[MEASURE] 4. <i>In vitro</i> detection of transformed cells - Preliminary studies of Digital soft agar colony formation assay -

• Kiyoko BANDO

  Sumitomo Dainippon Pharma Co., Ltd.

• Shinji KUSAKAWA

  National Institute of Health Sciences

• Junichiro SAITO

  Astellas Pharma Inc.

• Hideki ADACHI

  Sumitomo Dainippon Pharma Co., Ltd.

• Takafumi YOTSUMOTO

  Asubio Pharma Co. Ltd.

• Kentaro TORIUMI

  GE Healthcare

• Ken-ichi YAMAMOTO

  LSI Medience Corporation

• Ayano DOI

  Mitsubishi Tanabe Pharma Corporation

• Atsushi KITANAKA

  Sumika Chemical Analysis Service, Ltd

• Takeshi WATANABE

  Takeda Pharmaceutical Company Limited

• Maya MORITA

  Axcelead Drug Discovery Partners, Inc

• Yoji SATO

  National Institute of Health Sciences

抄録

[Introduction] The differentiated cells derived from pluripotent stem cells (PSCs) have potentials to transform to malignant cells, and thus, the establishment of validated assays to detect these cells in cell therapy products (CTPs) is of importance for minimizing the risk of PSCs-based tumorigenicity. The soft agar colony formation (SACF) assay can detect transformed cells which show anchorage-independent cell growth. However, its sensitivity is not sufficient for quality control assessment of CTPs. The Digital SACF assay using an image analyzer has high sensitivity, but has not been generally used yet. Therefore, it was selected as the assay for validation in MEASURE. [Assay protocol] The malignant cells are spiked into normal cells (mimic the final product including malignant cells, e.g, HeLa/hMSC suspension), and cell suspension is mixed with medium and agar, and then placed on the bottom agar layer. After about 4-5 weeks of culture, dual fluorescence stained colonies are detected using image analyzers. [Progress] Preliminary experiments were conducted to evaluate efficiency of single cell-derived colony formation from a HeLa cell in soft agar culture with or without hMSCs in order to identify potential variation that affects the study results. The colony formation rates in HeLa/hMSCs tended to vary as compare with those of HeLa only. Next, the preliminary study, which intends to detect a HeLa cell spiked to 10⁶ hMSCs, is being conducted. [Future plan] The main study at the multiple facilities will be planned to evaluate the detection sensitivity and its reproducibility.

収録刊行物

• 日本毒性学会学術年会

  日本毒性学会学術年会 45.1 (0), P-216-, 2018

  日本毒性学会

詳細情報

• CRID: 1390282763028536448
• NII論文ID: 130007432568
• DOI: 10.14869/toxpt.45.1.0_p-216
• 本文言語コード: ja
• データソース種別:
  • JaLC
  • CiNii Articles
• 抄録ライセンスフラグ: 使用不可