ヒト人工多能性幹細胞由来移植細胞の製造管理のための in vitro造腫瘍性評価系の開発

抄録

  The goal of pharmaceutical sciences is to deliver effective and safe medicinal products to patients. To achieve this goal, we need to ensure the efficacy, safety and quality of the products. Currently, many attempts are made to utilize human induced pluripotent stem cells (hiPSCs) in regenerative medicine/cell therapy. There are significant obstacles, however, preventing the clinical use of hiPSC-derived products. One of the most obvious safety issues is the presence of residual undifferentiated cells that have tumorigenic potential. Therefore, the assessment and control of the tumorigenicity of hiPSC-derived products is essential in order to prevent tumor development by residual pluripotent stem cells after implantation. We recently examined three in vitro assay methods to detect undifferentiated cells: soft agar colony formation assay, flow cytometry assay and quantitative real-time polymerase chain reaction assay (qRT-PCR). Although the soft agar colony formation assay was unable to detect hiPSCs, the flow cytometry assay using anti-TRA-1-60 antibody detected 0.1% undifferentiated hiPSCs that were spiked in primary retinal pigment epithelial (RPE) cells. Moreover, qRT-PCR with a specific probe and primers was found to detect a trace amount of LIN28 mRNA, which is equivalent to that present in a mixture of a single hiPSC and 5.0×10⁴ RPE cells. Our findings provide highly sensitive and quantitative in vitro assays essential for facilitating safety profiling of hiPSC-derived RPE cells for their clinical use.