Fig. 5: Lipid nanoparticle delivery of PE.

a, Immunoprecipitation-encapsulation assay of PE2 RNP LNP. b, Particle-size distribution of PE2 RNP LNP prepared with 1.5% DMG-PEG 2000 and ionizable lipids SM102 (d = 270 nm, PdI = 0.068), CL4H6 (d = 298 nm, PdI = 0.024) or DODMA (d = 176 nm, PdI = 0.096), and with 2.5% DMG-PEG 2000 and ionizable lipid SM102 (d = 194 nm, PdI = 0.040). Average plots are shown, n = 3 replicates. c,d, Delivery of PE2 RNP LNP to rd12 reporter cells (c), analysed by flow cytometry (bottom) and fluorescence microscopy (top) (d). Representative microscopic image is shown for rd12 reporter cells treated with 20 nM PE2 RNP LNP with 2.5% DMG-PEG 2000. 20 nM corresponds to 6 µg ml⁻¹. Scale bar, 100 μm. Two biological replicates with two analytical replicates each, mean ± s.d. e, Cryoelectron-microscopy image of PE2 RNP LNP containing ionizable lipid SM102 and 2.5% DMG-PEG 2000. Scale bar, 100 nm. f, Mass spectrometric quantification of PE2 as protein, RNP and LNP. Three analytical replicates, mean ± s.d. g, PE-mediated rescue of expression of RPE65 in a cell line transformed with cDNA encoding Rpe65 rd12. LNP with 2.5% DMG-PEG 2000 was used, and RNP concentration was 20 nM, 6 µg ml⁻¹. h, Next-generation sequencing analysis of PE-editing outcome in the cells with cDNA encoding Rpe65 rd12. Three analytical replicates, mean ± s.d. i, Off-target analysis of Rpe65 rd12 cells treated with PE RNP LNP. j, Restoration of visual function in rd12 mice treated with 1.6 µM (CL4H6, 476 ng RNP per eye), 2.0 µM (SM102, DODMA, 596 ng RNP per eye) or 2.2 µM (SM102 with 2.5% DMG-PEG 2000, 655 ng RNP per eye) PE2 LNP, 1 µl per eye, as evidenced by ERG. At least 5 eyes, mean ± s.d. Kruskal–Wallis test with Dunn’s multiple comparisons test; *P < 0.05, **P < 0.01, ****P < 0.0001, ^NSP ≥ 0.05. Uncropped blots are available as Source data.

Source data