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Screening for small molecule competitors of the phosphorylated substrates of beta-TrCP and their application for the induction of targeted protein degradation

Publicly Offered Research

Project AreaNew frontier for ubiquitin biology driven by chemo-technologies
Project/Area Number 21H00295
Research Category

Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)

Allocation TypeSingle-year Grants
Review Section Complex systems
Research InstitutionInstitute of Physical and Chemical Research

Principal Investigator

渡邉 信元  国立研究開発法人理化学研究所, 環境資源科学研究センター, 嘱託職員 (90221689)

Project Period (FY) 2021-04-01 – 2023-03-31
Project Status Completed (Fiscal Year 2022)
Budget Amount *help
¥5,200,000 (Direct Cost: ¥4,000,000、Indirect Cost: ¥1,200,000)
Fiscal Year 2022: ¥2,600,000 (Direct Cost: ¥2,000,000、Indirect Cost: ¥600,000)
Fiscal Year 2021: ¥2,600,000 (Direct Cost: ¥2,000,000、Indirect Cost: ¥600,000)
Keywordsユビキチン化 / 小分子探索 / タンパク質分解 / タンパク質間相互作用 / タンパク質リン酸化 / Fボックスタンパク質 / 蛍光タンパク質 / ユビキチン
Outline of Research at the Start

β-TrCP(β-Transducin repeat Containing Protein)は、SCF型E3ユビキチン化酵素の基質認識サブユニットF-BOXタンパク質のひとつである。本研究計画では、β-TrCPの基質への結合に拮抗する小分子リガンドを見出し、標的タンパク質をプロテアソーム依存分解させるシステムを構築する研究を提案する。分解させたいタンパク質として、白血病の原因遺伝子産物Bcr-Abl、その特異的リガンドとしてGleevec (imatinib)を選び、β-TrCPリガンドとの結合キメラ化合物を合成し、ユビキチン化を利用した細胞内タンパク質分解誘導系の有効性を証明する。

Outline of Annual Research Achievements

これまでの研究で得られたβ-TrCP阻害物質について、その類縁体の解析を進めた。試験管内のリン酸化ペプチドへの結合、ユビキチン化活性に対する影響をみるふたつのアッセイによって絞り込んだ物質が細胞内で機能できる事を、β-TrCP複合体によるユビキチン化を介した細胞でのβカテニンの分解、IκBの分解への影響を解析することで行った。これらの解析によりさらに強い結合能を有する小分子を3種取得した。しかしながらその阻害活性は期待ほど強くはなくPROTAC化合物の合成に進めるには不十分であると判断した。化合物のβ-TrCPとの相互作用をin silico解析を行い、化合物とβ-TrCPの間の水素結合形成が阻害活性に重要であることを見出した。これらについての論文発表を行うことを進め投稿中である。
一方、その過剰発現ががん化およびがん細胞の増殖維持を誘導することが知られる c-Mycがん遺伝子に着目し、そのユビキチン化依存分解誘導系の構築も目指した。具体的にはすでに構築しているc-Myc活性を細胞内で解析できる系を用い、細胞内でc-Myc活性を阻害出来る物質探索を継続した。これまでの化合物ライブラリーおよび微生物二次代謝物の解析ですでにいくつかのヒット化合物を得ていたが、これらの中から、antimycin Aがc-Mycのユビキチン化依存分解を誘導することを見出した。また活性のある微生物二次代謝物からの活性物質精製も進めss49という物質もc-Mycのユビキチン化依存分解を誘導することを見出した。どちらの物質もミトコンドリアの酵素を阻害することで障害を与え活性酸素種(ROS)を発生させ、ROSがc-Mycをリン酸化する酵素であるGSK3を活性化することでc-Mycのユビキチン化依存分解を誘導することを見出した。これらについても論文発表を行うことを進め投稿中である。

Research Progress Status

令和4年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

令和4年度が最終年度であるため、記入しない。

Report

(2 results)
  • 2022 Annual Research Report
  • 2021 Annual Research Report
  • Research Products

    (27 results)

All 2022 2021 Other

All Int'l Joint Research (5 results) Journal Article (14 results) (of which Int'l Joint Research: 10 results,  Peer Reviewed: 13 results,  Open Access: 7 results) Presentation (6 results) (of which Int'l Joint Research: 2 results) Remarks (2 results)

  • [Int'l Joint Research] Salk Institute(米国)

    • Related Report
      2022 Annual Research Report
  • [Int'l Joint Research] Max Planck Institute(ドイツ)

    • Related Report
      2022 Annual Research Report
  • [Int'l Joint Research] Guangzhou Medical University(中国)

    • Related Report
      2022 Annual Research Report
  • [Int'l Joint Research] Guangzhou Medical Univerisity(中国)

    • Related Report
      2021 Annual Research Report
  • [Int'l Joint Research] Universiti Sains Malaysia(マレーシア)

    • Related Report
      2021 Annual Research Report
  • [Journal Article] ZMIZ2 promotes the development of triple-receptor negative breast cancer2022

    • Author(s)
      Zou Xiaopan、Liu Yan、Di Jun、Wei Wei、Watanabe Nobumoto、Li Jiang、Li Xiaomeng
    • Journal Title

      Cancer Cell International

      Volume: 22 Issue: 1 Pages: 52-52

    • DOI

      10.1186/s12935-021-02393-x

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Identification of a small molecule that enhances ferroptosis via inhibition of FSP1.2022

    • Author(s)
      Yoshioka H, Kawamura T, Muroi M, Kondoh Y, Honda K, Kawatani M, Aono H, Waldmann H, Watanabe N, Osada H.
    • Journal Title

      ACS Chemical Biology

      Volume: 17 Issue: 2 Pages: 483-491

    • DOI

      10.1021/acschembio.2c00028

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Apigenin, a Single Active Component of Herbal Extract, Alleviates Xerostomia via ERα-Mediated Upregulation of AQP5 Activation2022

    • Author(s)
      Wei Wei、Cao Tingting、Pathak Janak L.、Liu Xintong、Mao Tianjiao、Watanabe Nobumoto、Li Xiaomeng、Zhang Manli、Li Jiang
    • Journal Title

      Frontiers in Pharmacology

      Volume: 13 Pages: 818116-818116

    • DOI

      10.3389/fphar.2022.818116

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] New antimalarials identified by a cell-based phenotypic approach: Structure?activity relationships of 2,3,4,9-tetrahydro-1H-β-carboline derivatives possessing a 2-((coumarin-5-yl)oxy)alkanoyl moiety2022

    • Author(s)
      Cho Nobuo、Kikuzato Ko、Futamura Yushi、Shimizu Takeshi、Hayase Hiroki、Kamisaka Kikuko、Takaya Daisuke、Yuki Hitomi、Honma Teruki、Niikura Mamoru、Kobayashi Fumie、Watanabe Nobumoto、Osada Hiroyuki、Koyama Hiroo
    • Journal Title

      Bioorganic & Medicinal Chemistry

      Volume: 66 Pages: 116830-116830

    • DOI

      10.1016/j.bmc.2022.116830

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed
  • [Journal Article] Identifying Leptospira interrogans putative virulence factors with a yeast protein expression screen2022

    • Author(s)
      Lai Weng Yu、Wong Zhenpei、Chang Chiat Han、Samian Mohd Razip、Watanabe Nobumoto、Teh Aik-Hong、Noordin Rahmah、Ong Eugene Boon Beng
    • Journal Title

      Applied Microbiology and Biotechnology

      Volume: 106 Issue: 19-20 Pages: 6567-6581

    • DOI

      10.1007/s00253-022-12160-1

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Crystal structure of <i>Arabidopsis</i> DWARF14‐LIKE2 (DLK2) reveals a distinct substrate binding pocket architecture2022

    • Author(s)
      Burger Marco、Honda Kaori、Kondoh Yasumitsu、Hong Sharon、Watanabe Nobumoto、Osada Hiroyuki、Chory Joanne
    • Journal Title

      Plant Direct

      Volume: 6 Issue: 9

    • DOI

      10.1002/pld3.446

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Role of estrogen receptor signaling pathway-related genes in diffuse large B-cell lymphoma and identification of key targets via integrated bioinformatics analysis and experimental validation2022

    • Author(s)
      Chen Bo、Mao Tianjiao、Qin Xiuni、Zhang Wenqi、Watanabe Nobumoto、Li Jiang
    • Journal Title

      Frontiers in Oncology

      Volume: 12

    • DOI

      10.3389/fonc.2022.1029998

    • Related Report
      2022 Annual Research Report
  • [Journal Article] Lonidamine and domperidone inhibit expansion of transformed cell areas by modulating motility of surrounding nontransformed cells2022

    • Author(s)
      Aoyama Megumi、Ishikawa Kosuke、Nemoto Shuntaro、Hirano Hiroyuki、Watanabe Nobumoto、Osada Hiroyuki、Watanabe Shinya、Semba Kentaro
    • Journal Title

      Journal of Biological Chemistry

      Volume: 298 Issue: 12 Pages: 102635-102635

    • DOI

      10.1016/j.jbc.2022.102635

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] The Critical Biomarkers Identification of Insulin Signaling Involved in Initiating cAMP Signaling Mediated Salivary Secretion in Sjogren Syndrome: Transcriptome Sequencing in NOD Mice Model2022

    • Author(s)
      Chen Bo、Zhou Jiannan、Mao Tianjiao、Cao Tingting、Hu Shilin、Zhang Wenqi、Li Xueyang、Qin Xiuni、Liu Xintong、Watanabe Nobumoto、Li Jiang
    • Journal Title

      Biological Procedures Online

      Volume: 24 Issue: 1 Pages: 26-26

    • DOI

      10.1186/s12575-022-00189-5

    • Related Report
      2022 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Alleviatory effects of Danshen, Salvianolic acid A and Salvianolic acid B on PC12 neuronal cells and Drosophila melanogaster model of Alzheimer's disease2021

    • Author(s)
      Tan Florence Hui Ping、Ting Andrew Chung Jie、Leow Ban Guan、Najimudin Nazalan、Watanabe Nobumoto、Azzam Ghows
    • Journal Title

      Journal of Ethnopharmacology

      Volume: 279 Pages: 114389-114389

    • DOI

      10.1016/j.jep.2021.114389

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] Ethyl caffeate ameliorated amyloid‐beta42 protein‐associated toxicity in <scp>PC12</scp> cells and <i>Drosophila melanogaster</i>2021

    • Author(s)
      Tan Florence Hui Ping、Hadri Nurulhuda Athirah Binti、Najimudin Nazalan、Watanabe Nobumoto、Azzam Ghows
    • Journal Title

      Geriatrics Gerontology International

      Volume: 21 Issue: 12 Pages: 1125-1130

    • DOI

      10.1111/ggi.14296

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Int'l Joint Research
  • [Journal Article] A cesium tolerance-enhancing chemical promotes cesium tolerance in Arabidopsis thaliana by interacting with BETA-GLUCOSIDASE 23.2021

    • Author(s)
      Moon JY, Adams E, Miyazaki T, Kondoh Y, Muroi M, Watanabe N, Osada H, Shin R
    • Journal Title

      Scientific Reports

      Volume: 26 Issue: 1 Pages: 21109-21109

    • DOI

      10.1038/s41598-021-00564-4

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Open Access
  • [Journal Article] Identification of Tropical Plant Extracts That Extend Yeast Chronological Life Span2021

    • Author(s)
      Kwong Mandy Mun Yee、Lee Jee Whu、Samian Mohammed Razip、Wahab Habibah A.、Watanabe Nobumoto、Ong Eugene Boon Beng
    • Journal Title

      Cells

      Volume: 10 Issue: 10 Pages: 2718-2718

    • DOI

      10.3390/cells10102718

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Journal Article] Screening of selected ageing-related proteins that extend chronological life span in yeast Saccharomyces cerevisiae2021

    • Author(s)
      Lee Jee Whu、Ong Tee Gee、Samian Mohammed Razip、Teh Aik-Hong、Watanabe Nobumoto、Osada Hiroyuki、Ong Eugene Boon Beng
    • Journal Title

      Scientific Reports

      Volume: 11 Issue: 1 Pages: 24148-24148

    • DOI

      10.1038/s41598-021-03490-7

    • Related Report
      2021 Annual Research Report
    • Peer Reviewed / Open Access / Int'l Joint Research
  • [Presentation] c-Myc阻害小分子探索系の構築と探索2022

    • Author(s)
      渡辺信元、石川公輔、仙波憲太郎、長田裕之
    • Organizer
      がん分子標的治療学会学術集会
    • Related Report
      2022 Annual Research Report
  • [Presentation] ドーパミン拮抗薬ドンペリドンは需要対比依存的にがん細胞の拡大を抑制する2022

    • Author(s)
      青山愛、渡辺信元、長田裕之、仙波憲太郎
    • Organizer
      がん分子標的治療学会学術集会
    • Related Report
      2022 Annual Research Report
  • [Presentation] 正常―がん細胞間相互作用を制御する化合物のスクリーニングとその機能解析2022

    • Author(s)
      青山愛、平野弘之、渡辺信元、長田裕之、仙波憲太郎
    • Organizer
      第81回日本癌学会学術総会
    • Related Report
      2022 Annual Research Report
  • [Presentation] High-throughput screening system for ligands of β-TrCP and analyses of hit compounds2022

    • Author(s)
      Liu, X., Osada, H., Watanabe, N
    • Organizer
      第81回日本癌学会学術総会
    • Related Report
      2022 Annual Research Report
  • [Presentation] High-throughput screening system for ligands of β-TrCP and analyses of hit compounds.2022

    • Author(s)
      Liu, X., Sanada, E., Osada, H., Watanabe, N.
    • Organizer
      The International Symposium in Tokyo 2022, “Ubiquitin New Frontier from Neo-Biology to Targeted Protein Degradation”
    • Related Report
      2022 Annual Research Report
    • Int'l Joint Research
  • [Presentation] High-throughput screening identified antimycin A as an accelerator of ubiquitin-proteasome dependent degradation of c-Myc protein.2022

    • Author(s)
      Liu, Z., Sanada, E., Ishikawa, K., Semba, K., Osada, H., Watanabe, N.
    • Organizer
      The International Symposium in Tokyo 2022, “Ubiquitin New Frontier from Neo-Biology to Targeted Protein Degradation”
    • Related Report
      2022 Annual Research Report
    • Int'l Joint Research
  • [Remarks]

    • URL

      http://www.riken.jp/research/labs/csrs/riken_max_planck/bioprobe_app/

    • Related Report
      2021 Annual Research Report
  • [Remarks]

    • URL

      http://www.npd.riken.jp/crdu/jp/index.html

    • Related Report
      2021 Annual Research Report

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Published: 2021-04-28   Modified: 2023-12-25  

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