P001 Comparison of in vivo T cell homing to the inflamed gut with the etrolizumab surrogate antibody FIB504 and vedolizumab in a humanised mouse model
S. Zundler*, A. Fischer, R. Atreya, C. Neufert, I. Atreya, M. F. Neurath
University of Erlangen-Nuremberg, Department of Medicine I, Erlangen, Germany
The anti-α4β7 antibody vedolizumab (VDZ), which inhibits gut homing of lymphocytes via interaction of α4β7 with MAdCAM-1, has greatly increased therapeutic options in patients with IBD. However, lymphocyte homing may also occur via other homing molecules like the α4β1 integrin and considerable numbers of patients do not respond to VDZ therapy. The anti-β7 antibody etrolizumab (ETZ) is currently tested in phase III trials and additionally blocks the binding of αEβ7 to E-Cadherin, which is believed to mediate epithelial retention of homed lymphocytes.
AEβ7 expression was determined on peripheral blood and lamina propria lymphocyte subsets of UC and CD patients and healthy donors by flow cytometry or immunofluorescence staining, respectively. The regulation of αEβ7 expression upon lymphocyte stimulation and incubation with cytokines was studied. In in-vitro adhesion assays the adhesive capacities of lymphocytes to MAdCAM-1 and E-Cadherin and the inhibitory potential of VDZ and the ETZ surrogate antibody FIB504 (ETZs) were tested. Finally, lymphocytes from UC patients were treated with either of the compounds, fluorescence labelled and injected into the ileocolic artery of immunosuppressed mice. Gut homing was assessed by in-vivo confocal microscopy and flow cytometry of lamina propria cells.
AEβ7 expression is significantly higher on CD8+ lymphocytes than on CD4+ lymphocytes both in the peripheral blood and the gut. Between both subsets, αEβ7 expression is correlated with IL-9 secretion, whereas CD4+IL9+ cells express less α4β7 than other CD4+ subsets did. Moreover, CD8+ cells exhibit a notably greater potential to increase αEβ7 expression upon stimulation and TGF-β treatment. ETZs markedly inhibited binding of CD4+ and CD8+ lymphocytes to rhE-Cadherin and blocked the adhesion of CD4+ and CD8+ lymphocytes to rhMAdCAM-1 to a degree comparable with VDZ. Fewer lymphocytes bound to a mix of both ligands upon treatment with ETZs compared with VDZ. In our humanised mouse model, the portion of human CD8+ cells in the murine gut was significantly reduced 3 hours after injection when cells were treated with ETZs vs VDZ.
Amongst CD4+ cells, the fraction of PU.1+ cells was decreased.
Figure 1. "In vivo microscopy after injection of human CD8+ cells (green) treated with VDZ or ETZs. Red: vessels. Blue: murine cells. Lower panel: Quantitative analysis of intestinal infiltration of CD8+ cells."
VDZ may not equally cover all pathogenetically relevant lymphocyte subsets leading to insufficient therapeutic response in predisposed patients. ETZ seems to offer superior reduction of intestinal lymphocyte infiltration, especially concerning CD8+ and Th9 cells.