title: A novel approach to identify Duchenne muscular dystrophy patients for aminoglycoside antibiotics therapy.
abstract: Aminoglycoside antibiotics have been found to suppress nonsense mutations located in the defective dystophin gene in mdx mice, suggesting a possible treatment for Duchenne muscular dystrophy (DMD). However, it is very difficult to find patients that are applicable for this therapy, because: (1) only 5-13% of DMD patients have nonsense mutations in the dystrophin gene, (2) it is challenging to find nonsense mutations in the gene because dystrophin cDNA is very long (14 kb), and (3) the efficiency of aminoglycoside-induced read-through is dependent on the kind of nonsense mutation. In order to develop a system for identifying candidates that qualify for aminoglycoside therapy, fibroblasts from nine DMD patients with nonsense mutation of dystrophin gene were isolated, induced to differentiate to myogenic lineage by AdMyoD, and exposed with gentamicin. The dystrophin expression in gentamicin-exposed myotubes was monitored by in vitro dystrophin staining and western blotting analysis. The results showed that gentamicin was able to induce dystrophin expression in the differentiated myotubes by the read-through of the nonsense mutation TGA in the gene; a read-through of the nonsense mutations TAA and TAG did not occur and consequently did not lead to dystrophin expression. Therefore, it is speculated that the aminoglycoside treatment is far more effective for DMD patients that have nonsense mutation TGA than for patients that have nonsense mutation TAA and TAG. In this study, we introduce an easy system to identify patients for this therapy and report for the first time, that dystrophin expression was detected in myotubes of DMD patients using gentamicin.
authors.affiliation: Department of Child Development, Kumamoto University School of Medicine, 1-1-1 Honjou, Kumamoto 860-0811, Japan. kimusige@kaiju.medic.kumamoto-u.ac.jp
authors.forename: Shigemi
authors.initials: S
authors.lastname: Kimura
authors.forename: Kaori
authors.initials: K
authors.lastname: Ito
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Miyagi
authors.forename: Takashi
authors.initials: T
authors.lastname: Hiranuma
authors.forename: Kowasi
authors.initials: K
authors.lastname: Yoshioka
authors.forename: Shirou
authors.initials: S
authors.lastname: Ozasa
authors.forename: Makoto
authors.initials: M
authors.lastname: Matsukura
authors.forename: Makoto
authors.initials: M
authors.lastname: Ikezawa
authors.forename: Masafumi
authors.initials: M
authors.lastname: Matsuo
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Takeshima
authors.forename: Teruhisa
authors.initials: T
authors.lastname: Miike
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D015153:Blotting, Western; D001921:Brain; D002478:Cells, Cultured; D018389:Codon, Nonsense; D018388:Codon, Terminator; D016189:Dystrophin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D015870:Gene Expression / Q000187:drug effects; D005820:Genetic Testing / Q000379:methods*; D005839:Gentamicins / Q000494:pharmacology*; D006801:Humans; D007536:Isomerism; D018482:Muscle, Skeletal; D020388:Muscular Dystrophy, Duchenne / Q000188:drug therapy* / Q000235:genetics*; D011689:Purkinje Cells
chemical_list: D000900:Anti-Bacterial Agents; D018389:Codon, Nonsense; D018388:Codon, Terminator; C484258:DMD protein, human; D016189:Dystrophin; D005839:Gentamicins