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Adeno-Associated Virus 8 and 9 Myofibre Type/Size Tropism Profiling Reveals Therapeutic Effect of Microdystrophin in
Matthew J Burke1, Braiden M Blatt1,2, James A Teixeira1
1Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, Missouri, USA.
Background:
Adeno-associated virus (AAV) 8 and 9 are in clinical trials for treating neuromuscular diseases such as Duchenne muscular dystrophy (DMD). Muscle consists of myofibres of different types and sizes. However, little is known about the fibre type and fibre size tropism of AAV in large mammals.
Methods:
We evaluated fibre type- and size-specific transduction properties of AAV8 and AAV9 in 17 dogs that received systemic gene transfer (dose 1.94 ± 0.52 × 1014 vg/kg; injected at 2.86 ± 0.30 months; harvested at 20.79 ± 3.30 months). For AAV8, two DMD dogs and three carrier dogs received an alkaline phosphatase (AP) reporter vector, and five DMD dogs received a four-repeat microdystrophin (uDys) vector. For AAV9, one normal and one DMD dog received the AP vector, and five DMD dogs received a five-repeat uDys vector. Association between AAV transduction and the fibre type/size was studied in three muscles that showed mosaic transgene expression, including the biceps femoris, teres major and latissimus dorsi.
Results:
Transgene expression was detected in 30%-45% of myofibres. In the AP reporter vector-injected dogs, neither AAV8 nor AAV9 showed a statistically significant fibre type preference. Interestingly, AP expression was enriched in smaller fibres. In uDys-treated DMD dogs, slow and fast myofibres were equally transduced. Notably, uDys-expressing myofibres were significantly larger than uDys-negative myofibres irrespective of the AAV serotype (p < 0.0001). In AAV8 uDys vector-injected dogs, the mini-Feret diameter was 15%, 16% and 23% larger in uDys-positive slow, fast and hybrid fibres, respectively; the cross-sectional area was 30%, 34% and 46% larger in uDys-positive slow, fast and hybrid fibres, respectively. In AAV9 uDys vector-injected dogs, the mini-Feret diameter was 12%, 13% and 25% larger in uDys-positive slow, fast and hybrid fibres, respectively; the cross-sectional area was 25%, 28% and 59% larger in uDys-positive slow, fast and hybrid fibres, respectively.
Conclusions:
Our studies suggest that AAV8 and AAV9 transduce fast and slow myofibres at equivalent efficiency. Importantly, uDys therapy effectively prevented dystrophic myofibre atrophy. Our study provides important insight into systemic muscle AAV delivery in large mammals and supports further development of uDys gene therapy for DMD.
Insights
Adeno-associated virus (AAV) 8 and 9 effectively transduce muscle fibers without preference for type but show enrichment in smaller fibers. Micro-dystrophin therapy prevents muscle fiber atrophy in Duchenne muscular dystrophy (DMD) models.
Area of Science:
- Gene therapy
- Molecular and cellular biology
- Neuromuscular disease research
Background:
- Adeno-associated virus (AAV) serotypes 8 and 9 are investigated for treating neuromuscular diseases like Duchenne muscular dystrophy (DMD).
- Understanding AAV tropism for different muscle fiber types and sizes in large mammals is crucial for effective gene therapy.
- Limited data exists on AAV tropism in large animal models, hindering translation to human therapies.
Purpose of the Study:
- To evaluate the fiber type and size tropism of AAV8 and AAV9 in large mammals.
- To assess the efficacy of micro-dystrophin (uDys) gene therapy in preventing muscle fiber atrophy in a Duchenne muscular dystrophy (DMD) model.
- To provide insights into systemic AAV delivery for neuromuscular diseases.
Main Methods:
- Systemic gene transfer of AAV8 and AAV9 vectors carrying reporter genes or micro-dystrophin (uDys) was performed in 17 dogs.
- Muscle samples were analyzed for transgene expression, fiber type (slow, fast, hybrid), and fiber size (mini-Feret diameter, cross-sectional area).
- Statistical analysis was used to determine AAV tropism and the impact of uDys therapy on myofiber size.
Main Results:
- AAV8 and AAV9 demonstrated no significant preference for transducing slow or fast myofibers.
- Reporter gene expression was enriched in smaller myofibers.
- Micro-dystrophin (uDys) expressing myofibers were significantly larger than uDys-negative myofibers, indicating prevention of atrophy, irrespective of AAV serotype.
Conclusions:
- AAV8 and AAV9 exhibit equivalent transduction efficiency across slow and fast myofibers in large mammals.
- Micro-dystrophin gene therapy effectively counteracts myofiber atrophy in a Duchenne muscular dystrophy model.
- These findings support the further development of AAV-mediated micro-dystrophin gene therapy for Duchenne muscular dystrophy.
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