title: Effect of Readthrough Treatment in Fibroblasts of Patients Affected by Lysosomal Diseases Caused by Premature Termination Codons.
abstract: Aminoglycoside antibiotics, such as gentamicin, may induce premature termination codon (PTC) readthrough and elude the nonsense-mediated mRNA decay mechanism. Because PTCs are frequently involved in lysosomal diseases, readthrough compounds may be useful as potential therapeutic agents. The aim of our study was to identify patients responsive to gentamicin treatment in order to be used as positive controls to further screen for other PTC readthrough compounds. With this aim, fibroblasts from 11 patients affected by 6 different lysosomal diseases carrying PTCs were treated with gentamicin. Treatment response was evaluated by measuring enzymatic activity, abnormal metabolite accumulation, mRNA expression, protein localization, and cell viability. The potential effect of readthrough was also analyzed by in silico predictions. Results showed that fibroblasts from 5/11 patients exhibited an up to 3-fold increase of enzymatic activity after gentamicin treatment. Accordingly, cell lines tested showed enhanced well-localized protein and/or increased mRNA expression levels and/or reduced metabolite accumulation. Interestingly, these cell lines also showed increased enzymatic activity after PTC124 treatment, which is a PTC readthrough-promoting compound. In conclusion, our results provide a proof-of-concept that PTCs can be effectively suppressed by readthrough drugs, with different efficiencies depending on the genetic context. The screening of new compounds with readthrough activity is a strategy that can be used to develop efficient therapies for diseases caused by PTC mutations.
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain.
authors.forename: Leslie
authors.initials: L
authors.lastname: Matalonga
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain.
authors.forename: Ángela
authors.initials: Á
authors.lastname: Arias
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain.
authors.forename: Frederic
authors.initials: F
authors.lastname: Tort
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain.
authors.forename: Xènia
authors.initials: X
authors.lastname: Ferrer-Cortés
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain.
authors.forename: Judit
authors.initials: J
authors.lastname: Garcia-Villoria
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain.
authors.forename: Maria Josep
authors.initials: MJ
authors.lastname: Coll
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Gort
authors.affiliation: Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic, IDIBAPS, CIBER de Enfermedades Raras (CIBERER), Barcelona, Spain. aribes@clinic.ub.es.
authors.forename: Antonia
authors.initials: A
authors.lastname: Ribes
mesh_terms: D000118:Acetylglucosaminidase / Q000235:genetics / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D018389:Codon, Nonsense / Q000235:genetics*; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D005839:Gentamicins / Q000494:pharmacology; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D006025:Glycosaminoglycans / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016464:Lysosomal Storage Diseases / Q000235:genetics* / Q000473:pathology*; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D008297:Male; D009154:Mutation / Q000235:genetics; D010069:Oxadiazoles / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D013108:Sphingomyelin Phosphodiesterase / Q000235:genetics*
chemical_list: D018389:Codon, Nonsense; D005839:Gentamicins; D006023:Glycoproteins; D006025:Glycosaminoglycans; C018322:IDS protein, human; D010069:Oxadiazoles; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; C543066:SMPD1 protein, human; D013108:Sphingomyelin Phosphodiesterase; C033079:alpha-N-acetyl-D-glucosaminidase; D000118:Acetylglucosaminidase; C515878:ataluren
keywords: Gentamicin; PTC124; aminoglycoside; lysosomal disease; premature stop codon; readthrough treatment