5-Ethynyluridine: A Bio-orthogonal Uridine Variant for mRNA-Based Therapies and Vaccines

Sjors Maassen, Britt Coenen, Sara Dulk, Martijn van der Werff, Harry Warner, Fabio Spada, Thomas Frischmuth, Danny Incarnato, Geert van den Bogaart*

*Corresponding author for this work

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Abstract

The identification of pseudo- and N1-methylpseudo-uridine (Ψ and mΨ, respectively) as immunosilent uridine analogues has propelled the development of mRNA-based vaccines and therapeutics. Here, we have characterised another uridine analogue, 5-ethynyluridine (EU), which has an ethynyl moiety. We show that this uridine analogue does not cause immune activation in human macrophages, as it does not induce interleukin-6 secretion or expression of the inflammatory and antiviral genes MX1, PKR, and TAP2. Moreover, EU allows for prolonged expression, as shown with mRNA coding for yellow fluorescent protein (YFP). Side-by-side comparisons of EU with unmodified, Ψ, and mΨ revealed that EU-modified mRNA is expressed at lower levels, but confers similar stability and low immunogenicity to the other uridine analogues. Furthermore, structure analysis of modified mRNAs suggests that the observed phenotype is largely independent of RNA folding. Thus, EU is a potential candidate for RNA-based vaccines and therapeutics.

Original languageEnglish
Article number202200658
Number of pages7
JournalChemBioChem
Volume24
Early online date3-Jan-2023
DOIs
Publication statusPublished - 1-Mar-2023

Keywords

  • macrophage
  • mRNA therapy
  • uridine analogues
  • viral immunity

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