Session Type: Abstract Submissions (ACR)
Background/Purpose: Transformed synovial fibroblasts (SF) mediate joint-specific damage in rheumatoid arthritis (RA) by expressing integrins and metalloproteinase that promote adhesion to and invasion of cartilage. The mechanism of SF transformation is unknown, but is critical for the rational design of specific therapies to prevent joint erosion in RA. We recently identified aberrant BRAF splice variants in synovial fibroblasts from some RA patients and demonstrated their role in RA fibroblast proliferation, results that suggest a primary role for oncogenic transformation of RA SF. The current studies were designed to further evaluate the role of oncogenesis in RA SF transformation.
Methods: Aberrant BRAF splice variants and mutations in KRAS were identified in RA SF by RT-PCR. The function of aberrant BRAF splice variants was evaluated in NIH-3T3 fibroblasts transfected with an expression vector containing cDNA of BRAF splice variants. Mitogen-activated protein kinase (MAPK) activation in transfected NIH-3T3 cells was determined by phosphorylation of MEK and ERK. The role of BRAF and CRAF in SF transformation was determined by RNAi, and Membrane-Type 1 Matrix Metalloproteinase (MT1-MMP) was identified in cells with MT1-MMP-specific antibodies. Collagen invasion by transfected NIH-3T3 cells was evaluated in an in vitro collagen invasion assay.
Results: SF from 6/9 RA patients had kinase “Dead” aberrant BRAF splice variants. NIH-3T3 cells transfected with aberrant BRAF splice variants constitutively activate MAPK, produce MT1-MMP, and invade collagen. Since “Dead” BRAF forms dimers with CRAF to activate MAPK we evaluated the role of CRAF in MAPK activation by RNAi. MAPK activation was inhibited by siRNA specific for BRAF and CRAF. Since MAPK activation by “Dead” BRAF also requires activated KRAS we looked for KRAS mutations in RA SF and identified mutations in 7/9 RA patients.
Conclusion: Our results suggest that joint-specific oncogenesis is responsible for synovial fibroblast transformation in some patients with RA.
N. L. Farmani,
K. K. Colburn,
R. H. Weisbart,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/oncogenic-activation-of-mapk-in-rheumatoid-arthritis-synovial-fibroblasts/