Abstract Background Perianal Crohn’s disease (pCD) is an aggressive IBD phenotype characterised by fistula formation. Anti-TNF agents are the gold standard therapy; however janus kinase (JAK) inhibitors, particularly the JAK1 inhibitor upadacitinib, have emerged as promising alternatives.1 We present: 1) the transcriptomic impact of upadacitinib on perianal fistula immune cells; 2) the spatial distribution of activated JAK1 in fistula; and 3) a case series demonstrating promising results of upadacitinib in pCD patients. Methods Single-cell RNA sequencing (scRNA-seq): CD45+ cells were isolated from fistula curettings from 12 pCD patients and 7 idiopathic fistula controls. In 5 pCD donors, paired cultures were incubated with or without 1μM upadacitinib prior to sequencing. Cell types were annotated, cytokine responses inferred from organoid transcriptomic signatures,2 and pathway activity assessed by PROGENy and gene set enrichment analysis. Immunohistochemistry (IHC): pCD resections (n = 3) were stained for activated JAK1 and assessed by a specialist histopathologist. Case series: 20 patients with upadacitinib-treated pCD at St. Mark’s Hospital and Mount Sinai were reviewed with local institutional approvals. Outcomes were assessed retrospectively by clinic notes and imaging. Clinical remission was defined by complete resolution of fistula discharge; clinical response was defined by an improvement in fistula discharge or pain; and radiological outcomes were defined according to recent TOpClass definitions.3 Results scRNA-seq revealed 18 immune cell populations. Interferon-γ response signatures were enriched in pCD versus controls across T-cell, B-cell, and myeloid subsets. PROGENy suggested heightened JAK-STAT activity in pCD, which was attenuated by upadacitinib. Downregulation of interferon-α/γ related gene sets was observed with treatment (Figure 1). IHC demonstrated dense JAK1 staining in the rectal mucosa and deeper fistula tract in pCD resections, suggesting JAK1-associated inflammation present throughout fistula tracts (Figure 2). In our series, 9 patients (45%) achieved clinical remission, and a further 5 patients (25%) clinical response, with upadacitinib over a median follow-up period of 15-months (range 6.0–33.5). Radiological healing or response was seen in 11 (58%) of the 19 patients with pre- and post- treatment MRIs. Conclusion Our data highlight interferon-driven, JAK-STAT–mediated immune activation in pCD. Upadacitinib suppressed these pathways at single-cell resolution, with IHC validating activated JAK1 within fistulae, and our case series suggesting real-world benefit. Together, these findings support JAK1 inhibition as a promising strategy in pCD, warranting further clinical evaluation. References: 1)Colombel JF, Lacerda AP, Irving PM, et al. Efficacy and Safety of Upadacitinib for Perianal Fistulizing Crohn’s Disease: A Post Hoc Analysis of 3 Phase 3 Trials. Clin Gastroenterol Hepatol. 2025 May;23(6):1019-1029.2. 2)Pavlidis P, Tsakmaki A, Treveil A, et al. Cytokine responsive networks in human colonic epithelial organoids unveil a molecular classification of inflammatory bowel disease. Cell Rep. 2022 Sep 27;40(13):111439. 3)Anand E, Devi J, Ballard DH, et al. Defining Radiological Healing in Perianal Fistulizing Crohn’s Disease: a TOpClass Global Expert Delphi Consensus. Clin Gastroenterol Hepatol. 2025 Apr 8:S1542-3565(25)00248-4. Conflict of interest: Hanna, Luke: No conflict of interest Constable, Laura: No conflict of interest Cozzetto, Domenico: No conflict of interest Anandabaskaran, Sulak: Received European Crohn’s Colitis Organisation Research Grant in 2022 and has received conference attendance support from AbbVie, Johnson & Johnson, Dr Falk and Ferring. Shakweh, Eathar: None. Kudo, Hiromi: No conflict of interest Quraishi, Baseer: No conflict of interest Pelly, Theo: No conflict of interest Rekopoulou, Katerina: No conflict of interest Mattews, Nik: No conflict of interest Altinmakas, Emre: No conflict of interest Martinez, Adriana: No conflict of interest Goldin, Rob: None Sebastian, Shaji: Grant: Takeda, Tillots pharma, Biogen, Pfizer, Abbvie, Johnson & Johnson, Olympus -Odin Vision Personal Fees: Tillots, Johnson & Johnson, Olympus Odin Vision, AbbVie, Takeda, Merck, Pharmacosmos, Amgen, Eli Lilly, BMS, Odin Vision Non-financial Support: Tillots, Takeda, AbbVie, Celltrion, Johnson & Johnson, Eli Lilly, Alphasigma, Ferring Pharma Lung, Phillip FC: No conflict of interest Wong, Serre-Yu: Other: Takeda: Research agreement and funding Hart, Ailsa: Grant: Takeda Personal Fees: Abbvie, Amgen, Arena, AZ, Falk, Celltrion, Eli Lilly, Ferring, Genentech/ Roche, GSK, Pfizer, Takeda, Napp, Pharmacosmos, Janssen (J&J), Bristol-Myers Squibb, Gilead, Galapagos Tozer, Philip: Personal Fees: Takeda - speakers fees, member of Inspire, and advisory boards Ferring - speakers fees Falk - speakers fees Tillott’s - speakers fees J & J - speakers fees Abbvie - speakers fees Powell, Nick: Grant: Takeda, BMS, Pfizer, Astra-Zeneca Personal Fees: Abbvie, Abivax, Allergan, Astra-Zeneca, Bristol-Myers Squibb, Celgene, Celltrion, Dr Falk Pharma UK Ltd, Ferring, Galapagos, GSK, Janssen, MSD, Roche, Pfizer, Sobi, Takeda, Tillotts
Hanna et al. (Thu,) studied this question.