The Muscle Xenograft Model of Facioscapulohumeral Muscular Dystrophy: Development, Optimization, and Novel Applications

Abstract

Aberrant expression of DUX4 in human muscle causes Facioscapulohumeral Muscular dystrophy (FSHD) which affects about 1 in 8,333 individuals worldwide, yet the mechanism by which DUX4 causes muscle wasting is unknown. The DUX4 gene is unique to humans and transgenic animal models have largely failed to exhibit the dystrophic phenotype and endogenous molecular changes characteristic of FSHD. Therefore, studies of DUX4 signaling in human muscle have been limited to patient biopsies and cultures of myogenic cells in vitro. There are no therapies that target the mechanism of disease, thus there is a pressing need for studies of DUX4 in mature human muscle. We have developed a method to xenograft human-derived muscle precursor cells, isolated from patients with FSHD and controls, into the tibialis anterior of immune-deficient mice to form xenografts of pure human muscle tissue. The FSHD xenografts are robust, mature, and well organized. Human myofibers are innervated and associate with human satellite cells, making the xenografts structurally comparable to intact human skeletal muscle. The FSHD but not control xenografts express DUX4 and DUX4 gene targets and have 4q35 methylation profiles typical of FSHD. The FSHD grafts also display a novel biomarker of FSHD, SLC34A2, measurable by immunofluorescence, which will provide a quantifiable metric for future therapeutic studies. We have described several modifications to the engraftment strategy that can be used to answer complex mechanistic and functional questions regarding the FSHD pathophysiology. Finally, we report promising data from a collaborative study with Fulcrum Therapeutics in which we were able to repress the DUX4-signaling pathway by administering a targeted small molecule therapy to FSHD grafts. Ours is the first scalable and reproducible in vivo model of FSHD muscle. Future studies will include those aimed at continuing to delineate the molecular pathogenesis of FSHD, performing functional tests to define the pathophysiology of FSHD, and testing new and exciting therapeutic strategies aimed at reducing the DUX4 program in human FSHD xenografts.