J. R. Fraser Cummings

Confirmation of the role of ATG16l1 as a Crohn's disease susceptibility gene

Background: A German genome‐wide nonsynonymous single nucleotide polymorphism (nsSNP) association study identified ATG16L1 as a Crohns disease (CD) susceptibility gene. The association appeared to be confined to the nsSNP rs2241880 and was confirmed in 2 German independent case‐control collections (combined P = 4.0 × 10−8, odds ratio [OR] 1.45; 95% confidence interval [CI]: 1.21‐1.74), a CD transmission disequilibrium test (TDT) collection, and an independent UK cohort. A weak statistical interaction with CARD15 was demonstrated. No association with ulcerative colitis (UC) was demonstrated. The aims of the study were to replicate the association with CD, examine subphenotype associations and statistical interactions with CARD15, IL23R, and the IBD5 risk haplotype, as well as explore the association with UC. Methods: The study included 645 CD and 676 UC rigorously phenotyped patients recruited from a single UK center. Unaffected controls comprised either spouses of patients (141) or individuals recruited from well‐person clinics (1049). The nsSNP rs2241880 was genotyped using MassArray (Sequenom). Results: A strong association with CD was demonstrated (P = 2.33 × 10−7, OR 1.45 [1.25–1.67]), but no significant association was demonstrated with any subphenotype. We failed to replicate the reported interaction between rs2241880 and the CARD15 low‐risk haplotypes dd and Dd. No significant statistical interaction with the 3 known CD susceptibility genes was seen. No association with UC susceptibility (P = 0.37, OR 1.06 [0.93‐1.22]), or any UC subphenotype was identified. Conclusions: We confirmed the findings that ATG16L1 is a CD susceptibility gene and found no evidence of interaction with CARD15, IL23R, or IBD5.

Contribution of the novel inflammatory bowel disease gene IL23R to disease susceptibility and phenotype.

Background: A North American genome‐wide single nucleotide polymorphism (SNP) association study identified IL23R as a novel inflammatory bowel disease (IBD) susceptibility gene. Association was reported with multiple risk variants in the centromeric portion of IL23R in 3 large independent cohorts. The aims of this study were to replicate the association of IL23R with Crohns disease (CD), examine subphenotype relationships, and look for evidence of epistasis with the known CD susceptibility gene CARD15 and susceptibility haplotype IBD5 in a large collection of CD patients. We further investigated the relationship between IL23R and ulcerative colitis (UC). Methods: In all, 604 CD and 647 UC patients who had been rigorously phenotyped and who had been recruited from a single UK center were used in this study. Controls were either spouses of patients (141) or were recruited from well‐person clinics (993). Eight SNPs were genotyped using MassArray (Sequenom). All 8 SNPs genotyped were significantly associated with CD. Results: The association with the nonsynonymous SNP rs11209026 was confirmed (P = 6.65 × 10−6, odds ratio [OR], 0.43, 95% confidence interval [CI]: 0.29‐0.64). The most significant SNP in our study was rs7517847 (P = 4.9 × 10−9, OR 0.65, 0.56–0.75), which is statistically independent of rs11209026. Preliminary evidence suggests an epistatic interaction with the IBD5 risk haplotype. The effects of mutations in this IL23R appear weaker in UC (P = 0.008, OR 0.63, 0.45–0.89 and 0.005 OR, 0.81, 0.71–0.94, respectively). No subphenotype associations were identified. Conclusions: We confirmed the findings that IL23R is a susceptibility gene for IBD with suggestive epistasis with the IBD5 locus in the CD population.

Medical management of Crohn’s disease

#### Summary pointsnnCrohn’s disease is a chronic, relapsing and remitting inflammatory condition of the gastrointestinal tract. Treatment has changed radically over the past decade with the introduction of biological therapy and increased use of immunomodulators. Awareness of the therapeutic potential and associated adverse events is necessary both for offering benefit and for protecting patients from undue risks from these treatments.nnThe median population incidence of the disease is 6.7 (range 1.6 to 14.6) cases per 100u2009000 annually and prevalence is 140 (10-199) cases per 100u2009000 in the West.1 About 690u2009000 people in Europe, including about 90u2009000 people in the United Kingdom, have the disease, with estimated healthcare costs of €3.04bn (£2.4bn;

The genetics of NOD‐like receptors in Crohn's disease

4.8bn) and £300m a year respectively.2nnThe disease presents at any age, although usually at age 16-30 years; it has a disproportionate effect on economically active individuals. Common presenting symptoms include diarrhoea, abdominal pain, weight loss, and fatigue. The disease is characterised by transmural intestinal inflammation, with occasional extraintestinal features such as …

Association between genetic variants in myosin IXB and Crohn's disease.

The first Crohns disease (CD) susceptibility gene identified was CARD15, which is a member of the emerging NOD-like receptor (NLR) family. These function as intracellular cystosolic pattern recognition receptors (PRRs) and play a central role in the innate immune response. We studied other members of the NLR family using a gene-wide haplotype tagging approach in a well-characterised collection of 547 CD patients and 465 controls. Four single nucleotide polymorphisms (SNPs) in NLRP3 had P values < 0.05 and are in high linkage disequilibrium (LD) with each other (r(2) > 0.90 for all four SNPs). rs4925648 and rs10925019 were the most strongly associated with CD susceptibility (P = 0.001, odds ratio (OR) 1.62, 95% CI 1.2-2.18; and P = 6.5 x 10(-4), OR 1.65, 95% CI 1.23-2.19, respectively). rs1363758 located in NLRP11 was associated with CD susceptibility [P = 0.002 (1.64, 1.19-2.25)], which was weakly confirmed in an independent case-cohort collection on joint analysis [P = 0.05, (1.28, 1-1.64)]. On sub-phenotype analysis, an interesting association between NLRP1 and skin extra-intestinal manifestations and colonic, inflammatory CD was identified. None of these results was replicated in the Wellcome Trust Case Control Consortium study and therefore need replication in a further large cohort.

Clinical Implications of Inflammatory Bowel Disease Genetics on Phenotype

Background: Genetic variation in myosin IXB (MYO9B) was found to be associated with ulcerative colitis (UC) in a recent collaborative study. A nonsynonymous single nucleotide polymorphism (SNP) rs1545620 at the 3′ end of the gene was found to be significantly associated with UC and weakly associated with Crohns disease (CD). The aim of our current study was to replicate these findings in an independent UC cohort and to investigate association with CD. We also investigated subphenotype association and interactions with CARD15, IL23R, ATG16L1, and the IBD5 risk haplotype. Methods: In all, 652 CD patients, 650 UC patients, and 1190 controls were genotyped for 8 MYO9B SNPs. Haplotype testing, epistasis testing with known polymorphisms, and subphenotype analysis were performed. Results: An intronic SNP rs2305767 in the MYO9B gene was associated with inflammatory bowel disease (IBD) overall (corrected P‐value 0.002, odds ratio [OR] 0.76, 95% confidence interval [CI] 0.67–0.86). On individual disease analysis an association was found with CD (corrected P‐value 0.001, OR 0.62, 95% CI 0.53–0.73) but not with UC. Analysis of the common MYO9B haplotypes showed significant association for CD and UC alone and IBD overall. No subphenotypic association was found. These data support an association between CD and SNPs in MYO9B independent of the established effects of SNPs in CARD15, IL23R, ATG16L1, and the IBD5 haplotype. There was no evidence of epistasis between SNPs in MYO9B and these established genes. Conclusions: MYO9B variants may be involved in IBD pathogenesis

Two-stage candidate gene study of chromosome 3p demonstrates an association between nonsynonymous variants in the MST1R gene and Crohn's disease

&NA; The genetic revolution has been with us for over a decade now. We have yet to see this impacting the care of patients except in a few rare examples. However, progress has been made in the field of inflammatory bowel disease (IBD) that could soon be translated to the bedside, both in terms of predicting the disease course as well as in the response to therapy. IBD traditionally has been classified as ulcerative colitis and Crohns disease, with 10% of patients classified as having indeterminate colitis on the basis of clinical, radiologic, endoscopic, and histologic findings. However, this traditional view is now being challenged. Developments in genetics and serological markers, as well as an appreciation of the disease course, have led to an understanding that IBD is a heterogeneous group of diseases with some common genetic and environmental factors but different clinical manifestations in terms of disease behavior, location, and response to treatment. Data are now emerging that may allow us to more objectively select the correct therapy for the correct patient, rather than the current approach, which is based on clinical experience backed up by a less‐than‐perfect evidence base. In this article, we will review the evidence for this.

MicroRNAs in inflammatory bowel diseases: paradoxes and possibilities.

Background: Genomewide linkage studies identified chromosome 3p21 as an IBD locus. Genomewide association studies have supported this locus and the Wellcome Trust Case Control Consortium (WTCCC) study narrowed it to a 0.6 Mb region. Our objectives were to perform a 2‐stage candidate gene association study of the 3p locus and to identify linkage disequilibrium (LD) between significant single‐nucleotide polymorphisms (SNPs) and an Oxfordshire subset (n = 282) of the WTCCC as well as the HapMap SNPs. Methods: A total of 197 SNPs in 53 genes from the 3p locus were genotyped on the Illumina platform in a screening cohort of 469 Crohns disease (CD) patients and 461 controls. Significant associations were then genotyped on the iPLEX platform in the original as well as a second cohort of 139 CD patients, 670 ulcerative colitis (UC) patients, and 1131 controls. All cases and controls were Caucasian and from the Oxfordshire region of the UK. Results: An intronic SNP rs1128535 in the TRAIP gene was associated with CD in the screening and validation cohorts (combined [n = 608] P = 0.0004 [corrected 0.002], odds ratio [OR] 0.77, 95% confidence interval [CI], 0.67–0.89]). No association was seen for UC. Epistasis was seen with the common CARD15 mutations (P = 0.00003 [corrected 0.0006], OR 0.48, 95% CI, 0.34–0.68). No LD was demonstrated with the WTCCC SNPs. Strong LD was demonstrated with 2 nonsynonymous HapMap SNPs in the MST1R gene in an adjacent LD block to the peak WTCCC association, suggesting a distinct association signal. Conclusions: The LD with these functional MST1R variants implicate this gene as having a possible role in CD pathogenesis.

Mo1727 Drug Therapies in Ulcerative Colitis Influence the Expression of MicroRNAs and Cytokines in the Sigmoid Mucosa in an Ex Vivo Model

Abstract:MicroRNAs (miRNAs) are single-stranded RNA molecules, which influence the translation of messenger RNA and hence protein synthesis. The altered expression of miRNAs in disease states in cancer and autoimmune diseases including inflammatory bowel disease is providing new insights into disease pathogenesis. This understanding is leading to consideration of the utility of miRNAs in diagnostics, prognostics, and therapeutics in inflammatory bowel disease. A literature search was conducted using the MEDLINE/PubMed databases using search terms inflammatory bowel disease, miRNA, treatment, and biomarkers.

Mo1728 Cytokine and microRNA Expression in Colonic and Ileal Crohn's Disease Is Modified by Drug Therapy in an Ex Vivo Model

Introduction UC is thought to be characterised by a TH-2 predominant immune response. MicroRNAs (miRNA) are short 19–23 nucleotide long strands of single-stranded RNA which modify post transcriptional gene expression by degrading or inhibiting translation of mRNA. miR31 and miR155 are miRNA that directly and indirectly target genes in the IL-13 dependant pathway. Confounding factors in current IBD miRNA studies include disease activity and medication. The specific function of miRNAs in UC, how they might be influenced by medications, and potential clinical utility is yet to be investigated. In this study we aim to investigate how the expression of a number of miRNA and mRNA is influenced by medications and then explore these findings further using a human ex-vivo culture system. Method Sigmoid biopsies from 40 patients with untreated active and inactive UC, normal controls, and patients treated with 5-ASA, Azathioprine, Infliximab and Adalimumab were frozen in liquid nitrogen. RT-qPCR was performed to analyse the relative expression of miRNA and mRNA. An ex vivo model was designed. 6 biopsies were taken from 8 patients with active untreated UC, with a Mayo score of 6 or greater. Biopsies were placed in tissue culture media in 6 separate wells with either no treatment, 5-ASA, 6-TG, Infliximab, or Adalimumab, were incubated for 24hrs at 37 o C. RT-qPCR was performed to assess expression of miR31 and miR155 and mRNA of CCL18 and SOCS1 that are important in the TH-2 pathway. Results miR31 and miR155 expression was increased in patients with untreated active and inactive UC compared to normal mucosa. CCL18 expression was also increased in untreated active disease compared to inactive and normal mucosa. Azathioprine reduced miR31, miR155 and CCL18 expression to levels seen in quiescent disease. Infliximab treatment reduced miR155, and CCL18 expression to that of inactive disease, which was not seen in Adalimumab. Anti-TNFs did not reduce miR31 expression to the level of untreated inactive disease. 5ASAs had no impact on expression of the miRNA nor mRNA studied. Ex vivo treated samples confirmed down regulation of miR31 was confined to 6-TG and not seen in anti-TNF treatment, and that CCL18 expression is also reduced by 6-TG and Infliximab, but not Adalimumab. Conclusion Our data confirmed that miR31 miR155 and CCL18 are increased in active UC compared to normal mucosa and inactive disease. We demonstrate that thiopurine therapy reduces expression of miR31 and CCl18 in contrast to anti-TNF agents. This model may have utility in increasing understanding of the mechanisms of action of IBD medications. It highlights the importance of controlling for medication when analysing miRNA expression in whole tissue biopsies. Disclosure of interest None Declared.