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The Oncologist, Vol. 9, Suppl 4, 3–13, September 2004
© 2004 AlphaMed Press

Bisphosphonates: Preclinical Review

Jonathan R. Green

Novartis Pharma AG, Basel, Switzerland

Correspondence: Correspondence: Jonathan R. Green, Ph.D., Novartis Pharma AG, Klybeckstrasse 141, WKL-125.901, CH-4002 Basel, Switzerland. Telephone: 41-61-696-4415; Fax: 41-61-696-3849; e-mail: jonathan.green{at}pharma.novartis.com


   LEARNING OBJECTIVES
Top
 Learning Objectives
Abstract
 Introduction
 Mechanism of Action
 Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
 Conclusions and Future...
 References
 
After completing this course, the reader will be able to:

  1. Describe the mechanism of action of first-generation and nitrogen-containing bisphosphonates.
  2. Explain how the mechanism of action of the bisphosphonates might directly affect tumor growth.
  3. Discuss how the bisphosphonates might be incorporated into both the prevention and treatment of cancer.

Access and take the CME test online and receive 1 hour of AMA PRA category 1 credit at CME.TheOncologist.com


   ABSTRACT
Top
 Learning Objectives
 Abstract
Introduction
 Mechanism of Action
 Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
 Conclusions and Future...
 References
 
Bisphosphonates effectively inhibit osteoclast-mediated bone resorption and are integral in the treatment of benign and malignant bone diseases. The evolution of bisphosphonates over the past 30 years has led to the development of nitrogen-containing bisphosphonates (N-BPs), which have a mechanism of action different from that of the nonnitrogen-containing bisphosphonates. Studies conducted over the past decade have elucidated the mechanism of action and pharmacologic properties of the N-BPs. N-BPs exert their effects on osteoclasts and tumor cells by inhibiting a key enzyme in the mevalonate pathway, farnesyl diphosphate synthase, thus preventing protein prenylation and activation of intracellular signaling proteins such as Ras. Recent evidence suggests that N-BPs also induce production of a unique adenosine triphosphate analogue (Apppi) that can directly induce apoptosis. Our increased understanding of the pharmacologic effects of bisphosphonates is shedding light on the mechanisms by which they exert antitumor effects. As a result of their biochemical effects on protein prenylation, N-BPs induce caspase-dependent apoptosis, inhibit matrix metalloproteinase activity, and downregulate {alpha}vß3 and {alpha}vß5 integrins. In addition, zoledronic acid (Zometa®; Novartis Pharmaceuticals Corp.; East Hanover, NJ and Basel, Switzerland) exerts synergistic antitumor activity when combined with other anticancer agents. Zoledronic acid also inhibits tumor cell adhesion to the extracellular matrix and invasion through MatrigelTM and has antiangiogenic activity. A growing body of evidence from animal models demonstrates that zoledronic acid and other bisphosphonates can reduce skeletal tumor burden and prevent metastasis to bone. Further studies are needed to fully elucidate these biochemical mechanisms and to determine if the antitumor potential of bisphosphonates translates to the clinical setting.

Key Words. Bisphosphonates • Zoledronic acid • Bone resorption • Apoptosis • Antitumor effects • Mevalonate pathway


   INTRODUCTION
Top
 Learning Objectives
 Abstract
 Introduction
Mechanism of Action
 Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
 Conclusions and Future...
 References
 
Bisphosphonates are ideally suited for the treatment of metabolic bone disease because they bind avidly to the bone mineral at sites of active bone metabolism, where they achieve therapeutic concentrations. The pioneering work of Fleisch and colleagues showed that bisphosphonates not only inhibit dissolution of hydroxyapatite crystals, but also affect osteoclast metabolism and function [1–3]. Bisphosphonates are released during bone resorption and are internalized by osteoclasts, leading to inhibition of bone resorption and induction of osteoclast apoptosis [4–6].

There is now extensive preclinical evidence that bisphosphonates also have antitumor activity, as evidenced by reduced proliferation and viability of tumor cell lines in vitro and reduced skeletal tumor burden and slower progression of bone lesions in animal models. Several mechanisms have been proposed to explain these observations. Bisphosphonates may render the bone a less favorable microenvironment for tumor cell growth by reducing tumor-induced osteolysis and local release of growth factors, and bisphosphonates may also have direct antitumor effects. Bisphosphonates inhibit proliferation and induce apoptosis of a variety of human tumor cell lines in vitro [7–17]. Bisphosphonates also inhibit tumor cell adhesion to the extracellular bone matrix, inhibit invasion of tumor cells through MatrigelTM, and have antiangiogenic and immunomodulatory activities. Consistent with these findings, bisphosphonates have been shown to inhibit the formation or progression of bone metastases and/or reduce skeletal tumor burden in a variety of animal models. The mechanisms responsible for the observed antitumor effects of bisphosphonates are beginning to be elucidated.


   MECHANISM OF ACTION
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanism of Action
Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
 Conclusions and Future...
 References
 
Bisphosphonates accumulate in the mineralized bone matrix and are released during bone resorption. First-generation, nonnitrogen-containing bisphosphonates are metabolized by osteoclasts to nonhydrolyzable cytotoxic ATP analogues [18–21]. For example, clodronate is metabolized to AppCC12p, which, at high concentrations, inhibits mitochondrial ATP/adenosine diphosphate (ADP) translocase, thereby causing loss of the mitochondrial membrane potential and direct induction of apoptosis [22–24]. The high affinity of bisphosphonates for bone mineral and subsequent uptake by activated osteoclasts during bone resorption ensures that cytotoxic concentrations of these metabolites only accumulate within osteoclasts. However, they may also accumulate within tumor cells growing in the bone because tumor cells stimulate osteolysis.

In contrast, nitrogen-containing bisphosphonates (N-BPs), which include pamidronate (Aredia®; Novartis Pharmaceuticals Corp.; East Hanover, NJ), alendronate (Fosamax®; Merck and Company, Inc.; West Point, PA), ibandronate (Bondronat®; Hoffmann-La Roche Inc.; Nutley, NJ), risedronate (Actonel®; Proctor and Gamble Pharmaceuticals, Inc.; Cincinnati, OH), and zoledronic acid (Zometa®; Novartis Pharmaceuticals Corp.), affect osteoclast activity and survival through a different mechanism. After internalization, these compounds inhibit a key enzyme, farnesyl diphosphonate (FPP) synthase, in the biosynthetic mevalonate pathway (Fig. 1Go) [5, 25–28]. As a result, N-BPs interfere with a variety of cellular functions essential for the bone-resorbing activity and survival of osteoclasts [28, 29]. Several intermediates in this pathway, including farnesyl pyrophosphate and geranylgeranyl pyrophosphate, are required for the posttranslational modification (i.e., prenylation) of guanosine triphosphate-binding proteins such as Ras, Rho, and Rac [30]. These signaling molecules are involved in the regulation of cell proliferation, cell survival, and cytoskeletal organization [26, 28, 31, 32]. In particular, inhibition of protein prenylation and Ras signaling within osteoclasts leads to defects in intracellular vesicle transport [33]. As a result, osteoclasts cannot form a tight-sealing zone or ruffled borders, which are required for bone resorption.



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  		Figure 1. Schematic representation of the mevalonate pathway and the effects of nitrogen-containing bisphosphonates. Abbreviation: HMG CoA = 3-hydroxy-3-methylglutaryl coenzymeA. Adapted with permission from Gober et al. [25].

 
Recent studies have shown that N-BPs may have yet another mechanism of action. As shown in Figure 1Go [25, 34], N-BPs also induce production of an intracellular ATP analogue known as Apppi (triphosphoric acid 1-adenosin-5'-yl ester 3-[3-methylbut-3-enyl] ester), which may directly induce apoptosis similar to AppCC12p (i.e., a metabolite of clodronate) [34]. Inhibition of FPP synthase results in accumulation of isopentenyl diphosphonate, which can be metabolized to Apppi. The ability of various bisphosphonates (at 0.1-µM concentrations) to inhibit recombinant human FPP synthase activity (Fig. 2Go) [35] correlates well with their ability to induce production of Apppi in J774 cells. In this assay, clodronate serves as a negative control with little or no effect on FPP synthase activity, and it induces no Apppi production. Of the N-BPs tested, zoledronic acid demonstrated the highest potency in terms of both inhibition of FPP synthase activity (Fig. 2Go) [35] and production of Apppi [34]. The implication of this work is that N-BPs, by inhibiting FPP synthase, can potentially induce apoptosis of osteoclasts and tumor cells by at least two distinct pathways.



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  		Figure 2. The effects of equimolar concentrations (0.1 µm) of bisphosphonates on recombinant human FPP synthase activity. *p < 0.001. Adapted with permission from Dunford et al. [35].

 
It has been demonstrated that the potency of various N-BPs with respect to inhibition of FPP synthase activity also correlates well with their observed potency in terms of inhibiting bone resorption in vitro (Table 1Go) [35, 36]. Among the N-BPs tested, zoledronic acid was the most potent inhibitor of FPP synthase activity, followed by risedronate and ibandronate. The relative potencies of these three N-BPs in the FPP synthase assay correlate closely with their relative potencies in terms of inhibiting vitamin D3-induced calcium release from mouse calvaria cultures [36]. These data suggest that inhibition of FPP synthase is a central mechanism by which N-BPs inhibit osteoclast-mediated bone resorption. Moreover, osteoclasts isolated from animals treated with N-BPs show a profound suppression of FPP synthase activity [37]. Therefore, FPP synthase appears to be one of the key molecular targets of N-BPs. Inhibition of protein prenylation in cancer cells is also thought to be responsible for many of the observed antitumor effects of N-BPs.


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  		Table 1. Relative potencies of bisphosphonates with respect to inhibition of FPP synthase and bone resorption activity
 

   EVIDENCE OF ANTITUMOR EFFECTS
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanism of Action
 Evidence of Antitumor Effects
Mechanisms of Antitumor Effects
 Conclusions and Future...
 References
 
In Vitro Studies
Bisphosphonates have demonstrated direct antitumor activity against a variety of tumor cell lines at concentrations ranging from 5–2,000 µM [38]. Bisphosphonates cause dose- and time-dependent inhibition of proliferation and induce apoptosis of myeloma, breast cancer, prostate cancer, pancreatic cancer, lung cancer, and osteosarcoma cell lines in vitro [7-17, 39]. For example, in studies with the human MDA-MB-231 breast cancer cell line, clodronate, pamidronate, and zoledronic acid demonstrated dose-dependent effects on cell viability, with 50% inhibitory concentration (IC50) values of 700, 40, and 15 µM, respectively [12]. This effect was not caused by calcium chelation and appeared to be specific to tumor cells. Similar results have been observed with other breast cancer cell lines. For instance, dose-dependent increases in the proportion of apoptotic cells have been observed when Hs 578T cells were incubated with zoledronic acid [12]. Studies with MCF-7 cells have further shown that the effects of zoledronic acid on tumor cell viability and apoptosis could be reversed by incubation with geranylgeraniol, indicating that inhibition of protein prenylation can induce tumor cell apoptosis [39]. In vitro studies with several different prostate cancer cell lines, including PC-3 and LNCaP 23.1, have shown that zoledronic acid inhibits proliferation, induces apoptosis, and causes cell-cycle arrest in a dose-dependent manner [40, 41]. These are just a few examples of the antitumor activity of bisphosphonates against a wide variety of human and murine tumor cell lines, and in every instance where several bisphosphonates have been tested, zoledronic acid has demonstrated the most potent activity.

In vitro studies have also shown that combining N-BPs with a variety of standard anticancer agents results in additive or synergistic antitumor effects against a range of tumor cell lines [39, 42–49]. Jagdev et al. [39] were the first to show that the combination of zoledronic acid (10 µM) plus paclitaxel (Taxol®; Bristol-Myers Squibb; Princeton, NJ; 2 nM) enhanced apoptosis of MCF-7 breast cancer cells fourfold compared with either agent alone. Similar results were reported when ibandronate or zoledronic acid were combined with epirubicin (Ellence®; Pharmacia and Upjohn; Portage, MI) plus docetaxel (Taxotere®; Aventis Pharmaceuticals Inc.; Bridgewater, NJ) [50]. In cultures of primary breast cancer cells, the concentrations of epirubicin plus docetaxel were gradually reduced to suboptimal levels by serial dilution until there was little or no inhibition of tumor cell growth with these two drugs alone. However, the addition of 15 µM ibandronate or zoledronic acid resulted in 35% and 70% inhibitions of tumor cell growth, respectively. Combinations of zoledronic acid with taxanes also have demonstrated synergistic antitumor activity against prostate cancer cell lines. The combination of zoledronic acid (12.5 or 25 µM) with suboptimal concentrations of docetaxel (≤0.1 ng/ml) demonstrated additive and dose-dependent cytotoxic effects on PC-3 prostate cancer cells at 72 hours [45]. In addition, recent studies with DU-145 prostate cancer and MCF-18 breast cancer cell lines have demonstrated that the combination of zoledronic acid with the cyclooxygenase-2 inhibitor SC236 had additive inhibitory effects on tumor cell growth [48, 49]. These and many other studies have shown that N-BPs can enhance the cytotoxic and cytostatic effects of standard anticancer agents used to treat a variety of solid tumors. Recently, zoledronic acid also has been shown to possess antileukemic activity against a Philadelphia chromosome-positive cell line, and the combination of zoledronic acid with imatinib mesylate (Gleevec®; Novartis Pharmaceuticals Corp.) demonstrated synergistic antileukemic activity [51]. These findings are intriguing given that Bcr-Abl stimulates the Ras signaling pathway.

Two recent studies with breast cancer cell lines have begun to shed light on the possible mechanisms underlying the observed synergy between N-BPs and anticancer agents. In the first of these studies, MCF-7 breast cancer cells were incubated with zoledronic acid (25 µM) and doxorubicin (Adriamycin®; Pharmacia and Upjohn; Peapack, NJ; 0.05 µM), and the effects of different sequences and incubation periods were tested [43]. The only combination that resulted in synergistic enhancement of apoptosis was doxorubicin for 24 hours followed by zoledronic acid for 1 hour, whereas zoledronic acid administered either before doxorubicin or together with doxorubicin did not increase apoptosis (Fig. 3Go) [43]. This suggests that exposure to doxorubicin at a concentration that was not cytotoxic sensitized tumor cells to the cytotoxic effects of zoledronic acid. A similar but opposite effect of sequencing was also reported by the same group when zoledronic acid (25 µM) was combined with tumor necrosis factor (TNF)-related apoptosis inducing ligand (TRAIL) at a concentration of 10 ng/ml. Among five combinations tested, the only combination that produced synergistic apoptotic effects was zoledronic acid for 48 hours followed by TRAIL for 24 hours. This combination yielded 14.7% apoptotic cells compared with 0.7% for zoledronic acid alone and 2.7% for TRAIL alone (p < 0.001). The intriguing results of these in vitro studies suggest a novel approach to enhance the synergy between N-BPs and chemotherapeutic agents in the clinical setting.



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  		Figure 3. Synergistic apoptotic effect of zoledronic acid (ZOL) in combination with doxorubicin (DOX) on MCF-7 breast cancer cells in vitro. Adapted with permission from Neville-Webbe et al. [43].

 
Animal Models
These in vitro findings are supported by data from many animal models showing that the newer N-BPs can significantly reduce the number and size of osteolytic lesions in tumor-bearing mice, reduce skeletal tumor burden, induce tumor cell apoptosis in bone lesions, reduce serum levels of tumor markers, and prevent formation of bone metastases (Table 2Go) [9, 40, 52–60]. Some models have even shown effects on visceral tumors and an improvement in survival of tumor-bearing mice, but these findings have been less consistently observed. Animal studies have focused on models of multiple myeloma, breast cancer, and prostate cancer. Using radiographic, histologic, and histomorphometric techniques, these studies have shown that N-BPs can inhibit the formation or progression of bone metastases and/or reduce skeletal tumor burden. Bisphosphonates have been administered either at the time of tumor cell inoculation (i.e., prevention setting) or after bone metastases are established (i.e., treatment setting).


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  		Table 2. Animal models demonstrating antitumor effects of zoledronic acid
 
Several models of multiple myeloma have been established and have consistently shown reductions in tumor-induced osteolysis and skeletal tumor burden by N-BPs [52, 53, 59]. For example, in the 5T2 model of murine myeloma, treatment with zoledronic acid (120 µg/kg s.c. twice weekly) after osteolytic lesions were established significantly reduced development of new osteolytic lesions, reduced the bone surface occupied by osteoclasts and inhibited tumor-induced osteolysis, and reduced skeletal tumor burden as evidenced by serum paraprotein levels [53]. Moreover, treatment with zoledronic acid at the first sign of circulating paraprotein significantly prolonged disease-free survival in those mice.

Numerous studies in breast cancer models have also been reported. Zoledronic acid was shown to inhibit progression of established bone metastases and development of new bone metastases in two models of breast cancer [54, 55]. In nude mice injected with human MDA-MB-231 breast cancer cells and allowed to develop osteolytic lesions, mice treated with zoledronic acid (0.2, 1.0, or 5.0 µg/day s.c.) had significantly less radiographic bone lesion area, by >80%, than controls [54]. Ibandronate (1.0 µg/day) and alendronate (10 µg/day) resulted in nonsignificant reductions in bone lesion area (65% and 55% reductions, respectively). These findings were confirmed in another independent study using highly sensitive in vivo imaging of MDA-MB-231 cells genetically engineered to express green fluorescent protein [55]. Similar studies have been conducted with ibandronate in nude mice bearing MDA-MB-231 breast cancer cells, which typically form both adrenal and bone metastases. Treatment with ibandronate (4 µg/day s.c.) inhibited the radiographic progression of established osteolytic lesions and decreased skeletal tumor burden compared with controls [9]. However, administration of ibandronate at the time of tumor cell inoculation resulted in a twofold increase in adrenal tumor load [61]. In a murine breast cancer model, treatment with zoledronic acid (5 µg/day) for 7 days after injection of 4T1 murine mammary tumor cells (i.e., prevention setting) markedly decreased the formation of new bone metastases at day 28 [54]. The observed decrease in radiographic bone lesion area was accompanied by an increase in the number of apoptotic osteoclasts and apoptotic tumor cells in bone lesions.

Notably, the 4T1 mammary tumor model has provided the first in vivo evidence of synergy between zoledronic acid and chemotherapy. In animals with established orthotopic tumors, the combination of zoledronic acid (250 µg/kg) with 20 mg/kg/day of oral UFT (a combination of uracil and tegafur [4:1 molar ratio]) reduced skeletal tumor burden more effectively than either agent alone [56]. Similarly, the combination of ibandronate with doxorubicin (150 µg/day) has been investigated in the MDA-MB-231 model and was shown to have additive antitumor effects in bone [61].

Studies in a prostate cancer model have also recently been reported. In those studies PC-3 and LuCaP 23.1 cells were injected directly into the tibia of mice [40]; PC-3 cells form osteolytic lesions, and LuCaP 23.1 cells form osteoblastic lesions. The treatment group received zoledronic acid (5 µg s.c. twice weekly) either at the time of tumor cell injection or after tibial tumors were established (7 days for PC-3 tumors and 33 days for LuCaP 23.1 tumors). Treatment with zoledronic acid significantly inhibited growth of both osteolytic and osteoblastic metastases by radiographic analysis (Fig. 4Go) [40] and also reduced skeletal tumor burden as evidenced by a significant decrease in serum levels of prostate-specific antigen in animals bearing LuCaP 23.1 tumors [40]. The observed reduction in serum prostate-specific antigen levels provides compelling direct evidence of the antitumor activity of zoledronic acid in this animal model.



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  		Figure 4. The effects of zoledronic acid (5 µg s.c. twice weekly) on tumor volume (assessed radiographically) in mice with tibial tumors from PC-3 and LuCaP 23.1 prostate cancer cell lines. Zoledronic acid was administered either at the time of tumor cell injection (i.e., prevention) or after tibial tumors were established (i.e., treatment), which was 7 days postinjection for PC-3 and 33 days postinjection for LuCaP. *p < 0.03; **p < 0.001. Adapted with permission from Corey et al. [40].

 
The potential of zoledronic acid to prevent bone metastasis was also demonstrated in an animal model of prostate cancer [62]. In that model, mice were injected with PC-3 cells, and the incidence of bone metastases was studied in normal mice and in mice that were rendered androgen deficient by surgical castration. Androgen-deficient mice developed significantly more bone metastases than did intact control mice. This strongly suggests that excess bone resorption caused by androgen ablation can stimulate metastasis of PC-3 cells to the bone. This is consistent with current hypotheses of tumor cell metastasis to bone. Moreover, daily treatment with zoledronic acid significantly reduced the incidence of bone metastases in both normal and castrated mice.

Although the majority of animal models suggest that the primary antitumor effect of bisphosphonates is manifested in the bone, where they reach the highest concentrations, preliminary data from the 4T1 mammary tumor model have demonstrated that zoledronic acid can also inhibit visceral metastases [57]. 4T1 cells expressing luciferase were used to quantitate tumor burden in mice. After tumor cell inoculation, mice were treated with zoledronic acid (5.0 µg every 4 days), which resulted in lower tumor burdens than in controls not only in bone but also in the liver and lungs [57, 63]. Treatment with zoledronic acid also prolonged survival of tumor-bearing mice. In the same model, ibandronate (4 µg/day s.c.) had no effect on lung or liver metastases or on survival [61, 63]. As mentioned above, treatment of mice bearing 5T2 myeloma cells with zoledronic acid (120 µg/kg s.c. twice weekly) also resulted in a significantly longer disease-free survival time (median, 47 days versus 35 days for untreated controls; p < 0.01) [53].

These animal models provide convincing evidence of the potential of bisphosphonates, particularly the more potent N-BPs, to reduce tumor burden in bone and inhibit formation and progression of bone metastases in a variety of tumor models. It is less clear whether bisphosphonates have antitumor activity beyond the bone, but it appears that zoledronic acid may be sufficiently potent to inhibit extraskeletal tumor cell growth and to prolong survival in some animal models.


   MECHANISMS OF ANTITUMOR EFFECTS
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanism of Action
 Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
Conclusions and Future...
 References
 
The precise mechanisms responsible for the antitumor effects of bisphosphonates are beginning to be elucidated. Bisphosphonates appear to make the bone a less favorable site for tumor cell growth via the inhibition of osteoclast-mediated bone resorption and osteoclastogenesis, thereby reducing the release of growth factors that stimulate tumor growth in bone. In addition, bisphosphonates directly inhibit tumor cell growth and survival and the ability of tumor cells to colonize the bone. Either or both of these mechanisms may be at work.

Apoptosis
One of the primary mechanisms responsible for the direct antitumor activity of bisphosphonates is induction of tumor cell apoptosis. Both N-BPs and non-N-BPs appear to induce apoptosis of osteoclasts and tumor cells by activation of caspases [9, 12–14, 16, 17, 64, 65]. One mechanism by which bisphosphonates induce apoptosis is through production of ATP analogues (either as direct metabolites or as a result of inhibition of the mevalonate pathway), which can disrupt mitochondrial ATP/ADP translocase. A recent study investigating the mechanism by which zoledronic acid induced apoptosis in human breast cancer cell lines (MDA-MB-231 and MCF-7) indicated that this response was associated with cytochrome c release from the mitochondria and subsequent caspase-3 activation [66]. It appears that N-BPs may induce cytochrome c release by modulating expression of Bcl-2, a key antiapoptotic regulatory protein [66]. These events may be precipitated by inhibition of Ras activation, which requires protein prenylation (specifically farnesylation) [66].

Inhibition of Tumor Cell Adhesion and Invasion of the Extracellular Bone Matrix
Bisphosphonates have also been shown to inhibit adhesion of tumor cells to extracellular matrix (ECM) proteins and to inhibit the process of tumor cell invasion and metastasis [67–70]. Using an in vitro MatrigelTM-based invasion assay, Boissier et al. have shown that bisphosphonates inhibit the ability of human breast and prostate cancer cells to invade the ECM [67]. In this assay, zoledronic acid and ibandronate caused dose-dependent inhibition of cell invasion through the MatrigelTM at extremely low concentrations (≤10–10 M) that did not inhibit tumor cell motility or induce significant apoptosis. Clodronate was approximately six orders of magnitude less potent in the same assay. Furthermore, the combination of ibandronate with taxanes enhanced the inhibitory effect on MDA-MB-231 cell invasion [69].

One contributory mechanism may be the inhibition of matrix metalloproteinase (MMP) activity, which is necessary for tumor cell invasion of the ECM [67, 71, 72]. Bisphosphonates have been shown to inhibit the activity of MMPs produced by tumor cell lines, and this seems to correlate with reduced invasiveness in the MatrigelTM assay [71, 72]. For example, zoledronic acid was shown to inhibit the production of MMP-2 and MMP-9 by PC-3 cells [40]. These data suggest a potential mechanism by which N-BPs may inhibit tumor cell invasion of the bone but cannot explain the apparent dependence of this effect on protein prenylation.

Recent studies suggest that inhibition of tumor cell adhesion to ECM proteins and invasion through MatrigelTM is dependent on inhibition of protein prenylation. In one study, zoledronic acid was shown to dose-dependently inhibit adhesion of MCF-7 and MDA-MB-231 cells to a variety of matrix proteins (Fig. 5Go) [73], and this inhibitory effect was overcome by addition of either farnesol or geranylgeraniol or by the addition of a broad spectrum caspase inhibitor [73]. Similar findings have been reported for alendronate. The inhibitory effect of alendronate on tumor cell invasion through MatrigelTM was reversed by the addition of geranylgeraniol and trans-trans-farnesol [74]. Therefore, inhibition of the mevalonate pathway and induction of caspase activity are important for the inhibitory effects of N-BPs on tumor cell adhesion to the ECM and on invasiveness. Further, it has been shown that an activating Ras mutation enhances adhesion of a normal breast epithelial cell line to ECM proteins, suggesting that increased Ras activation in response to growth factor receptor signaling may increase the metastatic potential of breast cancer cells [73]. Thus, by inhibiting protein prenylation and Ras signaling, zoledronic acid should reduce the metastatic potential of tumor cells.



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  		Figure 5. Percent adherent cells versus controls when MDA-MB-231 human breast cancer cells were incubated with zoledronic acid for 24 hours (0.1 or 100 µM) before culture on plates coated with various extracellular matrix proteins. Adapted with permission from Pickering et al. [73].

 
Antiangiogenic Effects
In vitro and in vivo studies have further demonstrated that zoledronic acid has antiangiogenic effects. In vitro assays with human umbilical vein endothelial cells (HUVECs) have shown that zoledronic acid dose-dependently inhibited the proliferation of HUVECs induced by fetal calf serum and basic fibroblast growth factor (bFGF), and these findings have been confirmed in vivo [75]. Systemic administration of zoledronic acid to mice resulted in potent inhibition of angiogenesis induced by s.c. implants impregnated with bFGF, with a dose of 3 µg/kg producing a 50% efficacy (ED50) [75]. It has also been reported that zoledronic acid can reduce bone-tumor-associated angiogenesis in the murine 5T2 myeloma model [53]. In another series of experiments, zoledronic acid, as well as ibandronate, risedronate, and clodronate, inhibited formation of capillary-like tubules by HUVECs in vitro [76]. In vivo, zoledronic acid and ibandronate, but not clodronate, decreased revascularization (as measured by vessel area) of the ventral prostate gland in castrated rats treated with testosterone [76].

The inhibitory effect of zoledronic acid on endothelial cell adhesion and migration appears to be mediated, at least in part, by modulation of integrins (e.g., {alpha}vß3 and {alpha}vß5) that are involved in angiogenesis [77, 78]. Interestingly, {alpha}vß3 integrin is also required for osteoclasts to adhere tightly to the bone and form resorption lacunae during active bone resorption, and {alpha}vß3 expression confers on tumor cells a greater propensity to metastasize to bone [79]. In fact, a small molecule inhibitor of {alpha}vß3 was recently shown to effectively prevent metastasis of MDA-MB-435 breast cancer cells to bone [80]. Therefore, effects on {alpha}vß3 could have pleiotropic effects on bone resorption and tumor metastasis. In addition, it has recently been reported that zoledronic acid decreases the survival of HUVECs by sensitizing them to TNF-induced programmed cell death [78]. Zoledronic acid also appears to modulate serum levels of proangiogenic growth factors such as vascular endothelial growth factor and bFGF in cancer patients [81]. These studies suggest a variety of potential mechanisms to account for the observed antiangiogenic effects of bisphosphonates.


   CONCLUSIONS AND FUTURE DIRECTIONS
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanism of Action
 Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
 Conclusions and Future...
References
 
There is now extensive in vitro and in vivo preclinical evidence that bisphosphonates, particularly the more potent N-BPs, have antitumor activity and can reduce skeletal tumor burden. The evidence that they have antitumor activity outside the bone is more tenuous. A variety of potential mechanisms to explain these observed antitumor effects have been proposed, including indirect effects on tumor cell growth in bone via inhibition of bone resorption and osteoclastogenesis. In addition, bisphosphonates clearly have the potential to directly induce apoptosis of tumor cells, inhibit tumor cell adhesion to the ECM, reduce the metastatic potential of tumor cells, and inhibit angiogenesis. Further research is ongoing to fully elucidate the molecular mechanisms involved and to determine the most effective dose and schedule of bisphosphonates to maximize their antitumor potential in the clinical setting, either alone or in combination with standard antineoplastic agents.


   ACKNOWLEDGMENT
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanism of Action
 Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
 Conclusions and Future...
 References
 
Jonathan Green is a full time employee of Novartis Pharma AG and holds stock in the company.


   References
Top
 Learning Objectives
 Abstract
 Introduction
 Mechanism of Action
 Evidence of Antitumor Effects
 Mechanisms of Antitumor Effects
 Conclusions and Future...
 References
 

  1. Fleisch H. Development of bisphosphonates. Breast Cancer Res 2002;4:30–34.[Medline]
  2. Russell RG, Croucher PI, Rogers MJ. Bisphosphonates: pharmacology, mechanisms of action and clinical uses. Osteoporos Int 1999;9(suppl 2):S66–S80.
  3. Stronski SA, Bettschen-Camin L, Wetterwald A et al. Bisphosphonates inhibit 1,25-dihydroxyvitamin D3-induced increase of osteocalcin in plasma of rats in vivo and in culture medium of rat calvaria in vitro. Calcif Tissue Int 1988;42:248–254.[Medline]
  4. Flanagan AM, Chambers TJ. Inhibition of bone resorption by bisphosphonates: interactions between bisphosphonates, osteoclasts, and bone. Calcif Tissue Int 1991;49:407–415.[Medline]
  5. Rogers MJ, Gordon S, Benford HL et al. Cellular and molecular mechanisms of action of bisphosphonates. Cancer 2000;88(suppl 12):2961–2978.[CrossRef][Medline]
  6. Sato M, Grasser W, Endo N et al. Bisphosphonate action. Alendronate localization in rat bone and effects on osteoclast ultrastructure. J Clin Invest 1991;88:2095–2105.
  7. Aparicio A, Gardner A, Tu Y et al. In vitro cytoreductive effects on multiple myeloma cells induced by bisphosphonates. Leukemia 1998;12:220–229.[CrossRef][Medline]
  8. Derenne S, Amiot M, Barille S et al. Zoledronate is a potent inhibitor of myeloma cell growth and secretion of IL-6 and MMP-1 by the tumoral environment. J Bone Miner Res 1999;14:2048–2056.[CrossRef][Medline]
  9. Hiraga T, Williams PJ, Mundy GR et al. The bisphosphonate ibandronate promotes apoptosis in MDA-MB-231 human breast cancer cells in bone metastases. Cancer Res 2001;61:4418–4424.[Abstract/Free Full Text]
  10. Lee MV, Fong EM, Singer FR et al. Bisphosphonate treatment inhibits the growth of prostate cancer cells. Cancer Res 2001;61:2602–2608.[Abstract/Free Full Text]
  11. Mackie PS, Fisher JL, Zhou H et al. Bisphosphonates regulate cell growth and gene expression in the UMR 106-01 clonal rat osteosarcoma cell line. Br J Cancer 2001;84:951–958.[CrossRef][Medline]
  12. Senaratne SG, Pirianov G, Mansi JL et al. Bisphosphonates induce apoptosis in human breast cancer cell lines. Br J Cancer 2000;82:1459–1468.[Medline]
  13. Shipman CM, Rogers MJ, Apperley JF et al. Bisphosphonates induce apoptosis in human myeloma cell lines: a novel anti-tumour activity. Br J Haematol 1997;98:665–672.[CrossRef][Medline]
  14. Shipman CM, Croucher PI, Russell RG et al. The bisphosphonate incadronate (YM175) causes apoptosis of human myeloma cells in vitro by inhibiting the mevalonate pathway. Cancer Res 1998;58:5294–5297.[Abstract/Free Full Text]
  15. Sonnemann J, Eckervogt V, Truckenbrod B et al. The bisphosphonate pamidronate is a potent inhibitor of human osteosarcoma cell growth in vitro. Anticancer Drugs 2001;12:459–465.[CrossRef][Medline]
  16. Takahashi R, Shimazaki C, Inaba T et al. A newly developed bisphosphonate, YM529, is a potent apoptosis inducer of human myeloma cells. Leuk Res 2001;25:77–83.[CrossRef][Medline]
  17. Tassone P, Tagliaferri P, Viscomi C et al. Zoledronic acid induces antiproliferative and apoptotic effects in human pancreatic cancer cells in vitro. Br J Cancer 2003;88:1971–1978.[CrossRef][Medline]
  18. Rogers MJ, Russell RG, Blackburn GM et al. Metabolism of halogenated bisphosphonates by the cellular slime mould Dictyostelium discoideum. Biochem Biophys Res Commun 1992;189:414–423.[CrossRef][Medline]
  19. Rogers MJ, Ji X, Russell RG et al. Incorporation of bisphosphonates into adenine nucleotides by amoebae of the cellular slime mould Dictyostelium discoideum. Biochem J 1994;303:303–311.
  20. Pelorgeas S, Martin JB, Satre M. Cytotoxicity of dichloromethane diphosphonate and of 1-hydroxyethane-1,1-diphosphonate in the amoebae of the slime mould Dictyostelium discoideum. A 31P NMR study. Biochem Pharmacol 1992;44:2157–2163.[CrossRef][Medline]
  21. Frith JC, Mönkkönen J, Blackburn GM et al. Clodronate and liposome-encapsulated clodronate are metabolized to a toxic ATP analog, adenosine 5'-(beta, gamma-dichloromethylene) triphosphate, by mammalian cells in vitro. J Bone Miner Res 1997;12:1358–1367.[CrossRef][Medline]
  22. Lehenkari PP, Kellinsalmi M, Napankangas JP et al. Further insight into mechanism of action of clodronate: inhibition of mitochondrial ADP/ATP translocase by a nonhydrolyzable, adenine-containing metabolite. Mol Pharmacol 2002;61:1255–1262.[Abstract/Free Full Text]
  23. Rogers MJ, Watts DJ, Russell RGG. Overview of bisphosphonates. Cancer 1997;80(suppl 8):1652–1660.[CrossRef][Medline]
  24. Selander KS, Monkkonen J, Karhukorpi EK et al. Characteristics of clodronate-induced apoptosis in osteoclasts and macrophages. Mol Pharmacol 1996;50:1127–1138.[Abstract]
  25. Gober HJ, Kistowska M, Angman L et al. Human T cell receptor gammadelta cells recognize endogenous mevalonate metabolites in tumor cells. J Exp Med 2003;197:163–168.[Abstract/Free Full Text]
  26. Luckman SP, Hughes DE, Coxon FP et al. Nitrogen-containing bisphosphonates inhibit the mevalonate pathway and prevent post-translational prenylation of GTP-binding proteins, including Ras. J Bone Miner Res 1998;13:581–589.[CrossRef][Medline]
  27. Rogers MJ, Frith JC, Luckman SP et al. Molecular mechanisms of action of bisphosphonates. Bone 1999;24(suppl 5):73S–79S.[Medline]
  28. Russell RG, Rogers MJ, Frith JC et al. The pharmacology of bisphosphonates and new insights into their mechanisms of action. J Bone Miner Res 1999;14(suppl 2):53–65.
  29. Hughes DE, Wright KR, Uy HL et al. Bisphosphonates promote apoptosis in murine osteoclasts in vitro and in vivo. J Bone Miner Res 1995;10:1478–1487.[Medline]
  30. Luckman SP, Coxon FP, Ebetino FH et al. Heterocycle-containing bisphosphonates cause apoptosis and inhibit bone resorption by preventing protein prenylation: evidence from structure-activity relationships in J774 macrophages. J Bone Miner Res 1998;13:1668–1678.[CrossRef][Medline]
  31. Oliff A. Farnesyltransferase inhibitors: targeting the molecular basis of cancer. Biochim Biophys Acta 1999;1423:C19–C30.[Medline]
  32. Zhang FL, Casey PJ. Protein prenylation: molecular mechanisms and functional consequences. Annu Rev Biochem 1996;65:241–269.[CrossRef][Medline]
  33. Alakangas A, Selander K, Mulari M et al. Alendronate disturbs vesicular trafficking in osteoclasts. Calcif Tissue Int 2002;70:40–47.[CrossRef][Medline]
  34. Mönkkönen H, Lehenkari PP, Kellinsalmi M et al. A new mechanism of action for bisphosphonates: apppi dedicated cytotoxicity of N-BPs. Bone 2004;34:S66–S67.
  35. Dunford JE, Thompson K, Coxon FP et al. Structure-activity relationships for inhibition of farnesyl diphosphate synthase in vitro and inhibition of bone resorption in vivo by nitrogen-containing bisphosphonates. J Pharmacol Exp Ther 2001;296:235–242.[Abstract/Free Full Text]
  36. Green JR, Müller K, Jaeggi KA. Preclinical pharmacology of CGP 42'446, a new, potent, heterocyclic bisphosphonate compound. J Bone Miner Res 1994;9:745–751.[Medline]
  37. Fisher JE, Rodan GA, Reszka AA. In vivo effects of bisphosphonates on the osteoclast mevalonate pathway. Endocrinology 2000;141:4793–4796.[Abstract/Free Full Text]
  38. Clezardin P. The antitumor potential of bisphosphonates. Semin Oncol 2002;29(suppl 21):33–42.[Medline]
  39. Jagdev SP, Coleman RE, Shipman CM et al. The bisphosphonate, zoledronic acid, induces apoptosis of breast cancer cells: evidence for synergy with paclitaxel. Br J Cancer 2001;84:1126–1134.[CrossRef][Medline]
  40. Corey E, Brown LG, Quinn JE et al. Zoledronic acid exhibits inhibitory effects on osteoblastic and osteolytic metastases of prostate cancer. Clin Cancer Res 2003;9:295–306. Erratum in: Clin Cancer Res 2003;9:1574–1575.[Abstract/Free Full Text]
  41. Oades GM, Senaratne SG, Clarke IA et al. Nitrogen containing bisphosphonates induce apoptosis and inhibit the mevalonate pathway, impairing Ras membrane localization in prostate cancer cells. J Urol 2003;170:246–252.[CrossRef][Medline]
  42. Matsumoto S, Kimura S, Segawa H et al. Efficacy of combining the third generation bisphosphonate, zoledronate with imatinib mesylate in suppressing small cell lung cancer cell line proliferation. Proc Am Soc Clin Oncol 2003;22:684.
  43. Neville-Webbe HL, Evans CA, Coleman RE et al. Zoledronic acid (ZOL) in combination with doxorubicin (DOX) has synergistic effects on apoptosis of breast cancer cells (BCCs) in vitro [abstract]. Presented at the IVth International Conference on Cancer-Induced Bone Diseases, December 7–9, 2003, San Antonio, TX.
  44. Tassone P, Forciniti S, Galea E et al. Growth inhibition and synergistic induction of apoptosis by zoledronate and dexamethasone in human myeloma cell lines. Leukemia 2000;14:841–844.[CrossRef][Medline]
  45. Ullén A, Lennartsson L, Hjelm-Eriksson M et al. Additive/synergistic anti-tumoral effects on prostate cancer cells in vitro following treatment with a combination of docetaxel and zoledronate [poster]. Presented at the European Winter Oncology Conference (EWOC-8), January 19–24, 2003, Flims, Switzerland.
  46. Ullén A, Lennartsson L, Hjelm-Eriksson M et al. Additive/synergistic anti-tumoral effect on prostate cancer cells in vitro following treatment with a combination of gemcitabine and zoledronic acid. Proc Am Soc Clin Oncol 2003;22:432.
  47. Vogt U, Wassmann K, Bosse U et al. Enhancement of breast tumor growth inhibition (BTGI) in vitro through combination of CMF, epirubicin/cyclophosphamide (EC), epirubicin/paclitaxel (ET) and epirubicin/docetaxel (EDoc) with ibandronate (IB) or zoledronic acid (ZOL). Proc Am Soc Clin Oncol 2003;22:55.
  48. Witters L, Crispino J, Javeed M et al. Inhibition of growth of a human prostate cancer cell line with the combination of zoledronic acid and a COX-2 inhibitor. Proc Am Soc Clin Oncol 2002;21:5b.
  49. Witters LM, Crispino J, Fraterrigo T et al. Effect of the combination of docetaxel, zoledronic acid, and a COX-2 inhibitor on the growth of human breast cancer cell lines. Am J Clin Oncol 2003;26:S92–S97.[CrossRef][Medline]
  50. Schlotter CM, Vogt U, Bosse U et al. Enhancement of breast tumor growth inhibition (BTGI) in vitro through combination of CMF, epirubicin/cyclophosphamide (EC), and epirubicin/paclitaxel (ET) with ibandronate (IB) or zoledronic acid (ZOL). J Bone Miner Res 2001;16(suppl 1):S191.
  51. Kuroda J, Kimura S, Segawa H et al. The third-generation bisphosphonate zoledronate synergistically augments the anti-Ph+ leukemia activity of imatinib mesylate. Blood 2003;102:2229–2235.[Abstract/Free Full Text]
  52. Yaccoby S, Pearse RN, Johnson CL et al. Myeloma interacts with the bone marrow microenvironment to induce osteoclastogenesis and is dependent on osteoclast activity. Br J Haematol 2002;116:278–290.[CrossRef][Medline]
  53. Croucher PI, De Raeve H, Perry MJ et al. Zoledronic acid treatment of 5T2MM-bearing mice inhibits the development of myeloma bone disease: evidence for decreased osteolysis, tumor burden and angiogenesis, and increased survival. J Bone Miner Res 2003;18:482–492.[CrossRef][Medline]
  54. Green J, Gschaidmeier H, Yoneda T et al. Zoledronic acid potently inhibits tumour-induced osteolysis in two models of breast cancer metastasis to bone. Ann Oncol 2000;11(suppl 4):14.
  55. Peyruchaud O, Winding B, Pecheur I et al. Early detection of bone metastases in a murine model using fluorescent human breast cancer cells: application to the use of the bisphosphonate zoledronic acid in the treatment of osteolytic lesions. J Bone Miner Res 2001;16:2027–2034.[CrossRef][Medline]
  56. Hiraga T, Ueda A, Tamura D et al. Effects of oral UFT combined with or without zoledronic acid on bone metastasis in the 4T1/luc mouse breast cancer. Int J Cancer 2003;106:973–979.[CrossRef][Medline]
  57. Nobuyuki H, Hiraga T, Williams PJ et al. The bisphosphonate zoledronic acid inhibits metastases to bone and liver with suppression of osteopontin production in mouse mammary tumor. J Bone Miner Res 2001;16(suppl 1):S191.
  58. Green JR. Bisphosphonates in cancer therapy. Curr Opin Oncol 2002;14:609–615.[CrossRef][Medline]
  59. Cruz JC, Alsina M, Craig F et al. Ibandronate decreases bone disease development and osteoclast stimulatory activity in an in vivo model of human myeloma. Exp Hematol 2001;29:441–447.[CrossRef][Medline]
  60. Alvarez E, Westmore M, Galvin RJ et al. Properties of bisphosphonates in the 13762 rat mammary carcinoma model of tumor-induced bone resorption. Clin Cancer Res 2003;9:5705–5713.[Abstract/Free Full Text]
  61. Michigami T, Hiraga T, Williams PJ et al. The effect of the bisphosphonate ibandronate on breast cancer metastasis to visceral organs. Breast Cancer Res Treat 2002;75:249–258.[CrossRef][Medline]
  62. Padalecki SS, Carreon M, Grubbs B et al. Androgen deprivation enhances bone loss and prostate cancer metastases to bone: prevention by zoledronic acid. Oncology 2003;17(suppl):32.[Medline]
  63. Yoneda T, Hashimoto N, Hiraga T. Bisphosphonate actions on cancer. Calcif Tissue Int 2003;73:315–318.[CrossRef][Medline]
  64. Benford HL, McGowan NWA, Helfrich MH et al. Visualization of bisphosphonate-induced caspase-3 activity in apoptotic osteoclasts in vitro. Bone 2001;28:465–473.[Medline]
  65. Fromigue O, Lagneaux L, Body JJ. Bisphosphonates induce breast cancer cell death in vitro. J Bone Miner Res 2000;15:2211–2221.[CrossRef][Medline]
  66. Senaratne SG, Mansi JL, Colston KW. The bisphosphonate zoledronic acid impairs Ras membrane [correction of impairs membrane] localisation and induces cytochrome c release in breast cancer cells. Br J Cancer 2002;86:1479–1486. Erratum in: Br J Cancer 2002;87:1340.[CrossRef][Medline]
  67. Boissier S, Ferreras M, Peyruchaud O et al. Bisphosphonates inhibit breast and prostate carcinoma cell invasion, an early event in the formation of bone metastases. Cancer Res 2000;60:2949–2954.[Abstract/Free Full Text]
  68. Boissier S, Magnetto S, Frappart L et al. Bisphosphonates inhibit prostate and breast carcinoma cell adhesion to unmineralized and mineralized bone extracellular matrices. Cancer Res 1997;57:3890–3894.[Abstract/Free Full Text]
  69. Magnetto S, Boissier S, Delmas PD et al. Additive antitumor activities of taxoids in combination with the bisphosphonate ibandronate against invasion and adhesion of human breast carcinoma cells to bone. Int J Cancer 1999;83:263–269.[CrossRef][Medline]
  70. van der Pluijm G, Vloedgraven H, van Beek E et al. Bisphosphonates inhibit the adhesion of breast cancer cells to bone matrices in vitro. J Clin Invest 1996;98:698–705.[Medline]
  71. Teronen O, Heikkila P, Konttinen YT et al. MMP inhibition and downregulation by bisphosphonates. Ann NY Acad Sci 1999;878:453–465.[CrossRef][Medline]
  72. Heikkila P, Teronen O, Moilanen M et al. Bisphosphonates inhibit stromelysin-1 (MMP-3), matrix metalloelastase (MMP-12), collagenase-3 (MMP-13) and enamelysin (MMP-20), but not urokinase-type plasminogen activator, and diminish invasion and migration of human malignant and endothelial cell lines. Anticancer Drugs 2002;13:245–254.[CrossRef][Medline]
  73. Pickering LM, Mansi JL, Colston KW. Adhesion of breast cancer cells to extracellular matrices is inhibited by zoledronic acid and enhanced by aberrant Ras signalling. Proc Am Soc Clin Oncol 2003;22:863.
  74. Virtanen SS, Väänänen HK, Härkönen PL et al. Alendronate inhibits invasion of PC-3 prostate cancer cells by affecting the mevalonate pathway. Cancer Res 2002;62:2708–2714.[Abstract/Free Full Text]
  75. Wood J, Bonjean K, Ruetz S et al. Novel antiangiogenic effects of the bisphosphonate compound zoledronic acid. J Pharmacol Exp Ther 2002;302:1055–1061.[Abstract/Free Full Text]
  76. Fournier P, Boissier S, Filleur S et al. Bisphosphonates inhibit angiogenesis in vitro and testosterone-stimulated vascular regrowth in the ventral prostate in castrated rats. Cancer Res 2002;62:6538–6544.[Abstract/Free Full Text]
  77. Bonjean K, Bellahcene A, Locigno R et al. Zoledronate modulates endothelial cell surface receptors involved in angiogenesis. Proc Am Assoc Cancer Res 2001;42:106.
  78. Bezzi M, Hasmim M, Bieler G et al. Zoledronate sensitizes endothelial cells to tumor necrosis factor-induced programmed cell death. Evidence for the suppression of sustained activation of focal adhesion kinase and protein kinase B/Akt. J Biol Chem 2003;278:43603–43614.[Abstract/Free Full Text]
  79. Pécheur I, Peyruchaud O, Serre CM et al. Integrin alpha(v)beta3 expression confers on tumor cells a greater propensity to metastasize to bone. FASEB J 2002;16:1266–1268.[Abstract/Free Full Text]
  80. Welch DR, Harms JF, Samant RS et al. The small molecule {alpha}vß3 antagonist (S247) inhibits MDA-MB-435 breast cancer metastasis to bone. Oncology 2003;17(suppl):18.[Medline]
  81. Santini D, Vincenzi B, Dicuonzo G et al. Zoledronic acid induces significant and long-lasting modifications of circulating angiogenic factors in cancer patients. Clin Cancer Res 2003;9:2893–2897.[Abstract/Free Full Text]
Received July 19, 2004; accepted for publication August 3, 2004.




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