title: Developing gene therapy for Duchenne muscular dystrophy using adeno associated virus (AAV) expressing Estrogen related receptor gamma (ERRγ)
Creators: name: Al-Siyabi, Muzna
Publisher: name: University of Reading
Year: 2018

description: Skeletal muscle is the most abundant tissue in the body, exhibiting major metabolic activity by contributing up to 40% of the resting metabolic rate in adults. One of the most remarkable trait of skeletal muscle is its great adaptability to numerous environmental and physiological challenges by changing its phenotype profile in terms of size and composition that are brought about by changes in gene expression, biochemical and metabolic properties. Amongst these adaptations is the increase in oxidative metabolism that is supported by increase in blood flow and capillary density. Estrogen related receptor gamma (ERRγ) belongs to a family of orphan nuclear receptors. It is considered to be a master switch for both oxidative and angiogenic factors. ERRγ is down-regulated in mdx along with its target metabolic and angiogenic genes. Transgenic overexpressing ERRγ in mdx mice, improve sarcolemmal integrity and muscle perfusion with restoration of metabolic and angiogenic genes. Duchenne muscular dystrophy (DMD) is an X-linked, recessive neuromuscular disorder caused by the loss of dystrophin that causes progressive loss of muscle fibre leading to cardiac and respiratory failure and ultimately death within the third decade. It is characterized by sarcolemmal fragility, impaired blood perfusion, calcium dysregulation, impaired mitochondrial function and oxidative stress. Following transcriptomic analysis we hypothesize that postnatal over-expression of ERRγ might improve oxidative capacity and angiogenesis in mdx mice. We first examined the effect of intramuscular administration of (5X1010 vg) AAV8-ERRγ into tibialis anterior (TA) muscle of two cohorts of mdx male mice, initiating the experiment at either 6 or 12 weeks of age. There was no difference in mass, cross sectional area of muscle fibres nor was there a difference in myosin heavy chain fibre typing in the TAs of either cohort; however, succinate dehydrogenase analysis (SDH) was significantly increased in both cohorts and H&amp;E analysis demonstrated a 10% reduction of centrally nucleated fibres, but this reduction was restricted to the 6 week-old cohort only. This gave proof of principle data of an improvement in metabolic and pathological parameters in dystrophic muscle following ERRγ over-expression and leading to systemic administration protocols. Surprisingly, systemic administration of (1x1012 vg) of AAV8-ERRγ into 6 week-old mdx male mice showed no functional improvement of EDL muscle and no difference in any examined markers. Therefore, we carried out an earlier systemic administration of (2 X1012 vg) of AAV8-ERRγ into three week-old mdx mice for longer time and resulted in improved specific force of EDL muscle while the eccentric contraction-induced force deficit was unaffected. EDL muscles showed improvements in SDH activity, a reduction in centrally nucleated fibres, with no change in IgG positive fibres or CK assay. EDL muscles showed an increase in capillary density, increased transcript levels of PGC-1α, angiogenesis, inflammation markers and a reduction in transcript levels of GADD34 and 4EBP1. Collectively, this data provides prima facia evidence that ERRγ over-expression at an early time point is a potential therapeutic strategy to overcome metabolic and pathological problems associated with dystrophic skeletal muscles.