ARTICLE IN PRESS

THE BREAST

0960-9776/$ - se

doi:10.1016/j.br

Abbreviations

sphate; HCM,

teinases; NSCL

events; VEGF,�Tel.: +44 11

E-mail addr

The Breast 16 (2007) 21–27

www.elsevier.com/locate/breast

Original Article

On the horizon: Can bisphosphonates prevent bone metastases?

R. Coleman�

Weston Park Hospital, Cancer Research Centre, Academic Unit of Clinical Oncology, Sheffield, UK

Abstract

Skeletal complications of bone metastases increase the risk of death and undermine patients’ functional independence and quality of

life. Although bisphosphonates are integral in the treatment regimen of patients with metastatic bone disease and have demonstrated

efficacy in delaying the onset and reducing the incidence of skeletal-related events, there is great interest in developing treatments to

prevent metastasis to bone. Emerging evidence indicates that the potential benefits of bisphosphonate therapy extend beyond the

treatment of metastatic bone lesions. Data from preclinical studies suggest that bisphosphonates may have antitumour activity and may

prevent bone metastasis. The mechanisms of these antitumour effects are currently under investigation and may include induction of

apoptosis, inhibition of tumour cell invasion and angiogenesis, and tumour growth reduction. Therefore, patients with early-stage disease

may benefit from early bisphosphonate therapy, before bone metastasis develops, and investigations are ongoing to determine the clinical

utility of bisphosphonates in this setting.

r 2007 Elsevier Ltd. All rights reserved.

Keywords: Antitumour; Bisphosphonates; Bone metastases; Skeletal-related events

Introduction

Bone metastases are a major source of skeletal morbidityin the form of skeletal-related events (SREs) includingpathologic fractures, spinal cord compression, severe bonepain requiring strong analgesics or palliative radiotherapy,surgery to bone, and hypercalcaemia of malignancy(HCM).1,2 These events can dramatically erode a patient’ssense of well-being and significantly undermine functionalindependence and quality of life. In addition, treatment ofSREs can result in an economic burden on families and thehealthcare system. Therefore, there is great interest indeveloping treatments to prevent metastasis to bone, andbisphosphonates are under investigation in this setting.

Bisphosphonates are inhibitors of osteoclast-mediatedbone resorption. The current indications for bisphospho-nates include the treatment of metabolic disorders of bonemetabolism (e.g., osteoporosis and Paget’s disease), the

e front matter r 2007 Elsevier Ltd. All rights reserved.

east.2007.10.006

: FGF, fibroblast growth factor; FPP, farnesyl dipho-

hypercalcaemia of malignancy; MMP, matrix metallopro-

C, non-small cell lung cancer; SREs, skeletal-related

vascular endothelial growth factor

4 226 5213; fax: +44 114 226 5678.

ess: r.e.coleman@sheffield.ac.uk

treatment of HCM, and the prevention of SREs frommalignant bone disease. Clinical trials also confirm thatbisphosphonates, and zoledronic acid in particular, canprevent bone loss from cancer treatment.3 There is alsoemerging evidence that the benefits of bisphosphonatetherapy in the oncology setting are more extensive. Forexample, data from preclinical studies demonstrate thatbisphosphonates may have antitumour activity, as evi-denced in vitro by reduced proliferation rates of tumourcell lines and in vivo by slower bone lesion progression inanimal models.4,5 The mechanisms responsible for theseantitumour effects are currently under investigation. It islikely that the multiple effects of bisphosphonates oncancer cells, as summarised herein, may be beneficial forpatients with early-stage disease.

Preclinical evidence for antitumour effects of

bisphosphonates

There is a strong rationale from preclinical assessmentsand preliminary clinical data to support the hypothesis thatbisphosphonates may affect the course of cancer progres-sion, especially with regard to the development of bonemetastases. In addition to their inhibitory effects on

ARTICLE IN PRESSR. Coleman / The Breast 16 (2007) 21–27S22

osteoclast-mediated osteolysis, bisphosphonates can alsoexert direct and indirect effects on osteoblasts, macro-phages, and cancer cells.6 In preclinical assays and modelsystems, the more active nitrogen-containing bisphospho-nates have been found to exert anticancer effects andinhibit multiple steps necessary for metastasis to bone.7 Forexample, zoledronic acid induces apoptosis in humanbreast and prostate cancer cell lines and exerts synergisticantitumour effects with both anthracyclines and taxanes.8,9

Clinically relevant concentration of doxorubicin andzoledronic acid-induced sequence- and dose-dependentapoptosis of breast and prostate cancer cell lines, whereasthe replacement of zoledronic acid with a non-nitrogen-containing bisphosphonate did not increase levels ofapoptosis (Fig. 1).8 Similar sequential effects of paclitaxeland zoledronic acid have also been observed in breastcancer cells.9 Furthermore, in mouse model systems ofbreast cancer, zoledronic acid not only delayed thedevelopment of metastatic lesions in the bone and viscera(Fig. 2),10 but also had synergistic effects in combinationwith doxorubicin for suppressing tumour growth andvascularisation.11

Nitrogen-containing bisphosphonates have also beenshown to reduce tumour-induced osteolysis and diseaseprogression in bone in mouse models of human cancerssuch as multiple myeloma, breast cancer, and prostatecancer.5,12–14 Furthermore, the use of radiographic,histologic, and histomorphometric techniques in variousmodels has demonstrated that nitrogen-containing bispho-sphonates can inhibit the formation or progression of bonemetastases and/or reduce skeletal tumour burden indepen-dent of the timing of bisphosphonate administration.4

In these models, bisphosphonates were effective whenadministered at the time of tumour cell inoculation as

Fig. 1. Sequential treatment of human cancer cells with doxorubicin and zoled

(murine fibroblastic) cells were treated with doxorubicin (0.05 mM for 24 h) an

maintained in fresh medium for 48 h, and percent apoptotic cells determined.bPp0.03 for doxorubicin then zoledronic acid compared with drugs alone. A

well as after bone metastases were already established,suggesting that they may play a role in both the preventionand treatment of bone metastases.

Inhibition of new blood vessel formation

In animal model systems, bisphosphonates have beenshown to inhibit cancer-associated increases in angiogen-esis,15–21 an important early step in tumour growth andmetastasis.22 In mice, bisphosphonate treatment reducedthe microvessel density of osteolytic lesions from multiplemyeloma bone disease,12 and zoledronic acid was found topotently inhibit fibroblast growth factor (FGF)-inducedangiogenesis.19 The FGF family of proteins has beenreported to play a role in the development of the mammarygland and to act as oncogenic factors during the develop-ment of breast cancer.23 The effects of bisphosphonates onangiogenesis may be clinically meaningful because in recentpilot studies in patients with advanced tumours metastaticto bone, both pamidronate and zoledronic acid were foundto lower serum levels of vascular endothelial growth factor(VEGF).24,25 Similarly, in a pilot study of patients withmetastatic bone disease (N=26) who were treated with 4weekly doses of 1mg zoledronic acid followed by standarddoses (4mg every 28 days), basal levels of VEGF weresignificantly decreased after 7 days of treatment(P=0.038).26 This reduction in circulating VEGF levelspersisted throughout the entire 84-day study (Po0.01 forall time points) and demonstrated the potential andpersistent antiangiogenic properties of zoledronic acid.26

Moreover, in a recent study in women with bonemetastases secondary to breast cancer, zoledronic acidlowered serum VEGF levels in most treated patients, andreductions in VEGF levels correlated with prolonged times

ronic acid enhances apoptosis. MCF-7 (breast), PC-3 (prostate), and 3T3

d then zoledronic acid (25 mM [MCF-7 or 1mM [PC-3 and 3T3] for 1 h),

Dox ¼ doxorubicin; Zol ¼ zoledronic acid; Ctl ¼ control. aP ¼ 0.004 and

dapted with permission from Neville-Webbe et al.8

ARTICLE IN PRESS

Fig. 2. Zoledronic acid inhibits bone, lung and liver metastases in a murine model of breast cancer. Histomorphometric analysis of tumour burden in bone

(mm2/mouse) and lung (% tumour area). Metastatic tumour burden in the lung and liver determined by luciferase activity (% luc). Data are mean7SE.

Representative histologic views of lung metastases treated with (ZOL) or without (Vehicle) zoledronic acid (5mg/mouse); asterisk within photo micrograph

indicates tumour location, bar ¼ 500mm; aindicates significant difference from untreated group (Po0.05). Adapted with permission from Hiraga et al.10

R. Coleman / The Breast 16 (2007) 21–27 S23

to the following clinical events: first SRE (P ¼ 0.0002),bone lesion progression (P ¼ 0.0024), and first decrease inperformance status (P ¼ 0.0352).21

Inhibition of invasion and attachment

In addition to their inhibitory effects on osteolysis,bisphosphonates can impede cancer cell invasion throughthe extracellular matrix and attachment to the capillarybeds in bone.27,28 Zoledronic acid has been shown toreduce the motility and migration of a variety of humancancer cell lines, including highly motile non-small cell lungcancer (NSCLC) cell lines, in vitro.29 Heikkila et al.30

reported that the activities of matrix metalloproteinases(MMPs), which facilitate tumour cell invasion, are alsoimpeded by nitrogen-containing bisphosphonates at clini-cally relevant concentrations. Moreover, these agents alsoimpeded invasion and migration by human breast cancercell lines in in vitro assay systems (e.g., MatrigelTM assays;BD Biosciences, Franklin Lakes, New Jersey) at concen-trations that were similar to those required for inhibitingMMP activity. In addition to blocking the activity ofMMPs, bisphosphonates may also reduce levels of MMPsecretion. In a small study in 30 patients with painful bonemetastases from breast or prostate cancer,31 zoledronicacid treatment reduced plasma levels of MMPs and otherproteinases. Therefore, both the level and relative activitiesof these factors may be reduced by bisphosphonatetherapy.

Direct antitumour effects

Nitrogen-containing bisphosphonates inhibit farnesyldiphosphate (FPP) synthase, an enzyme in the mevalonate

pathway that is required for the processing of key signaltransduction intermediates such as Ras and Rho. Many ofthese proteins are involved in oncogenesis; therefore,uptake of nitrogen-containing bisphosphonates by cancercells could impede tumour growth. Indeed, bisphospho-nates have been shown to inhibit the proliferation of andinduce apoptosis in a broad range of human cancer celllines and primary tumour isolates.4,32–34 Zoledronic acidhas inhibited the growth of all cancer cell lines in which ithas been studied. The mechanisms behind these effects arecurrently unknown, and it is likely that multiple cellularpathways are affected downstream of FPP synthase.4,32,35

For example, in a study of zoledronic acid in lung cancercell lines, antiproliferative effects significantly correlatedwith inhibition of the mevalonate pathway, as assessed byaccumulation of an unprenylated pathway intermediate(Rap1), and were reversed by introduction of geranylger-aniol,29 an intermediate downstream of FPP synthase.4

Immunomodulatory effects

The intravenous administration of nitrogen-containingbisphosphonates is associated with an acute-phase reactioncharacterised by flu-like symptoms in approximately onethird of patients receiving intravenous bisphosphonates forthe first time.36 The recently elucidated mechanism occursthrough bisphosphonate-mediated effects on immune cells.Preclinical and clinical data indicate that bisphosphonatesmay affect levels of circulating lymphocytes and antigen-presenting monocytes.37,38 Specifically, patients who ex-perience acute-phase reactions had increased levels ofcirculating gd T cells, a subset of T lymphocytes that canindependently recognise antigens and may have anticanceractivity.39 Bisphosphonates have been shown to stimulate

ARTICLE IN PRESSR. Coleman / The Breast 16 (2007) 21–27S24

gd T cells through their FPP synthase inhibitory activity,and these effects may contribute to treatment benefits.39

For example, when cancer cells were pretreated withzoledronic acid, the gd T cells recognised and lysed thecancer cells.40,41 Further studies are needed to determinethe physiologic relevance of the gd T-cell responses inpatients with cancer and to determine if acute-phasereactions correlate with bisphosphonate-mediated activa-tion of gd T cells.

Bisphosphonates modulate the interactions between tumour

and bone

Bisphosphonates have demonstrated clinical utility indelaying the onset and reducing the incidence of SREs inpatients with established bone metastases through theinhibition of osteoclast-mediated osteolysis, which inter-rupts the vicious cycle of tumour growth and bonedestruction.22 In addition, bisphosphonates may alsorender the bone microenvironment less conducive forthe development of metastatic tumours.42 Specifically,bisphosphonate treatment may lower the levels of growthfactors required for metastatic tumour growth that arereleased from the bone matrix when osteoclast-mediatedosteolysis is inhibited. In a mouse model system ofandrogen-independent prostate cancer, orchiectomy, whichresults in increased levels of osteolysis and decreased bonemineral density, resulted in a higher rate of metastasis tobone.43 Treatment with zoledronic acid, which inhibitsosteolysis and preserves bone mineral density, reduced theincidence of metastasis to bone in this animal model.43

It will be interesting to see if the bone-protective effectsseen in animal models translate to clinical benefits inhumans.

Can bisphosphonates prevent metastases in the clinic?

Although data from the definitive trials of the new-generation bisphosphonate zoledronic acid in this settinghave not yet matured, there is preliminary evidencesuggesting that patients with early breast cancer mayexperience benefits from bisphosphonate therapy. Forexample, the relatively weak first-generation bisphospho-nate clodronate has been investigated in three randomisedplacebo-controlled trials in patients with stage I–III breastcancer.44–46 Results from these studies have suggested thatbisphosphonates may prevent bone metastases in patientswith breast cancer. Only one of these studies showed asignificant positive skeletal effect.45 Interestingly, thestudies by Diel and Powles reported improved overallsurvival, while the Saarto study had no effect on overallsurvival, a reduced disease-free survival, and an increase inextraskeletal metastases.44–46 Although the results of theseclodronate trials are variable and inconclusive, they suggestthat the full antitumour effects of bisphosphonates may berealised with optimised treatment schedules or use ofanother more efficacious bisphosphonate.

In an open-label, small prospective pilot study, patientswith recurrent or metastatic advanced cancer without bonemetastases (N=40) were randomised to receive zoledronicacid or no bisphosphonate treatment.47 Zoledronic acidappeared to delay the onset of bone metastasis. Theproportion of patients who remained free from bonemetastasis at 12 months was 60% for patients treated withzoledronic acid compared with 10% for patients who didnot receive bisphosphonate therapy, and this differenceremained significant at 18 months (Po0.0005).47 Inanother study, treatment with zoledronic acid for 6 monthswas able to eliminate all detectable tumour cells in the bonemarrow of patients with early breast cancer, whereaspersisting tumour cells were detected in 29% of patients inthe control group.48 Taken together, these studies suggestthat zoledronic acid may have significant antitumourproperties in the clinical setting and thereby supply theimpetus for further studies.

Ongoing studies with bisphosphonates in the adjuvant setting

The majority of clinical data currently available onbisphosphonates clearly indicate that these agents reduceskeletal complications in patients with advanced metastaticbone disease. In addition to these treatment benefits,preliminary evidence suggests that bisphosphonate therapymay alter the course of disease progression when adminis-tered to patients early in the course of disease.Ongoing studies (i.e., NSABP-B-34, AZURE, SUC-

CESS, and S0307) are further exploring the use ofbisphosphonates as adjuvant therapy in patients withbreast cancer. The AZURE study49 will determine whetherzoledronic acid in combination with (neo)adjuvant che-motherapy and/or (neo)adjuvant endocrine therapy issuperior to (neo)adjuvant therapy alone in improvingdisease-free and bone metastasis-free survival in womenwith stage II/III breast cancer (Fig. 3).50 The first safetyanalyses of the AZURE trial indicate that the combinationof adjuvant chemotherapy and zoledronic acid is welltolerated with no significant difference in the profile orseverity of adverse events between groups.50 The NSABP-B-34 trial51 is evaluating the disease-free survival ofpatients with stage I/II breast cancer receiving clodronate(1600mg daily) or placebo for 3 years. Similarly, theSUCCESS trial, in which over 3000 patients have beenenrolled, is a German study aimed at measuring disease-free survival in patients with breast cancer who will receiveeither 2 or 5 years of zoledronic acid therapy. The primaryendpoint of the S0307 trial52 of adjuvant bisphosphonatesin breast cancer is to evaluate the disease-free survival inpatients receiving oral clodronate (1600mg daily) versusoral ibandronate (50mg daily), or patients receivingintravenous zoledronic acid (4mg once a month for6 months, then every 3 months) for 3 years.The effects of zoledronic acid (4mg every 3–4 weeks for

up to 24 months) on metastasis of NSCLC to bone and onoverall disease progression are currently being investigated

ARTICLE IN PRESS

Fig. 3. Zoledronic acid for the prevention of bone metastases in women with stage II/III breast cancer (AZURE trial). A total of 3360 patients have been

enrolled and randomised to receive standard chemotherapy or standard chemotherapy plus zoledronic acid (4mg intravenous [IV] 15min). Patients in the

standard therapy plus zoledronic acid group will receive zoledronic acid once every (q) 3–4 weeks (six doses), then once every 3 months (eight doses),

followed by once every 6 months (five doses) for a total of 5 years. Patients will be followed for 10 years to assess disease recurrence and survival. The first

efficacy data are anticipated in 2008. ER ¼ estrogen receptor; N ¼ lymph node. Reprinted from Coleman et al.50

R. Coleman / The Breast 16 (2007) 21–27 S25

in a large-scale international study53 in patients with stageIIIA or IIIB NSCLC (targeted accrual, n ¼ 445). A similarEuropean bone-metastasis prevention study54 is ongoing inpatients with prostate cancer to determine the proportionof patients with bone metastases at 48 months in patientsreceiving standard therapy plus zoledronic acid (4mg every3 months) or placebo (ZEUS; targeted accrual, n ¼ 1300).

Discussion

In summary, bone metastases compromise bone integ-rity, which results in a variety of skeletal complications thatundermine patients’ functional independence and qualityof life. Bisphosphonates are effective inhibitors of osteo-clast-mediated osteolysis and are integral in the treatmentof metastatic bone disease to delay the onset and reduce theincidence of SREs. Their potential for prevention of bonemetastasis is now an area of intense study. Data frompreclinical and preliminary clinical investigations areemerging and indicate that the therapeutic benefits ofbisphosphonates may include antitumour activity and theinhibition of metastasis. In addition to mediating theinhibition of osteolysis by osteoclasts, bisphosphonates canalso exert direct and indirect effects on other cells includingosteoblasts, macrophages, and cancer cells. Preclinicalstudies have demonstrated that bisphosphonates inhibitcancer-associated increases in angiogenesis, and these resultshave recently been validated in preliminary clinical trialswith both pamidronate and zoledronic acid. In addition tothe inhibition of angiogenesis, preclinical evidence indicatesthat bisphosphonates can impede cancer cell invasion andattachment and stimulate specific subsets of immune cells.

Although clinical data are limited regarding the efficacyof bisphosphonates to prevent tumour metastasis to boneand inhibit disease progression through antitumour activ-ity, the results from at least six different ongoing studieswill provide more conclusive information on the use of

bisphosphonates in this setting, and interim results in thebreast cancer setting are anticipated in 2008.

Conflict of Interest Statement

Dr. Coleman has received honoraria from and acted as aconsultant for Novartis Pharmaceuticals Corporation.

Role of funding source

This article was supported by an unrestricted edu-cational grant provided by Novartis PharmaceuticalsCorporation.

Acknowledgements

Financial support for medical editorial assistance wasprovided by Novartis Pharmaceuticals Corporation. Wethank Catherine Browning, PhD, ProEd Communications,Inc.,s for her medical editorial assistance with thismanuscript.

References

1. Coleman RE. Metastatic bone disease: clinical features, pathophysiol-

ogy and treatment strategies. Cancer Treat Rev 2001;27:165–76.

2. Hei YJ, Saad F, Coleman RE, Chen YM. Fractures negatively affect

survival in patients with bone metastases from breast cancer [poster].

Presented at: 28th Annual San Antonio Breast Cancer Symposium;

December 8–11, 2005; San Antonio, Texas. Abstract 6036.

3. Gnant MF, Mlineritsch B, Luschin-Ebengreuth G, et al. Zoledronic

acid prevents cancer treatment-induced bone loss in premenopausal

women receiving adjuvant endocrine therapy for hormone-responsive

breast cancer: a report from the Austrian Breast and Colorectal

Cancer Study Group. J Clin Oncol 2007;25:820–8.

4. Green JR. Bisphosphonates: preclinical review. Oncologist 2004;

9(suppl 4):3–13.

5. Peyruchaud O, Winding B, Pecheur I, Serre CM, Delmas P,

Clezardin P. Early detection of bone metastases in a murine model

ARTICLE IN PRESSR. Coleman / The Breast 16 (2007) 21–27S26

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;16:2027–34.

6. Body JJ, Mancini I. Bisphosphonates for cancer patients: why, how,

and when? Support Care Cancer 2002;10:399–407.

7. Saad F, Schulman CC. Role of bisphosphonates in prostate cancer.

Eur Urol 2004;45:26–34.

8. Neville-Webbe HL, Rostami-Hodjegan A, Evans CA, Coleman RE,

Holen I. Sequence- and schedule-dependent enhancement of zole-

dronic acid induced apoptosis by doxorubicin in breast and prostate

cancer cells. Int J Cancer 2005;113:364–71.

9. Neville-Webbe HL, Evans CA, Coleman RE, Holen I. Mechanisms of

the synergistic interaction between the bisphosphonate zoledronic acid

and the chemotherapy agent paclitaxel in breast cancer cells in vitro.

Tumour Biol 2006;27:92–103.

10. Hiraga T, Williams PJ, Ueda A, Tamura D, Yoneda T. Zoledronic

acid inhibits visceral metastases in the 4T1/luc mouse breast cancer

model. Clin Cancer Res 2004;10:4559–67.

11. Ottewell PD, Jones M, Coleman RE, Holen I. Synergistic effects of

cytotoxic drugs and anti-resorptive agents in vitro and in vivo [poster].

Presented at: 29th Annual San Antonio Breast Cancer Symposium;

December 14–17, 2006; San Antonio, Texas. Abstract 6102.

12. Croucher PI, De Hendrik R, 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;18:482–92.

13. 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;75:249–58.

14. Yoneda T, Michigami T, Yi B, Williams PJ, Niewolna M, Hiraga T.

Actions of bisphosphonate on bone metastasis in animal models of

breast carcinoma. Cancer 2000;88:2979–88.

15. 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;62:6538–44.

16. Budman DR, Calabro A. Zoledronic acid (Zometa) enhances the

cytotoxic effect of gemcitabine and fluvastatin: in vitro isobologram

studies with conventional and nonconventional cytotoxic agents.

Oncology 2006;70:147–53.

17. Zimering MB. Effect of intravenous bisphosphonates on release of

basic fibroblast growth factor in serum of patients with cancer-

associated hypercalcemia. Life Sci 2002;70:1947–60.

18. Heikkila P, Teronen O, Hirn MY, et al. Inhibition of matrix

metalloproteinase-14 in osteosarcoma cells by clodronate. J Surg

Res 2003;111:45–52.

19. Wood J, Bonjean K, Ruetz S, et al. Novel antiangiogenic effects of the

bisphosphonate compound zoledronic acid. J Pharmacol Exp Ther

2002;302:1055–61.

20. Bezzi M, Hasmim M, Bieler G, Dormond O, Ruegg C. Zoledronate

sensitizes endothelial cells to tumor necrosis factor-induced pro-

grammed cell death: evidence for the suppression of sustained

activation of focal adhesion kinase and protein kinase B/Akt. J Biol

Chem 2003;278:43603–14.

21. Vincenzi B, Santini D, Dicuonzo G, et al. Zoledronic acid-related

angiogenesis modifications and survival in advanced breast cancer

patients. J Interferon Cytokine Res 2005;25:144–51.

22. Guise TA, Mundy GR. Cancer and bone. Endocr Rev 1998;19:18–54.

23. Dillon C, Spencer-Dene B, Dickson C. A crucial role for fibroblast

growth factor signaling in embryonic mammary gland development.

J Mammary Gland Biol Neoplasia 2004;9:207–15.

24. Santini D, Vincenzi B, Avvisati G, et al. Pamidronate induces

modifications of circulating angiogenetic factors in cancer patients.

Clin Cancer Res 2002;8:1080–4.

25. 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;9:2893–7.

26. Santini D, Vincenzi B, Battistoni F, et al. In vivo prospective study

about the effects of weekly low dose administration of zoledronic acid

(ZA) on angiogenesis [abstract]. J Clin Oncol 2007;25(suppl):152 s

Abstract 3558.

27. 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;60:2949–54.

28. 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;57:3890–4.

29. Berger W, Kubista B, Elbling L, Sutterluty H, Micksche M. The N-

containing bisphosphonate zoledronic acid exerts potent anticancer

activity against non-small cell lung cancer cells by inhibition of protein

geranylgeranylation [poster]. Presented at: 96th American Association

for Cancer Research Annual Meeting; April 16–20, 2005; Anaheim/

Orange County, California. Abstract 4981.

30. Heikkila P, Teronen O, Moilanen M, et al. Bisphosphonates inhibit

stromelysin-1 (MMP-3), matrix metalloelastase (MMP-12), collage-

nase-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;

13:245–54.

31. Leto G, Badalamenti G, Arcara C, et al. Effects of zoledronic acid on

proteinase plasma levels in patients with bone metastases. Anticancer

Res 2006;26:23–6.

32. Clezardin P. Anti-tumour activity of zoledronic acid. Cancer Treat

Rev 2005;31(suppl 3):1–8.

33. Ural AU, Avcu F. Additive/synergistic anti-tumoral effects of the

combination of docetaxel and zoledronic acid on prostate cancer cells:

possible mechanisms? Acta Oncol 2006;45:491–2.

34. Ural AU, Avcu F, Candir M, Guden M, Ozcan MA, In vitro

synergistic cytoreductive effects of zoledronic acid and radiation on

breast cancer cells, Breast Cancer Res 2006;8:R52. Doi:10.1186/

bcr543.

35. Green JR. Antitumor effects of bisphosphonates. Cancer 2003;

97:840–7.

36. Conte P, Guarneri V. Safety of intravenous and oral bisphosphonates

and compliance with dosing regimens. Oncologist 2004;9(suppl 4):

28–37.

37. Pecherstorfer M, Jilch R, Sauty A, et al. Effect of first treatment with

aminobisphosphonates pamidronate and ibandronate on circulating

lymphocyte subpopulations. J Bone Miner Res 2000;15:147–54.

38. Sansoni P, Passeri G, Fagnoni F, et al. Inhibition of antigen-

presenting cell function by alendronate in vitro. J Bone Miner Res

1995;10:1719–25.

39. Thompson K, Rogers MJ. Bisphosphonates and gamma-delta T-cells:

New insights into old drugs. BoneKEy-Osteovision 2006;3:5–13.

40. Sato K, Kimura S, Segawa H, et al. Cytotoxic effects of gammadelta T

cells expanded ex vivo by a third generation bisphosphonate for

cancer immunotherapy. Int J Cancer 2005;116:94–9.

41. Das H, Wang L, Kamath A, Bukowski JF. Vgamma2Vdelta2 T-cell

receptor-mediated recognition of aminobisphosphonates. Blood 2001;

98:1616–8.

42. van der Pluijm G, Que I, Sijmons B, et al. Interference with the

microenvironmental support impairs the de novo formation of bone

metastases in vivo. Cancer Res 2005;65:7682–90.

43. Padalecki SS, Carreon M, Grubbs B, Cui Y, Guise TA. Androgen

deprivation enhances bone loss and prostate cancer metastases to

bone: prevention by zoledronic acid. Oncology (Huntington) 2003;

17(suppl 4):32.

44. Diel IJ, Solomayer EF, Costa SD, et al. Reduction in new metastases

in breast cancer with adjuvant clodronate treatment. N Engl J Med

1998;339:357–63.

45. Powles T, Paterson A, McCloskey E, et al. Reduction in bone relapse

and improved survival with oral clodronate for adjuvant treatment of

operable breast cancer. ISRCTN83688026] Breast Cancer Res 2006;8:

R13. Doi:0.1186/bcr384.

ARTICLE IN PRESSR. Coleman / The Breast 16 (2007) 21–27 S27

46. Saarto T, Vehmanen L, Virkkunen P, Blomqvist C. Ten-year

follow-up of a randomized controlled trial of adjuvant clodronate

treatment in node-positive breast cancer patients. Acta Oncol 2004;43:

650–6.

47. Mystakidou K, Katsouda E, Parpa E, Kelekis A, Galanos A, Vlahos

L. Randomized, open label, prospective study on the effect of

zoledronic acid on the prevention of bone metastases in patients with

recurrent solid tumors that did not present with bone metastases at

baseline. Med Oncol 2005;22:195–201.

48. Rack W, Janni A, Schoberth C, et al. Effect of zoled-

ronate on persisting isolated tumor cells (ITC) in the bone

marrow (BM) of patients without recurrence of early breast cancer

[abstract]. Proc Am Soc Clin Oncol 2004;23(suppl):834 Abstract

9515.

49. US National Institutes of Health. Clinical trial: Chemotherapy and/or

hormone therapy with or without zoledronate in treating women

with stage II or stage III breast cancer. Available at: http://

clinicaltrials.gov/ct/show/NCT00072020?order=1. Accessed Septem-

ber 12, 2007.

50. Coleman R, Thorpe H, Cameron D, et al. Zoledronic acid is well

tolerated and can be safely administered with adjuvant chemother-

apy—first safety data from the AZURE trial (BIG01/04) [poster].

Presented at: 29th Annual San Antonio Breast Cancer Symposium;

San Antonio, Texas; December 14–17, 2006. Abstract 2080.

51. US National Institutes of Health. Clinical trial: Clodronate with or

without chemotherapy and/or hormonal therapy in treating women

with stage I or stage II breast cancer. Available at: http://

clinicaltrials.gov/ct/show/NCT00009945?order=1. Accessed Septem-

ber 12, 2007.

52. US National Institutes of Health. Clinical trial: Zoledronate,

clodronate, or ibandronate in treating women who have undergone

surgery for stage I, stage II, or stage III breast cancer. Available

at: http://clinicaltrials.gov/ct/show/NCT00127205?order=1. Accessed

September 12, 2007.

53. US National Institutes of Health. Clinical trial: A study to evaluate

the safety and efficacy of zoledronic acid in the prevention or delaying

of bone metastasis in patients with stage IIIA and IIIB non-small

cell lung cancer. Available at: http://clinicaltrials.gov/ct/show/

NCT00172042?order=1. Accessed September 12, 2007.

54. Nederlands Trial Register. Trial information: ZEUS study. Available

at: www.trialregister.nl/trialreg/admin/rctview.asp?TC=355. Accessed

September 12, 2007.