Session Title: Systemic Sclerosis, Fibrosing Syndromes, and Raynaud's II
Session Type: Abstract Submissions (ACR)
Systemic sclerosis (SSc) or scleroderma, like many fibrotic disorders, has no effective therapy. Deposition of collagen and excess extracellular matrix depends on the transition of dermal fibroblasts into myofibroblasts. Current drug development has focused on targeting the initial inflammatory stimuli and specific receptors involved in fibrosis. Recent evidence, however, indicates that a transcriptional program regulated by Rho GTPase is critical for myofibroblast transition. This could be a more effective convergent target for interrupting pathological fibrosis regardless of the initial inflammatory or fibrotic stimulus. We recently identified novel inhibitors of this transcription mechanism including CCG-203971 (Bell et al, Bioorg Med Chem Lett. 23:3826, 2013). Here we assess their effectiveness in human SSc dermal fibroblasts.
Primary human dermal fibroblasts were obtained from normal individuals and patients with diffuse SSc. Cells were passaged three times in DMEM containing 10% fetal bovine serum prior to studies which were carried out at passage 4 or 5. mRNA for connective tissue growth factor (CTGF), alpha-smooth muscle actin (ACTA2) and collagen type 1 alpha 1 (COL1α1) were quantified by qRT-PCR. The fraction of cells positive for the myofibroblast maker alpha-smooth muscle actin (α-SMA) was determined by immunocytochemistry. Inhibitors of Rho/MRTF/SRF-regulated gene expression (CCG-203971) and pirfenidone, the only approved antifibrotic therapy, were tested for their ability to suppress these markers of myofibroblast transition and fibrosis.
SSc dermal fibroblasts express significantly more mRNA for CTGF (3.7-fold) and ACTA2 (4.6-fold) than do normal control fibroblasts. Similarly the percentage of cells staining positively for α-SMA was significantly greater for SSc (77±9%) than for normal (27±18%) dermal fibroblasts (p<0.001, n=6). CTGF,ACTA2, and COL1α1 mRNA in SSc fibroblasts were inhibited at 24 hours by CCG-203971 with an IC50 of ~10 μM. Treatment of SSc fibroblasts for 72 hours with 10 µM CCG-203971 reduced the percentage of α-SMA positive cells to control levels (26±18%) while pirfenidone at 300 µM was only marginally effective (41±16%).
Targeting the Rho/MRTF/SRF transcriptional pathway strongly suppresses myofibroblast activation and fibrosis-related gene expression in human SSc dermal fibroblasts. This could represent a novel targeted approach to disrupt a key genetic switch involved in fibrosis mechanisms and may provide a broad spectrum approach to treatment for systemic sclerosis and other disorders of fibrosis.
P. S. Tsou,
D. A. Fox,
S. D. Larsen,
R. R. Neubig,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/modulating-myofibroblast-transition-of-human-scleroderma-fibroblasts-through-inhibition-of-rho-guanine-nucleotidase-regulated-gene-transcription/