Vedolizumab (Entyvio)

Vedolizumab does not bind to the majority of memory CD4+ T lymphocytes (60%), neutrophils, and most monocytes. The highest level of vedolizumab binding is to a subset (25%) of human peripheral blood memory CD4+ T lymphocytes that include gut-homing interleukin 17 T-helper lymphocytes. Vedolizumab also binds to eosinophils at high levels, and to naive T-helper lymphocytes, naive and memory cytotoxic T lymphocytes, B lymphocytes, natural killer cells, and basophils at lower levels; vedolizumab binds to memory CD4+ T and B lymphocytes with subnanomolar potency (EC50=0.3-0.4 nM). Vedolizumab selectively inhibits adhesion of alpha4beta7-expressing cells to mucosal addressin cell adhesion molecule 1 (IC50=0.02-0.06 g/mL) and fibronectin (IC50=0.02 g/mL), but not vascular cell adhesion molecule 1.

Kinase Assay: Vedolizumab inhibition of high-affinity binding of MAdCAM-1 to human peripheral blood memory CD4+ T lymphocytes is tested. Peripheral blood (90 uL) is incubated with a saturating concentration (3 ug/mL) of MAdCAM-1-murine-Fc fusion protein and 4 mM MnCl2 in a final volume of 100 uL for 1 h at room temperature, in the presence or absence of vedolizumab. After washing with assay buffer, the cells are stained with fluorescentlabeled anti-mouse IgG for 15 min at room temperature. After washing again, cells are incubated with mouse serum for 10 min at room temperature, followed by staining with anti-CD4 and anti-CD45RO antibodies for 15 min at room temperature. After washing, red blood cells are lysed with BD FACS lysing solution and analyzed by flow cytometry in a FACSCalibur with CellQuest Pro software.