Multiple steps are involved in the metastasis of cancer cells from primary sites to distant organs. These steps should be considered in the design of pharmacologic approaches to prevent or inhibit the metastatic process. In the present study, we have compared the effects of inhibiting several steps involved in the bone metastatic process individually with inhibition of both together. The steps we chose were matrix metalloproteinase (MMP) secretion, likely involved in tumor cell invasion, and osteoclastic bone resorption, the final step in the process. We used an experimental model in which inoculation of human estrogen-independent breast cancer MDA-231 cells into the left cardiac ventricle of female nude mice causes osteolytic lesions in bone. To inhibit cancer invasiveness, the tissue inhibitor of the MMP-2 (TIMP-2), which is a natural inhibitor of MMPs, was overexpressed in MDA-231 cells. To inhibit bone resorption, a potent bisphosphonate, ibandronate (4 microg/mouse) was daily administered subcutaneously. Nude mice received either; (a) nontransfected MDA-231 cells; (b) nontransfected MDA231 cells and ibandronate; (c) TIMP-2-transfected MDA-231 cells; or (d) TIMP-2-transfected MDA-231 cells and ibandronate. In mice from group a, radiographs revealed multiple osteolytic lesions. However, in mice from group b or group c, osteolytic lesions were markedly decreased. Of particular note, in animals from group d receiving both ibandronate and TIMP-2-transfected MDA-231 cells, there were no radiologically detectable osteolytic lesions. Survival rate was increased in mice of groups c and d. There was no difference in local enlargement in the mammary fat pad between nontransfected and TIMP-2-transfected MDA-231 cells. These results suggest that inhibition of both MMPs and osteoclastic bone resorption are more efficacious treatment for prevention of osteolytic lesions than either alone, and suggest that when therapies are designed based on the uniqueness of the bone microenvironment and combined with several common steps in the metastatic process, osteolytic bone metastases can be more efficiently and selectively inhibited.