Molecular Analysis of Corrector Compound Effects in Sarcoglycan-Mutant Muscle Cells

Sarcoglycanopathies are a group of rare autosomal recessive limb-girdle muscular dystrophies caused by mutations in genes encoding sarcoglycans. These proteins form a tetrameric complex, a key component of the dystrophin-associated protein complex (DAPC). In a relevant subset of cases, particularly those caused by missense variants, mutant sarcoglycans may be folding-defective and prematurely removed by cellular quality-control machineries through endoplasmic reticulum-associated degradation (ERAD). The resulting loss or destabilization of the sarcoglycan complex compromises sarcolemmal integrity and contributes to progressive muscle degeneration. At present, no curative treatment is available, and clinical management remains largely supportive. A promising therapeutic approach is based on the use of small molecules originally developed as cystic fibrosis transmembrane conductance regulator (CFTR) correctors, which can promote the stabilization and trafficking of folding-defective proteins. Among these compounds, C17 has emerged as a candidate corrector in sarcoglycanopathy models. However, the comprehensive molecular response associated with C17 treatment remains incompletely defined. The aim of this thesis was to characterize the molecular changes associated with C17 treatment in patient-derived sarcoglycan-mutant muscle cells. Mass-spectrometry-based proteomics was used to compare global protein-abundance profiles across three treatment conditions: dimethyl sulfoxide (DMSO), used as the vehicle control; C17, the compound under investigation; and Tosylate (Tos), an inactive derivative of C17, included as a structurally related comparator. Differential-abundance analysis, hierarchical clustering, and functional enrichment were applied to identify treatment-associated proteins and biological pathways. Selected findings were further examined by Western blotting and transcript-level qPCR analysis. C17 treatment was associated with a reproducible increase in several components of the sarcoglycan complex, particularly α-, δ-, and γ-sarcoglycan. Beyond this target-proximal response, treatment-associated changes were identified in proteins related to ER/SR proteostasis, intracellular trafficking, mitochondrial processes, and muscle structural remodeling. Proteomic analysis also revealed a coordinated decrease in multiple enzymes involved in cholesterol biosynthesis, which was largely confirmed by follow-up qPCR analysis. Hierarchical clustering and enrichment analyses additionally identified a mitochondria-related proteomic signature associated with C17 treatment. This study provides a system-level characterization of the molecular response associated with C17 treatment in sarcoglycan-mutant muscle cells. The reproducible increase in disease-relevant sarcoglycan proteins represents the strongest treatment-associated finding, while the accompanying changes in proteostasis, trafficking, cholesterol biosynthesis, mitochondrial biology, and muscle structural proteins identify additional processes for future mechanistic investigation. Although these signatures require further functional validation, they provide a framework for defining the broader cellular response to C17 and prioritizing candidate pathways for subsequent studies.