Background and Aims: Primary sclerosing cholangitis (PSC) is a heterogeneous cholangiopathy characterized by progressive biliary fibrosis. RNA sequencing of liver tissue from patients with PSC (n = 74) enrolled in a 96-week clinical trial was performed to identify associations between biological pathways that were independent of fibrosis and clinical events. Approach and Results: The effect of fibrosis was subtracted from gene expression using a computational approach. The fibrosis-adjusted gene expression patterns were associated with time to first PSC-related clinical event (e.g., cholangitis, hepatic decompensation), and differential expression based on risk groups and Ingenuity Pathway Analysis were performed. Baseline demographic data were representative of PSC: median age 48 years, 71% male, 49% with inflammatory bowel disease, and 44% with bridging fibrosis or cirrhosis. The first principle component (PC1) of RNA-sequencing data accounted for 18% of variance and correlated with fibrosis stage (ρ = −0.80; P < 0.001). After removing the effect of fibrosis-related genes, the first principle component was not associated with fibrosis (ρ = −0.19; P = 0.11), and a semisupervised clustering approach identified two distinct patient clusters with differential risk of time to first PSC-related event (P < 0.0001). The two groups had similar fibrosis stage, hepatic collagen content, and α-smooth muscle actin expression by morphometry, Enhanced Liver Fibrosis score, and serum liver biochemistry, bile acids, and IL-8 (all P > 0.05). The top pathways identified by Ingenuity Pathway Analysis were eukaryotic translation inhibition factor 2 (eIF2) signaling and regulation of eIF4/p70S6K signaling. Genes involved in the unfolded protein response, activating transcription factor 6 (ATF6) and eIF2, were differentially expressed between the PSC clusters (down-regulated in the high-risk group by log-fold changes of −0.18 [P = 0.02] and −0.16 [P = 0.02], respectively). Clinical events were enriched in the high-risk versus low-risk group (38% [12/32] vs. 2.4% [1/42], P < 0.0001). Conclusions: Removing the contribution of fibrosis-related pathways uncovered alterations in the unfolded protein response, which were associated with liver-related complications in PSC.
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