Fig. 1: RNP + AAV-based targeted insertion in CYBA.
a Schematic representation of the targeted insertion of an SFFV-GFP cassette into the CYBA c.287+1 locus. b Percentage of indel formation in CD34+ HSPCs from healthy donors using different sgRNAs targeting sites close to the CYBA c.287+1 G > T variant. c Targeted integration levels and viabilities of treated HSPCs 4 days after nucleofection. Targeted integration levels were determined as the percentage of GFPhigh cells. d Proliferation of treated HSPCs measured by total cell count up to 8 days after nucleofection. e Flow cytometry-based CFU assay of treated HSPCs showing total colony counts and distribution of HSPC-derived colonies. f The percentage of GFP+ colonies observed in the CFU assay. g Human chimerism in injected mice was measured in the blood, spleen, and bone marrow (BM) 16 weeks after injection. Experiments were conducted using HSPCs from 4 individual healthy donors. h The distribution of lineages within the human graft of untreated HSPCs (n = 4) and CYBA gene-edited HSPCs (n = 8). i Targeted integration levels in the human graft 16 weeks after injection of untreated HSPCs (n = 4) and CYBA gene-edited HSPCs (n = 8). Data represented as mean (n = 3 biological replicates) ± standard deviation unless otherwise specified. Statistical significance was determined by one-way ANOVA with Tukey’s multiple comparisons test. For the CFU assay, total colony numbers were used for the ANOVA test. Source data are provided as a Source Data file.
