Metabolic engineering generates a transgene-free safety switch for cell therapy

Nat Biotechnol. 2020 Dec;38(12):1441-1450. doi: 10.1038/s41587-020-0580-6. Epub 2020 Jul 13.


Safeguard mechanisms can ameliorate the potential risks associated with cell therapies but currently rely on the introduction of transgenes. This limits their application owing to immunogenicity or transgene silencing. We aimed to create a control mechanism for human cells that is not mediated by a transgene. Using genome editing methods, we disrupt uridine monophosphate synthetase (UMPS) in the pyrimidine de novo synthesis pathway in cell lines, pluripotent cells and primary human T cells. We show that this makes proliferation dependent on external uridine and enables us to control cell growth by modulating the uridine supply, both in vitro and in vivo after transplantation in xenograft models. Additionally, disrupting this pathway creates resistance to 5-fluoroorotic acid, which enables positive selection of UMPS-knockout cells. We envision that this approach will add an additional level of safety to cell therapies and therefore enable the development of approaches with higher risks, especially those that are intended for limited treatment durations.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Cell Proliferation
  • Cell- and Tissue-Based Therapy*
  • Gene Editing
  • Gene Targeting
  • Genome, Human
  • Humans
  • K562 Cells
  • Male
  • Metabolic Engineering*
  • Mice
  • Multienzyme Complexes / genetics
  • Orotate Phosphoribosyltransferase / genetics
  • Orotic Acid / analogs & derivatives
  • Orotic Acid / pharmacology
  • Orotidine-5'-Phosphate Decarboxylase / genetics
  • Pluripotent Stem Cells / drug effects
  • Pluripotent Stem Cells / metabolism
  • Transgenes*
  • Uridine / biosynthesis


  • Multienzyme Complexes
  • Orotic Acid
  • uridine 5'-monophosphate synthase
  • 5-fluoroorotic acid
  • Orotate Phosphoribosyltransferase
  • Orotidine-5'-Phosphate Decarboxylase
  • Uridine