Gentamicin-induced readthrough of stop codons in duchenne muscular dystrophy

• Published in: Annals of neurology Vol. 67; no. 6; pp. 771 - 780
• Main Authors: Malik, Vinod, Rodino-Klapac, Louise R., Viollet, Laurence, Wall, Cheryl, King, Wendy, Al-Dahhak, Roula, Lewis, Sarah, Shilling, Christopher J., Kota, Janaiah, Serrano-Munuera, Carmen, Hayes, John, Mahan, John D., Campbell, Katherine J., Banwell, Brenda, Dasouki, Majed, Watts, Victoria, Sivakumar, Kumaraswamy, Bien-Willner, Ricardo, Flanigan, Kevin M., Sahenk, Zarife, Barohn, Richard J., Walker, Christopher M., Mendell, Jerry R.
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.06.2010; Wiley-Liss
• Subjects: Adolescent; Audiometry - methods; Biological and medical sciences; Child; Child, Preschool; Codon, Terminator - drug effects; Codon, Terminator - genetics; Cohort Studies; Creatine Kinase - blood; Diseases of striated muscles. Neuromuscular diseases; Enzyme-Linked Immunosorbent Assay - methods; Gentamicins - therapeutic use; Humans; Medical sciences; Muscle Cells - pathology; Muscular Dystrophy, Duchenne - blood; Muscular Dystrophy, Duchenne - genetics; Muscular Dystrophy, Duchenne - pathology; Mutation - genetics; Neurology; Protein Synthesis Inhibitors - therapeutic use; T-Lymphocytes - drug effects; T-Lymphocytes - pathology; Time Factors; Neuromuscular diseases; Nervous system diseases; Codon; Duchenne muscular dystrophy; Gentamicin; Genetic disease
• ISSN: 0364-5134; 1531-8249; 1531-8249
• DOI: 10.1002/ana.22024

Objective The objective of this study was to establish the feasibility of long‐term gentamicin dosing to achieve stop codon readthrough and produce full‐length dystrophin. Mutation suppression of stop codons, successfully achieved in the mdx mouse using gentamicin, represents an important evolving treatment strategy in Duchenne muscular dystrophy (DMD). Methods Two DMD cohorts received 14‐day gentamicin (7.5mg/kg/day): Cohort 1 (n = 10) stop codon patients and Cohort 2 (n = 8) frameshift controls. Two additional stop codon DMD cohorts were gentamicin treated (7.5mg/kg) for 6 months: Cohort 3 (n = 12) dosed weekly and Cohort 4 (n = 4) dosed twice weekly. Pre‐ and post‐treatment biopsies were assessed for dystrophin levels, as were clinical outcomes. Results In the 14‐day study, serum creatine kinase (CK) dropped by 50%, which was not seen in frameshift DMD controls. After 6 months of gentamicin, dystrophin levels significantly increased (p = 0.027); the highest levels reached 13 to 15% of normal (1 in Cohort 3, and 2 in Cohort 4), accompanied by reduced serum CK favoring drug‐induced readthrough of stop codons. This was supported by stabilization of strength and a slight increase in forced vital capacity. Pretreatment stable transcripts predicted an increase of dystrophin after gentamicin. Readthrough efficiency was not affected by the stop codon or its surrounding fourth nucleotide. In 1 subject, antigen‐specific interferon‐γ enzyme‐linked immunospot assay detected an immunogenic dystrophin epitope. Interpretation The results support efforts to achieve drug‐induced mutation suppression of stop codons. The immunogenic epitope resulting from readthrough emphasizes the importance of monitoring T‐cell immunity during clinical studies that suppress stop codons. Similar principles apply to other molecular strategies, including exon skipping and gene therapy. ANN NEUROL 2010;67:771–780