bisphosphonates: mode of action and pharmacologyof free, non–mineral-bound bisphosphonate as a...

SUPPLEMENT ARTICLE

Bisphosphonates: Mode of Action and PharmacologyR. Graham G. Russell, MD, PhD

Botnar Research Centre and Oxford University Institute of Musculoskeletal Sciences, Oxford, United Kingdom

The author has indicated he has no financial relationships relevant to this article to disclose.

ABSTRACT

The profound effects of the bisphosphonates on calcium metabolism were discov-ered over 30 years ago, and they are now well established as the major drugs usedfor the treatment of bone diseases associated with excessive resorption. Theirprincipal uses are for Paget disease of bone, myeloma, bone metastases, andosteoporosis in adults, but there has been increasing and successful application inpediatric bone diseases, notably osteogenesis imperfecta. Bisphosphonates arestructural analogues of inorganic pyrophosphate but are resistant to enzymatic andchemical breakdown. Bisphosphonates inhibit bone resorption by selective ad-sorption to mineral surfaces and subsequent internalization by bone-resorbingosteoclasts where they interfere with various biochemical processes. The simpler,non–nitrogen-containing bisphosphonates (eg, clodronate and etidronate) can bemetabolically incorporated into nonhydrolysable analogues of adenosine triphos-phate (ATP) that may inhibit ATP-dependent intracellular enzymes. In contrast,the more potent, nitrogen-containing bisphosphonates (eg, pamidronate, alendro-nate, risedronate, ibandronate, and zoledronate) inhibit a key enzyme, farnesylpyrophosphate synthase, in the mevalonate pathway, thereby preventing thebiosynthesis of isoprenoid compounds that are essential for the posttranslationalmodification of small guanosine triphosphate (GTP)-binding proteins (which arealso GTPases) such as Rab, Rho, and Rac. The inhibition of protein prenylation andthe disruption of the function of these key regulatory proteins explains the loss ofosteoclast activity. The recently elucidated crystal structure of farnesyl diphosphatereveals how bisphosphonates bind to and inhibit at the active site via their criticalnitrogen atoms. Although bisphosphonates are now established as an importantclass of drugs for the treatment of many bone diseases, there is new knowledgeabout how they work and the subtle but potentially important differences thatexist between individual bisphosphonates. Understanding these may help to ex-plain differences in potency, onset and duration of action, and clinicaleffectiveness.

www.pediatrics.org/cgi/doi/10.1542/peds.2006-2023H

doi:10.1542/peds.2006-2023H

KeyWordsbone resorption, osteoclasts,bisphosphonates, osteoporosis

AbbreviationsPPi—inorganic pyrophosphateATP—adenosine triphosphateFPP—farnesyl diphosphateGGPP—geranylgeranyl diphosphateFPPS—farnesyl pyrophosphate synthase

Accepted for publication Oct 5, 2006

Address correspondence to R. Graham G.Russell, MD, PhD, Botnar Research Centre,Nuffield Department of Orthopaedic Surgery,University of Oxford, Headington, Oxford OX37LD, United Kingdom. E-mail: [email protected]

PEDIATRICS (ISSN Numbers: Print, 0031-4005;Online, 1098-4275); published in the publicdomain by the American Academy ofPediatrics

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THE DISCOVERY AND development of the bisphospho-nates as a major class of drugs for the treatment of

bone diseases was recently reviewed1 and represents afascinating story that has its origins in studies of biolog-ical calcification processes. There are many books andreview articles available that describe the chemistry,pharmacology, and clinical applications of bisphospho-nates.2–5

It had been known since the 1930s that traceamounts of polyphosphates were capable of acting aswater softeners by inhibiting the crystallization of cal-cium salts, such as calcium carbonate,6 and in the 1960sFleisch et al7,8 showed that inorganic pyrophosphate, anaturally occurring polyphosphate and a known byprod-uct of many biosynthetic reactions in the body, waspresent in serum and urine and could prevent calcifica-tion by binding to newly forming crystals of hydroxyap-atite. It was proposed that inorganic pyrophosphate (PPi)might be the body’s own natural “water softener” thatnormally prevents calcification of soft tissues and regu-lates bone mineralization. It subsequently became clearthat calcification disorders might be linked to distur-bances in PPi metabolism. The first example was aninherited disorder, hypophosphatasia, in which lack ofalkaline phosphatase is associated with mineralizationdefects of the skeleton and elevated PPi levels,9 indicat-ing that alkaline phosphatase is probably the key extra-cellular enzyme responsible for hydrolyzing pyrophos-phate.

Attempts to exploit these concepts by using pyro-phosphate and polyphosphates to inhibit ectopic calcifi-cation in blood vessels, skin, and kidneys in laboratoryanimals were successful only when the compounds wereinjected.10 Orally administered pyrophosphate andpolyphosphates were inactive because of their hydrolysisin the gastrointestinal tract. During the search for morestable analogues of pyrophosphate that might also havethe antimineralization properties of pyrophosphate butwould be resistant to hydrolysis, several different chem-ical classes were studied. The bisphosphonates (at thattime called diphosphonates), characterized by P-C-P mo-tifs, were among these classes.11–13

Like pyrophosphate, bisphosphonates had high affin-ity for bone mineral14 and were found to prevent calci-fication both in vitro and in vivo but, unlike pyrophos-phate, were also able to prevent experimentally inducedpathologic calcification when given orally to rats invivo.15 This property of being active by mouth was key totheir future use in humans.

In these early studies bisphosphonates were shownnot only to prevent the experimentally induced calcifi-cation of many soft tissues, including skin, kidneys, andblood vessels in vivo but, with some of the compounds(eg, etidronate), to also inhibit mineralization of ectopicbone as well of normal calcified tissues such as bone andcartilage.16 Bisphosphonates seem to prevent calcifica-

tion by physicochemical mechanisms that produce directimpairment of the calcification process by acting as crys-tal poisons after adsorption to mineral surfaces ratherthan by effects on the deposition of matrix.

Perhaps the most important step toward the futureuse of bisphosphonates occurred when we found thatbisphosphonates, as we had already shown for PPi,17 alsohad the novel property of being able to inhibit the dis-solution of hydroxyapatite crystals.18 This finding led tostudies to determine if they might also inhibit boneresorption, which they did in many different experimen-tal models.19,20 In growing intact rats, the bisphospho-nates block the removal of both bone and cartilage, thusretarding the modeling of the metaphysis, which be-comes club-shaped and radiologically denser than nor-mal. This effect is the basis of the Schenk model21 and isa phenomenon of interest in pediatrics because it is alsoobserved in children who are treated with high doses ofbisphosphonates.

The bisphosphonates are also effective in preventingbone destruction in a number of animal models of hu-man disease, such as immobilization osteoporosis,22 andthe prevention of bone loss associated with ovariectomy.If not given in excess, bisphosphonates do not impairbone growth and can maintain or improve the biome-chanical properties of bone in both normal animals andexperimental models of osteoporosis.23

In general, there is a good correlation between po-tency and structure-activity relationships in vitro and invivo.24 In the presence of bisphosphonates, isolated os-teoclasts form fewer and smaller erosion cavities on var-ious mineralized matrices in vitro.25

PHARMACOLOGY AND CELLULAR ACTIONSEtidronate was the first bisphosphonate to be used inhumans for fibrodysplasia ossificans progressiva andPaget disease.26 Once the potential clinical value ofbisphosphonates had been appreciated, research effortswere devoted to the development of compounds with amore powerful antiresorptive activity but without a cor-responding ability to inhibit mineralization. With com-pounds such as etidronate there was only a 10- to 100-fold difference between doses that inhibit mineralizationcompared with doses that reduce bone resorption. En-hancing this window was readily achieved, and manyhundreds of bisphosphonates have been synthesized;more than a dozen have been used in humans. With thedevelopment of bisphosphonates that were more potentinhibitors of bone resorption, these dose differences wid-ened to several orders of magnitude, which meant thatinhibition of skeletal mineralization observed with etidr-onate ceased to be a major clinical concern. The grada-tion of potency evaluated in the animal models corre-sponded quite well with that found in humans, althoughthe differences in potency are much smaller in humans.

Bisphosphonates accumulate in bone, so it is impor-

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tant to know what happens during long-term adminis-tration. From a clinical point of view, it is reassuring thatthe inhibition of bone resorption reaches a new steady-state level rather than becoming progressively lower,even when the compounds are given continuously.27

The level of suppression depends on the administereddose and has also been observed in humans.28 Thereseems to be no progression of the antiresorptive effectwith time, which suggests that the bisphosphonate bur-ied in the bone is inactive for at least as long as it remainsburied there. This also means that within the therapeu-tic-dosage range, there is little risk of a continuous andprogressive decrease in bone turnover in the long runthat might lead to an increase in bone fragility. Anadditional important pharmacologic property of bisphos-phonates is that the total dose administered is a majordeterminant of their effects. This has been well studiedfor ibandronate29 and zoledronate.30 In both cases thesame inhibition of bone resorption has been docu-mented regardless of whether the bisphosphonate wasgiven in small frequent (eg, daily) doses compared withlarger doses given less frequently. This was the basis forthe development of intermittent-dosing regimens in hu-mans.

The pronounced selectivity of bisphosphonates forbone rather than other tissues is the basis for their valuein clinical practice. Their preferential uptake by andadsorption to mineral surfaces in bone bring them intoclose contact with osteoclasts. During bone resorption,bisphosphonates are probably internalized by endocyto-sis along with other products of resorption. Many studieshave shown that bisphosphonates can affect osteoclast-mediated bone resorption in a variety of ways, includingeffects on osteoclast recruitment, differentiation, and re-sorptive activity, and may induce apoptosis.

Because mature, multinucleated osteoclasts areformed by the fusion of mononuclear precursors of he-matopoietic origin, bisphosphonates could also inhibitbone resorption by preventing osteoclast formation, inaddition to affecting mature osteoclasts. In vitro,bisphosphonates can inhibit dose-dependently the for-mation of osteoclast-like cells in long-term cultures ofhuman bone marrow.31 In organ culture, also, somebisphosphonates can inhibit the generation of matureosteoclasts, possibly by preventing the fusion of oste-oclast precursors.32,33

It is likely that bisphosphonates are selectively inter-nalized by osteoclasts rather than other cell types be-cause of their accumulation in bone and the endocyticactivity of osteoclasts. During the process of bone resorp-tion, the subcellular space beneath the osteoclast is acid-ified by the action of vacuolar-type proton pumps in theruffled border of the osteoclast membrane.34 The acidicpH of this microenvironment causes dissolution of thehydroxyapatite bone mineral, whereas the breakdownof the extracellular bone matrix is brought about by the

action of proteolytic enzymes, including cathepsin K.Because bisphosphonates adsorb to bone mineral, espe-cially at sites of bone resorption where the mineral ismost exposed,35,36 osteoclasts are the cell type in bonemost likely to be exposed to the highest concentrationsof free, non–mineral-bound bisphosphonate as a resultof the release of the bisphosphonate from bone mineralin the low-pH environment beneath osteoclasts. It hasbeen estimated that pharmacologic doses of alendronatethat inhibit bone resorption in vivo could give rise tolocal concentrations as high as 1 mM alendronate in theresorption space beneath an osteoclast, which is muchhigher than the concentrations of bisphosphonates re-quired to affect osteoclast morphology and cause oste-oclast apoptosis in vitro.37

In contrast to their ability to induce apoptosis in os-teoclasts, which contributes to the inhibition of resorp-tive activity, some experimental studies suggest thatbisphosphonates may protect osteocytes and osteoblastsfrom apoptosis induced by glucocorticoids.38 Recent ev-idence suggests that the inhibition of osteocyte apoptosisby bisphosphonates is mediated through the opening ofconnexion 43 hemichannels and activation of extracel-lular signal-regulated kinases.39 The possibility thatbisphosphonates used clinically may get access to osteo-cytes differentially depending on their mineral-bindingaffinities and inherent structural properties needs to bestudied.

STRUCTURE-ACTIVITY RELATIONSHIPS ANDMECHANISMOFACTIONThe features of the bisphosphonate molecule necessaryfor biological activity were well defined in the earlystudies. The P-C-P moiety is responsible for the strongaffinity of the bisphosphonates for binding to hydroxy-apatite and allows for a number of variations in structureon the basis of substitution in the R1 and R2 positions onthe carbon atom (Fig 1). The ability of the bisphospho-nates to bind to hydroxyapatite crystals and to preventboth crystal growth and dissolution was enhanced whenthe R1 side chain (attached to the geminal carbon atomof the P-C-P group) was a hydroxyl group (as in eti-dronate) rather than a halogen atom such as chlorine (asin clodronate). The presence of a hydroxyl group at theR1 position increases the affinity for calcium (and, thus,bone mineral) because of the ability of bisphosphonatesto chelate calcium ions by tridentate rather than biden-tate binding.40

The ability of bisphosphonates to inhibit bone resorp-tion in vitro and in vivo also requires the P-C-P struc-ture. Monophosphonates (eg, pentane monophospho-nate) or P-C-C-P or P-N-P compounds are ineffective asinhibitors of bone resorption. Furthermore, the antire-sorptive effect cannot be accounted for simply by adsorp-tion of bisphosphonates to bone mineral and preventionof hydroxyapatite dissolution. It became clear that


bisphosphonates must inhibit bone resorption by cellulareffects on osteoclasts rather than simply by physico-chemical mechanisms.

After the successful clinical use of clodronate andetidronate in the 1970s and 1980s, more potent antire-sorptive bisphosphonates, which had different R2 sidechains but in which R1 was unaltered, were studied. Inparticular, bisphosphonates containing a basic primaryamino-nitrogen atom in an alkyl chain (as in pamidr-onate and alendronate) were found to be 10- to 100-foldmore potent than etidronate and clodronate. Then, inthe 1980s, there was a phase in which synthesis of novelcompounds took place specifically to determine theirpossible effects on calcium metabolism, with the resultthat compounds highly effective as inhibitors of boneresorption were identified and studied.

These compounds, especially those that contain a ter-tiary amino-nitrogen (such as ibandronate41 and olpad-ronate42), were even more potent at inhibiting boneresorption. Among this generation of compounds that

were synthesized to optimize their antiresorptive effects,the most potent antiresorptive bisphosphonates werethose containing a nitrogen atom within a heterocyclicring (as in risedronate43 and zoledronate44), which are upto 10 000-fold more potent than etidronate in someexperimental systems (Fig 2).

The analysis of structure-activity relationships al-lowed the spatial features of the active pharmacophoreto be defined in considerable detail even before themolecular mechanism of action was fully elucidated. Formaximal potency, the nitrogen atom in the R2 side chainmust be a critical distance away from the P-C-P groupand in a specific spatial configuration.45 This principlewas used successfully for predicting the features requiredin the chemical design of new and more active com-pounds.

Although the structure of the R2 side chain is themajor determinant of antiresorptive potency, both phos-phonate groups are also required for the drugs to bepharmacologically active.

FIGURE 1The generic structure of pyrophosphate comparedwith bisphos-phonates and their functional domains.

FIGURE 2Structures of the bisphosphonates used in clinical studies classi-fied according to their biochemical mode of action.


In summary, studies of the relationships betweenbisphosphonate structure and antiresorptive potencysuggested that the ability of bisphosphonates to inhibitbone resorption depend on 2 separate properties of thebisphosphonate molecule. The 2 phosphonate groups,together with a hydroxyl group at the R1 position,46

impart high affinity for bone mineral and act as a “bonehook,” which allows rapid and efficient targeting ofbisphosphonates to bone mineral surfaces. Once local-ized within bone, the structure and three-dimensionalconformation of the R2 side chain (as well as the phos-phonate groups in the molecule) determine the biolog-ical activity of the molecule and influence the ability ofthe drugs to interact with specific molecular targets. Ourunderstanding of what these molecular targets might behas become much clearer as a result of recent work.

Over the years there have been many efforts toexplain how bisphosphonates work on cells, especiallyvia inhibitory effects on enzymes (eg, by direct or in-direct inhibition of the osteoclast proton-pumpingH�ATPase,47,48 phosphatases, or lysosomal enzymes49,50).

Because osteoclasts are highly endocytic, bisphospho-nate present in the resorption space is likely to be inter-nalized by endocytosis and thereby affect osteoclasts di-rectly.25 The uptake of bisphosphonates by osteoclasts invivo has been confirmed by using radiolabeled51 andfluorescently labeled alendronate, which was internal-ized into intracellular vacuoles. After cellular uptake, acharacteristic morphologic feature of bisphosphonate-treated osteoclasts is the lack of a ruffled border, theregion of invaginated plasma membrane facing the re-sorption cavity. Bisphosphonates also disrupt the cy-toskeleton of the osteoclast.52 Early explanations forthese effects invoked the inhibition of protein kinases orphosphatases that regulate cytoskeletal structure, suchas protein tyrosine phosphatases.53–56 However, a morelikely mechanism by which the cytoskeleton may beaffected involves loss of function of small GTPases suchas Rho and Rac.

Since the early 1990s there has been a systematiceffort by our group and others to elucidate the molecularmechanisms of action of bisphosphonates,5,57,58 and wehave proposed that bisphosphonates can be classified

into at least 2 major groups with different modes ofaction (Fig 3). The first group comprises the non–nitro-gen-containing bisphosphonates that perhaps mostclosely resemble pyrophosphate, such as clodronate andetidronate, and these can be metabolically incorporatedinto nonhydrolyzable analogues of adenosine triphos-phate (ATP) by reversing the reactions of aminoacyl–transfer RNA synthetases.59 The resulting metabolitescontain the P-C-P moiety in place of the �,�-phosphategroups of ATP, thus resulting in nonhydrolyzable(AppCp) nucleotides.60–63 It is likely that intracellularaccumulation of these metabolites within osteoclasts in-hibits their function and may cause osteoclast cell death.The AppCp-type metabolites of bisphosphonates are cy-totoxic when internalized and cause similar changes inmorphology to those observed in clodronate-treatedcells, possibly by interference with mitochondrial ATPtranslocases.64 Overall, this group of bisphosphonates,therefore, seem to act as prodrugs, being converted toactive metabolites after intracellular uptake by oste-oclasts in vivo.

In contrast, the second group contains the more po-tent, nitrogen-containing bisphosphonates such as alen-dronate, risedronate, and zoledronate. Members of thisgroup interfere with other metabolic reactions, notablyin the mevalonate biosynthetic pathway, and affect cel-lular activity and cell survival by interfering with proteinprenylation and, therefore, the signaling functions ofkey regulatory proteins. These mechanisms have beenreviewed in detail elsewhere.1 The mevalonate pathwayis a biosynthetic route responsible for the production ofcholesterol, other sterols, and isoprenoid lipids such asisopentenyl diphosphate (also known as isopentenyl py-rophosphate), as well as farnesyl diphosphate (FPP) andgeranylgeranyl diphosphate (GGPP). FPP and GGPP arerequired for the posttranslational modification (prenyla-tion) of small GTPases such as Ras, Rab, Rho, and Rac,which are prenylated at a cysteine residue in character-istic C-terminal motifs.65 Small GTPases are importantsignaling proteins that regulate a variety of cell processesimportant for osteoclast function, including cell mor-phology, cytoskeletal arrangement, membrane ruffling,trafficking of vesicles, and apoptosis.66–69 Prenylation is

FIGURE 3Two distinct molecular mechanisms of action of bisphospho-nates (BPs) that both inhibit osteoclasts (see text for details). GTPindicates guanosine triphosphate.


required for the correct function of these proteins be-cause the lipid prenyl group serves to anchor the pro-teins in cell membranes and may also participate inprotein-protein interactions.70

Many observations point to the importance of themevalonate pathway for osteoclast function andstrengthen the proposal that the nitrogen-containingbisphosphonates inhibit osteoclastic bone resorptionpredominantly by inhibition of this pathway. Thesebisphosphonates inhibit the synthesis of mevalonate me-tabolites including FPP and GGPP, and thereby impairthe prenylation of proteins,71 and cause alteration offunction of small GTPases. There is a strong structure-activity relationship such that changes to the structure ofthe nitrogen-containing R2 side chain or to the phospho-nate groups, which alter antiresorptive potency, alsoinfluence the ability to inhibit protein prenylation to acorresponding degree.72 An important verification of thecritical importance of this pathway has come from show-ing that the addition of intermediates of the mevalonatepathway (such as FPP and GGPP) could overcomebisphosphonate-induced apoptosis and other events inmany cell systems. Another prediction was that if inhi-bition of the mevalonate pathway could account for theantiresorptive effects of bisphosphonates, then the statindrugs should also inhibit bone resorption. Statins areinhibitors of 3-hydroxy-3-methylglutaryl coenzyme A(HMG-CoA) reductase, one of the first steps in the me-valonate pathway. They proved to be even more potentthan bisphosphonates at inhibiting osteoclast formationand bone resorption in vitro,73,74 an effect that could alsobe overcome by the addition of geranylgeraniol (whichcan be used for protein geranylgeranylation) but notfarnesol (which is used for protein farnesylation). Hence,it seems that although nitrogen-containing bisphospho-nates can prevent both farnesylation and geranylgera-nylation of proteins (probably by inhibiting enzymesrequired for synthesis of FPP and GGPP), loss of gera-nylgeranylated proteins in osteoclasts is of greater con-sequence than loss of farnesylated proteins. This is con-sistent with the known role of geranylgeranylatedproteins such as Rho, Rac, and Rab in processes that arefundamental to osteoclast formation and function (eg,cytoskeletal rearrangement, membrane ruffling, and ve-sicular trafficking75), and further work has confirmedthis, particularly the importance of Rab proteins.

The comparison between bisphosphonates and statinsis interesting. The statins are widely used as cholesterol-lowering drugs (they are able to lower cholesterol bio-synthesis by inhibiting 3-hydroxy-3-methylglutaryl co-enzyme A reductase). Despite several studies, there is nosubstantial evidence that statins have effects on bonewhen used clinically, perhaps because they are selec-tively taken up by the liver rather than bone, which isthe converse of the case for bisphosphonates. Therefore,

this is an excellent example of how drug specificity isachieved by highly selective tissue targeting.

The exact enzymes of the mevalonate pathway thatare inhibited by individual bisphosphonates have beenpartially elucidated. Several enzymes of the pathway useisoprenoid diphosphates as a substrate (isopentenyl py-rophosphate isomerase, FPP synthase, GGPP synthase,squalene synthase) and, thus, are likely to have similarsubstrate-binding sites. Thus, if nitrogen-containingbisphosphonates act as substrate analogues of an isopre-noid diphosphate, it is possible that these bisphospho-nates will inhibit more than 1 of the enzymes of themevalonate pathway. Early studies revealed that incad-ronate and ibandronate, but not other bisphosphonates,are inhibitors of squalene synthase, an enzyme in themevalonate pathway that is required for cholesterol bio-synthesis.76,77 Inhibition of squalene synthase, however,would not lead to inhibition of protein prenylation.

However, it is now clear that farnesyl pyrophosphatesynthase (FPPS) is a major site of action of the nitrogen-containing bisphosphonates (N-bisphosphonates).78

FPPS catalyzes the successive condensation of isopente-nyl pyrophosphate with dimethylallyl pyrophosphateand geranyl pyrophosphate. There is a strong relation-ship among individual bisphosphonates between inhibi-tion of bone resorption and inhibition of FPPS, with themost potent bisphosphonates having IC50 values (con-centration that inhibits response by 50%) in the nano-molar range.79 Modeling studies have provided a molec-ular rationale for bisphosphonate binding to FPPS.80 Ourrecent studies using protein crystallography, enzyme ki-netics, and isothermal titration calorimetry led to thefirst published high-resolution radiograph structures ofthe human enzyme in complexes with risedronate andzoledronate.81 These agents bind to the dimethylallyl/geranyl pyrophosphate ligand pocket and induce a con-formational change. The interactions of the N-bisphos-phonate cyclic nitrogen with Thr201 and Lys200 suggestthat these inhibitors achieve potency by positioning theirnitrogen in a proposed carbocation82 binding site. Thisexplains how the nitrogen moiety is so important to thepotency of these bisphosphonates. Kinetic analyses re-veal that inhibition is competitive with geranyl pyro-phosphate and is of a slow, tight-binding character,which indicates that isomerization of an initial enzyme-inhibitor complex occurs after binding of the N-bisphos-phonate.

Taken together, these observations clearly indicatethat bisphosphonates can be grouped into 2 classes:those that can be metabolized into nonhydrolyzable an-alogues of ATP (the least potent bisphosphonates) andthose that are not metabolized but can inhibit proteinprenylation (the potent, nitrogen-containing bisphos-phonates). The identification of 2 such classes may helpto explain some of the other pharmacologic differencesbetween the 2 classes.


CLINICAL APPLICATIONS OF BISPHOSPHONATESAfter it was shown that bisphosphonates inhibited ex-perimentally induced calcification and bone resorption,their potential application to clinical disorders was obvi-ous, but it took many years for them to become wellestablished.

The earliest clinical applications of bisphosphonatesincluded use of etidronate as an inhibitor of calcificationin fibrodysplasia ossificans progressiva (formerly knownas myositis ossificans) and in patients who had under-gone total hip replacement surgery to prevent subse-quent heterotopic ossification and improve mobility.83

One of the other early clinical uses of bisphospho-nates was as agents for bone imaging, “bone scanning,”for which they still remain outstandingly useful for de-tecting bone metastases and other bone lesions. Theapplication of pyrophosphate and simple bisphospho-nates as bone-scanning agents depends on their strongaffinity for bone mineral, particularly at sites of in-creased bone turnover, and their ability to be linked to a�-emitting technetium isotope.84,85

The most impressive clinical application of bisphos-phonates has been as inhibitors of bone resorption, es-pecially for diseases in which no effective treatmentexisted previously. Thus, bisphosphonates became thetreatment of choice for a variety of bone diseases inwhich excessive osteoclast activity is an importantpathologic feature, including Paget disease of bone, met-astatic and osteolytic bone disease, and hypercalcemia ofmalignancy, as well as osteoporosis.

The clinical pharmacology of bisphosphonates is char-acterized by low intestinal absorption (�1%–4%) buthighly selective localization and retention in bone. Sig-nificant adverse effects of bisphosphonates are mini-mal.86–88 Although there are more similarities than dif-ferences between individual compounds and eachbisphosphonate is potentially capable of treating any ofthe disorders of bone resorption in which they are used,in practice different compounds have come to be favoredfor the treatment of different diseases. There are cur-rently at least 10 bisphosphonates (etidronate, clodr-onate, tiludronate, pamidronate, alendronate, risedr-onate, zoledronate, and ibandronate and, to a limitedextent, olpadronate and neridronate) that have beenregistered for various clinical applications in variouscountries. To a major extent, the diseases in which theyare used reflects the history of their clinical developmentand the degree of commercial interest in and sponsor-ship of the relevant clinical trials.

Paget disease was the first clinical disorder in which adose-dependent inhibition of bone resorption could bedemonstrated by using bisphosphonates in humans.89,90

Bisphosphonates have become the most important drugsused in the treatment of Paget disease.91 For many yearspamidronate given by intravenous infusion was usedextensively,92 but the newer and more potent bisphos-

phonates can produce even more profound suppressionof disease activity than was possible with the bisphos-phonates available in previous years.93,94 The latest ad-vance is with zoledronate,95 which, when given as asingle 5-mg infusion, produced a greater and longer-lasting suppression of excess bone turnover than evenoral risedronate given at 30 mg/day over 2 months,hitherto one of the most effective treatments.

In terms of commercial success, the use of bisphos-phonates in oncology has been preeminent. Many can-cers in humans are associated with hypercalcemia(raised blood calcium) and/or increased bone destruc-tion. Bisphosphonates are remarkably effective in thetreatment of bone problems associated with malignan-cy96 and are now the drugs of choice.97–99 Clinical trialsthat investigate the benefit of bisphosphonate therapyuse a composite end point defined as a skeletal-related orbone event, which typically includes pathologic fracture,spinal cord compression, radiation or surgery to bone,and hypercalcemia of malignancy. Bisphosphonates sig-nificantly reduce the incidence of these events in my-eloma100 and in patients with breast cancer metasta-ses101,102 and in metastatic prostate cancer,103 lung cancer,renal cell carcinoma, and other solid tumors. The goalsof treatment for bone metastases are also to preventdisease-related skeletal complications, palliate pain, andmaintain quality of life. Zoledronate,104 pamidronate,clodronate, and ibandronate105,106 have demonstrated ef-ficacy compared with placebo.

There is the important possibility that the survival ofpatients may be prolonged107–109 in some groups of pa-tients. Recently, osteonecrosis of the jaw110 was identi-fied as a potential complication of high-dose bisphospho-nate therapy in malignant diseases.

The other area of outstanding commercial successwith bisphosphonates has been in the therapy of osteo-porosis, which is a major public health problem.111,112 Upuntil the 1990s, there were few treatments for osteopo-rosis. As a drug class the bisphosphonates have emergedin the past few years as the leading effective treatmentsfor postmenopausal and other forms of osteoporosis.Etidronate was the first of these,113–115 followed by alen-dronate116–118 and then risedronate.119,120 All have beenapproved as therapies in many countries and can in-crease bone mass and reduce fracture rates at the spineby 30% to 50% and at other sites in postmenopausalwomen.121 The reduction in fractures may be related notonly to the increase in bone mass arising from the inhi-bition of bone resorption and reduced activation fre-quency of bone-remodeling units but also to enhancedosteon mineralization.122 These bisphosphonates alsoprevent bone loss associated with glucocorticosteroid ad-ministration.123,124

Among the newer bisphosphonates, ibandronate125

was introduced recently as a once-monthly tablet. Inaddition to formulations to be taken by mouth weekly or


monthly, new routes of administration are being stud-ied, especially periodic (eg, 3 monthly) injections withibandronate and once-yearly treatment with zoledr-onate.126 This has the attraction of delivering a defineddose without the variability associated with oral admin-istration as well as avoiding potential gastrointestinalintolerance. If these approaches are accompanied bygreater compliance and convenience, they are likely tobecome popular methods of treatment.

Other clinical issues under consideration withbisphosphonates include the choice of therapeutic regi-men (eg, the use of intermittent dosing rather thancontinuous, intravenous versus oral therapy), the opti-mal duration of therapy, the combination with otherdrugs such as teriparatide, and their extended use inrelated indications (eg, glucocorticosteroid-associatedosteoporosis, male osteoporosis, childhood osteopenicdisorders, arthritis, and other disorders). Therefore,there is much that needs to be done to improve the wayin which existing drugs can be used and to introducenew ones.

In pediatrics, pamidronate has proved remarkably ef-fective in increasing bone in children with the inherited“brittle-bone” disorder, osteogenesis imperfecta.127,128

SOME CURRENT ISSUESWITH BISPHOSPHONATES: BONEARCHITECTURE, STRUCTURE, AND STRENGTH, AND ON BONEHEALING AND FRACTURE REPAIRMany experimental and clinical studies show thatbisphosphonates conserve bone architecture andstrength.129–133 However, there have been concerns aboutwhether the use of prolonged high doses of bisphospho-nates may impair bone turnover to such an extent thatbone strength is impaired. High doses in animals areassociated with increased microdamage134,135 and evenfractures.136 It has been suggested that bisphosphonatesmight prevent naturally occurring microscopic cracks inbone from healing. There have been isolated reports ofadynamic bone associated with bisphosphonate usage,137

but long-term use of the bisphosphonates in the therapyof osteoporosis seems to be safe.138 Case reports of in-duction of osteopetrosis-like lesions in children whowere treated with excessive doses of pamidronate havebeen published.139

A question often asked is whether bisphosphonatesinhibit fracture repair. By reducing bone turnover onemight expect bisphosphonates to interfere with fracturehealing. However, a recent long-term study in a beagledog model that simulated fracture repair has demon-strated that ibandronate treatment did not adverselyaffect normal bone healing.140 Studies of repair processesafter creating drill-hole defects in dogs also showed noimpairment with ibandronate.141

Several other recent studies raised the intriguing pos-sibility that bisphosphonates may enhance fracture re-pair and related processes.142 In studies of the osseointe-gration of metal implants in ovariectomized rats,treatment with ibandronate resulted in improved os-seointegration rather than impairment of the healingprocess.143 Potential applications of bisphosphonates inorthopedics include protection against loosening of pros-theses,144 better integration of biomaterials and implants,improved healing in distraction osteogenesis,145 and con-serving bone architecture after osteonecrosis146,147 and inPerthes disease.148

There are potentially important differences betweenclinically useful bisphosphonates regarding their po-tency and duration of action. Efficacy is closely related toaffinity for bone mineral and ability to inhibit FPP syn-thase. Recent studies showing that there are markeddifferences among bisphosphonates in binding to hy-droxyapatite149 may explain the variations in retentionand persistence of effect that have been observed inanimal and clinical studies. In the case of zoledronic acid,in particular, the remarkable magnitude of effect andprolonged duration of action can be explained in part bythese new observations. In explaining the long durationof action, it has been proposed that there is continual

FIGURE 4Bisphosphonate (BP) uptake and detachment from bone surfac-es: effect of binding affinity on recirculation of bisphosphonateon and off bone surfaces. The differences in mineral-binding af-finity may affect distribution into different bone compartmentsand persistence of drug action at bone surfaces. (Adapted fromNancollas GH, Tang R, Phipps RJ, et al. Bone. 2006;38:617–627.)


recycling of bisphosphonate off and back onto the bonesurface. This notion is supported by observations thatbisphosphonates can be found in plasma and urinemany months after dosing (Fig 4).

There are numerous examples of bisphosphonateshaving effects on cells and tissues outside the skeleton.The effects on osteoclast precursors, tumor cells, macro-phages, and �,�-T cells are examples and in all cases areprobably explained by sufficient bisphosphonates enter-ing cells to inhibit the mevalonate pathway. A well-recognized adverse effect of the nitrogen-containingbisphosphonates is that they cause an acute-phase re-sponse in vivo,150,151 which can lead to induction of feverand “flu-like” symptoms in patients. These effects aretransient and occur predominantly on first exposure tothe drug, especially with intravenous administration.The mechanism has been attributed to release of proin-flammatory cytokines, and the mechanism has been fur-ther unraveled by showing that it involves selective re-ceptor-mediated activation of �,�-T cells, leading to theirproliferation and activation.152 The bisphosphonate ef-fect involves the mevalonate pathway in vitro and canbe overcome by using statins.153

Another interesting aspect of these nonskeletal effectsare the observations made on protozoan parasites, thegrowth of which can be inhibited by bisphosphonatesacting on FPPS.154,155

In the future, other clinical indications ripe for futurestudy include the prevention of bone loss and erosions inrheumatoid arthritis, possible applications in other jointdiseases, the reduction of bone loss associated with peri-odontal disease, and loosening of joint prostheses.

The recent elucidation of the likely mode of action ofbisphosphonates within cells opens up the possibility ofexploiting the subtle and potentially important differ-ences between the classes of bisphosphonates and indi-vidual compounds.

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