GYS1 Antisense Therapy Prevents Disease-Driving Aggregates and Epileptiform Discharges in a Lafora Disease Mouse Model

Neurology Biochemistry Basic Medical Sciences Life Sciences Immunology Medicine Cell Biology
GYS1 antisense oligonucleotide epilepsy Lafora disease glycogen synthesis neuroinflammation neurodegeneration

GYS1 Antisense Therapy Inhibits Pathogenic Aggregates and Epileptiform Discharges in a Mouse Model of Lafora Disease

Background and Objectives

Lafora disease (LD) is a devastating autosomal recessive genetic disorder characterized by epilepsy and rapidly progressive dementia in adolescence. The disease primarily involves mutations in the EPM2A or EPM2B genes, encoding the glycogen phosphatase laforin and the E3 ubiquitin ligase malin, respectively. These enzymes are crucial in regulating glycogen storage throughout the body. Loss of their function leads to the accumulation of abnormal glycogen molecules, termed Lafora bodies (LBs), particularly in the brain. These aggregates cause seizures and neurodegeneration. Currently, there is no effective treatment, and patients typically succumb to the disease within 11 years of onset.

Recent research demonstrated that reducing glycogen synthesis can effectively decrease LB accumulation and associated pathogenic effects in mouse models. This study focuses on the development and testing of an antisense oligonucleotide (ASO) targeting glycogen synthase 1 (GYS1), a key enzyme in glycogen synthesis, to assess its therapeutic efficacy in an EPM2B knockout (KO) mouse model.

Research Team and Publication Source

The study was conducted by a collaborative team from various institutions, including the Department of Molecular and Cellular Biochemistry at the University of Kentucky, Ionis Pharmaceuticals’ Antisense Drug Discovery Department, and the Department of Neuroscience at the University of Florida. The findings were published in Neurotherapeutics (2023, Volume 20), marking significant progress in LD research.

Methods and Procedures

The study employed an EPM2B KO mouse model to investigate the distribution and efficacy of GYS1-ASO therapy. Key experimental steps included:

  1. ASO Administration: Mice received stereotactic intracerebroventricular injections of GYS1-ASO at 4, 7, and 10 months of age. Each group comprised 10 mice (5 males and 5 females). Mice were sacrificed at 13 months for tissue analysis.

  2. mRNA and Protein Expression Analysis: Brain tissue RNA was extracted for real-time quantitative PCR (qRT-PCR) to measure GYS1 mRNA levels. Protein expression was assessed via Western blotting and immunohistochemistry.

  3. Glycogen Accumulation Analysis: LB and glycogen levels were evaluated using periodic acid-Schiff (PAS) staining, glycogen-specific immunohistochemistry, and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry.

  4. Neuronal Excitability Testing: Multiple electrode arrays (MEA) recorded spontaneous firing (SF) rates and epileptiform discharge (ED) events to assess neuronal excitability.

  5. Statistical Analysis: Data were analyzed using ANOVA and mixed-effects models to compare treatment groups.

Key Findings

  1. Significant Reduction in GYS1 Expression: GYS1-ASO treatment significantly reduced GYS1 mRNA and protein levels. Immunohistochemistry revealed decreased GYS1 expression across all brain regions.

  2. Reduced Glycogen Aggregation: GYS1-ASO markedly decreased glycogen accumulation and LB formation, particularly in the hippocampus and cerebellum.

  3. Decreased Epileptiform Discharges: MEA recordings showed significantly fewer ED events in GYS1-ASO-treated mice compared to controls, with discharge frequencies approaching normal levels.

  4. No Significant Impact on Neuroinflammation: GYS1-ASO did not significantly alter levels of neuroinflammatory markers (e.g., CCL5, CXCL10), likely due to the late treatment start.

Significance and Implications

  1. Innovative Therapeutic Strategy: The study provides the first evidence that targeting glycogen synthesis with an ASO can effectively slow disease progression in a mouse model of LD, paving the way for potential human application.

  2. Broad Applicability: GYS1-ASO proved effective in both EPM2A and EPM2B mutation models, indicating its potential utility across all LD patients.

  3. Combination Therapy Potential: Although GYS1-ASO does not clear existing LBs, it can be combined with other therapies targeting LB degradation for synergistic effects.

  4. Safety Profile: Reducing glycogen synthesis by approximately 50% was well-tolerated in healthy mice, supporting clinical translation.

Highlights of the Study

  • GYS1-ASO achieved broad brain distribution and significantly suppressed glycogen synthase expression.
  • The therapy reduced glycogen accumulation and normalized neuronal excitability, validated through multiple experimental methods.
  • This study provides critical proof-of-concept data for ASO-based treatments for LD.

Future Directions

Further research could optimize treatment timing and evaluate GYS1-ASO’s effects on peripheral tissues. Combining GYS1-ASO with LB-degrading therapies could provide comprehensive disease management. These findings offer new hope for LD treatment and potential insights for addressing other glycogen storage disorders.