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Introduction Diffuse cutaneous Systemic Sclerosis (SSc) is a life-limiting autoimmune disease with minimal treatment options. Autologous hematopoietic stem cell transplantation (AHSCT) is a potent disease-modifying therapy in dcSSc; however, its effects on fibroblasts are unknown. We recently showed that dermal fibroblasts (DF) in patients with dcSSc develop a cancer-like phenotype characterized by genomic instability and increased double-stranded DNA breaks (DSB). However, little is known about the mechanisms promoting DF survival following the accumulation of genomic mutations.We hypothesized that in dcssc df genomic instability results in the accumulation of genomic mutations; and that this results in the activation of protein kinase r-like er kinase (perk) and transcription of forkhead box1 (foxo1) promoting resistance-to-apoptosis and fibrosis (figure 1).Material and Methods We used whole exome sequencing (WES) to characterize the mutational frequencies and their associated signatures in dcSSc patients who did not undergo AHSCT (dcSSc, N=35) and those who did (post-AHSCT, N=9). We also generated DFs from dcSSc, post-AHSCT (or age/sex matched healthy controls (HC), (N=8-10 patients/group), and quantified the frequency of DSBs via gamma-H2AX levels (immunoblot (IB)), and DSB nuclear foci (confocal microscopy). We measured the relative ROS levels, mitochondrial membrane potential, and phospho-PERK (active) in DFs to mechanistically link DSB with PERK activation using flow cytometry and/or IB, respectively. We also determined the downstream effects of PERK activation on mitochondria (e.g. mitochondrial dynamics and biogenesis). Then, we measured FOXO1 activation via nuclear translocation, IB, and expression of its downstream mRNA target SOD2. Finally, mitochondrial-dependent resistance-to-apoptosis was determined at baseline, and following treatment with cyclophosphamide, a PERK or a FOXO1-inhibitor using TUNEL and cleaved caspase 9/3 levels (IB).Results dcSSc patients’ DFs had increased genomic instability and DSBs compared to patients treated with AHSCT. dcSSc DFs had increased indicators associated with PERK activation (phospho-PERK, ROS, and mitochondrial membrane potential). This was associated with increased mitochondrial remodelling, mitochondrial biogenesis, and mitochondrial fusion. Importantly, FOXO1 was exclusively activated in dcSSc, but not HC or post-AHSCT DFs. Inhibition of PERK or FOXO1 resulted in increased mitochondrial-dependent apoptosis.Conclusions Our study highlights a novel mechanism whereby genotoxic signals in dcSSc promote cell survival via a PERK/FOXO1-dependent axis and associated metabolic remodeling ( figure 1). It also provides mechanistic insights related to how AHSCT reduces the mutational landscape and pro-fibrotic signals in DF. Future studies targeting this dysregulated pathway may provide an additional rationale for exploring it therapeutically in patients with dcSSc.Funding CIHR/National Scleroderma Foundation/GlycoNET/Arthritis SocietyAbstract P.021 Figure 1Proposed mechanism promoting genomic instability in dcSSc