The aminoglycoside geneticin permits translational readthrough of the CTNS W138X nonsense mutation in fibroblasts from patients with nephropathic cystinosis

• Published in: Pediatric nephrology (Berlin, West) Vol. 34; no. 5; pp. 873 - 881
• Main Authors: Brasell, Emma J., Chu, LeeLee, El Kares, Reyhan, Seo, Jung Hwa, Loesch, Robin, Iglesias, Diana M., Goodyer, Paul
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer Berlin Heidelberg 01.05.2019; Springer; Springer Nature B.V
• Subjects: Amino Acid Transport Systems, Neutral - genetics; Aminoglycosides; Care and treatment; Codon, Nonsense; Codons; Cystine - metabolism; Cystinosis; Cystinosis - drug therapy; Cystinosis - genetics; Diagnosis; Fanconi syndrome; Fibroblasts; Fibroblasts - drug effects; Fibroblasts - metabolism; Gene mutation; Genetic aspects; Genetic Vectors - genetics; Geneticin; Gentamicins - pharmacology; Gentamicins - therapeutic use; HEK293 Cells; Heterozygotes; Humans; Kidney transplantation; Lysosomes; Medicine; Medicine & Public Health; Mutation; Nephrology; Nonsense mutation; Original Article; Patients; Pediatrics; Peptide Chain Termination, Translational - drug effects; Peptide Chain Termination, Translational - genetics; Plasmids - genetics; Proteins; Recombinant Proteins - genetics; RNA, Messenger - analysis; Transfection; Translation; Urology; Cystinosis; Geneticin; Nonsense mutation; Aminoglycoside; Translational readthrough
• ISSN: 0931-041X; 1432-198X
• DOI: 10.1007/s00467-018-4094-0

Background Cystinosis is an ultrarare disorder caused by mutations of the cystinosin ( CTNS ) gene, encoding a cystine-selective efflux channel in the lysosomes of all cells of the body. Oral therapy with cysteamine reduces intralysosomal cystine accumulation and slows organ deterioration but cannot reverse renal Fanconi syndrome nor prevent the eventual need for renal transplantation. A definitive therapeutic remains elusive. About 15% of cystinosis patients worldwide carry one or more nonsense mutations that halt translation of the CTNS protein. Aminoglycosides such as geneticin (G418) can bind to the mammalian ribosome, relax translational fidelity, and permit readthrough of premature termination codons to produce full-length protein. Methods To ascertain whether aminoglycosides permit readthrough of the most common CTNS nonsense mutation, W138X, we studied the effect of G418 on patient fibroblasts. Results G418 treatment induced translational readthrough of CTNS W138X constructs transfected into HEK293 cells and expression of full-length endogenous CTNS protein in homozygous W138X fibroblasts. Conclusions Reduction in intracellular cystine indicates that the CTNS protein produced is functional as a cystine transporter. Interestingly, similar effects were seen even in W138X compound heterozygotes. These studies establish proof-of-principle for the potential of aminoglycosides to treat cystinosis and possibly other monogenic diseases caused by nonsense mutations.