RECOMBINANT HUMAN INSULIN-LIKE GROWTH-FACTOR (IGF) BINDING PROTEIN-3 STIMULATES PROSTATE CARCINOMA CELL-PROLIFERATION VIA AN IGF-DEPENDENT MECHANISM - ROLE OF SERINE PROTEASES

Insulin-like growth factor-binding proteins (IGFBPs) modulate IGF acti on at cellular level, through either inhibition or potentiation, and t hey also have intrinsic activity that is independent of their binding to IGFs. In prostate carcinoma (PC-3) cells, which are capable of grow th for several days in serum-free medium, non-glycosylated recombinant human IGFBP-3 (rhIGFBP-3) had a biphasic mitogenic effect, stimulatio n being dose-dependent up to 20 ng/ml, followed by progressive depress ion down to zero stimulation at 150-200 ng/ml. This mitogenic effect w as not intrinsic activity, but involved IGF-II secreted by the cells, since stimulation was abolished in the presence of anti-type 1 IGF rec eptor antibody (alpha IR-3). Western ligand- and immunoblot analysis o f the culture media revealed several IGFBP species, in particular IGFB P-3 which exhibited an electrophoretic profile characteristic of limit ed proteolysis. The amounts of the proteolytic fragments increased in parallel with the concentrations of added rhIGFBP-3, but a large amoun t of intact protein remained at the highest concentrations added. When a serine protease inhibitor, 4-(2-aminoethyl)-benzenesulphonyl fluori de (Pefabloc SC), was added at concentrations demonstrated to be non-t oxic to the cells, IGFBP-3 proteolysis was diminished and rhIGFBP-3-in duced stimulation of proliferation was suppressed. Conversely, in the presence of plasminogen transformed to plasmin by urokinase secreted b y the cells, proliferation stimulated by rhIGFBP-3 and its proteolysis were enhanced. Our results suggest that the biphasic mitogenic effect of rhIGFBP-3 on PC-3 cells reflects changes in the availability to th e cells of the IGF-II they secrete, This availability depends on the e xtent of IGFBP-3 proteolysis (which promotes release of bound IGF-II) and on the proportion of intact forms (which sequestrate secreted IGF- II).