Figure 6.

Dasatinib decreases SFK activation and blocks cell migration promoted by activation of the uPA-uPAR signaling system. (a) U373MG cells that express EGFRvIII under the control of a Dox repressible promoter were treated with Dox or with vehicle for 4 days and then with 0.3 µM Dasatinib (+) or vehicle (DMSO) (−) in SFM for 4 h. Immunoblot analysis was performed to detect phosphorylated Tyr-416 in SFKs and tubulin as a control for load. (b) U373MG cells were treated with Dox or vehicle for 4 days. The cells were then added to Transwells in the presence of 0.3 µM Dasatinib (grey bar) or vehicle (DMSO, black bar). Cells were allowed to migrate for 18 h. Serum (10% FBS) was added only to the lower chamber. The number of migrating cells was determined and standardized against that observed with vehicle-treated U373MG cells (mean ± SEM, n=3, *, p<0.05). (c) ESC cell lines (ESC1, ESC2 and ESC5) were treated with 0.3 µM Dasatinib (+) or vehicle (DMSO) (−) in SFM for 4 h. Immunoblot analysis was performed to detect phosphorylated Tyr-416 in SFKs, phosphop130Cas, and tubulin as a control for load. (d) Escaper cells (three separate cell lines) were treated with Dasatinib (gray bars) or DMSO (black bars) by adding drug to the top and bottom chambers of Transwells. Cell migration was then studied (mean ± SEM, n=3. **, p<0.01). (e) EGFRvIII-expressing U373MG cells were treated with Gefitinib (1 µM) or vehicle (DMSO) for 4 days. These cells were then treated with 0.3 µM Dasatinib (grey bar) or vehicle (black bar) by adding drug to the top and bottom chambers of Transwells. Cells were allowed to migrate for 18 h. The number of migrating cells was standardized to that observed with vehicle-treated U373MG cells (mean ± SEM, n=3, *, p<0.05).