Project Abstract: Pompe disease is a devastating myopathy resulting from the deficiency of the lysosomal enzyme acid alpha- glucosidase (GAA),resulting in accumulation of glycogen in lysosomes and cytoplasm. The standard of care is enzyme replacement therapy (ERT) with recombinant human (rh) GAA. Patients with Pompe disease often have poor outcomes due to limited efficacy of the drug in clearing muscle glycogen stores. Substrate Reduction Therapy (SRT) is effective in other lysosomal storage diseases such as Gaucher and Fabry disease. We propose that by knocking down glycogen synthase (GSY1), we will reduce glycogen synthesis in muscle, thus reducing the amount of substrate requiring to be cleared by enzyme. Our long-term goal is to develop a potent therapy to stop/reduce the progression of the muscle disease in Pompe disease. Major gaps: Despite ERT, patients continue to accumulate glycogen in lysosomes leading to muscle fibrosis and fatty replacement. Effective therapy for the progressive muscle weakness of Pompe disease is thus a huge unmet need. Preliminary results: Antisense Oligonucleotides (ASO) technology has emerged as a powerful direct therapeutic alternative to conventional small molecule approaches or gene replacement strategies for the treatment of genetic disorders such as spinal muscular atrophy. ASOs are short, synthetic single-stranded DNA sequences designed to target RNA by well-characterized Watson-Crick base pairing, and once bound to the target RNA, can modulate RNA function through a degradative mechanism using RnaseH1. Ionis Pharmaceuticals designed and screened ASOs targeting mouse GYS1 in vitro to identify the most efficacious ASO. The lead ASOs from the screen were validated in a dose response study and the top 10 ASOs were screened for testing in wild type mice. Three GYS1 ASOs showed the best tolerability and efficacy profile leading to knock down of GYS1 mRNA by approximately 50% of control. Preclinical studies of the effects of the GYS1 ASOs 25 mg/kg/weekly on muscle glycogen, histology, and muscle function in 1 month and 3-month-old Pompe mice indicated that GYS1 ASOs were effective in knocking down GYS1 by approximately 85%, with approximately 50% improvement in clearing muscle glycogen levels. We therefore believe that early reduction of glycogen substrate or a combination therapy with ERT may be the key to successful treatment in Pompe disease. Hypothesis: We propose that GYS1 ASOs in Pompe mice will reduce glycogen production and prevent muscle weakness, either as monotherapy if started early, or in combination with ERT, to obtain maximum therapeutic benefit. Specific Aims: Aim 1: Identification of the most effective ASO-mediated glycogen synthase (GYS1) knock down to prevent glycogen accumulation in a mouse model of Pompe disease. Aim 2: Correction of muscle glycogen and weakness using GYS1 antisense oligonucleotides in combination with Enzyme Replacement Therapy (ERT) in Pompe mice. Success with this translati...