Preclinical assessment of GNEwt/bi-shRNA-GNEM743T lipoplex product development for GNE myopathy
Le résumé fourni par la source
GNE myopathy is a heredity disease of unmet medical need associated with progressive skeletal muscle wasting, atrophy and weakness caused by mutations in the GNE gene. GNE plays a pivotal role in sialic acid production. Sialic acid is a critical part of glycoprotein, ganglioside and glycolipid cell-cell interaction which is necessary for normal skeletal muscle function. Previously we demonstrated safety and efficacy of the GNEwt gene lipoplex in one patient. We engineered GNEM743T and dual function GNEwt/bi-shRNA plasmids to evaluate GNEM743T specific knockdown and GNEwt expression. Knockdown efficiency, protein expression, and functional rescue were assessed. A dose range study in mice quantified plasmid delivery and human GNE expression in muscle. We demonstrate effective plasmid function via knockdown of the GNEM743T gene mutation and concurrent expression of GNEwt gene in a dose dependent manner. Similar in vitro increase in sialic acid production is shown between prior single function plasmid and GNEwt/bi-shRNA-GNEM743T dual function plasmid. Moreover, we demonstrate murine in vivo muscle delivery and expression of GNEwt mRNA from the GNEwt/bi-shRNA-GNEM743T plasmid delivered via DOTAP-Cholesterol lipoplex following intravenous injection. These results encourage future studies, potentially leading toward clinical testing of GNEwt/bi-shRNA-GNEM743T lipoplex for GNE myopathy. GNEwt/bi-shRNA-GNEM743T dual function plasmid expresses GNE protein and knocks down GNEM743T.Transfection with GNEwt/bi-shRNA-GNEM743T results in increased sialic acid expression in vitro.GNEwt/bi-shRNA-GNEM743T plasmid demonstrates delivery and expression in murine model. GNEwt/bi-shRNA-GNEM743T dual function plasmid expresses GNE protein and knocks down GNEM743T. Transfection with GNEwt/bi-shRNA-GNEM743T results in increased sialic acid expression in vitro. GNEwt/bi-shRNA-GNEM743T plasmid demonstrates delivery and expression in murine model.
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