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Localizing the RPGR protein along the cilium: a new method to determine efficacies to treat RPGR mutations


Da Costa, Romain; Glaus, Esther; Tiwari, Amit; Kloeckener-Gruissem, Barbara; Berger, Wolfgang; Neidhardt, John (2015). Localizing the RPGR protein along the cilium: a new method to determine efficacies to treat RPGR mutations. Gene Therapy, 22(5):413-420.

Abstract

Retinal dystrophies constitute a group of clinically and genetically heterogeneous diseases that cause visual impairment. As treatments are not readily available, readout assays performed in patient-derived cells can aid in the development and comparative analysis of therapeutic approaches. We describe a new method with which the localization of the retinitis pigmentosa GTPase regulator (RPGR) protein along the cilium can be used as a measure for treatment efficacy. In a patient-derived fibroblast cell line, we found that the RPGR protein is mislocalized along the ciliary axoneme. The patient carried a point mutation that leads to skipping of RPGR exon 10. We confirmed that this skipping is causative for the impaired localization of RPGR using a U7 small nuclear RNA (U7snRNA)-based antisense approach in control cells. Treatment of the patient-derived fibroblasts with therapeutic U1snRNA significantly corrected the proteins’ mislocalization. In this proof of principle study, we show that detecting the RPGR protein along the cilium provides a reliable and quantifiable readout assay to evaluate the efficacy of therapies intended to correct or silence RPGR gene mutations. This method opens the possibility to compare different therapeutic agents, and thus facilitate the identification of treatment options for the clinically and molecularly complex RPGR-associated diseases.

Abstract

Retinal dystrophies constitute a group of clinically and genetically heterogeneous diseases that cause visual impairment. As treatments are not readily available, readout assays performed in patient-derived cells can aid in the development and comparative analysis of therapeutic approaches. We describe a new method with which the localization of the retinitis pigmentosa GTPase regulator (RPGR) protein along the cilium can be used as a measure for treatment efficacy. In a patient-derived fibroblast cell line, we found that the RPGR protein is mislocalized along the ciliary axoneme. The patient carried a point mutation that leads to skipping of RPGR exon 10. We confirmed that this skipping is causative for the impaired localization of RPGR using a U7 small nuclear RNA (U7snRNA)-based antisense approach in control cells. Treatment of the patient-derived fibroblasts with therapeutic U1snRNA significantly corrected the proteins’ mislocalization. In this proof of principle study, we show that detecting the RPGR protein along the cilium provides a reliable and quantifiable readout assay to evaluate the efficacy of therapies intended to correct or silence RPGR gene mutations. This method opens the possibility to compare different therapeutic agents, and thus facilitate the identification of treatment options for the clinically and molecularly complex RPGR-associated diseases.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Center for Integrative Human Physiology
04 Faculty of Medicine > Institute of Medical Molecular Genetics
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:2015
Deposited On:03 Nov 2015 15:26
Last Modified:05 Apr 2016 19:28
Publisher:Nature Publishing Group
ISSN:0969-7128
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1038/gt.2014.128
Official URL:http://www.ncbi.nlm.nih.gov/pubmed/25630948

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