Fig. 7: sgRNA-dependent DNA off-target activity of the base-editing system.

a–c sgRNA-dependent off-target DNA sites, as evaluated by GUIDE-seq analysis, of LRF_bs_3 (LRF 4C) (a), LRF_bs_2 (LRF 8C) (b), and KLF1_bs_1 (LRF 2T and KLF1) (c) sgRNAs in HEK293T cells. sgRNAs were coupled with a Cas9-nuclease corresponding to the Cas9 nickase included in the base editor (Cas9-SpRY for LRF_bs_3 and LRF_bs_2 sgRNAs, and Cas9 for KLF1_bs_1 sgRNA). The protospacer targeted by each sgRNA and the PAM are reported on top of each panel, followed by the off-target sites and their mismatches with the on-target (highlighted in color). The number of sequencing reads, the chromosomal coordinates (Human GRCh37/hg19), and the site of each off-target are reported. d–f Frequency of C-G to T-A (d and e) or A-T to G-C (f) base-editing conversion at on-target and off-target (OT) sites, for control and LRF 4C (d), LRF 8C (e), LRF 2T (f), and KLF1 (f) samples, as measured by targeted NGS sequencing. Data are expressed as individual values and median (n = 3 biologically independent experiments, 3 donors). *P = 0.0107 for d, or P = 0.0299 for f; **P = 0.0022; ****P ≤ 0.0001 (two-way ANOVA with Sidak (d and f) or Tukey (f) correction for multiple comparisons). g Venn diagrams showing the overlapping of C > T or A > G single-nucleotide variants in exons, in control (Ctr), CBE-SpRY-OPT2-, or ABEmax- treated HSPCs obtained from three different HD. Source data are provided as a Source Data file.