UROKINASE-TYPE PLASMINOGEN-ACTIVATOR - A PARACRINE FACTOR REGULATING THE BIOAVAILABILITY OF IGFS IN PA-III CELL-INDUCED OSTEOBLASTIC METASTASES

The transplantation of PA-III rat prostate cancer cells onto rat skele ton produces osteoblastic metastases. Therefore we studied the paracri ne interactions between the PA-III cells and osteoblast-derived osteos arcoma cells (UMR 106 cells). A serine protease secreted by PA-III cel ls hydrolyzed IGF-binding protein-1 and IGF-binding protein-2 (IGFBP-1 and IGFBP-2) detected in the cell culture media (CM) of OMR 106 cells by western ligand blotting. The serine protease of PA-III cell CM was purified using a benzamidine affinity column. This protease was a pro tein of 45-50 kDa on polyacrylamide gel electrophoresis under non-redu cing conditions but generated two protein bands under reducing conditi ons; a) one of 33-35 kDa possessing protease activity and b) another o f 20-25 kDa which was proteinolytically inactive. Sequence analysis id entified the amino acid sequence of the a-chain (20-25 kDa band) and o f the b-chain (33-35 kDa band) of rat urokinase-type plasminogen activ ator molecule. Urokinase purified from PA-III cell CM hydrolyzed IGFBP s of UMR 106 cells and stimulated the proliferation of UMR 106 cells i n serum-free cultures. Its protease activity was abolished by benzamid ine and aprotinin. Its mitogenic activity for osteoblasts was inhibite d by anti-IGF-I monoclonal antibody. Northern blot analysis documented the expression of the urokinase-type plasminogen activator-gene in th e mRNA extracted from PA-III cells. Unokinasse expression was inhibite d by dexamethasone. Therefore, we conclude that urokinase-type plasmin ogen activator stimulates osteoblasts via an IGF-I dependent mechanism . Hydrolysis of the IGFBOPs at the sites of PA-III cell-induced bone t umors account for an increased bioavailability of IGFs. This may facil itate the development and the growth of PA-III cell-induced bone tumor and can also mediate the subsequent local osteoblastic reaction.