Deciphering the Mechanism of RUNX1 in Systemic Sclerosis (SSc) Dermal Fibrosis - PROJECT SUMMARY Systemic sclerosis (SSc) is an autoimmune disease associated with high mortality rates. Current disease- modifying therapies have had limited success in improving clinical outcomes and symptoms. The main clinical manifestation of SSc is skin fibrosis, which results from complex changes in transcriptional and signaling pathways in the skin. Through transcription factor activity network analyses using genome-wide data from the skin, the runt-related transcription factor 1 (RUNX1) has been identified as a key regulator in the skin of individuals with diffuse cutaneous systemic sclerosis (dcSSc). RUNX1 is overexpressed in various human cancers, autoimmune diseases, and fibrotic conditions; however, the specific contribution of RUNX1 to the pathogenesis of SSc skin fibrosis remains unknown. Notably, an association between the severity of dermal fibrosis and increased RUNX1 expression levels has been found in skin biopsies of individuals with SSc. Single- cell RNA sequencing (scRNA-seq) data from the skin of patients with SSc demonstrates enrichment of RUNX1 in fibroblast. We developed a 3D tissue model — called self-assembled Skin Equivalent (saSE) — that incorporates patient-derived fibroblasts and monocytes and shows overexpression of RUNX1 similar to that observed in patients’ biopsies. Our central hypothesis is that RUNX1 is necessary for the cellular transition that results in the generation of the profibrotic myofibroblasts population in SSc. The primary objectives of this study are: (i) to establish the function and molecular mechanism(s) of RUNX1 in SSc-specific fibroblast populations, with the aim of elucidating how RUNX1 drives the activation of dermal fibroblasts, leading to matrix remodeling, increased matrix deposition, and enhanced contractility; and (ii) to assess the impact of inhibiting RUNX1 in a novel, in-vitro SSc skin equivalent model (saSE) in order to determine the effect of RUNX1 inhibition on profibrotic phenotypes and SSc-specific fibroblast populations through single-cell sequencing. The successful completion of this study will provide valuable insights into the mechanistic role of RUNX1 in SSc skin fibrosis. Anticipated outcomes include understanding how RUNX1 influences SSc-specific fibroblast populations, evaluating the effectiveness of inhibiting RUNX1 in reducing pro-fibrotic characteristics in a patient-derived skin equivalent model, and uncovering cellular heterogeneity and gene expression patterns through single-cell sequencing. By combining bioinformatic analyses, an innovative in-vitro 3D skin-like tissue model, and advanced sequencing technologies, this research will shed new light on the role of RUNX1 and its therapeutic potential in SSc dermal fibrosis. The environment at Dartmouth College is ideal for the proposed research, with an innovative, collaborative, and well-equipped research infrastructure that promotes cutting-edge scientific inquiry. Together with the outlined training plan, the proposed work will support my career plan of becoming an accomplished researcher in the field of molecular systems pharmacology and translational therapeutics.
