Clémentine Perrier

Etrolizumab as induction therapy for ulcerative colitis: a randomised, controlled, phase 2 trial

BACKGROUND Etrolizumab is a humanised monoclonal antibody that selectively binds the β7 subunit of the heterodimeric integrins α4β7 and αEβ7. We aimed to assess etrolizumab in patients with moderately-to-severely active ulcerative colitis. METHODS In this double-blind, placebo-controlled, randomised, phase 2 study, patients with moderately-to-severely active ulcerative colitis who had not responded to conventional therapy were recruited from 40 referral centres in 11 countries. Eligible patients (aged 18-75 years; Mayo Clinic Score [MCS] of 5 of higher [or ≥6 in USA]; and disease extending 25 cm or more from anal verge) were randomised (1:1:1) to one of two dose levels of subcutaneous etrolizumab (100 mg at weeks 0, 4, and 8, with placebo at week 2; or 420 mg loading dose [LD] at week 0 followed by 300 mg at weeks 2, 4, and 8), or matching placebo. The primary endpoint was clinical remission at week 10, defined as MCS of 2 or less (with no individual subscore of >1), analysed in the modified intention-to-treat population (mITT; all randomly assigned patients who had received at least one dose of study drug, had at least one post-baseline disease-activity assessment, and had a centrally read screening endoscopic subscore of ≥2). This study is registered with ClinicalTrials.gov, number NCT01336465. FINDINGS Between Sept 2, 2011, and July 11, 2012, 124 patients were randomly assigned, of whom five had a endoscopic subscore of 0 or 1 and were excluded from the mITT population, leaving 39 patients in the etrolizumab 100 mg group, 39 in the etrolizumab 300 mg plus LD group, and 41 in the placebo group for the primary analyses. No patients in the placebo group had clinical remission at week 10, compared with eight (21% [95% CI 7-36]) patients in the etrolizumab 100 mg group (p=0·0040) and four (10% [0·2-24]) patients in the 300 mg plus LD group (p=0·048). Adverse events occurred in 25 (61%) of 41 patients in the etrolizumab 100 mg group (five [12%] of which were regarded as serious), 19 (48%) of 40 patients in the etrolizumab 300 mg plus LD group (two [5%] serious), and 31 (72%) of 43 patients in the placebo group (five [12%] serious). INTERPRETATION Etrolizumab was more likely to lead to clinical remission at week 10 than was placebo. Therefore, blockade of both α4β7 and αEβ7 might provide a unique therapeutic approach for the treatment of ulcerative colitis, and phase 3 studies have been planned. FUNDING Genentech.

Butyricicoccus pullicaecorum in inflammatory bowel disease

Objective Many species within the phylum Firmicutes are thought to exert anti-inflammatory effects. We quantified bacteria belonging to the genus Butyricicoccus in stools of patients with ulcerative colitis (UC) and Crohns disease (CD). We evaluated the effect of Butyricicoccus pullicaecorum in a rat colitis model and analysed the ability to prevent cytokine-induced increases in epithelial permeability. Design A genus-specific quantitative PCR was used for quantification of Butyricicoccus in stools from patients with UC or CD and healthy subjects. The effect of B pullicaecorum on trinitrobenzenesulfonic (TNBS)-induced colitis was assessed and the effect of B pullicaecorum culture supernatant on epithelial barrier function was investigated in vitro. Results The average number of Butyricicoccus in stools from patients with UC and CD in active (UC: 8.61 log10/g stool; CD: 6.58 log10/g stool) and remission phase (UC: 8.69 log10/g stool; CD: 8.38 log10/g stool) was significantly lower compared with healthy subjects (9.32 log10/g stool) and correlated with disease activity in CD. Oral administration of B pullicaecorum resulted in a significant protective effect based on macroscopic and histological criteria and decreased intestinal myeloperoxidase (MPO), tumour necrosis factor α (TNFα) and interleukin (IL)-12 levels. Supernatant of B pullicaecorum prevented the loss of transepithelial resistance (TER) and the increase in IL-8 secretion induced by TNFα and interferon γ (IFN gamma) in a Caco-2 cell model. Conclusions Patients with inflammatory bowel disease have lower numbers of Butyricicoccus bacteria in their stools. Administration of B pullicaecorum attenuates TNBS-induced colitis in rats and supernatant of B pullicaecorum cultures strengthens the epithelial barrier function by increasing the TER.

Predictive value of epithelial gene expression profiles for response to infliximab in Crohn's disease.

Background: Infliximab (IFX) has become the mainstay of therapy of refractory Crohns disease (CD). However, a subset of patients shows incomplete or no response to this agent. In this study we investigated whether we could identify a mucosal gene panel to predict (non)response to IFX in CD. Methods: Mucosal biopsies were obtained during endoscopy from 37 patients with active CD (19 Crohns colitis [CDc] and 18 Crohns ileitis [CDi]) before and after first IFX treatment. Response was defined based on endoscopic and histologic findings. Total RNA was analyzed with Affymetrix Human Genome U133 Plus 2.0 Arrays. Quantitative real‐time reverse‐transcription polymerase chain reaction (RT‐PCR) was used to confirm microarray data. Results: At baseline, significant gene expression differences were found between CDc and CDi. For predicting response in CDc, comparative analysis of CDc pretreatment expression profiles identified 697 significant probe sets between CDc responders (n = 12) and CDc nonresponders (n = 7). Class prediction analysis of CDc top 20 and top 5 significant genes allowed complete separation between CDc responders and CDc nonresponders. The CDc top 5 genes were TNFAIP6, S100A8, IL11, G0S2, and S100A9. Only one patient with CDi completely healed the ileal mucosa. Even using less stringent response criteria, we could not identify a predictive gene panel for IFX responsiveness in CDi. Conclusions: This study identified a 100% accurate predictive gene signature for (non)response to IFX in CDc, whereas no such a predictive gene set could be identified for CDi. Inflamm Bowel Dis 2010

Cytokine blockade in inflammatory bowel diseases

Inflammatory bowel diseases (IBDs) are chronic disabling diseases with significant morbidity. A deregulated immune response towards the intestinal microbiota is thought to play an important role in the pathogenesis of IBD, and thus biological therapies targeting key molecules such as cytokines have been designed. Several anti-TNF-α agents are currently being used to treat Crohns disease and ulcerative colitis. Although these molecules dramatically improved the treatment of patients, side effects and the development of antidrug antibodies limits their application. There is thus an urgent need for alternative approaches to decrease inflammation and limit immunogenicity. Small neutralizing molecules, active immunization, gene silencing, selective transcription inhibitors and delivery of agents through the oral route are some of the currently developed strategies to meet these needs. In parallel, neutralizing antibodies targeting other pathways of the immune system have been developed and tested. Antibodies targeting IL-12/IL-23 pathways, and proinflammatory cytokines such as IFN-γ, IL-17A, IL-2 and IL-6 often showed an initial promising result, but for none of these agents efficacy has unequivocally been established. Administration of the regulatory cytokines IL-10 and IL-11 also failed to induce reproducible clinical effects. This article focuses on the anti-TNF therapies and the current challenges with monoclonal antibody therapies, discusses the innovative strategies targeting cytokine pathways to decrease inflammation in the bowel, and summarizes the recently developed agents neutralizing proinflammatory cytokines.

Neutralization of membrane TNF, but not soluble TNF, is crucial for the treatment of experimental colitis.

Background:Agents neutralizing membrane tumor necrosis factor (mTNF) and soluble TNF (sTNF) are widely used for the treatment of inflammatory bowel disease (IBD). Neutralization of mTNF, however, is associated with increased susceptibility to infectious diseases. The aim of this study was to determine whether neutralization of sTNF exclusively, by the use of a dominant negative mutant of TNF (XENP1595), could reduce the severity of colitis in mice. Methods:Colitis was induced in immunodeficient mice by transfer of CD45RBhi CD25− T-cells. Once the disease had developed, mice were treated twice a week with XENP1595, phosphate-buffered saline (PBS), anti-TNF monoclonal antibody (mAb), or isotype control. The anti-TNF mAb blocks both mTNF and sTNF. Weights, disease activity index, macroscopic inflammation of the colon, and histological sections were evaluated. T-cell populations from the colon were analyzed by flow cytometry. Results:Treatment of mice with XENP1595 did not change the course of the disease, whereas mice treated with anti-TNF mAb recovered weight soon after the first treatment dose. Inflammation in the colon was reduced in mice treated with anti-TNF mAb compared to isotype control-treated animals. Mice treated with XENP1595 had a similar degree of inflammation in the colon as PBS-treated animals. The number of effector and regulatory T-cells in the colon remained unaffected by all treatments. Conclusions:Neutralization of sTNF exclusively was unable to induce remission in T-cell-mediated colitis, suggesting that neutralization of mTNF is crucial for the treatment of IBD.

Unique gene expression and MR T2 relaxometry patterns define chronic murine dextran sodium sulphate colitis as a model for connective tissue changes in human Crohn's disease.

Introduction Chronically relapsing inflammation, tissue remodeling and fibrosis are hallmarks of inflammatory bowel diseases. The aim of this study was to investigate changes in connective tissue in a chronic murine model resulting from repeated cycles of dextran sodium sulphate (DSS) ingestion, to mimic the relapsing nature of the human disease. Materials and Methods C57BL/6 mice were exposed to DSS in drinking water for 1 week, followed by a recovery phase of 2 weeks. This cycle of exposure was repeated for up to 3 times (9 weeks in total). Colonic inflammation, fibrosis, extracellular matrix proteins and colonic gene expression were studied. In vivo MRI T 2 relaxometry was studied as a potential non-invasive imaging tool to evaluate bowel wall inflammation and fibrosis. Results Repeated cycles of DSS resulted in a relapsing and remitting disease course, which induced a chronic segmental, transmural colitis after 2 and 3 cycles of DSS with clear induction of fibrosis and remodeling of the muscular layer. Tenascin expression mirrored its expression in Crohn’s colitis. Microarray data identified a gene expression profile different in chronic colitis from that in acute colitis. Additional recovery was associated with upregulation of unique genes, in particular keratins, pointing to activation of molecular pathways for healing and repair. In vivo MRI T2 relaxometry of the colon showed a clear shift towards higher T2 values in the acute stage and a gradual regression of T2 values with increasing cycles of DSS. Conclusions Repeated cycles of DSS exposure induce fibrosis and connective tissue changes with typical features, as occurring in Crohn’s disease. Colonic gene expression analysis revealed unique expression profiles in chronic colitis compared to acute colitis and after additional recovery, pointing to potential new targets to intervene with the induction of fibrosis. In vivo T2 relaxometry is a promising non-invasive assessment of inflammation and fibrosis.

Effects of T cell-induced colonic inflammation on epithelial barrier function†

Background: Epithelial barrier disturbance is thought to contribute to the pathogenesis of inflammatory bowel diseases; however, it remains unclear whether it is a primary defect participating to the onset of inflammation or only a consequence of sustained inflammation. Methods: A time course study of epithelial barrier functions and immune mediators was performed in the CD4+CD45RBhi T cell transfer model of colitis using Ussing chambers. Results: In nonreconstituted severe combined immunodeficiency (SCID) mice, no epithelial dysfunction was observed. However, after transfer of CD4+CD45RBhi T cells or total CD4+ T cells, colon of SCID mice displayed a decreased epithelial resistance, even before overt microscopic inflammation had occurred. Sustained colitis of CD4+CD45RBhi T cell reconstituted mice was also associated with enhanced subepithelial resistance, enhanced paracellular permeability, and decreased net ion transport. All these reflect a disturbance of barrier function and may contribute to diarrhea. Epithelial resistance was positively correlated with interleukin 10 (IL‐10) and transforming growth factor &bgr; (TGF‐&bgr;) levels and net ion transport inversely correlated with tumor necrosis factor alpha (TNF‐&agr;) levels, pointing to the protective effect of IL‐10 and TGF‐&bgr; and to a damaging effect of TNF‐&agr;. Indomethacin, a nonselective COX inhibitor, decreased epithelial resistance independent of T cells and inflammation, but its effect was more pronounced in inflamed colon. Conclusions: Induction of colitis by transfer of CD4+CD45RBhi T cells in SCID mice leads to changes in the colonic epithelium before colitis develops. Decreased epithelium resistance might contribute to the development of colitis; however, it is not sufficient to lead to chronic inflammation. (Inflamm Bowel Dis 2010)

New drug therapies on the horizon for IBD

IBD are diseases of the GI tract causing important morbidity. Several anti-TNF-α agents are currently used to treat Crohn’s disease and ulcerative colitis. Although these molecules have dramatically improved the treatment of IBD, up to half of the patients have no sustained benefit due to nonresponse, loss of response or intolerance. New anti-TNF strategies are under development. Novel therapies targeting other immune pathways are under study, such as antibodies targeting the IL-12/IL-23 pathway, and have shown interesting preliminary results. In parallel, antiadhesion therapies limiting the recruitment of cells to the gut will reach the clinic in the coming years. Small molecules inhibiting the production of proinflammatory cytokines are already used in the clinic for rheumatoid arthritis, and Tofacitinib, a Janus kinase inhibitor, seems to have great potential to treat ulcerative colitis. In this review we focus on the novel molecules that are likely to reach the clinic in the near future for the treatment of IBD.

Effects of haptoglobin polymorphisms and deficiency on susceptibility to inflammatory bowel disease and on severity of murine colitis

Background Haptoglobin (Hp) is a haemoglobin-binding protein with immunomodulatory properties. Its gene (16q22) harbours a common polymorphism with two different alleles: Hp1 and Hp2. Genotype Hp22 has been shown to be over-represented in different immune diseases. Results in Crohns disease (CD) are contradictory. Aims To determine whether Hp plays a role in inflammatory bowel disease, both genetically and functionally. Methods 1061 patients with CD, 755 with ulcerative colitis (UC) and 152 with primary sclerosing cholangitis, as well as 452 healthy controls, were genotyped using touch-down PCR. To confirm association results, 464 CD trios and 151 UC trios were genotyped. Serum Hp concentrations were determined in 62 individuals of different genotype. Colitis was induced in mice with dextran sulphate sodium (DSS) and oxazolone (Oxa). Cytokine production was evaluated by mRNA quantification in colonic tissue and ELISA on supernatants of mesenteric lymph node cells. Results Prevalence of Hp2 was higher in CD and UC than in controls. In the confirmatory cohorts, Hp2 was over-transmitted to the affected offspring. Serum Hp concentrations were higher in individuals with genotypes Hp11 and Hp21 than in those with Hp22 (1.38 vs 0.89 g/l). DSS- and Oxa-induced colitis were more severe in Hp-deficient mice than in control mice and accompanied by higher concentrations (although not statistically significantly different) of tissue mRNA for cytokines. Interleukin-17 production was significantly higher in the presence of Hp-deficient serum compared with wild-type serum. Conclusions The Hp gene may play a role in susceptibility to inflammatory bowel disease. Its implication in other immune diseases underscores the common pathways between these diseases. Experimental models of colitis showed that Hp has a protective role in inflammatory colitis, most likely by inhibiting the production of Th1 and Th17 cytokines.

Interleukin‐15 receptor α expression in inflammatory bowel disease patients before and after normalization of inflammation with infliximab

Interleukin‐15 (IL‐15) is a pro‐inflammatory cytokine thought to contribute to the inflammation in inflammatory bowel diseases (IBD). The specific receptor chain IL‐15Rα can be expressed as a transmembranous signalling receptor, or can be cleaved by a disintegrin and metalloprotease domain 17 (ADAM17) into a neutralizing, soluble receptor (sIL‐15Rα). The aim of this study is to evaluate the expression of IL‐15Rα in ulcerative colitis (UC) and Crohns disease (CD) patients before and after infliximab (IFX) therapy. Gene expression of IL‐15Rα, IL‐15 and ADAM17 was measured at the mRNA level by quantitative reverse transcription‐PCR in mucosal biopsies harvested before and after first IFX therapy. Concentrations of sIL‐15Rα were measured in sera of patients by ELISA and IL‐15Rα protein was localized in the gut by immunohistochemistry and immunofluorescence. Mucosal expression of IL‐15Rα is increased in UC and CD patients compared with controls and it remains elevated after IFX therapy in both responder and non‐responder patients. The concentration of sIL‐15Rα in serum is also increased in UC patients when compared with controls and does not differ between responders and non‐responders either before or after IFX. CD patients have levels of sIL‐15Rα comparable to healthy controls before and after therapy. In mucosal tissues, IL‐15Rα+ cells closely resemble activated memory B cells with a pre‐plasmablastic phenotype. To conclude, IBD patients have an increased expression of IL‐15Rα mRNA in the mucosa. Expression is localized in B cells, suggesting that IL‐15 regulates B‐cell functions during bowel inflammation. No change in release of sIL‐15Rα is observed in patients treated with IFX.