohashi, W., Kimura, S., Iwanaga, T., Furusawa, Y., Irie, T., Izumi, H., Watanabe, T., Hijikata, A., Hara, T., Ohara, O., Koseki, H., Sato, T., Robine, S., Mori, H., Hattori, Y., Watarai, H., Mishima, K., Ohno, H., Hase, K., Fukada, T.
  • Journal Title: PLoS Genet. Volume: 12
  • DOI: 10.1371/journal.pgen.1006349
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Mule Regulates the Intestinal Stem Cell Niche via the Wnt Pathway and Targets EphB3 for Proteasomal and Lysosomal Degradation (2016)
  • Author(s): Dominguez-Brauer C, Hao Z, Elia AJ, Fortin JM, Nechanitzky R, Brauer PM, Sheng Y, Mana MD, Chio II, Haight J, Pollett A, Cairns R, Tworzyanski L, Inoue S, Reardon C, Marques A, Silvester J, Cox MA, Wakeham A, Yilmaz OH, Sabatini DM, van Es JH, Clevers H, Sato T, Mak TW
  • Journal Title: Cell Stem Cell Volume: 19 Pages: 205-216
  • DOI: 10.1016/j.stem.2016.04.002
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] A Colorectal Tumor Organoid Library Demonstrates Progressive Loss of Niche Factor Requirements during Tumorigenesis (2016)
  • Author(s): Fujii M, Shimokawa M, Date S, Takano A, Matano M, Ohta Y, Nanki K, Kawasaki K, Nakazato Y, Uraoka T, Watanabe T, Kanai T, Sato T.
  • Journal Title: Cell stem cell Volume: 18 Pages: 827-38
  • DOI: 10.1016/j.stem.2016.04.003
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] SETD7 controls intestinal regeneration and tumorigenesis by regulating Wnt/beta-catenin and Hippo/YAP signaling. (2016)
  • Author(s): Oudhoff MJ, Braam MJS, Freeman SA, Wong D, Rattray DG, Want J, Antignano F, Snyder K, Refaeli I, Hughes MR, McNagny KM, Gold MR, Arrowsmith CH, Sato T, Rossi FMV, Tatlock JT, Owen D, Brown PJ, Zaph C
  • Journal Title: Developmental Cell Volume: 37 Pages: 47-57
  • DOI: 10.1016/j.devcel.2016.03.002
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Active and water-soluble form of lapidated Wnt protein is maintained by a serum glycoprotein afamin/ alpha-almumin. (2016)
  • Author(s): Mihara, E., Hirai, H., Yamamoto, H., Tamura-Kawakami, K., Matano, M., Kikuchi, A., Sato, T., and Takagi, J.
  • Journal Title: Elife. Volume: 5 Pages: 1-19
  • DOI: 10.7554/elife.11621
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] EHBP1L1 coordinates Rab8 and Bin1 to regulate apical-directed transport in polarized epithelial cells. (2016)
  • Author(s): Nakajo A, Yoshimura SI, Togawa H, Kunii M, Iwano T, Izumi A, Noguchi Y, Watanabe A, Goto A, Sato T, Harada A.
  • Journal Title: The Journal of Cell Biology. Volume: 3 Pages: 1474-1485
• [Journal Article] Effcient gene engineering of human intestinal organoid using electroporation. (2015)
  • Author(s): Fujii M, Matano M, Nanki K, Sato T*.
  • Journal Title: Nature protocol Volume: 3 Pages: 1474-1485
• [Journal Article] Combination Therapy with Infliximab and Thiopurine Compared to Infliximab Monotherapy in Maintaining Remission of Postoperative Crohn's Disease. (2015)
  • Author(s): Sakuraba A, Okamoto S, Matsuoka K, Sato T, Naganuma M, Hisamatsu T, Iwao Y, Ogata H, Kanai T, Hibi T
  • Journal Title: Digestion Volume: 91 Pages: 233-238
  • DOI: 10.1159/000375302
  • Peer Reviewed / Open Access
• [Journal Article] Ink4a/Arf-dependent Loss of Parietal Cells Induced by Oxidative Stress Promotes CD44-dependent Gastric Tumorigenesis. (2015)
  • Author(s): Seishima R, Wada T, Tsuchihashi K, Okazaki S, Yoshikawa M, Oshima H, Oshima M, Sato T, Hasegawa H, Kitagawa Y, Goldenring JR, Saya H, Nagano O.
  • Journal Title: Cancer prevention research Volume: 8:492-501 Pages: 492-501
  • DOI: 10.1158/1940-6207.capr-15-0025-t
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Modeling colorectal cancer using CRISPR-Cas9– mediated engineering of human intestinal organoids (2015)
  • Author(s): Matano M,Date S, Shimokawa M, Takano A, Fujii M, Ohta Y, Watanabe T, Kanai T, Sato T.
  • Journal Title: Nat Med Volume: 21(3) Pages: 256-262
  • DOI: 10.1038/nm.3802
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Classical Th1 cells obtain colitogenicity by co-existence of RORγt-expressing T cells in experimental colitis. (2014)
  • Author(s): Saigusa K, Hisamatsu T, Handa T, Sujino T, Mikami Y, Hayashi A, Mizuno S, Takeshita K, Sato T, Matsuoka K, Kanai T
  • Journal Title: Inflamm Bowel Dis. Volume: 20 Pages: 1820-1827
  • DOI: 10.1097/mib.0000000000000149
  • Peer Reviewed
• [Journal Article] Cross-talk between RORγt+ innate lymphoid cells and intestinal macrophages induces mucosal IL-22 production in Crohn's disease. (2014)
  • Author(s): 1.Mizuno S, Mikami Y, Kamada N, Handa T, Hayashi A, Sato T, Matsuoka K, Matano M, Ohta Y, Sugita A, Koganei K, Sahara R, Takazoe M, Hisamatsu T, Kanai T.
  • Journal Title: Inflamm Bowel Dis. Volume: 20(8) Pages: 1426-1434
  • DOI: 10.1097/mib.0000000000000105
  • Peer Reviewed
• [Presentation] ミニ組織培養技術の応用: がんと幹細胞ニッチ． (2016)
  • Author(s): 佐藤俊朗
  • Organizer: 東京大学医科学研究所学友会セミナー
  • Place of Presentation: 東京大学医科学研究所（東京都港区）
  • Year and Date: 2016-03-24
• [Presentation] Functional Dynamics of Human Colon Cancer Stem Cells. Cellular Dynamics in Tissues. (2016)
  • Author(s): Toshiro Sato
  • Organizer: National Institute for Basic Biology
  • Place of Presentation: Main Conference Room, Yamate-Campus（愛知県岡崎市）
  • Year and Date: 2016-01-21
• [Presentation] 加齢・炎症ストレスの及ぼす腸管上皮幹細胞の分子遺伝学的変化．1PS18 ステムセルエイジング：老化の謎は解明できるか？ (2016)
  • Author(s): 佐藤俊朗，南木康作，下川真理子
  • Organizer: 第39回日本分子生物学会年会
  • Place of Presentation: パシフィコ横浜(神奈川県横浜市）
• [Presentation] Visualization of Human Colon Cancer Stem Cells Using Organoid Technology. Princess Takamatsu Cancer Research Fund. (2016)
  • Author(s): Toshiro Sato
  • Organizer: Current Status and Perspective of Cancer Stem Cell Research
  • Place of Presentation: Tokyo Palace Hotel(東京都千代田区)
• [Presentation] Easy manipulation of the genome: CRISPR technology. Translational Basic Science Novel models for GI disease (2016)
  • Author(s): Toshiro Sato
  • Organizer: UEGW 2016
  • Place of Presentation: Ｖienna （Ａustria）
• [Presentation] Disease modeling of human colorectal cancer. EMBO (2016)
  • Author(s): Toshiro Sato
  • Organizer: EMBL Symposium
  • Place of Presentation: Heidelberg（Germany）
• [Presentation] Gut environments and human colorectal carcinogenesis. Symposium 17. The intestinal microflora and cancer (2016)
  • Author(s): Toshiro Sato, Shinya Sugimoto, Mami Matano
  • Organizer: The 75th Annual Meeting of the Japanese Cancer Association
  • Place of Presentation: PACIFICO Yokohama (Yokohama,Kanagawa)
• [Presentation] ヒト大腸がん発がん・がん幹細胞のゲノム編集を用いた機能解析 (2016)
  • Author(s): 佐藤俊朗．太田悠木，股野麻未，藤井正幸，下川真理子
  • Organizer: 第89回日本生化学会大会
  • Place of Presentation: 仙台国際センター（宮城県仙台市）
• [Presentation] Disease Modeling of Colorectal Cancer Using Orgnaoids,Concurrent IV Disease modeling II. (2016)
  • Author(s): Toshiro Sato
  • Organizer: ISSCR 2016
  • Place of Presentation: San Francisco（America)
• [Presentation] Disease modeling of colorectal cancer using organoids (2016)
  • Author(s): Toshiro Sato
  • Organizer: The 41st Naito Conference
  • Place of Presentation: CHATERAISE Gateaux Kingdom SAPPORO（Sapporo-shi, Hokkaido）
• [Presentation] 腸管上皮オルガノイド培養の確立とその応用．特別企画２ 日本から世界に向けて発信されたトップ研究 (2016)
  • Author(s): 佐藤俊朗
  • Organizer: 第102回日本消化器病学会大会
  • Place of Presentation: 京王プラザホテル（東京都新宿区）
• [Presentation] Intestinal Stem Cell Niche Signaling in Homeostatic and Diseased Epithelium. Session2. Gut homeostasis and IBD (2016)
  • Author(s): Toshiro Sato
  • Organizer: The 5th International Forum of the 102nd General Meeting of the Japanese Society of Gastroenterology
  • Place of Presentation: 京王プラザホテル（東京都新宿区）
• [Presentation] Dynamics of stem cell system in intestinal epithelium. (2015)
  • Author(s): Toshiro Sato
  • Organizer: Biochemistry and Molecular Biology 2015
  • Place of Presentation: Symposium 3S4 Cell and Time（兵庫県神戸市）
  • Year and Date: 2015-12-03
• [Presentation] Modeling Colorectal Cancer with Oragnoid Culture System. (2015)
  • Author(s): Toshiro Sato
  • Organizer: Till&McCulloch Meeting 2015
  • Place of Presentation: Till&McCulloch Meeting 2015 Toronto(Canada)
  • Year and Date: 2015-10-27
• [Presentation] Modeling of Colorectal Cancer in a dish using an organoid culture system. (2015)
  • Author(s): Toshiro Sato
  • Organizer: 112th International Titisee conference
  • Place of Presentation: Schwarzwaldhotel Titisee(Germany)
  • Year and Date: 2015-10-22
• [Presentation] ミニ組織培養技術の応用：がんと幹細胞ニッチ． (2015)
  • Author(s): 佐藤俊朗
  • Organizer: 日本食品免疫学会
  • Place of Presentation: 伊藤謝恩ホール（東京都文京区）
  • Year and Date: 2015-10-16
• [Presentation] ゲノム編集技術を用いた人工大腸がんの作成と発がんメカニズム研究． (2015)
  • Author(s): 佐藤俊朗
  • Organizer: JBICバイオ関連基盤技術研究会
  • Place of Presentation: TIME24ビル（東京都江東区）
  • Year and Date: 2015-09-29
• [Presentation] ミニ組織培養技術の応用：がんと幹細胞ニッチ.第39回阿蘇シンポジウム． (2015)
  • Author(s): 佐藤俊朗
  • Organizer: 第39回阿蘇シンポジウム
  • Place of Presentation: 阿蘇リゾートグランヴィリオホテル（熊本県阿蘇市）
  • Year and Date: 2015-07-31
• [Presentation] オルガノイド技術のがん・再生領域におけるインパクトと挑戦． (2015)
  • Author(s): 佐藤俊朗
  • Organizer: 第14回SKIP Seminar/第81回FRONTIERS in Medical Seience Seminar Series．
  • Place of Presentation: 慶應義塾大学信濃町キャンパス総合医科学研究棟（東京都新宿区）
  • Year and Date: 2015-07-30
• [Presentation] Intestinal Stem Cell Niche Signaling in Homeostatic and Diseased Epithelium. (2015)
  • Author(s): Toshiro Sato
  • Organizer: GI Researh Academy 2015.
  • Place of Presentation: Royal Park Hotel（東京都中央区）
  • Year and Date: 2015-06-12
• [Presentation] 大腸癌オルガノイド培養のインパクト． (2015)
  • Author(s): 佐藤俊朗
  • Organizer: 第19回GI Cell Biology研究会．
  • Place of Presentation: ホテルメトロポリタンエドモント（東京都千代田区）
  • Year and Date: 2015-06-11
• [Presentation] 3次元オルガノイド培養を用いた幹細胞・がん研究へのアプローチ． (2015)
  • Author(s): 佐藤俊朗
  • Organizer: 第14回バイオテクノロジー展（BIO tech 2015）
  • Place of Presentation: 東京ビックサイト（東京都江東区）
  • Year and Date: 2015-05-14
• [Presentation] Modelling of colorectal cancer using CRISPR/Cas9 mediated engineering of human intestinal organoids. (2015)
  • Author(s): Toshiro Sato
  • Organizer: American Association of Cancer Research (AACR) annual meeting 2015.
  • Place of Presentation: Pennsylvania Convention Center Philadelphia（America）
  • Year and Date: 2015-04-18
• [Presentation] 消化器幹細胞の培養技術確立と再生医療への応用． (2015)
  • Author(s): 佐藤俊朗
  • Organizer: 第29回日本医学会総会 2015 関西．
  • Place of Presentation: 京都国際会議場（京都府京都市）
  • Year and Date: 2015-04-12
• [Presentation] ミニ組織培養技術の大腸がん研究への応用 (2015)
  • Author(s): 佐藤俊朗
  • Organizer: 第１回TRC がん幹細胞シリーズセミナー
  • Place of Presentation: 愛媛大学
  • Year and Date: 2015-02-17
• [Presentation] ヒト腸管上皮感細胞培養の応用：ニッチシグナルと大腸がん. (2014)
  • Author(s): 佐藤俊朗
  • Organizer: 第1回六甲医学研究会
  • Place of Presentation: 六甲山ホテル
  • Year and Date: 2014-11-07
• [Presentation] The impact of intestinal Organoid culture system on colorectal cancer research. (2014)
  • Author(s): Toshiro Sato
  • Organizer: 第73回日本癌学会学術総会
  • Place of Presentation: パシフィコ横浜
  • Year and Date: 2014-09-26
• [Presentation] Establishment and Applications of Intestinal Stem Cell Culture system; Stem Cell Niche and Cancer. (2014)
  • Author(s): Toshiro Sato
  • Organizer: The 33rd Sapporo International Cancer Symposium
  • Place of Presentation: 札幌
  • Year and Date: 2014-06-27
• [Presentation] 細胞外環境による腸管上皮幹細胞制御とがん化機構 (2014)
  • Author(s): 佐藤俊朗, 太田悠木
  • Organizer: 第14回日本抗加齢医学会総会
  • Place of Presentation: 大阪国際会議場
  • Year and Date: 2014-06-08
• [Presentation] Establishment and Applications of Intestinal Stem Cell Culture system; Stem Cell Niche and Cancer. (2014)
  • Author(s): Toshiro Sato
  • Organizer: GI Researh Academy
  • Place of Presentation: 東京（ロイヤルパークホテル）
  • Year and Date: 2014-06-06
• [Presentation] The Self-renewing Mechanism of Intestinal Stem Cell: Niche and Cancer (2014)
  • Author(s): Toshiro Sato, Yuki Ohta, Ai Takano, Shoichi Date, Mamiko Matano, Mariko Shimokawa
  • Organizer: 47th Annual Meeting of the Japanese Society of Developmental Biologists and Asia Pacific Developmental Biology Network
  • Place of Presentation: WINC愛知（名古屋）
  • Year and Date: 2014-05-28
• [Presentation] Establishment and Applications of Intestinal Stem Cell Culture System. (2014)
  • Author(s): Toshiro Sato
  • Organizer: Annual International Workshop onMucosal Immunology and Vaccine for Young Investigators.
  • Place of Presentation: 東京大学
  • Year and Date: 2014-05-23
• [Presentation] 腸管幹細胞とそのニッチ (2014)
  • Author(s): 佐藤俊朗
  • Organizer: 京都大学再生医科学研究所公開シンポジウム
  • Place of Presentation: 京都大学
  • Year and Date: 2014-05-13
• [Presentation] 消化管上皮幹細胞培養の基盤と応用 (2014)
  • Author(s): 佐藤俊朗，Hans Clevers, 金井隆典
  • Organizer: 第100回日本消化器病学会総会
  • Place of Presentation: 東京国際フォーラム
  • Year and Date: 2014-04-25
• [Presentation] 消化器ミニ組織培養の開発とその再生医療応用 (2014)
  • Author(s): 佐藤俊朗
  • Organizer: 第51回日本臨床分子医学会学術集会
  • Place of Presentation: 東京国際フォーラム
  • Year and Date: 2014-04-11
• [Book] Mini-Gut Organoids: Reconstitution of the Stem Cell Niche. (2015)
  • Author(s): Date S, Sato T*.
  • Total Pages: 21
  • Publisher: Annual Review of Cell and Developmental Biology.
• [Book] Back to 2D Culture for Ground State of Intestinal Stem Cells. (2015)
  • Author(s): Shimokawa M, Sato T*.
  • Total Pages: 3
  • Publisher: Cell Stem Cell.
• [Book] SnapShot: Growing Organoids from Stem Cells. (2015)
  • Author(s): Sato T*, Clevers H*.
  • Total Pages: 2
  • Publisher: Cell
• [Book] Wntシグナルによる消化器上皮幹細胞制御 (2014)
  • Author(s): 佐藤俊朗
  • Publisher: 中外医学社
• [Book] ヒト消化器上皮培養によるオーダーメイド治療への可能性 (2014)
  • Author(s): 太田悠木，佐藤俊朗
  • Publisher: 先端医学社