The protein factor β2-microglobulin (β2M), purified from the conditioned medium of human prostate cancer cell lines, stimulated growth and enhanced osteocalcin (OC) and bone sialoprotein (BSP) gene expression in human prostate cancer cells by activating a cyclic AMP (cAMP)-dependent protein kinase A signaling pathway. When β2M was overexpressed in prostate cancer cells, it induced explosive tumor growth in mouse bone through increased phosphorylated cAMP-responsive element binding protein (CREB) and activated CREB target gene expression, including OC, BSP, cyclin A, cyclin D1, and vascular endothelial growth factor. Interrupting the β2M downstream signaling pathway by injection of the β2M small interfering RNA liposome complex produced an effective regression of previously established prostate tumors in mouse bone through increased apoptosis as shown by immunohistochemistry and activation of caspase-9, caspase-3, and cleavage of poly(ADP-ribose) polymerase. These results suggest that β2M signaling is an attractive new therapeutic target for the treatment of lethal prostate cancer bone metastasis. ©2006 American Association for Cancer Research.