PCDH15 dual-AAV gene therapy for deafness and blindness in Usher syndrome type 1F models
Maryna V. Ivanchenko, … , Bence György, David P. Corey
Maryna V. Ivanchenko, … , Bence György, David P. Corey
Published October 23, 2024
Citation Information: J Clin Invest. 2024;134(23):e177700. https://doi.org/10.1172/JCI177700.
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Research Article Ophthalmology Otology

PCDH15 dual-AAV gene therapy for deafness and blindness in Usher syndrome type 1F models

Abstract

Usher syndrome type 1F (USH1F), resulting from mutations in the protocadherin-15 (PCDH15) gene, is characterized by congenital lack of hearing and balance, and progressive blindness in the form of retinitis pigmentosa. In this study, we explore an approach for USH1F gene therapy, exceeding the single AAV packaging limit by employing a dual–adeno-associated virus (dual-AAV) strategy to deliver the full-length PCDH15 coding sequence. We demonstrate the efficacy of this strategy in mouse USH1F models, effectively restoring hearing and balance in these mice. Importantly, our approach also proves successful in expressing PCDH15 protein in clinically relevant retinal models, including human retinal organoids and nonhuman primate retina, showing efficient targeting of photoreceptors and proper protein expression in the calyceal processes. This research represents a major step toward advancing gene therapy for USH1F and the multiple challenges of hearing, balance, and vision impairment.

Authors

Maryna V. Ivanchenko, Daniel M. Hathaway, Eric M. Mulhall, Kevin T.A. Booth, Mantian Wang, Cole W. Peters, Alex J. Klein, Xinlan Chen, Yaqiao Li, Bence György, David P. Corey

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Figure 6

Localization of endogenous and dual-AAV–delivered PCDH15 in NHP retina.

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Localization of endogenous and dual-AAV–delivered PCDH15 in NHP retina. ...
(A) Dual AAV vectors were injected subretinally into the eye of a green monkey (C. sabaeus). At 9 weeks, the treated retina was assayed for expression and localization by histology. (B) Immunofluorescence labeling of endogenous PCDH15 (green) in the retina of a cynomolgus monkey (M. fascicularis). An antibody against ARR3 (magenta) marks cone photoreceptors, and an antibody against Rho (blue) marks OSs of rods. PCDH15 (green) is located at the junction between ISs and OSs and along the calyceal processes (arrows) in both cones and rods (n = 4 punches). (C) Scanning electron micrograph of the inner/outer segment junction in a cone photoreceptor (right) and rod photoreceptors (left) in a cynomolgus monkey (n = 4 punches). The calyceal processes (arrows) protrude from the apical ISs of photoreceptors to surround the OSs. (D) Immunofluorescence labeling of endogenous PCDH15 (green) in the vehicle-injected control retina of a green monkey (n = 3 punches). PCDH15 (green) is located at the junction between ISs and OSs and along the calyceal processes (arrows) in both cones and rods. White asterisk indicates the OS of a cone photoreceptor. BODIPY labels all membranes of photoreceptors. (E) Representative confocal microscopy images of cryosections from the bleb area created by the dual-AAV injection. An anti-HA signal was detected in the photoreceptors injected with dual AAVs, seen at low magnification (left) and higher magnification (right two panels). (F) Transduction efficiency in rod and cone photoreceptors injected with dual AAVs. Data are presented as mean ± SEM. (G) The HA label was not seen in the eye injected with the vehicle (n = 3 blebs). Scale bars: 5 μm (B, D, and G), 1 μm (C and E).