Introduction

The gut microbiome has emerged as a significant contributor to the pathophysiology of many human ailments, including Inflammatory Bowel Disease (1). However, the potential for exploring and manipulating the microbiome for the identification of new treatment avenues is also gaining traction.
The microbiome immunomodulatory paradigm which currently exists suggests that bacteria within the gut are associated with disease through sequencing techniques, while the functional capacity of the microbiome is under-examined; currently microbes are broadly divided into pro-inflammatory or anti-inflammatory.
“Pathobiont” was a term coined to describe microbes which can cause harm when in certain conditions (2). Bacteroides species for example have been implicated in both Crohn’s’ disease (CD) and ulcerative colitis (UC) through sequencing analyses, and have been demonstrated to produce compounds which increase expression of pro-inflammatory cytokines in enterocyte and goblet cell lines (3). Bacteroides spp. are however, some of the most predominant anaerobes within the gastrointestinal tract (4).
Of the many immune pathways which contribute to the pathogenesis of IBD, the IL-23- or IL-6- JAK/STAT signalling pathways are emerging as targets for drug development (5).
We hypothesised that compounds produced by healthy gut bacteria, including such Bacteroides spp., modulate this important immune axis in IBD and produce an attractive treatment opportunity that is yet to be fully harnessed.

Aims & Methods

The suppressive effects of culture supernatants (CS) from 475 anaerobic isolates, collected from 6 healthy control stool samples, were tested on human embryonic kidney STAT3 reporter cell lines. 6 suppressive stool isolates were chosen for further characterisation of secreted product composition, prior to whole genome shotgun sequenced and biochemical comparison with closely related commercially available type strains. CS was also tested on murine immune cells and explants, and PBMCs isolated from the whole blood of UC, CD, and non-IBD controls. Finally, isolates HS183 and HS811 were given by rectal gavage to Winnie mice, which model spontaneous colitis in vivo.

Results

CS from stool isolates demonstrated a range of immunomodulatory capacities in STAT3 reporter cell lines. 6 isolates were selected for further experimentation; of the 6 isolates, CS contained suppressive compounds in both the <3kDa and >3kDa fractions, with varying sensitivity to proteinase K treatment, suggesting multiple immunomodulatory bioactives may be secreted by the same isolate. When sequenced, the 6 isolates were found to be closely related to Bacteroides spp. Biochemically, stool strains and type strains behaved similarly when grouped by closely related species. The selected CS also suppressed proinflammatory gene expression in murine and human cells ex vivo, with interpatient variability. Finally, CS from isolates HS183 and HS811 reduced disease activity and modulated pro-inflammatory markers in following rectal gavage in Winnie mice.

Conclusion

We found that bacteria traditionally considered to be pathobionts, contributing to the pathogenesis of IBD, also have the capacity to secrete anti-inflammatory compounds which can be harnessed as potential novel therapeutics. Our pipeline allows for the rapid identification of bacterially secreted bioactives which modulate this specific signalling pathway. Finally, our in vivo and ex vivo experiments using models of colitis and patient samples, further exemplifies the potential for personalised medicine approaches for new medications in the future.

References

1. Franzosa EA, Sirota-Madi A, Avila-Pacheco J, Fornelos N, Haiser HJ, Reinker S, et al. Gut microbiome structure and metabolic activity in inflammatory bowel disease. Nat Microbiol. 2019;4(2):293-305.
2. Rehaume LM, Matigian N, Mehdi AM, Lachner N, Bowerman KL, Daly J, et al. IL-23 favours outgrowth of spondyloarthritis-associated pathobionts and suppresses host support for homeostatic microbiota. Ann Rheum Dis. 2019.
3. Ó Cuív P, de Wouters T, Giri R, Mondot S, Smith WJ, Blottiere HM, et al. The gut bacterium and pathobiont Bacteroides vulgatus activates NF-kappaB in a human gut epithelial cell line in a strain and growth phase dependent manner. Anaerobe. 2017;47:209-17.
4. Wexler HM. Bacteroides: the good, the bad, and the nitty-gritty. Clinical microbiology reviews. 2007;20(4):593-621.
5. Soendergaard C, Bergenheim FH, Bjerrum JT, Nielsen OH. Targeting JAK-STAT signal transduction in IBD. Pharmacol Ther. 2018;192:100-11.

Introduction

A marked decrease of Faecalibacterium prausnitzii is a well-established hallmark of Crohn’s disease (CD)-associated dysbiosis and predicts disease relapse (1-2). Several strains of F. prausnitzii exhibit anti-inflammatory effects in vitro and in rodents colitis models, positioning this species as a promising therapeutic candidate (3-4). However, it has never been administered to humans. Here, we present the development and first-in-human evaluation of F. prausnitzii strain EXL01 for CD treatment.

Aims & Methods

F. prausnitzii EXL01, isolated from a healthy donor, was selected based on strict safety and preliminary in vitro efficacy criteria. Its anti-inflammatory effect was tested in trinitrobenzenesulfonic acid (TNBS)-induced colitis in rats and dextran sodium sulfate (DSS)-induced colitis in mice. After favorable toxicological studies, a first-in-human, open-label, single-arm study (Part A) was conducted in patients with mild to moderate ileal CD (CDAI 180–350), following clinical remission (CDAI <150) or response (CDAI decrease ≥70) after corticosteroids (CS). EXL01 (10¹⁰ total cell count/day) was administered orally for 24 weeks, with a 6-week CS taper. Colonoscopies were performed at baseline and week 24. The primary endpoint was safety; secondary endpoints included clinical efficacy, blood/fecal biomarkers, transcriptomic analysis, and gut microbiota profiling. Part B, initially planned as a placebo-controlled phase, was not pursued due to low recruitment.

Results

According to classical evaluation endpoints, EXL01 demonstrated dose-dependent anti-inflammatory effects in both TNBS and DSS models. Between March 2023 and October 2024, 8 patients were enrolled across Belgium and Poland. Baseline characteristics were consistent with the target CD population. After corticosteroid induction, the parameters were as follow: CDAI = 78.1 (42-105); SES-CD= 6.9 (2-17); CRP = 5.7mg/l (3-13). EXL01 was well tolerated; no treatment-related serious adverse events or safety signals were observed. Six participants (75%) completed the 24-week treatment; two discontinued due to disease flare. Compared to patients who remained in remission, those who flared during the study showed higher baseline systemic inflammation markers (CRP, neutrophils, platelets) and innate immunity gene expression, with limited corticosteroid response. While gut microbiota composition remained largely stable, transcriptomic analyses revealed distinct changes in ileal gene expression following EXL01 treatment, notably modulation of immune-related genes and upregulation of energy metabolism pathways, similar to what we saw previously in vitro (bioRxiv 2024.10.06.616495).

Conclusion

EXL01, administered during and after CS tapering, was safe and well tolerated for up to 24 weeks in patients with CD. Though limited by small sample size, the study supports the biological activity of EXL01, notably through its impact on ileal gene expression. A phase 2 randomized controlled trial is ongoing to further assess efficacy.

References

1. Rajca S, Grondin V, Louis E, et al. Alterations in the intestinal microbiome (dysbiosis) as a predictor of relapse after infliximab withdrawal in Crohn's disease. Inflamm Bowel Dis. 2014 Jun;20(6):978-86. doi: 10.1097/MIB.0000000000000036. PMID: 24788220.
2. Pascal V, Pozuelo M, Borruel N, et al. A microbial signature for Crohn's disease. Gut. 2017;66(5):813-22.
3. Sokol H, Pigneur B, Watterlot L, et al. Faecalibacterium prausnitzii is an anti-inflammatory commensal bacterium identified by gut microbiota analysis of Crohn disease patients. Proc Natl Acad Sci U S A. 2008;105(43):16731-6.
4. Martin R, Chain F, Miquel S, et al. The commensal bacterium Faecalibacterium prausnitzii is protective in DNBS-induced chronic moderate and severe colitis models. Inflamm Bowel Dis. 2014;20(3):417-30.

Disclosure

HS, PhL and BH are co-founder of Exeliom. PR, DS, BH are employees of Exeliom Biosciences

Introduction

Many prospective studies conducted in healthy volunteers have shown that diet is associated with the risk of developing IBD. By contrast, data are scarce on the role of diet in disease activity in patients with IBD. The aim of the present study was to find dietary factors associated with the risk of IBD relapses.

Aims & Methods

The “Mikinautes” e-cohort is a nationwide French prospective study recruiting IBD patients older than 6 years at inclusion and followed in private practice and public hospitals. French Crohn and Colitis Patients’ association was also involved in the design, funding and recruitment. Patients filled in online questionnaires about their dietary habits and health status. Validated food frequency questionnaires (FFQ) recorded average intakes of 230 food items over the past 12 months. Patients were followed for at least one year and medical records were prospectively collected. Relapse was defined by i) an increased disease activity reported monthly by the patients and/or a clinical report mentioning a relapse, together with ii) a documented consultation with a doctor and iii) a change in IBD therapy, either drug, enteral nutrition and/or surgery).
A posteriori dietary patterns (DPs) were identified using principal component analysis (PCA) performed on 31 food groups. Primary analysis was a Cox model among patients in remission at inclusion to reduce potential reverse causality. Secondary analyses included: i) a Cox model conducted among all patients, which measured the time between the onset of the last relapse before inclusion (or the ongoing relapse at inclusion) and the first relapse during follow-up and ii) a logistic model comparing inactive versus active patients (defined as patients who had at least one relapse at inclusion or over the first year of follow-up). Models were adjusted for age, sex, smoking status and energy intake.

Results

After exclusion of 14 patients with implausible dietary intakes, 693 patients were retained for analyses, comprising 66% of females, 59% Crohn Disease (CD) patients, 34% ulcerative colitis (UC) patients and 7% indeterminate colitis (IC). Median follow-up [Q1-Q3] was 2.1 years [1.6;2.7]. 77% of patients were in remission at inclusion. Their cumulative probability (95% CI) of flare were 18% [15%; 21%] at 1 year and 25% [22%; 29%] at 2 years. In total, 36% of patients had an active disease after one year of follow-up (including active patients at inclusion).
Four DPs were identified through PCA. In CD, primary Cox analysis revealed an association between the second DP and disease activity (HR 95% CI: 0.42 [0.22;0.80] for 3^rdvs 1^st tertile of consumption, p-trend=0.050). This DP was characterized by high intakes of vegetables, fruit, nuts, legumes, cereals, eggs and unsweetened beverages and by a low consumption of dairy products and desserts, bread, sweet/fat or salty/fat products. Secondary analyses confirmed this association (HR: 0.37 [0.18;0.78] for 3^rdvs 1^st tertile, p-trend=0.010; OR: 0.42 [0.22;0.80] for 3^rdvs 1^st tertile, p-trend=0.010). In UC, fruit consumption was protective toward relapse in primary (HR: 0.46 [0.26;0.82] for 3^rdvs 1^st tertile, p-trend=0.009) together with secondary analyses; no dietary pattern was associated with relapse.

Conclusion

A diet characterized by a high consumption of fruit, vegetables, legumes and nuts is associated with a reduction in the risk of relapse by half in patients with CD. High fruit consumption is associated with the same decreased risk of relapse in patients with UC. These results inform dietary counseling in IBD.

References

Acknowledgements: This study was funded by the French association AFA Crohn-RCH France. It was supported by the networks of the Pediatric GETAID, the GETAID, the CREGG, the ANGH and EPIMAD. We thank Nutrinet-Santé, IBD patients and their doctors who contributed to this study.

Disclosure

FC reports receiving research grants from Alpha Wassermann, MaaT pharma, Mayoly Spindler, Medtronic, Nestlé; receiving payment for lectures from AbbVie, AlphaSigma, Ferring, Janssen, Eli Lilly,
Nestlé Health Sciences, Pierre Fabre, Pileje, Takeda, Tillots, Viatris ; receiving consulting fees from AbbVie, Janssen, MaaT pharma, Tillots. HS, SA, CGR, AB, LD and JPH have nothing to declare.