Expression of insulin-like growth factor-binding protein 6 complementary DNA alters neuroblastoma cell growth

Insulin-like growth factors I and II (IGF-I and IGF-II) are actively involved in neuroblastoma cell growth. In all biological fluids, they are noncovalently bound to high-affinity binding proteins. At least six species of these IGF-binding proteins (IGFBPs) have been identified, but their precise roles remain unclear. One of them, IGFBP-6, is produced by neuroblastoma cells in culture under certain experimental conditions and seems to be associated with the arrest of cell growth. We stably transfected IGR-N-91 and SK-N-SH neuroblastoma cells with an expression vector comprising IGFBP-6 cDNA, whose expression was placed under the control of the constitutive and ubiquitous cytomegalovirus promoter. Analyses of the cell cycle (flux cytofluorometry), mitogenic activity (radiolabeled thymidine incorporation), and the number of viable cells (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide test) showed that the mitogenic effects of serum, IGF-I, IGF-II, and des (1-3) IGF-I, a truncated IGF-I analogue with no affinity for IGFBP-6, were depressed in both transfected cell lines. With s.c. injection of transfected IGR-N-91 cells into nude mice, tumors developed in only 50-70% of cases, 1 or 2 weeks after those in controls, and were 60-90% smaller. Our findings show that IGFBP-6 influences neuroblastoma cell growth, both in vitro and in experimental xenograft development.