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KONO-S ANASTOMOSIS REDUCES ENDOSCOPIC AND SURGICAL POST-OPERATIVE RECURRENCE IN CROHN’S DISEASE: THE SUPREME-CD TRIAL UPDATE
Aconitate decarboxylase 1 (ACOD1; also known as IRG-1) is the ubiquitous source of the metabolite itaconate, which dampens inflammasome activation by preventing HIF1α production of IL-1β and prevents TNF production through NRF2 activation. We reasoned that expression of ACOD1 is an inhibitory pathway within monocytes to curtail their inflammatory function, which could be manipulated in IBD therapy.
Methods
We performed single-cell RNA sequencing of monocytes from colonoscopic biopsies of treatment naïve IBD patients, as well as complementary analysis of DSS treated C57BL/6J (WT) mice, using single-cell protein-mRNA (Primeflow) technologies for validation. To determine the role of ACOD1 in colitis, Acod1-/- mice were administered 2% DSS in drinking water for 6 days (acute colitis) or 4 days followed by 14 day recovery period (inflammation resolution) and assessed the colon myeloid compartment by multi-parameter flow cytometry. This was complemented with competitive bone marrow chimeric mice to assess the intrinsic role of ACOD1 controlling monocyte survival and proinflammatory status. Transcriptional profiling (Nanostring) was used to determine the molecular pathways influenced by ACOD1.
Results
We found that ACOD1/Acod1 represented an evolutionarily conserved of feature of inflammation-associated monocytes, with expression present in mucosal but not blood monocytes. Acod1-/- mice demonstrated increased susceptibility to acute intestinal injury with DSS compared to co-housed WT controls, characterised by increased weight loss and colon shortening with significant accumulation of colonic neutrophils and CD64+Ly6C+ MHC-II+/– monocytes. Consistent with Acod1 limiting inflammasome activation, Acod1-/- monocytes demonstrated significantly greater IL-1b producing capabilities than WT controls.
To assess Acod1 deficient and sufficient haemopoietic cells in in the same environment, we generated competitive bone marrow chimeric mice. Acod1 deficiency conferred a significant advantage to monocyte recruitment to the colon, characterised by a 10fold increase in Ly6C+ MHC-II–colon monocytes in colitis.
Transcriptional profiling of WT and Acod1-/- colonic monocytes revealed significantly greater expression of molecules required for monocyte extravasation, including Sell (CD62L), Icam1, Ccr2 and Itgal (CD11a). In addition, expression of genes indicative of macrophage maturation such as Itgax (CD11c), Cd163 and Cx3cr1 were lower in the context of Acod1 deficiency, suggesting that normal maturation of these cells may be perturbed by loss of Acod1.
Conclusion
ACOD1/Acod1 represents an evolutionarily conserved, monocyte specific immuno-metabolic pathway that acts to limit monocyte recruitment, fate and pro-inflammatory function within the inflamed intestine.

% change in starting weight of Acod1-/- or C57BL6/JCrl (WT) mice after 6 days of 2% DSS (A) or 4 days plus 14 days normal drinking water (B). (C) Flow cytometry plots of colon lamina propria CD64+ cells isolated from (B) with relative proportions of Ly6C+ MHC-II- monocytes (P1) Ly6C+ MHC-II+ monocytes (P2) and Ly6C- MHC-II+ macrophages (P3/4) and the proportion of IL-1β expressing P1 cells observed after 2hours incubation with 1ug/ml Momensin (D).Congenic CD45.1/CD45.2+ mice were lethally irradiated and reconstitiuted with WT CD45.1 or Acod1-/- CD45.2 BM, before treatment with 6 days 2% of DSS or normal drinking water. (E) Chimerism of colon CD64+ cells relative to blood monocytes from gating strategy in (C). (F) CD45.1 and CD45.2 P1 and P2 moncytes from DSS treated mice were purified by flow cytometry, RNA extracted and profiled by Nanostring.
Methods:
Anti-αvβ6 autoantibodies were measured in 4 longitudinal serum samples collected from 82 subjects who later developed UC and 82 matched controls (HC) from a Department of Defense pre-clinical cohort (PREDICTS). The longitudinal samples were collected at the following timepoints: Sample A at the time of UC diagnosis, Sample B at a median of 2 years before Sample A, Sample C at a median of 4 years before Sample A and Sample D at a median of 10 years before Sample A. In a distinct, external validation cohort (GEM), we tested 12 pre-UC subjects and 49 matched controls. Further, anti-αvβ6 were measured in 2 incident UC cohorts (COMPASS n=55 and OSCCAR n=104) and associations between anti-αvβ6 and UC-related outcomes were defined using Cox proportional-hazards model.
Results:
Anti-αvβ6 autoantibodies were significantly higher among individuals who developed UC compared to controls up to 10 years before diagnosis in PREDICTS (Figure 1A), and this was validated in the GEM cohort in which samples were collected at a median of 4.2 years before diagnosis (Figure 1B). During the pre-clinical phase (Sample D to A), the proportion of anti-αvβ6 seropositive subjects increased from 12.2% (Sample D) to 20.7% (Sample C) to 30.5% (Sample B) to 52.4% (Sample A) of subjects who developed UC (Chi-square test for trend p<0.0001), compared with a mean of 2.7% in HC group across the 4 timepoints in the PREDICTS cohort (Figure 1C). Anti-αvβ6 predicted UC development with an AUC of at least 0.8 up to 10 years before diagnosis (Figure 1D). Finally, high levels of anti-αvβ6 were associated with a composite of adverse UC-outcomes including hospitalization, disease extension, colectomy, systemic steroid use and/or escalation to biologic therapy in both cohorts of recently diagnosed UC patients (Figure 1E).
Conclusion:
Anti-integrin αvβ6 autoantibodies precede the clinical diagnosis of UC by up to 10 years and are associated with adverse UC-related outcomes. The presence of anti-αvβ6 autoantibodies before UC diagnosis suggests that a pre-clinical phase, potentially amenable to therapeutic intervention may indeed predate UC diagnosis.

The serum metabolome contains a plethora of endogenously produced and environmentally absorbed biomarkers that could serve as Crohn’s disease(CD) markers. However, most metabolomic studies have used post-diagnosis samples, which may be a response to CD disease activity as opposed to a predictor of disease evolution. Moreover, the association between pre-diagnosis CD risk metabolites and other biomarkers remains unknown. We aimed to assess serum metabolite alterations in the pre-clinical phase of CD and assess the correlation of these alterations with other known risk factors
Methods
In a cohort of prospectively followed healthy first-degree relatives(FDRs) enrolled in the Genetics Environment Microbial Project(CCC-GEM), subjects who later developed CD(n=78) were matched up to 1:4 with control FDRs that remained healthy(n=312). At enrollment, we measured untargeted serum metabolites using the combination of four ultrahigh performance liquid chromatography-tandem mass spectroscopy; subclinical inflammation using fecal calprotectin(FCP) assay; gut barrier function using urinary fractional excretion of lactulose to mannitol ratio(LMR) assay. We used conditional logistic regression to identify serum metabolites associated with CD onset and used marginal correlation analysis to assess the association of specific CD-associated metabolites with CD risk factors described above. Finally, random forests were performed to assess individual pre-diagnostic risk biomarkers' predictive potential for CD and their area under the curve(AUC) based on 5-fold cross validation were compared using the Delong test. Associations with false discovery rate-adjusted P values(q)<0.05 were considered significant
Results
In total, 63 of 1001 pre-diagnostic serum metabolites were associated with CD onset(1×10-4<q<0.05), including enrichments for sphingomyelin and aspartate, and depletions for isoleucine, leucine and valine. Among the identified serum metabolites, 20 were correlated with increased FCP(2×10-5<q<0.05) and two with elevated LMR(0.002<q<0.02). A metabolome-based model to predict CD yielded an AUC of 0.89(95% CI 0.85-0.92), which was significantly superior to the LMR-based model with an AUC of 0.48(95% CI 0.40-0.56, p<0.001) and the FCP-based model having an AUC of 0.70(95% CI 0.59-0.81, p<0.001)
Conclusion
Our study revealed alterations in serum metabolites in pre-diagnostic CD, some of these were related to gut barrier function and subclinical inflammation. The model based on serum metabolites performed better than LMR and FCP in predicting CD, suggesting metabolomic pathways other than intestinal permeability and inflammation that contribute to CD development. Identified serum metabolites may provide potential insights into disease pathogenesis, and the prediction model's good performance may offer a novel opportunity for CD prevention and early diagnosis
AIM: This trial aimed to provide randomized controlled data comparing Kono-S anastomosis and conventional stapled ileocolic side-to-side anastomosis in patients with CD.
METHODS: Randomized controlled trial (RCT) at a tertiary referral centre. The primary endpoints are endoscopic recurrence rate (Rutgeerts score > i2) after 6 months and surgical recurrence at 60 months. Secondary endpoints are clinical recurrence (CR) rate after 24 and 36 months, and SR after 24 and 36 months. Statistics was performed by using standard analyses. Furthermore, a sample size able to consent the detection of a reduction >30% in total endoscopic recurrence (when assuming a 60% to 65% ER expected rate in the control group and a 30% in the case group) was calculated; to allow a 10% drop-out of patients, 36 patients per group were needed (72 patients in the entire population).
RESULTS: A total of 119 CD patients were included in the study. In all, 51 ileocolic CD patients were randomized in the Kono group and 68 in the conventional side-to-side group.
At 6 months, 17 (33.3%) patients in the Kono group and 51 (75%) patients in the conventional group presented an ER (Rutgeerts score>i2) at colonoscopy (p<0.001). A severe post-operative ER (Rutgeerts score>i3) was found in 8 (15.7%) patients in the Kono group versus 23 (33.8%) patients in the conventional group (p=0.03).
At 24 months, CR rate was 19.6% in the Kono group versus 30.9% in the conventional group (p=0.2), while SR rate was 0% in the Kono group versus 4.4% (3 patients) in the standard group (p=0.2).
At 36 months, CR rate was 19.6% in the Kono-s group versus 33.8% in the conventional group (p=0.1). At 36 months, no patients (0%) in the Kono group experienced SR, while 6 patients (8.8%) in the conventional group did (p=0.03).
Furthermore, when considering a median follow-up of 54 (42-60) months at interim analysis of SR, the difference between the two groups was confirmed to be significant (0% vs 8.8%; p=0.029).
CONCLUSIONS: This is the first RCT comparing Kono-S anastomosis and conventional side-to-side anastomosis in CD. The results demonstrate a significant reduction in postoperative endoscopic and surgical recurrence rates, no safety issues, and a favourable trend for lower clinical recurrence rate in patients who underwent Kono-S anastomosis compared to the conventional side-to-side anastomosis.

6-Months Endoscopic Recurrence Rate (Rutgeerts score>i2) and Severe Endoscopic Recurrence Rate (Rutgeerts score>i3) in Side-to-Side VS Kono groups.

Surgical view of the Kono-S anastomosis