Fibroblasts-dependent maturation and phenotype exacerbation of dystrophic hiPSC-derived MYOtissues enables muscle strength evaluation for gene therapy screening

Abstract

SUMMARY The yet incurable Duchenne muscular dystrophy (DMD) is caused by the absence of dystrophin, a protein essential to preserve muscle integrity continuously challenged by contractions. Adeno- associated virus (AAV) delivery of truncated forms of dystrophin is currently the most promising therapeutic approach. However, patient outcomes differed from animal studies, emphasizing the necessity for models predictive of human response. Here, we describe the generation of MYOrganoids, a 3D muscle platform derived from human induced pluripotent stem cells (iPSC), whose structural and functional maturation is enhanced by fibroblasts incorporation. Importantly, a pro-fibrotic microenvironment reproduced by incorporation of dystrophic fibroblasts, was pivotal to exacerbate muscle force loss and fatiguability of DMD MYOrganoids, enabling their use as therapeutic readouts. Remarkably, efficient gene transfer of the gold standard microdystrophin in DMD MYOrganoids, failed to fully restore membrane dystroglycan components and partially rescued muscle strength, in line with the marginal correction of the DMD transcriptional signature achieved. This study highlights the potential of human MYOrganoids to unravel the limitations of current treatments under aggravated conditions and accelerate the discovery of more effective strategies.