Integration of transcriptomics and metabonomics: improving diagnostics, biomarker identification and phenotyping in ulcerative colitis

A systems biology approach to multi-faceted diseases has provided an opportunity to establish a holistic understanding of the processes at play. Thus, the current study merges transcriptomics and metabonomics data in order to improve diagnostics, biomarker identification and to explore the possibilities of a molecular phenotyping of ulcerative colitis (UC) patients. Biopsies were obtained from the descending colon of 43 UC patients (22 active UC and 21 quiescent UC) and 15 controls. Genome-wide gene expression analyses were performed using Affymetrix GeneChip Human Genome U133 Plus 2.0. Metabolic profiles were generated using (1)H Nuclear magnetic resonance spectroscopy (Bruker 600 MHz, Bruker BioSpin, Rheinstetten, Germany). Data were analyzed with the use of orthogonal-projection to latent structure-discriminant analysis and a multivariate logistic regression model fitted by lasso. Prediction performance was evaluated using nested Monte Carlo cross-validation. The prediction performance of the merged data sets and that of relative small (<20 variables) multivariate biomarker panels suggest that it is possible to discriminate between active UC, quiescent UC, and controls; between patients with or without steroid dependency, as well as between early or late disease onset. Consequently, this study demonstrates that the novel approach of integrating metabonomics and transcriptomics combines the better of the two worlds, and provides us with clinical applicable candidate biomarker panels. These combined panels improve diagnostics and more importantly also the molecular phenotyping in UC and provide insight into the pathophysiological processes at play, making optimized and personalized medication a possibility. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11306-013-0580-3) contains supplementary material, which is available to authorized users

Influence of Smoking on Colonic Gene Expression Profile in Crohn's Disease

BACKGROUND: The development and course of Crohn's disease (CD) is related to both genetic and environmental factors. Smoking has been found to exacerbate the course of CD by increasing the risk of developing fistulas and strictures as well as the need for surgery, possibly because of an interaction between smoking or nicotine on macrophage function and the intestinal microvasculature. Several genes are involved in the pathogenesis of CD, and in this study the gene expression differences of the descending colonic mucosa were investigated in CD (smokers or never smokers) and controls (smokers or never smokers). AIM: To identify any difference in gene expression of the descending colonic mucosa between smoking and never-smoking CD patients (and controls) by determining genetic expression profiles from microarray analysis. METHODS: Fifty-seven specimens were obtained by routine colonoscopy from the included material: CD smokers (n = 28) or never-smokers (n = 14) as compared to fifteen healthy controls (8 smokers and 7 never-smokers). RNA was isolated and gene expression assessed with Affymetrix GeneChip Human Genome U133 Plus 2.0. Data were analyzed by principal component analysis (PCA), Wilcoxon rank sum test and multiple linear regressions. Real-time (RT) PCR was subsequently applied to verify microarray results. RESULTS: The PCA analysis showed no intrinsic clustering of smokers versus never-smokers. However, when Wilcoxon rank sum test corrected with Q values were performed, six known genes were significantly expressed differently in the inflamed CD smokers as compared to the inflamed CD never-smokers: ring finger protein 138 (RNF138), metalothionein 2A (MT2A) and six transmembrane epithelial antigen of the prostate 3 (STEAP3), SA hypertension-associated homolog, PGM2L1 and KCNJ2. The subsequent RT-PCR-analyses verified, however, that only RNF138, MT2A and STEAP3 were significantly up-regulated in CD smokers in specimens with inflammatory activity of the descending colon. CONCLUSIONS: The present study demonstrates that the genes, RNF138, MT2A, and STEAP3 are differently expressed in the inflamed descending colon of smoking versus never-smoking CD patients, which might be of relevance for the poorer clinical course among CD smokers. Many gastroenterologists are still not totally aware of the benefits of smoking cessation in relation to CD, and do not put much effort into getting the patients to quit, therefore more information on the negative effects of smoking, seems warranted

Metabonomics uncovers a reversible proatherogenic lipid profile during infliximab therapy of inflammatory bowel disease

Background: One-third of inflammatory bowel disease (IBD) patients show no response to infliximab (IFX) induction therapy, and approximately half of patients responding become unresponsive over time. Thus, identification of potential treatment response biomarkers are of great clinical significance. This study employs spectroscopy-based metabolic profiling of serum from patients with IBD treated with IFX and healthy subjects (1) to substantiate the use of spectroscopy as a semi-invasive diagnostic tool, (2) to identify potential biomarkers of treatment response and (3) to characterise the metabolic changes during management of patients with tumour necrosis factor-α inhibitors.Methods: Successive serum samples collected during IFX induction treatment (weeks 0, 2, 6 and 14) from 87 IBD patients and 37 controls were analysed by 1H nuclear magnetic resonance (NMR) spectroscopy. Data were analysed with principal components analysis and orthogonal projection to latent structures discriminant analysis using SIMCA-P+ v12 and MATLAB.Results: Metabolic profiles were significantly different between active ulcerative colitis and controls, active Crohn's disease and controls, and quiescent Crohn's disease and controls. Metabolites holding differential power belonged primarily to lipids and phospholipids with proatherogenic characteristics and metabolites in the pyruvate metabolism, suggestive of an intense inflammation-driven energy demand. IBD patients not responding to IFX were identified as a potentially distinct group based on their metabolic profile, although no applicable response biomarkers could be singled out in the current setting.Conclusion: 1H NMR spectroscopy of serum samples is a powerful semi-invasive diagnostic tool in flaring IBD. With its use, we provide unique insights into the metabolic changes taking place during induction treatment with IFX. Of distinct clinical relevance is the identification of a reversible proatherogenic lipid profile in IBD patients with active disease, which partially explains the increased risk of cardiovascular disease associated with IBD

Low-dose azathioprine and allopurinol versus azathioprine monotherapy in patients with ulcerative colitis (AAUC)::An investigator-initiated, open, multicenter, parallel-arm, randomised controlled trial

BACKGROUND: Retrospective studies suggest that for patients with ulcerative colitis (UC) combination therapy with low-dose azathioprine and allopurinol (L-AZA/ALLO) may result in higher remission rates than monotherapy with azathioprine (AZA). We prospectively investigated the effects of these drugs for remission in patients with moderate-to-severe UC. METHODS: Open-label, unblinded, randomised, controlled, investigator-initiated, multicentre study conducted at eight hospital sites in Denmark. Adult patients with established UC, who were steroid dependent/refractory, thiopurine naïve, had a normal thiopurine methyltransferase, and achieved remission with steroids or infliximab were eligible for inclusion. Patients were randomly assigned by the investigators (1:1) to 52 weeks of treatment with once daily oral AZA (median dose 50 mg) combined with ALLO 100 mg versus AZA monotherapy (median dose 200 mg), using a computer-generated randomisation list with blocks of six. The trial was open without masking. All randomised patients who received at least one dose of study drug were included in primary and safety analyses (intention to treat population). The primary outcome was steroid and infliximab free remission after 52 weeks, defined as a Mayo Score of ≤1 and no rectal bleeding. The trial is completed and is registered in ClinicalTrials.gov (ClinicalTrials.gov NCT03101800). FINDINGS: Between January 9, 2017 and February 10, 2021, 47 patients were randomised to l-AZA/ALLO and 42 to AZA and received at least one dose of the study drug. After 52 weeks, 20 of 47 (43%) patients in the l-AZA/ALLO group and nine of 42 (21%) patients in the AZA group achieved remission (odds ratio 2·54 [95% CI 1·00 to 6.78, p < 0·048]). Fourteen patients (30%) in the l-AZA/ALLO group and 16 (38%) in the AZA group were withdrawn from the study due to adverse events. INTERPRETATION: This study suggests that after one year l-AZA/ALLO therapy may be associated with a beneficial effect on steroid- and infliximab-free clinical remission in patients with moderate-to-severe UC and should be considered as first line therapy. FUNDING: Funding for AAUC was provided by The Capital Region of Denmark (Regionernes Medicinpulje (6062/16))