Breast cancer is the most frequent malignancy in women and often leads to osteolytic bone metastases. Amino-bisphosphonates are anti-resorptive drugs and currently used as adjuvant therapy for affected patients. Similarly to statins, they inhibit the mevalonate pathway, which is essential for the farnesylation and geranylgeranylation of proteins. Here, we demonstrate that this inhibition by atorvastatin and zoledronic acid evokes synergistic anti-tumour effects in human breast cancer cells. Human triple-negative MDA-MB-231 breast cancer cells were treated with 1 μM atorvastatin and 10 μM zoledronic acid. Whereas individual treatments only elicited weak effects, the combination induced a three-fold increase of caspases 3/7 activation and DNA fragmentation as well as a decrease of cell vitality by 60%. The synergistic effects were underlined by the induction of poly ADP ribose polymerase (PARP) cleavage, the reduced phosphorylation of the survival mediator protein kinase B, and a significant reduction of the anti-apoptotic genes B-cell lymphoma 2 (BCL-2) and Survivin (SVV). Furthermore, the treatments significantly reduced the expression and secretion of interleukins 6, 8 and 11 as well as of Dickkopf-1 representing major players in osteolytic bone metastases. Finally, supplementing the cells with either farnesylpyrophosphate (FPP) or geranylgeranylpyrophosphate (GGPP) upon the combination treatment clearly revealed that only GGPP completely abrogated the synergistic anti-tumour effects. These observations show that geranylgeranylation is indispensable for the survival of MDA-MB-231 cells. As functional small Rho-GTPases are geranylated proteins, we assumed an inactivation by the combination treatment. Surprisingly, a concomitant treatment with a Rho-GTPases activator led to even higher anti-tumour effects. In line, a pull down assay revealed an accumulation of the active GTP-bound forms of the Rho GTPases RhoA and CDC42 pointing to a possible anti-tumour mechanism. Our results indicate the synergistic use of mevalonate pathway inhibitors as a new attractive therapeutic approach for the treatment of breast cancer-induced bone metastases.
Disclosure: The authors declared no competing interests. This work was supported by grants from the Deutsche Forschungsgemeinschaft (RA 2151/1-1 and RA 2151/2-1).