Development of sialic acid replacement therapy for GNE myopathy and towards further therapeutic approaches

Abstract

GNE myopathy is a progressive myopathy caused by pathogenic variants in the GNE gene encoding UDP–GlcNAc 2–epimerase/ManNAc kinase, which is responsible for the first enzymatic reaction in sialic acid biosynthesis. Patient muscles are characterized by the presence of so–called ‘rimmed vacuoles,’ which are associated with the intracellular accumulation of various insoluble proteins within the myofibres and with autophagic vacuoles surrounding them. Since the identification of the causative GNE gene mutation, questions and concerns have arisen regarding the relationship between sialic acid synthesis and the development of muscle disease. We first showed that the enzyme activities of the GNE products with the patient variants are reduced, again indicating that reduced activity of this enzyme is the pathogenic factor, and we also found reduced sialylation of the patient cells. Then we found that addition of sialic acid into the medium allowed the cells to take up sialic acid and restore cellular sialylation. Based on these findings, a mouse model was generated for preclinical studies with sialic acid compounds in vivo : Gne gene KO mouse was crossed with transgenic mouse with the human D176V mutant GNE. The given mice showed reduced sialic acid levels in all organs other than the brain from birth while also exhibiting age–related myopathic symptoms, replicating the pathology of human patients. The most difficult aspect of sialic acid supplementation studies in mice was the rapid excretion of exogenous sialic acids, which could be overcome by appropriate administration or the use of several sialic acid compounds. Administration of sialic acid compounds before the onset of the disease completely prevented the onset of the disease, and administration to the affected muscle successfully restored the muscle symptoms. These results indicate that (1) frequent administration of free sialic acid, (2) use of sialic acid compounds with a long blood duration and (3) use of a compound with a high cellular uptake capacity are effective in achieving high therapeutic effects. On the other hand, developing a treatment for recovery of affected muscle was aimed at ameliorating myofiber atrophy and inhibiting protein accumulation and inducing their degradation in muscle fibers. (1) We found that sialic acid has an antioxidant effect in the skeletal muscle of model mice and that antioxidant drugs are effective in inhibiting accumulation, and (2) chaperone–inducing drugs are effective in inducing degradation.