Engineering a precise adenine base editor with minimal bystander editing

Chen, Liang; Zhang, Shun; Xue, Niannian; Hong, Mengjia; Zhang, Mengjie; Zhang, Dan; Yang, Jing; Bai, Sijia; Huang, Yifan; Meng, Haowei; Wu, Hao; Luan, Changming; Zhu, Biyun; Ru, Gaomeng; Gao, Hongyi; Zhong, Liping; Liu, Meizhen; Liu, Mingyao; Cheng, Yiyun; Yi, Chengqi; Wang, Liren; Zhao, Yongxiang; Song, Gaojie; Li, Dali

doi:10.1038/s41589-022-01163-8

Cited by 93 publications

(58 citation statements)

References 43 publications

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“…Adenine deaminase engineering has been also applied to reduce the RNA off-target effects through introducing critical amino acid mutations 13 , 14 . Recently, we have shown that through introduction of N108Q and L145T substitutions, ABE9 exhibits 1–2 nucleotides editing window, eliminating preferentially cytosine bystander editing and minimizing off-targeting effects on DNA and RNA 15 . However, the A-to-G base editing efficiency, especially near the PAM region, is still limited and impedes its broader applications.…”

Section: Introductionmentioning

confidence: 99%

Improving adenine and dual base editors through introduction of TadA-8e and Rad51DBD

et al. 2023

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Base editors, including dual base editors, are innovative techniques for efficient base conversions in genomic DNA. However, the low efficiency of A-to-G base conversion at positions proximal to the protospacer adjacent motif (PAM) and the A/C simultaneous conversion of the dual base editor hinder their broad applications. In this study, through fusion of ABE8e with Rad51 DNA-binding domain, we generate a hyperactive ABE (hyABE) which offers improved A-to-G editing efficiency at the region (A10-A15) proximal to the PAM, with 1.2- to 7-fold improvement compared to ABE8e. Similarly, we develop optimized dual base editors (eA&C-BEmax and hyA&C-BEmax) with markedly improved simultaneous A/C conversion efficiency (1.2-fold and 1.5-fold improvement, respectively) compared to A&C-BEmax in human cells. Moreover, these optimized base editors catalyze efficiently nucleotide conversions in zebrafish embryos to mirror human syndrome or in human cells to potentially treat genetic diseases, indicating their great potential in broad applications for disease modeling and gene therapy.

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Section: Introductionmentioning

confidence: 99%

Improving adenine and dual base editors through introduction of TadA-8e and Rad51DBD

et al. 2023

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“…In CBEs, these variants include SECURE-BE3 150 , eA3A-BE3 152 , 166 , hA3G-CBE 167 , AALN-BE4 148 , YE1-BE4 148 , transformer BE (tBE) 168 , AID-2S/AID-3S 169 , and other next-generation CBEs 170 , 171 . In ABEs, SECURE-ABE 135 , ABE7.10-F148A 152 , ABEmaxAW or ABEmaxQW 151 , ABE8e-V106W 136 , ABE-del153 172 and ABE9 (N108Q/L145T) 173 were created to reduce RNA off-targeting and narrow the editing window. Of note, many of these mutations may reduce the deaminase activity.…”

Section: Dna Base Editorsmentioning

confidence: 99%

Assessing and advancing the safety of CRISPR-Cas tools: from DNA to RNA editing

2023

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CRISPR-Cas gene editing has revolutionized experimental molecular biology over the past decade and holds great promise for the treatment of human genetic diseases. Here we review the development of CRISPR-Cas9/Cas12/Cas13 nucleases, DNA base editors, prime editors, and RNA base editors, focusing on the assessment and improvement of their editing precision and safety, pushing the limit of editing specificity and efficiency. We summarize the capabilities and limitations of each CRISPR tool from DNA editing to RNA editing, and highlight the opportunities for future improvements and applications in basic research, as well as the therapeutic and clinical considerations for their use in patients.

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“…In addition, concerning the base editing window, two aspects are worthy of further optimization. (i) Narrow the editing window of BE to a single base, reduce the by-products of unintended editing, for example, a more precise adenine base editor ABE9 (Figure 1L), which was developed recently by introducing two mutations L145T and N108Q in ABE8e, maintained the editing activity and minimized the editing window to position 5-6 in mammalian cells (Chen et al, 2022b). Furthermore, by combining with PAM-less Cas proteins, it will be possible to achieve accurate single base editing at any target sites in the genome.…”

Section: Optimization Of the Existing Bes And Exploitation Of Novel Besmentioning

confidence: 99%

Plant base editing and prime editing: The current status and future perspectives

Zhang

et al. 2023

JIPB

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Precise replacement of an allele with an elite allele controlling an important agronomic trait in a predefined manner by gene editing technologies is highly desirable in crop improvement. Base editing and prime editing are two newly developed precision gene editing systems which can introduce the substitution of a single base and install the desired short indels to the target loci in the absence of double-strand breaks and donor repair templates, respectively. Since their discoveries, various strategies have been attempted to optimize both base editor (BE) and prime editor (PE) in order to improve the precise editing efficacy, specificity, and expand the targeting scopes. Here, we summarize the latest development of various BEs and PEs, as well as their applications in plants. Based on these progresses, we recommend the appropriate BEs and PEs for both basic plant research and crop improvement. Moreover, we propose the perspectives for further optimization of these two editors. We envision that both BEs and PEs will become the routine and customized precise gene editing tools for both plant biological research and crop improvement in the near future.

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Engineering a precise adenine base editor with minimal bystander editing

Cited by 93 publications

References 43 publications

Improving adenine and dual base editors through introduction of TadA-8e and Rad51DBD

Improving adenine and dual base editors through introduction of TadA-8e and Rad51DBD

Assessing and advancing the safety of CRISPR-Cas tools: from DNA to RNA editing

Plant base editing and prime editing: The current status and future perspectives

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