Adenine base editing in an adult mouse model of tyrosinaemia

Abstract

In contrast to traditional CRISPR–Cas9 homology-directed repair, base editing can correct point mutations without supplying a DNA-repair template. Here we show in a mouse model of tyrosinaemia that hydrodynamic tail-vein injection of plasmid DNA encoding the adenine base editor (ABE) and a single-guide RNA (sgRNA) can correct an A>G splice-site mutation. ABE treatment partially restored splicing, generated fumarylacetoacetate hydrolase (FAH)-positive hepatocytes in the liver, and rescued weight loss in mice. We also generated FAH+ hepatocytes in the liver via lipid-nanoparticle-mediated delivery of a chemically modified sgRNA and an mRNA of a codon-optimized base editor that displayed higher base-editing efficiency than the standard ABEs. Our findings suggest that adenine base editing can be used for the correction of genetic diseases in adult animals.

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Fig. 1: Adenine base editing rescues liver disease phenotype in a mouse model of tyrosinaemia.
Fig. 2: Adenine base editing partially corrects the Fah mutation in mouse liver.
Fig. 3: Optimizing the coding sequence of ABE6.3 and adding an N-terminal nuclear localization sequence improves base editing.
Fig. 4: RA6.3 shows a higher editing efficiency compared to ABE6.3 and Cas9-mediated HDR at two genomic sites in HEK293T cells.
Fig. 5: RA6.3 increases editing efficiency in vivo compared to ABE6.3.

Data availability

The authors declare that all data supporting the findings of this study are available within the paper and its Supplementary Information. The deep-sequencing data are available from the Sequence Read Archive (SRA) under accession number PRJNA513076 and PRJNA513291.

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Acknowledgements

We thank C. Mello, P. Zamore, S. Wolfe and E. Sontheimer for discussions; J. Smith for editing the manuscript; M. Grompe (Oregon Health and Science University) for providing the Fah mice; Y. Liu and E. Kittler of the UMass Morphology and Deep Sequencing Cores for support. W.X. was supported by grants from the National Institutes of Health ((NIH) DP2HL137167, P01HL131471 and UG3HL147367), American Cancer Society (129056-RSG-16-093), the Lung Cancer Research Foundation, Hyundai Hope on Wheels, UMass CCTS and ALS Association. This work was supported by DARPA HR0011-17-2-0049; US NIH RM1 HG009490, R01 EB022376, U01 AI142756 and R35 GM118062; and HHMI (to D.R.L.), and R01 CA195787; K22 CA181280 (to L.E.D.). This work was supported in part by the Marble Center for Cancer Nanomedicine and a Cancer Center Support (core) grant P30-CA14051 from the National Cancer Institute. M.R. was supported by the HHMI Hanna H. Gray Fellowship. L.W.K. is an NSF Graduate Research Fellow and was supported NIH Training Grant T32 GM095450. H.Y. was supported by the National Natural Science Foundation of China 31871345, the Young Thousand Talented Program from Wuhan University and startup funding from Wuhan University.

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C.Q.S., T.J., M.R., D.R.L., H.Y. and W.X. designed the study. C.Q.S., T.J., M.R., L.H.R., L.W.K., M.P.Z., E.M.S., J.L.D., Y.C., L.J.Z., L.E.D. and D.G.A. performed experiments and analysed data. C.Q.S., T.J., H.Y. and W.X. wrote the manuscript with comments from all authors.

Corresponding authors

Correspondence to David R. Liu or Hao Yin or Wen Xue.

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Competing interests

D.R.L. is a consultant and co-founder of Editas Medicine, Pairwise Plants and Beam Therapeutics, which are companies that use genome editing. The other authors declare no competing interests.

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Supplementary information

Supplementary Information

Supplementary figures and tables.

Reporting Summary

Supplementary Dataset 1

Guide-seq off-target sites.

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Song, C., Jiang, T., Richter, M. et al. Adenine base editing in an adult mouse model of tyrosinaemia. Nat Biomed Eng 4, 125–130 (2020). https://doi.org/10.1038/s41551-019-0357-8

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