OP28 Defective STAT3 signaling in refractory Very Early Onset Inflammatory Bowel Disease is associated with a transcriptional signature which predicts response to anti-IL23-based therapies

Abstract Background Very early onset inflammatory bowel disease (VEOIBD) has a monogenic cause in >5% of cases. IL10R deficiency is among the more common causes of monogenic IBD and presents with severe disease refractory to conventional therapies. For most VEOIBD patients, the molecular basis of disease is unknown. We employed flow cytometry and bulk RNA-sequencing to localize pathway-level dysfunction in VEOIBD patients with unknown molecular basis. Methods VEOIBD patients prospectively enrolled in a biorepository were screened for IL10R deficiency using a flow cytometry assay. Patient peripheral blood mononuclear cells (PBMCs) were stimulated with IL10 and control cytokine IL21 and pSTAT3 activation was measured. RNA-sequencing was performed on 115 whole blood samples, comprising VEOIBD patients with either confirmed (n=35, IL10R deficient n=4), or unknown monogenic causes (n=70), and controls (n=10). Transcriptome analysis focused on patients with abnormal STAT3 activation as determined by flow cytometry. Results We identified 6 patients with refractory VEOIBD and defective phosphorylation of STAT3 upon stimulation with both IL10 and IL21 by flow cytometry (Fig 1A), a result confirmed by Western blot (Fig 1B). All six "STAT3-defective" (STAT3-def) patients lacked deleterious mutations in known VEOIBD genes as determined by whole exome sequencing. Principal component analysis of blood transcriptome data showed the STAT3-def and IL10R deficient patients generally clustered together (Fig 1C). Cell-type de-convolution analysis of the blood transcriptomes was used to infer cell-type abundance and cell-type specific differentially expressed genes (DEGs) for the STAT3-def and IL10R deficient groups relative to the rest. Genes downregulated in estimated M1 macrophages from the STAT3-def group were enriched in the Hallmark pathway "IL6-JAK-STAT3 signaling." STAT3-def M1 macrophage DEGs were then projected on an adult colonic Crohn’s disease Bayesian network and extended out a path length of 1. The resulting STAT3-def subnetwork of 440 genes included the known VEOIBD genes STAT3, STAT1, TYMP, and TRIM22 (Fig 1D) and the top enriched signaling pathways included IL23, IL6, and IL12. This is of significant interest given three STAT3-def patients with disease refractory to anti-TNF and vedolizumab achieved remission with anti-IL23-based therapies (Table 1). Conclusion We identified 6 patients with refractory VEOIBD and a shared biochemical phenotype and blood transcriptional signature. Network analysis revealed signaling pathways targeted by existing IBD medications, including anti-IL23-based therapies. Molecular analysis of IBD patient blood has exciting potential to localize pathway-level dysfunction and guide precision medicine approaches.

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