Figure 6.

Metalloprotease activity is required for thrombin-induced epidermal growth factor (EGF) receptor (EGFR) and ErbB2 transactivation and cellular invasion. (a and b) Serum-deprived MDA-MB-231 cells were preincubated with 0.1% dimethyl sulfoxide (DMSO) (vehicle control), 10 µM GM6001 active or inactive agent for 1 h at 37 °C and then treated with or without 10 nM thrombin for 5 min at 37 °C. EGFR and ErbB2 tyrosine phosphorylation was detected as described in Figure 1. The data are expressed as fold increase over control and are representative of three different experiments. (c) MDA-MB-231 cells were pretreated for 1 h with 10 mM GM6001 active or inactive control agent then incubated with 1 pM thrombin at 37 °C. Cells were then added to the upper well and cellular invasion toward conditioned medium (CM) containing inactive or active GM6001 was assessed. (d) Serum-starved MDA-MB-231 cells were preincubated with 20 µg/ml of CRM 197, 100 ng/ml heparin or vehicle control for 30 min at 37 °C and then treated in the absence (Ctrl) or presence of 10 nM Th or 16 nM EGF ligand for 5 min at 37 °C. ErbB tyrosine phosphorylation was then examined as described above.