Susan J. Connor

Multidonor intensive faecal microbiota transplantation for active ulcerative colitis: a randomised placebo-controlled trial

BACKGROUND The intestinal microbiota is implicated in the pathogenesis of ulcerative colitis. Faecal microbiota transplantation is a novel form of therapeutic microbial manipulation, but its efficacy in ulcerative colitis is uncertain. We aimed to establish the efficacy of intensive-dosing, multidonor, faecal microbiota transplantation in active ulcerative colitis. METHODS We conducted a multicentre, double-blind, randomised, placebo-controlled trial at three hospitals in Australia. We randomly allocated patients with active ulcerative colitis (Mayo score 4-10) in a 1:1 ratio, using a pre-established randomisation list, to either faecal microbiota transplantation or placebo colonoscopic infusion, followed by enemas 5 days per week for 8 weeks. Patients, treating clinicians, and other study staff were unaware of the assigned treatment. Faecal microbiota transplantation enemas were each derived from between three and seven unrelated donors. The primary outcome was steroid-free clinical remission with endoscopic remission or response (Mayo score ≤2, all subscores ≤1, and ≥1 point reduction in endoscopy subscore) at week 8. Analysis was by modified intention-to-treat and included all patients receiving one study dose. We performed 16S rRNA stool analysis to assess associated microbial changes. This trial is registered with ClinicalTrials.gov, number NCT01896635. The trial has ended; this report presents the final analysis. FINDINGS From November, 2013, to May, 2015, 85 patients were enrolled to our trial, of whom 42 were randomly assigned faecal microbiota transplantation and 43 were allocated placebo. One patient assigned faecal microbiota transplantation and three allocated placebo did not receive study treatment and were excluded from the analysis. The primary outcome was achieved in 11 (27%) of 41 patients allocated faecal microbiota transplantation versus three (8%) of 40 who were assigned placebo (risk ratio 3·6, 95% CI 1·1-11·9; p=0·021). Adverse events were reported by 32 (78%) of 41 patients allocated faecal microbiota transplantation and 33 (83%) of 40 who were assigned placebo; most were self-limiting gastrointestinal complaints, with no significant difference in number or type of adverse events between treatment groups. Serious adverse events occurred in two patients assigned faecal microbiota transplantation and in one allocated placebo. Microbial diversity increased with and persisted after faecal microbiota transplantation. Several bacterial taxa were associated with clinical outcome; in particular, the presence of Fusobacterium spp was associated with lack of remission. INTERPRETATION Intensive-dosing, multidonor, faecal microbiota transplantation induces clinical remission and endoscopic improvement in active ulcerative colitis and is associated with distinct microbial changes that relate to outcome. Faecal microbiota transplantation is, thus, a promising new therapeutic option for ulcerative colitis. Future work should focus on precisely defining the optimum treatment intensity and the role of donor-recipient matching based on microbial profiles. FUNDING Broad Medical Research Program, Gastroenterological Society of Australia, Mount Sinai (New York) SUCCESS fund, University of New South Wales.

HLA-DQA1-HLA-DRB1 variants confer susceptibility to pancreatitis induced by thiopurine immunosuppressants

Pancreatitis occurs in approximately 4% of patients treated with the thiopurines azathioprine or mercaptopurine. Its development is unpredictable and almost always leads to drug withdrawal. We identified patients with inflammatory bowel disease (IBD) who had developed pancreatitis within 3 months of starting these drugs from 168 sites around the world. After detailed case adjudication, we performed a genome-wide association study on 172 cases and 2,035 controls with IBD. We identified strong evidence of association within the class II HLA region, with the most significant association identified at rs2647087 (odds ratio 2.59, 95% confidence interval 2.07–3.26, P = 2 × 10−16). We replicated these findings in an independent set of 78 cases and 472 controls with IBD matched for drug exposure. Fine mapping of the HLA region identified association with the HLA-DQA1*02:01–HLA-DRB1*07:01 haplotype. Patients heterozygous at rs2647087 have a 9% risk of developing pancreatitis after administration of a thiopurine, whereas homozygotes have a 17% risk.

CCR2 expressing CD4+ T lymphocytes are preferentially recruited to the ileum in Crohn’s disease

Background and aims: Chemokine receptors are key determinants of leucocyte trafficking. While the chemokine receptor CCR9 and its chemokine ligand CCL25 (TECK) mediate lymphocyte homing to the healthy small intestine, the chemokine receptors important for recruitment during intestinal inflammation are undefined. Animal studies have suggested potential roles for CCR2 and CCR5 in inflammatory bowel disease (IBD). The aim of this study was to understand the role of CCR2 in human IBD. Methods: Resections of ileum or colon were obtained from patients undergoing surgery for small bowel Crohn’s disease (SBCD; n = 10), Crohn’s colitis (n = 5), ulcerative colitis (n = 6), and non-IBD related conditions (control ileum n = 11; control colon n = 11). Expression of CCR2 by lamina propria lymphocytes (LPLs) was determined by both flow cytometry and immunohistochemistry. As a functional correlate, chemotaxis assays using the CCR2 ligand, CCL2 (MCP-1), were performed. Expression of CCR2 by peripheral blood lymphocytes was determined by flow cytometry. Results: There were greater than 30-fold more CCR2+ LPLs in SBCD than in control ileum (29.3% (19.9–55.1) v 0.9% (0.4–11.5); p = 0.0007). Specifically, CCR2+CD4+ LPLs were increased (p = 0.002) whereas CCR2+CD8+ LPLs were not. Increased expression included both memory (CD45RO+; p = 0.005) and naïve (CD45RO−; p = 0.01) CCR2+ populations. The increase in CCR2+ LPLs in SBCD was confirmed by both immunohistochemistry (p = 0.0002) and enhanced chemotactic responses to CCL2. CCR2 expression was not increased in the peripheral blood of patients with SBCD, suggesting ongoing recruitment of the CCR2+ population to the ileum. In contrast with SBCD, there was no significant increase in CCR2+ LPLs in Crohn’s colitis or ulcerative colitis samples. Conclusions: The chemokine receptor CCR2 appears to be an important contributor to accumulation of CD4+ T lymphocytes in the ileum in small bowel Crohn’s disease. Blockade of CCR2 may provide a novel therapeutic alternative.

Improved survival in young women with colorectal cancer.

BACKGROUND:Studies have reported the effect of gender in the context of assessing predictors of survival from colorectal cancer (CRC); however, few have specifically addressed the impact of gender on the clinical and pathological outcomes of CRC. Appreciation of gender disparities may assist in the implementation of measures to address these differences, and improve the overall outcomes of patients with CRC.METHODS:The South Western Sydney Colorectal Tumour Group registry, which encompasses a population in excess of 800,000, prospectively collects data on new patients with CRC. Data from 1997 to 2004 were collected, including demography, site, grade, histopathology, stage, treatment, and survival.RESULTS:In total, 2,050 consecutive patients (44% women) with CRC were analyzed. Compared to men, women were older (median 69 yr, range 27–95 yr vs 67, range 22–92 yr, P= 0.001), had more emergency surgery for CRC-related complications (18.8% vs 15.1%, P= 0.03), had more proximal cancers (42.2% vs 31.5%, P < 0.001), had more poorly differentiated cancers (16.9% vs 12.9%, P= 0.01), and had fewer radiotherapy treatments for Dukes B and C rectal cancers (36.4% vs 48.1%, P= 0.02). Young women (aged 50 yr and below) had significantly better overall survival compared to young men; in this group, female gender predicted improved overall survival independent of age, emergency surgery, site, grade, and stage (hazard ratio [HR] 0.46, 95% confidence interval [CI] 0.25–0.86, P= 0.01). Similarly, young women had significantly better cancer-specific survival (HR 0.46, 95% CI 0.25–0.85, P= 0.01). However, older women (aged over 50 yr) had worse survival independent of age, emergency surgery, site, grade, and stage (HR 1.38, 95% CI 1.14–1.68, P= 0.001). There were no gender differences in screening, histopathology, stage, or utilization of chemotherapy.CONCLUSIONS:This study demonstrated an opposing effect of gender on overall and cancer-specific survival at either side of the age of 50 yr. The protective effect of estrogen on CRC may be an important factor. Women had a greater proportion of emergency surgery, which was related to the predominance of proximal cancers in this gender. Women also had more proximal cancers, thereby limiting flexible sigmoidoscopy as a screening test.

Cutting Edge: Vasoactive Intestinal Peptide Acts as a Potent Suppressor of Inflammation In Vivo by Trans-Deactivating Chemokine Receptors

Chemokines mediate trafficking of leukocytes to sites of inflammation and immune responses through activation of G protein-coupled receptors, which thereby provide appealing targets for novel anti-inflammatory agents. Vasoactive intestinal peptide (VIP) is an immunosuppressive neurotransmitter. We show that VIP inhibited the function of chemokine receptors on monocytes and CD4+ T lymphocytes, with impaired chemotaxis and calcium flux in response to the cognate chemokine ligands CXCL12, CCL3, CCL4, and CCL5. This was mediated by VIP receptor type 1 and was not caused by chemokine receptor internalization. However, VIP caused dose-dependent phosphorylation of the chemokine receptor CCR5. This trans-deactivation process was studied in a murine model of delayed-type hypersensitivity: continuous infusion of VIP resulted in significant abrogation of monocyte and lymphocyte infiltration. Circulating mononuclear cells from VIP-infused mice were unable to respond to chemokines. VIP may provide a novel approach to treatment of inflammatory diseases through inhibition of chemokine-dependent leukocyte recruitment.

Donor Recruitment for Fecal Microbiota Transplantation.

Background:Increasing demand for fecal microbiota transplantation (FMT) has created a need for stool banks sourced from long-term healthy donors. Here, we describe our experience in recruiting and screening fecal donors. Methods:Mailbox, newspaper, and online advertisements were used. Potential donors were required to satisfy a prescreen telephone conversation, pass blood and stool investigations, then undertake a screening interview including medical history, physical examination, and evaluation of donor selection criteria. Results:One hundred sixteen potential donors were prescreened of whom 74 failed—47 declined based on study donation requirements (primarily related to frequency and duration of donations), 13 had medical comorbidities, 6 variant Creutzfeldt–Jakob disease risk factors, 8 for other reasons. Thirty-eight completed stool and blood testing—1 failed blood testing (indeterminate hepatitis C serology), whereas 15 failed stool investigations (5 Dientamoeba fragilis, 5 Blastocystis hominis, 1 B. hominis and D. fragilis, 1 Giardia intestinalis plus D. fragilis, 1 Norovirus plus Clostridium difficile toxin positive, and 2 leucocytes or erythrocytes on stool microscopy). Of the 18 potential donors proceeding to screening interview, 6 were excluded (3 body mass index >30, 1 illicit drug use, 1 uncontrolled anxiety and concerns regarding compliance, 1 irregular bowel movements after new medication commencement). In total, only 12 of 116 (10%) potential donors were enrolled as study donors. Conclusions:Recruitment of fecal donors for FMT is challenging with only a small percentage ultimately serving as donors. Many were unable or unwilling to meet the donor commitment requirements. A surprisingly large proportion of healthy asymptomatic donors failed stool testing, primarily due to gastrointestinal parasites.

Vasoactive intestinal peptide impairs leucocyte migration but fails to modify experimental murine colitis

Inflammatory bowel diseases are chronic inflammatory disorders of the gastrointestinal tract. Vasoactive intestinal peptide (VIP) is a neuropeptide with known anti‐inflammatory activity. We have demonstrated previously that administration of VIP inhibits leucocyte migration in a murine model of delayed‐type hypersensitivity, and anti‐inflammatory efficacy is supported by other studies. The aim of this study was to investigate the VIP effects in a murine model of intestinal inflammation. Colitis was induced in BALB/c mice by a 2·5 mg enema of 2,4,6‐trinitrobenzenesulphonic acid (TNBS) and the mice were killed on day 7. Mice were administered either a 3‐day (therapeutic) or 7‐day (prophylactic) constant infusion of VIP by subcutaneously implanted mini‐osmotic pumps, or intraperitoneal (i.p.) injection of VIP on alternate days over 7 days. Clinical disease scores, weight changes, histopathology of colon tissues, plasma VIP levels, cytokine levels and chemotaxis of peripheral blood mononuclear cells were evaluated. After administration of TNBS, mice quickly developed severe colitis accompanied by dramatic body weight loss (20% by day 6) and high mortality (30%). Prophylactic treatment using high‐dose VIP abrogated leucocyte chemotaxis; however, it failed to ameliorate the weight loss and mortality. Moreover, VIP delivered either by constant infusion or i.p. failed to modify the clinical, histological or cytokine markers of disease. Our studies show that, despite an ability to inhibit chemokine‐induced chemotaxis of mononuclear cells, VIP was unable to modulate TNBS‐induced colitis. This contrasts with the efficacy of VIP in models of mild inflammatory disease and suggests that VIP is unlikely to provide a useful model for novel anti‐IBD therapy.

Inflammatory Bowel Disease Environmental Risk Factors: A Population-Based Case–Control Study of Middle Eastern Migration to Australia

BACKGROUND & AIMS The incidences of the inflammatory bowel diseases (IBDs) Crohns disease (CD) and ulcerative colitis (UC) are increasing, indicating gene-environment interactions. Migrants from low-IBD-prevalence countries to a high-prevalence country may help identify the relative contribution of environmental risk factors compared with native Caucasians. METHODS This prospective case-control study evaluated IBD environmental risk factors of Middle Eastern migrants (MEM) in Australia compared with matched Caucasian IBD subjects, MEM controls, Caucasian controls, and controls in the Middle East using adjusted odds ratios (aOR). RESULTS A total of 795 subjects were recruited: 154 MEM cases (75 CD; 79 UC), 153 MEM controls, 162 Caucasian cases (85 CD; 77 UC), 173 Caucasian controls, and 153 controls in Lebanon. Smoking increased CD risk in MEM and Caucasians and reduced UC risk in Caucasians (aOR, 0.77; 95% CI, 0.41-0.98) but not MEM (aOR, 1.45; 95% CI, 0.80-2.62). Antibiotic use reduced the risk of MEM CD (aOR, 0.27; 95% CI, 0.11-0.67) and UC (aOR, 0.38; 95% CI, 0.18-0.80), but increased the risk in Caucasians (CD: aOR, 5.24; 95% CI, 2.13-12.90; and UC: aOR, 6.82; 95% CI, 2.67-17.38). Most hygiene markers (rural dwelling, pet ownership, pet feeding, and farm animal contact) reduced CD and UC risk in MEM (P < .05). In contrast, in Caucasians these hygiene markers lacked significance. Other significant risk factors include IBD family history, appendectomy, tonsillectomy, and breastfeeding. CONCLUSIONS Differential IBD environmental risk factors exist between migrants and native Caucasians, indicating a dynamic interplay between environmental factors and IBD risk for immigrants that is distinct to those factors most relevant in native Caucasians.

The impact of short-term ranitidine use on the precision of the 13C-urea breath test in subjects infected with Helicobacter pylori.

BACKGROUND The 13C-urea breath test (13C-UBT) is a very accurate method of Helicobacter pylori diagnosis with a false-negative rate of 1-3%. However, the accuracy of the 13C-UBT is affected by potent acid inhibition with proton-pump inhibitors, which may suppress H. pylori and cause false-negative results. It is not known whether this occurs with less potent acid inhibition by H2-antagonists and any effect may be important clinically. OBJECTIVE To determine the kinetics of 13CO2 excretion in H. pylori infected subjects during and after short-term ranitidine use. METHODS Volunteers underwent a baseline 13C-UBT (positive: delta13CO2 > or = 5.0; negative: < or = 3.5; indeterminate: > 3.5 to < 5.0). Infected subjects took ranitidine 300 mg each evening for up to 28 days. 13C-UBTs were performed at weekly intervals and then every other day after ranitidine was ceased. If the 13C-UBT remained positive after 14 days, ranitidine was continued for a further 14 days. RESULTS Thirty-one subjects were studied (mean age 40.4 +/- 2.1 years; 23 female/8 male; mean baseline delta13CO2 27.3 +/- 2.5). In 28 subjects the 13C-UBT remained positive during ranitidine use. The mean delta13CO2 rose to 124% (P< 0.06) and 121% (P < 0.05) of baseline at 14 and 28 days respectively. In two subjects, the delta13CO2 became indeterminate at day 7 (delta13CO2 4.3 and 3.8). In one of these, return to a positive value (delta13CO2 13.6; 103% of baseline) occurred while still on ranitidine. The other subject became positive again by day 3 off ranitidine (17.8; 119% of baseline). One subject had a transiently negative test after 21 days and this became positive again while still taking ranitidine. CONCLUSIONS Ranitidine has a minimal effect on the 13C-UBT. The rate of indeterminate or false-negative tests is no greater than in patients on no anti-secretory medication.

Implementing guidelines on the prevention of opportunistic infections in inflammatory bowel disease

INTRODUCTION Opportunistic infections are a key safety concern in the management of patients with inflammatory bowel disease (IBD). Despite the existence of international guidelines, many gastroenterologists have not adopted routine screening and vaccination. The aim of this study was to modify clinical behaviour by use of a simple screening tool. METHODS A screening and vaccination proforma for hepatitis B, varicella, Influenza, Pneumococcus, human papillomavirus, tuberculosis, hepatitis C and HIV was provided to each participating gastroenterologist. Gastroenterologists were surveyed for awareness of vaccine recommendations and current practice prior to and following the introduction of the proforma. Rates of immunity and the proportion of patients receiving the recommended screening and vaccinations were documented. RESULTS 30 gastroenterologists at 8 different IBD centres took part in the assessment. A total of 919 patients were included (55% female, 65% Crohns, 33% ulcerative colitis, 2% indeterminate IBD). Introduction of the proforma increased self-reported gastroenterologist screening from 47% to 97% pre- and post-intervention respectively, p<0.001. After the proforma was applied, vaccination against hepatitis B, varicella, Influenza, and Pneumococcus was recommended in 67%, 2.5%, 75% and 69% of the patients respectively. Of these, 42%, 39%, 66% and 49% patients followed the recommendations and were vaccinated. Cervical smears were recommended in 31%, with 62% of these obtaining the recommended cervical smear. CONCLUSIONS Implementation of a screening and vaccination proforma significantly changed gastroenterologist self-reported behaviour. Patient compliance with these recommendations was not optimal and suggests the need for further patient education, in addition to other forms of support.