Postsynthetic regulation of insulin-like growth factor-binding protein-3 by MCF-7 human breast cancer cells in culture

Breast cancer cells are exposed to insulin-like growth factors (IGFs) which stimulate their proliferation, and to IGF-binding proteins (IGFBPs) which sequester and modulate IGF action. The primary circulatory IGFBP is IGFBP-3. In the present study, cultured MCF-7 breast cancer cells regulated clearance of IGFBP-3 via both cell association and proteolysis. Exogenously added IGFBP-3 was significantly cleared from the medium over time yielding the formation of smaller sized immunodetected fragments. Clearance was inhibited by IGF-I and -II. In contrast, clearance was not affected by growth factors and an IGF-analog having mitogenic activity but not binding to IGFBPs. In fact, activity of the IGFs and analogs paralleled their degree of binding to the IGFBP, suggesting that the IGF-binding altered IGFBP-3 making it less susceptible to clearance. Qualitatively similar results were obtained when these experiments were conducted using cell-free conditioned medium, thus suggesting the presence of secreted protease(s). However, level of proteolytic activity was much less than that found in the presence of cells. Clearance of rhIGFBP-3 also involved binding to the cell. Disappearance of rhIGFBP-3 was shown to be attenuated by heparin, which blocks cell surface binding sites. In contrast, compounds which block internalization did not inhibit IGFBP-3 clearance.