Azathioprine is effective for oral involvement in Crohn's disease but not for orofacial granulomatosis alone.

BACKGROUND: There have been no previous reports assessing the effectiveness of azathioprine (AZA) in the treatment of orofacial granulomatosis (OFG). This report is a review of patients receiving AZA for active OFG with or without concomitant gut Crohn's disease (CD) in a specialist tertiary referral centre. METHODS: Clinical response was defined by Global Physician Assessment at 4-, 12- and 24-month follow-up and a standardised oral disease activity score (ODAS). RESULTS: Sixty of 215 patients seen with OFG in our clinic over a 12-year period were treated with AZA. Of these, 22 had concomitant CD. The proportion of patients responding to AZA with a diagnosis of CD/OFG vs. OFG only at 4, 12 and 24 months were 54% vs. 21% (P = 0.03), 59% vs. 21% (P = 0.003) and 41% vs. 24% (P = 0.16), respectively. A statistically significant difference was seen between starting and follow-up ODAS scores at 4 months in the CD/OFG group which was not observed in the OFG only group. Factors predicting a need for AZA included a diagnosis of intestinal CD, sulcal swelling, sulcal ulcers and upper lip involvement. The factor predicting response to treatment was a diagnosis of CD at 12 months of follow-up. No difference in the number of adverse effects was observed between the two groups of patients. CONCLUSIONS: AZA is significantly more effective in the treatment of oral disease with a concurrent diagnosis of CD rather than in the treatment of OFG alone.

A practical guide to the use of thiopurines in oral medicine.

Thiopurines are widely used as first-line immunosuppressive therapies in the management of chronic inflammatory oral disease. However, despite over half a century of clinical experience, the evidence base for their use is limited. The aims of this paper were to review the evidence for the use of thiopurines in oral medicine and provide a contemporary model of thiopurine metabolism and mechanism of action and a rationale for clinical use and safe practice.

Defective macrophage handling of Escherichia coli in Crohn's disease.

BACKGROUND AND AIM: Escherichia coli can be isolated from lamina propria macrophages in Crohn's disease (CD), and their intramacrophage persistence may provide a stimulus for inflammation. To further determine the contributions of macrophage dysfunction and E. coli pathogenicity to this, we aimed to compare in vitro functioning of macrophages from patients with CD and healthy controls (HC) in response to infection with CD-derived adherent-invasive E. coli (AIEC) and less pathogenic E. coli strains. METHODS: Monocyte-derived macrophages were cultured from patients with CD and HC. Intramacrophage survival of E. coli strains (CD-derived adherent-invasive [AI] and non-AI strains and laboratory strain K-12) was compared. Macrophage cytokine release (tumor necrosis factor alpha [TNFα], interleukin [IL]-23, IL-8 and IL-10) and monocyte phagoctyosis and respiratory burst function were measured after E. coli infection. For CD patients, laboratory data were correlated with clinical phenotype, use of immunomodulation, and CD risk alleles (NOD2, IL-23R, ATG16L1 and IRGM). RESULTS: Attenuated TNFα and IL-23 release from CD macrophages was found after infection with all E. coli strains. There was prolonged survival of CD-derived AIEC, CD-derived non-AIEC and E. coli K-12 in macrophages from CD patients compared to within those from HC. No abnormality of monocyte phagocytosis or respiratory burst function was detected in CD. Macrophage dysfunction in CD was not influenced by phenotype, use of immunomodulation or genotype. CONCLUSIONS: CD macrophage responses to infection with E. coli are deficient, regardless of clinical phenotype, CD genotype or E. coli pathogenicity. This suggests host immunodeficiency is an important contributor to intramacrophage E. coli persistence in CD.

Pharmacogenetics of pemetrexed combination therapy in lung cancer: pathway analysis reveals novel toxicity associations.

Identification of polymorphisms that influence pemetrexed tolerability could lead to individualised treatment regimens and improve quality of life. Twenty-eight polymorphisms within eleven candidate genes were genotyped using the Illumina Human Exome v1.1 BeadChip and tested for their association with the clinical outcomes of non-small cell lung cancer and mesothelioma patients receiving pemetrexed/platinum doublet chemotherapy (n=136). GGH rs11545078 was associated with a reduced incidence of grade ⩾3 toxicity within the first four cycles of therapy (odds ratio (OR) 0.25, P=0.018), as well as reduced grade ⩾3 haematological toxicity (OR 0.13, P=0.048). DHFR rs1650697 conferred an increased risk of grade ⩾3 toxicity (OR 2.14, P=0.034). Furthermore, FOLR3 rs61734430 was associated with an increased likelihood of disease progression at mid-treatment radiological evaluation (OR 4.05, P=0.023). Polymorphisms within SLC19A1 (rs3788189, rs1051298 and rs914232) were associated with overall survival. This study confirms previous pharmacogenetic associations and identifies novel markers of pemetrexed toxicity.

Pharmacogenetic variants in the DPYD, TYMS, CDA and MTHFR genes are clinically significant predictors of fluoropyrimidine toxicity.

BACKGROUND: Fluoropyrimidine drugs are extensively used for the treatment of solid cancers. However, adverse drug reactions are a major clinical problem, often necessitating treatment discontinuation. The aim of this study was to identify pharmacogenetic markers predicting fluoropyrimidine toxicity. METHODS: Toxicity in the first four cycles of 5-fluorouracil or capecitabine-based chemotherapy were recorded for a series of 430 patients. The association between demographic variables, DPYD, DPYS, TYMS, MTHFR, CDA genotypes, and toxicity were analysed using logistic regression models. RESULTS: Four DPYD sequence variants (c.1905+1G>A, c.2846A>T, c.1601G>A and c.1679T>G) were found in 6% of the cohort and were significantly associated with grade 3-4 toxicity (P<0.0001). The TYMS 3'-untranslated region del/del genotype substantially increased the risk of severe toxicity (P=0.0123, odds ratio (OR)=3.08, 95% confidence interval (CI): 1.38-6.87). For patients treated with capecitabine, a MTHFR c.1298CC homozygous variant genotype predicted hand-foot syndrome (P=4.1 × 10⁻6, OR=9.99, 95% CI: 3.84-27.8). The linked CDA c.-92A>G and CDA c.-451C>T variants predicted grade 2-4 diarrhoea (P=0.0055, OR=2.3, 95% CI: 1.3-4.2 and P=0.0082, OR=2.3, 95% CI: 1.3-4.2, respectively). CONCLUSION: We have identified a panel of clinically useful pharmacogenetic markers predicting toxicity to fluoropyrimidine therapy. Dose reduction should be considered in patients carrying these sequence variants.

Dietary intervention for oral allergy syndrome as a treatment in orofacial granulomatosis: a new approach?

BACKGROUND: Orofacial granulomatosis (OFG) is a chronic granulomatous condition of the mouth, face and lips. Recent work demonstrates a high rate of atopy and silver birch sensitisation from skin prick testing (SPT). Oral allergy syndrome (OAS) is an acute oro-pharyngeal IgE mediated reaction, triggered by foods that cross react with pollens, most commonly silver birch. The aim of this study was to determine if patients with OFG and positive SPT to common OAS associated pollens responded to avoidance of cross reactive foods. METHODS: Patients with OFG and positive SPT to silver birch, grass, mugwort, ragweed and latex were required to avoid cross reacting foods, for 6 weeks and, in those who responded, for a total of 12 weeks. All had standardized oral examinations and were given severity scores (SS) at each appointment. RESULTS: Twenty two of 47 (47%) patients had one or more positive SPT and 13/22 completed 6 weeks on the diet. No difference was seen in SS between weeks 0 (14.62 ± 11.16) and 6 (13.31 ± 10.33; P = 0.656). Six of 14 (43%) had significantly improved SS (week 0; 19.17 ± 12.95, week 6; 10.83 ± 4.99, P = 0.027). Five completed 12 weeks and no further improvement was seen (week 6; 11 ± 5.57, week 12; 10.4 ± 9.94; P = 0.068). Two patients required no further treatments. CONCLUSIONS: On an intention to treat basis, only 2/14 patients improved and required no further intervention. Whilst this diet cannot be recommended routinely, the improvement seen in some patients raises questions about the role of OAS in patients with OFG.

Development of a low phenolic acid diet for the management of orofacial granulomatosis.

BACKGROUND: Orofacial granulomatosis (OFG) is a rare disease of unknown cause. A cinnamon- and benzoate-free diet is successful in up to 72% of patients. Phenolic acids are among the chemical constituents restricted in this diet, which avoids some but not all of these structurally similar compounds. The present study aimed to: (i) develop a novel diet low in phenolic acids; (ii) implement this in a small clinical trial; and (iii) assess its nutritional adequacy. METHODS: A literature review identified 10 papers quantifying phenolic acids from which 91 10-mg phenolic acid exchanges were devised. A phenolic acid exclusion diet with precautionary micronutrient supplementation was designed and implemented in 10 patients. Phenolic acids were excluded for 6 weeks and were reintroduced at a rate of one exchange every second day for 6 weeks. Wilcoxon matched pairs tests analysed disease outcomes measured by an oral disease severity scoring tool at weeks 0, 6 and 12. Nutritional adequacy was assessed, excluding micronutrient supplementation, at weeks 0 and 6, and compared intakes with dietary reference values. RESULTS: The diet was nutritionally inadequate for a range of micronutrients. Seven of 10 patients responded. Mean [standard deviation (SD)] severity scores improved from week 0-6 [20.8 (9.39) and 10.1 (5.72); P = 0.009] and were maintained in five patients who completed the reintroduction [6.6 (3.13) and 7.2 (5.54); P = 0.713]. CONCLUSIONS: A low phenolic acid diet with micronutrient supplementation holds promise of a novel dietary treatment for OFG. Further work is required in larger studies to determine long-term outcomes.

Assessment of thiopurine methyltransferase enzyme activity is superior to genotype in predicting myelosuppression following azathioprine therapy in patients with inflammatory bowel disease.

BACKGROUND: Myelosuppression occurs in 2-7% of inflammatory bowel disease (IBD) patients treated with azathioprine, and can be associated with reduced activity of thiopurine methyltransferase (TPMT) in some patients. It has been proposed that pretreatment assessment of TPMT status reduces the incidence of toxicity and is cost-effective. AIMS: To determine if screening for TPMT status predicts side-effects to azathioprine in patients with IBD and to ascertain whether screening by TPMT enzyme activity or genotype is superior. METHODS: Sequential IBD patients were identified and azathioprine tolerance recorded. Blood was collected for measurement of TPMT activity and TPMT*3C, TPMT*3A and TPMT*2 genotypes. RESULTS: Of 130 patients, 25% stopped azathioprine because of toxicity. Four patients experienced severe myelosuppression (WCC < 2). Eleven of 17 patients with reduced TPMT activity were heterozygotes, including one patient with marked TPMT deficiency who experienced severe myelosuppression. There was no association between intermediate TPMT deficiency and any side-effect. CONCLUSIONS: Moderate reduction of TPMT activity in heterozygotes was not associated with toxicity, but very low TPMT activity caused severe myelosuppression in one patient. This would have been predicted by measuring TPMT activity but not by genotyping. Measurement of TPMT activity may therefore be superior to genotype in predicting severe myelosuppression.

Utilisation of anti-TNF levels in a UK tertiary IBD centre.

OBJECTIVE: To ascertain how anti-tumour necrosis factor (TNF) drug and anti-drug antibody levels testing is used in a 'real-world' setting to optimise inflammatory bowel disease (IBD) treatment. DESIGN: Retrospective cohort study of prospectively collected patient data. SETTING: Tertiary IBD centre in London, UK. PATIENTS: All patients at Guy's and St Thomas' Hospitals on anti-TNF who had levels measured between the start of testing in 2012 and October 2014. INTERVENTIONS: Anti-TNF drug and anti-drug antibody levels as part of routine monitoring. MAIN OUTCOME MEASURES: Indication for measuring levels and changes in management made as a result of the levels. RESULTS: 330 infliximab levels were carried out in 199 patients and 143 adalimumab levels were carried out in 103 patients. Levels were primarily done in those with evidence of loss of response; 37% of infliximab levels and 52% of adalimumab levels. Levels resulted in a change in management in 26% of patients in infliximab group and 25% of patients in adalimumab group; however, this was greater in those with loss of response, 62% and 61% respectively. Anti-drug antibodies were detected in 7% of patients. CONCLUSIONS: Our early experience has demonstrated that measuring anti-TNF drug and anti-drug antibody levels can be useful in the optimisation of IBD management. In an increasing number of patients, particularly those with evidence of loss of response, it allows early decisions to be made regarding changing therapy. It also offers the potential for significant cost-saving by preventing pointless dose escalation in the context of therapeutic levels or when high-level anti-drug antibodies are present.

Infliximab and adalimumab drug levels in Crohn's disease: contrasting associations with disease activity and influencing factors.

BACKGROUND: Discriminative drug level thresholds for disease activity endpoints in patients with Crohn's disease. have been consistently demonstrated with infliximab, but not adalimumab. AIMS: To identify threshold concentrations for infliximab and adalimumab in Crohn's disease according to different disease endpoints, and factors that influence drug levels. METHODS: We performed a cross-sectional service evaluation of patients receiving maintenance infliximab or adalimumab for Crohn's disease. Serum drug levels were at trough for infliximab and at any time point for adalimumab. Endpoints included Harvey-Bradshaw index, C-reactive protein and faecal calprotectin. 6-tioguanine nucleotide (TGN) concentrations were measured in patients treated with thiopurines. RESULTS: A total of 191 patients (96 infliximab, 95 adalimumab) were included. Differences in infliximab levels were observed for clinical (P=.081) and biochemical remission (P=.003) and faecal calprotectin normalisation (P<.0001) with corresponding thresholds identified on ROC analysis of 1.5, 3.4 and 5.7 µg/mL. Adalimumab levels were similar between active disease and remission regardless of the endpoint assessed. Modelling identified that higher infliximab dose, body mass index and colonic disease independently accounted for 31% of the variation in infliximab levels, and weekly dosing, albumin and weight accounted for 23% of variation in adalimumab levels. TGN levels did not correlate with drug levels. CONCLUSIONS: Infliximab drug levels are associated with the depth of response/remission in patients with Crohn's disease, but no such relationship was observed for adalimumab. More data are needed to explain the variation in drug levels.

Immunohistochemical analysis of ageing human B and T cell populations reveals an age-related decline of CD8 T cells in spleen but not gut-associated lymphoid tissue (GALT).

It is thought that senescence of the immune system is responsible, at least in part, for many health problems associated with ageing. Previous studies on changes in lymphocyte composition have used flow cytometry to study peripheral blood lymphocytes (PBL's), or cells isolated from rodent tissue, and have yielded conflicting results. We have used immunohistochemistry to determine whether the B and T cells in human tissue from spleen and gut are affected by age. Areas of germinal centre, mantle zone and marginal zone of B cell follicles were measured. In addition, CD4 and CD8 T cells in T cell areas and in B cell follicles were counted. We observed a striking age-related decrease in the proportion of CD8+ T cells in the T cell zones of the spleen. This decrease was not apparent in the T cell population that occupies splenic B cell areas, or in GALT. Further differences, in CD4+ cells, were seen between T cell populations in the T cell zones and those in B cell areas. These findings highlight differences between lymphocyte populations in different lymphoid tissues, and different compartments within each tissue, which may be of importance in future studies of the ageing immune system.

Analysis of feeding patterns in normal and vagotomized rabbits.

Spontaneous meal sizes, intermeal intervals, and 24 hr feeding rhythms were monitored in normal and 60 day recovered vagotomized rabbits fed solid laboratory chow. Mean sizes of meals and intermeal intervals, and the circadian distribution of food intake did not differ between the two groups, but vagotomy was associated with increased frequencies of both smaller and larger than average meals. Positive meal to postmeal interval correlations were evident in intact but not vagotomized animals, whereas vagotomized animals displayed a meal to premeal interval correlation in the light phase that was not present in normal rabbits.

Gastro-oesophageal reflux and duodenogastric reflux before and after eradication in Helicobacter pylori gastritis.

OBJECTIVE: Helicobacter pylori and duodenogastric reflux (DGR) are both associated with chronic gastritis, peptic ulcer and gastric cancer. The nature of their interrelationship remains unclear. H. pylori eradication has also been reported to result in new or worsening acid gastro-oesophageal reflux (GOR). The aim of this study was to investigate the relationship between GOR, DGR and H. pylori infection. METHOD: 25 patients with H. pylori gastritis underwent ambulatory 24-hour oesophageal and gastric pHmetry and gastric bilirubin monitoring before and 12 weeks after H. pylori eradication, confirmed by 14C urea breath testing (UBT). Ten healthy subjects served as a control group. RESULTS: There were no differences between patient and control groups for gastric alkaline exposure or gastric bilirubin exposure (P> 0.25 in all categories). Oesophageal acid reflux was higher in the study group (P< 0.02). No differences were detected in oesophageal acid reflux, gastric alkaline exposure, or gastric bilirubin exposure (P = 0.35, 0.18 and 0.11, respectively) before and after eradication. CONCLUSIONS: Acid GOR is not increased by H. pylori eradication. DGR in patients with H. pylori gastritis is similar to that in healthy, non-infected subjects. H. pylori eradication produces no change in GOR or DGR. In patients with chronic gastritis, H. pylori infection and DGR appear to be independent of each other.

Vagally mediated feeding responses to phloridzin infusion in the rabbit.

Rabbits were infused with the plant glycoside, phloridzin, which blocks absorption of glucose across a number of bodily tissues. Feeding was dramatically increased in the first 0.5 h following phloridzin infusion into either the duodenum or the hepatic-portal vein of intact rabbits. Food intake covaried inversely with glycemic levels after phloridzin infusion into the hepatic-portal vein, but rabbits did not show a systematic relationship between blood glucose levels and food intake following duodenal infusion of phloridzin. When administered into the general circulation via the jugular vein, phloridzin did not elicit feeding. Finally, vagotomized rabbits did not show the hyperphagic response to phloridzin that was observed in intact rabbits. It was concluded that the feeding response to phloridzin is vagally mediated and appears to be induced by glucose transport inhibition at some peripheral site.

Mechanism of allopurinol induced TPMT inhibition.

Up to 1/5 of patients with wildtype thiopurine-S-methyltransferase (TPMT) activity prescribed azathioprine (AZA) or mercaptopurine (MP) demonstrate a skewed drug metabolism in which MP is preferentially methylated to yield methylmercaptopurine (MeMP). This is known as thiopurine hypermethylation and is associated with drug toxicity and treatment non-response. Co-prescription of allopurinol with low dose AZA/MP (25-33%) circumvents this phenotype and leads to a dramatic reduction in methylated metabolites; however, the biochemical mechanism remains unclear. Using intact and lysate red cell models we propose a novel pathway of allopurinol mediated TPMT inhibition, through the production of thioxanthine (TX, 2-hydroxymercaptopurine). In red blood cells pre-incubated with 250 µM MP for 2h prior to the addition of 250 µM TX or an equivalent volume of Earle's balanced salt solution, there was a significant reduction in the concentration of MeMP detected at 4h and 6h in cells exposed to TX (4 h, 1.68, p=0.0005, t-test). TX acts as a direct TPMT inhibitor with an apparent Ki of 0.329 mM. In addition we have confirmed that the mechanism is relevant to in vivo metabolism by demonstrating raised urinary TX levels in patients receiving combination therapy. We conclude that the formation of TX in patients receiving combination therapy with AZA/MP and allopurinol, likely explains the significant reduction of methylated metabolites due to direct TPMT inhibition.

Distinct microbial populations exist in the mucosa-associated microbiota of sub-groups of irritable bowel syndrome.

BACKGROUND: There is increasing evidence to support a role for the gastrointestinal microbiota in the etiology of irritable bowel syndrome (IBS). Given the evidence of an inflammatory component to IBS, the mucosa-associated microbiota potentially play a key role in its pathogenesis. The objectives were to compare the mucosa-associated microbiota between patients with diarrhea predominant IBS (IBS-D), constipation predominant IBS (IBS-C) and controls using fluorescent in situ hybridization and to correlate specific bacteria groups with individual IBS symptoms. METHODS: Forty-seven patients with IBS (27 IBS-D and 20 IBS-C) and 26 healthy controls were recruited to the study. Snap-frozen rectal biopsies were taken at colonoscopy and bacterial quantification performed by hybridizing frozen sections with bacterial-group specific oligonucleotide probes. KEY RESULTS: Patients with IBS had significantly greater numbers of total mucosa-associated bacteria per mm of rectal epithelium than controls [median 218 (IQR - 209) vs 128 (121) P = 0.007], and this was chiefly comprised of bacteroides IBS [69 (67) vs 14 (41) P = 0.001] and Eubacterium rectale-Clostridium coccoides [52 (58) vs 25 (35) P = 0.03]. Analysis of IBS sub-groups demonstrated that bifidobacteria were lower in the IBS-D group than in the IBS-C group and controls [24 (32) vs 54 (88) vs 32 (35) P = 0.011]. Finally, amongst patients with IBS, the maximum number of stools per day negatively correlated with the number of mucosa-associated bifidobacteria (P < 0.001) and lactobacilli (P = 0.002). CONCLUSIONS & INFERENCES: The mucosa-associated microbiota in patients with IBS is significantly different from healthy controls with increases in bacteroides and clostridia and a reduction in bifidobacteria in patients with IBS-D.

Review article: cinnamon- and benzoate-free diet as a primary treatment for orofacial granulomatosis.

BACKGROUND: Orofacial granulomatosis is a rare chronic granulomatous inflammatory disease of the lips, face and mouth. The aetiology remains unclear but may involve an allergic component. Improvements have been reported with cinnamon- and benzoate-free diets. AIMS: To explore the prevalence of compound and food sensitivity and examine the dietary treatments used in orofacial granulomatosis. METHODS: A comprehensive literature search was carried out and relevant studies from January 1933 to January 2010 were identified using the electronic database search engines; AGRIS 1991-2008, AMED 1985-2008, British Nursing and Index archive 1985-2008, EMBASE 1980-2008, evidence based medicine review databases (e.g. Cochrane DSR), International Pharmaceutical and Medline 1950-2008. RESULTS: Common sensitivities identified, predominantly through patch testing, were to benzoic acid (36%) food additives (33%), perfumes and flavourings (28%), cinnamaldehyde (27%), cinnamon (17%), benzoates (17%) and chocolate (11%). The cinnamon- and benzoate-free diet has been shown to provide benefit in 54-78% of patients with 23% requiring no adjunctive therapies. A negative or positive patch test result to cinnamaldehyde, and benzoates did not predict dietary outcome. The most concentrated source of benzoate exposure is from food preservatives. Use of liquid enteral formulas can offer a further dietary therapy, particularly in children with orofacial granulomatosis. CONCLUSION: Management of orofacial granulomatosis is challenging but cinnamon- and benzoate-free diets appear to have a definite role to play.

Management of inflammatory bowel disease in pregnancy.

Inflammatory bowel disease (IBD) affects body image, relationships, family planning, fertility and pregnancy outcomes. However, the common misconception that IBD is a contraindication, or serious concern, in pregnancy is essentially a myth. Most patients with IBD can expect to have uneventful pregnancies. We present an overview of the management of IBD during pregnancy, including management in those planning pregnancy, the suitability of relevant medication during pregnancy and breast feeding, investigation and monitoring of IBD during pregnancy, surgical management and considerations relating to delivery. While there are some definite alterations required in the management of IBD during pregnancy, management is essentially unchanged. With close attention to aspects such as nutrition and smoking cessation, and optimal disease control in the run-up to and during pregnancy, we have an opportunity to help our patients with IBD achieve good pregnancy outcomes.

Treating irritable bowel syndrome with probiotics: the evidence.

Irritable bowel syndrome (IBS) is a disorder of chronic abdominal pain, altered bowel habit and abdominal distension. It is the commonest cause of referral to gastroenterologists in the developed world and yet current therapeutic strategies are often unsatisfactory. There is now increasing evidence linking alterations in the gastrointestinal (GI) microbiota and IBS. Changes in faecal and mucosa-associated microbiota, post-infectious IBS, a link with small intestinal bacterial overgrowth and an up-regulation of the GI mucosal immune system all suggest a role for the GI microbiota in the pathogenesis of IBS. Given this evidence, therapeutic alteration of the GI microbiota by probiotic bacteria could be beneficial. The present paper establishes an aetiological framework for the use of probiotics in IBS and comprehensively reviews randomised placebo-controlled trials of probiotics in IBS using multiple electronic databases. It highlights safety concerns over the use of probiotics and attempts to establish guidelines for their use in IBS in both primary and secondary care.