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Insulin-like Growth Factor Binding Protein (IGFBP)-3 Protease Activity Secreted by MCF-7 Breast Cancer Cells: Inhibition by IGFs Does Not Require IGF-IGFBP Interaction1
The proliferative action of insulin-like growth factors (IGFs) on breast cancer cells is regulated by IGF binding proteins (IGFBPs). This study characterizes the proteolysis of IGFBP-3 by an enzyme secreted by MCF-7 human breast cancer cells. Proteolysis of IGFBP-3 by incubation at 37 C with serum-f...
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Published in: | Endocrinology (Philadelphia) 1997-04, Vol.138 (4), p.1683-1690 |
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Main Authors: | , , |
Format: | Article |
Language: | English |
Online Access: | Get full text |
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Summary: | The proliferative action of insulin-like growth factors (IGFs) on
breast cancer cells is regulated by IGF binding proteins (IGFBPs). This
study characterizes the proteolysis of IGFBP-3 by an enzyme secreted by
MCF-7 human breast cancer cells. Proteolysis of IGFBP-3 by incubation
at 37 C with serum-free medium from MCF-7 cells was maximal at pH
5.0–5.5, with no activity detected below pH 4.5. This enzyme activity
resulted in the disappearance of the 40- to 45- and 30-kDa bands of
pure plasma-derived IGFBP-3, detectable by immunoblotting after
SDS-PAGE, and the appearance of a single 21-kDa immunoreactive species.
The 21-kDa protein did not bind IGF-I or IGF-II by ligand blotting. The
enzyme activity appeared at 25- to 30-kDa by gel chromatography at pH
6.5 and was inhibited by EDTA and leupeptin, an inhibitor of cysteine
and serine proteases, but not by the serine protease inhibitors
aprotinin and benzamidine. IGFBP-3 protease activity was inhibited
in medium conditioned by cells incubated with 50 ng/ml IGF-I. A similar
inhibitory effect was seen under cell-free conditions by adding IGF-I
to medium harvested from cells incubated without IGFs. The cell-free
inhibition of IGFBP-3 proteolysis by IGFs did not require IGF
interaction with the binding protein, because [long
Arg3]IGF-I, which binds to IGFBP-3 with less than 0.2% of
the potency of IGF-I, inhibited IGFBP-3 proteolysis with 20% of the
potency of IGF-I. These results suggest that IGFs may regulate their
own activity in breast cancer cells, preventing IGFBP-3 proteolysis by
a mechanism that is not receptor mediated and does not require
IGF-IGFBP interaction. |
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ISSN: | 0013-7227 1945-7170 |
DOI: | 10.1210/endo.138.4.5064 |