bluuebunny/crossref_metadata_2025 huggingface dataset:
Temporal regulation of renewable supply for electrolytic hydrogen
Authors: Zeyen Elisabeth , Riepin Iegor , Brown Tom
Published: January 2024

Electrolytic hydrogen produced using renewable electricity can help lower carbon dioxide emissions in sectors where feedstocks, reducing agents, dense fuels or high temperatures are required. This study investigates the implications of various standards being proposed to certify that the grid electricity used is renewable. The standards vary in how strictly they match the renewable generation to the electrolyser demand in time and space. Using an energy system model, we compare electricity procurement strategies to meet a constant hydrogen demand for selected European countries in 2025 and 2030. We compare cases where no additional renewable generators are procured with cases where the electrolyser demand is matched to additional supply from local renewable generators on an annual, monthly or hourly basis. We show that local additionality is required to guarantee low emissions. For the annually and monthly matched case, we demonstrate that baseload operation of the electrolysis leads to using fossil-fuelled generation from the grid for some hours, resulting in higher emissions than the case without hydrogen demand. In the hourly matched case, hydrogen production does not increase system-level emissions, but baseload operation results in high costs for providing constant supply if only wind, solar and short-term battery storage are available. Flexible operation or buffering hydrogen with storage, either in steel tanks or underground caverns, reduces the cost penalty of hourly versus annual matching to 7%–8%. Hydrogen production with monthly matching can reduce system emissions if the electrolysers operate flexibly or the renewable generation share is large. The largest emission reduction is achieved with hourly matching when surplus electricity generation can be sold to the grid. We conclude that flexible operation of the electrolysis should be supported to guarantee low emissions and low hydrogen production costs.
Mechanical stretch regulates macropinocytosis in Hydra vulgaris
Authors: Skokan Taylor D. , Hobmayer Bert , McKinley Kara L. , Vale Ronald D.
Published: January 2024

Cells rely on a diverse array of engulfment processes to sense, exploit, and adapt to their environments. Among these, macropinocytosis enables indiscriminate and rapid uptake of large volumes of fluid and membrane, rendering it a highly versatile engulfment strategy. Much of the molecular machinery required for macropinocytosis has been well established, yet how this process is regulated in the context of organs and organisms remains poorly understood. Here, we report the discovery of extensive macropinocytosis in the outer epithelium of the cnidarian Hydra vulgaris. Exploiting Hydra’s relatively simple body plan, we developed approaches to visualize macropinocytosis over extended periods of time, revealing constitutive engulfment across the entire body axis. We show that the direct application of planar stretch leads to calcium influx and the inhibition of macropinocytosis. Finally, we establish a role for stretch-activated channels in inhibiting this process. Together, our approaches provide a platform for the mechanistic dissection of constitutive macropinocytosis in physiological contexts and highlight a potential role for macropinocytosis in responding to cell surface tension.
Corticocortical connections of the rostral forelimb area in rats: a quantitative tract-tracing study
Authors: Urban III Edward T , Hudson Heather M , Li Yanming , Nishibe Mariko , Barbay Scott , Guggenmos David J , Nudo Randolph J
Published: January 2024

The rostral forelimb area (RFA) in the rat is a premotor cortical region based on its dense efferent projections to primary motor cortex. This study describes corticocortical connections of RFA and the relative strength of connections with other cortical areas. The goal was to provide a better understanding of the cortical network in which RFA participates, and thus, determine its function in sensorimotor behavior. The RFA of adult male Long-Evans rats (n = 6) was identified using intracortical microstimulation techniques and injected with the tract-tracer, biotinylated dextran amine (BDA). In post-mortem tissue, locations of BDA-labeled terminal boutons and neuronal somata were plotted and superimposed on cortical field boundaries. Quantitative estimates of terminal boutons in each region of interest were based on unbiased stereological methods. The results demonstrate that RFA has dense connections with primary motor cortex and frontal cortex medial and lateral to RFA. Moderate connections were found with insular cortex, primary somatosensory cortex (S1), the M1/S1 overlap zone, and lateral somatosensory areas. Cortical connections of RFA in rat are strikingly similar to cortical connections of the ventral premotor cortex in non-human primates, suggesting that these areas share similar functions and allow greater translation of rodent premotor cortex studies to primates.
OP02 IL23R-CAR-Tregs: creating a therapeutic breakthrough for Crohn’s Disease
Authors: Cui Y , Boulakirba S , David M , Bouchareychas L , Rouquier S , Sajuthi S , Ayrault M , Navarin C , Lafon A , Lara G , Menardi A , Saviane G , Dumont C , Abel T , Fenard D , Lu H , de la Rosa M , Gertner-Dardenne J
Published: January 2024

Background

Regulatory T cells (Tregs) are key immunomodulators to maintain immune homeostasis and self-tolerance, including in the intestine. Disrupted immune homeostasis is associated with Crohn’s Disease (CD) pathogenesis. As a living drug, Treg therapy is a promising treatment for restoring immune balance and achieving durable tissue tolerance in the diseased intestine. By targeting a disease-relevant protein, Interleukin-23 receptor (IL23R), we engineered Tregs expressing IL23R-chimeric antigen receptor (CAR) to create IL23R-CAR-Tregs for treating CD.

Methods

IL23R protein expression in CD intestine was verified by immunohistochemical analysis. Tonic signaling and CAR-specific activation were quantified during the IL23R-CAR screening. The IL23R-CAR-Treg phenotype was assessed by monitoring the expression of FoxP3 and Helios transcription factors, and the demethylation status of FoxP3 Treg-specific demethylated region (TSDR). The suppressive function of IL23R-CAR-Tregs was evaluated using in vitro assays and a colitis mouse model. The functionality of IL23R-CAR-Tregs were further assessed using intestinal biopsy-derived cells from CD patients in an in vitro activation assay. Transcriptomic and proteomic analyses were performed to associate molecular profiles with IL23R-CAR-Treg activation.

Results

Our highly potent IL23R-CAR lead candidate displays negligible tonic signalling and a strong signal-to-noise ratio. The resulting IL23R-CAR-Tregs maintain their regulatory phenotype with high FoxP3 and Helios expression and TSDR demethylation during in vitro expansion, CAR-specific stimulation, and in the presence of proinflammatory cytokines. In accordance with their regulatory phenotype, IL23R-CAR-Tregs exhibit CAR-dependent suppressive activity, promote a tolerogenic environment, and protect mice from colitis. The expression of IL23R was also confirmed in the intestinal mucosa from CD patients. Importantly, IL23R-CAR-Tregs elicit activation following exposure to intestinal biopsy-derived cells from CD, demonstrating the engagement and potency of IL23R-CAR towards their target in CD. We also identified molecular profiles that correlated with IL23R-CAR activation and, more broadly, showed their potential to monitor IL23R-CAR-Treg activity in CD patients.

Conclusion

Our results demonstrate that an IL23R-CAR-Treg product could be a promising therapy for CD patients, with the potential long-term benefits of immune tolerance.
OP18 Efficacy and safety of darvadstrocel treatment in patients with complex perianal fistulas and Crohn’s Disease: results from the global ADMIRE-CD II phase 3 study
Authors: Serclova Z , Garcia-Olmo D , Chen S T , Wexner S , Panés J , Wu C , Fleshner P , Zhang B , Colombel J F , Song M , McKay C , Nazarey P , Wright E , Raffals L
Published: January 2024

Background

Complex perianal fistulas are a serious complication in patients with Crohn’s disease (CD). Darvadstrocel (DVS), a suspension of expanded adult allogeneic adipose-derived mesenchymal stem cells, is approved in Europe and Japan for treatment of complex Crohn’s perianal fistulas (CPF). The global ADMIRE-CD II phase 3 randomized double-blind placebo-controlled study evaluated the efficacy and safety of DVS for treatment of complex CPF.

Methods

Patients aged 18–75 years with clinically controlled, inactive or mildly active CD and complex CPF (≤2 internal openings [IO] and ≤3 external openings [EOs]) who had an inadequate, or a loss of response to immunosuppressive agents or biologics were included. Patients were randomized 1:1 to receive a local injection of DVS as a single dose (120 x 106 cells/24 mL) or placebo. In all patients, fistula preconditioning included vigorous curettage and seton placement 2–3 weeks before treatment, and seton removal, a further curettage and closure of IOs immediately before treatment. The primary endpoint was combined remission (closure of all treated EOs that were draining at baseline, despite gentle finger compression, and absence of collections >2 cm confirmed by MRI) at 24 weeks. Secondary endpoints included combined remission at 52 weeks, clinical remission (closure of all treated EOs without MRI confirmation) at 24 and 52 weeks, and time to clinical remission at 24 weeks. Safety was monitored up to 52 weeks.

Results

From 19 Oct 2017 to 26 Jul 2023, 568 patients received DVS (n = 283) or placebo (n = 285) with fistula preconditioning; 56.3% of patients enrolled in Europe and Israel, and 43.7% in North America. Mean (SD) age, sex and race were similar in the DVS and placebo arms (38.4 [11.9] vs 37.7 [10.8] years; 42.8% vs 45.6% female; 85.9% vs 89.1% White). Combined remission rates at 24 weeks did not statistically differ between treatments (48.8% DVS vs 46.3% placebo) and there were no differences in secondary endpoints (Table 1; Figure 1). Based on health authority guidelines, post hoc analyses of patients randomized before COVID-19 (11 March 2020; n = 141 DVS, n = 143 placebo) were performed: combined remission rates at 24 weeks were 46.8% (DVS) and 38.5% (placebo). The safety profile for DVS was consistent with prior studies with no new safety signals (Table 1).

Conclusion

The efficacy outcomes assessed did not statistically differ between DVS and placebo, and the placebo response rate (with fistula preconditioning) was higher than expected. Post hoc analyses revealed lower placebo response rates in patients randomized before COVID-19 (similar to the pivotal ADMIRE CD I study) than the overall placebo arm. DVS was well tolerated.
OP33 Mechanistic insights on the role of ultra processed foods as a trigger/fuel for IBD
Authors: Guedelha Sabino J
Published: January 2024

Background

Ultra processed foods (UPFs) are a heterogeneous category of products that contain food additives and/or have undergone several industrial processes. The higher intake of UPFs seen in industrialized countries has been associated with increased incidence of inflammatory bowel diseases (IBD), mainly Crohn’s disease (CD). However, the specific mechanisms through which the UPFs can induce or maintain intestinal inflammation are still unknown. Therefore, most dietary interventions for IBD essentially exclude all UPFs from the menu, increasing the difficulty of long-term compliance of these diets. The main focus of my research is the study of which components may be detrimental in UPFs, in the hope that we can limit the number of UPFs that we need to exclude from our diets.

Methods

To understand the mechanism of action of UPFs, a translational approach is needed with complementary pre-clinical and clinical studies. We used organoid cultures from patients with CD to test the impact of food additives on the intestinal epithelium (e.g. permeability, pro-inflammatory effect). These organoid cultures are more physiologically relevant than immortalized cell lines, which lack several important aspects (e.g. genetic background, complex assembly of different cells). The results of the first tests with this platform have been submitted to ECCO (EC24-1418). Additionally, we established several protocols and collaborations to run well designed dietary intervention studies to explore the mechanisms of action of these food additives in humans. In the FOAM study, we explored the effects of 5 emulsifiers in an double blind randomized placebo controlled trial. The results of the FOAM study have also been submitted to ECCO (EC24-0858). We also explored the acceptance, feasibility, and compliance of an emulsifier free diet in healthy volunteers (EC24-1181).

Results

Our results support the need to better understand the role of food additives and UPFs in IBD, and the need for better evidence supporting the detrimental role of food additives.

Conclusion

Compliance is an important issue to consider in all dietary interventions, more than in pharmacological interventions. Strategies focusing on improving the compliance of dietary interventions can have a big impact in the applicability of these interventions in therapeutic and/or prophylactic settings. However, the current knowledge on the possible detrimental role of UPFs does not allow to pinpoint which UPFs are detrimental and which might have no effect on intestinal inflammation. Hence, there is an unmet need for pre-clinical and clinical studies focusing on the mechanisms through which these UPFs might play a role in intestinal inflammation.
DOP17 Tofacitinib versus Vedolizumab Among Bio-naïve Patients With Ulcerative Colitis: A Real-World Propensity-Weighted Comparison
Authors: Gros B , Constantine-Cooke N , Kennedy J , Elford A T , O’Hare C , Noble C , Jones G R , Arnott I D , Plevris N , Lees C W
Published: January 2024

Background

Over the last decade, treatment options for moderate-to-severe ulcerative colitis (UC) have expanded. However, comparative studies between these agents are limited, especially among biologic naïve patients. There are sparse data comparing biologics with small molecules for IBD. We aimed to compare the efficacy, safety and persistence of tofacitinib and vedolizumab as the first advanced treatment for patients with UC.

Methods

Patients who received tofacitinib or vedolizumab as first advanced therapy for UC in NHS Lothian were included. We capped the upper age limit at 65 years to take into account regulatory guidelines for the use of JAKi as first line therapies across IMIDs. To allow treatment effect to be assessed outside of a randomized trial, we used inverse probability of treatment weighting (IPTW). This approach takes the probability of treatment assignment into account without potentially drastically reducing the analysable cohort size. The probability of treatment assignment was calculated via logistic regression using age, gender, IBD duration, Montreal extent, CRP, concomitant corticosteroids and partial Mayo score at drug commencement. Missing data for these variables were imputed using multivariate imputation by chained equations. Confounder-adjusted survival curves were created using Kaplan-Meier estimates weighted via IPTW. The Pepe and Fleming test was used to compare survival curves from drug commencement to day 1000 of treatment.

Results

We included 158 patients of whom 81 (51.2%) received vedolizumab and 77 (48.7%) tofacitinib. Median follow-up for patients on vedolizumab was 3.1 (1.6-4.8) years and for tofacitinib 1.5 (0.34-2.3) years. Baseline demographics were comparable except for disease extent (Table 1). At drug commencement there were no differences in steroid prescription (60.5% vedolizumab versus 57.1% tofacitinib), partial Mayo score, CRP or faecal calprotectin. At week 12, steroid free clinical remission was more frequent in the vedolizumab group (69% vs 51.4%, p=0.030) Figure 1B.

Vedolizumab persistence was superior to tofacitinib (p=0.005) Figure 1A. Primary non-response and secondary loss of response were 9.9% and 17.3% for vedolizumab and 23.4% and 13% for tofacitinib respectively. There were no differences in the frequency of adverse events (11 [13.6%] vedolizumab vs 19 [24.7%] tofacitinib, p=0.629). However, adverse events resulting in temporary discontinuation of the drug occurred in 1 (1.2%) vedolizumab versus 11 (14.3%) tofacitinib, p=0.013.

Conclusion

We found that the persistence and tolerability of vedolizumab was superior to tofacitinib in bionaive UC, although the rates of clinical and biomarker remission were comparable. These data may help inform positioning of advanced therapy in UC.
DOP38 A rare KRT17+ population emerges from the epithelium in the inflamed intestinal mucosa of patients with Ulcerative Colitis
Authors: Dotti I , Melón-Ardanaz E , Sanzo Á , Gudiño V , Corraliza A M , Teubel I , Esteller M , Ricart E , Salas A
Published: January 2024

Background

KRT17 is an epithelial "alarmin" that contributes to the early host innate immune response to insults. We previously found that KRT17 transcription, not detectable in the healthy colon, is up-regulated in the intestinal mucosa of patients with active ulcerative colitis (UC). Here, we aimed to characterize the epithelial cells expressing KRT17 in UC mucosa and explore the contribution of the intestinal environment in promoting the emergence of this cell population.

Methods

We conducted single-cell RNAseq (scRNA-seq) on the epithelial compartment of intestinal biopsies from a cohort of non-inflammatory bowel disease (IBD) controls and patients with active UC. Analysis was performed using our own-curated pipeline in Seurat1. Spatial protein localization was analysed by immunofluorescence on fixed intestinal samples. To measure the modulation of KRT17 and associated markers, ex vivo air-liquid interface (ALI) epithelial cultures derived from human non-IBD organoids were used. The signatures of UC-derived organoids2 were mapped onto the scRNA-seq dataset.

Results

ScRNA-seq analysis identified a rare KRT17+ epithelial cell population emerging in active UC characterized by the expression of KRT17 (Fig. 1A) and other alarmins (KRT6A, KRT16, S100A8), metalloproteinases (MMP1, MMP10), and inflammatory markers (SAA1, DUOX2…). KRT17 protein, absent in non-IBD colon, was expressed in the epithelium of the inflamed UC mucosa exhibiting a patchy pattern, and marked epithelial clusters with both columnar and squamous-like morphology (Fig. 1B). The ALI culture closely recapitulated the differentiated intestinal epithelium in homeostasis by showing a columnar aspect, high expression of differentiation markers, and low or undetectable expression of stem/proliferation or KRT17+-cell markers. Exposure of the ALI culture to hypoxic stress (by re-submersion in expansion medium) induced the expression of KRT17+ cell-specific and inflammation markers (Fig. 1C), and the acquisition of a squamous morphology (Fig. 1D). IBD-related IFN-ɣ and IL-22 cytokines could not fully replicate this phenotype (Fig. 1E). Lastly, we observed that genes up-regulated in UC, compared to non-IBD organoids, were predominantly expressed within the KRT17+ population (Fig. 1F).

Conclusion

Our results suggest that a novel KRT17+ epithelial population may arise from the intestinal cells of inflamed UC mucosa under the influence of specific environmental factors. This population could contribute to the persistence of the disease over time.

References

1. Garrido A. et al, Nat Commun, 2023

2. Dotti I. et al, Gut, 2017
DOP51 Risk of kidney failure in patients with Inflammatory Bowel Disease undergoing colectomy: a nationwide cohort study
Authors: Yang Y , Ludvigsson J F , Forss A , Faucon A L , Faye A S , Olen O , Sjölander A , Carrero J J
Published: January 2024

Background

Inflammatory bowel disease (IBD) is frequently accompanied by kidney complications. Potential triggers or subpopulations at high-risk of kidney problems are not well elucidated. We hypothesized that surgical interventions, specifically colectomy, might in part explain this risk.

Methods

A nationwide Swedish cohort study comprising 82,052 individuals with biopsy-proven IBD diagnosed during 1965-2017, with follow-up until 2019. We investigated the association between incident colectomy (time-varying exposure) and future risk of kidney failure (diagnosis of end-stage kidney disease or death due to chronic kidney disease) using Cox proportional hazard models. We also examined the impact of partial vs. total colectomy and the presence of a stoma. Covariates included age, sex, education level, and selected comorbidities.

Results

Over a median follow-up of 14 years, 15,922 individuals underwent colectomy, and 952 kidney failure events occurred. Colectomy was associated with an increased relative risk of kidney failure (adjusted hazard ratio [aHR] 1.60; 95% CI 1.39-1.84). Compared to periods without colectomy, undergoing total colectomy (aHR 1.82; 95% CI 1.54-2.15) and colectomy with the presence of a stoma (aHR 2.40; 95% CI 1.75-3.28) showed higher risks versus partial colectomy or colectomy without a stoma, respectively. Subgroup analyses revealed higher kidney failure risk in patients with ulcerative colitis and those aged ≥40 years at IBD diagnosis. In absolute risks, at 30-year after colectomy, 8.9% of patients with IBD had kidney failure, with an absolute risk difference of 4.5% (95% CI 1.9-7.0%).

Conclusion

In people with IBD, rates of kidney failure are higher among those undergoing colectomy, particularly following total colectomy or colectomy with a stoma. This study identifies a high-risk population that may benefit from established protocols for kidney function monitoring/surveillance and referral to nephrologist care.
DOP81 Rosnilimab, a novel PD-1 agonist monoclonal antibody, reduces T cell proliferation, inflammatory cytokine secretion, and PD-1high expressing CD4 and CD8 T cells: Results from a Phase 1 healthy volunteer clinical trial
Authors: Luu K , Dahl M , Hare E , Sibley C , Lizzul P , Randazzo B
Published: January 2024

Background

Programmed cell death protein 1 (PD-1) is expressed on activated T cells and is a key co-inhibitory node in immune regulation. By targeting regulatory mechanisms to modulate immune cells driving disease, there is an opportunity to restore immune balance. Rosnilimab is a PD-1 agonist antibody designed to reduce T cell proliferation and inflammatory cytokine secretion and deplete PD-1high T follicular helper, T peripheral helper, and T effector cells. It is being studied for ulcerative colitis (UC), where PD-1+ T cells are prevalent in inflamed lamina propria (>40%) and the periphery. The primary objective of this healthy volunteer Phase 1 study was to assess the safety and tolerability of single ascending doses (SAD) and multiple ascending doses (MAD) of rosnilimab. Findings from pharmacokinetic (PK) and pharmacodynamic (PD) assessments are summarized.

Methods

This single-centre study included 14 cohorts in SAD and 3 cohorts in MAD. Each cohort had 8 participants (6 active, 2 placebo [PBO]). Cohorts were enrolled sequentially in each phase. Intravenous (IV) and subcutaneous (SC) administration were assessed in SAD; SC route was assessed in MAD.

Results

A total of 144 participants were enrolled; 90 to active SAD cohorts, 18 to active MAD cohorts, and 30 and 6 to SAD and MAD PBO cohorts, respectively. Rosnilimab was well tolerated with no dose-limiting toxicities or deaths. Two serious adverse events were reported in SAD (deemed unrelated to treatment) and 0 in MAD. PD-1 expressing cells were reduced by ~50% in both CD4+ and CD8+ subsets, in a dose-dependent manner and in correlation with full receptor occupancy (RO) through Day 30 in SAD. PD activity was rapid with sustained reduction in PD-1+ T cells, and ex-vivo stimulation resulted in reduced T cell functional activity. This reduction was maximized on PD-1high expressing T cells: ~90% reduction vs baseline. There was no significant impact on the overall total T cell or regulatory T (Treg) cell numbers, resulting in restoration of T cell composition to a less activated state and a positive shift in the Treg:Teff ratio. An antigen-specific functional T cell assay measuring ex vivo interferon-gamma release in response to antigen challenge was inhibited up to ~90% vs baseline and the response lasted for more than 30 days following a single dose. Rosnilimab had a favourable PK profile consistent with full RO, a 2-week half-life, and dose-proportional exposure in IV and SC dosing.

Conclusion

Rosnilimab demonstrated favourable safety, PK, and PD activity. The role of PD-1 in UC pathophysiology coupled with these results and translational data, demonstrate proof of mechanism and support progression into a Phase 2 study of rosnilimab in UC. (NCT06127043)
P005 Exosome miR-132-3p derived from Adipose Stem Cells in Creeping fat Regulates RASA1/ERK1/2-dependant Lymphatic Functions and Improve Colitis and Mesenteritis
Authors: Wang X , Shu W , Wang Y
Published: January 2024

Background

Creeping fat (CrF) is an important pathological characteristics of Crohn’s disease (CD) and characterized by lymphangiogenesis and abnormal lymphatic functions. Adipose tissue derived stem cells (ADSCs) can regulate surrounding cells function by delivering exosomes (exo). However, whether CrF-ADSC-exo could affect lymphatic endothelium to regulate lymphatic functions, and further influence procession of mesenteritis and colitis remains elusive.

Methods

Paired CrF and uninvolved mesenteric adipose tissue (uMAT) samples were collected from CD patients during operation, and exosomes were separated by ultra-centrifugation. Human lymphatic endothelial cells (HLECs) were cultured in vitro and evaluated the abilities in proliferation, migration and tube formation. IL-10 KO mice was used for assessing the treatment of CrF-ADSC-exo in experimental colitis and mesenteritis.

Results

ADSC-derived exosomes from paired CrF and uMAT samples were confirmed to transfer to LEC by immunofluorescence microscope. In vitro, CrF-ADSC-exo effectively stimulated HLECs in proliferation, migration and tube formation. In vivo, CrF-ADSC-exo injection increased lymphangiogenesis in MAT and improved lymphatic drainage, and further alleviated colitis and mesenteritis in IL-10 KO mice. We next adopted high-throughput sequencing of paired CrF-ADSC-exo and uMAT-ADSC-exo, and validated significantly high expression of miR-132-3p in CrF-ADSC-exo. In vitro, upregulation or downregulation of miR-132-3p in HLECs stimulated or inhibited the lymphatic abilities in proliferation, migration and tube formation, respectively. Mechanically, we demonstrated that CrF-ADSC-exo regulated lymphatic function through miR-132-3p-RASA1-ERK1/2 axis by western blot. CrF-ADSC-exo miR-132-3p improved lymphatic drainage function in MAT and reduced inflammatory colon and mesentery in IL-10 KO mice. The higher level of miR-132-3p in CrF from CD patients was tightly correlated with inflammatory mediators expressions when compared with uMAT and healthy MAT.

Conclusion

Our study discovers that CrF-ADSC-exo improves lymphatic functions and alleviates colitis and mesenteritis through miR-132-3p/RASA1/ERK1/2 axis, which discloses new role of CrF involved in CD procession and provides novel insight for CD treatment.
P009 IL-17 mediates visceral hypersensitivity by sensitising colonic nociceptors
Authors: Paine L , Higham J , Barker K , Welsh F , St John Smith E , Bulmer D
Published: January 2024

Background

Comparison of cytokine expression in colitic tissue with respective cytokine receptor expression in pain sensing neurons highlights a likely contribution of IL-17 to the production of abdominal pain during flare.

To explore this pro-nociceptive potential of IL-17 in colitis we examined the ability of IL-17 to stimulate sensory neurons and the colonic afferent response to noxious distension.

Methods

IL-17 mediated sensory neuron activation: changes in the intracellular [Ca2+]i within thoracolumbar (T12-L5 spinal segments) dorsal root ganglia sensory neurons were examined in cells cultured overnight and loaded with Fluo-4-AM (30min) prior to imaging. Responses were measured to IL-17 (10-300 ng/mL) alone and following pre-treatment with either TRPV1 or TRPA1 inhibitors. Afterwards cells were exposed to capsaicin (1 µM) followed by 50 mM KCl, to confirm the presence of nociceptors and neurons respectively by positive response. In a separate set of experiments the effect of IL-17 was also examine on a neuron pure cell culture in which non-neuronal cells were removes by magnet assisted cell sorting.

IL-17 mediated colonic nociceptor activation: electrophysiological recordings were made of lumbar splanchnic nerve activity and the response to noxious luminal distension of the isolated colorectum (ex-vivo), perfused with carbogenated Krebs solution, examined following pre-treatment with IL-17 (50 ng/mL) alone or in the presence of respective inhibitors.

All experiments were performed using tissue from male C57BL/6J mice euthanaised by rising concentration of CO2followed by exsanguination in accordance with Schedule 1 of the UK Animals Scientific Procedures act (1986).

Results

Consistent with the ability of IL-17 to stimulate sensory neurons including those classified as nociceptors by their co-sensitivity to capsaicin, pre-treatment with IL-17 also produced a significant sensitisation of the colonic afferent response to luminal distension at noxious distending pressures. The increase in [Ca2+]i produced by IL-17 in sensory neurons was blocked by pre-treament with a TRPV1 antagonist but not a TRPA1 antagonist. Furthermore, the proportion of neurons stimulated by IL-17 was comparable between standard DRG cell cultures and neuron pure cell cultures demonstrating a direct effect of IL-17 on sensory neurons. Finally, IL-17 mediated mechanosensitisation of colonic nociceptors was unaffected by TRPV1 inhibition.

Conclusion

Our findings demonstrate that IL-17 stimulates capsaicin-sensitive nociceptors and mediates visceral hypersensitivity in colonic nociceptors consistent with a causative role for IL-17 is the generation of abdominal pain in people with colitis.
P025 Obefazimod and its active metabolites ABX-464-N-Glu act by stabilizing protein-protein interaction among key RNA biogenesis partners, CBC and ARS2
Authors: Bron P , Martin K , Sloan S , Scherrer D , Tazi J
Published: January 2024

Background

Obefazimod is an oral, once-daily, small molecule in phase 3 clinical trials for the treatment of patients with ulcerative colitis (UC). Obefazimod selectively enhances the expression of a single micro-RNA (miRNA), miR-124. miRNAs function by binding to specific mRNA targets and reducing their translation into their respective proteins. Enhanced expression of miR-124 results in a decrease in cytokines and immune cells, helping to reduce inflammation and potentially controlling the progression of chronic conditions like UC. Obefazimod and its main active metabolite, ABX464-N-Glu, bind the nuclear Cap Binding Complex (CBC) that directs cellular RNA biogenesis by recognizing co-transcriptionally the cap structure of RNA polymerase II transcripts. The CBC complex is made of Cbp20 and Cbp80, and interacts with a key partner involved in microRNA biogenesis, ARS2. Here, we explore the structural basis for targeting the CBC complex.

Methods

Microscale thermophoresis analyses were performed to determine the binding affinities between the two components of CBC, Cbp20 and Cbp80, in presence or absence of obefazimod or its metabolite ABX464-N-Glu, and their interaction with ARS2 also in presence or absence of the two molecules. Cryo-electron microscopy (cryo-EM) datasets of CBC-apo, CBC-obefazimod, CBC-ABX464-N-Glu, CBC-ARS2-Apo, CBC-ARS2-obefazimod, CBC-ARS2-ABX464-N-Glu were processed and after 2D classification and 3D refinement the different structures were solved. To understand and explore the conformational dynamics of the complex, 3D multibody refinement was performed on CBC-Apo, CBC-obefazimod and CBC:ABX464-N-Glu.

Results

The cryo-EM structures showed that obefazimod or ABX464-N-Glu bind the same location at the interface of CBC complex to act as orthosteric stabilizers of Cbp20 and CBp80. The conformational change induced by the binding of obefazimod or ABX464-N-Glu to the CBC complex strengthens the interaction between the two subunits of CBC, resulting in a more rigid structure of CBC complex. Thermophoresis revealed that the binding of obefazimod or ABX464-N-Glu improves the interaction of CBC with ARS2, a key partner of CBC to establish the fate of RNA in the cell. The cryo-EM structures of CBC-ARS2, obefazimod-CBC-ARS2 or ABX464-N-Glu-CBC-ARS2 show that ARS2 passes through the site where Obefazimod and ABX464-NGlu bind. We propose that the orthosteric effect of obefazimod or ABX464-N-Glu on CBC is maintained even when these molecules leave their binding site, allowing for better affinity of ARS2 for CBC.

Conclusion

Our results identify a unique interfacial stabilizing mode of action for anti-inflammatory drugs targeting the CBC complex enhancing its interaction with ARS2 to generate specific microRNA.
P053 Pro-resolution Pathway in Experimental Colitis – A New Possible Therapeutic Approach for Inflammatory Bowel Diseases
Authors: Chaim F H M , Pascoal L B , Palma B B , Rodrigues B L , Castro M M , Fagundes J J , Milanski M , Lopes L R , Velloso L A , Leal R F
Published: January 2024

Background

Inflammatory bowel diseases (IBD) are idiopathic disorders characterized by chronic inflammation of the gastrointestinal tract. New therapeutic strategies involving the inflammatory process are being proposed because the usual therapies indicated for IBD may cause adverse reactions and clinical failure. Recent studies have highlighted the role of specialized pro-resolution lipid mediators (SPMs) in the active resolution of chronic inflammation. Therefore, this study aimed to investigate the SPM Resolvin D2 (RvD2) and its precursor, omega-3 polyunsaturated fatty acid, in experimental colitis induced by dextran sulfate sodium (DSS).

Methods

An experimental model with male mice of the C57BL/6J lineage was carried out. Mice were separated into a control group, and a DSS-induced colitis group (3% dilution in drinking water); both groups were subdivided and received either a standard diet or an experimental diet enriched with 20% of oil rich in omega-3. Another group of mice, after acquiring experimental DSS-induced colitis, was subdivided and treated with RvD2 or anti-TNFα. Throughout the experimental protocols, the body weight, food intake, Disease Activity Index (DAI), metabolic phenotype, and inflammatory profile of these animals were assessed by real-time PCR, ELISA, histology, Western blotting, and lipid analysis. The study was approved by the University of Campinas research ethics committee.

Results

The animals with experimental colitis induced by DSS showed an increase in TNFα and IL22 transcriptional expression besides a reduction in the enzymes involved in the endogenous biosynthesis of RvD2, such as PLA2, 15-LOX, 5-LOX, and its receptor GPR18. Dietary supplementation with omega-3-rich oil increased RvD2 and its precursor besides reducing the DAI, weight loss, colonic shortening, and the histologic colonic inflammation score. Treatment with RvD2 effectively attenuated experimental colitis, reducing TNFα transcriptional levels and p-JNK protein expression. Besides, RvD2 increased the immediate precursor of RvD2 (7,8-epoxy-17-HDHA). All of which indicates that this pathway is responsible for the observed effects.

Conclusion

Our study provided a better understanding of the molecular mechanisms involved in the exacerbated inflammatory response present in colitis. The beneficial effect of RvD2 and its precursor omega-3 may bring about a new therapeutic approach for IBD.
P073 Enhancing the MFSD2A protein for the treatment of colitis-associated cancer: novel insights for pathogenesis and treatment
Authors: Cagliani S , Massimino L , Facoetti A , Spanò S , Errico C , Parigi T L , Colasante G , Danese S , Ungaro F
Published: January 2024

Background

The intrinsic connection between inflammation and tumor promotion is well characterized and is a key pathogenic event in patients with colorectal cancer (CRC). A small fraction of patients with CRC suffers from genetic predisposition. However chronic inflammation, such as ulcerative colitis (UC), increases the risk of intestinal carcinogenesis, thus strengthening the connection between these conditions. A few years ago, our laboratory described that the intestinal endothelium isolated from actively inflamed UC patients displayed defective production of inflammation-resolving DHA-derived metabolites, by comparison with healthy controls and tissues in remission. We also reported that the Major Facilitator Superfamily Domain containing 2A (MFSD2A) participated in the production of the inflammation-resolving DHA-derived metabolites by the intestinal endothelium of patients with UC in remission, ameliorating colonic inflammation. Therefore, we hypothesized that the endothelial MFSD2A also has a role in preventing and/or counteracting cancer-associated inflammation.

Methods

Human Intestinal Microvascular Endothelial Cells (HIMEC) isolated from CRC and healthy samples were transduced with either MFSD2A protein-encoding lentiviral vectors (MFSD2A) or GFP as control, or MFSD2A targeting shRNA or its scramble, and were analyzed by lipidomics. Furthermore, Caco-2 cells co-cultured with HIMEC-MFSD2A were analyzed by FACS, evaluating their proliferation. Moreover, an orthotopic model of CRC was made intrarectally injecting CD1 nude mice with a cell mixture of Caco-2 and HUVEC overexpressing MFSD2A or GFP as control. Mice were gavaged with DHA-ethyl-ester or PBS. FACS analysis was performed on tumor and colonic samples.

Results

Lipidomic analysis revealed that the loss of MFSD2A in CRC is detrimental because there is a reduced production of pro-resolving lipids confirming that, as for UC, MFSD2A is important for the maintenance of an adequate balance between pro-resolving and pro-inflammatory phenotype. Moreover, enhancing MFSD2A expression at endothelial level limits Caco-2 cell proliferation, supporting its beneficial role. In addition, DHA administration in mice ameliorated clinical symptoms of inflammation upon intestinal endothelial MFSD2A overexpression, supporting the beneficial role of MFSD2A in vivo.

Conclusion

Tumor-associated inflammation remains a crucial hallmark of CRC. Our study seeks to identify the role of MFSD2A in the resolution phase of inflammation, pointing out alternative therapies for CRC treatment. These data suggested that MFSD2A might contrast the tumor-associated inflammation and ensure the correct balance between pro-inflammatory and pro-resolving milieu.
P077 The Immunometabolic Bimodal Mechanism of NLRX1 Agonist NX-13 in a Pig Model of Ulcerative Colitis
Authors: Verstockt B , Mosig R , Dubinsky M , Cataldi F , Danese S
Published: January 2024

Background

In mouse models, NX-13 is an orally active, gut-restricted NLRX1 agonist that has been shown to reduce severity of disease in multiple UC models through a novel immunometabolic, bimodal mechanism. Most preclinical UC studies rely upon rodent models, the human gastrointestinal physiology, microbiome, and immune system bear a greater similarity to that of pigs. Therefore, we sought to expand and validate our preclinical NX-13 results in mice using the dextran sulfate sodium (DSS) pig model of colitis. Here, we describe the disease severity, cellular, and biomarker results, and confirm the immunometabolic MOA observed in other species.

Methods

Pigs were randomized by weight (n=6 per group) and were administered tablets of excipient alone (placebo), NX-13 10 mg, NX-13 50 mg, or NX-13 100mg once daily. Pigs were challenged with 1% DSS in drinking water for 6 days and monitored daily for weight gain (loss). Composite disease activity score (0-12) was taken based on the sum of three subscores (0-4) for rectal bleeding, stool consistency and overall health/activity. Necropsy was conducted on day 7. Feces were collected for fecal calprotectin (FCP) quantification. Colonic tissue was macroscopically scored and collected for analysis by flow cytometry, histopathology and gene expression.

Results

Oral NX-13 treatment protected against weight loss and reduced colitis development with differences as early as day 2, becoming significant between days 4 and 6 as disease worsens in placebo treated pigs (Fig1A). NX-13 yielded a dose-dependent improvement in macroscopic lesion severity and microscopic immune cell infiltration (Fig1B). Specifically, Th1 cells (Fig1C) and TNF producing myeloid cells in the colonic lamina propria were reduced by NX-13, as well as FCP levels in stool (Fig1D) and mRNA expression in the colon (Fig1E). Oral NX-13 treatment reduced inflammatory markers including TNF, monocyte and neutrophil chemo attractants, IL-6, and IL-23 (Fig3F). Additionally, NX-13 treatment affected markers specific to the bimodal immunometabolic MOA of NLRX1 activation, namely upregulation of NLRX1 and mitochondrial metabolism gene COX3 (Fig1G), and reduction of NFkB and NLRP3 (Fig1H).

Conclusion

NX-13 demonstrated fast and clinically meaningful improvement in disease activity, and significantly impacted on key inflammatory markers in a DSS colitis model in pigs. These results align with previous murine results and observations in the phase 1b clinical program. Therefore, they can be used to generate additional mechanistic hypotheses for translational studies in the ongoing NEXUS phase 2 trial in moderate to severe UC.
P095 Accelerated degradation of gut barrier-protecting 3-aminobenzoic acid exacerbate colitis in Inflammatory Bowel Disease
Authors: Tanaka M , Toyonaga T , Nakagawa F , Sumiyoshi N , Shibuya N , Iwamoto T , Minemura A , Ariyoshi T , Matsumoto A , Oka K , Shimoda M , Saruta M
Published: January 2024

Background

Disruption of intestinal epithelial cell (IEC) barrier function is crucial for the development of inflammatory bowel disease (IBD). Decreased diversity and compositional changes in intestinal bacteria, namely dysbiosis, impair IEC barrier function in IBD, but the underlying mechanism is not fully understood. This study aims to identify the impact of dysbiosis-driven changes in intestinal bacterial metabolites on IEC permeability in patients with IBD.

Methods

Caco2 cells were cultured on the trans-well inserts and stimulated with a library of intestinal bacterial metabolites. Proportional changes of trans-epithelial electrical resistance (TEER) were calculated at 48 hours after stimulation (ΔTEER%) and the permeation of 70-kDa FITC-dextran was evaluated using a Qubit fluorometer. Fecal samples were collected from 10 IBD patients with active colonic inflammation (5 ulcerative colitis and 5 Crohn’s disease) and 5 healthy controls (non-IBD, NIBD). Fecal bacteria-derived metabolites were quantified by LC-MS/MS and compared between IBD patients and NIBD controls. 16SrRNA-based metagenomic analysis was performed on the same fecal samples using QIIME2 followed by a PICRUSt2-based functional analysis of microbial communities. Anti-inflammatory effect of 3-aminobenzoic acid (3-ABA) was examined in C57BL/6 mice using a dextran sodium sulfate (DSS)-induced colitis model.

Results

Among 119 intestinal bacterial metabolites, 4-ABA decreased epithelial permeability the most with the highest ΔTEER% value (control vs. 4-ABA, 1.8 vs. 30.9 %, p<0.05). 3-ABA, one of three structural isomers of ABA, also decreased epithelial permeability in a dose dependent manner with higher ΔTEER% value than 4-ABA and reduced permeation of FITC-dextran (control vs. 3-ABA, 8.3 vs 3.4 nM, p<0.05). Fecal level of 3-ABA was significantly lower in IBD patients than in NIBD controls (7.0 vs. 505.0 μM). Functional prediction of intestinal microbiota demonstrated that IBD patients were significantly more enriched with the bacteria involved in the degradation of ABA than NIBD controls. Rectal administration of 3-ABA significantly improved body-weight loss, shortening of colon length, histologic scores, and the expression of inflammatory cytokines in mice with DSS-induced colitis (control vs. 3-ABA; body-weight loss,15.33 vs. 18.91 g; colon length, 6.11 vs. 7.35 cm; histologic score, 2.14 vs 0.60).

Conclusion

Accelerated degradation of luminal 3-ABA by intestinal bacteria may disrupt intestinal epithelial barrier function and exacerbate colitis in patients with IBD.
P105 Mendelian Randomization screening for JAK inhibitor related potential serious adverse events
Authors: Miguel González J A , Pavlidis P , Marigorta U M
Published: January 2024

Background

JAK inhibitors (JAKi) suppress the JAK/STAT signalling pathway through inhibition of the four Janus kinase proteins JAK1, JAK2, JAK3 and TYK2. JAKi are licensed for the treatment of a wide range of immune mediated inflammatory diseases (IMIDs) including inflammatory bowel disease. Concerns have been raised for the potential of this class of medication to associate with serious adverse events, particularly in those older than 65 years of age or with previous cardiovascular comorbidities. Mendelian randomization (MR) uses genetic variants as instrumental variables (IVs) to examine the causal effects of an exposure (e.g. inhibition of a drug target) on an outcome (e.g. the risk of developing a disease and/or potential adverse effects due to alteration of gene activity). We carried out a comprehensive MR evaluation to explore the therapeutic potential of JAKi for a range of IMIDs, and to study the unintended drug effects that might be caused by this therapeutic strategy.

Methods

The basis of MR relies on the use of genetic variants as IVs that are reliably related to the risk factor and that are not susceptible of reverse causation and confounding. We used genetic variants (SNPs) associated with protein levels and gene expression (pQTLs and eQTLs, respectively) as IVs to study the effects of inhibiting the genes in the JAK/STAT signalling pathway. The underlying assumption is that these IVs serve as proxies of the effects of JAKi. As a source of the effect estimates on the selected outcomes, we used summary statistics from a compiled list of genome-wide association studies.

Results

We used the MR approach to chart the range of possible outcomes associated with JAKi treatment. First, we explored the therapeutic potential of JAKi for IMIDs. Second, we evaluated a range of potential adverse events associated with JAKi, focusing particularly on cardiovascular, infectious and cancer-related malignancies. Our MR results confirm a general tendency towards JAK/STAT signalling pathway involvement in IMIDs risk, implying the suitability of JAKi for IMIDs. Regarding potential adverse outcomes, we carried out stratification-based analyses to evaluate whether potential adverse events are enhanced in individuals at particular high risk. The MR analyses highlighted potential risk of JAKi for non-IMIDs traits, particularly when targeting JAK2. This includes already known associations with cardiovascular disease and specific cancer types.

Conclusion

We performed an outcome-wide exploration based on MR to characterize the therapeutic potential and range of adverse effects expected for JAKi. We recapitulate the known potential of these drugs to treat IMIDs, while uncovering specific severe adverse effects associated with this therapeutic target.