Abstract
Purpose :
Mutations within the retinitis pigmentosa GTPase regulator (RPGR) are the most frequent cause of X-Linked Retinitis Pigmentosa (XLRP), a common and severe form of inherited retinal disease. XLRP is characterised by the progressive degeneration and loss of photoreceptors, leading to visual loss and, ultimately, bilateral blindness. Unfortunately, treatments for RPGR-associated XLRP are non-existent. Therefore, we sought to investigate the efficacy of RPGRORF15 gene supplementation in human RPGR-deficient retinal organoids (ROs).
Methods :
Isogenic RPGR knockout (KO) induced pluripotent stem cells (iPSCs) were generated using established CRISPR/Cas9 gene editing methods. RPGR-KO iPSCs were differentiated into 3D ROs, which were utilised to test the RPGR clinical vector construct. Molecular, structural, and functional readouts included RPGR quantification, RPGR photoreceptor localization, RPGR glutamylation and ciliation assessment.
Results :
Successful differentiation of RPGR-KO iPSCs was confirmed by qPCR and immunocytochemistry of major retinal and phototransduction markers. Viral transduction of RPGR-KO ROs with AAV-RPGR led to restoration of RPGR expression in human rods and cones. RPGR was localised at the photoreceptor cilium and led to marked improvements in several molecular readouts.
Conclusions :
The RPGR transgene was correctly expressed, processed, and localised in human rods and cones following viral transduction of RPGR-deficient human ROs. These data agree with the reported Phase I/II trial positive results in patients with RPGR-associated XLRP.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.