Single Nuclei Transcriptome Analysis of DMD to Reveal Disease-Modifying Targets
Abstract
Skeletal muscle is a cellularly complex and plastic tissue that responds to local injury by recruiting and expanding immune cells, fibroblasts, and other muscle resident cells to guide satellite cell-mediated repair of damaged multinucleated myofibers. In Duchenne muscular dystrophy (DMD), lack of dystrophin leads to chronic muscle damage, immune infiltration, dysregulation of repair, with loss of muscle function and replacement with fat and fibrosis. Our understanding of the cell and molecular mechanisms regulating dystrophy and repair remain incomplete, especially in humans. Here we propose to study all of the cells within muscle and their gene expression response with dystrophin loss or restoration that occurs naturally or with genetic therapies. We have developed relatively less invasive biopsy methods, coupled with improvements in sample handling that allow us to study about 5,000 nuclei from each person’s muscle biopsy and sequence about 20,000 genes from each nucleus. We have analyzed 8 healthy muscles, 9 DMD muscles with some dystrophin expression, and 43 DMD samples from across the disease severity and stage of disease. This has created an unprecedented set of observations of how cells in muscle are responding to the lack of dystrophin or restoration of dystrophin. Here we propose to augment this database by the strategic collection on an additional 60 DMD/BMD muscles to broaden and clarify insights that can be gleaned from an unbiased assessment of gene expression in dystrophic muscle. We infer that genes that are differentially expressed in relation to disease severity or stage of disease highlight molecules that are potentially identifying therapeutic targets of prognostic markers. We will sample muscle from DMD patients as young as 2 years to as old as 27 years, and BMD patients from 5 years to 69 years, and 20 healthy controls. This data in aggregate will provide unprecedented insights into Duchenne and highlight otherwise cryptic pathways for disease mitigation and identify novel biomarkers of disease. All of the collected and organized data will be shared publicly without patient identifiers to permit ongoing investigation and serve as a reference for muscle studies with various DMD therapeutics in development to accelerate novel treatments. All patients with a dystrophinopathy may benefit from the output of this research. Within 1 year, we will develop a sensitive RNA-based method to observe transgene expression or exon skipping of mRNA from small muscle samples, and within 3 years we will identify promising therapeutic candidates most relevant for augmentation/combination/follow on treatment from therapies like exon skipping and gene therapy. This work is a translational study addressing the Duchenne Muscular Dystrophy Research Program Translational Research Award Focus Area of skeletal muscle to improve care for DMD by unbiased discovery of novel prognostic, age, and disease severity expression biomarker, which may serve as surrogate clinical markers. Further, this project will augment our understanding of the natural history of DMD and provide clues to the field for novel therapeutic targeting and combinational targeting of pathways.
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Jan 04, 2024
- Source ID
- HT94252310597
Entities
People
- Stanley Nelson
Organizations
- United States Army
- University of California, Los Angeles