Background and Aims The potassium channel Kv1.3 is a potentially attractive therapeutic target in T cell-mediated inflammatory diseases, as the activity of antigen-activated T cells is selectively impeded by Kv1.3 inhibition. In this study, we examined Kv1.3 as a potential therapeutic intervention point for ulcerative colitis (UC), and studied the efficacy of DES1, a small-molecule inhibitor of Kv1.3, in vitro and in vivo. Methods Kv1.3 expression on T cells in peripheral blood mononuclear cells (PBMCs) isolated from donors with and without UC was examined by flow cytometry. In biopsies from UC patients, Kv1.3-expressing CD4+ T cells were detected by flow cytometry and immunohistochemistry. In vitro, we determined the ability of DES1 to inhibit anti-CD3-driven activation of T cells. In vivo, the efficacy of DES1 was determined in a humanized mouse model of UC and compared to infliximab and tofacitinib in head-to-head studies. Results Kv1.3 expression was elevated in PBMCs from UC patients and correlated with the prevalence of TH1 and TH2 T cells. Kv1.3 expression was also detected on T cells from biopsies of UC patients. In vitro, DES1 suppressed anti-CD3-driven activation of T cells in a concentration-dependent manner. In vivo, DES1 significantly ameliorated inflammation in the UC model and most effectively so when PBMCs from donors with higher levels of activated T cells were selected for reconstitution. The efficacy of DES1 was comparable to that of either infliximab or tofacitinib. Conclusion Inhibition of Kv1.3 (by DES1, for instance) appears to be a potential therapeutic intervention strategy for UC patients.

With glucose being the preferred source of energy in activated T cells, targeting glycolysis has become an attractive therapeutic intervention point for chronic inflammatory bowel diseases (IBD). The switch to glycolysis is mediated by phosphoinositide‐3‐kinases (PI3K) which relay signals from surface receptors to the AKT pathway. We first confirmed by analysis of the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) that metabolism is shifted towards glycolysis in IBD patients as compared to non‐IBD donors. In contrast to non‐IBD donors, OCR correlated with ECAR (IBD: cor = 0.79, p = 2E‐10; non‐IBD: cor = 0.37, p = n.s.), in IBD patients. Second, we tested the PI3K inhibitor copanlisib as a potential therapeutic. Ex vivo, copanlisib suppressed the ECAR significantly in T cells activated by anti‐CD3 antibodies and significantly decreased ECAR rates in the presence of copanlisib (anti‐CD3: 58.24 ± 29.06; copanlisib: 43.16 ± 20.23, p < .000. In addition, copanlisib impaired the activation of CD4+ CD25+ T cells (anti‐CD3: 42.15 ± 21.46; anti‐CD3 + copanlisib: 26.06 ± 21.82 p = .013) and the secretion of cytokines (IFNγ: anti‐CD3: 6332.0 ± 5707.61 pmol/ml; anti‐CD3 + copanlisib: 6332.0 ± 5707.61, p = .018). In vivo, copanlisib significantly improved the histological scores (ethanol: 8.5 ± 3.81; copanlisib: 4.57 ± 2.82, p = .006) in the NSG‐UC mouse model. Orthogonal partial least square analysis confirmed the efficacy of copanlisib. These data suggest that the PI3K pathway provides an attractive therapeutic intervention point in IBD for patients in relapse. Targeting metabolic pathways have the potential to develop phase dependent therapies.

Introduction Crohn's disease (CD) is characterized by pronounced intestinal fibrosis and severe mucosal damage and conventional animal models are limited to reflect these pathological manifestations. The aim of this study was to examine whether the combination of patient immune‐profiling and preclinical studies in a mouse model based on NOD/scid IL‐2Rγnull (NSG) reconstituted with peripheral blood mononuclear cells (PBMC) from CD patients has the capacity to harmonize ex vivo human and in vivo animal studies. Methods Immunological profiles of CD (n = 24) and ulcerative colitis (UC) patients (n = 47) were established by flow cytometry of subgroups of immune cells and subjected to hierarchical cluster and estimation graphics analyses. Pathological phenotypes of NSG mice, which were reconstituted with PBMC from CD, UC, and non‐IBD donors (NSG‐CD, NSG‐UC, and NSG‐non‐IBD) were compared. Readouts were the clinical, colon, and histological scores; subtypes of immune cells from spleen and colon; and levels of inflammatory markers, such as c‐reactive protein (CRP), monocyte chemotactic protein (MCP)‐3, transforming growth factor‐beta (TGFß), and hepatocyte growth factor (HGF). Fibrocytes were identified by immunohistochemistry in colonic sections. Results CD patients were significantly clustered in a group characterized by increased levels of TH1, TH2 cells, and decreased levels of CD14+ CD163+ monocytes (p = .003). In contrast to NSG‐UC mice, NSG‐CD mice exhibited an immune‐remodeling phenotype characterized by enhanced collagen deposition, elevated levels of CD14+ CD163+ monocytes, HGF, and TGFß. This phenotype was further corroborated by the presence of human fibrocytes as components of fibrotic areas. Conclusion The NSG‐CD model partially reflects the human disease and allows for studying the development of fibrosis.

Background Recently, we have developed a mouse model that relies on NOD-scid IL-2Rγnull (NSG) mice reconstituted with peripheral blood mononuclear cells (PBMC) derived from patients with ulcerative colitis (UC, NSG-UC). In this model, symptoms of UC are induced by rectal challenge with ethanol. The objective of the study was to adapt this model to Crohn’s disease and to compare the phenotypes of the NSG-UC and NSG-CD mouse models. Methods NSG mice were reconstituted with PBMC from UC (n = 4) or CD (n = 3) patients. Mice were separated into two groups: unchallenged control and ethanol challenged mice. Readout were clinical-, colon and histological scores, analysis of frequencies of subgroups of human T cells, monocytes and B cells isolated from spleen and colon by flow cytometry, and the expression levels of the inflammatory markers TGFß, CRP, MCP-3, and IL-6 in the colon or serum by Luminex analysis. Results The pathological phenotype was markedly different in NSG-UC mice as compared with NSG-CD mice. Firstly, histological analysis revealed that NSG-UC mice exhibited more of a pro-inflammatory phenotype as indicated by a severe influx of inflammatory cells, oedema, crypt loss, crypt abscesses and epithelial hyperplasia. In contrast, NSG-CD mice displayed crypt loss and goblet cell atrophy and pronounced fibrosis indicating ongoing wound healing processes. These observations were corroborated by frequencies of splenic and colonic leucocytes. Antigen experienced CD4+ T (CD45RO+) cells and switched B cells (CD19+ CD27+ IgD-) were significantly increased in NSG-UC mice, whereas significantly higher levels of experienced CD8+ T cells and M1 (CD64+), M2 (CD163+) CD14+ monocytes and unswitched B cells (CD19+ CD27+ IgD+) indicated a monocyte driven inflammation in NSG-CD mice. This observation was also reflected in the colon of mice. Inflammation in UC was characterised by increased frequencies of neutrophils, activated CD4+ T cells (CD69+) and increased levels of CRP and MCP-3, whereas NSG-CD mice were signified by increased frequencies of M2 and M1 monocytes and of TGFß levels. In contrast to NSG-CD mice, NSG-UC mice also displayed higher serum levels of IL-6. Secondly, the impact of ethanol was more pronounced in the NSG-UC mice. In NSG-CD mice, the challenge did not evoke significant differences as compared with unchallenged control mice. Conclusion The comparison of pathological phenotypes of the NSG-UC and NSG-CD mouse models revealed differences, some of which reflect the respective human disease. The NSG-UC and NSG-CD mouse models may constitute powerful tools to get a better understanding of the different inflammatory processes in UC and CD.