AUTOCRINE REGULATION OF CELL-PROLIFERATION BY THE INSULIN-LIKE GROWTH-FACTOR (IGF) AND IGF BINDING PROTEIN-3 PROTEASE SYSTEM IN A HUMAN PROSTATE CARCINOMA CELL-LINE (PC-3)

PC-3 cells, whose growth is androgen-independent, were shown to be cap able of slow proliferation in serum-free medium and in the absence of added growth factor for 7 days. They secreted insulin-like growth fact or (IGF)-II but no detectable IGF-I. This IGF-II, although produced in small amounts, plays a role in their proliferation because growth cou ld be inhibited dose dependently by up to 80% in the presence of monoc lonal antibodies directed against IGFs or the type 1 IGF receptor. PC- 3 cells also secreted IGF binding proteins (IGFBPs) -2, -3, -4, and -6 . Immunoblot analysis revealed selective proteolysis of IG-FBP-3, yiel ding fragments of the same molecular size as those generated from IGFB P-3 in vivo. With the addition to the culture medium of a serine prote ase inhibitor, 4-(2-aminoethyl)-benzenesulfonyl fluoride (Pefabloc-SC) , at concentrations <0.2 mM that were nontoxic to the cells, cell prol iferation was dose dependently inhibited up to 80% and, at the same ti me, proteolysis of the IGFBP-3 secreted by the cells was depressed. Ur okinase activity detected in the conditioned media was depressed by Pe fabloc, suggesting that the urokinase-type plasminogen activator was i nvolved in the proteolysis of IGFBP-3. In addition, 0.01-5 mu g/ml pla sminogen induced a dose-dependent increase in both proliferation and t he proportions of proteolysed IG-FBP-3 in the media. The stimulation o f proliferation was totally blocked in the presence of anti-type 1 IGF receptor antibody. Recombinant human IGF-II (5-200 ng/ml) added to ce ll-free medium conditioned by 48 h of culture dose dependently stimula ted PC-3 cell proliferation. At concentrations less than or equal to 1 00 ng/ml, its mitogenic action was potentiated when medium had been co nditioned by cells cultured in the presence of plasminogen but inhibit ed when medium had been conditioned by cells cultured in the presence of Pefabloc. We conclude from these results 1) that IGF-II is involved in the autocrine control of PC-3 cell proliferation via the type 1 IG F receptor; and 2) that this proliferation is directly dependent on IG F-II bioavailability that itself is modulated by the limited IGFBP-3 p roteolysis induced, at least in part, by urokinase-type plasminogen ac tivator and plasmin.