Background: Givinostat, a histone deacetylase (HDAC) inhibitor, is the first epigenetic drug approved for the treatment of Duchenne muscular dystrophy (DMD). DMD features progressive muscle degeneration with replacement by fibrotic matrix, driven mainly by fibro–adipogenic progenitor cells (FAPs) and skeletal muscle fibroblasts (SMFs). HDAC enzymes regulate these pathogenic stromal responses, and their inhibition restores homeostatic conditions.
Objective: To use integrated RNA-seq and ATAC-seq to define how givinostat modulates TGFβ-induced fibrotic programs in DMD-derived FAPs and SMFs.
Results: In both cell types, TGFβ induced a fibrotic transformation, activating epithelial–mesenchymal transition (EMT) and extracellular matrix (ECM) pathways, upregulating core effectors such as FN1, collagens, CDH2, and ACTA2, and extensive opening of EMT/ECM-associated regulatory regions. In DMD FAPs, TGFβ additionally suppressed immune and chemotaxis signatures, shifting cells toward a matrix-producing, immunosuppressed phenotype. Givinostat strongly counteracted these changes, overturning EMT/ECM signatures, reducing expression and accessibility at matrix genes, and reversing myofibroblast marker induction. EMT transcription factors SNAI1 and ZEB1 were suppressed, while programs mediated by CXCL1/2/3, CXCL8/IL8, and CXCR4 were re-engaged, indicating chemotactic signaling restoration. Our multi-omics approach revealed cell-type–specific mechanisms. In FAPs, givinostat reversed fibrotic nodes, including TNC, LOXL2, and MMP9, and dampened NF-κB–linked chemokine axes (CCL22/CCL20/IL18), yet broad TGFβ-induced immune suppression remained partially corrected. In SMFs, TGFβ induced a prominent IL11 autocrine loop and proliferative/stress programs.
Conclusions: For the first time, we demonstrate that givinostat potently reduced IL11, downregulated E2F/G2–M/Myc cell-cycle modules, the unfolded protein response, and shifted the SMAD3–TEAD3–SRF EMT/mechanotransduction hub toward a more quiescent state. These findings indicate that givinostat exerts a common anti-fibrotic effect on TGFβ-driven EMT/ECM across the DMD stromal populations examined, while rewiring distinct upstream regulatory circuits in FAPs and SMF fibroblasts. All the targets identified here provide a mechanistic framework for optimizing HDAC inhibition and for developing biomarkers of anti-fibrotic response in DMD.