Thiopurine metabolite ratios for monitoring therapy in pediatric Crohn disease.

OBJECTIVES: Thiopurines (azathioprine, 6-mercaptopurine) are a mainstay of treatment in Crohn disease (CD). Monitoring intracellular metabolite (6-thioguanine nucleotides [6-TGN] and 6-methylmercaptopurine [6-MMP]) levels can help optimize therapeutic efficacy and minimize potential toxicity. Determination of 6-MMP/6-TGN ratios may provide additional useful information, such as the identification of individuals with excessive thiopurine methyltransferase activity and disadvantageous 6-MMP overproduction. These patients are at increased risk of therapeutic failure and hepatotoxicity. The aim of the study was to evaluate the correlation of 6-MMP/6-TGN ratios with therapeutic efficacy and risk of hepatotoxicity in CD. METHODS: The present study was a single-center cross-sectional study including pediatric patients with CD studied prospectively with clinical and laboratory assessments along with serial measurements of 6-MMP and 6-TGN. Clinical response was determined using established clinical indices. RESULTS: The study included 238 pediatric patients with CD with a total of 1648 evaluation points. The patients were in steroid-free remission at 59.1% of the evaluation points. 6-MMP/6-TGN ratios of 4 to 24 were protective against relapse (odds ratio [OR] 0.52, 95% confidence interval [CI] -0.39 to 0.69, P = 0.001). Hepatotoxicity was associated with high 6-MMP levels (>3919  pmol/8 × 10 red blood cell count: OR 7.65, 95% CI 3.7-15.9, P = 0.001) and high 6-MMP/6-TGN ratios (>24: OR 5.35, 95% CI -3.43 to 8.43, P = 0.001). CONCLUSIONS: We observed significant associations between 6-MMP/6-TGN ratios and clinical response, and risk of hepatotoxicity. Our results suggest that determination of thiopurine metabolite ratios is a valuable tool for identification of patients at increased risk of therapeutic failure and hepatotoxicity.

Phenotypic Variation in Paediatric Inflammatory Bowel Disease by Age: A Multicentre Prospective Inception Cohort Study of the Canadian Children IBD Network.

BACKGROUND AND AIMS: Incidence of paediatric inflammatory bowel disease [IBD] in Canada is among the highest worldwide, and age of onset may be decreasing. In a multicentre nationwide inception cohort study, we examined variation in phenotype of IBD throughout the paediatric age spectrum. METHODS: Children aged ≥2 years [y] and <17y [A1 age at diagnosis], with new onset IBD, were systematically evaluated at sites of the Canadian Children IBD Network. Prospectively recorded phenotypic data were compared between age groups. RESULTS: Among 1092 children (70% Caucasian; 64% Crohn's disease [CD], 36% ulcerative colitis/inflammatory bowel disease unclassified [UC/IBD-U]; median age 13 y, interquartile range [IQR] 11-15 y), 210 [19%] were diagnosed before the age of age 10 y [Paris A1a] and 43 [4%] before age 6 y (very-early-onset [VEO-IBD]). CD was less common in younger children [42%, 56%, 66%, respectively, of VEO-IBD, A1a; A1b]. Colon-only IBD [UC/IBDU or CD-colon] was present in 81% of VEO-IBD and 65% of A1a; ileal disease increased progressively, reaching plateau at age 10 y. CD location was ileocolonic [L3] in 53% overall. Ileitis [L1] increased with age [6% of VEO-IBD; 13% of A1a; 21% of A1b], as did stricturing/penetrating CD [4% of A1a; 11% of A1b]. At all ages UC was extensive [E3/E4] in >85%, and disease activity moderate to severe according to Physician's Global Assessment [PGA] and weighted Paediatric Crohn's Disease Activity Index/Paediatric Ulcerative Colitis Activity Index [wPCDAI/PUCAI] in >70%. Heights were modestly reduced in CD [mean height z score -0.30 ± 1.23], but normal in UC/IBD-U. CONCLUSIONS: Paris classification of age at diagnosis is supported by age-related increases in ileal disease until age 10 years. Other phenotypic features, including severity, are similar across all ages. Linear growth is less impaired in CD than in historical cohorts, reflecting earlier diagnosis.

13C urea breath testing to diagnose Helicobacter pylori infection in children.

The casual relationship between Helicobacter pylori colonization of the gastric mucosa and gastritis has been proven. Endoscopy and subsequent histological examination of antral biopsies have been regarded as the gold standard for diagnosing H pylori gastritis. The C urea breath test is a noninvasive test with a high specificity and sensitivity for H pylori colonization. Increasingly, it is becoming an important tool for use in diagnosing H pylori infection in pediatric populations. This test is particularly well suited for epidemiological studies evaluating reinfection rates, spontaneous clearance of infection and eradication rates after therapy. However, few groups have validated the test in the pediatric age group. The testing protocol has not yet been standardized. Variables include fasting state, dose of urea labelled with 13C, delta cutoff level of 13C carbon dioxide, choice of test meal and timing of collection of expired breath samples. Further studies are urgently needed to evaluate critically the impact of H pylori infection in the children. The 13C urea breath test should prove very useful in such prospective studies.

Perforated duodenal ulcer in a pediatric patient with eosinophilic gastroenteritis.

An 11-year-old boy with eosinophilic gastroenteritis treated by an elimination diet alone presented with a perforated gastroduodenal ulcer subsequent to blunt trauma to the abdomen. Peripheral eosinophilia, chronic iron deficiency, chronic hypoalbuminemia and severe failure to thrive had been present since age 2 years. Immunological work-up revealed food allergies, documented by skin tests. A review of the literature since 1966 revealed only six other cases of perforation of the gastrointestinal tract, one of whom was also a child.

Susceptibility loci reported in genome-wide association studies are associated with Crohn's disease in Canadian children.

BACKGROUND: Recent genome-wide association studies based on adult and paediatric populations have implicated >30 genes/loci as susceptibility loci for Crohn's disease (CD). AIMS: To investigate whether reported genes/loci were also associated with CD in Canadian children. DESIGN AND METHODS: A case-control design was implemented at three paediatric gastroenterology clinics in Canada. Children < or =18 years of age with a confirmed diagnosis of CD were recruited along with controls. Single nucleotide polymorphisms (SNPs) in five genome-wide association studies reported genes/loci were genotyped. Associations between individual SNPs and CD were examined. RESULTS: A total of 406 cases and 415 controls were studied. The mean (+/-s.d.) age of the cases was 12.3 (+/-3.2) years. Most cases were male (56.6%), had ileo-colonic disease (L3 +/- L4, 52.0%) and inflammatory behaviour (B1 +/- p, 86.9%) at diagnosis. Allelic association analysis (two-tailed) showed that three of the five targeted SNPs were significantly associated with overall susceptibility for CD (ZNF365, r10995271, P = 0.001; PTPN2, rs1893217, P = 0.005; STAT3, rs744166, P = 0.01). Associations with SNP rs4613763 in the PTGER4 locus were marginally nonsignificant (P = 0.07). The ZNF365 and STAT3 SNPs were predominantly associated with ileal disease with or without colonic involvement. CONCLUSION: The identified susceptibility genes/loci for adult-onset CD also confer risk for paediatric-onset CD.

Interleukin 10 (IL-10) gene variants and susceptibility for paediatric onset Crohn's disease.

BACKGROUND: A recent genome-wide association study in adult patients with ulcerative colitis (UC) has implicated the interleukin 10 (IL-10) gene as an important candidate gene. Moreover, a UC-associated single nucleotide polymorphism (SNP) rs3024405 was also significantly associated with adult Crohn's disease (CD). AIMS: To examine whether IL-10-CD associations extended to paediatric-onset CD. METHODS: We implemented the case-control design at three paediatric gastroenterology clinics in Canada. CD patients (

Gene-centric association mapping of chromosome 3p implicates MST1 in IBD pathogenesis.

Association mapping and candidate gene studies within inflammatory bowel diseases (IBD) linkage regions, as well as genome-wide association studies in Crohn's disease (CD) have led to the discovery of multiple risk genes, but these explain only a fraction of the genetic susceptibility observed in IBD. We have thus been pursuing a region on chromosome 3p21-22 showing linkage to CD and ulcerative colitis (UC) using a gene-centric association mapping approach. We identified 12 functional candidate genes by searching for literature cocitations with relevant keywords and for gene expression patterns consistent with immune/intestinal function. We then performed an association study composed of a screening phase, where tagging single nucleotide polymorphisms (SNPs) were evaluated in 1,020 IBD patients, and an independent replication phase in 745 IBD patients. These analyses identified and replicated significant association with IBD for four SNPs within a 1.2 Mb linkage disequilibrium region. We then identified a non-synonymous coding variant (rs3197999, R689C) in the macrophage-stimulating 1 (MST1) gene (P-value 3.62 x 10(-6)) that accounts for the association signal, and shows association with both CD and UC. MST1 encodes macrophage-stimulating protein (MSP), a protein regulating the innate immune responses to bacterial ligands. R689C is predicted to interfere with MSP binding to its receptor, suggesting a role for this gene in the pathogenesis of IBD.

Modulation of cytokine release from colonic explants by bacterial antigens in inflammatory bowel disease.

The intestinal flora play an important role in experimental colitis and inflammatory bowel disease (IBD). Using colonic explant cultures from 132 IBD and control subjects, we examined tumour necrosis factor-alpha (TNF-alpha), interleukin (IL)-1 and interleukin-1 receptor antagonist (IL-1RA) production in vitro in response to bacterial activators. Unstimulated TNF-alpha release was increased significantly in rectal biopsies from involved IBD tissue, correlating with inflammation severity. Whereas lipopolysaccharide (LPS) only moderately stimulated TNF-alpha production from inflamed tissue, pokeweed mitogen (PWM) induced its release in all groups, with a stronger response in involved IBD tissue. Superantigen staphylococcal enterotoxin A (SEA) had a similar, but weaker effect. SEB was observed to be the strongest inducer of TNF-alpha for all groups, again with a more marked response in inflamed tissue. Stimulated release of IL-1 was considerably less than for TNF-alpha. The superantigens' superior potency over LPS was not as marked for IL-1 as it was for TNF-alpha. In addition to IL-1, IL-1RA release was also triggered by the bacterial products. The net effect of activation on the IL-1RA/IL-1 ratio was relatively modest. Release of the proinflammatory cytokines TNF-alpha and IL-1, as well as that of the anti-inflammatory cytokine IL-1RA was increased by incubation of colonic tissue with bacterial factors. TNF-alpha production and release was increased significantly in involved colonic explants from IBD. SEB was even capable of inducing TNF-alpha release from uninvolved colonic tissue.

Accuracy of the [13C]-urea breath test in diagnosing Helicobacter pylori gastritis in pediatric patients.

BACKGROUND: The causal association between Helicobacter pylori (H. pylori) colonization of the gastric mucosa and gastritis is now well established. Histologic examination of endoscopic biopsy specimens has long been regarded as the gold standard for diagnosis. However, the changes can be focal in nature and presence of the organism may be missed in nonsampled areas. The urea breath test, which uses a stable isotope, offers distinct advantages, in that it is noninvasive and measures the activity of the micro-organism. It thus represents a potentially invaluable tool in the initial diagnosis of the infection and in verifying its eradication. METHODS: The study design was that of a prospective, blinded comparison of the [13C]-urea breath test with histologic assessment of antral biopsy specimens using the Warthin-Starry stain, to diagnose H. pylori infection in a group of 79 consecutive pediatric patients. RESULTS: Patients classified as negative by histology (n=67) had breath 13C enrichment of 0.97+/-0.07 delta per thousand (mean+/-SEM), with a range of -0.20 and 2.83 delta per thousand. In contrast, those with a positive histologic results (n=12) had an enrichment of 25.41+/-5.01 delta per thousand (range, 3.43-58.80; p < 0.001). At the chosen cutoff point of 3 delta per thousand, the sensitivity and specificity as well as the positive and negative predictive values of the breath test were uniformly 100%. CONCLUSION: The [13C]-urea breath test is a highly reliable, noninvasive method for the diagnosis of H. pylori gastritis in children and adolescents.