Inhibition of bone resorption by alendronate and risedronate does not require osteoclast apoptosis
Introduction
Bisphosphonates (BPs) are the most potent and effective inhibitors of bone resorption in clinical use. Their P-C-P backbone structure: resembles pyrophosphate. BPs bind tightly to the bone mineral, and binding is enhanced when R1 (above) is a hydroxyl.33, 36 The R2 side chain defines individual BPs in this family and determines pharmacological activity. Based on their structure-dependent mode of action, BPs can be separated into two classes. The nitrogen-containing BPs (N-BPs), such as alendronate (ALN) and risedronate (RIS), contain one or more nitrogen atoms within R2. Non-N-BPs, such as etidronate (EHDP) or clodronate (CL2), have CH3 or Cl, respectively, at R2 or a nitrogen-free ring structure (tiludronate). Recent studies have shown that these features, in addition to the P-C-P backbone, confer specificity in intracellular action.
CL2, and to a lesser extent EHDP and tiludronate, but not N-BPs, are metabolized by macrophages into toxic adenosine triphosphate (ATP) analogs.11 Tiludronate also inhibits the vacuolar ATPase.7 N-BPs inhibit the mevalonate to cholesterol pathway enzyme farnesyldiphosphate (FPP) synthase and, consequently, geranylgeranylation.3, 8, 35 This action was shown to mediate N-BPs inhibition of osteoclast (Oc) formation and bone resorption and induction of apoptosis.2, 6, 10, 16, 26, 34 Recent data have also demonstrated in vivo inhibition of the mevalonate pathway selectively in the Oc by the N-BPs ALN, RIS, and ibandronate, but not by CL2 and EHDP.9
This study examines the relationship between Oc apoptosis and inhibition of bone resorption induced by N-BPs (ALN and RIS) and non-N-BPs (CL2 and EHDP). The findings suggest that, while inhibition of FPP synthase by N-BPs is responsible for suppression of bone resorption, as well as induction of osteoclast apoptosis and disruption of actin structure, apoptosis itself is not required for the other two inhibitory actions and may not occur at the lowest effective doses. In contrast, the non-N-BPs failed to inhibit bone resorption or disrupt the cytoskeleton when apoptosis was blocked. Therefore, it seems that apoptosis is required for inhibition of osteoclastic bone resorption by EHDP and CL2 but not by ALN or RIS.
Section snippets
Rabbit Oc bone resorption assay
Bone resorption assays were performed as previously described.26 Briefly, tibiae and femora were aseptically isolated and minced in α-MEM containing HEPES (10 mmol/L, pH 7.1) and penicillin/streptomycin (100 IU and 100 mg/mL, respectively). Cells in supernatant were isolated, supplemented with 10% fetal bovine serum (FBS), and plated onto 0.25-mm-thick slices of bovine bone. After 1 h, test compounds were added (at concentrations noted in the text) into triplicate wells in a volume of 200 μL
Alendronate and risedronate inhibit bone resorption in the absence of Oc cell death
In a previous study, ALN-induced Oc death and inhibition of bone resorption changed in the same direction, suggesting a possible link.16 However, in vivo N-BP treatment was shown to increase Oc number, at least initially, when inhibition of bone resorption clearly occurs.4, 10, 32 To investigate more closely the correlation between Oc death and inhibition of bone resorption, we examined the effects of ALN or RIS on bone resorption and cell number, across five doses between 0.06 and 6 nmol
Discussion
The findings of this study further support the separation of BPs into two classes based on their mode of action. The difference resides not only in their immediate intracellular targets, but, as indicated in this study, also in the events responsible for inhibition of bone resorption.
All BPs tested here and previously can clearly induce Oc apoptosis.2, 6, 13, 16, 26 Furthermore, Oc apoptosis would certainly reduce bone resorption and may occur as a result of treatment with any of the BPs.
References (38)
- et al.
Alendronate is a specific, nanomolar inhibitor of farnesyl diphosphate synthase
Arch Biochem Biophys
(2000) - et al.
Rho-A is critical for osteoclast podosome organization, motility, and bone resorption
J Biol Chem
(2000) - et al.
Integrin-dependent leukocyte adhesion involves geranylgeranylated protein(s)
J Biol Chem
(1999) - et al.
PRK1 is targeted to endosomes by the small GTPase, RhoB
J Biol Chem
(1998) - et al.
A possible mechanism of the specific action of bisphosphonates on osteoclastsTiludronate preferentially affects polarized osteoclasts having ruffled borders
Bone
(1995) - et al.
Tiludronate inhibits protein tyrosine phosphatase activity in osteoclasts
Bone
(1997) - et al.
Rho, rac, and cdc42 GTPases regulate the assembly of multimolecular focal complexes associated with actin stress fibers, lamellipodia, and filopodia
Cell
(1995) - et al.
Rac-GTPase, osteoclast cytoskeleton and bone resorption
Eur J Cell Biol
(1999) - et al.
Bisphosphonates act directly on the osteoclast to induce caspase cleavage of mst1 kinase during apoptosis. A link between inhibition of the mevalonate pathway and regulation of an apoptosis-promoting kinase
J Biol Chem
(1999) - et al.
The small GTP-binding protein rho regulates the assembly of focal adhesions and actin stress fibers in response to growth factors
Cell
(1992)
The small GTP-binding protein rac regulates growth factor-induced membrane ruffling
Cell
Farnesyl pyrophosphate synthase is the molecular target of nitrogen-containing bisphosphonates
Biochem Biophys Res Commun
Binding and antiresorptive properties of heterocycle-containing bisphosphonate analogsStructure-activity relationships
Bone
The Rap1 GTPase functions as a regulator of morphogenesis in vivo
Eur Med Biol Org J
Farnesol and geranylgeraniol prevent activation of caspases by aminobisphosphonatesBiochemical evidence for two distinct pharmacological classes of bisphosphonate drugs
Mol Pharmacol
Alendronate increases skeletal mass of growing rats during unloading by inhibiting resorption of calcified cartilage
J Bone Miner Res
Protein geranylgeranylation is required for osteoclast formation, function, and survivalInhibition by bisphosphonates and GGTI-298
J Bone Miner Res
The bisphosphonate tiludronate is a potent inhibitor of the osteoclast vacuolar H(+)-ATPase
J Bone Miner Res
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
Cited by (161)
Cellular and molecular actions of bisphosphonates
2021, Bone Cancer: Bone Sarcomas and Bone Metastases - From Bench to BedsideCrosstalk between Fas and S1P<inf>1</inf> signaling via NF-kB in osteoclasts controls bone destruction in the TMJ due to rheumatoid arthritis
2019, Japanese Dental Science ReviewLong-term therapy with intravenous zoledronate increases the number of nonattached osteoclasts
2017, Journal of Cranio-Maxillofacial SurgeryZoledronate prevents lactation induced bone loss and results in additional post-lactation bone mass in mice
2016, BoneCitation Excerpt :As we hypothesised, Zln prevented the lactation induced loss of BV/TV, vBMD, aBMD, and mechanical properties (except at the femoral neck) at all investigated bone sites. Zln's primary mechanism of action is inhibition of osteoclast function or survival [29–32], and in agreement with this we observed substantially lower circulating levels of TRAP and Oc.S/BS in Zln-treated lactating and recovering mice. The BFR/BS, Os.S/BS and circulating levels of ALP were also distinctively lower in lactating mice receiving Zln, presumably mediated by Zln's inhibition of the bone resorption and thereby bone remodelling [33,34].
Medication-related osteonecrosis of the jaw (MRONJ): a review of pathogenesis hypothesis and therapy strategies
2024, Archives of Toxicology