Fluorescent risedronate analogues reveal bisphosphonate uptake by bone marrow monocytes and localization around osteocytes in vivo

Bisphosphonates are effective antiresorptive agents owing to their bone-targeting property and ability to inhibit osteoclasts. It remains unclear, however, whether any non-osteoclast cells are directly affected by these drugs in vivo. Two fluorescent risedronate analogues, carboxyfluorescein-labeled risedronate (FAM-RIS) and Alexa Fluor 647–labeled risedronate (AF647-RIS), were used to address this question. Twenty-four hours after injection into 3-month-old mice, fluorescent risedronate analogues were bound to bone surfaces. More detailed analysis revealed labeling of vascular channel walls within cortical bone. Furthermore, fluorescent risedronate analogues were present in osteocytic lacunae in close proximity to vascular channels and localized to the lacunae of newly embedded osteocytes close to the bone surface. Following injection into newborn rabbits, intracellular uptake of fluorescently labeled risedronate was detected in osteoclasts, and the active analogue FAM-RIS caused accumulation of unprenylated Rap1A in these cells. In addition, CD14high bone marrow monocytes showed relatively high levels of uptake of fluorescently labeled risedronate, which correlated with selective accumulation of unprenylated Rap1A in CD14+ cells, as well as osteoclasts, following treatment with risedronate in vivo. Similar results were obtained when either rabbit or human bone marrow cells were treated with fluorescent risedronate analogues in vitro. These findings suggest that the capacity of different cell types to endocytose bisphosphonate is a major determinant for the degree of cellular drug uptake in vitro as well as in vivo. In conclusion, this study shows that in addition to bone-resorbing osteoclasts, bisphosphonates may exert direct effects on bone marrow monocytes in vivo. © 2010 American Society for Bone and Mineral Research

1. 1.

   Russell RGG, Watts NB, Ebetino FH, Rogers MJ. Mechanisms of action of bisphosphonates: similarities and differences and their potential influence on clinical efficacy. Osteoporos Int. 2008;19:733-759.

   Not in our library10.1007/s00198-007-0540-8 (opens in a new tab)

2. 2.

   Coxon FP, Thompson K, Roelofs AJ, Ebetino FH, Rogers MJ. Visualizing mineral binding and uptake of bisphosphonate by osteoclasts and non-resorbing cells. Bone. 2008;42:846-860.

   Not in our library10.1016/j.bone.2007.12.225 (opens in a new tab)

3. 3.

   Alendronate mechanism of action: geranylgeraniol, an intermediate in the mevalonate pathway, prevents inhibition of osteoclast formation, bone resorption, and kinase activation in vitro · 1999

   Not checked
4. 4.

   Kavanagh KL, Guo K, Dunford JE, et al. The molecular mechanism of nitrogen-containing bisphosphonates as anti-osteoporosis drugs. Proc Natl Acad Sci USA. 2006;103:7829-7934.

   Not in our library10.1073/pnas.0601643103 (opens in a new tab)

5. 5.

   Luckman SP, Hughes DE, Coxon FP, Russell RGG, Rogers MJ. Nitrogen-containing bisphosphonates inhibit the mevalonate pathway and prevent post-translational prenylation of GTP-binding proteins, including Ras. J Bone Miner Res. 1998;13:581-589.

   Not in our library10.1359/jbmr.1998.13.4.581 (opens in a new tab)

6. 6.

   Dunford JE, Rogers MJ, Ebetino FH, Phipps RJ, Coxon FP. Inhibition of protein prenylation by bisphosphonates causes sustained activation of Rac, Cdc42, and Rho GTPases. J Bone Miner Res. 2006;21:684-694.

   Not in our library10.1359/jbmr.060118 (opens in a new tab)

7. 7.

   Sato M, Grasser W, Endo N, et al. Bisphosphonate action: alendronate localization in rat bone and effects on osteoclast ultrastructure. J Clin Invest. 1991;88:2095-2105.

   Not in our library10.1172/jci115539 (opens in a new tab)

8. 8.

   Azuma Y, Sato H, Oue Y, et al. Alendronate distributed on bone surfaces inhibits osteoclastic bone resorption in vitro and in experimental hypercalcemia models. Bone. 1995;16:235-245.

   Not in our library10.1016/8756-3282(94)00035-x (opens in a new tab)

9. 9.

   Masarachia P, Weinreb M, Balena R, Godan GA. Comparison of the distribution of [3H]alendronate and [3H]etidronate in rat and mouse bones. Bone. 1996;19:281-290.

   Not in our library10.1016/8756-3282(96)00182-2 (opens in a new tab)

10. 10.

    Thompson K, Rogers MJ, Coxon FP, Crockett JC. Cytosolic entry of bisphosphonate drugs requires acidification of vesicles after fluid-phase endocytosis. Mol Pharmacol. 2006;69:1624-1632.

    Not in our library10.1124/mol.105.020776 (opens in a new tab)

11. 11.

    Roelofs AJ, Thompson K, Gordon S, Rogers MJ. Molecular mechanisms of action of bisphosphonates: current status. Clin Cancer Res. 2006;12:6222-6230s.

    Not in our library10.1158/1078-0432.ccr-06-0843 (opens in a new tab)

12. 12.

    Peripheral blood monocytes are responsible for γδ T cell activation induced by zoledronic acid through accumulation of IPP/DMAPP · 2008

13. 13.

    Kunzmann V, Bauer E, Feurle J, Weißinger F, Tony HP, Wilhelm M. Stimulation of gammadelta T cells by aminobisphosphonates and induction of antiplasma cell activity in multiple myeloma. Blood. 2000;96:384-392.

    Not in our library

14. 14.

    Gober HJ, Kistowska M, Angman L, Jenö P, Mori L, De Libero G. Human T cell receptor γδ cells recognize endogenous mevalonate metabolites in tumor cells. J Exp Med. 2003;197:163-168.

    Not in our library10.1084/jem.20021500 (opens in a new tab)

15. 15.

    Thompson K, Rogers MJ. Statins prevent bisphosphonate-induced γδ T cell proliferation and activation in vitro. J Bone Miner Res. 2004;19:278-288.

    Not in our library10.1359/jbmr.0301230 (opens in a new tab)

16. 16.

    Hewitt RE, Lissina A, Green AE, Slay ES, Price DA, Sewell AK. The bisphosphonate acute phase response: rapid and copious production of proinflammatory cytokines by peripheral blood γδ T cells in response to aminobisphosphonates is inhibited by statins. Clin Exp Immunol. 2005;139:101-111.

    Not in our library10.1111/j.1365-2249.2005.02665.x (opens in a new tab)

17. 17.

    Zaheer A, Lenkinsky RE, Mahmood A, Jones AG, Cantley LC, Frangioni JV. In vivo nearinfrared fluorescence imaging of osteoblastic activity. Nat Biotech. 2001;19:1148-1154.

    Not in our library10.1038/nbt1201-1148 (opens in a new tab)

18. 18.

    Kozloff KM, Weissleder R, Mahmood U. Noninvasive optical detection of bone mineral. J Bone Miner Res. 2007;22:1208-1216.

    Not in our library10.1359/jbmr.070504 (opens in a new tab)

19. 19.

    Kashemirov BA, Bala JLF, Chen X, et al. Fluorescently labeled risedronate and related analogues: “Magic linker” synthesis. Bioconjug Chem. 2008;19:2308-2310.

    Not in our library10.1021/bc800369c (opens in a new tab)

20. 20.

    Nancollas GH, Tang R, Phipps RJ, et al. Novel insights into actions of bisphosphonates on bone: differences in interactions with hydroxyapatite. Bone. 2006;38:617-627.

    Not in our library10.1016/j.bone.2005.05.003 (opens in a new tab)

21. 21.

    LeGeros RZ. Calcium phosphates in oral biology and medicine. Monogr Oral Sci. 1991;15:1-201.

    Not in our library

22. 22.

    Coxon FP, Frith JC, Benford HL, Rogers MJ. Isolation and purification of rabbit osteoclasts. Methods Mol Med. 2003;80:89-99.

    Not in our library10.1385/1-59259-366-6:89 (opens in a new tab)

23. 23.

    Coxon FP, Ebetino FH, Mules EH, Seabra MC, McKenna CE, Rogers MJ. Phosphonocarboxylate inhibitors of Rab geranylgeranyl transferase disrupt the prenylation and membrane localization of Rab proteins in osteoclasts in vitro and in vivo. Bone. 2005;37:349-358.

    Not in our library10.1016/j.bone.2005.04.021 (opens in a new tab)

24. 24.

    Roelofs AJ, Hulley PA, Meijer A, Ebetino FH, Russell RGG, Shipman CM. Selective inhibition of Rab prenylation by a phosphonocarboxylate analogue of risedronate induces apoptosis, but not S-phase arrest, in human myeloma cells. Int J Cancer. 2006;119:1254-1261.

    Not in our library10.1002/ijc.21977 (opens in a new tab)

25. 25.

    Henneman ZJ, Nancollas GH, Ebetino FH, Russell RG, Phipps RJ. Bisphosphonate binding affinity as assessed by inhibition of carbonated apatite dissolution in vitro. J Biomed Mater Res A. 2008;85:993-1000.

    Not in our library10.1002/jbm.a.31599 (opens in a new tab)

26. 26.

    Lin JH. Bisphosphonates: a review of their pharmacokinetic properties. Bone. 1994;18:75-85.

    Not in our library10.1016/8756-3282(95)00445-9 (opens in a new tab)

27. 27.

    Plotkin LI, Weinstein RS, Parfitt AM, Roberson PK, Manolagas SC, Bellido T. Prevention of osteocyte and osteoblast apoptosis by bisphosphonates and calcitonin. J Clin Invest. 1999;104:1363-1374.

    Not in our library10.1172/jci6800 (opens in a new tab)

28. 28.

    Follet H, Li J, Phipps RJ, Hui S, Condon K, Burr DB. Risedronate and alendronate suppress osteocyte apoptosis following cyclic fatigue loading. Bone. 2007;40:1172-1177.

    Not in our library10.1016/j.bone.2006.12.052 (opens in a new tab)

29. 29.

    Plotkin LI, Manolagas SC, Bellido T. Transduction of cell survival signals by connexin-43 hemichannels. J Biol Chem. 2002;277:8648-8657.

    Not in our library10.1074/jbc.m108625200 (opens in a new tab)

30. 30.

    Plotkin LI, Lezcano V, Thostenson J, Weinstein RS, Manolagas SC, Bellido T. Connexin 43 is required for the anti-apoptotic effect of bisphosphonates on osteocytes and osteoblasts in vivo. J Bone Miner Res. 2008;23:1712-1721.

    Not in our library10.1359/jbmr.080617 (opens in a new tab)

31. 31.

    Bonneville M, Scotet E. Human V(9V(2 T cells: promising new leads for immunotherapy of infections and tumors. Curr Opin Immunol. 2005;18:539-546.

    Not in our library10.1016/j.coi.2006.07.002 (opens in a new tab)

32. 32.

    Mantovani A, Schioppa T, Porta C, Allavena P, Sica A. Role of tumor-associated macrophages in tumor progression and invasion. Cancer Metastas Rev. 2006;25:315-322.

    Not in our library10.1007/s10555-006-9001-7 (opens in a new tab)

33. 33.

    Giraudo E, Inoue M, Hanahan D. An amino-bisphosphonate targets MMP-9-expressing macrophages and angiogenesis to impair cervical carcinogenesis. J Clin Invest. 2004;114:623-633.

    Not in our library10.1172/jci22087 (opens in a new tab)

34. 34.

    Yamagishi S, Abe R, Inagaki Y, et al. Minodronate, a newly developed nitrogen-containing bisphosphonate, suppresses melanoma growth and improves survival in nude mice by blocking vascular endothelial growth factor signalling. Am J Pathol. 2004;165:1865-1874.

    Not in our library10.1016/s0002-9440(10)63239-7 (opens in a new tab)

35. 35.

    Melani C, Sangaletti S, Barazzetta FM, Werb Z, Colombo MP. Amino-biphosphonate mediated MMP-9 inhibition breaks the tumor-bone marrow axis responsible for myeloid-derived suppressor cell expansion and macrophage infiltration in tumor stroma. Cancer Res. 2007;67:11438-11446.

    Not in our library10.1158/0008-5472.can-07-1882 (opens in a new tab)

36. 36.

    Löwik CWGM, Van der Pluijm G, Van der Wee-Pals LJA, Bloys vanTreslong-de, Groot H, Bijvoet OLM. Migration and phenotypic transformation of osteoclast precursors into mature osteoclasts: the effect of a bisphosphonate. J Bone Miner Res. 1988;3:185-192.

    Not in our library10.1002/jbmr.5650030210 (opens in a new tab)

37. 37.

    Hughes DE, MacDonald BR, Russell RGG, Gowen M. Inhibition of osteoclast-like cell formation by bisphosphonates in long-term cultures of human bone marrow. J Clin Invest. 1989;83:1930-1935.

    Not in our library10.1172/jci114100 (opens in a new tab)

38. 38.

    Van Beek ER, Löwik CWGM, Papapoulos SE. Bisphosphonates suppress bone resorption by a direct effect on early osteoclast precursors without affecting the osteoclastogenic capacity of osteogenic cells: the role of protein geranylgeranylation in the action of nitrogen-containing bisphosphonates on osteoclast precursors. Bone. 2002;30:64-70.

    Not in our library10.1016/s8756-3282(01)00655-x (opens in a new tab)

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