DOI QR코드

DOI QR Code

Obatoclax Regulates the Proliferation and Fusion of Osteoclast Precursors through the Inhibition of ERK Activation by RANKL

  • Oh, Ju Hee (Department of Biomedical Laboratory Science, College of Medical Sciences, Soonchunhyang University) ;
  • Lee, Jae Yoon (Department of Biomedical Laboratory Science, College of Medical Sciences, Soonchunhyang University) ;
  • Park, Jin Hyeong (Department of Biomedical Laboratory Science, College of Medical Sciences, Soonchunhyang University) ;
  • No, Jeong Hyeon (Department of Biomedical Laboratory Science, College of Medical Sciences, Soonchunhyang University) ;
  • Lee, Na Kyung (Department of Biomedical Laboratory Science, College of Medical Sciences, Soonchunhyang University)
  • Received : 2014.12.11
  • Accepted : 2015.01.05
  • Published : 2015.03.31

Abstract

Obatoclax, a pan-Bcl2 inhibitor, shows antitumor activities in various solid malignancies. Bcl2-deficient mice have shown the importance of Bcl2 in osteoclasts, as the bone mass of the mice was increased by the induced apoptosis of osteoclasts. Despite the importance of Bcl2, the effects of obatoclax on the proliferation and differentiation of osteoclast precursors have not been studied extensively. Here, we describe the anti-proliferative effects of obatoclax on osteoclast precursors and its negative role on fusion of the cells. Stimulation with low doses of obatoclax significantly suppressed the proliferation of osteoclast precursors in a dose-dependent manner while the apoptosis was markedly increased. Its stimulation was sufficient to block the activation of ERK MAP kinase by RANKL. The same was true when PD98059, an ERK inhibitor, was administered to osteoclast precursors. The activation of JNK1/2 and p38 MAP kinase, necessary for osteoclast differentiation, by RANKL was not affected by obatoclax. Interestingly, whereas the number of TRAP-positive mononuclear cells was increased by both obatoclax and PD98059, fused, multinucleated cells larger than $100{\pm}m$ in diameter containing more than 20 nuclei were completely reduced. Consistently, obatoclax failed to regulate the expression of osteoclast marker genes, including c-Fos, TRAP, RANK and CtsK. Instead, the expression of DC-STAMP and Atp6v0d2, genes that regulate osteoclast fusion, by RANKL was significantly abrogated by both obatoclax and PD98059. Taken together, these results suggest that obatoclax down-regulates the proliferation and fusion of osteoclast precursors through the inhibition of the ERK1/2 MAP kinase pathway.

Keywords

ERK1/2 MAP kinase;fusion;obatoclax;osteoclast precursors;proliferation

Acknowledgement

Supported by : National Research Foundation of Korea

References

  1. Anderson, D.M., Maraskovsky, E., Billingsley, W.L., Dougall, W.C., Tometsko, M.E., Roux, E.R., Teepe, M.C., DuBose, R.F., Cosman, D., and Galibert, L. (1997). A homologue of the TNF receptor and its ligand enhance T-cell growth and dendritic-cell function. Nature 390, 175-179 https://doi.org/10.1038/36593
  2. Blair, H.C. (1998). How the osteoclast degrades bone. Bioessays 20, 837-848 https://doi.org/10.1002/(SICI)1521-1878(199810)20:10<837::AID-BIES9>3.0.CO;2-D
  3. Boyle, W.J., Simonet, W.S., and Lacey, D.L. (2003). Osteoclast differentiation and activation. Nature 423, 337-342. https://doi.org/10.1038/nature01658
  4. Campa's, C., Cosialls, A.M., Barragn, M., Iglesias-Serret, D., Santidrian, A.F., Coll-Mulet, L., de Frias, M., Domingo, A., Pons, G., and Gil, J. (2006). Bcl-2 inhibitors induce apoptosis in chronic lymphocytic leukemia cells. Exp. Hematol. 34, 1663-1669 https://doi.org/10.1016/j.exphem.2006.07.008
  5. Choi, J., Choi, S.Y., Lee, S.Y., Lee, J.Y., Kim, H.S., Lee, S.Y., and Lee, N.K. (2013). Caffeine enhances osteoclast differentiation and maturation through p38 MAP kinase/Mitf and DCSTAMP/CtsK and TRAP pathway. Cell. Signal. 25, 1222-1227 https://doi.org/10.1016/j.cellsig.2013.02.015
  6. David, J.P., Sabapathy, K., Hoffmann, O., Idarraga, M.H., and Wagner, E.F. (2002). JNK1 modulates osteoclastogenesis through both c-Jun phosphorylation-dependent and-independent mechanisms. J. Cell Sci. 115, 4317-4325. https://doi.org/10.1242/jcs.00082
  7. de Vries, T.J., Schoenmaker, T., Beertsen, W., van der Neut, R., and Everts, V. (2005). Effect of CD44 deficiency on in vitro and in vivo osteoclast formation. J. Cell. Biochem. 94, 954-966. https://doi.org/10.1002/jcb.20326
  8. Harada, S., and Rodan, G.A. (2003). Control of osteoblast function and regulation of bone mass. Nature 423, 349-355. https://doi.org/10.1038/nature01660
  9. Hartgers, F.C., Vissers, J.L., Looman, M.W., van Zoelen, C., Huffine, C., Figdor, C.G., and Adema, G.J. (2000). DC-STAMP, a novel multimembrane-spanning molecule preferentially expressed by dendritic cells. Eur. J. Immunol. 30, 3585-3590. https://doi.org/10.1002/1521-4141(200012)30:12<3585::AID-IMMU3585>3.0.CO;2-Y
  10. Hockenbery, D.M., Oltvai, Z.N., Yin, X.M., Milliman, C.L., and Korsmeyer, S.J. (1993). Bcl-2 functions in an antioxidant pathway to prevent apoptosis. Cell 75, 241-251. https://doi.org/10.1016/0092-8674(93)80066-N
  11. Hwang, J.J., Kuruvilla, J., Mendelson, D., Pishvaian, M.J., Deeken, J.F., Siu, L.L., Berger, M.S., Viallet, J., and Marshall, J.L. (2010). Phase I dose finding studies of obatoclax (GX15-070), a small molecule pan-BCL-2 family antagonist, in patients with advanced solid tumors or lymphoma. Clin. Cancer Res. 16, 4038-4045. https://doi.org/10.1158/1078-0432.CCR-10-0822
  12. Ishii, M., Iwai, K., Koike, M., Ohshima, S., Kudo-Tanaka, E., Ishii, T., Mima, T., Katada, Y., Miyatake, K., Uchiyama, Y., and Saeki, Y. (2006). RANKL-induced expression of tetraspanin CD9 in lipid raft membrane microdomain is essential for cell fusion during osteoclastogenesis. J. Bone Miner. Res. 21, 965-976. https://doi.org/10.1359/jbmr.060308
  13. Kim, H.S., and Lee, N.K. (2014). Gene expression profiling in osteoclast precursors by insulin using microarray analysis. Mol. Cells 30, 827-832.
  14. Kim, K., Lee, S.H., Kim, J., Choi, Y., and Kim, N. (2008). NFATc1 induces osteoclast fusion via upregulation of Atp6v0d2 and the dendritic cell-specific transmembrane protein (DCSTAMP). Mol. Endocrinol. 22, 176-185. https://doi.org/10.1210/me.2007-0237
  15. Kim, P.S., Jochems, C., Grenga, I., Donahue, R.N., Tsang, K.Y., Gulley, J.L., Schlom, J., and Farsaci, B. (2014). Pan-Bcl-2 Inhibitor, GX15-070 (Obatoclax), decreases human T regulatory lymphocytes while preserving effector T lymphocytes: a rationale for its use in combination immunotherapy, J. Immunol. 192, 2622-2633. https://doi.org/10.4049/jimmunol.1301369
  16. Konopleva, M., Watt, J., Contractor, R., Tsao, T., Harris, D., Estrov, Z., Bornmann, W., Kantarjian, H., Viallet, J., Samudio, I., et al. (2008). Mechanisms of antileukemic activity of the novel Bcl-2 homology domain-3 mimetic GX15-070 (obatoclax). Cancer Res. 68, 3413-3420. https://doi.org/10.1158/0008-5472.CAN-07-1919
  17. Kukita, T., Wada, N., Kukita, A., Kakimoto, T., Sandra, F., Toh, K., Nagata, K., Iijima, T., Horiuchi, M., Matsusaki, H., et al. (2004). RANKL-induced DC-STAMP is essential for osteoclastogenesis. J. Exp. Med. 200, 941-946. https://doi.org/10.1084/jem.20040518
  18. Lee, J.Y., and Lee, N.K. (2014). Up-regulation of cyclinD1 and Bcl2A1 by insulin is involved in osteoclast proliferation. Life Sci. 114, 57-61. https://doi.org/10.1016/j.lfs.2014.07.006
  19. Lee, S.H., Rho, J., Jeong, D., Sul, J.Y., Kim, T., Kim, N., Kang, J.S., Miyamoto, T., Suda, T., Lee, S.K., et al. (2006). v-ATPase V0 subunit d2-deficient mice exhibit impaired steoclast fusion and increased bone formation. Nat. Med. 12, 1403-1409 https://doi.org/10.1038/nm1514
  20. Li, X., Udagawa, N., Itoh, K., Suda, K., Murase, Y., and Nishihara, T. (2002). p38 MAPK-mediated signals are required for inducing osteoclast differentiation but not for osteoclast function. Endocrinology 143, 3105-3113. https://doi.org/10.1210/endo.143.8.8954
  21. Mansky, K.C., Sankar, U., Han, J., and Ostrowski, M.C. (2002). Microphthalmia transcription factor is a target of the p38 MAPK pathway in response to receptor activator of NF-B ligand signaling. J. Biol. Chem. 277, 11077-11083. https://doi.org/10.1074/jbc.M111696200
  22. Nishi, T., Kawasaki-Nishi, S., and Forgac, M. (2003). Expression and function of the mouse V-ATPase d subunit isoforms. J. Biol. Chem. 278, 46396-46402. https://doi.org/10.1074/jbc.M303924200
  23. Paik, P.K., Rudin, C.M., Brown, A., Rizvi, N.A., Takebe, N., Travis, W., James, L., Ginsberg, M.S., Juergens, R., Markus, S., et al. (2010). A phase I study of obatoclax mesylate, a Bcl-2 antagonist, plus topotecan in solid tumor malignancies. Cancer Chemother. Pharmacol. 66, 1079-1085. https://doi.org/10.1007/s00280-010-1265-5
  24. Rho, J., Altmann, C.R., Socci, N.D., Merkov, L., Kim, N., So, H., Lee, O., Takami, M., Brivanlou, A.H., and Choi, Y. (2002). Gene expression profiling of osteoclast differentiation by combined suppression subtractive hybridization (SSH) and cDNA microarray analysis. DNA Cell. Biol. 21, 541-549 https://doi.org/10.1089/104454902320308915
  25. Rodan, G.A., and Martin, T.J. (2000) Therapeutic approaches to bone diseases. Science 289, 1508-1514. https://doi.org/10.1126/science.289.5484.1508
  26. Staege, H., Brauchlin, A., Schoedon, G., and Schaffner, A. (2001). Two novel genes FIND and LIND differentially expressed in deactivated and Listeria-infected human macrophages. Immunogenetics 53, 105-113. https://doi.org/10.1007/s002510100306
  27. Teitelbaum, S.L. (2000). Bone resorption by osteoclasts. Science 289, 1504-1508 https://doi.org/10.1126/science.289.5484.1504
  28. Trudel, S., Li, Z.H., Rauw, J., Tiedemann, R.E., Wen, X.Y., and Stewart, A.K. (2007). Preclinical studies of the pan-Bcl inhibitor obatoclax (GX015-070) in multiple myeloma. Blood 109, 5430-5438 https://doi.org/10.1182/blood-2006-10-047951
  29. Vignery, A. (2000). Osteoclasts and giant cells: macrophagemacrophage fusion mechanism. Int. J. Exp. Pathol. 81, 291-304. https://doi.org/10.1046/j.1365-2613.2000.00164.x
  30. Wei, S., Wang, M.W., Teitelbaum, S.L., and Ross, F.P. (2002). Interleukin-4 reversibly inhibits osteoclastogenesis via inhibition of NF-kappa B and mitogen-activated protein kinase signaling. J. Biol. Chem. 277, 6622-6630. https://doi.org/10.1074/jbc.M104957200
  31. Wong, B.R., Josien, R., Lee, S.Y., Sauter, B., Li, H.L., Steinman, R.M., and Choi, Y. (1997). TRANCE (tumor necrosis factor [TNF]-related activation-induced cytokine), a new TNF family member predominantly expressed in T cells, is a dendritic cellspecific survival factor. J. Exp. Med. 186, 2075-2080. https://doi.org/10.1084/jem.186.12.2075
  32. Wu, H., Xu, G., and Li, Y.P. (2009). Atp6v0d2 is an essential component of the osteoclast-specific proton pump that mediates extracellular acidification in bone resorption. J. Bone Miner. Res. 24, 871-885. https://doi.org/10.1359/jbmr.081239
  33. Xing, L., Xiu, Y., and Boyce, B.F. (2012). Osteoclast fusion and regulation by RANKL-dependent and independent factors. World J. Orthop. 3, 212-222. https://doi.org/10.5312/wjo.v3.i12.212
  34. Yamashita, J., Datta, N.S., Chun, Y.H., Yang, D.Y., Carey, A.A., Kreider, J.M., Goldstein, S.A., and McCauley, L.K. (2008). Role of Bcl2 in osteoclastogenesis and PTH anabolic actions in bone. J. Bone Miner. Res. 23, 621-632.
  35. Yang, J., Zhang, X., Wang, W., and Liu, J. (2010). Insulin stimulates osteoblast proliferation and differentiation through ERK and PI3K in MG-63 cells. Cell Biochem. Funct. 28, 334-341. https://doi.org/10.1002/cbf.1668
  36. Zhang, C., Dou, C.E., Xu, J., and Dong, S. (2014). DC-STAMP, the key fusion-mediating molecule in osteoclastogenesis. J. Cell. Physiol. 229, 1330-1335. https://doi.org/10.1002/jcp.24553

Cited by

  1. Insulin enhances RANKL-induced osteoclastogenesis via ERK1/2 activation and induction of NFATc1 and Atp6v0d2 vol.27, pp.12, 2015, https://doi.org/10.1016/j.cellsig.2015.09.002
  2. Design, synthesis and biological evaluation of 3-aryl-rhodanine benzoic acids as anti-apoptotic protein Bcl-2 inhibitors vol.25, pp.22, 2015, https://doi.org/10.1016/j.bmcl.2015.09.051
  3. The triptolide-induced apoptosis of osteoclast precursor by degradation of cIAP2 and treatment of rheumatoid arthritis of TNF-transgenic mice pp.0951418X, 2018, https://doi.org/10.1002/ptr.6224