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 6

AAV1.Follistatin increases TA muscle mass and absolute force in the presence of DUX4 expression.

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AAV1.Follistatin increases TA muscle mass and absolute force in the pres...
(A) Six- to 7-week-old TIC-DUX4 and control mice were i.m. injected with 1 × 1011 particles of AAV1.Follistatin. One week later, mice were treated with 5 mg/kg tamoxifen dosed 1×/week for 8 weeks. AAV1.Follistatin-injected TA muscles from tamoxifen-treated TIC-DUX4 were overtly larger than saline-injected TIC-DUX4 controls and WT muscles. Image magnification 4×. (B) H&E-stained cryosections show evidence of histopathology (myofibers with central nuclei) in tamoxifen-treated TIC-DUX4 TA muscles expressing follistatin and saline injected TIC-DUX4 controls. Tamoxifen-treated WT controls injected with saline appeared normal. Scale bar: 100μm. (C) Follistatin treatment significantly increased TA muscle weight compared with untreated controls (1-way ANOVA with Tukey’s multiple comparisons test; P < 0.001). Mean TA muscle weights: Follistatin-treated, induced TIC-DUX4 mice, 81 mg; saline-treated induced TIC-DUX4 mice, 29 mg; uninduced TIC-DUX4 mice, 37 mg; age-matched WT, 50 mg. n = 14 AAV1.Follistatin-treated, tamoxifen-induced TIC-DUX4 muscles; 10 saline-treated, tamoxifen-induced TIC-DUX4 muscles; 12 uninduced TIC-DUX4 muscles; 9 untreated C57/BL6 controls. (D) TA absolute force was significantly increased in induced TIC-DUX4 mice treated with AAV1.Follistatin compared with induced, saline-treated TIC-DUX4 controls (1-way ANOVA with Tukey’s multiple comparisons test; P < 0.0001). Mean TA absolute force: Follistatin-treated, induced TIC-DUX4 mice, 1,558 mN; saline-treated induced TIC-DUX4 mice, 601.3 mN; uninduced TIC-DUX4 mice, 1,349 mN; age-matched WT, 1,148 mN. (E) Specific force was determined by normalizing absolute force to muscle cross-sectional area. Follistatin treatment in induced TIC-DUX4 mice did not significantly improve specific force compared with all groups (1-way ANOVA with Tukey’s multiple comparisons test). n = 6 AAV1.Follistatin-treated, tamoxifen-induced TIC-DUX4 muscles; 5 saline-treated, tamoxifen-induced TIC-DUX4 muscles; 7 uninduced TIC-DUX4 muscles; 8 untreated WT controls.