Synthetic low-calcaemic vitamin D3 analogues inhibit secretion of insulin-like growth factor II and stimulate production of insulin-like growth factor-binding protein-6 in conjunction with growth suppression of HT-29 colon cancer cells

The aims of the present study were to compare the ability of various synthetic analogues of 1 alpha ,25-dihydroxyvitamin D-3 [1 alpha ,25-(OH)(2)D-3] to inhibit proliferation of HT-29 cells a human colon adenocarcinoma cell line. HT-29 cells were incubated for 144 h with various concentrations (0-100 nM) of 1 alpha ,25-(OH)2D3, or the analogues EB1089. CB1093 or 1 beta ,25-(OH)(2)D-3. All these analogues except 1 beta ,25-(OH)(2)D-3 inhibited cell proliferation, but relative potencies and efficacies of EB1089 and CB1093 were much greater than that of the native vitamin. Cells grew in serum-free medium, reaching a plateau density at day 10 of culture, and addition of 10 nM 1 alpha ,25-(OH)(2)D-3 or 1 beta .25-(OH)(2)D-3 did not alter the long-term growth characteristics of HT-29 cells. However, cells treated with 10 nM EB1089 or CB1093 grew at a rate slower than control and reached final densities that were 53 +/-1 and 36 +/-2% lower than control, respectively. Immunoblot analysis of serum-free conditioned medium using a monoclonal anti-insulin-like growth factor-(IGF)-II antibody showed that both 10 nM EB1089 and CB1093 markedly inhibited secretion of both mature 7500 M-r and higher M-r forms of IGF-II. Ligand blot and immunoblot analyses of conditioned media revealed the presence of IGFBPs of M-r 24.000 (IGFBP-4), 30,000 (glycosylated IGFBP-4), 35,000 (IGFBP-2) and 32,000-34,000 (IGFBP-6). The level of IGFBP-2 was decreased by 42 +/-8 and 49 +/-7% by 10 nM EB 1089 and CB1093, respectively, compared to controls. IGFBP-6 was increased approximately twofold by EB1089 and CB1093, and exogenously added IGFBP-6 inhibited HT-29 cell proliferation. These results suggest that inhibition of HT-29 cell proliferation by EB1089 and CB1093 may be attributed, at least in part, to the decreased secretion of IGF-II. The increase in IGFBP-6 concentration coupled with its high affinity for IGF-II may also contribute to decreased cellular proliferation by an indirect mechanism involving sequestration of endogenously produced IGF-II. (C) 2001 Elsevier Science Ireland Ltd. All rights reserved.