Evaluation of gene delivery mechanism based on quantitative visualization of cellular defense system against exogenous genes for high-efficiency gene therapy

• Project/Area Number: 24790036
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Physical pharmacy
• Research Institution: National Institute of Advanced Industrial Science and Technology (2014); Institute of Physical and Chemical Research (2013); Hokkaido University (2012)
• Principal Investigator: SASAKI Akira 独立行政法人産業技術総合研究所, バイオメディカル研究部門, 研究員 (30580162)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000); Fiscal Year 2014: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000); Fiscal Year 2013: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2012: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
• Keywords: 遺伝子デリバリー / 蛍光イメージング / 1分子計測 / 画像相関 / 酵素活性 / 画像相関分光法 / DNA分解酵素 / 蛍光相互相関分光法 / 生物物理 / DNA / １分子計測

Outline of Final Research Achievements

The Purpose of this study is to visualize intracellular diffusion and degradation of exogenous genes quantitatively. We have newly developed raster image cross-correlation spectroscopy (ccRICS) setup. Degradation activity of DNA in living cells was successfully monitored as a movie through a direct measurement by the developed ccRICS system. Our result shows that the DNA degradation activity in living cells differs between cell lines. The achievement reveals a defense machinery against exogenous DNA invasion and determine its activity. The work will contribute a development of nucleic acid medicine and gene therapy.

Research Products

• [Journal Article] Recombinant protein (EGFP-Protein G)-coated PbS quantum dots for in vitro and in vivo dual fluorescence (visible and second-NIR) imaging of breast tumors (2015)
  • Author(s): Sasaki A., Tsukasaki Y., Komatsuzaki A., Sakata T., Yasuda H. and Jin T.
  • Journal Title: Nanoscale Volume: 7 Issue: 12 Pages: 5115-5119
  • DOI: 10.1039/c4nr06480a
  • Peer Reviewed / Open Access
• [Journal Article] Fluorescence Imaging of Mitochondria in Living Cells Using a Novel Fluorene Derivative with a Large Two-Photon Absorption Cross-Section (2012)
  • Author(s): Tani S., Nakagawa K., Honda T., Saito H., Suzuki Y., Kawamata J., Uchida M., Sasaki A. and Kinjo M.
  • Journal Title: Curr. Pharm. Biotechnol. Volume: 13(14) Pages: 2649-2654
  • Peer Reviewed
• [Journal Article] Bovine serum albumin-coated quantum dots as a cytoplasmic viscosity probe in a single living cell (2012)
  • Author(s): Nakane Y.
  • Journal Title: Anal. Methods Volume: 4 Issue: 7 Pages: 1903-1905
  • DOI: 10.1039/c2ay25318f
  • Peer Reviewed
• [Journal Article] 蛍光相互相関分光法を駆使した生細胞内の分子状態解析 (2012)
  • Author(s): 佐々木章, 金城政孝
  • Journal Title: 生化学 Volume: 84(12) Pages: 1024-1027
  • NAID: 10031146000
  • Peer Reviewed
• [Presentation] ラスター画像相互相関分光法による生細胞内外来DNA分解活性の時空間的可視化 (2015)
  • Author(s): 佐々木章、山本条太郎、神隆、金城政孝
  • Organizer: 第67回日本細胞生物学会大会
  • Place of Presentation: タワーホール船堀
  • Year and Date: 2015-07-02
• [Presentation] Fluorescence correlation methods for detecting enzymatic activities in living cells (2015)
  • Author(s): Sasaki A.
  • Organizer: Indo-Japan Symposium on Advances in Biomedicine and Bioinformatics
  • Place of Presentation: IIT-Delhi, India
  • Year and Date: 2015-02-23
  • Invited
• [Presentation] 蛍光相互相関分光法による分子間相互作用解析 (2015)
  • Author(s): 佐々木章
  • Organizer: 第24回細胞生物学ワークショップ
  • Place of Presentation: 北海道大学
  • Year and Date: 2015-01-15
  • Invited
• [Presentation] 蛍光相互相関解析による細胞内外来DNA動態の可視化 (2012)
  • Author(s): 佐々木章
  • Organizer: 生化学会北海道支部、生物物理学会北海道支部、北海道分子生物学研究会共催 2012年度合同シンポジウム『生命現象の分子レベルでの解明』
  • Place of Presentation: 北海道大学（札幌市）
  • Invited
• [Presentation] Mapping of intracellular degradation of exogenous DNAs by using cross-correlation raster image correlation spectroscopy (ccRICS) (2012)
  • Author(s): Sasaki A. and Kinjo M.
  • Organizer: The 10th International Symposium for Future Drug Discovery and Medical Care
  • Place of Presentation: Hokkaido Univ. (Sapporo)
• [Presentation] Visualization of intracellular defense system against exogenous DNAs by using cross-correlation raster image correlation spectroscopy (ccRICS) (2012)
  • Author(s): 佐々木章, 金城政孝
  • Organizer: 日本生物物理学会第50回年会
  • Place of Presentation: 名古屋大学（名古屋市）