Congenital muscular dystrophy (CMD) is a group of genetically inherited neuromuscular disorders characterized by muscle weakness and degeneration. LAMA2-deficient CMD is a subtype caused by mutations in the laminin alpha-2 (LAMA2) gene, for which there is no treatment available. Upregulation of disease compensatory gene Lama1, achieved via CRISPR activation and delivered using AAV9, is a promising therapeutic approach for LAMA2-CMD. However, progress towards the clinical application of such strategy is hampered by the lack of comprehensive baseline immunophenotypes in LAMA2-CMD. Such knowledge is important because precise interventions on inflammatory pathways are crucial to minimize the risks and augment the success of gene therapy administrations, irrespective of the disease context. Therefore, we set out to fill such knowledge gap by investigating the dynamics of lymphocytes, myeloid cells, and circulating cytokines in skeletal muscles and blood from dyW, a mouse model of LAMA2-CMD.
We found increased leukocyte infiltration in muscles from the dyW mice as early as 2-weeks old, with pro-inflammatory macrophages (M1) and T regulatory cells (Tregs) as the predominant cell types. This suggests the involvement of these cells in the early stage of muscle pathology. We also observed increased inflammatory cytokines IL-6 and IFN-gamma in the dyW blood, indicating their role in initiating damage and inflammation signals from the circulation to the muscles. Subsequently, we evaluate how these immune cell profiles change in the mice treated with AAV9 carrying CRISPRa to upregulate disease compensatory gene Lama1. We observed a significant decrease in the levels of IL-6 and IFN-gamma cytokines in the serum of treated dyW mice. Additionally, the expression of activation markers PD1 and GITR on CD4+ T cells was reduced following the upregulation of Lama1.
Collectively, our study highlights the role of immune cells and inflammatory mediators in early pathology of LAMA2-CMD, which can be reduced following CRISPRa-mediated Lama1 upregulation. These findings are important to increase our understanding of the immunological changes associated with LAMA2-CMD and pave the path towards development of safe and effective therapy for this condition.