Add-on multiple submucosal injections of the RNA oligonucleotide GUT-1 to anti-TNF antibody treatment in patients with moderate-to-severe ulcerative colitis: an open-label, proof-of concept study.

BACKGROUND: Carbohydrate sulfotransferase 15 (CHST15) is an enzyme biosynthesizing matrix glycosaminoglycan that modulates tissue remodeling. We evaluated the efficacy of add-on submucosal injections of GUT-1, the RNA oligonucleotide inhibitor of CHST15, to ongoing anti-tumor necrosis factor (TNF) antibody treatment in patients with moderate-to-severe ulcerative colitis (UC). METHODS: This was an open-label study of 250 nM of GUT-1 by endoscopic submucosal injections at weeks 0, 2, 4 in five UC patients who lost response during maintenance treatment to anti-TNF antibodies. The primary endpoint was the rate of endoscopic improvement at week 6 and secondary endpoints included the rates of clinical remission by modified Mayo Score (mMS). Patients received follow-up observation with continuous maintenance treatment by the same anti-TNF antibody till the time of clinical recurrence or for overall 52 weeks. RESULTS: At week 6, rates of endoscopic improvement and clinical remission were 80% (n = 4/5) and 60% (n = 3/5), respectively. The mean Endoscopy Subscore was reduced from 2.4 (95%CI: 1.7 to 3.1) at baseline, to 1.0 (95%CI: 0.1 to 1.9) at week 6. The mean mMS was reduced from 7.8 (95%CI: 6.2 to 9.4) to 1.3 (95%CI: 2.9 to 4.3). GUT-1 was well tolerated. Three patients did not show clinical recurrence for 52 weeks. All three corticosteroid-dependent patients showed no corticosteroid exposure for at least 24 weeks after achieving clinical remission. Multiple dosing was also well tolerated. CONCLUSIONS: Add-on multiple injections of GUT-1 to ongoing anti-TNF antibody was able to induce rapid and durable clinical responses in UC patients who lost response to anti-TNF therapy. TRIAL REGISTRATION: Clinical trial Registration Number (Japan): UMIN000020900.

Clinical Pharmacist Counselling Improves Long-term Medication Safety and Patient-reported Outcomes in Anti-TNF-treated Patients With Inflammatory Bowel Diseases: The Prospective, Randomized AdPhaNCED Trial.

BACKGROUND: Antitumor necrosis factor (anti-TNF) antibody treatment has led to marked improvements in the management of patients with inflammatory bowel diseases (IBDs). Nevertheless, anti-TNF therapy is associated with potential adverse drug reactions (ADRs). Our prospective, randomized trial investigated the effect of intensified clinical pharmacist counselling in a multidisciplinary team on medication safety in anti-TNF-treated IBD patients. METHODS: Patients with IBD with ongoing anti-TNF treatment were enrolled in our tertiary center AdPhaNCED trial and randomized to either receive conventional standard of care (control group) or additional clinical pharmacist counselling (intervention group) over 12 months. The primary end point consisted of the number and severity of ADRs associated with anti-TNF therapy. Secondary end points included patient satisfaction with medication information and medication safety. RESULTS: One hundred twenty-seven IBD patients were included in this study. Anti-TNF-related ADRs were significantly lower in the intervention compared with the control group (0.20 vs 0.32 [mean] ADR/patient/month, P = .006) after 12 months. The risk of more severe ADRs (Common Terminology Criteria for Adverse Events [CTCAE] grade ≥2) was significantly higher in the control compared with the intervention group (hazard ratio, 0.34; P = .001). The probability of ADR resolution (hazard ratio, 2.02; P < .001) and patient satisfaction with medication information (14.82 vs 11.60; P < .001) were significantly higher in the intervention group compared with the control group. CONCLUSIONS: Our study results demonstrate that intensified pharmacist counselling significantly reduces the occurrence and severity of therapy-related ADRs and improves patient satisfaction. Clinical pharmacists should therefore be part of a holistic approach to IBD care delivered by a multidisciplinary team.

The prospective, randomized AdPhaNCED trial demonstrated that anti-TNF-treated IBD patients had diminished and less severe drug-related adverse reactions and higher patient satisfaction when they received intensified pharmacist counselling in comparison with conventional standard of care over 12 months.

Comparative efficacy and safety of subcutaneous infliximab and vedolizumab in patients with Crohn's disease and ulcerative colitis included in randomised controlled trials.

BACKGROUND: While indirect comparison of infliximab (IFX) and vedolizumab (VDZ) in adults with Crohn's disease (CD) or ulcerative colitis (UC) shows that IFX has better effectiveness during induction, and comparable efficacy during maintenance treatment, comparative data specific to subcutaneous (SC) IFX (i.e., CT-P13 SC) versus VDZ are limited. AIM: Pooled analysis of randomised studies to compare efficacy and safety with IFX SC and VDZ in moderate-to-severe inflammatory bowel disease. METHODS: Parallel-group, randomised studies evaluating IFX SC and VDZ in patients with moderate-to-severe CD or UC were identified. Eligible studies reported ≥ 1 prespecified outcome of interest at Week 6 (reflecting treatment during the induction phase) and/or at 1 year (Weeks 50-54; reflecting treatment during the maintenance phase). Prespecified efficacy and safety outcomes considered in this pooled analysis included the proportions of patients achieving disease-specific clinical responses, clinical remission, or discontinuing due to lack of efficacy, and the proportions of patients experiencing adverse events (AEs), serious AEs, infections, serious infections, or discontinuing due to AEs. Data from multiple studies or study arms were extracted and pooled using a random-effect model; comparative analyses were performed separately for patients with CD and UC. RESULTS: We identified three eligible CD trials and four eligible UC trials that assigned over 1200 participants per disease cohort to either IFX SC or VDZ. In patients with CD, intravenous induction therapy with IFX demonstrated better efficacy (non-overlapping 95% confidence intervals [CIs]) compared with VDZ; during the maintenance phase, IFX SC showed numerically better efficacy (overlapping 95% CIs) than VDZ. A lower proportion of IFX SC-treated patients discontinued therapy due to lack of efficacy over 1 year. In patients with UC, efficacy profiles were similar with IFX SC and VDZ during the induction and maintenance phases, and a lower proportion of IFX SC-treated patients discontinued therapy due to lack of efficacy over 1 year. In both cohorts, safety profiles for IFX SC and VDZ were generally comparable during 1 year. CONCLUSION: IFX SC demonstrated better efficacy than VDZ in patients with CD, and similar efficacy to VDZ in patients with UC; 1-year safety was comparable with IFX SC and VDZ.

Upadacitinib Reduces Crohn's Disease Symptoms Within the First Week of Induction Therapy.

BACKGROUND & AIMS: Upadacitinib (UPA), an oral Janus kinase inhibitor, is approved to treat moderately to severely active Crohn's disease (CD). Because symptomatic response is an important initial treatment goal for patients, we evaluated the rapidity of symptomatic improvement in patients with CD receiving UPA 45 mg once daily (UPA45) induction therapy. METHODS: This post hoc analysis included pooled data from 2 phase 3, multicenter, double-blind, 12-week induction trials (U-EXCEL and U-EXCEED) and 1 maintenance trial (U-ENDURE). Daily diary data for the first 15 days of UPA45 or placebo (PBO) treatment were used to analyze improvement in very soft/liquid stool frequency (SF) and abdominal pain score (APS). Clinical outcomes were evaluated at every study visit. RESULTS: Overall, 1021 patients (n = 674 UPA45; n = 347 PBO) were analyzed. UPA45 demonstrated greater efficacy vs PBO for SF <3 and APS ≤1, providing rapid relief by day 5 or 6, regardless of prior biologic exposure. Mean changes in SF and APS were greater with UPA45 beginning at week 2 (-2.0 and -0.5, respectively; P < .001) and were maintained through week 12 (-3.0 and -1.0, respectively; P < .001) vs PBO. The first achievement of daily SF/APS clinical remission occurred earlier with UPA45 (median, 13 d) vs PBO (median, 32 d), and patients treated with UPA45 showed improved rates of SF/APS clinical remission (21.1% UPA45 vs 8.9% PBO) and clinical response (58.8% UPA45 vs 37.9% PBO) starting at week 2 (both P ≤ .01). CONCLUSIONS: UPA45 provided rapid relief of clinical symptoms within the first week of treatment in patients with CD. CLINICALTRIALS: gov numbers: NCT03345849, NCT03345836, and NCT03345823.

Upadacitinib Achieves Clinical and Endoscopic Outcomes in Crohn's Disease Regardless of Prior Biologic Exposure.

BACKGROUND & AIMS: Upadacitinib, an oral Janus kinase inhibitor, achieved significantly higher rates of clinical remission and endoscopic response vs placebo during induction (U-EXCEL [NCT03345849], U-EXCEED [NCT03345836]) and maintenance (U-ENDURE [NCT03345823]) treatment in patients with moderate-to-severe Crohn's disease. Prior biologic failure is often associated with reduced responses to subsequent therapies. This post hoc analysis assessed upadacitinib efficacy by prior biologic failure status. METHODS: Patients were randomized to placebo or upadacitinib 45 mg (UPA45) for 12 weeks (induction). UPA45 clinical responders were enrolled in U-ENDURE and rerandomized to placebo, upadacitinib 15 mg, or upadacitinib 30 mg (UPA30) for 52 weeks. Assessments were by prior biologic failure. RESULTS: Of 1021 patients, 733 (71.8%) had prior biologic failure. Across outcomes and subgroups, upadacitinib-treated patients achieved higher rates vs placebo. During induction, upadacitinib had higher rates vs placebo for clinical remission based on stool frequency/abdominal pain score (without failure: 54.0% vs 28.3%; with failure: 42.2% vs 14.1%) and endoscopic response (without failure: 52.0% vs 16.2%; with failure: 35.7% vs 5.3%). In maintenance, the greatest treatment effect (upadacitinib vs placebo) was among patients with prior biologic failure treated with UPA30 (clinical remission without failure: 58.5% vs 32.7%; with failure: 42.5% vs 8.7%; endoscopic response without failure: 43.9% vs 17.9%; with failure: 38.9% vs 4.0%). Patients without vs with prior biologic failure had fewer adverse events. CONCLUSIONS: Upadacitinib led to higher absolutes rates of clinical and endoscopic outcomes in patients without vs with prior biologic failure. Patients treated with upadacitinib achieved greater rates of clinical and endoscopic improvements vs placebo, regardless of prior biologic exposure. CLINICALTRIALS: gov: NCT03345849, NCT03345836, NCT03345823.

Biomarkers for Personalizing IBD Therapy: The Quest Continues.

Despite recent advances in the understanding of the pathogenesis of inflammatory bowel diseases (IBD) and advent of multiple targeted therapies, approximately one-third of patients are primary non-responders to initiated treatment, and half of patients lose response over time. There is currently a lack of available biomarkers that would prognosticate therapeutic effectiveness of these advanced therapies. This is partly explained by insufficient characterization of the functional roles assumed by the chosen molecular targets during disease treatment. There is a dire need for validated objective biomarkers, which could be indicators of a biological process, that can be applied in clinical practice to assist us in assigning therapies to patients with the highest probability of response. An appropriate molecular and cellular characterization that accounts for the interindividual differences in drug efficacy and potential side effects would help to guide clinicians in the management of patients with IBD and represent a major step to tailor a more personalized approach to treatment. An appropriate combination of complementing biomarkers should ideally incorporate a multimodal analysis in which genetic, microbial, transcriptional, proteomic, metabolic, and immunologic data are combined to enable a truly personalized approach. This would classify patients into disease subgroups according to molecular characteristics, which would enable us to initiate the most appropriate therapeutic substance. Emergence of single-cell technologies to map the intestinal cellular landscape and multiomic approaches have helped to further dissect the pathogenic mechanisms of mucosal inflammation, but the clinical translation of potential biomarkers remains cumbersome, and an ongoing concerted effort by the IBD community is required.

Etrolizumab-s fails to control E-Cadherin-dependent co-stimulation of highly activated cytotoxic T cells.

Despite promising preclinical and earlier clinical data, a recent phase III trial on the anti-ß7 integrin antibody etrolizumab in Crohn's disease (CD) did not reach its primary endpoint. The mechanisms leading to this outcome are not well understood. Here we characterize the ß7+ T cell compartment from patients with CD in comparison to cells from individuals without inflammatory bowel disease. By flow cytometric, transcriptomic and functional profiling of circulating T cells, we find that triple-integrin-expressing (α4+ß7+ß1hi) T cells have the potential to home to the gut despite α4ß7 blockade and have a specific cytotoxic signature. A subset of triple-integrin-expressing cells readily acquires αE expression and could be co-stimulated via E-Cadherin-αEß7 interactions in vitro. Etrolizumab-s fails to block such αEß7 signalling at high levels of T cell stimulation. Consistently, in CD patients treated with etrolizumab, T cell activation correlates with cytotoxic signatures. Collectively, our findings might add one important piece to the puzzle to explain phase III trial results with etrolizumab, while they also highlight that αEß7 remains an interesting target for future therapeutic approaches in inflammatory bowel disease.

ATP citrate lyase (ACLY)-dependent immunometabolism in mucosal T cells drives experimental colitis in vivo.

OBJECTIVE: Mucosal T cells play a major role in inflammatory bowel disease (IBD). However, their immunometabolism during intestinal inflammation is poorly understood. Due to its impact on cellular metabolism and proinflammatory immune cell function, we here focus on the enzyme ATP citrate lyase (ACLY) in mucosal T cell immunometabolism and its relevance for IBD. DESIGN: ACLY expression and its immunometabolic impact on colitogenic T cell function were analysed in mucosal T cells from patients with IBD and in two experimental colitis models. RESULTS: ACLY was markedly expressed in colon tissue under steady-state conditions but was significantly downregulated in lamina propria mononuclear cells in experimental dextran sodium sulfate-induced colitis and in CD4+ and to a lesser extent in CD8+ T cells infiltrating the inflamed gut in patients with IBD. ACLY-deficient CD4+ T cells showed an impaired capacity to induce intestinal inflammation in a transfer colitis model as compared with wild-type T cells. Assessment of T cell immunometabolism revealed that ACLY deficiency dampened the production of IBD-relevant cytokines and impaired glycolytic ATP production but enriched metabolites involved in the biosynthesis of phospholipids and phosphatidylcholine. Interestingly, the short-chain fatty acid butyrate was identified as a potent suppressor of ACLY expression in T cells, while IL-36α and resolvin E1 induced ACLY levels. In a translational approach, in vivo administration of the butyrate prodrug tributyrin downregulated mucosal infiltration of ACLYhigh CD4+ T cells and ameliorated chronic colitis. CONCLUSION: ACLY controls mucosal T cell immunometabolism and experimental colitis. Therapeutic modulation of ACLY expression in T cells emerges as a novel strategy to promote the resolution of intestinal inflammation.

Long-term outcomes of cyclosporin induction and ustekinumab maintenance combination therapy in patients with steroid-refractory acute severe ulcerative colitis.

Background: Effective management of patients with acute severe ulcerative colitis (ASUC) is a major challenge and there remains a paucity of available maintenance treatment options after efficacious cyclosporin induction therapy. Objectives: We investigated the long-term effectiveness and safety of cyclosporin and ustekinumab combination therapy in patients with steroid refractory ASUC. Design: Monocentric, prospective study. Methods: We included patients with steroid refractory ASUC with multiple failed prior advanced therapies, who were treated with cyclosporin and ustekinumab combination therapy. Results: Among the 11 included patients, 10 had prior failure to infliximab and 8 failed at least three previous biological therapies. The mean baseline Mayo and Lichtiger scores were 10.9 (9-12) and 13.3 (11-14), respectively. Ustekinumab was initiated 3.2 weeks (1-8) after initiation of cyclosporin treatment and combination therapy was continued for a mean of 11.5 (4-20) weeks. Clinical response was achieved in six patients at week 16 and clinical steroid-free clinical remission in five patients at week 48. Endoscopic remission was achieved in five patients at week 16 and together with histological remission in five patients at week 52. Intestinal ultrasound demonstrated mean bowel wall thickening in the sigmoid colon of 5.5 mm at baseline and 3.5 mm at week 52, respectively. Two patients had to undergo colectomy (mean 4.5 months, range 3-6) and three stopped ustekinumab therapy due to ineffectiveness. Overall, combination therapy was well tolerated. Conclusion: Combination of cyclosporin and ustekinumab therapy allowed nearly half of ASUC patients to reach clinical and endoscopic remission after 52 weeks, warranting further studies. Trial registration: Not applicable.

Effects of cyclosporin and ustekinumab combination therapy in acute severe ulcerative colitis In this study, we looked at how to treat patients with a severe form of ulcerative colitis, a type of inflammatory bowel disease, when the usual treatments don't work. We tested a combination of two drugs, cyclosporine and ustekinumab, to see if it could help these patients in the long term. We included eleven patients who had already tried many other treatments and didn't get better. Most of them had also tried a drug called infliximab and had failed at least three other biological therapies. At the start, these patients were very sick, with high scores on disease activity measures. We gave them ustekinumab in addtion after a therapy with cyclosporin had been started before. The combination therapy continued for an average of almost 12 weeks. After 16 weeks, six patients showed improvement in their symptoms, and five were able to stop taking steroids. Five patients also had their colon lining looking healthy again when we looked inside with a scope after 16 weeks. And after 52 weeks, five patients had normal colon lining and healthy tissue under the microscope. Ultrasound showed that the thickness of their colon wall had decreased. Unfortunately, two patients had to have surgery to remove their colon, and three had to stop taking ustekinumab because it didn't help them. Overall, the combination therapy was safe and well-tolerated. In conclusion, combining cyclosporin and ustekinumab helped about half of the patients with severe ulcerative colitis get better and have healthy colon lining after 52 weeks. This suggests that more research is needed to understand the benefits of this treatment in these patients.

Blocking GPR15 counteracts integrin-dependent T cell gut homing in vivo.

BACKGROUND AND AIMS: The G protein coupled receptor GPR15 is expressed on and functionally important for T cells homing to the large intestine. However, the precise mechanisms by which GPR15 controls gut homing have been unclear. Thus, we aimed to elucidate these mechanisms as well as to explore the potential of targeting GPR15 for interfering with T cell recruitment to the colon in IBD. METHODS: We used dynamic adhesion and transmigration assays as well as a humanized in vivo model of intestinal cell trafficking to study GPR15-dependent effects on gut homing. Moreover, we analysed GPR15 and integrin expression in patients with and without IBD cross-sectionally and longitudinally. RESULTS: GPR15 controlled T cell adhesion to MAdCAM-1 and VCAM-1 upstream of α4ß7 and α4ß1 integrin, respectively. Consistently, high co-expression of these integrins with GPR15 was found on T cells from patients with IBD and GPR15 also promoted T cell recruitment to the colon in humanized mice. Anti-GPR15 antibodies effectively blocked T cell gut homing in vitro and in vivo. In vitro data as well as observations in a cohort of patients treated with vedolizumab suggest that this might be more effective than inhibiting α4ß7. CONCLUSIONS: GPR15 seems to have a broad, but organ-selective impact on T cell trafficking and is therefore a promising target for future therapy of IBD. Further studies are needed.

Myenteric Plexus Immune Cell Infiltrations and Neurotransmitter Expression in Crohn's Disease and Ulcerative Colitis.

BACKGROUND AND AIMS: Pain is a cardinal symptom in inflammatory bowel disease [IBD]. An important structure in the transduction of pain signalling is the myenteric plexus [MP]. Nevertheless, IBD-associated infiltration of the MP by immune cells lacks in-depth characterisation. Herein, we decipher intra- and periganglionic immune cell infiltrations in Crohn´s disease [CD] and ulcerative colitis [UC] and provide a comparison with murine models of colitis. METHODS: Full wall specimens of surgical colon resections served to examine immune cell populations by either conventional immuno-histochemistry or immunofluorescence followed by either bright field or confocal microscopy. Results were compared with equivalent examinations in various murine models of intestinal inflammation. RESULTS: Whereas the MP morphology was not significantly altered in IBD, we identified intraganglionic IBD-specific B cell- and monocyte-dominant cell infiltrations in CD. In contrast, UC-MPs were infiltrated by CD8+ T cells and revealed a higher extent of ganglionic cell apoptosis. With regard to the murine models of intestinal inflammation, the chronic dextran sulphate sodium [DSS]-induced colitis model reflected CD [and to a lesser extent UC] best, as it also showed increased monocytic infiltration as well as a modest B cell and CD8+ T cell infiltration. CONCLUSIONS: In CD, MPs were infiltrated by B cells and monocytes. In UC, mostly CD8+ cytotoxic T cells were found. The chronic DSS-induced colitis in the mouse model reflected best the MP-immune cell infiltrations representative for IBD.

NLRP3 Inhibition Leads to Impaired Mucosal Fibroblast Function in Patients with Inflammatory Bowel Diseases.

BACKGROUND AND AIMS: Inflammatory bowel diseases (IBD) are characterized by mucosal inflammation and sequential fibrosis formation, but the exact role of the hyperactive NLRP3 inflammasome in these processes is unclear. Thus, we studied the expression and function of the NLRP3 inflammasome in the context of inflammation and fibrosis in IBD. METHODS: We analysed intestinal NLRP3 expression in mucosal immune cells and fibroblasts from IBD patients and NLRP3-associated gene expression via single-cell RNA sequencing and microarray analyses. Furthermore, cytokine secretion of NLRP3 inhibitor treated blood and mucosal cells, as well as proliferation, collagen production, and cell death of NLRP3 inhibitor treated intestinal fibroblasts from IBD patients were studied. RESULTS: We found increased NLRP3 expression in the inflamed mucosa of IBD patients and NLRP3 inhibition led to reduced IL-1ß and IL-18 production in blood cells and diminished the bioactive form of mucosal IL-1ß. Single cell analysis identified overlapping expression patterns of NLRP3 and IL-1ß in classically activated intestinal macrophages and we also detected NLRP3 expression in CD163+ macrophages. In addition, NLRP3 expression was also found in intestinal fibroblasts from IBD patients. Inhibition of NLRP3 led to reduced proliferation of intestinal fibroblasts, which was associated with a marked decrease in production of collagen type I and type VI in IBD patients. Moreover, NLRP3 inhibition in intestinal fibroblasts induced autophagy, a cellular process involved in collagen degradation. CONCLUSIONS: In the presented study, we demonstrate that inhibiting NLRP3 might pave the way for novel therapeutic approaches in IBD, especially to prevent the severe complication of intestinal fibrosis formation.

p21 Prevents the Exhaustion of CD4+ T Cells Within the Antitumor Immune Response Against Colorectal Cancer.

BACKGROUND & AIMS: T cells are crucial for the antitumor response against colorectal cancer (CRC). T-cell reactivity to CRC is nevertheless limited by T-cell exhaustion. However, molecular mechanisms regulating T-cell exhaustion are only poorly understood. METHODS: We investigated the functional role of cyclin-dependent kinase 1a (Cdkn1a or p21) in cluster of differentiation (CD) 4+ T cells using murine CRC models. Furthermore, we evaluated the expression of p21 in patients with stage I to IV CRC. In vitro coculture models were used to understand the effector function of p21-deficient CD4+ T cells. RESULTS: We observed that the activation of cell cycle regulator p21 is crucial for CD4+ T-cell cytotoxic function and that p21 deficiency in type 1 helper T cells (Th1) leads to increased tumor growth in murine CRC. Similarly, low p21 expression in CD4+ T cells infiltrated into tumors of CRC patients is associated with reduced cancer-related survival. In mouse models of CRC, p21-deficient Th1 cells show signs of exhaustion, where an accumulation of effector/effector memory T cells and CD27/CD28 loss are predominant. Immune reconstitution of tumor-bearing Rag1-/- mice using ex vivo-treated p21-deficient T cells with palbociclib, an inhibitor of cyclin-dependent kinase 4/6, restored cytotoxic function and prevented exhaustion of p21-deficient CD4+ T cells as a possible concept for future immunotherapy of human disease. CONCLUSIONS: Our data reveal the importance of p21 in controlling the cell cycle and preventing exhaustion of Th1 cells. Furthermore, we unveil the therapeutic potential of cyclin-dependent kinase inhibitors such as palbociclib to reduce T-cell exhaustion for future treatment of patients with colorectal cancer.

Submucosal Injection of the RNA Oligonucleotide GUT-1 in Active Ulcerative Colitis Patients: A Randomized, Double-Blind, Placebo-Controlled Phase 2a Induction Trial.

BACKGROUND AND AIMS: Carbohydrate sulfotransferase 15 [CHST15] biosynthesizes sulphated matrix glycosaminoglycans and is implicated in intestinal inflammation and fibrosis. Here, we evaluate the efficacy and safety of the double-stranded RNA oligonucleotide GUT-1, a specific blocker of CHST15, as induction therapy in patients with ulcerative colitis [UC]. METHODS: In this randomized, double-blind, placebo-controlled, phase 2a study, we enrolled endoscopically active UC patients, refractory to conventional therapy, in five hospital centres across Germany. Patients were randomized 1:1:1 using a block randomized technique to receive a single dosing of 25 nM GUT-1, 250 nM GUT-1, or placebo by endoscopic submucosal injections. The primary outcome measure was improvement of endoscopic lesions at weeks 2 or 4. The secondary outcome measures included clinical and histological responses. Safety was assessed in all patients who received treatment. RESULTS: Twenty-eight patients were screened, 24 were randomized, and 21 were evaluated. Endoscopic improvement at weeks 2 or 4 was achieved by 71.4% in the GUT-1 250 nM, 0% in the GUT-1 25 nM, and 28.6% in the placebo group. Clinical remission was shown by 57.1% in the GUT-1 250 nM, 0% in the GUT-1 25 nM, and 14.3% in the placebo groups. Histological improvement was shown by 42.9% in the GUT-1 250 nM, 0% in the GUT-1 25 nM, and 0% in the placebo groups. GUT-1 250 nM reduced CHST15 expression significantly and suppressed mucosal inflammation and fibrosis. GUT-1 application was well tolerated. CONCLUSION: Single dosing by submucosal injection of GUT-1 repressed CHST15 mucosal expression and may represent a novel induction therapy by modulating tissue remodelling in UC.

IL-20 controls resolution of experimental colitis by regulating epithelial IFN/STAT2 signalling.

OBJECTIVE: We sought to investigate the role of interleukin (IL)-20 in IBD and experimental colitis. DESIGN: Experimental colitis was induced in mice deficient in components of the IL-20 and signal transducer and activator of transcription (STAT)2 signalling pathways. In vivo imaging, high-resolution mini-endoscopy and histology were used to assess intestinal inflammation. We further used RNA-sequencing (RNA-Seq), RNAScope and Gene Ontology analysis, western blot analysis and co-immunoprecipitation, confocal microscopy and intestinal epithelial cell (IEC)-derived three-dimensional organoids to investigate the underlying molecular mechanisms. Results were validated using samples from patients with IBD and non-IBD control subjects by a combination of RNA-Seq, organoids and immunostainings. RESULTS: In IBD, IL20 levels were induced during remission and were significantly higher in antitumour necrosis factor responders versus non-responders. IL-20RA and IL-20RB were present on IECs from patients with IBD and IL-20-induced STAT3 and suppressed interferon (IFN)-STAT2 signalling in these cells. In IBD, experimental dextran sulfate sodium (DSS)-induced colitis and mucosal healing, IECs were the main producers of IL-20. Compared with wildtype controls, Il20-/-, Il20ra-/- and Il20rb-/- mice were more susceptible to experimental DSS-induced colitis. IL-20 deficiency was associated with increased IFN/STAT2 activity in mice and IFN/STAT2-induced necroptotic cell death in IEC-derived organoids could be markedly blocked by IL-20. Moreover, newly generated Stat2ΔIEC mice, lacking STAT2 in IECs, were less susceptible to experimental colitis compared with wildtype controls and the administration of IL-20 suppressed colitis activity in wildtype animals. CONCLUSION: IL-20 controls colitis and mucosal healing by interfering with the IFN/STAT2 death signalling pathway in IECs. These results indicate new directions for suppressing gut inflammation by modulating IL-20-controlled STAT2 signals.

Tofacitinib Affects M1-like and M2-like Polarization and Tissue Factor Expression in Macrophages of Healthy Donors and IBD Patients.

BACKGROUND: Tofacitinib, as inhibitor of Janus kinases (JAK), interrupts the transmission of numerous pro-inflammatory cytokines involved in the pathogenesis of inflammatory bowel diseases (IBD). Therefore, tofacitinib provides a potent option to treat ulcerative colitis (UC). Besides the anti-inflammatory potential, inhibition of widespread JAKs carries the risk of side effects. Macrophages are involved in the form of different subtypes in inflammation, wound healing, and even coagulation. This study aimed to explore the balanced use of tofacitinib in M1-like as well as M2-like macrophages of healthy donors and patients with IBD. METHODS: Monocytes of healthy donors and patients with chronic courses of IBD were obtained from blood samples. Macrophage colony-stimulating factor (M-CSF)-derived macrophages were treated with tofacitinib (1 µM, 5 µM, 10 µM) and polarized with either lipopolysaccharide and interferon (IFN)-γ towards M1-like-phenotype or with interleukin (IL)-4 towards M2-like-phenotype. ELISA and flow cytometry were used to evaluate cytokine levels and surface molecules. RESULTS: Tofacitinib had a modulating effect on M1-like macrophages whereby the effect on pro-inflammatory cytokines (TNF-α, IL-6, IL-1ß, IL-12, IL-23) was less pronounced than the induction of anti-inflammatory IL-10. However, during M2-like polarization tofacitinib impaired the development of the corresponding phenotype becoming evident through decreased IL-10 levels and CD206 expression in treated macrophages. In both phenotypes, tofacitinib strongly downregulated the expression of immunostimulatory molecules (CD80, CD86, CD83, CD40). Furthermore, a dose-dependent correlation between treatment with tofacitinib and expressed tissue factor was noticed. CONCLUSIONS: Tofacitinib influences both polarizations (M1/M2) and the expression of tissue factor in a dose-dependent manner.

This study revealed a dose-dependent effect of tofacitinib on both M1-like and M2-like polarization, resulting in a decreased development of the corresponding phenotype. Furthermore, the applied dose of tofacitinib correlated with the expressed tissue factor in M1-like macrophages.

Healing of the epithelial barrier in the ileum is superior to endoscopic and histologic remission for predicting major adverse outcomes in ulcerative colitis.

Background: Achieving endoscopic remission is a key therapeutic goal in patients with ulcerative colitis (UC) that is associated with favorable long-term disease outcomes. Here, we prospectively compared the predictive value of endoscopic and/or histologic remission against ileal barrier healing for predicting long-term disease behavior in a large cohort of UC patients in clinical remission. Methods: At baseline, UC patients in clinical remission underwent ileocolonoscopy with assessment of ileal barrier function by confocal endomicroscopy. Endoscopic and histologic disease activity and ileal barrier healing were scored using validated scores. During subsequent follow-up (FU), patients were closely monitored for clinical disease activity and occurrence of major adverse outcomes (MAO) defined as the following: disease relapse; UC-related hospitalization; UC-related surgery; necessity for initiation or dose escalation of systemic steroids, immunosuppressants, small molecules or biological therapy. Results: Of the 73 UC patients included, 67% experienced MAO during a mean FU of 25 months. The probability of MAO-free survival was significantly higher in UC patients with endoscopic and/or histologic remission compared to patients with endoscopically and/or histologically active disease. Ileal barrier healing on endomicroscopy was highly accurate for predicting the further course of UC and outcompeted endoscopic and histologic remission for predicting MAO-free survival. Conclusion: Ileal barrier healing in clinically remittent UC patients can accurately predict future MAO development and is superior in its predictive capabilities than endoscopic and histologic remission. Ileal barrier healing therefore represents a novel and superior surrogate parameter for stratification of UC patients according to their risk for development of complicated disease behavior. Clinical trial registration: https://classic.clinicaltrials.gov/ct2/show/NCT05157750, identifier NCT05157750.

IL-3 receptor signalling suppresses chronic intestinal inflammation by controlling mechanobiology and tissue egress of regulatory T cells.

IL-3 has been reported to be involved in various inflammatory disorders, but its role in inflammatory bowel disease (IBD) has not been addressed so far. Here, we determined IL-3 expression in samples from patients with IBD and studied the impact of Il3 or Il3r deficiency on T cell-dependent experimental colitis. We explored the mechanical, cytoskeletal and migratory properties of Il3r -/- and Il3r +/+ T cells using real-time deformability cytometry, atomic force microscopy, scanning electron microscopy, fluorescence recovery after photobleaching and in vitro and in vivo cell trafficking assays. We observed that, in patients with IBD, the levels of IL-3 in the inflamed mucosa were increased. In vivo, experimental chronic colitis on T cell transfer was exacerbated in the absence of Il-3 or Il-3r signalling. This was attributable to Il-3r signalling-induced changes in kinase phosphorylation and actin cytoskeleton structure, resulting in increased mechanical deformability and enhanced egress of Tregs from the inflamed colon mucosa. Similarly, IL-3 controlled mechanobiology in human Tregs and was associated with increased mucosal Treg abundance in patients with IBD. Collectively, our data reveal that IL-3 signaling exerts an important regulatory role at the interface of biophysical and migratory T cell features in intestinal inflammation and suggest that this might be an interesting target for future intervention.