Small-molecule compounds boost genome-editing efficiency of cytosine base editor

Author:

Zhao Tianyuan1,Li Qing2,Zhou Chenchen1,Lv Xiujuan1,Liu Hongyan1,Tu Tianxiang1,Tang Na3,Cheng Yanbo2,Liu Xiaoyu1,Liu Changbao4,Zhao Junzhao4,Song Zongming5,Wang Haoyi3,Li Jinsong2,Gu Feng1ORCID

Affiliation:

1. School of Ophthalmology and Optometry, Eye Hospital, Wenzhou Medical University, State Key Laboratory and Key Laboratory of Vision Science, Ministry of Health and Zhejiang Provincial Key Laboratory of Ophthalmology and Optometry, Wenzhou, Zhejiang, China

2. State Key Laboratory of Cell Biology, Shanghai Key Laboratory of Molecular Andrology, Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, Shanghai, China

3. State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Beijing, China

4. The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, China

5. Henan Eye Hospital, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University, People's Hospital of Henan University, Zhengzhou, Henan, China

Abstract

Abstract Cytosine base editor (CBE) enables targeted C-to-T conversions at single base-pair resolution and thus has potential therapeutic applications in humans. However, the low efficiency of the system limits practical use of this approach. We reported a high-throughput human cells-based reporter system that can be harnessed for quickly measuring editing activity of CBE. Screening of 1813 small-molecule compounds resulted in the identification of Ricolinostat (an HDAC6 inhibitor) that can enhance the efficiency of BE3 in human cells (2.45- to 9.21-fold improvement). Nexturastat A, another HDAC6 inhibitor, could also increase BE3-mediated gene editing by 2.18- to 9.95-fold. Ricolinostat and Nexturastat A also boost base editing activity of the other CBE variants (BE4max, YE1-BE4max, evoAPOBEC1-BE4max and SpRY-CBE4max, up to 8.32-fold). Meanwhile, combined application of BE3 and Ricolinostat led to >3-fold higher efficiency of correcting a pathogenic mutation in ABCA4 gene related to Stargardt disease in human cells. Moreover, we demonstrated that our strategy could be applied for efficient generation of mouse models through direct zygote injection and base editing in primary human T cells. Our study provides a new strategy to improve the activity and specificity of CBE in human cells. Ricolinostat and Nexturastat A augment the effectiveness and applicability of CBE.

Funder

National Key Research and Development Program of China

National Natural Science Foundation of China

Natural Science Foundation of Zhejiang Province

Wenzhou Medical University

Lin He's Academician Workstation of New Medicine and Clinical Translation

National Postdoctoral Program for Innovative Talents

China Postdoctoral Science Foundation

Wenzhou City Key Innovation Team of Reproductive Genetics

Wenzhou city

Publisher

Oxford University Press (OUP)

Subject

Genetics

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