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Clinical and Translational Medicine May 2024
Topics: Humans; DNA Copy Number Variations; Female; Cohort Studies; Pregnancy; Dystrophin; Genetic Testing; Noninvasive Prenatal Testing; Prenatal Diagnosis
PubMed: 38797938
DOI: 10.1002/ctm2.1706 -
Cells May 2024In recent years, clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) protein have emerged as a revolutionary gene editing... (Review)
Review
In recent years, clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) protein have emerged as a revolutionary gene editing tool to treat inherited disorders affecting different organ systems, such as blood and muscles. Both hematological and neuromuscular genetic disorders benefit from genome editing approaches but face different challenges in their clinical translation. The ability of CRISPR/Cas9 technologies to modify hematopoietic stem cells ex vivo has greatly accelerated the development of genetic therapies for blood disorders. In the last decade, many clinical trials were initiated and are now delivering encouraging results. The recent FDA approval of Casgevy, the first CRISPR/Cas9-based drug for severe sickle cell disease and transfusion-dependent β-thalassemia, represents a significant milestone in the field and highlights the great potential of this technology. Similar preclinical efforts are currently expanding CRISPR therapies to other hematologic disorders such as primary immunodeficiencies. In the neuromuscular field, the versatility of CRISPR/Cas9 has been instrumental for the generation of new cellular and animal models of Duchenne muscular dystrophy (DMD), offering innovative platforms to speed up preclinical development of therapeutic solutions. Several corrective interventions have been proposed to genetically restore dystrophin production using the CRISPR toolbox and have demonstrated promising results in different DMD animal models. Although these advances represent a significant step forward to the clinical translation of CRISPR/Cas9 therapies to DMD, there are still many hurdles to overcome, such as in vivo delivery methods associated with high viral vector doses, together with safety and immunological concerns. Collectively, the results obtained in the hematological and neuromuscular fields emphasize the transformative impact of CRISPR/Cas9 for patients affected by these debilitating conditions. As each field suffers from different and specific challenges, the clinical translation of CRISPR therapies may progress differentially depending on the genetic disorder. Ongoing investigations and clinical trials will address risks and limitations of these therapies, including long-term efficacy, potential genotoxicity, and adverse immune reactions. This review provides insights into the diverse applications of CRISPR-based technologies in both preclinical and clinical settings for monogenic blood disorders and muscular dystrophy and compare advances in both fields while highlighting current trends, difficulties, and challenges to overcome.
Topics: Humans; Genetic Therapy; CRISPR-Cas Systems; Animals; Gene Editing; Muscular Dystrophy, Duchenne; Clinical Trials as Topic; Clustered Regularly Interspaced Short Palindromic Repeats
PubMed: 38786024
DOI: 10.3390/cells13100800 -
Biomolecules May 2024Chimerism-based strategies represent a pioneering concept which has led to groundbreaking advancements in regenerative medicine and transplantation. This new approach... (Review)
Review
Chimerism-based strategies represent a pioneering concept which has led to groundbreaking advancements in regenerative medicine and transplantation. This new approach offers therapeutic potential for the treatment of various diseases, including inherited disorders. The ongoing studies on chimeric cells prompted the development of Dystrophin-Expressing Chimeric (DEC) cells which were introduced as a potential therapy for Duchenne Muscular Dystrophy (DMD). DMD is a genetic condition that leads to premature death in adolescent boys and remains incurable with current methods. DEC therapy, created via the fusion of human myoblasts derived from normal and DMD-affected donors, has proven to be safe and efficacious when tested in experimental models of DMD after systemic-intraosseous administration. These studies confirmed increased dystrophin expression, which correlated with functional and morphological improvements in DMD-affected muscles, including cardiac, respiratory, and skeletal muscles. Furthermore, the application of DEC therapy in a clinical study confirmed its long-term safety and efficacy in DMD patients. This review summarizes the development of chimeric cell technology tested in preclinical models and clinical studies, highlighting the potential of DEC therapy in muscle regeneration and repair, and introduces chimeric cell-based therapies as a promising, novel approach for muscle regeneration and the treatment of DMD and other neuromuscular disorders.
Topics: Muscular Dystrophy, Duchenne; Humans; Regeneration; Animals; Cell- and Tissue-Based Therapy; Muscle, Skeletal; Dystrophin; Myoblasts
PubMed: 38785982
DOI: 10.3390/biom14050575 -
Disease Models & Mechanisms May 2024Absence of dystrophin results in muscular weakness, chronic inflammation and cardiomyopathy in Duchenne muscular dystrophy (DMD). Pharmacological corticosteroids are the...
Absence of dystrophin results in muscular weakness, chronic inflammation and cardiomyopathy in Duchenne muscular dystrophy (DMD). Pharmacological corticosteroids are the DMD standard of care; however, they have harsh side effects and unclear molecular benefits. It is uncertain whether signaling by physiological corticosteroids and their receptors plays a modifying role in the natural etiology of DMD. Here, we knocked out the glucocorticoid receptor (GR, encoded by Nr3c1) specifically in myofibers and cardiomyocytes within wild-type and mdx52 mice to dissect its role in muscular dystrophy. Double-knockout mice showed significantly worse phenotypes than mdx52 littermate controls in measures of grip strength, hang time, inflammatory pathology and gene expression. In the heart, GR deletion acted additively with dystrophin loss to exacerbate cardiomyopathy, resulting in enlarged hearts, pathological gene expression and systolic dysfunction, consistent with imbalanced mineralocorticoid signaling. The results show that physiological GR functions provide a protective role during muscular dystrophy, directly contrasting its degenerative role in other disease states. These data provide new insights into corticosteroids in disease pathophysiology and establish a new model to investigate cell-autonomous roles of nuclear receptors and mechanisms of pharmacological corticosteroids.
Topics: Animals; Receptors, Glucocorticoid; Mice, Inbred mdx; Mice, Knockout; Dystrophin; Myocardium; Muscular Dystrophy, Duchenne; Muscle, Skeletal; Myocytes, Cardiac; Mice; Cardiomyopathies; Mice, Inbred C57BL; Muscular Dystrophy, Animal; Phenotype; Systole
PubMed: 38770680
DOI: 10.1242/dmm.050397 -
BMC Musculoskeletal Disorders May 2024Duchenne muscular dystrophy (DMD) is a devastating X-linked neuromuscular disorder caused by various defects in the dystrophin gene and still no universal therapy. This...
OBJECTIVE
Duchenne muscular dystrophy (DMD) is a devastating X-linked neuromuscular disorder caused by various defects in the dystrophin gene and still no universal therapy. This study aims to identify the hub genes unrelated to excessive immune response but responsible for DMD progression and explore therapeutic siRNAs, thereby providing a novel treatment.
METHODS
Top ten hub genes for DMD were identified from GSE38417 dataset by using GEO2R and PPI networks based on Cytoscape analysis. The hub genes unrelated to excessive immune response were identified by GeneCards, and their expression was further verified in mdx and C57 mice at 2 and 4 months (M) by (RT-q) PCR and western blotting. Therapeutic siRNAs were deemed as those that could normalize the expression of the validated hub genes in transfected C2C12 cells.
RESULTS
855 up-regulated and 324 down-regulated DEGs were screened from GSE38417 dataset. Five of the top 10 hub genes were considered as the candidate genes unrelated to excessive immune response, and three of these candidates were consistently and significantly up-regulated in mdx mice at 2 M and 4 M when compared with age-matched C57 mice, including Col1a2, Fbn1 and Fn1. Furthermore, the three validated up-regulated candidate genes can be significantly down-regulated by three rational designed siRNA (p < 0.0001), respectively.
CONCLUSION
COL1A2, FBN1 and FN1 may be novel biomarkers for DMD, and the siRNAs designed in our study were help to develop adjunctive therapy for Duchenne muscular dystrophy.
Topics: Muscular Dystrophy, Duchenne; Animals; Mice, Inbred mdx; RNA, Small Interfering; Mice; Mice, Inbred C57BL; Disease Models, Animal; Male; Humans; Protein Interaction Maps
PubMed: 38762732
DOI: 10.1186/s12891-024-07206-6 -
The American Journal of Pathology May 2024Duchenne muscular dystrophy (DMD), caused by loss-of-function mutations in the dystrophin gene, results in progressive muscle weakness and early fatality. Impaired...
Leucyl-tRNA Synthetase Contributes to Muscle Weakness through Mammalian Target of Rapamycin Complex 1 Activation and Autophagy Suppression in a Mouse Model of Duchenne Muscular Dystrophy.
Duchenne muscular dystrophy (DMD), caused by loss-of-function mutations in the dystrophin gene, results in progressive muscle weakness and early fatality. Impaired autophagy is one of the cellular hallmarks of DMD, contributing to the disease progression. Molecular mechanisms underlying the inhibition of autophagy in DMD are not well understood. In the current study, the DMD mouse model mdx is used for the investigation of signaling pathways leading to suppression of autophagy. Mammalian target of rapamycin complex 1 (mTORC1) is found to be hyperactive in the DMD muscles, accompanying muscle weakness and autophagy impairment. Surprisingly, Akt, a well-known upstream regulator of mTORC1, is not responsible for mTORC1 activation or the dystrophic muscle phenotypes. Instead, leucyl-tRNA synthetase (LeuRS) is found to be overexpressed in mdx muscles compared with the wild type. LeuRS is known to activate mTORC1 in a noncanonical mechanism that involves interaction with RagD, an activator of mTORC1. Disrupting LeuRS interaction with RagD by the small-molecule inhibitor BC-LI-0186 reduces mTORC1 activity, restores autophagy, and ameliorates myofiber damage in the mdx muscles. Furthermore, inhibition of LeuRS by BC-LI-0186 improves dystrophic muscle strength in an autophagy-dependent manner. Taken together, our findings uncover a noncanonical function of the housekeeping protein LeuRS as a potential therapeutic target in the treatment of DMD.
PubMed: 38762116
DOI: 10.1016/j.ajpath.2024.04.006 -
Trends in Molecular Medicine May 2024Duchenne muscular dystrophy (DMD) is caused by mutations in the X-linked DMD gene, resulting in the absence of dystrophin, progressive muscle degeneration, and heart... (Review)
Review
Duchenne muscular dystrophy (DMD) is caused by mutations in the X-linked DMD gene, resulting in the absence of dystrophin, progressive muscle degeneration, and heart failure. Genetically tailored pig models resembling human DMD mutations recapitulate the biochemical, clinical, and pathological hallmarks of DMD with an accelerated disease progression compared to human patients. DMD pigs have been used to evaluate therapeutic concepts such as gene editing to reframe a disrupted DMD reading frame or the delivery of artificial chromosome vectors carrying the complete DMD gene. Moreover, DMD pigs have been instrumental in validating new diagnostic modalities such as multispectral optoacoustic tomography (MSOT) for non-invasive monitoring of disease progression. DMD pigs may thus help to bridge the gap between proof-of-concept studies in cellular or rodent models and clinical studies in patients.
PubMed: 38749865
DOI: 10.1016/j.molmed.2024.04.013 -
Research Square May 2024Current gene therapy for Duchenne muscular dystrophy (DMD) utilizes adeno-associated virus (AAV) to deliver miniaturized dystrophin (micro-dystrophin or µDys), which...
Current gene therapy for Duchenne muscular dystrophy (DMD) utilizes adeno-associated virus (AAV) to deliver miniaturized dystrophin (micro-dystrophin or µDys), which does not provide full protection for striated muscles as it lacks many important functional domains within full-length (FL) dystrophin. Here we develop a triple vector system to deliver FL-dystrophin into skeletal and cardiac muscles. We rationally split FL-dystrophin into three fragments (N, M, and C) linked to two orthogonal pairs of split intein, allowing efficient, unidirectional assembly of FL-dystrophin. The three fragments packaged in myotropic AAV (MyoAAV4A) restore FL-dystrophin expression in both skeletal and cardiac muscles in male mice. Dystrophin-glycoprotein complex components are also restored in the sarcolemma of dystrophic muscles. MyoAAV4A-delivered FL-dystrophin significantly improves muscle histopathology, contractility, and overall strength comparable to µDys, but unlike µDys, it also restores defective ERK signaling in heart. The FL-dystrophin gene therapy therefore promises to offer superior protection for DMD.
PubMed: 38746161
DOI: 10.21203/rs.3.rs-3867299/v1 -
Frontiers in Physiology 2024Duchenne muscular dystrophy (DMD) is a fatal striated muscle degenerative disease. DMD is caused by loss of dystrophin protein, which results in sarcolemmal instability...
Duchenne muscular dystrophy (DMD) is a fatal striated muscle degenerative disease. DMD is caused by loss of dystrophin protein, which results in sarcolemmal instability and cycles of myofiber degeneration and regeneration. Pathology is exacerbated by overactivation of infiltrating immune cells and fibroblasts, which leads to chronic inflammation and fibrosis. Mineralocorticoid receptors (MR), a type of nuclear steroid hormone receptors, are potential therapeutic targets for DMD. MR antagonists show clinical efficacy on DMD cardiomyopathy and preclinical efficacy on skeletal muscle in DMD models. We have previously generated myofiber and myeloid MR knockout mouse models to dissect cell-specific functions of MR within dystrophic muscles. Here, we compared skeletal muscle gene expression from both knockouts to further define cell-type specific signaling downstream from MR. Myeloid MR knockout increased proinflammatory and profibrotic signaling, including numerous myofibroblast signature genes. was the most highly upregulated fibrotic gene in myeloid MR-knockout skeletal muscle and is a component of fibrosis in dystrophic skeletal muscle. Surprisingly, (), canonically a collagen crosslinker, was increased in both MR knockouts, but did not localize to fibrotic regions of skeletal muscle. Lox localized within myofibers, including only a region of quadriceps muscles. (), another Lox family member, was increased only in myeloid MR knockout muscle and localized specifically to fibrotic regions. This study suggests that MR signaling in the dystrophic muscle microenvironment involves communication between contributing cell types and modulates inflammatory and fibrotic pathways in muscle disease.
PubMed: 38737833
DOI: 10.3389/fphys.2024.1322729 -
Translational Lung Cancer Research Apr 2024The identification of prognostic biomarkers is crucial for guiding treatment strategies in mesothelioma patients. The Duchenne muscular dystrophy () gene and its...
BACKGROUND
The identification of prognostic biomarkers is crucial for guiding treatment strategies in mesothelioma patients. The Duchenne muscular dystrophy () gene and its specific transcripts have been associated with patient survival in various tumours. In this study, we aimed to investigate the prognostic potential of gene expression and its transcripts in mesothelioma patients.
METHODS
We analysed The Cancer Genome Atlas (TCGA) mesothelioma RNAseq, mutation, and clinical data to assess the association between gene expression and its transcripts (Dp427, Dp71 splice variants) and mesothelioma survival. We also evaluated the specific Dp71 transcript as a unique prognostic biomarker across mesothelioma subtypes. Additionally, we performed differential gene expression analysis between high and low gene/transcript expression groups.
RESULTS
The analysis included 57 epithelioid, 23 biphasic, two sarcomatoid, and five not otherwise specified (NOS) histological subtypes of mesothelioma samples. Univariate analysis revealed that high expression of the gene and its Dp71 transcript was significantly associated with shorter survival in mesothelioma patients (P0.003 and P<0.001, respectively). In a multivariate analysis, the association between Dp71 expression and survival remained significant [hazard ratio (HR) 2.29, 95% confidence interval (CI): 1.24-4.23, P0.008] across all mesothelioma patients, and also among patients with mesotheliomas without deep deletions (HR 3.58, 95% CI: 1.31-9.80, P0.01). Pathway analysis revealed enrichment of cell cycle (P3.01×10) and homologous recombination (P0.01) pathways in differentially expressed genes (DEGs) between high and low Dp71 groups. Furthermore, there were correlations between Dp71 transcript expression and tumour microenvironment (TME) cells, including a weak positive correlation with macrophages (R=0.32, P0.002) specifically M2 macrophages (R=0.34, P0.001).
CONCLUSIONS
Our findings indicate that the differential expression of specific transcripts is associated with poor survival in mesothelioma patients. The specific Dp71 transcript can serve as a potential biomarker for predicting patient survival in diverse histological subtypes of mesothelioma. Further studies are needed to understand the role of specific dystrophin transcripts in cancer and TME cells, and their implications in the pathogenesis and progression of mesothelioma. Identifying patients at risk of poor survival based on transcript expression can guide treatment strategies in mesothelioma, informing decisions regarding treatment intensity, follow-up schedules, eligibility for clinical trials, and ultimately, end-of-life care planning.
PubMed: 38736495
DOI: 10.21037/tlcr-24-28