Limited Dose-Dependent Effects of Vedolizumab on Various Leukocyte Subsets

Abstract

Objectives: The anti-α4β7 integrin antibody vedolizumab (VDZ) is successfully used for the treatment of inflammatory bowel diseases. However, only a subgroup of patients respond to therapy. VDZ is administered at a fixed dose, leading to a wide range of serum concentrations in patients. Previous work from our group showed a dose-dependent preferential binding of VDZ to effector compared with regulatory CD4 + T cells. Therefore, we aimed to determine the dose-dependent binding profile of VDZ to other leukocyte subsets.

Methods: We characterized α4β7 integrin expression on CD8 + T cells, CD19 + B cells, CD14 + monocytes, natural killer cells, and eosinophils from patients with inflammatory bowel disease and healthy controls. We studied the binding of VDZ to these cells at different concentrations and investigated the functional consequences for dynamic adhesion and transmigration in vitro .

Results: The expression of α4β7 differed between the analyzed leukocyte subsets and was significantly higher on eosinophils from inflammatory bowel disease patients compared with controls. Almost all α4β7-expressing cells from these subsets were bound by VDZ at a concentration of 10 μg/mL. Dynamic cell adhesion was significantly impaired in all subsets, but there were no dose-dependent differences in the inhibition of adhesion.

Discussion: Our data suggest that α4β7-expressing CD8 + T cells, CD19 + B cells, CD14 + monocytes, natural killer cells, and eosinophils are a target of VDZ. However, there do not seem to be concentration-dependent differences, regarding the effects on these cells in the clinically relevant range. Thus, the reported exposure-efficacy characteristic of VDZ can probably mainly be attributed to CD4 + T-cell subsets.

Figure 1.

α4β7 integrin expression on different leukocyte subsets in healthy controls (Ctrl), Crohn's disease (CD), or ulcerative colitis (UC) patients. Representative (left) and quantitative (right) flow cytometry of CD8⁺ T cells (a), CD19⁺ B cells (b), natural killer (NK) cells (c), monocytes (d), and eosinophils (e). n = 3–11 per group. Results depicted as bar graphs with SEM and single data points. Significant outliers were identified using the Grubbs test (P = 0.05) and excluded from analysis. Statistical comparisons were performed using 1-way ANOVA with Tukey's multiple comparisons test. *P < 0.05; **P < 0.01; ***P < 0.001.

Figure 2.

Dose-dependent binding of vedolizumab (VDZ) to α4β7 integrin-expressing different leukocyte subsets. (a) Quantitative flow cytometry of α4⁺β7⁺VDZ⁺ cells in different leukocyte subsets incubated with the indicated concentrations of fluorescently labeled VDZ. n = 15–20 per group. (b) Quantitative flow cytometry of α4⁺β7⁺VDZ⁺ cells in different leukocyte subsets compared between healthy controls (Ctrl) and patients with inflammatory bowel disease (IBD). n = 5–12 per group. Results depicted as bar graphs with SEM and single data points. Significant outliers were identified using the Grubbs test (P = 0.05) and excluded from analysis. Statistical comparisons were performed using Kruskal-Wallis with Dunn's multiple comparisons test (a) and 2-way ANOVA with Sidak's multiple comparisons test (b). *P < 0.05; **P < 0.01; ***P < 0.001.

Figure 3.

Concentration-dependent adhesion of different leukocyte subsets to mucosal addressin cell adhesion molecule (MAdCAM-1). Dynamic adhesion of magnetic-activated cell sorting (MACS) purified CD8⁺ T cells (a), CD19⁺ B cells (b), natural killer (NK) cells (c), CD14⁺ monocytes (d), and eosinophils (e). Left panels: Representative microscopic images of adhered cells (overlay of counted high power fields); middle panels: quantification of the background-corrected number of cells incubated with or without the indicated concentrations of vedolizumab (VDZ) (adhering to MAdCAM-1 [sum of 8 counted high power fields]; right panels: relative inhibition of adhesion of cells to MAdCAM-1 after treatment with the indicated concentrations of VDZ [background-corrected]). n = 6–12. Results depicted as bar graphs with SEM and single data points. Significant outliers were identified using the Grubbs test (P = 0.05) and excluded from analysis. Statistical comparisons were performed using repeated measurement 1-way ANOVA with Tukey's multiple comparisons test or paired t-test. *P < 0.05; **P < 0.01; ***P < 0.001.

Figure 4.

Concentration-dependent mucosal addressin cell adhesion molecule (MAdCAM-1)-driven transmigration of different leukocyte subsets. Representative (left) and quantitative (right) flow cytometry of transmigrated CD8⁺ T cells (a), CD19⁺ B cells (b), natural killer (NK) cells (c), and eosinophils (d) after treatment with the indicated concentrations of vedolizumab (VDZ) or corresponding isotype control (untreated). n = 5–8. Results depicted as bar graphs with SEM and single data points. Significant outliers were identified using the Grubbs test (P = 0.05) and excluded from analysis. Statistical comparisons were performed using repeated measurement 1-way ANOVA with Tukey's multiple comparisons test. *P < 0.05; **P < 0.01; ***P < 0.001.