Skip to main content

Advertisement

SpringerLink
Log in

Deoxyhypusine synthase (DHPS) inhibitor GC7 induces p21/Rb-mediated inhibition of tumor cell growth and DHPS expression correlates with poor prognosis in neuroblastoma patients

  • Original Paper
  • Published:
Cellular Oncology Aims and scope Submit manuscript

Abstract

Purpose

Neuroblastoma (NB) is an aggressive pediatric malignancy that typically occurs in infants and children under the age of 5 years. High-stage tumors relapse frequently even after aggressive multimodal treatment, resulting in therapy resistance and eventually in patient death. Clearly, new biologically-targeted drugs are needed that more efficiently suppress tumor growth and prevent relapse. We and others previously showed that polyamines such as spermidine play an essential role in NB tumorigenesis and that DFMO, an inhibitor of the central polyamine synthesis gene ODC, is effective in vitro and in vivo, prompting its evaluation in NB clinical trials. However, the specific molecular actions of polyamines remain poorly defined. Spermidine and deoxyhypusine synthase (DHPS) are essential components in the hypusination-driven post-translational activation of eukaryotic initiation factor 5A (eIF5A).

Methods

We assessed the role of DHPS in NB and the impact of its inhibition by N 1-guanyl-1,7-diaminoheptane (GC7) on tumor cell growth using cell proliferation assays, Western blot, immunofluorescence microscopy, and Affymetrix micro-array mRNA expression analyses in NB tumor samples.

Results

We found that GC7 inhibits NB cell proliferation in a dose-dependent manner, through induction of the cell cycle inhibitor p21 and reduction of total and phosphorylated Rb proteins. Strikingly, high DHPS mRNA expression correlated significantly with unfavorable clinical parameters, including poor patient survival, in a cohort of 88 NB tumors (all P < 0.04).

Conclusions

These results suggest that spermidine and DHPS are key contributing factors in NB tumor proliferation through regulation of the p21/Rb signaling axis.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. G.M. Brodeur, Neuroblastoma: biological insights into a clinical enigma. Nat Rev Cancer 3, 203–216 (2003)

    Article  CAS  PubMed  Google Scholar 

  2. N.K. Cheung, M.A. Dyer, Neuroblastoma: developmental biology, cancer genomics and immunotherapy. Nat Rev Cancer 13, 397–411 (2013)

    Article  CAS  PubMed  Google Scholar 

  3. J.M. Maris, Recent advances in neuroblastoma. N Engl J Med 362, 2202–2211 (2010)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  4. J.M. Maris, M.D. Hogarty, R. Bagatell, S.L. Cohn, Neuroblastoma Lancet 369, 2106–2120 (2007)

    Article  CAS  Google Scholar 

  5. J.R. Park, A. Eggert, H. Caron, Neuroblastoma: biology, prognosis, and treatment. Hematol Oncol Clin North Am 24, 65–86 (2010)

    Article  PubMed  Google Scholar 

  6. M. Schwab, F. Westermann, B. Hero, F. Berthold, Neuroblastoma: biology and molecular and chromosomal pathology. Lancet Oncol 4, 472–480 (2003)

    Article  CAS  PubMed  Google Scholar 

  7. D.L. Baker, M.L. Schmidt, S.L. Cohn, J.M. Maris, W.B. London, A. Buxton, D. Stram, R.P. Castleberry, H. Shimada, A. Sandler, R.C. Shamberger, A.T. Look, C.P. Reynolds, R.C. Seeger, K.K. Matthay, G. Children’s Oncology, Outcome after reduced chemotherapy for intermediate-risk neuroblastoma. N Engl J Med 363, 1313–1323 (2010)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  8. S.L. Cohn, A.D. Pearson, W.B. London, T. Monclair, P.F. Ambros, G.M. Brodeur, A. Faldum, B. Hero, T. Iehara, D. Machin, V. Mosseri, T. Simon, A. Garaventa, V. Castel, K.K. Matthay, The International Neuroblastoma Risk Group (INRG) classification system: an INRG task force report. J Clin Oncol 27, 289–297 (2009)

    Article  PubMed Central  PubMed  Google Scholar 

  9. S.G. Kreissman, R.C. Seeger, K.K. Matthay, W.B. London, R. Sposto, S.A. Grupp, D.A. Haas-Kogan, M.P. Laquaglia, A.L. Yu, L. Diller, A. Buxton, J.R. Park, S.L. Cohn, J.M. Maris, C.P. Reynolds, J.G. Villablanca, Purged versus non-purged peripheral blood stem-cell transplantation for high-risk neuroblastoma (COG A3973): a randomised phase 3 trial. Lancet Oncol 14, 999–1008 (2013)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  10. D.R. Strother, W.B. London, M.L. Schmidt, G.M. Brodeur, H. Shimada, P. Thorner, M.H. Collins, E. Tagge, S. Adkins, C.P. Reynolds, K. Murray, R.S. Lavey, K.K. Matthay, R. Castleberry, J.M. Maris, S.L. Cohn, Outcome after surgery alone or with restricted use of chemotherapy for patients with low-risk neuroblastoma: results of children’s oncology group study P9641. J Clin Oncol 30, 1842–1848 (2012)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  11. A. Canete, M. Gerrard, H. Rubie, V. Castel, A. Di Cataldo, C. Munzer, R. Ladenstein, B. Brichard, J.D. Bermudez, J. Couturier, B. de Bernardi, A.J. Pearson, J. Michon, Poor survival for infants with MYCN-amplified metastatic neuroblastoma despite intensified treatment: the international society of paediatric oncology European neuroblastoma experience. J Clin Oncol 27, 1014–1019 (2009)

    Article  PubMed  Google Scholar 

  12. B.H. Kushner, K. Kramer, M.P. LaQuaglia, S. Modak, K. Yataghene, N.K. Cheung, Reduction from seven to five cycles of intensive induction chemotherapy in children with high-risk neuroblastoma. J Clin Oncol 22, 4888–4892 (2004)

    Article  CAS  PubMed  Google Scholar 

  13. A.S. Bachmann, The role of polyamines in human cancer: prospects for drug combination therapies. Hawaii Med J 63, 371–374 (2004)

    PubMed  Google Scholar 

  14. R.A. Casero Jr., L.J. Marton, Targeting polyamine metabolism and function in cancer and other hyperproliferative diseases. Nat Rev Drug Discov 6, 373–390 (2007)

    Article  CAS  PubMed  Google Scholar 

  15. A.E. Pegg, Polyamine metabolism and its importance in neoplastic growth and a target for chemotherapy. Cancer Res 48, 759–774 (1988)

    CAS  PubMed  Google Scholar 

  16. A.E. Pegg, D.J. Feith, Polyamines and neoplastic growth. Biochem Soc Trans 35, 295–299 (2007)

    Article  CAS  PubMed  Google Scholar 

  17. E.W. Gerner, F.L. Meyskens Jr., Polyamines and cancer: old molecules, new understanding. Nat Rev Cancer 4, 781–792 (2004)

    Article  CAS  PubMed  Google Scholar 

  18. D.L. Koomoa, L.P. Yco, T. Borsics, C.J. Wallick, A.S. Bachmann, Ornithine decarboxylase inhibition by {alpha}-difluoromethylornithine activates opposing signaling pathways via phosphorylation of both Akt/Protein Kinase B and p27Kip1 in Neuroblastoma. Cancer Res 68, 9825–9831 (2008)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  19. C.J. Wallick, I. Gamper, M. Thorne, D.J. Feith, K.Y. Takasaki, S.M. Wilson, J.A. Seki, A.E. Pegg, C.V. Byus, A.S. Bachmann, Key role for p27Kip1, retinoblastoma protein Rb, and MYCN in polyamine inhibitor-induced G1 cell cycle arrest in MYCN-amplified human neuroblastoma cells. Oncogene 24, 5606–5618 (2005)

    Article  CAS  PubMed  Google Scholar 

  20. D.L. Koomoa, T. Borsics, D.J. Feith, C.C. Coleman, C.J. Wallick, I. Gamper, A.E. Pegg, A.S. Bachmann, Inhibition of S-adenosylmethionine decarboxylase by inhibitor SAM486A connects polyamine metabolism with p53-Mdm2-Akt/protein kinase B regulation and apoptosis in neuroblastoma. Mol Cancer Ther 8, 2067–2075 (2009)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  21. D.L. Koomoa, D. Geerts, I. Lange, J. Koster, A.E. Pegg, D.J. Feith, A.S. Bachmann, DFMO/eflornithine inhibits migration and invasion downstream of MYCN and involves p27Kip1 activity in neuroblastoma. Int J Oncol 42, 1219–1228 (2013)

    CAS  PubMed Central  PubMed  Google Scholar 

  22. K. Samal, P. Zhao, A. Kendzicky, L.P. Yco, H. McClung, E. Gerner, M. Burns, A.S. Bachmann, G. Sholler, AMXT-1501, a novel polyamine transport inhibitor, synergizes with DFMO in inhibiting neuroblastoma cell proliferation by targeting both ornithine decarboxylase and polyamine transport. Int J Cancer 133, 1323–1333 (2013)

    Article  CAS  PubMed  Google Scholar 

  23. M.D. Hogarty, M.D. Norris, K. Davis, X. Liu, N.F. Evageliou, C.S. Hayes, B. Pawel, R. Guo, H. Zhao, E. Sekyere, J. Keating, W. Thomas, N.C. Cheng, J. Murray, J. Smith, R. Sutton, N. Venn, W.B. London, A. Buxton, S.K. Gilmour, G.M. Marshall, M. Haber, ODC1 Is a critical determinant of MYCN Oncogenesis and a therapeutic target in neuroblastoma. Cancer Res 68, 9735–9745 (2008)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  24. R.J. Rounbehler, W. Li, M.A. Hall, C. Yang, M. Fallahi, J.L. Cleveland, Targeting ornithine decarboxylase impairs development of MYCN-amplified neuroblastoma. Cancer Res 69, 547–553 (2009)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  25. G. Sholler, E. Currier, D.L. Koomoa, A.S. Bachmann, synergistic inhibition of neuroblastoma tumor development by targeting ornithine decarboxylase and topoisomerase II. In 14th Advances in Neuroblastoma Research (ANR) Conference, Stockholm, Sweden, POT74 (2010)

  26. G. Sholler, E. Gerner, G. Bergendahl, B.J. LaFleur, A. VanderWerff, T. Ashilkaga, W. Ferguson, W. Roberts, R.K. Wada, D. Eslin, J. Kraveka, J. Kaplan, D. Mitchell, N.S. Parikh, K. Neville, L. Sender, T. Higgins, M. Kawakita, K. Hiramatsu, S.S. Moriya, A.S. Bachmann, A phase I trial of DFMO as a single agent and in combination with etoposide in patients with refractory or recurrent neuroblastoma. In Annual Meeting of the American Association for Cancer Research (AACR), Washington, D.C., LB-179 (2013)

  27. A.S. Bachmann, D. Geerts, G. Sholler, Neuroblastoma: Ornithine decarboxylase and polyamines are novel targets for therapeutic intervention. In Pediatric Cancer, Neuroblastoma: Diagnosis, Therapy, and Prognosis, ed. by M.A. Hayat, Springer Publisher, 91–103 (2012)

  28. A.S. Bachmann, V.A. Levin, Clinical applications of polyamine-based therapeutics. In Polyamine Drug Discovery, ed. by P.M. Woster and R.A. Casero, Jr., Royal Society of Chemistry, 257–276 (2012)

  29. U. Bachrach, Naturally occurring polyamines: interaction with macromolecules. Curr Protein Pept Sci 6, 559–566 (2005)

    Article  CAS  PubMed  Google Scholar 

  30. A.E. Pegg, Mammalian polyamine metabolism and function. IUBMB Life 61, 880–894 (2009)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  31. H.M. Wallace, A.V. Fraser, A. Hughes, A perspective of polyamine metabolism. Biochem J 376, 1–14 (2003)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  32. M.H. Park, K. Nishimura, C.F. Zanelli, S.R. Valentini, Functional significance of eIF5A and its hypusine modification in eukaryotes. Amino Acids 38, 491–500 (2010)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  33. E.A. Paz, B. LaFleur, E.W. Gerner, Polyamines are oncometabolites that regulate the LIN28/let-7 pathway in colorectal cancer cells. Mol Carcinog 53(Suppl 1), E96–106 (2014)

    Article  CAS  PubMed  Google Scholar 

  34. E.W. Gerner, Cancer chemoprevention locks onto a new polyamine metabolic target. Cancer Prev Res 3, 125–127 (2010)

    Article  CAS  Google Scholar 

  35. J. Jakus, E.C. Wolff, M.H. Park, J.E. Folk, Features of the spermidine-binding site of deoxyhypusine synthase as derived from inhibition studies. Effective inhibition by bis- and mono-guanylated diamines and polyamines. J Biol Chem 268, 13151–13159 (1993)

    CAS  PubMed  Google Scholar 

  36. M.H. Park, E.C. Wolff, Y.B. Lee, J.E. Folk, Antiproliferative effects of inhibitors of deoxyhypusine synthase. Inhibition of growth of Chinese hamster ovary cells by guanyl diamines. J Biol Chem 269, 27827–27832 (1994)

    CAS  PubMed  Google Scholar 

  37. Y. Lee, H.K. Kim, H.E. Park, M.H. Park, Y.A. Joe, Effect of N1-guanyl-1,7-diaminoheptane, an inhibitor of deoxyhypusine synthase, on endothelial cell growth, differentiation and apoptosis. Mol Cell Biochem 237, 69–76 (2002)

    Article  CAS  PubMed  Google Scholar 

  38. A. Slack, Z. Chen, R. Tonelli, M. Pule, L. Hunt, A. Pession, J.M. Shohet, The p53 regulatory gene MDM2 is a direct transcriptional target of MYCN in neuroblastoma. Proc Natl Acad Sci U S A 102, 731–736 (2005)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  39. P. Fardin, A. Barla, S. Mosci, L. Rosasco, A. Verri, R. Versteeg, H.N. Caron, J.J. Molenaar, I. Ora, A. Eva, M. Puppo, L. Varesio, A biology-driven approach identifies the hypoxia gene signature as a predictor of the outcome of neuroblastoma patients. Mol Cancer 9, 185 (2010)

    Article  PubMed Central  PubMed  Google Scholar 

  40. I. Revet, G. Huizenga, A. Chan, J. Koster, R. Volckmann, P. van Sluis, I. Ora, R. Versteeg, D. Geerts, The MSX1 homeobox transcription factor is a downstream target of PHOX2B and activates the Delta-Notch pathway in neuroblastoma. Exp Cell Res 314, 707–719 (2008)

    Article  CAS  PubMed  Google Scholar 

  41. T. Barrett, D.B. Troup, S.E. Wilhite, P. Ledoux, D. Rudnev, C. Evangelista, I.F. Kim, A. Soboleva, M. Tomashevsky, K.A. Marshall, K.H. Phillippy, P.M. Sherman, R.N. Muertter, R. Edgar, NCBI GEO: archive for high-throughput functional genomic data. Nucleic Acids Res 37, D885–890 (2009)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  42. V. Bewick, L. Cheek, J. Ball, Statistics review 12: survival analysis. Crit Care 8, 389–394 (2004)

    Article  PubMed Central  PubMed  Google Scholar 

  43. T. Borsics, E. Lundberg, D. Geerts, D.L. Koomoa, J. Koster, K. Wester, A.S. Bachmann, Subcellular distribution and expression of prenylated Rab acceptor 1 domain family, member 2 (PRAF2) in malignant glioma: influence on cell survival and migration. Cancer Sci 101, 1624–1631 (2010)

    Article  CAS  PubMed  Google Scholar 

  44. J.E. Gawecka, D. Geerts, J. Koster, M.J. Caliva, F.J. Sulzmaier, J. Opoku-Ansah, R.K. Wada, A.S. Bachmann, J.W. Ramos, PEA15 impairs cell migration and correlates with clinical features predicting good prognosis in neuroblastoma. Int J Cancer 131, 1556–1568 (2012)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  45. D. Geerts, J. Koster, D. Albert, D.L. Koomoa, D.J. Feith, A.E. Pegg, R. Volckmann, H. Caron, R. Versteeg, A.S. Bachmann, The polyamine metabolism genes ornithine decarboxylase and antizyme 2 predict aggressive behavior in neuroblastomas with and without MYCN amplification. Int J Cancer 126, 2012–2024 (2010)

    CAS  PubMed Central  PubMed  Google Scholar 

  46. D. Geerts, C.J. Wallick, D.L. Koomoa, J. Koster, R. Versteeg, R.C. Go, A.S. Bachmann, Expression of prenylated Rab acceptor 1 domain family, member 2 (PRAF2) in neuroblastoma: correlation with clinical features, cellular localization, and cerulenin-mediated apoptosis regulation. Clin Cancer Res 13, 6312–6319 (2007)

    Article  CAS  PubMed  Google Scholar 

  47. I. Lange, D. Geerts, D.J. Feith, G. Mocz, J. Koster, A.S. Bachmann, Novel interaction of ornithine decarboxylase with sepiapterin reductase regulates neuroblastoma cell proliferation. J Mol Biol 426, 332–346 (2014)

    Article  CAS  PubMed  Google Scholar 

  48. L.P. Yco, D. Geerts, J. Koster, A.S. Bachmann, PRAF2 stimulates cell proliferation and migration and predicts poor prognosis in neuroblastoma. Int J Oncol 42, 1408–1416 (2013)

    CAS  PubMed  Google Scholar 

  49. C. Scuoppo, C. Miething, L. Lindqvist, J. Reyes, C. Ruse, I. Appelmann, S. Yoon, A. Krasnitz, J. Teruya-Feldstein, D. Pappin, J. Pelletier, S.W. Lowe, A tumour suppressor network relying on the polyamine-hypusine axis. Nature 487, 244–248 (2012)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  50. J. Lu, Z.P. Chen, Y.P. Yan, S. Knapp, H. Schugar, K.Y. Chen, Aminohexanoic hydroxamate is a potent inducer of the differentiation of mouse neuroblastoma cells. Cancer Lett 160, 59–66 (2000)

    Article  CAS  PubMed  Google Scholar 

  51. A.D. Luchessi, T.D. Cambiaghi, S.M. Hirabara, R.H. Lambertucci, L.R. Silveira, I.L. Baptista, A.S. Moriscot, C.M. Costa-Neto, R. Curi, Involvement of eukaryotic translation initiation factor 5A (eIF5A) in skeletal muscle stem cell differentiation. J Cell Physiol 218, 480–489 (2009)

    Article  CAS  PubMed  Google Scholar 

  52. M. Preukschas, C. Hagel, A. Schulte, K. Weber, K. Lamszus, H. Sievert, N. Pallmann, C. Bokemeyer, J. Hauber, M. Braig, S. Balabanov, Expression of eukaryotic initiation factor 5A and hypusine forming enzymes in glioblastoma patient samples: implications for new targeted therapies. PLoS One 7, e43468 (2012)

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  53. J.J. Molenaar, R. Domingo-Fernandez, M.E. Ebus, S. Lindner, J. Koster, K. Drabek, P. Mestdagh, P. van Sluis, L.J. Valentijn, J. van Nes, M. Broekmans, F. Haneveld, R. Volckmann, I. Bray, L. Heukamp, A. Sprussel, T. Thor, K. Kieckbusch, L. Klein-Hitpass, M. Fischer, J. Vandesompele, A. Schramm, M.M. van Noesel, L. Varesio, F. Speleman, A. Eggert, R.L. Stallings, H.N. Caron, R. Versteeg, J.H. Schulte, LIN28B induces neuroblastoma and enhances MYCN levels via let-7 suppression. Nat Genet 44, 1199–1206 (2012)

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We thank Dr. Jason Shohet (Texas Children’s Hospital, Houston, TX) and Dr. Giselle Sholler (Helen DeVos Children’s Hospital, Grand Rapids, MI) for providing neuroblastoma cell lines MYCN2 and Be(2)-C, respectively. Dr. Patrick Woster (Medical University of South Carolina, Charleston, SC) is thanked for providing us with DFMO. This work was supported by the National Cancer Institute R01 grant CA-111419 (André S. Bachmann), The Daniel K. Inouye College of Pharmacy internal funds (André S. Bachmann), the Dutch Cancer Society (“KWF Kankerbestrijding”) UVA2005-3665 (Dirk Geerts), and the European Union COST Action BM0805 (Dirk Geerts).

Conflict of interest

The authors have declared that no competing interest exists.

Author contributions

A.S.B. and A.B.. designed experiments; A.B., D.G., and J.K. performed experiments and analyzed raw data; and A.S.B., A.B.., and D.G. wrote the paper. All authors reviewed and edited the final manuscript.

Author information

Authors and Affiliations

  1. Department of Pharmaceutical Sciences, The Daniel K. Inouye College of Pharmacy, University of Hawaii at Hilo, 34 Rainbow Drive, Hilo, HI, 96720, USA

    Andrea Bandino & André S. Bachmann

  2. Department of Pediatric Oncology/Hematology, Sophia Children’s Hospital, Erasmus University Medical Center, 3015 GE, Rotterdam, The Netherlands

    Dirk Geerts

  3. Department of Oncogenomics, Academic Medical Center, University of Amsterdam, 1105 AZ, Amsterdam, The Netherlands

    Jan Koster

  4. Department of Cell and Molecular Biology, John A. Burns School of Medicine, University of Hawaii at Manoa, Honolulu, HI, 96813, USA

    André S. Bachmann

  5. Department of Pediatrics and Human Development, College of Human Medicine, Michigan State University, Grand Rapids, MI, 49503, USA

    André S. Bachmann

Authors
  1. Andrea Bandino
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Dirk Geerts
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jan Koster
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. André S. Bachmann
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to André S. Bachmann.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bandino, A., Geerts, D., Koster, J. et al. Deoxyhypusine synthase (DHPS) inhibitor GC7 induces p21/Rb-mediated inhibition of tumor cell growth and DHPS expression correlates with poor prognosis in neuroblastoma patients. Cell Oncol. 37, 387–398 (2014). https://doi.org/10.1007/s13402-014-0201-9

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13402-014-0201-9

Keywords

Search

Navigation