AAV-mediated follistatin gene therapy improves functional outcomes in the TIC-DUX4 mouse model of FSHD
Carlee R. Giesige, Lindsay M. Wallace, Kristin N. Heller, Jocelyn O. Eidahl, Nizar Y. Saad, Allison M. Fowler, Nettie K. Pyne, Mustafa Al-Kharsan, Afrooz Rashnonejad, Gholamhossein Amini Chermahini, Jacqueline S. Domire, Diana Mukweyi, Sara E. Garwick-Coppens, Susan M. Guckes, K. John McLaughlin, Kathrin Meyer, Louise R. Rodino-Klapac, Scott Q. Harper
Carlee R. Giesige, Lindsay M. Wallace, Kristin N. Heller, Jocelyn O. Eidahl, Nizar Y. Saad, Allison M. Fowler, Nettie K. Pyne, Mustafa Al-Kharsan, Afrooz Rashnonejad, Gholamhossein Amini Chermahini, Jacqueline S. Domire, Diana Mukweyi, Sara E. Garwick-Coppens, Susan M. Guckes, K. John McLaughlin, Kathrin Meyer, Louise R. Rodino-Klapac, Scott Q. Harper
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Research Article Muscle biology Therapeutics

AAV-mediated follistatin gene therapy improves functional outcomes in the TIC-DUX4 mouse model of FSHD

Abstract

Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant or digenic disorder linked to derepression of the toxic DUX4 gene in muscle. There is currently no pharmacological treatment. The emergence of DUX4 enabled development of cell and animal models that could be used for basic and translational research. Since DUX4 is toxic, animal model development has been challenging, but progress has been made, revealing that tight regulation of DUX4 expression is critical for creating viable animals that develop myopathy. Here, we report such a model — the tamoxifen-inducible FSHD mouse model called TIC-DUX4. Uninduced animals are viable, born in Mendelian ratios, and overtly indistinguishable from WT animals. Induced animals display significant DUX4-dependent myopathic phenotypes at the molecular, histological, and functional levels. To demonstrate the utility of TIC-DUX4 mice for therapeutic development, we tested a gene therapy approach aimed at improving muscle strength in DUX4-expressing muscles using adeno-associated virus serotype 1.Follistatin (AAV1.Follistatin), a natural myostatin antagonist. This strategy was not designed to modulate DUX4 but could offer a mechanism to improve muscle weakness caused by DUX4-induced damage. AAV1.Follistatin significantly increased TIC-DUX4 muscle mass and strength even in the presence of DUX4 expression, suggesting that myostatin inhibition may be a promising approach to treat FSHD-associated weakness. We conclude that TIC-DUX4 mice are a relevant model to study DUX4 toxicity and, importantly, are useful in therapeutic development studies for FSHD.

Authors

Carlee R. Giesige, Lindsay M. Wallace, Kristin N. Heller, Jocelyn O. Eidahl, Nizar Y. Saad, Allison M. Fowler, Nettie K. Pyne, Mustafa Al-Kharsan, Afrooz Rashnonejad, Gholamhossein Amini Chermahini, Jacqueline S. Domire, Diana Mukweyi, Sara E. Garwick-Coppens, Susan M. Guckes, K. John McLaughlin, Kathrin Meyer, Louise R. Rodino-Klapac, Scott Q. Harper

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Figure 5

Progressive pathology in TIC-DUX4 mice treated with a low-dose tamoxifen regimen.

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Progressive pathology in TIC-DUX4 mice treated with a low-dose tamoxifen...
Data for this figure were generated with 8- to 12-week-old mice induced with 5 mg/kg tamoxifen given 1×/week for 1–4 months, as indicated. (A) H&E-stained cryosections (20×) from indicated muscles of TIC-DUX4 mice induced for 4 weeks to 16 weeks show progressive muscle degeneration and regeneration with time. WT littermates were treated with this tamoxifen regimen for 16 weeks and show normal muscle histology. (B) Total activity and rearing behavior were measured in all mice throughout the study. Top graph shows total cage activity from tamoxifen-treated TIC-DUX4 mice and WT littermates mice, measured monthly for 3 months, where baseline represents a pretreatment data point. The bottom graph shows rearing frequency in the same cohort over the same time course. There was a significant and progressive decline in Tam-induced TIC-DUX4 mouse activity and rearing that reached significance at 1 and 2 months, compared with TIC-DUX4 baseline (2-way ANOVA repeated measures; P < 0.05). In contrast, activity and rearing behavior in WT animals was not significantly different across the time course. n = 7 animal per group. When comparing TIC-DUX4 with WT animals at each time point, there was a significant difference between the groups only at the 2 month time point, for both overall activity and rearing behavior (Tukey’s post-hoc test; P < 0.01) (C and D) Low-dose Tam treatment caused significant absolute force deficits in the TA (P < 0.0001) and gas (P < 0.01) muscles and significantly decreased specific force in the gas (P < 0.05) compared with uninduced TIC-DUX4 mice. Test results were obtained with 1-way ANOVA using Tukey’s multiple comparisons test. For TA absolute force: Induced TIC-DUX4 mice, 601.3 mN; uninduced TIC-DUX4, 1,349 mN; WT, 1,148 mN). TA specific force: Induced TIC-DUX4 mice, 212.2 mN/mm2; uninduced TIC-DUX4, 274.9 mN/mm2; WT, 259.4 mN/mm2. For gas absolute force: Induced TIC-DUX4 mice, 693.6 mN; uninduced TIC-DUX4, 2,184 mN; WT, 2,356 mN). Gas specific force: Induced TIC-DUX4 mice, 103.9 mN/mm2; uninduced TIC-DUX4, 280.9 mN/mm2; WT, 253.3 mN/mm2. For TA muscles, n = 5 tamoxifen-induced TIC-DUX4 legs, 7 uninduced TIC-DUX4 legs, and 8 WT littermate legs. For gas, n = 5 tamoxifen-induced TIC-DUX4 legs, 8 uninduced legs, and 7 WT littermate legs.