Duchenne muscular dystrophy (DMD) is an X-linked recessive neuromuscular disorder with a prevalence of approximately 1 in 3500–5000 males. DMD manifests as childhood-onset muscle degeneration, followed by loss of ambulation, cardiomyopathy, and death in early adulthood due to a lack of functional dystrophin protein. Out-of-frame mutations in the dystrophin gene are the most common underlying cause of DMD. Gene editing via the clustered regularly interspaced short palindromic repeats (CRISPR) system is a promising therapeutic for DMD, as it can permanently correct DMD mutations and thus restore the reading frame, allowing for the production of functional dystrophin. The specific mechanism of gene editing can vary based on a variety of factors such as the number of cuts generated by CRISPR, the presence of an exogenous DNA template, or the current cell cycle stage. CRISPR-mediated gene editing for DMD has been tested both in vitro and in vivo, with many of these studies discussed herein. Additionally, novel modifications to the CRISPR system such as base or prime editors allow for more precise gene editing. Despite recent advances, limitations remain including delivery efficiency, off-target mutagenesis, and long-term maintenance of dystrophin. Further studies focusing on safety and accuracy of the CRISPR system are necessary prior to clinical translation.

Antisense oligonucleotide–mediated (AO-mediated) therapy is a promising strategy to treat several neurological diseases, including spinal muscular atrophy (SMA). However, limited delivery to the CNS with AOs administered intravenously or subcutaneously is a major challenge. Here, we demonstrate a single subcutaneous administration of cell-penetrating peptide DG9 conjugated to an AO called phosphorodiamidate morpholino oligomer (PMO) reached the CNS and significantly prolonged the median survival compared with unconjugated PMO and R6G-PMO in a severe SMA mouse model. Treated mice exhibited substantially higher expression of full-length survival of motor neuron 2 in both the CNS and systemic tissues compared with nontreated and unmodified AO–treated mice. The treatment ameliorated the atrophic musculature and improved breathing function accompanied by improved muscle strength and innervation at the neuromuscular junction with no signs of apparent toxicity. We also demonstrated DG9-conjugated PMO localized in nuclei in the spinal cord and brain after subcutaneous injections. Our data identify DG9 peptide conjugation as a powerful way to improve the efficacy of AO-mediated splice modulation. Finally, DG9-PMO is a promising therapeutic option to treat SMA and other neurological diseases, overcoming the necessity for intrathecal injections and treating body-wide tissues without apparent toxicity.

Duchenne muscular dystrophy (DMD) is an X-linked recessive neuromuscular disorder with a prevalence of approximately 1 in 3,500-5,000 males. DMD manifests as childhood-onset muscle degeneration, followed by loss of ambulation, cardiomyopathy, and death in early adulthood due to a lack of functional dystrophin protein. Out-of-frame mutations in the dystrophin gene are the most common underlying cause of DMD. Gene editing via the clustered regularly interspaced short palindromic repeats (CRISPR) system is a promising therapeutic for DMD, as it can permanently correct DMD mutations and thus restore the reading frame, allowing for the production of functional dystrophin. The specific mechanism of gene editing can vary based on a variety of factors such as the number of cuts generated by CRISPR, the presence of an exogenous DNA template, or the current cell cycle stage. CRISPR-mediated gene editing for DMD has been tested both in vitro and in vivo, with many of these studies discussed herein. Additionally, novel modifications to the CRISPR system such as base or prime editors allow for more precise gene editing. Despite recent advances, limitations remain including delivery efficiency, off-target mutagenesis, and long-term maintenance of dystrophin. Further studies focusing on safety and accuracy of the CRISPR system are necessary prior to clinical translation.

Syndromic cardiac malformations can result in morbidity, yet their genetic etiology is only understood for a subset of individuals. Genome sequencing efforts in congenital anomaly cohorts may identify disease-associated variants in previously unrecognized genes. Through international matchmaking efforts, we identified eighteen individuals in total with de novo or loss-of-function variants in EIF3A (n = 4) or EIF3B (n = 14). The clinical phenotype varied but predominantly included cardiac defects, craniofacial dysmorphisms, mild developmental delays, and behavioral abnormalities. These genes encode core subunits of the eukaryotic initiation factor 3 (eIF3) complex, which plays a critical role in binding mRNA transcripts to the 40S ribosomal subunit during translation initiation. Both genes are highly constrained against loss of function, and animal models have demonstrated that disruptions in the eIF3 complex result in a range of developmental defects, including cardiovascular malformations. Additionally, EIF3B is located within the minimally overlapping region implicated in cardiac anomalies associated with 7p22.3 microdeletions. We sought to further study the role of these genes in syndromic congenital heart disease. To explore their functional impact, we generated zebrafish models with mutations in the orthologous eif3s10 and eif3ba genes, which resulted in developmental abnormalities, including thin heart tubes, lack of craniofacial cartilage, and embryonic lethality. We propose that pathogenic variants in EIF3A, as well as pathogenic variants or microdeletions involving EIF3B, cause a distinct autosomal-dominant neurodevelopmental syndrome characterized by cardiovascular and craniofacial manifestations.