Introduction

Differential diagnosis of Inflammatory Bowel Disease Unclassified (IBDU) patients to Crohn’s Disease (CD) or Ulcerative Colitis (UC) at diagnosis is challenging. Personalized disease-specific strategies are becoming state of art and therefore it is of importance for outcome of the patients and treatment decision to be taken by the clinicians to have a precise diagnosis.

Aims & Methods

Whole transcriptome RNA sequencing was performed using AmpliSeq human gene expression panel on formalin-fixed paraffin-embedded (FFPE) colonic mucosal biopsies of IBD patients (CD, n=16, UC, n=17) and non-IBD controls (n=10) to study differential gene expression between CD and UC samples. Top 100 of the most significant differentially expressed genes were then tested in two independent external GEO datasets. Functional enrichment analysis of 10 successful candidates were performed and 7 genes were selected for further validation. RT-qPCR analyses were performed on an independent patient cohort of 45 CDs and 53 UCs along with 12 non-IBD controls. The cDNA samples were submitted to quality restriction during the statistical analysis resulting in a cut-off of Ct values below 29 for the endogenous control allowing 46 patient samples to enter the analysis. In situ hybridization (ISH) was performed using RNAscope on 11 selected cases for describing tissue localisation and expression.

Results

Sequencing results showed 308 genes differentially expressed between CD and UC from a total of 14190 genes. The genes MTCL1, SH3PXD2A-AS1, CLCF1, and CD180 were under the limit of detection with RT-qPCR and were excluded from further analysis. Relative expression levels from RT-qPCR for the genes PI3, ANXA1, and VDR, together with the age and sex, resulted in an area under the receiver-operating characteristic curve (AUC) of 0.84 (P = 0.02) in discriminating CD from UC. Significantly higher expression of VDR alone was observed in CD samples with AUCs of 0.77 (P = 0.01). ISH analyses showed expression of PI3, ANXA1, and VDR mRNAs localized to mucosal epithelial cells.

Conclusion

Using whole transcriptome analysis of FFPE sample-derived RNA, we have identified genes that are differentially expressed in CD and UC, and we have developed an RT-qPCR assay including a panel of 3 genes (PI3, ANXA1, VDR) for the differential diagnosis among IBD unclassified (IBDU) patients. We propose that the RT-qPCR assay could be used in a clinical routine setup for distinguishing UC from CD.

Introduction

Inflammatory bowel diseases are chronic disabling conditions with a complex and multifactorial aetiology, which is still not completely understood. OCTN1, an organic cation transporter could have a role in modulating the inflammatory response, whose genetic polymorphisms were associated to increased risk of inflammatory bowel diseases. Up to now, limited information exists on its role in predicting/modulating response to therapies.

Aims & Methods

The aim of this study was to evaluate the role of OCTN1 in modifying gut microbiota and immune composition to Infliximab therapy in murine colitis. Dextran sodium sulphate model of colitis on C57BL/6 mice and ocnt1 gene knockout mice was used to assess clinical efficacy of infliximab administered intravenously. Stool, colon and mesenteric lymph node samples were collected to assess differences in gut microbiota composition, histology for Rachmilewitz score and T cell populations respectively.

Results

Octn1^-/-influences microbiota profile and is associated with a worse dysbiosis in colitis mice. Infliximab treatment attenuates colitis-associated dysbiosis with increase of bacterial richness and evenness in both strains. In comparison with wild type, octn1^-/- mice have a higher baseline percentage of Treg, Tmemory, Th2 and Th17.

Conclusion

Our data support murine model to study OCTN1 genetic asset in inflammatory bowel diseases. This could be the first step to consider this membrane transporter as a biomarker in inflammatory conditions and as a predictor of response to therapies.

Introduction

Small bowel activity detected by endoscopy is known to be associated with unfavourable clinical outcome in Crohn’s disease, regardless of symptoms. The treat-to-target approach whereby endoscopic remission is set as a treatment target is recommended. Imaging modalities for monitoring the small bowel are limited, as well as being invasive.

Aims & Methods

In this study, our objective was to investigate which of the existing biomarkers for Crohn’s disease best indicates endoscopic small bowel activity, and whether combining the biomarkers can improve accuracy. 104 consecutive patients with ileal and ileocolonic type Crohn’s disease who underwent balloon assisted enteroscopy (BAE) at our hospital from October 2021 to August 2022 were enrolled, with clinical and laboratory data prospectively collected and analysed. BAE was evaluated using the modified Simple Endoscopic Score for CD (modified SES-CD). Ulcers were defined as those ≧ 0.5 cm.

Results

Haemoglobin, platelet count, C-reactive protein (CRP), leucine-rich alpha-2 glycoprotein (LRG), faecal calprotectin, and faecal haemoglobin values all demonstrated a significant difference in those with ulcers found on BAE. All biomarkers significantly correlated with modified SES-CD. LRG and faecal calprotectin showed the highest areas under the curve (0.841 and 0.853) by ROC analysis for detecting small bowel ulcers. LRG showed a sensitivity of 78% and specificity of 80% at a cut-off value of 13 μg/mL, whereas faecal calprotectin showed a sensitivity of 91% and specificity of 67% at a cut-off value of 151 μg/g. Dual positivity for LRG and faecal calprotectin, as well as LRG and faecal haemoglobin, both predicted the presence of ulcers with a further improved specificity of 92% and 100%. A positive result for either LRG or faecal calprotectin/haemoglobin showed an improved sensitivity of 96% and 91%. Positivity for LRG and either of the faecal biomarkers was associated with poor clinical outcome in terms of hospitalisation, surgery, and relapse.

Conclusion

This is the first study comparing the major existing biomarkers for Crohn’s disease in terms of monitoring for small bowel activity. LRG, faecal calprotectin, and faecal haemoglobin are accurate biomarkers, especially when used in combination. Patients who are dual-positive for LRG and faecal calprotectin or faecal haemoglobin should be assessed by early endoscopic evaluation of the small bowel and considered for change in treatment, as they are at higher risk for hospitalisation, relapse, and surgery.

Introduction

Carbohydrate sulfotransferase 15 (CHST15) is an enzyme biosynthesizing sulfated matrix glycosaminoglycan that modulates tissue remodeling.

Aims & Methods

We evaluated the efficacy of add-on submucosal injections of GUT-1, the RNA oligonucleotide inhibitor of CHST15, to ongoing tumor necrosis factor (TNF) antagonist treatment in patients with moderate-to-severe ulcerative colitis (UC) and insufficient response to TNF antagonists.
This was an open-label study of 250 nM of GUT-1, which was applied by endoscopic submucosal injections at weeks 0, 2 and 4 in five UC patients who lost response during maintenance treatment to TNF antagonists. The primary endpoint was the rate of endoscopic improvement at week 6 and secondary endpoints included the rates of clinical remission. All patients received follow-up observation with continuous maintenance treatment by the same TNF antagonists till the time of clinical recurrence, defined by rectal bleeding and endoscopic examination, or for overall 48 weeks (Clinical trial Registration Number: UMIN000020900).

Results

At baseline, all included patients showed insufficient clinical and endoscopic response to ongoing TNF antagonist treatment (3 for infliximab and 2 for adalimumab), as measured by modified Mayo Score (mMS) of 7.8 (95%CI: 6.2 to 9.4), Endoscopic Subscore of 2.4 (95%CI: 1.7 to 3.1), Stool Frequency Subscore of 2.8 (95%CI: 2.2 to 3.4) and Rectal Bleeding Subscore of 2.6 (95%CI: 1.9 to 3.3). At week 6, Endoscopic Subscore reduced to 1.0 (95%CI: 0.1 to 1.9) and rate of endoscopic improvement was 80% (n=4/5). mMS reduced to 1.3 (95%CI: 0.4 to 3.2) and rate of clinical remission was 60% (n=3/5). Exploratory, Nancy histological index reduced to 1.8 (95%CI: 0.2 to 3.4) at week 6 from 3.6 (95%CI: 2.9 to 4.3) at baseline. Three patients did not show any signals of clinical recurrence for 48 weeks. All three corticosteroid-dependent patients showed no corticosteroid exposure for at least 20 weeks after achieving clinical remission. During the follow-up period, clinical recurrence was observed in one patient at week 36 and another at week 44.

Conclusion

Add-on multiple injections of GUT-1 to ongoing TNF antagonist treatment was able to induce rapid and durable clinical responses in patients with TNF antagonist-refractory UC. GUT-1 was well tolerated by the patients.

Introduction

Ulcerative colitis (UC) is a refractory inflammatory bowel disease of the colon with repeated relapses and remissions that requires lifelong treatment. Previous studies indicated that proinflammatory cytokines involved the onsets and developments of UC. New treatments for targeting these cytokines, such as TNF-α, IL-12 and IL-23 have been developed in a recent decade, underlying that immune abnormalities are the core for the etiology of UC.
Platelets play a role not only in hemostatic coagulation but in inflammatory mediator by interacting with various cells in the innate and adaptive immunity. Chronic inflammatory response confers platelets activation leading to elevated platelet count and hypercoagulability. Moreover, platelets aggregates with leukocyte in active inflammatory milieu. In fact, thrombocytosis, thromboembolism and upregulation of platelet-monocyte complex (PMC) in UC patients have been reported, implying that platelets could be novel biomarker, therapeutic target and clues for elucidating the pathology of UC. However, the clinical significance of platelet activation and PMC formation in UC is not fully elucidated and few studies were conducted to assess how platelet activation contributes to disease aggravations of UC. Therefore, we comprehensively analyzed platelet activation and platelet-monocyte complex by focusing on a correlation with clinical severity of UC.

Aims & Methods

Patients with UC (n=70) including MES0-2 group (n=34) and MES3 group (n=27), diagnosed according to a combination of symptoms, endoscopic findings, histologic findings, and the absence of alternative diagnoses, and age and sex-matched healthy controls (HCs) (n =19) were enrolled for this study. We analyzed the surface markers such as CD62P, CD40L, CD63, PAC-1 and Annexin V on CD61+ platelets by flowcytometry (FCM) in patients with UC in comparison with those in healthy adults. We also analyzed the proportion of CD61+CD14+ platelet-monocyte complexes and quantified the activation of PMCs by FCM. Moreover, we also investigated the correlation among surface markers, proportion of platelet-monocyte aggregates and Mayo endoscopic score (MES) in UC patients.

Results

The expression of CD62P but not other surface markers on platelets was significantly upregulated in UC when compared to HCs (p<0.01), and was significantly upregulated in in MES3 group when compared to the MES0-2 group (P<0.001). The proportion of PMC was also higher in MES3 groups when compared to MES0-2 group (P<0.05), and was markedly reduced by treatment (P<0.05). In addition, CD16+ monocytes, phenotypically mature and pro-inflammatory cell population, were increased in patients with UC when compared to HCs (P<0.01). The expression of CD16 on monocytes aggregated with platelets was significantly higher when compered to that without platelet (P<0.001).

Conclusion

Our study suggests that platelet activation accelerates mucosal inflammation via adhesion with monocytes in UC and contributes to the aggravation of disease.

References

Semple JW, et al. Platelets and the immune continuum.
Nat Rev Immunol. 2011 Apr;11(4):264-74.
Collins CE, et al. Platelets circulate in an activated state in inflammatory bowel disease.
Gastroenterology. 1994 Apr;106(4):840-5.

Introduction

Inflammatory Bowel Diseases (IBD) are chronic inflammatory disorders characterised by alterations of acute relapses and remission. The past decades have brought substantial advances in the pharmacological management of IBD by the introduction of immunosuppressive agents, biologics and lately small molecules. Although, the sustained efficacy of the currently available anti-inflammatory therapies is still far from optimal. Therefore, the development of novel therapies represents a major unmet clinical need in this field. Recently the enrichment of the coagulation pathway genes and among these, plasminogen activator inhibitor 1 (PAI-1) was described in IBD, whereas PAI-1 emerged as a link between the epithelium and inflammation. Moreover, the genetic deletion of PAI-1 impaired the severity of experimental colitis in mice.

Aims & Methods

Based on these, our aim was to develop and test novel PAI-1 inhibitors for the treatment of IBD. Novel, peptide-based PAI-1 inhibitors were designed and screened in silico, then the selected ones were synthetized using the ixPeptide® platform. The inhibitory potential of the peptides was determined by an in vitro PAI-1 activity assay. Cytotoxicity was monitored in vitro in Hela cells with the CellTiter Glo assay. Based on the in vitro results LDN-071 was selected for further analysis. Experimental colitis was induced in mice with 3% DSS and after the induction of colitis 10 mg/bwkg LDN-071 was administrated per os for four consecutive days. The changes of body weight, spleen weight and colon length were measured. Haematoxylin eosin (HE) and Periodic acid-Schiff (PAS) staining were performed for histology. ELISA and qRT-PCR measurements were applied to analyse the mucosal cytokine expressions. In vivo toxicity of LDN-071 was determined in mice.

Results

LDN-071 significantly inhibited PAI-1 in vitro without decreasing the cell viability in a concentration range of 1µM-1mM. The administration of 10 mg/bwkg LDN-071 in the DSS-treated mice prevented the body weight loss, the development of bloody diarrhoea, the decrease of colon length and increase of the spleen weight, which were displayed by the DSS+vehicle-treated animals. The histological analysis of the colon revealed that DSS treatment induced a severe transmural inflammation, ulceration, loss of crypt structures and goblet cell depletion. Importantly, all of these parameters were significantly impaired by the administration of 10 mg/bwkg LDN-071. Additionally, mucosal gene and protein expressions of TNF-α, IL-1β and IL-6 were significantly impaired by PAI-1 inhibition. Importantly, no signs of toxicity were LDN-071 observed in mice exposed to 100 mg/bwkg LDN-071 for 7 days.

Conclusion

Our results showed that LDN-071, a novel peptide-based inhibitor of PAI-1. It significantly reduces the severity of experimental colitis in mice. We propose that the inhibition of PAI-1 could be a potential novel therapeutic strategy in IBD.

Disclosure

LDN-071 PAI-1 inhibitor is owned by Ladon Therapeutics Ltd.

Introduction

Filgotinib (FIL) is a once-daily, oral, Janus kinase 1 preferential inhibitor approved for the treatment of ulcerative colitis (UC). Previous analyses of data from the phase 2b/3 SELECTION trial (NCT02914522) demonstrated that FIL 200 mg (FIL200) improved health-related quality of life (HRQoL) and led to comprehensive disease control (comprising clinical, biomarker, HRQoL and endoscopic improvements) in patients with UC.¹

Aims & Methods

We evaluated individual Inflammatory Bowel Disease Questionnaire (IBDQ) item data from SELECTION to assess which items drive the benefits of FIL treatment. In SELECTION, adults with moderately to severely active UC were randomized 2:2:1 to receive FIL200, FIL 100 mg or PBO once daily for 11 weeks in Induction Studies A and B (biologic-naive and -experienced patients, respectively). Patients who responded to FIL were rerandomized 2:1 to continue their induction FIL dose in the 47-week Maintenance Study. In this post hoc analysis, standardized effect sizes (SES) were calculated to evaluate the relative magnitude of the difference in mean scores of individual IBDQ items at week 10 (W10) between FIL200 versus PBO-treated patients, and at week 58 (W58) between patients who continued FIL200 treatment (FIL200→FIL200) versus those who were rerandomized to PBO (FIL200→PBO). SES were also calculated for the mean change in score of individual IBDQ items from baseline to W10, and from maintenance baseline to W58. SES were categorized by Cohen’s thresholds (‘trivial’ [<0.2]; ‘small’ [≥0.2–<0.5]; ‘moderate’ [≥0.5–<0.8]; and ‘large’ [≥0.8]). A last observation carried forward imputation approach was used for missing data.

Results

At W10, patients treated with FIL200 overall (n=506) had improvements in all individual IBDQ items versus those treated with PBO (n=277) and had moderate to large increases from baseline (SES ≥0.5) in all individual IBDQ items. The IBDQ items with the five largest SES for FIL200 versus PBO at W10 (analysed by induction study), and for FIL200→FIL200 (n=199) versus FIL200→PBO (n=98) at W58 are summarized in the Table. FIL200 treatment consistently resulted in the largest benefits in items related to pain (‘abdominal pain’ and ‘abdominal cramps’), ability to engage socially (‘satisfied with life’, ‘depressed or discouraged’, ‘social engagement’ and ‘problems with activities’) and bowel urgency (‘loose bowel movements’ and ‘finding a washroom’) versus PBO at W10, and versus FIL200→PBO at W58

Table. Top five individual IBDQ item scores ranked by SES for FIL200 versus PBO groups at W10 and FIL200→FIL200 versus FIL200→PBO groups at W58.

Conclusion

FIL200 treatment provides improvements in HRQoL versus PBO, with large SES seen for most IBDQ items, and the greatest improvements in items related to pain, social engagement and bowel urgency over 58 weeks. This analysis reveals the specific IBDQ items that drive the benefits to HRQoL of FIL and adds further evidence to that showing that FIL contributes to restoration of health through comprehensive disease control for patients with UC.¹

References

1. Schreiber S et al. J Crohns Colitis 2023; https://doi.org/10.1093/ecco-jcc/jjad018

Disclosure

BF reports grants and personal fees from AbbVie, Amgen, AstraZeneca, Bristol Myers Squibb, Janssen Biotech/Centocor, Johnson & Johnson/Janssen, Pfizer, Receptos and Takeda; and personal fees from Ablynx, ActoGeniX, AdMIRx, Inc., Akebia Therapeutics, Inc., Allergan, Atlantic Pharmaceuticals, Avaxia Biologics, Inc., Avir Pharma, Baxter Healthcare Corporation, Biogen Idec, BiomX Israel, Boehringer Ingelheim, Boston Pharmaceuticals, Calypso Biotech, Celgene, Elan/Biogen, Eli Lilly, enGene, Ferring Pharmaceuticals, Galapagos, Genentech/Roche, gIcare Pharma, Gilead, Given Imaging, Gossamer Bio, GSK, Inception IBD, Inc., Ironwood Pharmaceuticals, Japan Tobacco Company, Kyowa Hakko Kirin Co. Ltd, Lexicon Pharmaceuticals, Lycera, Mesoblast, Millennium Pharmaceuticals, MSD, Nestlé, Nextbiotix, Novartis, Novo Nordisk, Par’Immune, Progenity, Prometheus Therapeutics and Diagnostics, Protagonist, Qu Biologics, Salix Pharmaceuticals, Shire, Sienna Biopharmaceuticals, Sigmoid Pharma, Synergy Pharma, Teva Pharmaceuticals, TiGenix, Tillotts Pharma, UCB, Vertex Pharmaceuticals, VHsquared, Vivelix Pharmaceuticals, Wyeth, Zealand Pharma and Zyngenia.
SV reports grants from AbbVie, Galapagos, J&J, Pfizer and Takeda; consulting and/or speaker fees from AbbVie, AbolerIS Pharma, AgomAb, Alimentiv, Arena Pharmaceuticals, AstraZeneca, Avaxia Biologics, Bristol Myers Squibb, Boehringer Ingelheim, Celgene, CVasThera, Cytoki Pharma, Dr Falk Pharma, Ferring Pharmaceuticals, Galapagos, Genentech/Roche, Gilead, GSK, Hospira, Imidomics, Janssen, J&J, Lilly, Materia Prima, MiroBio, Morphic, MrMHealth, Mundipharma, MSD, Pfizer, Prodigest, Progenity, Prometheus, Robarts Clinical Trials, Second Genome, Shire, Surrozen, Takeda, Theravance Biopharma, Tillotts Pharma and Zealand Pharma.
LP-B reports speaking fees from AbbVie, Amgen, Arena Pharmaceuticals, Biogen, Celltrion, Eli Lilly, Ferring Pharmaceuticals, Galapagos, Genentech, Gilead, Janssen Pharmaceuticals, Medac Pharma, MSD, Pfizer, Sandoz, Takeda, Tillotts Pharma, Viatris and Vifor Pharma;grants from Celltrion, Fresenius Kabi and Takeda; and consulting fees from AbbVie, Abivax, Alimentiv, Alma Bio Therapeutics, Amgen, Applied Molecular Transport, Arena Pharmaceuticals, Biogen, Bristol Myers Squibb, Celltrion, CONNECT Biopharma, Cytoki Pharma, Eli Lilly, Enthera, Ferring Pharmaceuticals, Fresenius Kabi, Galapagos, Genentech, Gilead, Gossamer Bio, GSK, H.A.C. Pharma, IAG Image Analysis, Index Pharmaceuticals, Inotrem, Janssen Pharmaceuticals, medac Pharma, Mopac, Morphic, MSD, Norgine, Novartis, OM Pharma, ONO Pharmaceutical, OSE