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In vivo cytidine base editing of hepatocytes without detectable off-target mutations in RNA and DNA


Villiger, Lukas; Rothgangl, Tanja; Witzigmann, Dominik; Oka, Rurika; Lin, Paulo J C; Qi, Weihong; Janjuha, Sharan; Berk, Christian; Ringnalda, Femke; Beattie, Mitchell B; Stoffel, Markus; Thöny, Beat; Hall, Jonathan; Rehrauer, Hubert; van Boxtel, Ruben; Tam, Ying K; Schwank, Gerald (2021). In vivo cytidine base editing of hepatocytes without detectable off-target mutations in RNA and DNA. Nature Biomedical Engineering, 5(2):179-189.

Abstract

Base editors are RNA-programmable deaminases that enable precise single-base conversions in genomic DNA. However, off-target activity is a concern in the potential use of base editors to treat genetic diseases. Here, we report unbiased analyses of transcriptome-wide and genome-wide off-target modifications effected by cytidine base editors in the liver of mice with phenylketonuria. The intravenous delivery of intein-split cytidine base editors by dual adeno-associated viruses led to the repair of the disease-causing mutation without generating off-target mutations in the RNA and DNA of the hepatocytes. Moreover, the transient expression of a cytidine base editor mRNA and a relevant single-guide RNA intravenously delivered by lipid nanoparticles led to ~21% on-target editing and to the reversal of the disease phenotype; there were also no detectable transcriptome-wide and genome-wide off-target edits. Our findings support the feasibility of therapeutic cytidine base editing to treat genetic liver diseases.

Abstract

Base editors are RNA-programmable deaminases that enable precise single-base conversions in genomic DNA. However, off-target activity is a concern in the potential use of base editors to treat genetic diseases. Here, we report unbiased analyses of transcriptome-wide and genome-wide off-target modifications effected by cytidine base editors in the liver of mice with phenylketonuria. The intravenous delivery of intein-split cytidine base editors by dual adeno-associated viruses led to the repair of the disease-causing mutation without generating off-target mutations in the RNA and DNA of the hepatocytes. Moreover, the transient expression of a cytidine base editor mRNA and a relevant single-guide RNA intravenously delivered by lipid nanoparticles led to ~21% on-target editing and to the reversal of the disease phenotype; there were also no detectable transcriptome-wide and genome-wide off-target edits. Our findings support the feasibility of therapeutic cytidine base editing to treat genetic liver diseases.

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Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Institute of Pharmacology and Toxicology
07 Faculty of Science > Institute of Pharmacology and Toxicology

04 Faculty of Medicine > Functional Genomics Center Zurich
04 Faculty of Medicine > University Children's Hospital Zurich > Medical Clinic
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Scopus Subject Areas:Life Sciences > Biotechnology
Physical Sciences > Bioengineering
Health Sciences > Medicine (miscellaneous)
Physical Sciences > Biomedical Engineering
Physical Sciences > Computer Science Applications
Language:English
Date:1 February 2021
Deposited On:19 Feb 2021 16:30
Last Modified:06 Feb 2022 07:45
Publisher:Springer
ISSN:2157-846X
OA Status:Green
Publisher DOI:https://doi.org/10.1038/s41551-020-00671-z
PubMed ID:33495639

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