The first bisphosphonate was synthesized in 1865 [16
]. Bisphosphonates are a further development of the pyrophosphates which are well-known in the chemical industry. However, in contrast to pyrophosphates they are not inactivated in vivo
. In 1968, it was shown that bisphosphonates effectively inhibit osteoclast-mediated bone resorption and since then they have become an integral part in the management of benign and cancer-induced bone disease [17
A wealth of preclinical studies demonstrates that bisphosphonates exhibit direct antitumor effects. In vitro
studies have clearly shown that bisphosphonates, in particular Zol, induce tumour cell apoptosis and inhibit tumour cell adhesion, invasion and proliferation and angiogenesis [9
Up to now the antiproliferative and proapoptotic effect of Zol was mainly proven in human breast cancer cell lines. For the first time we have investigated the ex vivo effect of Zol in freshly resected human breast specimen and compared it to the effect of commonly used chemotherapeutic regimen, i.e. FEC and TAC. The results revealed that Zol was at least equal or even superior in its antitumoric effect when compared to both combined regimens. The significance of the found correlation between the IndexSums obtained with FEC, TAC and Zol with T-, N- and ER-status of the tumors remains currently unknown.
Since our experiments have been performed with fresh breast tumor samples which provide only limited quantity of cells after disaggregation we were not able to apply alternative test systems such as MTT or XTT assay to validate the outcome. However, we have tested three breast cancer cell lines in response to the cytotoxic effect of Zol using the ATP-TCA with similar results.
The effector mechanisms of Zol have been investigated by several groups. Their data point towards reversing effects of Zol on cell proliferation and upregulation of pro-apoptotic combined with downregulation of anti-apoptotic proteins and survival factors [19
]. The mevalonate pathway which is a target of Zol may be important for these observations [20
These and our results indicate that Zol and probably all bisphosphonates may exert a direct antitumoric effect on breast cancer cells beyond their antimetastatic effect on bone matrix. Thus the use of bisphosphonate treatment as adjuvant therapy in women with breast cancer may have clinical significance.
This hypothesis is supported by some animal and clinical studies investigating the use of Zol beyond the action on cancer-induced bone loss. In a mouse model Zol demonstrated a significant inhibition of breast cancer tumor growth [17
]. Further in vivo
studies revealed synergistic antiproliferative effects of Zol in combination with doxorubicin [20
], docetaxel [21
] as well as paclitaxel [23
Intermittent addition of Zol can significantly reduce serum levels of vascular endothelial growth factor (VEGF) as compared to basal levels [24
In ABCSG-12 the addition of Zol to endocrine therapy was associated with a significant 36% improvement in disease free survival (DFS) compared with endocrine treatment alone, the median follow-up was 48
]. Furthermore, antitumour benefits of zoledronic acid were observed outside bone.
Data from several pilot studies show that Zol potentially clears disseminated tumour cells (DTCs) from the bone marrow of patients with early breast cancer [26
] or in the neoadjuvant setting with locally advanced breast cancer [29
]. The presence of DTCs in the bone marrow is a recognized risk factor for the subsequent development of bone metastases, but longer term data are required to establish the clinical relevance of these findings.
The first subgroup analysis (n
205) of the AZURE-study (Does Adjuvant Zoledronic Acid RedUce Recurrence in Breast Cancer) of patients with neoadjuvant chemotherapy revealed a synergistic effect of the combination of chemotherapy with Zol on primary tumor size [15
]. The parameter residual invasive tumor size
(RITS) was significantly reduced by 43% (CT alone 27.4
mm vs. CT + Zol 15.5
0,006) and pathological complete response rate
improved by 69% (CT alone 6.9% vs. CT + Zol 11.7%; p
0.146), which, however, was not statistically significant.
The mechanism(s) by which Zol may trigger a direct antitumoric action is currently not known. It inhibits farnesyl diphosphate synthase within the mevalonate pathway and, through this mechanism, is a potent inhibitor of osteoclast-mediated bone resorption. The mevalonate pathway is known to be probably involved in cancer development.
Our data for the first time suggest a positive correlation between the efficacy of Zol and the ER status. A possible association between the efficacy of Zol and the presence of ER on inhibition of tumor cell growth could be shown in two in vitro
experiments that revealed that ER-negative cell lines are inhibited in their growth by Zol worse than ER-positive cell lines [10
]. However, in other experimental studies the efficacy of Zol was similar in ER-negative and ER-positive cell lines [20
] or even more pronounced in the ER-negative cell line [30
Interestingly, in the Women’s Health Initiative trial on bisphosphonate use and breast cancer incidence
a significant lower incidence of about 30% of ER-positive breast cancers was found in bisphosphonate users than in nonusers [31
]. The mechanism(s) of a possible dependence of Zol from ER status remains unknown so far. Hints might be derived from prostate cancer, since in the prostate a dysregulated cellular growth is mediated by inhibiting the rate-limiting pathway step in cholesterol synthesis, thereby decreasing isoprenylate intermediates, decreasing cholesterol rich cellular membrane domains, and down-regulating androgen and estrogen receptors [32
]. We therefore speculate that expression of ER might point towards high activity of cholesterol synthesis pathway and therefore high effect of blocking the mevalonate pathway by Zol. Further research is necessary to reveal this topic in more detail.
The Zol concentrations used in the present study were in the range of 0.625 to 20
μg/ml and the IC50 value was 1
μg/ml. The physiological plasma concentrations (Cmax) after infusion of Zol are in the range of 0.2 to 0.4
μg/ml within 24
]. Thus the effective in vitro Zol concentrations were comparable to in vivo plasma concentrations.