Research for hepatocarcinogenesis using iPS cells with genetic mutations and information of HBV integration

• Project/Area Number: 19K17453
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 53010:Gastroenterology-related
• Research Institution: Tokyo Medical and Dental University
• Principal Investigator: Kawai-Kitahata Fukiko 東京医科歯科大学, 医学部附属病院, 特任助教 (00755580)
• Project Period (FY): 2019-04-01 – 2021-03-31
• Project Status: Completed (Fiscal Year 2020)
• Budget Amount: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000); Fiscal Year 2020: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000); Fiscal Year 2019: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
• Keywords: 肝細胞癌 / B型肝炎ウイルス / ヒトiPS細胞 / 肝細胞 / 遺伝子変異 / HBV integration / iPS細胞

Outline of Research at the Start

本研究では、肝細胞癌におけるNGSを用いた癌関連遺伝子変異、viral integrationの解析技術とヒトiPS細胞を用いたHBV感染・培養系の構築を発展的に融合させ、核酸アナログ内服によるHBV複製制御下の肝細胞癌においてHBV integrationが関わる肝発癌・進展機構を解明し、さらに遺伝子改変ヒトiPS細胞技術を応用し探索されたゲノム要因の機能解析、すなわちHBV integrationと癌関連遺伝子変異をゲノム編集により導入したiPS細胞の樹立とHBV integration過程を再現するin vitro HBV感染培養系の構築により臨床病態に則した発癌機構の詳細を解明する。

Outline of Final Research Achievements

We investigated cancer-related genes and viral integration using next generation sequencing in HCC tissues with HBV suppression by nucleotide/nucleoside analogues.
In HBs antigen positive patients with chronic persistent HBV infection, gene mutational profile and HBV integration breakpoints were similar to those in HBs antigen positive patients with normal ALT. In patients with prior HBV infection, HBV integration were found in common cancer-related genes although a total number of HBV integration breakpoints was small compared with those in chronic persistent HBV infection. Based on these results, we also established human induced pluripotent stem cell model with HBV integration breakpoints at TERT and KMT locus.

Academic Significance and Societal Importance of the Research Achievements

世界的に蔓延するB型肝炎ウイルス(HBV)感染症において、HBV制御下におけるB型肝癌の癌関連遺伝子やウイルスDNAのヒトゲノムへの挿入を調べ、さらにHBV breakpointsを再現してiPS細胞株への挿入変異を導入した。HBV制御状態により発癌過程が共通するまたは異なる可能性を示したことにより今後癌進展機構など臨床的な知見となり得る。またiPS細胞を用いた新たな応用研究分野を開発する知的・技術的基盤の確立に貢献することにつながる。

Research Products

• [Journal Article] Mac-2 binding protein glycosylation isomer as a novel predictive biomarker for patient survival after hepatitis C virus eradication by DAAs (2020)
  • Author(s): Nakagawa M, Nawa N, Takeichi E, Shimizu T, Tsuchiya J, Sato A, Miyoshi M, Kawai-Kitahata F, Murakawa M, Nitta S, Itsui Y, Azuma S, Kakinuma S, Fujiwara T, Watanabe M, Tanaka Y, Asahina Y; Ochanomizu Liver Conference Study Group.
  • Journal Title: Journal of Gastroenterology Volume: 55 Issue: 10 Pages: 990-999
  • DOI: 10.1007/s00535-020-01715-6
  • Peer Reviewed / Open Access
• [Journal Article] Time course alterations of virus sequences and immunoglobulin titers in a chronic hepatitis E patient (2020)
  • Author(s): Nitta Sayuri、Takahashi Kazuaki、Kawai‐Kitahata Fukiko、Tsuchiya Jun、Sato Ayako、Miyoshi Masato、Murakawa Miyako、Itsui Yasuhiro、Nakagawa Mina、Azuma Seishin、Kakinuma Sei、Watanabe Mamoru、Asahina Yasuhiro
  • Journal Title: Hepatology Research Volume: 50 Issue: 4 Pages: 524-531
  • DOI: 10.1111/hepr.13480
  • Peer Reviewed
• [Journal Article] Vasoactive Intestinal Peptide Derived From Liver Mesenchymal Cells Mediates Tight Junction Assembly in Mouse Intrahepatic Bile Ducts (2020)
  • Author(s): Sato A, Kakinuma S, Miyoshi M, Kamiya A, Tsunoda T, Kaneko S, Tsuchiya J, Shimizu T, Takeichi E, Nitta S, Kawai-Kitahata F, Murakawa M, Itsui Y, Nakagawa M, Azuma S, Koshikawa N, Seiki S, Nakauchi H, Asahina Y, Watanabe M.
  • Journal Title: Hepatology Communications Volume: 4 Issue: 2 Pages: 235-254
  • DOI: 10.1002/hep4.1459
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Comprehensive genetic analysis of cholangiolocellular carcinoma with a coexistent hepatocellular carcinoma‐like area and metachronous hepatocellular carcinoma (2019)
  • Author(s): Kawai-Kitahata F, Asahina Y, Kaneko S, Tsuchiya J, Sato A, Miyoshi M, Tsunoda T, Inoue-Shinomiya E, Murakawa M, Nitta S, Itsui Y, Nakagawa M, Azuma S, Kakinuma S, Tanabe M, Sugawara E, Takemoto A, Ojima H, Sakamoto M, Muraoka M, Takano S, Maekawa S, Enomoto N and Watanabe M.
  • Journal Title: Hepatology Research Volume: 49 Issue: 12 Pages: 1466-1474
  • DOI: 10.1111/hepr.13403
  • Peer Reviewed / Open Access
• [Journal Article] Loss of Fibrocystin Promotes Interleukin-8-Dependent Proliferation and CTGF Production of Biliary Epithelium. (2019)
  • Author(s): Tsunoda T, Kakinuma S, Miyoshi M, Kamiya A, Kaneko S, Sato A, Tsuchiya J, Nitta S, Kawai-Kitahata F, Murakawa M, Itsui Y, Nakagawa M, Azuma S, Sogo T, Komatsu H, Mukouchi R, Inui A, Fujisawa T, Nakauchi H, Asahina Y, Watanabe M.
  • Journal Title: Journal of Heptatology Volume: 印刷中 Issue: 1 Pages: 143-152
  • DOI: 10.1016/j.jhep.2019.02.024
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Impact of novel NS5A resistance-associated substitutions of hepatitis C virus detected in treatment-experienced patients (2019)
  • Author(s): Nitta S, Asahina Y, Kato T, Tsuchiya J, Inoue-Shinomiya E, Sato A, Tsunoda T, Miyoshi M, Kawai-Kitahata F, Murakawa M, Itsui Y, Nakagawa M, Azuma S, Kakinuma S, Hikita H, Takehara T, Watanabe M.
  • Journal Title: Scientific Reports Volume: 9 Issue: 1 Pages: 5722-5722
  • DOI: 10.1038/s41598-019-42114-z
  • Peer Reviewed / Open Access
• [Presentation] B型肝炎ウイルスの制御状態による肝細胞癌の 癌関連遺伝子変異とviral integrationの解析 (2020)
  • Author(s): 北畑富貴子、朝比奈靖浩、柿沼 晴
  • Organizer: 第56回日本肝臓学会総会
• [Presentation] Comprehensive analysis of cancer-related genes and aav/hepatitis b virus integration into genome on development of hepatocellular carcinoma in patients with prior hepatitis b virus infection (2020)
  • Author(s): Fukiko Kawai-Kitahata, Yasuhiro Asahina , Sei Kakinuma, Miyako Murakawa, Sayuri Nitta, Masato Miyoshi, Ayako Sato, Jun Tsuchiya, Taro Shimizu, Eiko Takeichi, Mina Nakagawa, Yasuhiro Itsui, Seishin Azuma, Shinji Tanaka , Minoru Tanabe, Shinya Maekawa, Nobuyuki Enomoto and Mamoru Watanabe
  • Organizer: EASL The Digital International Liver Congress 2020
  • Int'l Joint Research
• [Presentation] 背景肝の線維化からみた肝細胞癌における癌関連遺伝子変異とviral integrationの検討 (2019)
  • Author(s): 北畑富貴子、朝比奈靖浩、村川美也子、新田沙由梨、柿沼晴、角田知之 、三好正人、四宮恵美、佐藤綾子、土屋淳、井津井康浩、中川美奈、東正新、田中真二、田邉稔、前川伸哉、榎本信幸、渡辺守
  • Organizer: 第55回日本肝臓学会総会
• [Presentation] 細胆管細胞癌内の異なる表現型及び肝細胞癌における癌関連遺伝子の網羅的解析 (2019)
  • Author(s): 北畑富貴子、朝比奈靖浩、金子俊、村川美也子、新田沙由梨、柿沼晴、三好正人、佐藤綾子、井津井康浩、中川美奈、東正新、田邉稔、菅原江美子、竹本暁、坂元亨宇、尾島英知、村岡優、前川伸哉 、榎本信幸 、渡辺守
  • Organizer: 第55回日本肝癌研究会
• [Presentation] Comprehensive analysis of cancer gene mutations and viral integration in hepatocellular carcinoma arising from non-fibrotic liver (2019)
  • Author(s): Yasuhiro Asahina, Fukiko Kawai-Kitahata, Miyako Murakawa, Sayuri Nitta, Yasuhiro Itsui, Mina Nakagawa, Seishin Azuma, Sei Kakinuma
  • Organizer: The Liver Meeting 2019
  • Int'l Joint Research