Sustained inhibition of tumor growth and prolonged survival following sequential administration of doxorubicin and zoledronic acid in a breast cancer model

Abstract

Combination therapy, using agents that target the microenvironment as well as the cancer cells, is common in the treatment of advanced breast cancer. Here, we show that a 6-week course of weekly sequential administration of the cytotoxic drug doxorubicin (2 mg/kg), followed 24 hr later by the antiresorptive agent zoledronic acid (100 lg/kg), causes substantial inhibition of subcutaneous MDA-MB-436 breast tumor growth in immunocompromised mice, leading to significantly increased survival. Tumor growth did not resume following withdrawal of treatment after 6 weeks, with 60% of the animals in this group surviving for more than 160 days. In comparison, animals receiving single-agent therapy all died within 50 days. Molecular analysis of the tumors showed no effect on cell cycle or apoptosis following administration of 100 lg/kg zoledronic acid or 2 mg/kg doxorubicin alone. When doxorubicin was administered 24 hr before zoledronic acid, tumors displayed decreased expression of CYCLINS E1, B, D1 and D3 as well as CDK2, CDC2, CDK4 and CDK7, indicative of cell-cycle inhibition. Tumors from animals receiving sequential treatment also showed induction of both intrinsic-and extrinsic-apoptotic pathways, with increased expression of BAX, decreased expression of BCL-2 and activation of CASPASE 3, 8 and 9. Accumulation of the unprenylated form of RAP1a, a surrogate marker for uptake of zoledronic acid, was only detected in tumors from animals treated with doxorubicin 24 hr before zoledronic acid. Our data are the first to show a sustained antitumor effect in vivo following a limited course of sequential administration of doxorubicin followed by zoledronic acid.

Mechanisms of action of bisphosphonates in oncology: a scientific concept evolving from antiresorptive to anticancer activities

“…It has been proposed that administering chemotherapy before zoledronate provides more effective anticancer activity because there is a higher uptake of the N-BP in tumor cells in vivo when chemotherapy is given first. 5 Results obtained with this sequential treatment are reminescent of those obtained in the neoadjuvant cohort of the AZURE trial, showing that in breast cancer patients receiving neoadjuvant chemotherapy, the addition of zoledronate further reduced the residual invasive tumor size and improved by twofold the complete pathological response, when compared with chemotherapy alone, suggesting an antitumor effect of zoledronate. 3 Targeting the premetastatic niche.…”

“…1,3 Conversely, it has been shown that a sequential treatment with doxorubicin followed 24 h later by zoledronate (used at clinically relevant dosage) reduces subcutaneous growth of MDA-MB-436 breast tumors in animals. 1,5 Importantly, accumulation of unprenylated Rap1A could be only detected in MDA-MB-436 tumors from animals treated with doxorubicin followed by zoledronate, which was indicative of the cellular uptake of zoledronate within subcutaneous tumors. 5 Mechanisms responsible for this synergy between zoledronate and doxorubicin are poorly understood.…”

“…1,5 Importantly, accumulation of unprenylated Rap1A could be only detected in MDA-MB-436 tumors from animals treated with doxorubicin followed by zoledronate, which was indicative of the cellular uptake of zoledronate within subcutaneous tumors. 5 Mechanisms responsible for this synergy between zoledronate and doxorubicin are poorly understood. It has been proposed that administering chemotherapy before zoledronate provides more effective anticancer activity because there is a higher uptake of the N-BP in tumor cells in vivo when chemotherapy is given first.…”

The effects of adding zoledronic acid to neoadjuvant chemotherapy on tumour response: exploratory evidence for direct anti-tumour activity in breast cancer

“…The reduction in subcutaneous tumour growth was accompanied by increased tumour cell apoptosis, reduction in tumour cell proliferation and tumour angiogenesis. The molecular mechanisms behind this sequence-dependent synergy remain to be established, but giving sequenced treatment elicited changes in the expression of a number of genes and proteins associated with cell-cycle progression and apoptosis not occurring with single-agent doxorubicin or ZOL (Ottewell et al, 2010). These data therefore suggest that low serum concentrations of ZOL, in combination with cytotoxic drugs, are sufficient to exert anti-tumour effects in peripheral tissues and lead to the hypothesis that there may be beneficial antitumour effects in patients with early breast cancer.…”

Final 5‐year results of Z‐FAST trial

“…2,3,[13][14][15][16][17] Recently, preclinical and clinical data have also demonstrated anticancer effects of bisphosphonates in breast and other cancer types. [18][19][20][21][22][23][24][25][26][27][28][29][30][31][32] The prevention and treatment of bone loss with upfront (initiated simultaneously with letrozole) versus delayed (initiated with a decrease in T score to <À2 or occurrence of clinical nontraumatic fracture) zoledronic acid in early breast cancer patients receiving letrozole was evaluated in 3 similarly designed, geographically diverse studies (Z-FAST; ZO-FAST; E-ZO-FAST) and in the similarly-designed N03CC study. 15 Interim (<36 months follow-up) results from the former 3 studies indicate that upfront rather than delayed-start zoledronic acid is significantly more effective in preventing bone loss.…”