Insulin-like Growth Factor (IGF)-binding Protein-3 Mutants That Do Not Bind IGF-I or IGF-II Stimulate Apoptosis in Human Prostate Cancer Cells

Abstract

The nonbinding 6m-hIGFBP-3 mutant still was able to inhibit DNA synthesis in a mink lung epithelial cell line in which inhibition by wild-type hIGFBP-3 previously had been shown to be exclusively IGF-independent. 6m-hIGFBP-3 only can act by IGF-independent mechanisms since it is unable to form complexes with the IGFs that inhibit their action. We next compared the ability of wild-type and 6m-hIGFBP-3 to stimulate apoptosis in serum-deprived PC-3 human prostate cancer cells. PC-3 cells are known to synthesize and respond to IGF-II, so that IGFBP-3 could potentially act by either IGF-dependent or IGF-independent mechanisms. In fact, 6m-hIGFBP-3 stimulated PC-3 cell death and stimulated apoptosisinduced DNA fragmentation to the same extent and with the same concentration dependence as wild-type hIG-FBP-3. These results indicate that IGF-independent mechanisms are major contributors to IGFBP-3-induced apoptosis in PC-3 cells and may play a wider role in the antiproliferative and antitumorigenic actions of IGFBP-3.

Promotion of Cancer Cell Migration

“…These results suggest that the high affinity binding site of IGFBP-6 for IGFs is similar to those of IGFBP-3 and -5 (5,6,24), and that mIGFBP-6 can be used as a tool for studying IGF-independent actions of IGFBP-6.…”

“…A recent crystal structure of a complex comprising the N-domain of IGFBP-4 and IGF-I confirmed this binding site (4). Substitution of these hydrophobic residues in IGFBP-3 and -5 decreased their IGF-I binding affinities Ͼ1000-fold and diminished their effects on IGF actions (5,6). However, another study showed residual IGF binding after similar mutations in IGFBP-3, suggesting other regions are involved in IGF binding (7).…”

“…1), based on previous structural and mutation studies of IGFBP-3 and IGFBP-5 (5,6,24). Yields of 8 mg/liter (wtIGFBP-6) and 10 mg/liter (mIGFBP-6) were achieved after Ni-NTA affinity purification.…”

Tissue Transglutaminase Has Intrinsic Kinase Activity

“…Interestingly, IGFBP-3 has been shown to be pro-apoptotic in a variety of cell lines (4,6,24). This process is thought to be an IGF-independent effect of IGFBP-3 mediated by binding of IGFBP-3 to a surface receptor (25).…”

“…IGFBP-3 can both enhance and inhibit the effects of IGF-I in vivo and in vitro depending upon experimental conditions (2)(3)(4)(5)(6). Enhancement of IGF-I action may result from enhanced delivery of IGF-I to its plasma membrane receptor, whereas inhibition may be a consequence of competition between IG-FBP-3 and the type 1 IGF receptor for binding of IGF-I (1,2).…”