{
  "abstract": "Introduction Unlike some advanced therapies for Ulcerative Colitis (UC), upadacitinib (upa), a selective JAK1 inhibitor, is effective in biologic-exposed patients (Danese et al. 2022). IL-22 is a key pathogenic cytokine that signals via JAK1. High IL-22 activity, characterised by increased neutrophil chemotaxis and extracellular matrix (ECM) dysfunction, is associated with ustekinumab and anti-TNF resistance (Pavlidis et al. 2022). Its receptor is expressed mainly on epithelial cells in the gut. We used an epithelial colonic organoid model to explore the effect of upa on these resistance mechanisms.Methods Left-sided colon biopsies were sampled from inactive UC and healthy control patients (HC) undergoing diagnostic endoscopy. Crypts were isolated using published protocols, suspended in Matrigel (Corning, US) and incubated at 37°C (Dotti et al. 2022). Before experimentation, the resulting organoids were differentiated for 72 hours in Wnt-free media. They were exposed for 24 hours to: 0.001% DMSO vehicle control, DMSO + IL-22 10 ng/ml, IL-22 10 ng/ml + 5 μM or 1 μM or 0.1 μM upa. RNA was extracted with Qiagen MiniKit and profiled with Next Generation Sequencing. Differentially expressed genes (DEGs) and pathways were determined. Proteins in the supernatant were quantified using Luminex multiplex panels for immune and fibrosis markers. Analyses were performed in R 4.3.2.Results Following IL-22 stimulation of HC organoids (n=10), there were 2102 DEGs, with 1035 upregulated. These related to pathways involved in protein translation and interferon alpha/beta signalling. IL-22 induced a more pronounced transcriptional response in UC organoids (n=7), including 4900 DEGs, of which 2400 were upregulated. These related to interferon alpha/beta signalling, IL-10 and IL-1 signalling, innate immune processes like complement cascade, and ECM mechanisms like keratinisation. There were 969 and 971 shared up-/downregulated genes, respectively. UC organoid genes were perturbed to a higher degree, up to 8 log fold change and -4 log fold change, respectively, compared to HC. In a dose-dependent manner, upa strongly inhibited the expression of key proinflammatory and treatment resistance transcripts, such as metalloproteinases, that were induced by IL-22 in our model. Similar findings were seen at proteomic level, where upa suppressed key IL-22-inducible proteins, including the chemokines CXCL1 and CXCL10.Conclusions Upa inhibits IL-22-induced resistance mechanisms originating in colonic epithelial cells in UC, potentially contributing to its relative effectiveness in biologic-refractory patients. This complements recent single cell analyses that found upregulation of interferon pathways in anti-TNF non-responders with UC (Thomas et al. 2024). The breadth and degree of transcriptomic changes is more marked in UC compared to HC organoids after cytokine stimulation, suggesting they should be used in organoid research for UC as they more closely mimic in vivo conditions.",
  "authors": [
    {
      "affiliations": [
        "Imperial College London, United Kingdom",
        "St Mark’s Academic Institute, United Kingdom",
        "Maidstone and Tunbridge Wells NHS Trust, United Kingdom"
      ],
      "name": "Aamir Saifuddin"
    },
    {
      "affiliations": [
        "Imperial College London, United Kingdom"
      ],
      "name": "Hajir Ibraheim"
    },
    {
      "affiliations": [
        "Imperial College London, United Kingdom"
      ],
      "name": "Domenico Cozzetto"
    },
    {
      "affiliations": [
        "Imperial College London, United Kingdom"
      ],
      "name": "Neha Panneer-Selvam"
    },
    {
      "affiliations": [
        "Lincoln County Hospital, United Kingdom"
      ],
      "name": "Sahil Misri"
    },
    {
      "affiliations": [
        "Imperial College London, United Kingdom"
      ],
      "name": "Jake Riley"
    },
    {
      "affiliations": [
        "King’s College Hospital NHS Foundation Trust, United Kingdom"
      ],
      "name": "Chronis Pavlidis"
    },
    {
      "affiliations": [
        "St Mark’s Academic Institute, United Kingdom"
      ],
      "name": "Ailsa Hart"
    },
    {
      "affiliations": [
        "Imperial College London, United Kingdom"
      ],
      "name": "Nick Powell"
    }
  ],
  "title": "P175 IL-22-driven inflammatory and drug resistance pathways are JAK1-dependent in a human 3D mini-gut organoid system",
  "uid": "dab9bf33-f71b-580a-9889-f9dae0670812"
}