Abstract:Myofibrillar protein catabolic rate was calculated in 50 young patients with Duchenne muscular dystrophy from the amount of 3-methylhistidine excreted in the urine, and was found to be about seven times that of a control series, expressed as the percentage of myofibrillar protein catabolized per day. This wastage of myofibrillar protein is a consequence of Duchenne muscular dystrophy and inhibition of protein degradation appears to be one possible approach in the treatment of this disease.
“…also have shown that catabolism in DMD muscles is increased. 45 A relatively higher level of BUN is reported in mdx mice compared with WT C57BL/6 mice. 46 The reduced BUN level with no significant change in the creatinine level may result from amelioration of muscle degeneration by the treatment.…”
Antisense-mediated exon skipping, which can restore the reading frame, is a most promising therapeutic approach for Duchenne muscular dystrophy. Remaining challenges include the limited applicability to patients and unclear function of truncated dystrophin proteins. Multiexon skipping targeting exons 45–55 at the mutation hotspot of the dystrophin gene could overcome both of these challenges. Previously, we described the feasibility of exons 45–55 skipping with a cocktail of Vivo-Morpholinos in vivo; however, the long-term efficacy and safety of Vivo-Morpholinos remains to be determined. In this study, we examined the efficacy and toxicity of exons 45–55 skipping by intravenous injections of 6 mg/kg 10-Vivo-Morpholino cocktail (0.6 mg/kg each vPMO) every 2 weeks for 18 weeks to dystrophic exon-52 knockout (mdx52) mice. Systemic skipping of the entire exons 45–55 region was induced, and the Western blot analysis exhibited the restoration of 5–27% of normal levels of dystrophin protein in skeletal muscles, accompanied by improvements in histopathology and muscle strength. No obvious immune response and renal and hepatic toxicity were detected at the end-point of the treatment. We demonstrate our new regimen with the 10-Vivo-Morpholino cocktail is effective and safe for long-term repeated systemic administration in the dystrophic mouse model.
“…also have shown that catabolism in DMD muscles is increased. 45 A relatively higher level of BUN is reported in mdx mice compared with WT C57BL/6 mice. 46 The reduced BUN level with no significant change in the creatinine level may result from amelioration of muscle degeneration by the treatment.…”
Antisense-mediated exon skipping, which can restore the reading frame, is a most promising therapeutic approach for Duchenne muscular dystrophy. Remaining challenges include the limited applicability to patients and unclear function of truncated dystrophin proteins. Multiexon skipping targeting exons 45–55 at the mutation hotspot of the dystrophin gene could overcome both of these challenges. Previously, we described the feasibility of exons 45–55 skipping with a cocktail of Vivo-Morpholinos in vivo; however, the long-term efficacy and safety of Vivo-Morpholinos remains to be determined. In this study, we examined the efficacy and toxicity of exons 45–55 skipping by intravenous injections of 6 mg/kg 10-Vivo-Morpholino cocktail (0.6 mg/kg each vPMO) every 2 weeks for 18 weeks to dystrophic exon-52 knockout (mdx52) mice. Systemic skipping of the entire exons 45–55 region was induced, and the Western blot analysis exhibited the restoration of 5–27% of normal levels of dystrophin protein in skeletal muscles, accompanied by improvements in histopathology and muscle strength. No obvious immune response and renal and hepatic toxicity were detected at the end-point of the treatment. We demonstrate our new regimen with the 10-Vivo-Morpholino cocktail is effective and safe for long-term repeated systemic administration in the dystrophic mouse model.
“…Moreover, 8 weeks after the treatment, when the estimated level of dystrophin-positive fibres was around 10%, CK levels were higher in treated than in non-treated mdx mice. Different behaviour of the MYOM3 fragments and CK after partial restoration of dystrophin expression may reflect the capacity of these biomarkers to differentially reveal intracellular process such as microparticle turnover ( 44 ) or increased myofibrillar protein catabolism ( 45 , 46 ). Figure 7.…”
Therapy-responsive biomarkers are an important and unmet need in the muscular dystrophy field where new treatments are currently in clinical trials. By using a comprehensive high-resolution mass spectrometry approach and western blot validation, we found that two fragments of the myofibrillar structural protein myomesin-3 (MYOM3) are abnormally present in sera of Duchenne muscular dystrophy (DMD) patients, limb-girdle muscular dystrophy type 2D (LGMD2D) and their respective animal models. Levels of MYOM3 fragments were assayed in therapeutic model systems: (1) restoration of dystrophin expression by antisense oligonucleotide-mediated exon-skipping in mdx mice and (2) stable restoration of α-sarcoglycan expression in KO-SGCA mice by systemic injection of a viral vector. Following administration of the therapeutic agents MYOM3 was restored toward wild-type levels. In the LGMD model, where different doses of vector were used, MYOM3 restoration was dose-dependent. MYOM3 fragments showed lower inter-individual variability compared with the commonly used creatine kinase assay, and correlated better with the restoration of the dystrophin-associated protein complex and muscle force. These data suggest that the MYOM3 fragments hold promise for minimally invasive assessment of experimental therapies for DMD and other neuromuscular disorders.
Muscle wasting in myotonic dystrophy may result from decreased muscle anabolic processes rather than from increased catabolism. Male patients with myotonic dystrophy often have low levels of circulating androgens, and androgen administration has been shown to increase their muscle mass. We have studied the effect of testosterone enanthate administration (3 mg/kg weekly for 3 months) on muscle and whole body protein synthesis in 6 male patients with myotonic dystrophy. Muscle protein synthesis was estimated from the rate of isotope incorporation into muscle protein obtained by quadriceps muscle biopsy during a primed continuous infusion of L-[1-13C]leucine. Testosterone administration resulted in a significant increase in muscle protein synthesis in all patients. Whole body protein synthesis did not increase, indicating that protein synthesis in other tissues may have declined. Muscle ribonucleic acid content rose significantly in response to testosterone administration, suggesting that testosterone initiated its effect by hormone receptor interaction with muscle nuclei.
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