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Updated: Jan 3, 2026
Adeno-Associated Virus-Mediated Delivery of CRISPR for Cardiac Gene Editing in Mice
Published on: August 2, 2018
Titin splicing regulates cardiotoxicity associated with calpain 3 gene therapy for limb-girdle muscular dystrophy
William Lostal1, Carinne Roudaut1, Marine Faivre1
1Généthon INSERM, U951, INTEGRARE Research Unit, Evry F-91002, France.
Abstract:
Limb-girdle muscular dystrophy type 2A (LGMD2A or LGMDR1) is a neuromuscular disorder caused by mutations in the calpain 3 gene (CAPN3). Previous experiments using adeno-associated viral (AAV) vector-mediated calpain 3 gene transfer in mice indicated cardiac toxicity associated with the ectopic expression of the calpain 3 transgene. Here, we performed a preliminary dose study in a severe double-knockout mouse model deficient in calpain 3 and dysferlin. We evaluated safety and biodistribution of AAV9-desmin-hCAPN3 vector administration to nonhuman primates (NHPs) with a dose of 3 × 1013 viral genomes/kg. Vector administration did not lead to observable adverse effects or to detectable toxicity in NHP. Of note, the transgene expression did not produce any abnormal changes in cardiac morphology or function of injected animals while reaching therapeutic expression in skeletal muscle. Additional investigation on the underlying causes of cardiac toxicity observed after gene transfer in mice and the role of titin in this phenomenon suggest species-specific titin splicing. Mice have a reduced capacity for buffering calpain 3 activity compared to NHPs and humans. Our studies highlight a complex interplay between calpain 3 and titin binding sites and demonstrate an effective and safe profile for systemic calpain 3 vector delivery in NHP, providing critical support for the clinical potential of calpain 3 gene therapy in humans.
Insights
Calpain 3 gene therapy shows promise for limb-girdle muscular dystrophy. Adeno-associated viral vector delivery in nonhuman primates was safe and effective, unlike in mice, suggesting species-specific differences.
Area of Science:
- Molecular Biology
- Genetics
- Neuromuscular Disorders
Background:
- Limb-girdle muscular dystrophy type 2A (LGMD2A) is caused by mutations in the calpain 3 gene (CAPN3).
- Previous AAV-mediated CAPN3 gene transfer in mice showed cardiac toxicity.
- Species-specific differences in CAPN3 buffering capacity and titin splicing may explain toxicity in mice.
Purpose of the Study:
- To evaluate the safety and biodistribution of AAV9-desmin-hCAPN3 vector in nonhuman primates (NHPs).
- To investigate potential cardiac toxicity associated with CAPN3 gene therapy.
- To support the clinical potential of CAPN3 gene therapy for LGMD2A.
Main Methods:
- A preliminary dose study using AAV9-desmin-hCAPN3 vector administration in a severe double-knockout mouse model.
- Safety and biodistribution assessment of AAV9-desmin-hCAPN3 vector (3 × 10^13 viral genomes/kg) in NHPs.
- Evaluation of cardiac morphology, function, and transgene expression in skeletal muscle.
Main Results:
- AAV9-desmin-hCAPN3 vector administration in NHPs showed no observable adverse effects or detectable toxicity.
- Transgene expression in NHPs did not cause abnormal cardiac changes, with therapeutic levels achieved in skeletal muscle.
- Mice exhibit reduced capacity for buffering calpain 3 activity compared to NHPs and humans.
Conclusions:
- Systemic delivery of CAPN3 via AAV vector is safe and effective in NHPs.
- Species-specific titin splicing and buffering capacity influence CAPN3 toxicity.
- These findings provide critical support for the clinical application of CAPN3 gene therapy in human LGMD2A.
