Targeted insertion of an optimized donor DNA is effective in a humanized mouse model of dominant retinitis pigmentosa
9 affiliations
- 1Telethon Institute of Genetics and Medicine (TIGEM), Pozzuoli, Italy
- 2Institute for Transfusion Medicine and Gene Therapy, Medical Center – University of Freiburg, Freiburg, Germany
- 3Center for Cell and Gene Therapy Freiburg (CGF), Faculty of Medicine, University of Freiburg, Freiburg, Germany
- 4Institute of Medical Bioinformatics and Systems Medicine, Medical Center – University of Freiburg, Freiburg, Germany
- 5German Cancer Consortium (DKTK), Partner site Freiburg, a partnership between DKFZ and Medical Center - University of Freiburg, Freiburg, Germany
- 6Department of Advanced Biomedical Sciences and Department of Translational Medicine, University of Naples “Federico II”, Naples, Italy
- 7Medical Genetics, Department of Advanced Biomedical Sciences, University of Naples “Federico II”, Naples, Italy
- 8Scuola Superiore Meridionale (SSM), Genomics and Experimental Medicine Program, Naples, Italy
- 9Gene Therapy Joint Lab, Department of Advanced Biomedical Sciences and Department of Translational Medicine, University of Naples “Federico II”, Naples, Italy
Abstract
Retinitis pigmentosa (RP) affects 1 in 3,000 individuals worldwide, with 30%–40% of cases inherited as autosomal dominant (AD). Mutations in RHO (RP4) are the most common cause of ADRP. Because most RHO mutations exert gain-of-function or dominant-negative effects, conventional gene supplementation is insufficient, requiring mutant allele inactivation. Allele-specific editing is impractical, as each mutation requires a unique therapeutic strategy. We present a mutation-agnostic, RHO-specific approach using adeno-associated viral vector-mediated homology-independent targeted integration (AAV-HITI). Optimized donor DNA design enables targeted integration and efficient transgene expression from the endogenous RHO locus. In a humanized RP4 mouse model harboring the RHO P23H mutant allele alongside an endogenous wild-type mouse Rho allele, AAV-HITI significantly improves retinal structure, function, and visual acuity up to 1 year post-treatment. Comprehensive molecular analyses characterize on-target editing in mouse retina and off-target editing in a human cell line. These findings establish an effective, human-centric AAV-HITI platform for RP4 and support its evaluation in this and other dominant genetic conditions.
Related
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- Generation of retinitis pigmentosa patient-derived hiPSC lines (IDVi007-A, IDVi007-B) carrying the RHO c.68C > A variant (p.P23H) and CRISPR/Cas9-corrected isogenic hiPSC lines (IDVi007-A-1, IDVi007-A-2, IDVi007-A-3)
BibTeX
@article{esposito2026targeted,
title = {{Targeted insertion of an optimized donor DNA is effective in a humanized mouse model of dominant retinitis pigmentosa}},
author = {Federica Esposito and Arjun Padmanabhan and Manuel Rhiel and Mariangela Lupo and Elena Marrocco and Stefano Auricchio and Xabier Bujanda Cundin and Simona Pellecchia and Tatjana I. Cornu and Geoffroy Andrieux and Manel Llado-Santaeularia and Enrico Maria Surace and Ivana Trapani and Pasqualina Colella and Gennaro Gambardella and Toni Cathomen and Alberto Auricchio},
journal = {Cell Reports Medicine},
year = {2026},
month = sep,
doi = {10.1016/j.xcrm.2026.103047},
url = {https://doi.org/10.1016/j.xcrm.2026.103047},
note = {CC BY-NC-ND 4.0},
}
RIS
TY - JOUR
TI - Targeted insertion of an optimized donor DNA is effective in a humanized mouse model of dominant retinitis pigmentosa
AU - Federica Esposito
AU - Arjun Padmanabhan
AU - Manuel Rhiel
AU - Mariangela Lupo
AU - Elena Marrocco
AU - Stefano Auricchio
AU - Xabier Bujanda Cundin
AU - Simona Pellecchia
AU - Tatjana I. Cornu
AU - Geoffroy Andrieux
AU - Manel Llado-Santaeularia
AU - Enrico Maria Surace
AU - Ivana Trapani
AU - Pasqualina Colella
AU - Gennaro Gambardella
AU - Toni Cathomen
AU - Alberto Auricchio
PY - 2026
DA - 2026/09/11
JO - Cell Reports Medicine
DO - 10.1016/j.xcrm.2026.103047
UR - https://doi.org/10.1016/j.xcrm.2026.103047
AB - Retinitis pigmentosa (RP) affects 1 in 3,000 individuals worldwide, with 30%–40% of cases inherited as autosomal dominant (AD). Mutations in RHO (RP4) are the most common cause of ADRP. Because most RHO mutations exert gain-of-function or dominant-negative effects, conventional gene supplementation is insufficient, requiring mutant allele inactivation. Allele-specific editing is impractical, as each mutation requires a unique therapeutic strategy. We present a mutation-agnostic, RHO-specific approach using adeno-associated viral vector-mediated homology-independent targeted integration (AAV-HITI). Optimized donor DNA design enables targeted integration and efficient transgene expression from the endogenous RHO locus. In a humanized RP4 mouse model harboring the RHO P23H mutant allele alongside an endogenous wild-type mouse Rho allele, AAV-HITI significantly improves retinal structure, function, and visual acuity up to 1 year post-treatment. Comprehensive molecular analyses characterize on-target editing in mouse retina and off-target editing in a human cell line. These findings establish an effective, human-centric AAV-HITI platform for RP4 and support its evaluation in this and other dominant genetic conditions.
SN - 2666-3791
ER -