We developed a systematic evaluation approach to assess the off-target effects of base editors (BE3, YE1-BE3-FNLS, and ABE7.10(F148A)) in transgenic mice. The study found that BE3 expression generated de novo mutations in the offspring of transgenic mice. Transcriptome analysis revealed that both BE3 and YE1-BE3-FNLS induced transcriptome-wide single nucleotide variants (SNVs), and the number of RNA SNVs was positively correlated with CBE expression levels. On the other hand, ABE7.10(F148A) showed no detectable off-target DNA or RNA SNVs. Importantly, long-term monitoring revealed abnormal phenotypes such as obesity and developmental delay in mice with permanent genomic BE3 overexpression, highlighting potential overlooked side effects of BE3 in vivo.
Base editors have been reported to induce off-target mutations in cultured cells, mouse embryos and rice, but their long-term effects in vivo remain unknown. Here, we develop a Systematic evaluation Approach For gene Editing tools by Transgenic mIce (SAFETI), and evaluate the off-target effects of BE3, high fidelity version of CBE (YE1-BE3-FNLS) and ABE (ABE7.10(F148A)) in similar to 400 transgenic mice over 15 months. Whole-genome sequence analysis reveals BE3 expression generated de novo mutations in the offspring of transgenic mice. RNA-seq analysis reveals both BE3 and YE1-BE3-FNLS induce transcriptome-wide SNVs, and the numbers of RNA SNVs are positively correlated with CBE expression levels across various tissues. By contrast, ABE7.10(F148A) shows no detectable off-target DNA or RNA SNVs. Notably, we observe abnormal phenotypes including obesity and developmental delay in mice with permanent genomic BE3 overexpression during long-time monitoring, elucidating a potentially overlooked aspect of side effects of BE3 in vivo.
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