Cabozantinib, an inhibitor of vascular endothelial growth factor and hepatocyte growth factor signaling, decreases bone lesions in patients with prostate cancer. To determine direct effects of cabozantinib on bone, resorption in neonatal mouse bone organ culture and on gene expression, proliferation, and phenotypic markers in osteoblast and osteoclast cell lines were examined. Cabozantinib, 0.3 and 3 µM, prevented PTHrP-stimulated calcium release from neonatal mouse calvaria. Since the effect on resorption could reflect effects on osteoblasts to prevent osteoclast activation, or direct inhibition of osteoclasts, responses in osteoblastic and osteoclast precursor cell lines were examined. Twenty-four-hour treatment of osteoblastic MC3T3-E1 cells with 3 µM cabozantinib decreased expression of receptor activator of NFkB ligand (RANKL) and alkaline phosphatase. Forty-eight-hour treatment of MC3T3-E1 cells with 3 µM cabozantinib inhibited cell proliferation and decreased MTT activity. Effects on alkaline phosphatase activity were biphasic, with small stimulatory effects at concentrations below 3 µM. When RAW 264.7 osteoclast precursor cells differentiated with 20 ng/ml RANKL were co-treated for 24 h with 3 µM cabozantinib, expression of RANK, TRAP, cathepsin K, alpha v or beta 3 integrin, or NFATc1 were unaffected. Five-day treatment of RANKL-treated RAW 264.7 cells with 3 µM cabozantinib decreased TRAP and MTT activity. The results suggest that the osteoblast could be the initial target, with subsequent direct and indirect effects on osteoclastogenesis leading to decreased resorption. The multiple effects of cabozantinib on the cell microenvironment of bone are consistent with its effectiveness in reducing lesions from prostate cancer metastases. J. Cell. Biochem. 115: 2033–2038, 2014. © 2014 Wiley Periodicals, Inc.