DNAとRNAシークエンシングデータを用いた統合変異解析ツールの開発

• Project/Area Number: 21K12117
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 62010:Life, health and medical informatics-related
• Research Institution: National Cancer Center Japan
• Principal Investigator: 上野 敏秀 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 (40381446)
• Project Period (FY): 2021-04-01 – 2025-03-31
• Project Status: Granted (Fiscal Year 2022)
• Budget Amount: ¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000); Fiscal Year 2024: ¥650,000 (Direct Cost: ¥500,000、Indirect Cost: ¥150,000); Fiscal Year 2023: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2022: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2021: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
• Keywords: バイオインフォマティクス / 融合遺伝子検出 / 変異解析 / ゲノム解析 / DNA・RNAシークエンシング

Outline of Research at the Start

DNA由来サンプルから変異解析やコピー数解析、RNA由来サンプルから発現量解析や融合遺伝子検出などのそれぞれの単体の解析を行うだけでなく、変異の有無の精度向上や疾患に起こっている現象を統合的に捉える工夫をDNAとRNAのデータから取得し、両方の結果を横断的・統合的に解析するパイプラインの開発を行う。

Outline of Annual Research Achievements

DNAとRNAサンプルからのゲノム異常検出を行う統合的解析パイプラインの開発のうち、本年度はRNAサンプルから融合遺伝子検出プログラムの作成を行った。STAR-FusionやArribaなど融合遺伝子検出ツールが数多く公開されている現在、定型的な解析であれば困らないが、融合点に介在配列を含むような融合遺伝子を検出したい、とか、Internal Tandem Duplication (ITD)も検出したい（一部のツールは検出可能である）、など既存ツールでは対応できない解析もできるよう拡張を行った。リファレンス配列は、ゲノムではなく、前処理を行ったトランスクリプト配列を使用し、偽陽性や偽陰性を防ぐ工夫をしている。融合遺伝子検出は融合点が定まるものを対象とし、多くのサンプルで検出されるような候補についてはブラックリスト化して除去する機能も設計した。検出感度（検出リード数）は、既存のツールたちよりも良好であるが、計算時間は既存ツールよりも多めなため、高速化に向けて改善を行いたい。結果リストの中には、実際に検出されたリードを載せているので、配列ベースでの確認はUCSCのblatのページなどから手動で確認可能することができるが、それとは別に、検出した融合遺伝子候補のリファレンスをダイナミックに作成し、Integrative Genomics Viewer (IGV)を通して融合遺伝子配列にリードがマップしているキャプチャー画像を取得できる機能も開発した。検出状況を視覚的に確認することは、偽陽性排除や信頼度を上げることに役立つものである。

Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

DNAとRNAデータの統合的変異解析パイプラインの構築を目標としているうちのRNA部分の融合遺伝子検出について開発できたので、おおむね順調に進んでいると考えている。

Strategy for Future Research Activity

融合遺伝子検出プログラムの計算時間が既存ツールに比べると多くなっているので、高速化に取り組みたい。

Research Products

• [Journal Article] Transcriptomic Profiling of MSI-H/dMMR Gastrointestinal Tumors to Identify Determinants of Responsiveness to Anti?PD-1 Therapy (2022)
  • Author(s): Chida K、Kawazoe A、Suzuki T、Kawazu M、Ueno T、Takenouchi K、Nakamura Y、Kuboki Y、Kotani D、Kojima T、Bando H、Mishima S、Kuwata T、Sakamoto N、Watanabe J、Mano H、Ikeda M、Shitara K、Endo I、Nakatsura T、Yoshino T
  • Journal Title: Clinical Cancer Research Volume: 28 Issue: 10 Pages: 2110-2117
  • DOI: 10.1158/1078-0432.ccr-22-0041
  • Peer Reviewed
• [Journal Article] Tumor mutational burden measurement using comprehensive genomic profiling assay (2022)
  • Author(s): Kage Hidenori、Kohsaka Shinji、Tatsuno Kenji、Ueno Toshihide、Ikegami Masachika、Zokumasu Koichi、Shinozaki-Ushiku Aya、Nagai Sumimasa、Aburatani Hiroyuki、Mano Hiroyuki、Oda Katsutoshi
  • Journal Title: Japanese Journal of Clinical Oncology Volume: 52 Issue: 8 Pages: 925-929
  • DOI: 10.1093/jjco/hyac063
  • Peer Reviewed
• [Journal Article] Exploration of predictive biomarkers for postoperative recurrence of stage II/III colorectal cancer using genomic sequencing (2022)
  • Author(s): Kishigami Fumishi、Tanaka Yosuke、Yamamoto Yoko、Ueno Toshihide、Kojima Shinya、Sato Kazuhito、Inoue Satoshi、Sugaya Saori、Ishihara Soichiro、Mano Hiroyuki、Kawazu Masahito
  • Journal Title: Cancer Medicine Volume: 11 Issue: 18 Pages: 3457-3470
  • DOI: 10.1002/cam4.4710
  • Peer Reviewed / Open Access
• [Journal Article] High-Throughput Functional Evaluation of MAP2K1 Variants in Cancer (2022)
  • Author(s): Mizuno Sho、Ikegami Masachika、Koyama Takafumi、Sunami Kuniko、Ogata Dai、Kage Hidenori、Yanagaki Mitsuru、Ikeuchi Hiroshi、Ueno Toshihide、Tanikawa Michihiro、Oda Katsutoshi、Osuga Yutaka、Mano Hiroyuki、Kohsaka Shinji
  • Journal Title: Molecular Cancer Therapeutics Volume: 22 Issue: 2 Pages: 227-239
  • DOI: 10.1158/1535-7163.mct-22-0302
  • Peer Reviewed / Open Access
• [Journal Article] Mixed Response to Cancer Immunotherapy is Driven by Intratumor Heterogeneity and Differential Interlesion Immune Infiltration (2022)
  • Author(s): T. Morinaga, T. Inozume, Y. Togashi et al.
  • Journal Title: Cancer Research Communications Volume: 2 Issue: 7 Pages: 739-753
  • DOI: 10.1158/2767-9764.crc-22-0050
  • Peer Reviewed / Open Access
• [Journal Article] PD-1 blockade therapy promotes infiltration of tumor-attacking exhausted T cell clonotypes (2022)
  • Author(s): Nagasaki Joji、Inozume Takashi、Yosuke Togashi et al.
  • Journal Title: Cell Reports Volume: 38 Issue: 5 Pages: 110331-110331
  • DOI: 10.1016/j.celrep.2022.110331
  • NAID: 120007192058
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Lactic acid promotes PD-1 expression in regulatory T cells in highly glycolytic tumor microenvironments (2022)
  • Author(s): Kumagai S, Koyama S, Itahashi K, Tanegashima T, Lin YT, Togashi Y, Kamada T, Irie T, Okumura G, Kono H, Ito D, Fujii R, Watanabe S, Sai A, Aokage K, Suzuki J, Ishii G, Kuwata T, Sakamoto N, Kawazu M, Ueno T, Mori T, Yamazaki N, Tsuboi M, Yatabe Y, Kinoshita T, Doi T, Shitara K, Mano H, Nishikawa H
  • Journal Title: Cancer Cell Volume: 40 Issue: 2 Pages: 201-218
  • DOI: 10.1016/j.ccell.2022.01.001
  • Peer Reviewed / Open Access
• [Journal Article] HLA Class I Analysis Provides Insight Into the Genetic and Epigenetic Background of Immune Evasion in Colorectal Cancer With High Microsatellite Instability (2022)
  • Author(s): Kawazu M, Ueno T, Saeki K, Sax N, Togashi Y, Kanaseki T, Chida K, Kishigami F, Sato K, Kojima S, Tanegashima T, Matsubara D, Tane K, Tanaka Y, Iinuma H, Hashiguchi Y, Hazama S, Khor SS, Tokunaga K, Tsuboi M, Niki T, Eto M, Shitara K, Torigoe T, Ishihara S, Aburatani H, Haeno H, Nishikawa H, Mano H
  • Journal Title: Gastroenterology Volume: 162 Issue: 3 Pages: 799-812
  • DOI: 10.1053/j.gastro.2021.10.010
  • Peer Reviewed / Open Access
• [Journal Article] Multi-omic profiling of peritoneal metastases in gastric cancer identifies molecular subtypes and therapeutic vulnerabilities (2021)
  • Author(s): Tanaka Yosuke、Chiwaki Fumiko、Kojima Shinya、Kawazu Masahito、Komatsu Masayuki、Ueno Toshihide、Inoue Satoshi、Sekine Shigeki、Matsusaki Keisuke、Matsushita Hiromichi、Boku Narikazu、Kanai Yae、Yatabe Yasushi、Sasaki Hiroki、Mano Hiroyuki
  • Journal Title: Nature Cancer Volume: 2 Issue: 9 Pages: 962-977
  • DOI: 10.1038/s43018-021-00240-6
  • Peer Reviewed / Open Access
• [Journal Article] MicroSEC; Sequence error filtering pipeline for formalin-fixed and paraffin-embedded samples. (2021)
  • Author(s): Ikegami M, Kohsaka S, Hirose T, Ueno T, Inoue S, Konomata N, Yamauchi H, Mori T, Sekine S, Inamoto Y, Yatabe Y, Kobayashi H, Tanaka S, Mano H.
  • Journal Title: Commun Biol Volume: 4 Issue: 1 Pages: 1396-1396
  • DOI: 10.1038/s42003-021-02930-4
  • Peer Reviewed / Open Access
• [Journal Article] Comparative Study on the Efficacy and Exposure of Molecular Target Agents in Non?small Cell Lung Cancer PDX Models with Driver Genetic Alterations (2021)
  • Author(s): Jo Hitomi、Yagishita Shigehiro、Hayashi Yoshiharu、Ryu Shoraku、Suzuki Mikiko、Kohsaka Shinji、Ueno Toshihide、Matsumoto Yuji、Horinouchi Hidehito、Ohe Yuichiro、Watanabe Shun-ichi、Motoi Noriko、Yatabe Yasushi、Mano Hiroyuki、Takahashi Kazuhisa、Hamada Akinobu
  • Journal Title: Molecular Cancer Therapeutics Volume: 21 Issue: 2 Pages: 359-370
  • DOI: 10.1158/1535-7163.mct-21-0371
  • Peer Reviewed / Open Access
• [Journal Article] Transcript-targeted analysis reveals isoform alterations and double-hop fusions in breast cancer (2021)
  • Author(s): Namba Shinichi、Ueno Toshihide、Kojima Shinya、Kobayashi Kenya、Kawase Katsushige、Tanaka Yosuke、Inoue Satoshi、Kishigami Fumishi、Kawashima Shusuke、Maeda Noriko、Ogawa Tomoko、Hazama Shoichi、Togashi Yosuke、Ando Mizuo、Shiraishi Yuichi、Mano Hiroyuki、Kawazu Masahito
  • Journal Title: Communications Biology Volume: 4 Issue: 1 Pages: 1320-1320
  • DOI: 10.1038/s42003-021-02833-4
  • Peer Reviewed / Open Access
• [Journal Article] TIGIT/CD155 axis mediates resistance to immunotherapy in patients with melanoma with the inflamed tumor microenvironment (2021)
  • Author(s): Kawashima S, Inozume T, Kawazu M, Ueno T, Nagasaki J, Tanji E, Honobe A, Ohnuma T, Kawamura T, Umeda Y, Nakamura Y, Kawasaki T, Kiniwa Y, Yamasaki O, Fukushima S, Ikehara Y, Mano H, Suzuki Y, Nishikawa H, Matsue H, Togashi Y.
  • Journal Title: Journal for ImmunoTherapy of Cancer Volume: 9 Issue: 11 Pages: e003134-e003134
  • DOI: 10.1136/jitc-2021-003134
  • NAID: 120007175403
  • Peer Reviewed / Open Access
• [Journal Article] Highly sensitive fusion detection using plasma cell-free RNA in non-small-cell lung cancers. (2021)
  • Author(s): Hasegawa N, Kohsaka S, Kurokawa K, Shinno Y, Takeda Nakamura I, Ueno T, Kojima S, Kawazu M, Suehara Y, Ishijima M, Goto Y, Kojima Y, Yonemori K, Hayashi T, Saito T, Shukuya T, Takahashi F, Takahashi K, Mano H.
  • Journal Title: Cancer Sci Volume: 112 Issue: 10 Pages: 4393-4403
  • DOI: 10.1111/cas.15084
  • Peer Reviewed / Open Access
• [Journal Article] Comprehensive functional evaluation of variants of fibroblast growth factor receptor genes in cancer. (2021)
  • Author(s): Takeda Nakamura I, Kohsaka S, Ikegami M, Ueno T, Li K, Beyett T, Koyama T, Shimizu T, Yamamoto N, Takahashi F, Takahashi K, Eck M, Mano H.
  • Journal Title: NPJ Precision Oncology Volume: 16 Issue: 1 Pages: 66-66
  • DOI: 10.1038/s41698-021-00204-0
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] A Low Tumor Mutational Burden and <i>PTEN</i> Mutations Are Predictors of a Negative Response to PD-1 Blockade in MSI-H/dMMR Gastrointestinal Tumors (2021)
  • Author(s): Chida Keigo, Kawazoe Akihito, Kawazu Masahito et al.
  • Journal Title: Clinical Cancer Research Volume: 27 Issue: 13 Pages: 3714-3724
  • DOI: 10.1158/1078-0432.ccr-21-0401
  • NAID: 130008144997
  • Peer Reviewed / Open Access
• [Journal Article] Rapid reprogramming of tumour cells into cancer stem cells on double-network hydrogels (2021)
  • Author(s): Suzuka J, Tsuda M, Wang L, Kohsaka S, Kishida K, Semba S, Sugino H, Aburatani S, Frauenlob M, Kurokawa T, Kojima S, Ueno T, Ohmiya Y, Mano H, Yasuda K, Gong JP, Tanaka S
  • Journal Title: Nat Biomed Eng Volume: － Issue: 8 Pages: 914-925
  • DOI: 10.1038/s41551-021-00692-2
  • Peer Reviewed / Open Access
• [Journal Article] Development of an optimal protocol for molecular profiling of tumor cells in pleural effusions at single-cell level (2021)
  • Author(s): Takeda Nakamura I, Ikegami M, Hasegawa N, Hayashi T, Ueno T, Kawazu M, Yagishita S, Goto Y, Shinno Y, Kojima Y, Takamochi K, Takahashi F, Takahashi K, Mano H, Kohsaka S
  • Journal Title: Cancer Sci Volume: － Issue: 5 Pages: 2006-2019
  • DOI: 10.1111/cas.14821
  • Peer Reviewed / Open Access