Fig. 5: Retinal disease correction in vivo with PE-eVLPs.
a, An optimized PE3b strategy to correct a 4-bp deletion in Mfrp in the rd6 mice. The mutation (red), the epegRNA spacer (light blue) and the ngRNA spacer sequence (dark blue) are highlighted. b, Schematic of subretinal injection of rd6 mice. c, Prime editing efficiency and the associated indel frequency in the genomic DNA collected from the rd6 mice. Data are represented as mean values ± s.e.m. Each dot represents an individual mouse for n = 4 (untreated) or n = 11 (v3 PE3b-eVLP treated). d,e, Western blot of RPE tissue protein extracts (d) and immunohistochemistry blot on RPE flatmounts (e) from wild-type C57BL/6J mice, untreated rd6 mice and v3 PE3-eVLP-treated rd6 mice. Flatmounts were stained with anti-MFRP antibody (green) and anti-ZO-1 antibody (red). f,g, Analysis of PE-dependent editing (f) and indel byproducts (g) at the on-target site and top ten CIRCLE-seq nominated off-target sites associated with the rd6 epegRNA (f) and ngRNA (g) sequence. For f and g, data are represented as mean values ± s.e.m. Each dot represents an individual mouse for n = 3 (untreated) or n = 3 (v3 PE3b-eVLP treated). h, The R44X mutation in Rpe65 exon 3 in the rd12 mouse model and the optimized PE3b strategy used for correction of this mutation. The mutation (red), the epegRNA spacer (light blue) and the ngRNA spacer sequence (dark blue) are highlighted. i, Prime editing efficiency of Rpe65 R44X mutation correction and the associated indel frequency in genomic DNA collected from rd12 mice. j, Prime editing efficiency of Rpe65 R44X mutation correction in Rpe65 RNA collected from rd12 mice. k, Western blot of RPE tissue protein extracts from wild-type C57BL/6J mice, untreated rd12 mice and v3 PE3b-eVLP-treated rd12 mice. l, Scotopic A- and B-wave amplitudes measured by ERG following overnight dark adaptation. For i, j and l, data are represented as mean values ± s.e.m. Each dot represents an individual mouse for n = 4 (untreated) or n = 4 (v3 PE3b-eVLP treated). m, Representative ERG waveforms from wild-type C57BL/6J mice, untreated rd12 mice and v3 PE3-eVLP-treated rd12 mice. Each mouse received approximately 4.2 × 1010 eVLPs per eye. WT, wild type.
