Regulation of the Plasminogen Activator System in the MDA‐MB‐231 Breast Cancer Cell Line

Abstract

The objective of this study was to determine the contribution of the urokinase‐type plasminogen activator (uPA) and plasminogen activator inhibitor 1 (PAI‐1) to invasiveness of the MDA‐MB‐231 breast cancer cell line, and to study control of uPA and PAI‐1 expression. Amiloride, a small molecule inhibitor of uPA, and a uPA blocking antibody inhibited invasion of the 231 cells by 30% while a PAI‐1 blocking antibody increased invasion by 50%. Treatment with the PI3K inhibitor LY294002 increased PAI‐1 protein levels in conditioned media while differences in uPA were not detected. However, another PI3K inhibitor, wortmannin, decreased uPA in conditioned media. Semi‐quantitative RT‐PCR indicated LY294002 and wortmannin increased PAI‐1 mRNA levels with a concomitant decrease in uPA mRNA. Amiloride treatment decreased PI3K activity resulting in increased PAI‐1 in conditioned media. Treatment with the receptor associated peptide (RAP), which blocks activity of the low density like lipoprotein receptor responsible for binding and clearing uPA‐PAI‐1 complexes from the cell surface, decreased uPA mRNA and protein levels. These results confirm uPA and PAI‐1 are important contributors to MDA‐MB‐231 invasiveness, PI3K positively regulates uPA while negatively regulating PAI‐1, and the PI3K pathway which regulates uPA and PAI‐1 expression may be activated by the recognition of uPA‐PAI‐1 complexes at the cell surface.