Stereo-random oligonucleotides enable efficient recruitment of ADAR in vitro and in vivo
Site-directed RNA editing is a promising and potentially safer alternative to genome editing. Previous methods have been developed that recruit the endogenously and ubiquitously expressed ADAR enzymes to initiate site-specific A-to-I edits, but often suffer from low efficacy or dependency on viral delivery. Chemically modified oligonucleotides may be a promising alternative, but the approach still lacks systematic in-depth studies. Furthermore, the best characterized platform uses stereo-pure backbone chemistry, which is not widely used, commercially unavailable and challenging to manufacture. Here, we report on single-stranded oligonucleotides of 30-60 nt length, which are fully chemically stabilized by applying commercially available, classical RNA drug modifications, like 2´-O-methyl, 2´-fluoro, and DNA on a stereo-random phosphate/phosphorothioate backbone. We demonstrate our so-called RESTORE 2.0 oligonucleotides to induce the correction of pathogenic point mutations, efficacy after GalNAc-mediated uptake into human primary hepatocytes, and proof of in-vivo efficacy in mice upon lipid nanoparticle-mediated delivery. The discovered design principles may increase the accessibility of site-directed RNA base editing to expand and support further research in this field.
Authors
- Laura S. Pfeiffer (ORCID: https://orcid.org/0000-0002-2487-4138)
- Ngadhnjim Latifi (ORCID: https://orcid.org/0000-0002-9792-3113)
- Daniel Tobias Hofacker (ORCID: https://orcid.org/0000-0001-5880-0690)
- Feijie Zhang
- Mark A. Kay (ORCID: https://orcid.org/0000-0002-2799-2615)
- Inga Jarmoskaite (ORCID: https://orcid.org/0000-0001-5847-5867)
- Jonathan M. Geisinger (ORCID: https://orcid.org/0000-0003-2764-102X)
- Lisa Groß
- Paul Vogel
- Marco Herrera‐Barrera (ORCID: https://orcid.org/0000-0001-7593-907X)
- Thorsten Stafforst (ORCID: https://orcid.org/0000-0001-9359-3439)
- Jin Billy Li (ORCID: https://orcid.org/0000-0003-0713-1399)
- Tobias Merkle
- Clemens Lochmann (ORCID: https://orcid.org/0009-0005-3869-8242)
- Stefanie Gackstatter
- Carolin Schlitz
- Davide Fumagalli
- Vanessa Deisling
Institutions
- California Institute for Regenerative Medicine (US)
- Research Institute for Work, Technology and Culture (DE)
- University of Tübingen (DE)
- Stanford University (US)
Publication Details
- Journal
- Nature Communications
- Published
- 2025-10-03
- DOI
- https://doi.org/10.1038/s41467-025-64434-7
- Citations
- 1
- Primary Topic
- RNA regulation and disease
- Type
- article
- Field-Weighted Citation Impact
- 0.81
Funders
- Deutsche Forschungsgemeinschaft
- Eberhard Karls Universität Tübingen
- Deutsche Akademie der Naturforscher Leopoldina - Nationale Akademie der Wissenschaften
- National Institutes of Health
- National Cancer Institute
- National Institute of General Medical Sciences
- National Institute of Allergy and Infectious Diseases