Recent advances in understanding the mechanism of action of bisphosphonates

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Bisphosphonates (BPs) are widely used in the treatment of diseases associated with excessive osteoclast-mediated bone resorption, such as osteoporosis. Although several years ago the molecular target of the potent nitrogen-containing BPs (N-BPs) was identified as farnesyl diphosphate synthase, an enzyme in the mevalonate pathway, recent data have shed new light on the precise mechanism of inhibition and demonstrated that the acute-phase reaction, an adverse effect of N-BPs, is also caused by inhibition of this enzyme. In addition, the identification of BP analogues that inhibit different enzymes in the mevalonate pathway could lead to the development of novel inhibitors of bone resorption with potential applications in the treatment of bone disease.

Introduction

Bisphosphonates (BPs) are the most widely used and effective antiresorptive agent for the treatment of diseases in which there is an increase in osteoclastic resorption, including post-menopausal osteoporosis, Paget's disease and tumour-associated osteolysis. These drugs are synthetic analogues of inorganic pyrophosphate (PPi), consisting of two phosphonate groups linked by nonhydrolysable phosphoether bonds to a central carbon atom, to which are attached two covalently bonded sidechains (R1 and R2). This P–C–P backbone structure enables BPs to bind avidly to divalent metal ions such as Ca2+ [1], and as a result BPs bind to bone mineral surfaces in vivo [2]. This bone-targeting property, followed by their localized release during osteoclast-mediated resorption, explains why BPs appear to have a highly selective effect on osteoclasts in vivo [1].

BPs can be separated into two groups according to their molecular mechanism of action. The simple BPs that most closely resemble PPi, such as clodronate and etidronate, are metabolized intracellularly into methylene-containing analogues of ATP by aminoacyl-tRNA synthetase enzymes, with the nonhydrolysable P–C–P group of the BP replacing P–O–P (i.e. the β,γ-phosphate groups) in ATP [3]. These AppCp-type metabolites accumulate in the cytosol of osteoclasts and induce cell death [4], probably by inhibiting ATP-utilizing enzymes such as adenine nucleotide translocase (ANT), a component of the mitochondrial permeability transition pore [5]. Inhibition of ANT by the AppCCl2p metabolite of clodronate causes hyperpolarization of the mitochondrial inner membrane [5], which can lead to breakdown of the mitochondrial membrane potential, release of cytochrome C and induction of apoptosis [1].

The more potent N-BPs, which have bulkier side-chains characterized by a nitrogen moiety either in an alkyl chain (e.g. alendronate and ibandronate) or within a heterocyclic structure (e.g. risedronate and zoledronate), are not metabolized. Rather, N-BPs act by inhibiting farnesyl diphosphate synthase (FPP synthase), an enzyme of the mevalonate pathway, thereby depleting cells of the isoprenoid lipids FPP and geranylgeranyl diphosphate (GGPP), which are required for the post-translational prenylation of small GTPases, such as those of the Ras, Rho and Rab families [6, 7, 8]. Because prenylation is required for the localization of these GTPases to subcellular membranes, N-BPs are thought to inhibit resorption by disrupting the localization and function of small GTPases that are essential for osteoclast activity and survival [1].

This review focuses on recent advances that have further clarified the mechanism of action of N-BPs, from the initial binding to bone and uptake into osteoclasts through to the mechanism of inhibition of FPP synthase and the consequences of inhibiting this enzyme.

Section snippets

Bone targeting of bisphosphonates and uptake by osteoclasts

Although the bone-targeting property of BPs is known to be related to their ability to chelate Ca2+ ions through bidentate or tridentate binding via the phosphonate groups and R1 sidechain [1], recent studies suggest that the R2 sidechain of N-BPs can also influence overall bone affinity as a result of the ability of the nitrogen moiety to interact with the crystal surface of bone mineral. For example, risedronate and ibandronate appear to have a lower affinity for bone mineral than does

Mechanism of inhibition of FPP synthase by N-BPs

The exact mechanism by which N-BPs inhibit FPP synthase is only just becoming clear. The recent generation of X-ray crystal structures of human FPP synthase co-crystallized with N-BPs [18••, 19••] indicates that the BPs appear to bind in one of the two isoprenoid lipid-binding pockets (the GPP/DMAPP pocket) in the enzyme active site, with the R2 sidechain positioned in the hydrophobic cleft that normally accommodates the isoprenoid lipid, and the phosphonate groups bonding to lysine residues

Consequences of inhibiting FPP synthase

Small GTPases, such as those of the Ras, Rho and Rab families, are crucial signalling proteins that regulate a variety of cell processes necessary for osteoclast function, including cytoskeletal arrangement, membrane ruffling, trafficking of intracellular vesicles and cell survival [25]. It has been assumed that N-BPs, by preventing protein prenylation, disrupt the interaction of these small GTPases with cell membranes, thereby interfering with these processes. For example, loss of prenylation

Molecular basis for the acute-phase reaction to bisphosphonates

The major adverse effect of intravenous N-BP administration is a flu-like acute-phase reaction, which typically occurs in about one-third of patients receiving N-BPs for the first time. Although this phenomenon was first described 20 years ago [38], the molecular mechanism involved has only recently become clear. A study by Kunzmann et al. [39] reported that patients who suffered an acute-phase reaction to pamidronate had increased circulating levels of γδ T-cells in peripheral blood. Unlike

Potential of new bisphosphonate analogues

Recently, it has become clear that changes to the structure of N-BPs can give rise to compounds capable of inhibiting other enzymes of the mevalonate pathway that use isoprenoid lipids. For example, we found that a weakly antiresorptive phosphonocarboxylate (PC) analogue of risedronate, in which one of the phosphonate groups of risedronate is replaced with a carboxylate group (3-PEHPC, previously called NE10790), has no effect on FPP synthase but specifically inhibits the protein prenyl

Conclusions

FPP synthase is the major pharmacological target of N-BPs, accounting for not only the antiresorptive activity of these drugs, but also their ability to induce an acute-phase reaction, via the activation of γδ T cells. Recent studies have clarified the exact mechanism by which N-BPs inhibit this enzyme, thus finally explaining the relationship between chemical structure and antiresorptive potency of these blockbuster drugs. However, further studies are needed to understand the subsequent

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

References (46)

  • A. Cordle et al.

    Mechanisms of statin-mediated inhibition of small G-protein function

    J Biol Chem

    (2005)
  • H. Monkkonen et al.

    A new endogenous ATP analog (ApppI) inhibits the mitochondrial adenine nucleotide translocase (ANT) and is responsible for the apoptosis induced by nitrogen-containing bisphosphonates

    Br J Pharmacol

    (2006)
  • J.E. Fisher et al.

    Alendronate mechanism of action: geranylgeraniol, an intermediate in the mevalonate pathway, prevents inhibition of osteoclast formation, bone resorption, and kinase activation in vitro

    Proc Natl Acad Sci USA

    (1999)
  • V. Kunzmann et al.

    Gamma/delta T-cell stimulation by pamidronate

    N Engl J Med

    (1999)
  • K. Thompson et al.

    Statins prevent bisphosphonate-induced γ,δ-T-cell proliferation and activation in vitro

    J Bone Miner Res

    (2004)
  • M.J. Rogers

    New insights into the molecular mechanisms of action of bisphosphonates

    Curr Pharm Des

    (2003)
  • J.C. Frith 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)
  • J.C. Frith et al.

    The molecular mechanism of action of the anti-resorptive and anti-inflammatory drug clodronate: evidence for the formation in vivo of a metabolite that inhibits bone resorption and causes osteoclast and macrophage apoptosis

    Arthritis Rheum

    (2001)
  • P.P. Lehenkari 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)
  • J.E. Dunford 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)
  • Nancollas GH, Tang R, Phipps RJ, Henneman Z, Gulde S, Wu W, Mangood A, Russell RG, Ebetino FH: Novel insights into...
  • Leu CT, Luegmayr E, Freedman LP, Rodan GA, Reszka AA: Relative binding affinities of bisphosphonates for human bone and...
  • F.H. Ebetino et al.

    Mechanisms of action of etidronate and other bisphosphonates

    Rev Contemp Pharmacother

    (1998)
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