A tissue selective rapamycin analog inhibits MDA-MB-231 breast cancer cell-induced osteolytic bone metastases in nude mice

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Bone metastases, frequent consequences of common malignancies such as breast, lung, and prostate cancer, involve complex molecular processes and are extremely difficult to treat. In the bone microenvironment, metastatic cancer cells produce osteoclast activating factors (e.g., PTHrP) that stimulate bone resorption. Bone-derived growth factors (e.g., TGF-β and IGF1) are subsequently released, promoting cancer-cell proliferation and the amplification of a cycle that produces net osteolytic (bone destructive) consequences. AP24170 is a rapamycin analog with a synthetically linked bone-targeted (BT) moiety. The bone-targeting functionality is designed to accumulate BT rapalogs such as AP24170 on bone, enhancing the anti-tumor and anti-osteolytic effects of mTOR inhibition at bone, and minimizing effects outside bone, compared to non-targeted rapamycin analogs. In this study, we tested this concept by examining the effects of AP24170 and rapamycin in an intracardiac MDA-MB-231 tumor cell injection model of bone osteolytic metastasis in nude mice. Three days post intracardiac cell inoculation, AP24170 and rapamycin were administered intravenously (biw and qw) to mice using equimolar doses (14 and 10 mg/kg, respectively) for a period of 3 weeks. Both AP24170 treated groups had a lower percentage of limb bones with cortical osteolytic lesions (24-21%) and fewer osteolytic lesions per mouse (1 met, av) than the comparable rapamycin treated groups (44-52% and 2-3 mets/mouse, av) as assessed by whole animal radiography. The vehicle treated mice developed osteolytic lesions in 85% of the bones examined and averaged 5 osteolytic limb lesions per mouse. The difference in potency between AP24170 and rapamycin was the opposite of that observed in prior in vitro cell assays, consistent with effective bone-targeting in vivo. In a separate study, we examined systemic mTOR inhibitory effects by administering equimolar doses of each compound to mice, harvesting abdominal muscle 24 hr later, and assessing mTOR activity by measuring levels of phosphorylated S6 (pS6) by western blot. Rapamycin treatment elicited a clear dose-responsive reduction in pS6 levels, as expected, but AP24170 caused no apparent reduction of pS6, suggesting that the inhibitory effects of AP24170 are selective for bone. The tissue (bone) selective nature of AP24170 and the potent in vivo effects observed in this and other models of bone cancer make this class of compounds promising candidates for the treatment of both bone metastases and primary bone cancer.