4-[3-(2-Chlorophenyl)-2-propen-1-ylidene]-1-phenyl-3,5-pyrazolidinedione

IC50: 22.8 µM (GEF activity of DOCK2 ^DHR-2 for Rac1)

CPYPP binds to DOCK2 DHR-2 domain in a reversible manner and inhibited its catalytic activity in vitro. When lymphocytes are treated with CPYPP, both chemokine receptor- and antigen receptor-mediated Rac activation are blocked, resulting in marked reduction of chemotactic response and T cell activation.
Although overexpression of DOCK2 induces Rac activation in HEK293T cells, this activation is markedly suppressed by treating the cells with CPYPP at 100 µM for 1 hr before assay.

When 2.5 mg/kg of CPYPP is administrated intravenously, the plasma concentration of CPYPP is only 2.4 µM at 30 min. However, by intraperitoneally injecting 250 mg/kg of CPYPP into mice, the plasma concentration of CPYPP reached to 11.3 µM at 30 min and 10.9 µM at 1 hr, respectively.
The adoptively transferred spleen cells from mice that has been made by a ‘‘knock-in’’ strategy to express endogenous DOCK2 as a fusion protein with green fluorescent protein (GFP). Intraperitoneal injection of CPYPP (5 mg per mouse) 1 hr before adoptive transfer reduces the percentage of the migrated T cells to <25% of the control level.