Insulin-like growth factors (IGF), polypeptides that regulate growth, differentiation, and survival in cells and tissues, were found to enhance gene expression from both heterologous and homologous promoters in the presence of constitutively active STAT5. This highly conserved 700-bp DNA region contains two closely located consensus STAT5-binding sites. Hypoxia regulates the IGF-1 gene expression through the STAT5b. We confirmed STAT5b is up-regulated under hypoxic conditions, and the increased STAT5b binds strongly to the STAT5-binding sites 1 and 2 contained within the distal 5'-flanking region of IGF-1 gene in HepG2 cells. EMSA studies showed that STAT5-binding activities to the IGF-1 promoter distinctly increased under hypoxia in STAT5b-transfected COS-7 cells. The IGF-1 gene expression was also increased by hypoxia in HepG2 cells. STAT5b expression was inhibited by siRNA experiments leading to decreased IGF-1. These results provide a basis of molecular targets for cancer treatment via the STAT5b-IGF-1 pathway in solid tumor cells.