Increased cytotoxicity of herpes simplex virus thymidine kinase expression in human induced pluripotent stem cells

Chizuru Iwasawa, Ryota Tamura, Yuki Sugiura, Sadafumi Suzuki, Naoko Kuzumaki, Minoru Narita, Makoto Suematsu, Masaya Nakamura, Kazunari Yoshida, Masahiro Toda, Hideyuki Okano, Hiroyuki Miyoshi

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Human induced pluripotent stem cells (iPSCs) hold enormous promise for regenerative medicine. The major safety concern is the tumorigenicity of transplanted cells derived from iPSCs. A potential solution would be to introduce a suicide gene into iPSCs as a safety switch. The herpes simplex virus type 1 thymidine kinase (HSV-TK) gene, in combination with ganciclovir, is the most widely used enzyme/prodrug suicide system from basic research to clinical applications. In the present study, we attempted to establish human iPSCs that stably expressed HSV-TK with either lentiviral vectors or CRISPR/Cas9-mediated genome editing. However, this task was difficult to achieve, because high-level and/or constitutive expression of HSV-TK resulted in the induction of cell death or silencing of HSV-TK expression. A nucleotide metabolism analysis suggested that excessive accumulation of thymidine triphosphate, caused by HSV-TK expression, resulted in an imbalance in the dNTP pools. This unbalanced state led to DNA synthesis inhibition and cell death in a process similar to a “thymidine block”, but more severe. We also demonstrated that the Tet-inducible system was a feasible solution for overcoming the cytotoxicity of HSV-TK expression. Our results provided a warning against using the HSV-TK gene in human iPSCs, particularly in clinical applications.

Original languageEnglish
Article number810
JournalInternational journal of molecular sciences
Volume20
Issue number4
DOIs
Publication statusPublished - 2019 Feb 2

Keywords

  • Cytotoxic
  • Genome editing
  • Herpes simplex virus type 1 thymidine kinase
  • Induced pluripotent stem cells
  • Lentiviral vector
  • Nucleotide metabolism

ASJC Scopus subject areas

  • Catalysis
  • Molecular Biology
  • Spectroscopy
  • Computer Science Applications
  • Physical and Theoretical Chemistry
  • Organic Chemistry
  • Inorganic Chemistry

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