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Background and Aims To inform their future use in regulated clinical trials to evaluate treatment efficacy hypotheses, the measurement properties of three histologic indices, Geboes Score (GS), Robarts Histopathology Index (RHI), and Nancy Index (NI), were evaluated among patients with ulcerative colitis. Methods Analyses were conducted on data from a Phase 3 clinical trial of adalimumab (M14-033, n=491) and focused on evaluating the measurement properties of the GS, RHI, and NI. Specifically, internal consistency and inter-rater reliability, convergent, discriminant, and known-groups validity, and sensitivity to change were assessed at Baseline, and Weeks 8 and 52. Results Internal consistency for the RHI showed lower alpha (α) values at Baseline (α=0.62) relative to Weeks 8 (α=0.82) and 52 (α=0.81). The inter-rater reliability values of RHI (0.91), NI (0.64), and GS (0.53) were excellent, good, and fair, respectively. Regarding validity, Week 52 correlations were moderate to strong between full and partial Mayo scores and Mayo subscale scores and the RHI and GS, and were weak to moderate for the NI. Significant differences between mean scores of all three histologic indices were observed across known-groups based on Mayo endoscopy subscores and full Mayo scores at Weeks 8 and 52 (p Conclusions The GS, RHI, and NI are each capable of producing reliable and valid scores that are sensitive to changes in disease activity over time, in patients with moderately to severely active ulcerative colitis. While all three indices demonstrated relatively acceptable measurement properties, the GS and RHI performed better than the NI.

Intestinal permeability and neutrophil activity are closely linked to inflammatory bowel disease (IBD) pathophysiology. Here we discuss two techniques for assessing permeability and neutrophil activity in mouse IBD models using near infrared (NIR) detection. To address the limitation of visible light readouts—namely high background—IRDye 800CW was used to enable rapid, non-terminal measurements of intestinal permeability. The increased sensitivity of NIR readouts for colon permeability is shown using dextran sulfate sodium (DSS) and anti-CD40 murine colitis models in response to interleukin-22 immunoglobulin Fc (IL22Fc) fusion protein and anti-p40 monoclonal antibody treatments, respectively. In addition to enhanced permeability, elevated levels of neutrophil elastase (NE) have been reported in inflamed colonic mucosal tissue. Activatable NIR fluorescent probes have been extensively used for disease activity evaluation in oncologic animal models, and we demonstrate their translatability using a NE-activatable reagent to evaluate inflammation in DSS mice. Confocal laser endomicroscopy (CLE) and tissue imaging allow visualization of spatial NE activity throughout diseased colon as well as changes in disease severity from IL22Fc treatment. Our findings with the 800CW dye and the NE probe highlight the ease of their implementation in preclinical IBD research.

Ly108, a glycoprotein of the signaling lymphocytic activation molecule family of cell surface receptors expressed by T, B, NK, and APCs has been shown to have a role in NK cell cytotoxicity and T cell cytokine responses. In this study, we describe that CD4+ T cells from mice with a targeted disruption of exons 2 and 3 of Ly108 (Ly108ΔE2+3) produce significantly less IL-4 than wild-type CD4+ cells, as judged by in vitro assays and by in vivo responses to cutaneous infection with Leishmania mexicana. Surprisingly, neutrophil functions are controlled by Ly108. Ly108ΔE2+3 mice are highly susceptible to infection with Salmonella typhimurium, bactericidal activity of Ly108ΔE2+3 neutrophils is defective, and their production of IL-6, IL-12, and TNF-α is increased. The aberrant bactericidal activity by Ly108ΔE2+3 neutrophils is a consequence of severely reduced production of reactive oxygen species following phagocytosis of bacteria. Thus, Ly108 serves as a regulator of both innate and adaptive immune responses.

Originally described by Uhlig et. al., the anti-CD40 agonist acute model of gut inflammation is a useful tool for interrogating myeloid-driven gut inflammation. Previous studies have shown that this model is sensitive to intervention with anti-TNFα, anti-IFNγ and anti-IL12/ 23p40. Here we present data that extend previous findings in this model and demonstrate an anti-CD40 dose dependent severity of disease pathogenesis as well as dose response curves for treatment with anti-TNFα and anti-IL12/23p40. In addition, we have identified both circulating and fecal biomarkers of disease. Methods: RAG2 deficient mice on the C57BL/6 background were injected i.p. with a single dose of anti-CD40 agonist antibody (FGK45) on day 0 and monitored daily for body weight loss until day 7. Prophylactic treatment with either anti-TNFα or anti-IL12/23p40 was performed using i.p. injections on day -1 and day 2 prior to sacrifice at day 7. Spleens were collected and weights recorded, plasma and feces were also collected for drug exposure levels and biomarker analysis. The colon was removed, flushed with PBS and buffered formalin prior to histological sectioning and scoring. Results: Mice treated with anti-CD40 (single dose 1.25-20mpk) exhibited a dose-dependent increase in splenomegaly, colonic inflammation and liver pathology with an induction ED50 of 2.5mpk. In addition, plasma levels of CXCL-10, MCP-1, MIG and other inflammatory mediators were also increased dose-dependently with CXCL-10 levels highly correlating with histology scores (r2 = 0.85). Fecal calprotectin (S100A8-S100A9 dimer), a biomarker routinely used in the clinic, also increased dose-dependently and correlated well with histologic scores (r2 = 0.72 and 0.68 respectively). As significant liver necrosis was observed at doses of anti-CD40 above 5mpk, this dose was selected for intervention studies. Prophylactic treatment with either anti-IL12/23p40 or anti-TNFα attenuated colonic pathology in a dose-dependent manner at day 7 of disease with an EC50 of 100 μg/mL for anti-p40. Therapeutic intervention (i.e. a single injection given at day 3 of disease) with anti-TNFα also significantly reduced colon histopathology scores while antiIL-12/23p40 only showed therapeutic significance at the highest dose (EC50 of 7 and 1.3 μg/mL respectively). Reduced colonic inflammation in all cases was accompanied by a reduction in circulating CXCL-10 levels. In summary, we have shown that the anti-CD40 acute model of gut inflammation is a useful drug discovery model for interrogating myeloid mechanisms involved in colitis and have identified circulating CXCL-10 and fecal calprotectin as biomarkers related to both pathogenesis and intervention in this model. We have also generated dose-efficacy relationships for clinically validated therapies such as anti-TNF and anti-IL12/23p40.