旭硝子財団助成研究成果報告2020

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32木村　剛25Tsuyoshi KIMURA吉井　達之26Tatsuyuki YOSHII細胞移動によるがん免疫抑制解除を目指した免疫細胞誘引・捕獲マテリアルの開発 （2018採択）Development of a material that lure and capture immune cells for cancellation of cancer immunosuppression through cell migration(Project 2018)人工オルガネラと化学プローブによる細胞内微量化学種の検出 （2018採択）Highly sensitive detection of intracellular chemicals using engineered artificial organelle and chemosensor(Project 2018)旭硝子財団　助成研究成果報告（2020）The biological olfactory system can detect and discriminate a variety of chemical compounds with remarka-ble precision. In the present study, we attempted to generate olfactory receptor-based biosensors by generat-ing olfactory sensory neurons expressing olfactory receptors in vitro. Based on gene expression profile of de-veloping olfactory sensory neurons, we narrowed down candidate transcription factors which might be important for differentiation of olfactory sensory neurons. We are currently trying to differentiate fibroblasts into olfactory sensory neurons through lentivirus-mediated direct lineage reprogramming.がん微小環境においては，免疫抑制性の制御性T細胞（Treg）が集積し，がん増殖的かつ免疫抑制的な状態にある．本研究では，がん免疫抑制状態を解除し腫瘍抑制するため，腫瘍選択的にTreg除去する免疫細胞誘引・捕獲マテリアルの開発を行った．Tregに対する特異的抗体である抗CD25抗体を非細胞接着性材料に固定化した抗Treg抗体固定化材料を調製した．様々な形状の材料への抗体固定化が可能であった．抗Treg抗体固定化材料を用いたTregの選択的捕獲がin vitroおよびin vivoで示され，また，捕獲したTregの機能も維持されていた．さらに，担癌モデル動物での腫瘍抑制試験では，約30%の腫瘍抑制が示された．以上より本マテリアルによる免疫抑制解除を介した腫瘍抑制の可能性が示された．For cancer microenvironment, immunosuppressive regulatory T cells (Tregs) accumulate and suppress the activation of tumor-antigen-specific T cells. In this study, we developed material for attracting and capturing Tregs in order to selectively removes Tregs from tumor and release the immunosuppressed state. An anti-Treg antibody-immobilized material was prepared by immobilizing an anti-CD25 antibody, which is a specific an-tibody against Treg, on a non-cell adhesive material. It was possible to immobilize the antibody on materials of various shapes. Selective capture of Tregs using the anti-Treg antibody-immobilized material was demon-strated in vitro and in vivo, and the function of the captured Tregs was maintained. Furthermore, a tumor sup-pression test in a tumor-bearing model animal showed about 30% tumor suppression. From these results, it is suggested that the anti-Treg antibody-immobilized material could release the tumor immunosuppressive state.本研究では，蛍光プローブを生きた細胞内の局所に濃縮する技術を創製し，それを用いた微量化学種の超高感度検出システムを構築することを目的とする．そのためにタグとなるタンパク質（DHFR）と結合して蛍光を発するプローブ分子の開発を行った．開発したプローブ分子はin vitroでの評価でDHFRに結合して蛍光を発することが明らかとなった．このプローブは細胞内に発現させたDHFRのライブイメージングも可能であった．また，細胞内にタンパク質の自己集合体（人工オルガネラ）を作製した．スクリーニングの結果，細胞内に相分離構造を形成するキメラタンパク質を見出した．The aim of this study is to create a highly sensitive fluorescent detection system of intracellular chemicals such as ROS and RNS by condensing fluorescent probes in the specific spot in cell. We designed and synthe-sized fluorescent probe which shows fluorescence upon binding to DHFR. The synthesized probe showed