148 The Use of Human iPSC Derived Cardiomyocytes for Systematic Comparison of Cardiac Transduction Efficiency of AAV Serotypes Molecular Therapy Volume 22, Supplement 1, May 2014 Copyright © The Ameri[.]
CELL AND GENE THERAPY FOR MYOGENIC DISORDERS Phosphorodiamidate Morpholino Oligonucleotide Improves Systemic Delivery and Dystrophin Restoration in mdx Mice Jason D Tucker,1 Mingxing Wang,1 Bo Wu,1 Pei Lu,1 Qi Lu.1 Neurology, Carolinas Healthcare System, Charlotte, NC The stability and low toxicity of phosphorodiamidate morpoholino oligonucleotide (PMO), one of the most promising antisense oligonucleotide chemistry currently available, with studies in animal models and current clinical trials have demonstrated clear benefits to subjects, a rare success in pharmaco-genetic interventions However, low efficiency, and non-specific delivery remain critical barriers of oligonucleotide therapy in DMD and other diseases To overcome these critical barriers, we have developed new polymers toward more effective delivery systems without increased toxicity Synthesized poly(ester amine)s (PEAs) through cross-linking low molecular weight polyethylenimine and tris[2-(acryloyloxy)ethyl] isocyanurate have been previously demonstrated as effective gene and oligonucleotide delivery systems in vitro as well as in vivo These polymers effectively condense with anionic cargo to particles resistant to physiological conditions sufficient to improve transfection and reporter gene expression upon intramuscular administration as compared to pDNA alone but also PEI 25k The aim of this project has been to evaluate conjugation of these new PEA polymers to directly impart cationic charge to otherwise charge-neutral PMO toward more effective delivery while attempting to address inherent toxicities of cationic oligonucleotide conjugates Our results demonstrate the potential for synthetic polymer conjugates on in vitro transfection, as well as intramuscular and systemic delivery of antisense PMO for the restoration of dystrophin expression in the mdx mouse 147 Parathyroid Hormone Administration and Parathyroid Hormone Type Receptor Activation Accelerate Muscle Differentiation and Muscle Regeneration in Muscular Dystrophy Shigemi Kimura,1 Kowasi Yoshioka.1 Department of Child Development, Kumamoto University Graduate School, Kumamoto, Japan We have established genetically engineered embryonic stem cells (ZHTc6-MyoD) that harbor a tetracycline-regulated expression vector encoding myogenic transcriptional factor MyoD After removal of tetracycline, almost all of the cells differentiated into myotubes, but a few cells later re-formed colonies DNA microarray analysis showed that the expression of parathyroid hormone type receptor (PTH1R) in the colonies that differentiated into muscle lineage cells is higher in the colonies of undifferentiated ZHTc6-MyoD cells In addition to the PTH1R required for the muscle differentiation of ZHTc6-MyoD cells, parathyroid hormone (1-34) [PTH (1-34)] further accelerated muscle differentiation In human and mouse skeletal muscle tissue, most PTH1R-positive cells also expressed Pax7 and CD34, indicating that these cells originated from satellite cells The administration of 60 μg/kg per day of PTH (1-34) to mdx mice (dystrophin-deficient mice) was started at weeks of age The four limbs hanging wire test was performed on day 10 The mean hang time of PTH-treated mdx mice (n=6) was 132±61.6 sec In contrast, the mean hang time of saline-injected mdx mice (n=7) was significantly shorter, at 49.5±44.9 sec (P=0.03) The treadmill test was performed on day 43 Mice were forced to run on a treadmill with the belt parallel to the floor at a speed of 25 cm/sec for 100 The distance that they ran was recorded and compared All C57BL/10 mice (n=6) ran for 100 min, and the mean distance was 1486±7.4 m PHT-treated mdx (n=6) mice ran a mean distance of 843.6±352 m, although one of the PHT-treated mice did run for 100 None of the saline-injected mdx mice (n=7) ran for 100 min, and the mean Molecular Therapy Volume 22, Supplement 1, May 2014 Copyright © The American Society of Gene & Cell Therapy distance covered was 425±190 m The C57BL/10 mice (normal mice) ran significantly longer than the saline-injected mdx mice (P