Signal Transduction Pathways Mediated by Secreted and Non-secreted Forms of intact Insulin-like Growth Factor Binding Protein-3 (IGFBP-3) and its 1-97 N-terminal Fragment in PC-3 Human Prostate Cancer Cells.

Our previous results indicated that both the secreted and the intracellular form of full length and 1-97 N-terminal fragment of IGFBP-3 induces apoptosis in PC-3 human prostate cancer cells in an IGF-dependent and independent manner. This study was undertaken to delineate possible down-stream signaling pathways that are involved in this process. Intact IGFBP-3 and its N-terminal 1-97 fragments with or without a signal pro-peptide was fused to YFP and expressed in PC-3 human prostate cancer cells. In some cases, the putative IGF-binding site present in full length IGFBP-3 and its N-terminal fragment was also mutated. Extent of apoptosis was quantified using FACS. Up-regulation of total Stat-1 and activation of phospho-Stat-1 was shown by western blot. TGF-β signal was measured by luciferase reporter assay. Results from inhibitor studies indicated that both the Caspase 8 and caspase 9 pathways are involved in IGFBP-3 (non-secreted form) induced apoptosis in PC-3 cells. Exogenous addition of IGFBP-3 to PC-3 cells increased Stat-1 protein expression/tyrosine phosphorylation. Interestingly, results also showed that knockdown of Stat-1 by siRNA potentiated the IGFBP-3 induced apoptosis in PC-3 cells. In addition, both full-length IGFBP-3 and its 1-97 N-terminal fragments inhibited TGFβ signaling in these cells. This is the first report that compares the signal transduction pathways involved in apoptotic pathways mediated by IGFBP-3 in PC-3 human prostate cancer cells. Non-secreted form of full length IGFBP-3 and its N-terminal fragments induced apoptosis in PC-3 cells via activation of caspase 8 and caspase 9. We noted that both secreted and non-secreted forms of IGFBP-3 are involved in modulating Stat-1 and TGF-β pathways to induce apoptotic actions in PC-3 cells. Surprisingly, only non-secreted form of IGFBP-3 and its N-terminal fragments are involved in the induction of apoptosis in PC-3 cells via caspase 8 and caspase 9 activation. These studies clearly demonstrate that secreted and non-secreted FL and its 1-97 N-terminal fragments induce apoptosis in PC-3 cells by regulating different mechanistic pathways.