Heterocycle-Containing Bisphosphonates Cause Apoptosis and Inhibit Bone Resorption by Preventing Protein Prenylation: Evidence from Structure-Activity Relationships in J774 Macrophages

Abstract

Recent evidence suggests that bisphosphonates (BPs) may inhibit bone resorption by mechanisms that lead to osteoclast apoptosis. We have previously shown that BPs also reduce cell viability and induce apoptosis in the macrophage-like cell line J774. To determine whether BPs inhibit osteoclast-mediated bone resorption and affect J774 macrophages by the same molecular mechanism, we examined the potency to reduce J774 cell viability of pairs of nitrogen-containing BPs that differ slightly in the structure of the heterocycle-containing side chain but that differ markedly in antiresorptive potency. In all cases, the most potent antiresorptive BP of each pair also caused the greatest loss of J774 viability, while the less potent antiresorptive BPs were also less potent at reducing J774 cell viability. Similarly, the bisphosphinate, phosphonoalkylphosphinate and monophosphonate analogs of BPs (in which one or both phosphonate groups are modified, giving rise to much less potent or inactive antiresorptive agents) were much less potent or inactive at reducing J774 cell viability. Thus, the structure-activity relationships of BPs for inhibiting bone resorption match those for causing loss of cell viability in J774 cells, indicating that BPs inhibit osteoclast-mediated bone resorption and reduce J774 macrophage viability by the same molecular mechanism. Loss of J774 cell viability after treatment with BPs was associated with a parallel increase in apoptotic cell death. We have recently proposed that nitrogen-containing BPs reduce cell viability and cause J774 apoptosis as a consequence of inhibition of enzymes of the mevalonate pathway and hence loss of prenylated proteins. In this study, the BPs that were potent inducers of J774 apoptosis and potent antiresorptive agents were also found to be effective inhibitors of protein prenylation in J774 macrophages, whereas the less potent BP analogs did not inhibit protein prenylation. This provides strong evidence that BPs with a heterocyclic, nitrogencontaining side chain, such as risedronate, inhibit osteoclast-mediated bone resorption and induce J774 apoptosis by preventing protein prenylation. (J Bone Miner Res 1998;13:1668-1678)

Bisphosphonates induce apoptosis in human breast cancer cell lines

“…Therefore, the ability of pamidronate to down-regulate bcl-2 expression further suggests the involvement of caspase activation with changes in mitochondrial function in this pathway. Luckman and colleagues have reported that aminobisphosphonates induce apoptosis in a mouse macrophage cell line via inhibition of the mevalonate pathway and inhibition of protein prenylation (Luckman et al, 1998). A similar pathway has been identified for induction of apoptosis in human myeloma cells by YM 175 (Shipman et al, 1998).…”

Identification of a Novel Phosphonocarboxylate Inhibitor of Rab Geranylgeranyl Transferase That Specifically Prevents Rab Prenylation in Osteoclasts and Macrophages

“…Assessment of Viable Cell Number by MTT Assay-The number of viable J774 macrophage cells was determined by MTT assay as previously described (23). J774 cells were seeded at a density of 10 4 cells/well into 96-well plates and then treated with RIS, NE10790, or NE10485 the following day in replicates of six wells.…”

Bisphosphonates: Mode of Action and Pharmacology

“…There is a strong structureactivity relationship such that changes to the structure of the nitrogen-containing R 2 side chain or to the phosphonate groups, which alter antiresorptive potency, also influence the ability to inhibit protein prenylation to a corresponding degree. 72 An important verification of the critical importance of this pathway has come from showing that the addition of intermediates of the mevalonate pathway (such as FPP and GGPP) could overcome bisphosphonate-induced apoptosis and other events in many cell systems. Another prediction was that if inhibition of the mevalonate pathway could account for the antiresorptive effects of bisphosphonates, then the statin drugs should also inhibit bone resorption.…”