CRISPR/Cas9 genome-wide knockout screening for identifying the novel gene to overcome the tumor-immune escape

• Project/Area Number: 20K07695
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 50020:Tumor diagnostics and therapeutics-related
• Research Institution: Osaka University
• Principal Investigator: Morimoto Soyoko 大阪大学, 大学院医学系研究科, 寄附講座助教 (10649023)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Project Status: Completed (Fiscal Year 2022)
• Budget Amount: ¥4,420,000 (Direct Cost: ¥3,400,000、Indirect Cost: ¥1,020,000); Fiscal Year 2022: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2021: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2020: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
• Keywords: CRISPR-guide RNA library / Cas9 / CD8T cell / CD27 / memory / CRISPR-gRNA library / メモリー / 免疫逃避

Outline of Research at the Start

CRISPR-guide RNA libraryによる網羅的遺伝子ノックアウトと次世代シーケンサーを活用し、「癌の免疫逃避に耐性で品質が低下しない特性をT細胞に付与できる標的遺伝子」および「癌の免疫原性を高める標的遺伝子」を網羅的に探索・同定する。さらに、得られた成果を腫瘍局所で認められる「癌－免疫逃避相」を克服する革新的な癌免疫療法の開発に発展させることを目的として本研究課題を遂行する。

Outline of Final Research Achievements

Genome-wide knockout screening using CRISPR-guide RNA library/Cas9 system identified the novel gene X to maintain the expression of CD27 in CD8T cells under conditions that reduce CD27 expression. Repression of gene X expression promoted proliferation and remained cytokine production and cytotoxicity in addition to retaining CD27 expression in CD8T cells. To clarify the proliferation mechanism, now we analyze and integrate information of ATAC-seq and RNA-seq between control-CD8T cells and gene X-repressed CD8T cells.

Academic Significance and Societal Importance of the Research Achievements

免疫チェックポイント阻害療法やCAR-T細胞療法は、強力な抗腫瘍効果が期待できる癌免疫療法である。しかしながら、腫瘍攻撃の中核を担う細胞傷害性T細胞の「品質」の低下が原因で、治療を受けた患者のすべてに強い抗腫瘍効果が発揮されるわけではない。強い細胞傷害活性と高い増殖能は、T細胞に本来備わっている性質としては同時に成立しない。そこで、本研究成果で得られた「細胞傷害活性の維持と驚異的な増殖能を発揮する癌抗原特異的T細胞」は、現在の癌免疫療法が直面する最大の課題を解決することが大いに期待される。

Research Products

• [Journal Article] Cellular and Humoral Immune Responses Induced by an HLA Class I-restricted Peptide Cancer Vaccine Targeting WT1 Are Associated With Favorable Clinical Outcomes in Advanced Ovarian Cancer. (2022)
  • Author(s): Nishida S, Morimoto S, Oji Y, Morita S, Shirakata T, Enomoto T, Tsuboi A, Ueda Y, Yoshino K, Shouq A, Kanegae M, Ohno S, Fujiki F, Nakajima H, Nakae Y, Nakata J, Hosen N, Kumanogoh A, Oka Y, Kimura T, Sugiyama H.
  • Journal Title: J Immunother. Volume: 45 Issue: 1 Pages: 55-66
  • DOI: 10.1097/cji.0000000000000405
  • Peer Reviewed / Open Access
• [Journal Article] Identification of two distinct populations of WT1-specific cytotoxic T lymphocytes in co-vaccination of WT1 killer and helper peptides. (2021)
  • Author(s): Fujiki F, Tsuboi A, Morimoto S, Hashimoto N, Inatome M, Nakajima H, Nakata J, Nishida S, Hasegawa K, Hosen N, Oka Y, Oji Y, Sogo S, Sugiyama H.
  • Journal Title: Cancer Immunol Immunother. Volume: 70 Issue: 1 Pages: 253-263
  • DOI: 10.1007/s00262-020-02675-9
  • Peer Reviewed / Open Access
• [Journal Article] Reader-free ELISPOT assay for immuno-monitoring in peptide-based cancer vaccine immunotherapy (2020)
  • Author(s): Hayashi Sae、Imanishi Rin、Adachi Mayuko、Ikejima Sayaka、Nakata Jun、Morimoto Soyoko、Fujiki Fumihiro、Nishida Sumiyuki、Tsuboi Akihiro、Hosen Naoki、Nakajima Hiroko、Hasegawa Kana、Oka Yoshihiro、Sugiyama Haruo、Oji Yusuke
  • Journal Title: Biomedical Reports Volume: 12 Pages: 244-250
  • DOI: 10.3892/br.2020.1289
  • Peer Reviewed / Open Access