CRISPR-Cas9 based screening for essential tumor suppressors in liver carcinogenesis

• Project/Area Number: 17K19625
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Research Field: Tumor biology and related fields
• Research Institution: National Cancer Center Japan
• Principal Investigator: Yamamoto Yusuke 国立研究開発法人国立がん研究センター, 研究所, 主任研究員 (60768117)
• Co-Investigator(Kenkyū-buntansha): 松崎 潤太郎 国立研究開発法人国立がん研究センター, 研究所, 研究員 (60464864)
• Project Period (FY): 2017-06-30 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥6,500,000 (Direct Cost: ¥5,000,000、Indirect Cost: ¥1,500,000); Fiscal Year 2018: ¥2,600,000 (Direct Cost: ¥2,000,000、Indirect Cost: ¥600,000); Fiscal Year 2017: ¥3,900,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥900,000)
• Keywords: がん抑制遺伝子 / 上皮細胞 / CRISPR-Cas9 / 多段階発がん / CRISPR/Cas9 / 癌 / 発現制御

Outline of Final Research Achievements

A large number of research projects identified genetic mutations in the process of carcinogenesis, and there have been some useful findings of studies using cancer cell lines. With next generation sequencing, genetic variants in a variety of cancers have been identified and shown to have an impact on carcinogenesis; however, it is still unclear how the mutations essentially influenced the carcinogenesis and their metastatic potential. In this research project, we introduced and quantitatively analyzed the contribution of genetic mutations involved in the carcinogenic process by CRISPR-Cas9, in order to knockout the tumor suppressor genes in primary epithelial cells, and examined the effect of each gene on carcinogenesis. Also, we tried to recapitulate the multi-step carcinogenesis in vitro. This study enables us to accurately reflect and reproduce genomic aberrations in normal cells, and to achieve validation of the actual function of driver gene mutations.

Academic Significance and Societal Importance of the Research Achievements

CRISPR-Cas9システムのガイドRNAの作製の後、ガイドRNAの細胞への導入と遺伝子欠損株の単離に成功している。正常細胞の条件検討の段階として、がん細胞を用いた実験を行なっており実験系の確立を目指している。多数の遺伝子に対して同様の手法を適応することができ、研究の進行については加速度的に対応することが可能となる。その上で、遺伝子変異を導入した細胞の安定培養、動物モデルの作製についても準備を行なっていく。近年開発されている一塩基変異を誘導するCRISPR-Cas9遺伝子編集技術との融合、比較実験の検証により更なる精密医療へと繋がっていくと考えられる。

Research Products

• [Journal Article] Synthetic Lethality in Lung Cancer-From the Perspective of Cancer Genomics. (2019)
  • Author(s): Shimomura I, Yamamoto Y, Ochiya T.
  • Journal Title: Medicines (Basel) Volume: ;6(1) Issue: 1 Pages: 38-38
  • DOI: 10.3390/medicines6010038
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] ゲノム編集によるがん抑制因子のスクリーニング系の開発 (2018)
  • Author(s): 山本雄介
  • Journal Title: Precision Medicine Volume: 1 Pages: 102-106
  • Peer Reviewed
• [Journal Article] Extracellular microRNAs and oxidative stress in liver injury: a systematic mini review (2018)
  • Author(s): Matsuzaki J, Ochiya T
  • Journal Title: Journal of Clinical Biochemistry and Nutrition Volume: 63 Issue: 1 Pages: 6-11
  • DOI: 10.3164/jcbn.17-123
  • NAID: 130007387134
  • ISSN: 0912-0009, 1880-5086
  • Peer Reviewed / Open Access
• [Presentation] Serum microRNAs for the early-stage detection of hepatocellular carcinoma. (2017)
  • Author(s): Juntaro Matsuzaki, Shunsuke Kondo, Minoru Esaki, Takuji Okusaka, Kazuaki Shimada, Ken Kato, Yoshiaki Aoki, Makiko Ichikawa, Satoko Takizawa, Hiromi Sakamoto, Shumpei Niida, Fumitaka Takeshita, and Takahiro Ochiya
  • Organizer: Cold Spring Harbor Asia “Liver Biology, Diseases &Cancer”
  • Int'l Joint Research