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Generation of binary aptamesr that works cooperatively in close proximity

Research Project

Project/Area Number 19K05545
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeMulti-year Fund
Section一般
Review Section Basic Section 34020:Analytical chemistry-related
Research InstitutionKumamoto University

Principal Investigator

Kitamura Yusuke  熊本大学, 大学院先端科学研究部(工), 助教 (80433019)

Co-Investigator(Kenkyū-buntansha) 勝田 陽介  熊本大学, 大学院先端科学研究部(工), 助教 (50632460)
Project Period (FY) 2019-04-01 – 2022-03-31
Project Status Completed (Fiscal Year 2021)
Budget Amount *help
¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000)
Fiscal Year 2021: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2020: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2019: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
Keywords核酸 / アプタマー / 分子協働性 / バイオセンサー / PD-L1 / がん免疫療法 / 核酸アプタマー / バイナリープローブ / 光化学反応
Outline of Research at the Start

標的タンパク上で両者が近接した場合にのみ、選択的に連結される仕組みを構築すれば、その後、連結体として共にプルダウンすることで第二のアプタマーを取得することが可能である。従来のアプタマー取得法では、単純に結合親和性が低い候補核酸を淘汰していく仕組みであるが、本手法では、互いの近接度(距離)という新しい概念の淘汰圧をかけ、合理的に近くに結合するアプタマーを探索する。よって、両アプタマーを同時併用する 際(バイナリー化する際)、既に互いに協同的に作用し得る間隔で結合しているため、刺激に応答した連結や協同的発シグナルなど様々な高機能化への誘導が容易に行える。

Outline of Final Research Achievements

PD-L1, a membrane protein that is highly expressed on the cell membrane of tumor cells, is focused as a target. At first, from the initial DNA libraries with a diversity of 1.0E + 12, only those that bind to PD-L1 were selected by the usual SELEX method (preliminary selection). During the amplification of selected libraries by PCR, an anthracene-modified library (Ant-Lib) was generated by using a forward primer with their 5' end modified with anthracene. Using these libraries, we selected an aptamer (secondary aptamer) that would closely bind to the previously reported anti-PD-L1 aptamer (primary aptamer) with its 3' end modified with anthracene through the photochemical dimerization of anthracenes at the termini of primary and secondary aptamers (Proximity SELEX).

Academic Significance and Societal Importance of the Research Achievements

近接位選択を10ラウンドを終えている。最終的に得らえたライブラリーをベクターに組み込み、大腸菌にトランスフェクションし、得られたコロニーから回収したベクターの配列をサンガーシーケンシング法により配列を解読する予定である。近接位に結合可能なセカンダリーアプタマーが取得できていた場合、他のアプタマーにおいても同様にセカンダリーアプタマーを取得することによってバイナリー化が可能であると考えられ、既存のアプタマーの機能や性能を自在に拡張、改変可能な汎用の手法となることが期待される。

Report

(3 results)
  • 2021 Final Research Report ( PDF )
  • 2020 Research-status Report
  • 2019 Research-status Report
  • Research Products

    (24 results)

All 2021 2020 2019

All Journal Article (12 results) (of which Int'l Joint Research: 1 results,  Peer Reviewed: 12 results) Presentation (12 results) (of which Int'l Joint Research: 3 results)

  • [Journal Article] Catalytic Amplification of Electrochemical Signal in Homogeneous Solution Using an Entropy-driven DNA Circuit2021

    • Author(s)
      Yusuke Kitamura, Keisuke Yoshimura, Ryo Kuramoto, Yousuke Katsuda, Toshihiro Ihara
    • Journal Title

      Analytical Sciences

      Volume: 37 Issue: 3 Pages: 533-537

    • DOI

      10.2116/analsci.20SCN04

    • NAID

      130007996267

    • ISSN
      0910-6340, 1348-2246
    • Year and Date
      2021-03-10
    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] Detection of cancer cells in whole blood using a dynamic deformable microfilter and a nucleic acid aptamer2021

    • Author(s)
      Soichiro Fukuyama, Seitaro Kumamoto, Seiya Nagano, Shoma Hitotsuya, Keiichiro Yasuda, Yusuke Kitamura, Masaaki Iwatsuki, Hideo Baba, Toshihiro Ihara, Yoshitaka Nakanishi, Yuta Nakashima
    • Journal Title

      Talanta

      Volume: 228 Pages: 122239-122239

    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] Electrochemical Molecular Beacon for Nucleic Acid Sensing in a Homogeneous Solution2020

    • Author(s)
      Yusuke KITAMURA, Kotaro MISHIO, Pelin ARSLAN, Boui IKEDA, Chiharu IMOTO, Yousuke KATSUDA, Toshihiro IHARA
    • Journal Title

      Analytical Sciences

      Volume: 36 Issue: 8 Pages: 959-964

    • DOI

      10.2116/analsci.19P456

    • NAID

      130007886432

    • ISSN
      0910-6340, 1348-2246
    • Year and Date
      2020-08-10
    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] Xylitol Separation from a Polyol Mixture Using Lanthanide Ion-loaded Resins2020

    • Author(s)
      Yusuke Kitamura, Rika Shobu, Hirotaka Matsuura, Akinori Jyo, Toshihiro Ihara
    • Journal Title

      Analytical Sciences

      Volume: 36 Issue: 6 Pages: 769-773

    • DOI

      10.2116/analsci.19N032

    • NAID

      130007853445

    • ISSN
      0910-6340, 1348-2246
    • Year and Date
      2020-06-10
    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] A RuO<sub>2</sub> Nanosheet as a Novel Quencher-free Platform for the Detection of Nucleic Acids in a Homogeneous Solution2020

    • Author(s)
      KITAMURA Yusuke、TANIGUCHI Takaaki、TSUTSUMI Miwako、NURDIWIJAYANTO Leanddas、MATSUO Tomoya、KATSUDA Yousuke、IHARA Toshihiro
    • Journal Title

      Analytical Sciences

      Volume: 36 Issue: 4 Pages: 397-400

    • DOI

      10.2116/analsci.20C004

    • NAID

      130007830180

    • ISSN
      0910-6340, 1348-2246
    • Year and Date
      2020-04-10
    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] Catalytic formation of luminescent lanthanide complexes using an entropy-driven DNA circuit2020

    • Author(s)
      Yusuke Kitamura, Yukina Azuma, Yousuke Katsuda, Toshihiro Ihara
    • Journal Title

      Chemical communications

      Volume: 56 Issue: 27 Pages: 3863-3863

    • DOI

      10.1039/d0cc00602e

    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] G-quadruplexes in mRNA: A key structure for biological function2020

    • Author(s)
      Kamura Takuto、Katsuda Yousuke、Kitamura Yusuke、Ihara Toshihiro
    • Journal Title

      Biochemical and Biophysical Research Communications

      Volume: 526 Issue: 1 Pages: 261-266

    • DOI

      10.1016/j.bbrc.2020.02.168

    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] Detection of prostate-specific antigen in semen using DNA aptamers: an application of nucleic acid aptamers in forensic body fluid identification2020

    • Author(s)
      Tetsuya Satoh, Seiya Kouroki, Yusuke Kitamura, Toshihiro Ihara, Kazutoshi Matsumura, Susumu Iwase
    • Journal Title

      Analytical methods

      Volume: 12 Issue: 21 Pages: 2703-2703

    • DOI

      10.1039/d0ay00371a

    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] A dynamically deformable microfilter for selective separation of specific substances in microfluidics2020

    • Author(s)
      Seitaro Kumamoto, Kenshiro Nakatake, Souichiro Fukuyama, Keiichiro Yasuda, Yusuke Kitamura, Masaaki Iwatsuki, Hideo Baba, Toshihiro Ihara, Yoshitaka Nakanishi, Yuta Nakashima
    • Journal Title

      Biomicrofluidics

      Volume: 14 Issue: 6 Pages: 064113-064113

    • DOI

      10.1063/5.0025927

    • Related Report
      2020 Research-status Report
    • Peer Reviewed
  • [Journal Article] A novel cholinesterase assay for the evaluation of neurotoxin poisoning based on the electron-transfer promotion effect of thiocholine on an Au electrode2019

    • Author(s)
      Hiroshi Shimada, Yuki Kiyozumi, Yuki Koga, Yoko Ogata, Yousuke Katsuda, Yusuke Kitamura, Masaaki Iwatsuki, Katsuhiko Nishiyama, Hideo Baba, and Toshihiro Ihara
    • Journal Title

      Sens. Actuators B Chem.,

      Volume: 298 Pages: 126893-126893

    • DOI

      10.1016/j.snb.2019.126893

    • Related Report
      2019 Research-status Report
    • Peer Reviewed
  • [Journal Article] Catalytic Formation of Luminescent Complex Clusters Based on Autonomous Strand Exchange Reaction of DNA2019

    • Author(s)
      Yusuke Kitamura,* Akihiro Nozaki, Rie Ozaki, Yousuke Katsuda, and Toshihiro Ihara*
    • Journal Title

      ACS Appl. Bio Mater.

      Volume: 2 Issue: 35 Pages: 2988-2993

    • DOI

      10.1002/anie.200902000

    • Related Report
      2019 Research-status Report
    • Peer Reviewed
  • [Journal Article] Generalized Preparation of Two-Dimensional Quasi-nanosheets via Self-assembly of Nanoparticles2019

    • Author(s)
      Ren Cai, Dan Yang, Keng-Te Lin, Yifan Lyu, Bowen Zhu, Zhen He, Lili Zhang, Yusuke Kitamura, Liping Qiu, Xigao Chen, Yuliang Zhao, Zhuo Chen and Weihong Tan
    • Journal Title

      J. Am. Chem. Soc.

      Volume: 141 Issue: 4 Pages: 1725-1734

    • DOI

      10.1021/jacs.8b12415

    • Related Report
      2019 Research-status Report
    • Peer Reviewed / Int'l Joint Research
  • [Presentation] アントラセン光二量化反応を利用した鎖乗り換え型三本鎖核酸の形成2020

    • Author(s)
      小嶋 美鈴, 西村 健人, 北村 裕介, 勝田 陽介, 井原 敏博
    • Organizer
      第14回バイオ関連化学シンポジウム2020
    • Related Report
      2020 Research-status Report
  • [Presentation] 核酸の連鎖的鎖交換反応を利用したシグナル増幅型腫瘍細胞検出法の開発2020

    • Author(s)
      北村 裕介, 工藤 悠暉, 勝田 陽介, 中島 雄太, 安田 敬一郎, 熊本 清太郎, 岩槻 政晃, 馬場 秀夫, 中西 義孝, 井原 敏博
    • Organizer
      日本分析化学会第69年会
    • Related Report
      2020 Research-status Report
  • [Presentation] ルテニウム-白金複合錯体を鋳型特異的に放出するDNAプローブの合成とその遺伝子解析への応用2020

    • Author(s)
      小嶋 美鈴, 北村 裕介, 勝田 陽介, 井原 敏博
    • Organizer
      日本分析化学会第69年会
    • Related Report
      2020 Research-status Report
  • [Presentation] 電気化学モレキュラービーコンの開発とその均一溶液中における核酸検出への応用2020

    • Author(s)
      北村裕介、見汐航太郎、池田朋生、勝田陽介、井原敏博
    • Organizer
      日本化学会第100春季年会
    • Related Report
      2019 Research-status Report
  • [Presentation] Fabrication of anti-EpCAM aptamer-modified gold substrate for capturing cancer cells2019

    • Author(s)
      Ayase Tashima, Yuki Kudo, Yusuke Kitamura, Yuta Nakashima, Keiichiro Yasuda, Masaaki Iwatsuki, Yousuke Katsuda, Kenshiro Nakatake, Hideo Baba, Yoshitaka Nakanishi, Toshihiro Ihara
    • Organizer
      The 14th International Student Conference on Advanced Science and Technology (ICAST) Kumamoto
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research
  • [Presentation] Amplified detection of cancer cells by DNA circuit2019

    • Author(s)
      Yusuke Kitamura, Yuki Kudo, Ayase Tashima, Yuta Nakashima, Keiichiro Yasuda, Masaaki Iwatsuki, Yousuke Katsuda, Hideo Baba, Yoshitaka Nakanishi, Toshihiro Ihara
    • Organizer
      The 46th International Symposium on Nucleic Acids Chemistry
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research
  • [Presentation] DNAサーキットを利用した発光性希土類金属錯体クラスターの触媒的生成2019

    • Author(s)
      北村 裕介、野崎 晃広、尾崎 理衣、勝田 陽介、井原 敏博
    • Organizer
      日本分析化学会 第68年会
    • Related Report
      2019 Research-status Report
  • [Presentation] 抗EpCAMアプタマー修飾金基板上へのがん細胞の捕捉2019

    • Author(s)
      田島 彩瀬、工藤 悠暉、北村 裕介、中島 雄太、安田 敬一郎、岩槻 政晃、勝田 陽介、馬場 秀夫、中西 義孝、井原 敏博
    • Organizer
      第13回バイオ関連化学シンポジウム2019
    • Related Report
      2019 Research-status Report
  • [Presentation] クロスオーバーSELEX による抗 CD24DNA アプタマーの取得とその機能評価2019

    • Author(s)
      北村 裕介、後藤 広志、林田 泰起、勝田 陽介、井原 敏博
    • Organizer
      第13回バイオ関連化学シンポジウム2019
    • Related Report
      2019 Research-status Report
  • [Presentation] DNAサーキットによるシグナル増幅を利用したがん細胞の 検出に関する基礎的検討2019

    • Author(s)
      工藤 悠暉, 北村 裕介、田島 彩瀬、勝田 陽介、安田 敬一郎、中島 雄太、岩槻 政晃、馬場 秀夫、中西 義孝、井原 敏博
    • Organizer
      第37回九州分析化学若手の会 夏季セミナー
    • Related Report
      2019 Research-status Report
  • [Presentation] DNAサーキットによるシグナル増幅を利用したがん細胞の検出2019

    • Author(s)
      工藤 悠暉, 北村 裕介、田島 彩瀬、勝田 陽介、安田 敬一郎、中島 雄太、岩槻 政晃、馬場 秀夫、中西 義孝、井原 敏博
    • Organizer
      第56回化学関連支部合同九州大会
    • Related Report
      2019 Research-status Report
  • [Presentation] Fabrication of EpCAM aptamer-modified gold substrate and specific capture of cancer cells on it2019

    • Author(s)
      Ayase Tashima, Yusuke Kitamura, Yuki, Kudo, Yuta Nakashima, Keiichiro Yasuda, Masaaki Iwatsuki, Yousuke Katsuda, Hideo Baba, Yoshitaka Nakanishi, Toshihiro Ihara
    • Organizer
      第24回日本化学会九州支部 韓国化学会釜山支部合同セミナー
    • Related Report
      2019 Research-status Report
    • Int'l Joint Research

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Published: 2019-04-18   Modified: 2025-03-27  

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