1 alpha, 25-dihydroxy-16-ene-23-yne-26,27-hexafluorocholecalciferol (Ro24-5531) modulation of insulin-like growth factor-binding protein-3 and induction of differentiation and growth arrest in a human osteosarcoma cell line.
1 alpha,25-Dihydroxycholecalciferol [1,25-(OH)2D3] is a potent differentiating agent in a variety of tumor cell lines. However, the induction of severe hypercalcemia has limited its clinical use. Several analogs have been synthesized that retain the antiproliferative differentiating effects of 1,25-(OH)2D3, but do not have the calcitropic effect of the parent compound. One such analog, 1 alpha,25(OH)2-16-ene-23-yne-26,27-hexafluorocholecalciferol (Ro24-5531), can induce differentiation in HL-60 cells and does not induce hypercalcemia in animal models. We, therefore, evaluated the effect of Ro24-5531 on a human osteosarcoma cell line, MG-63. Compared with 1,25-(OH)2D3, the analog Ro24-5531 is 10-100 times more potent as an inhibitor of MG-63 cell proliferation, as determined by [3H]thymidine incorporation and/or 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The inhibition in cell growth is accompanied by a decrease in the expression of p34cdc2 (> 4-fold), a protein critically involved in cell cycle regulation. Ro24-5531 treatment of MG-63, at a concentration of 10(-8) mol/L, induced expression of the bone differentiation markers biglycan and osteocalcin, as determined by Northern analysis. These data suggest that Ro24-5531 treatment induces growth arrest coupled with differentiation. To begin to evaluate the mechanisms by which Ro24-5531 may exert an effect, we evaluated the effect of Ro24-5531 on components of the insulin-like growth factor I (IGF-I) signaling pathway, an important regulator of normal bone growth and differentiation. The expression of IGF-binding protein (IGFBP), IGFBP-3 messenger ribonucleic acid, and protein levels are increased 20-fold after 72 h of treatment with Ro24-5531 and are associated with a marked increase in detectable binding of ligand to binding protein, as measured by RRA. These data suggest an association between Ro24-5531-induced growth arrest and increased expression of IGFBP-3.
Molecular Oncology Section, National Cancer Institute, Bethesda, Maryland 20892, USA.
M C
MC
Velez-Yanguas
T
T
Kalebic
M
M
Maggi
C C
CC
Kappel
J
J
Letterio
M
M
Uskokovic
L J
LJ
Helman
D001846:Bone Development / Q000187:drug effects; D002117:Calcitriol / Q000031:analogs & derivatives* / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000378:metabolism; D012516:Osteosarcoma / Q000473:pathology*; D012333:RNA, Messenger / Q000032:analysis
D018972:Insulin-Like Growth Factor Binding Protein 3; D012333:RNA, Messenger; C086492:Ro 24-5531; D007334:Insulin-Like Growth Factor I; D002117:Calcitriol