Fig. 5

Retinal uptake, efficacy, and duration of action of mQR-421a upon intravitreal (IVT) injection in WT mice (A) Representative sections of untreated and mQR-421a-treated mouse retina. WT C57BL/6J mice received a single bilateral IVT injection of 50 μg of mQR-421a and were sacrificed to visualize the presence of mQR-421a at 7 and 259 days post-injection. mQR-421a was visualized using a Cy5-labeled FISH assay (red signal); cell nuclei were stained with DAPI (blue signal). GC, ganglion cell layer; INL, inner nuclear layer; RPE, retinal pigment epithelium. (B) Bioanalyzer (PCR) analysis of Ush2a exon 12 skipping in untreated and mQR-421a-treated mice. Ush2a transcripts were amplified using primers binding to exons 10 and 14. The top 1,164-base pair band corresponds to Ush2a transcripts with exon 12, and the bottom 522-base pair band corresponds to transcripts without exon 12. (C) Dose-dependent Ush2a exon 12 skipping by mQR-421a in WT C57BL/6J mice. Mice received a single bilateral IVT of 7.5, 15, 60, or 90 μg per eye of mQR-421a or 30 μg control AON. Ush2a transcripts with or without exon 12 were quantified by ddPCR and normalized by total Ush2a levels measured at exon 8-9, and percentage of exon 12 skipping was calculated. Mean ± SEM, n = 6 animals per dose per group. (D) Long duration of action of mQR-421a in mice retina. Mice received a single bilateral IVT of 30 μg mQR-421a per eye and followed for 1, 2, 14, 28, 56, 103, or 203 days. Ush2a transcripts with or without exon 12 were quantified by ddPCR and normalized by total Ush2a levels measured at exon 8-9, and percentage of exon 12 skipping was calculated. Mean ± SEM, n = 12 (1, 2, 14, and 28 day time points) or n = 8 (56, 103, and 203 day time point) eyes.