Figure 7. Performance of AAV2-7m8-PR1.7 vector–promoter combination in human cones.

(A–C) GFP expression in human induced pluripotent stem cell–derived (iPSC-derived) retinal organoids (n = 10 organoids) infected with AAV2-7m8-PR1.7-GFP. (A) Brighfield, (B) epifluorescence, and (C) confocal images of 43-day-old whole mount organoids infected with AAV2.7m8-PR1.7-GFP at day 28 with a dose of 5 × 10¹⁰ vg/organoid. Scale bar: 200 μm in A and B, and 250 μm in C. Outline in C represents the edges of the organoids (D–F) Retinal organoid cryosections for visualization of GFP expression (cyan). Transduced cones are visualized by superimposition of GFP (cyan) and human cone arrestin (hCAR) immunostaining (magenta). Scale bar: 20 μm in D–F. Arrows represent colocalization of GFP and hCAR stainings. (G–I) Efficient and specific transduction of human cones in postmortem retinal explants. (G) Postmortem human retinal explant placed in culture. Dashed circle shows the approximate area where 1 × 10¹⁰ viral particles were deposited onto the explant (n = 2 explants from 2 eyes of a single donor). (H) Close-up of the transduced area showing high-level GFP fluorescence in region of the explant in contact with the vector. Scale bar: 100 μm. (I) GFP expression (cyan) is restricted to the photoreceptor layer as shown by DAPI (blue) staining. (J) GFP is expressed in cones as shown by colocalization of GFP staining of cone markers, namely M/L opsin. Scale bar: 50 μm in I–J. Arrows represent colocalization of GFP and M/L opsin stainings. AAV, adeno-associated virus; vg, viral genome; PR1.7, promoter of 1.7 kilobases in length, based on the human red opsin gene enhancer and promoter sequences.