Résumé :
|
Glycogen storage disease type II (GSDII) or Pompe disease is an autosomal recessive disorder caused by defects in the lysosomal acid alpha-glucosidase (GAA) gene. It is characterized by glycogen accumulation, especially in skeletal muscle and heart, leading to death in the infantile form. Enzyme replacement therapy (ERT) has recently demonstrated its efficacy on motor strength and cardiac function. However, it has been shown that type II muscle fibers are more resistant to therapy, a phenomenon probably due to large autophagic areas and altered intracellular traffic of the recombinant enzyme. Therefore, our aim is to develop a novel therapeutic approach based on the reduction of glycogen loading, especially in type II fibers. Small interfering RNAs targeted to the two major genes for glycogen synthesis (glycogenin and glycogen synthase) were designed and selected on C2C12 cells. Two lentiviral vectors (siRNA/GN2 and siRNA/GYS2) were tested on primary myoblasts from GSDII mice showing a significant decrease in GN/GYS expression and in glycogen synthesis (80-90% in differentiated cells). Enlarged lysosomes were not found in siRNA/GN or siRNA/GYS cells compared to non-transduced cells. AAV/siRNA/GYS was then constructed and administered in GSDII mice to perform a conditional disruption of the glycogen synthase. A single intramuscular injection induced a reduction in GYS mRNA expression (50%) and glycogen accumulation in the injected gastrocnemius compared to the contralateral muscle. In parallel, a double knock-out (lacking GYS and GAA) was created and used to determine the long-term effects of a permanent glycogen synthase defect. In this model, glycogen accumulation and autophagosomes were not detected in muscle. These results suggest that substrate reduction could be a potential therapeutic strategy in GSDII in association with ERT especially in type II fibers.
|