Effect of GW654652 on MM cell migration. (A-B) Growth factor-deprived MM.1S cells were pretreated with 10 μg/mL GW654652 (A) or 1 and 10 μg/mL GW654652 (B) or were left untreated. Cells were then plated on a fibronectin-coated polycarbonate membrane (8-μm pore size) in a modified Boyden chamber and were exposed for 4 hours to 5 to 50 ng/mL VEGF (A) or 5 ng/mL VEGF (B) in the lower chamber. (C) Growth factor-deprived MM.1R, RPMI-Dox40 (Dox40), U266, and RPMI cells were pretreated with 10 μg/mL GW654652 or were left untreated, plated on a fibronectin-coated polycarbonate membrane (8-μm pore size) in a modified Boyden chamber, and exposed to 5 ng/mL VEGF for 4 hours. (D) VEGF-triggered cell migration was determined in MM.1S cells after GW654652 removal (GW re) compared with migration in cells treated with GW654652 (GW) or left untreated (contr). At the end of treatment, cells on the lower part of the membrane were counted using a Coulter counter ZBII. Data represent mean ± SD for duplicate samples. Results shown are representative of 3 independent experiments.