Insulin-like growth factor binding protein-3 (IGFBP-3) is one of the six known IGF binding proteins present in human serum and is comprised of 264-amino acid mature protein, with a 27-amino acid signal peptide [1] . IGFBP-3 is expressed in a wide variety of tissues. Epidemiological studies have clearly suggested that there is an inverse relationship between IGFBP-3 levels and occurrence of cancers [2] . Increased expression levels of serum IGFBP-3 result in decreased prevalence of prostate [3] [4] and colorectal [5] [6] cancers. It has also been shown that IGFBP-3 over expression results in decreased tumor formation in xenografts of non-small lung cancer cells [7] and M12 human prostate cancer cells [8] . Patient samples with hepatocellular and non-small cell lung carcinoma have been shown to have decreased expression of IGFBP-3 [9] [10] . Besides, it has also been reported that there is an increased level of IGFBP-3 in senescence [11] . In addition, IGFBP-3 expression level was decreased in cells immortalized with the human papillomavirus type 16 oncoprotein E7 due to proteasomal degradation [12] . Although IGFBP-3 is a secretory protein, but active protein could also be found in nucleus and cytoplasm [13] . Besides its full length form, IGFBP-3 could also be found in an N-terminal truncated form [14] [15] .

IGFBP-3 has been known to play an important role in cell proliferation by inducing its anti-proliferative and pro-apoptotic effects in breast and prostate cancer cells [16] -[18] . It has also been shown that IGFBP-3 not only triggers growth inhibitory effects by inducing apoptosis, but also can function in G1 cell cycle arrest in human breast, kidney and lung cancer cells [19] [20] . IGFBP-3 may also play an important role in mediating inhibitory effects of transforming growth factor (TGF)-β (Figure 1), retinoic acid, tumor necrosis factor (TNF)-α and p53 [16] [21] -[26] , but modulation in IGFBP-3 synthesis or action could regulate these anti-proliferative effects.

IGF dependent studies revealed that Insulin-like growth factor binding protein (IGFBP)-3 could induce apoptosis by binding to IGF-I and form a binary complex with IGF-I and prevent it to activate (IGF-IR) IGF-1 receptor (Figure 2) to stimulate cell proliferation and survival [27] [28] . In addition, Insulin-like growth factor binding protein (IGFBP)-3 has been shown to potently inhibit cell proliferation and induce apoptosis in an insulin-like growth factor (IGF)-independent manner [13] [18] [29] . IGFBP-3 in the media may exert its pro-apoptotic effects by binding to some cell surface receptors resulting in the activation of various signal transduction pathways.

Alternatively, IGFBP-3 may possibly enter into the cell by endocytosis (Figure 3). It has also been reported in some other studies that IGFBP-3 binds to RXRα (Figure 1) and induce its pro-apoptotic effects in PC-3 human prostate cancer cells [30] . Surprisingly, recent studies have shown that IGFBP-3 fail to gain entry or be inter- nalized upon binding to IGF-1 in the extracellular media to induce its pro-apoptotic effects in non-transformed mammary epithelial cells [31] .

IGFBP-3 action results in the activation of various signal transduction pathways [32] and Stat-1 (signal transducer and activator of transcription 1) has been known to have a functional role in IGFBP-3-induced apoptosis (Figure 1) in rat chondroprogenitor cells [33] , although our previous studies [34] showed a protective role of Stat-1 on IGFBP-3 induced apoptosis in PC-3 human prostate cancer cells, implying the fact that role of Stat-1 may be cell type dependent.

Some other downstream signaling molecules of IGFBP-3 have also been reported in various other studies [16] [35] [36] . It has been shown from these studies that IGFBP-3 can bind to transforming growth factor-β (TGF-β) cell surface receptors and one of the studies have shown direct interaction of IGFBP-3 with TGF-β-receptor typeV (TGF-β-RV) in mink lung epithelial cells [37] .

IGFBP-3 has also been known to bind and activate intracellular signaling by forming a hetero-meric complex with other TGF-β receptors (TGF-β RII and TGF-β RI) in T47D breast cancer cells [35] [36] . Besides, addition of IGFBP-3 resulted in the activation of Smad 2 phosphorylation and cell growth inhibition in these cells. It has also been shown that IGFBP-3 mediates its pro-apoptotic effects via TGF-β in PC-3 human prostate cancer cells [16] . Results from our previous studies indicated the inhibition of TGF-β signaling in presence of IGFBP-3 in PC-3 human prostate cancer cells [34] , although we did not study the Smad activation in these cells.

It has also been reported that IGFBP-3 down-regulates Akt activity in human epidermal growth factor receptor-2 (HER-2) over expressed MCF-7 breast cancer cells [38] , in this regard recent studies have shown that IGFBP-3 induces apoptosis in MCF-7 breast cancer cells by inhibiting IGF-I/Akt survival pathway [39] . Previous studies also indicated that IGFBP-3 can bind to a new cell death receptor (IGFBP-3R), a single-span membrane protein which specifically binds to IGFBP-3 but doesn’t bind to other IGFBP’s. In vivo studies using prostate and breast cancer xenografts in athymic nude mice, showed anti-tumor effects of IGFBP-3R [40] . It was shown from the in vitro studies that IGFBP-3R triggers IGFBP-3 induced apoptosis in various cancer cells via a caspase-8 dependent pathway. IGFBP-3R directly interacts and activates caspase-8 in inducing apoptosis and knockdown of caspase-8 expression or activity can lead to inhibition of IGFBP-3/IGFBP-3R induced apoptosis. All these studies clearly indicate that IGFBP-3 induces its anti-proliferative and pro-apoptotic effects either by sequestering IGF-1 to prevent it to bind to Insulin-like growth factor-I receptor (IGF-IR) in an IGF dependent manner or by activating several candidate molecules via signal transduction pathway independent of IGF binding. Although insulin-like growth factor binding protein-3 receptor (IGFBP-3R) has been known to be directly involved in IGFBP-3 action and may prove to be an important target molecule for the treatment of cancer, further studies are still needed to clarify the role of this receptor or other IGFBP-3 cell surface binding protein(s) in IGFBP-3 mediated cell signaling events.

IGFBP-3 is known to activate various downstream signaling molecules. Some of these signaling events could be initiated after binding to its receptor independent of IGF-I binding, to induce anti-proliferative and pro-apoptotic effects in a certain type of cancer cells. On the other hand, IGFBP-3 can also bind to IGF-1 to modulate its ability to bind to IGF-1 receptor to inhibit cell proliferation and cell survival, resulting in cell death.

This work was supported by an intramural grant from NIDDK, NIH.

^*Corresponding author.