Review ArticleStem cell-based therapies for Duchenne muscular dystrophy
Section snippets
Pathological features of DMD
DMD is due to mutations in the dystrophin gene. The dystrophin gene is translated into a 427 kDa protein, which is part of the Dystrophin Glycoprotein Complex (DGC) that provides a structural and signaling link between the cytoskeleton of the muscle fiber and the extracellular matrix (Ervasti, 2007). In healthy individuals, the dystrophin protein stabilizes the plasma membrane of the striated muscle fibers. However, in patients with DMD or the allelic Becker muscular dystrophy (BMD), mutations
Skeletal muscle determination in the vertebrate embryo
Muscle commitment and differentiation are mainly controlled by the regulated spatio-temporal expression of a set of four proteins (Myf5, MyoD1, Myogenin, and Mrf4) termed the muscle regulatory factors (MRFs) (Buckingham and Rigby, 2014). The MRFs are transcription factors that drive the expression of a multitude of genes regulating establishment and maintenance of the myogenic fate.
In embryonic development, myogenic cells originate from mesodermal precursors that colonize the paraxial mesoderm
Cell therapy for DMD
Cell therapy is based on the heterologous, or autologous, transplantation of cells, with the goal of regenerating the damaged tissue or organ of the patient, and replenishing specific stem cell populations. In the case of DMD, the main goal is to reconstitute the satellite cell pool with dystrophin competent cells, and thereby restore muscle function due to the presence of dystrophin expressing muscle fibers. The source of the therapeutic cells can be healthy, histo-compatible donors, or
Pluripotent stem cells
Vertebrate pluripotent stem cells (PSCs) retain their ability to differentiate into the three germ layers of the embryo: ectoderm, mesoderm, and endoderm. Typical PSCs are the embryonic stem cells (ESCs) (Evans and Kaufman, 1981; Martin, 1981; Thomson et al., 1998), and the induced PSC (iPSCs) (Takahashi et al., 2007; Takahashi and Yamanaka, 2006).
The generation of iPSCs opened up new avenues in stem cell therapy, and solved many problems associated with use of ESCs. For example, while human
Muscle linage specification systems
One of the strategies to achieve a direct myogenic specification of PSCs is to replicate in the culture dish the inductive stimuli which underlie the muscle determination in the developing embryos. To accomplish this goal, one approach is for monolayer PSCs to be treated in vitro with the specific cytokines and growth/morphogenetic factors that orchestrate the specification of the mesoderm in vivo, the somitogenesis, and the commitment of the early muscle progenitors (Chal et al., 2015). A
Genetic engineering of hiPSCs to restore functional dystrophin expression
In order to generate dystrophin expressing muscle fibers, hiPSCs derived from DMD patients can be genetically corrected to express functional dystrophin for autologous cell replacement therapy. CRISPR-Cas9 mediated gene editing is currently being investigated as a tool to perform such correction. It involves two components: a single guide RNA (sgRNA) and the Cas9 endonuclease. Cas9 endonuclease associates with the sgRNA at the genomic target sequence to create DNA double strand breaks leading
Potential limitations of using hiPSCs for DMD treatment
The use of hiPSCs for treatment of muscular dystrophies is very promising. However, before proceeding to clinical trials, four key limitations must be overcome, and potential safety issues addressed. (i) We have to identify the patients' best somatic cell type from which to generate the hiPSCs, and we have to improve the hiPSCs muscle commitment protocols, for example by generating the myogenic cells more quickly in vitro, and by using culture media free of animal factors. (ii) We have to
Future directions of the hiPSCs therapy development for the MDs
To develop clinically applicable hiPSCs-based therapies, researchers have focused on deriving cells that have high potency in terms of regenerating and self-renew, i.e. cells that have similar features to those of adult satellite cells (Incitti et al., 2019) (Fig. 1). Based on this, progenitor cells can acquire a higher clinical potential. As discussed earlier, hiPSCs-derived myogenic progenitors have a molecular profile that is similar to fetal-stage myoblasts. Therefore, one of the most
Conclusion
Stem cells, due to their advantageous regeneration capability, bring the promise for cell transplantation therapy (Fig. 1 summary of cells that can be used or tested for stem cell-based therapies for muscular dystrophies). hiPSCs that can be derived from patients open the avenue for autologous cell therapy. With the rapid development of serum-free lineage specification protocols, expandable myogenic progenitor cells can be differentiated from hiPSCs. This population of cells has similar
Funding
This work was supported by National Institutes of Health R01AR070751 (G.L.), the Maryland Stem Cell Research Fund (MSCRF; G.L. and C.S.), and the Muscular Dystrophy Association (MDA: G.L.)
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