Growth inhibition by insulin-like growth factor-binding protein-3 in T47D breast cancer cells requires transforming growth factor-β (TGF-β) and the type II TGF-β receptor

This study explores the relationship between anti-proliferative signaling by transforming growth factor-p (TGF-beta) and insulin-like growth factor-binding protein-3 (IGFBP-3) in human breast cancer cells. In MCF-7 cells, the expression of recombinant IGFBP-3 inhibited proliferation and sensitized the cells to further inhibition by TGF-beta1. To investigate the mechanism, we used T47D cells that lack type II TGF-P receptor (TGF-beta RII) and are insensitive to TGF-beta1. After introducing the TGF-beta RII by transfection, the basal proliferation rate was significantly decreased. Exogenous TGF-beta1 caused no further growth inhibition, but immunoneutralization of endogenous TGF-PI restored the proliferation rate almost to the control level. The addition of IGFBP-3 did not inhibit the proliferation of control cells but caused dose-dependent inhibition in TGF-beta RII-expressing cells when exogenous TGF-beta1 was also present. Similarly, receptor-expressing cells showed dose-dependent sensitivity to exogenous TGF-beta 1only in the presence of exogenous IGFBP-3. This indicates that in these cells, antiproliferative signaling by exogenous IGFBP-3 requires both the TGF-beta RII and exogenous TGF-beta1. To investigate this synergism, the phosphorylation of TGF-beta signaling intermediates, Smad2 and Smad3, was measured. Phosphorylation of each Smad was stimulated by TGF-beta1 and, independently, by IGFBP-3 with the two agents together showing a cumulative effect. These data suggest that IGFBP-3 inhibitory signaling requires an active TGF-beta signaling pathway and implicate Smad2 and Smad3 in IGFBP-3 signal transduction.