Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Manuscript
  • Published:

Signal Transduction in AML

Constitutive NF-κB DNA-binding activity in AML is frequently mediated by a Ras/PI3-K/PKB-dependent pathway

Leukemia volume 18, pages 103–112 (2004)Cite this article

Abstract

In the present study, we aimed to elucidate the mechanism responsible for constitutive NF-κB DNA-binding activity in AML cells. Intervening in aberrant signaling pathway provides a rational approach for in vivo targeting of AML cells. Constitutive NF-κB DNA-binding activity was observed in 16 of 22 (73%) investigated AML cases and was, in general, associated with resistance to spontaneous apoptosis. Indeed, inhibition of NF-κB activity by the NF-κB inhibitor SN-50 peptide resulted in enhanced chemotherapy-induced apoptosis. In the majority of cases, constitutive NF-κB activity was mediated by a Ras/PI3 kinase (PI3-K)/protein kinase B (PKB)-mediated pathway. The PI3-K inhibitor Ly294002 and the Ras inhibitor L-744832 both inhibited PKB phosphorylation and NF-κB DNA-binding activity. The constitutive activation of Ras GTP-ase was caused by mutations in the gene encoding for N-Ras in 29% of the cases. The constitutive NF-κB activity could so far not be ascribed to the autocrine production of growth factors or to mutations in the Flt3 receptor, since anti-GM-CSF, -IL-1, -IL6, -TNFα or the tyrosine kinase inhibitor AG1296 did not affect the NF-κB DNA-binding activity. The present study demonstrates that Ras activation is an important pathway for triggering the NF-κB pathway in AML cells.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  1. Baldwin Jr AS . The NF-kappa B and I kappa B proteins: new discoveries and insights. Annu Rev Immunol 1996; 14: 649–683.

    Article  CAS  Google Scholar 

  2. Ghosh S, May MJ, Kopp EB . NF-kappa B and Rel proteins: evolutionarily conserved mediators of immune responses. Annu Rev Immunol 1998; 16: 225–260.

    Article  CAS  Google Scholar 

  3. Baeuerle PA, Baichwal VR . NF-kappa B as a frequent target for immunosuppressive and anti-inflammatory molecules. Adv Immunol 1997; 65: 111–137.

    Article  CAS  Google Scholar 

  4. Mayo MW, Baldwin AS . The transcription factor NF-kappaB: control of oncogenesis and cancer therapy resistance. Biochim Biophys Acta 2000; 1470: M55–M62.

    CAS  Google Scholar 

  5. Guzman ML, Neering SJ, Upchurch D, Grimes B, Howard DS, Rizzieri DA et al. Nuclear factor-kappaB is constitutively activated in primitive human acute myelogenous leukemia cells. Blood 2001; 98: 2301–2307.

    Article  CAS  Google Scholar 

  6. Akira S, Kishimoto T . NF-IL6 and NF-kappa B in cytokine gene regulation. Adv Immunol 1997; 65: 1–46.

    Article  CAS  Google Scholar 

  7. Thanos D, Maniatis T . NF-kappa B: a lesson in family values. Cell 1995; 80: 529–532.

    Article  CAS  Google Scholar 

  8. Lee JI, Burckart GJ . Nuclear factor kappa B: important transcription factor and therapeutic target. J Clin Pharmacol 1998; 38: 981–993.

    Article  CAS  Google Scholar 

  9. Maniatis T . Catalysis by a multiprotein IkappaB kinase complex. Science 1997; 278: 818–819.

    Article  CAS  Google Scholar 

  10. Stancovski I, Baltimore D . NF-kappaB activation: the I kappaB kinase revealed? Cell 1997; 91: 299–302.

    Article  CAS  Google Scholar 

  11. Zandi E, Rothwarf DM, Delhase M, Hayakawa M, Karin M . The IkappaB kinase complex (IKK) contains two kinase subunits, IKKalpha and IKKbeta, necessary for IkappaB phosphorylation and NF-kappaB activation. Cell 1997; 91: 243–252.

    Article  CAS  Google Scholar 

  12. Baumgartner B, Weber M, Quirling M, Fischer C, Page S, Adam M et al. Increased IkappaB kinase activity is associated with activated NF-kappaB in acute myeloid blasts. Leukemia 2002; 16: 2062–2071.

    Article  CAS  Google Scholar 

  13. Baumann B, Weber CK, Troppmair J, Whiteside S, Israel A, Rapp UR et al. Raf induces NF-kappaB by membrane shuttle kinase MEKK1, a signaling pathway critical for transformation. Proc Natl Acad Sci USA 2000; 97: 4615–4620.

    Article  CAS  Google Scholar 

  14. Tuyt LM, Dokter WH, Birkenkamp K, Koopmans SB, Lummen C, Kruijer W et al. Extracellular-regulated kinase 1/2, Jun N-terminal kinase, and c-Jun are involved in NF-kappa B-dependent IL-6 expression in human monocytes. J Immunol 1999; 162: 4893–4902.

    CAS  PubMed  Google Scholar 

  15. Bar-Sagi D . Mechanisms of signal transduction by Ras. Semin Cell Biol 1992; 3: 93–98.

    Article  CAS  Google Scholar 

  16. Beaupre DM, Kurzrock R . RAS and leukemia: from basic mechanisms to gene-directed therapy. J Clin Oncol 1999; 17: 1071–1079.

    Article  CAS  Google Scholar 

  17. Kiyoi H, Naoe T, Nakano Y, Yokota S, Minami S, Miyawaki S et al. Prognostic implication of FLT3 and N-RAS gene mutations in acute myeloid leukemia. Blood 1999; 93: 3074–3080.

    CAS  Google Scholar 

  18. Lee Jr JT, McCubrey JA . The Raf/MEK/ERK signal transduction cascade as a target for chemotherapeutic intervention in leukemia. Leukemia 2002; 16: 486–507.

    Article  CAS  Google Scholar 

  19. Downward J . Mechanisms and consequences of activation of protein kinase B/Akt. Curr Opin Cell Biol 1998; 10: 262–267.

    Article  CAS  Google Scholar 

  20. Shields JM, Pruitt K, McFall A, Shaub A, Der CJ . Understanding Ras: ‘it ain't over ‘til it's over’. Trends Cell Biol 2000; 10: 147–154.

    Article  CAS  Google Scholar 

  21. Irani K, Xia Y, Zweier JL, Sollott SJ, Der CJ, Fearon ER et al. Mitogenic signaling mediated by oxidants in Ras-transformed fibroblasts. Science 1997; 275: 1649–1652.

    Article  CAS  Google Scholar 

  22. O'Gorman DM, McKenna SL, McGahon AJ, Knox KA, Cotter TG . Sensitisation of HL60 human leukaemic cells to cytotoxic drug-induced apoptosis by inhibition of PI3-kinase survival signals. Leukemia 2000; 14: 602–611.

    Article  CAS  Google Scholar 

  23. Bennett JM, Catovsky D, Daniel MT, Flandrin G, Galton DA, Gralnick HR et al. Proposed revised criteria for the classification of acute myeloid leukemia. A report of the French–American–British Cooperative Group. Ann Intern Med 1985; 103: 620–625.

    Article  CAS  Google Scholar 

  24. Schreiber E, Matthias P, Muller MM, Schaffner W . Rapid detection of octamer binding proteins with ‘mini-extracts’, prepared from a small number of cells. Nucleic Acids Res 1989; 17: 6419.

    Article  CAS  Google Scholar 

  25. M'Rabet L, Coffer PJ, Wolthuis RM, Zwartkruis F, Koenderman L, Bos JL . Differential fMet-Leu-Phe- and platelet-activating factor-induced signaling toward Ral activation in primary human neutrophils. J Biol Chem 1999; 274: 21847–21852.

    Article  CAS  Google Scholar 

  26. Birkenkamp KU, Geugien M, Lemmink HH, Kruijer W, Vellenga E . Regulation of constitutive STAT5 phosphorylation in acute myeloid leukemia blasts. Leukemia 2001; 15: 1923–1931.

    Article  CAS  Google Scholar 

  27. Hayes VM, Westra JL, Verlind E, Bleeker W, Plukker JT, Hofstra RM et al. New comprehensive denaturing-gradient-gel-electrophoresis assay for KRAS mutation detection applied to paraffin-embedded tumours. Genes Chromosomes Cancer 2000; 29: 309–314.

    Article  CAS  Google Scholar 

  28. Mizuki M, Fenski R, Halfter H, Matsumura I, Schmidt R, Muller C et al. Flt3 mutations from patients with acute myeloid leukemia induce transformation of 32D cells mediated by the Ras and STAT5 pathways. Blood 2000; 96: 3907–3914.

    CAS  PubMed  Google Scholar 

  29. Hayakawa F, Towatari M, Kiyoi H, Tanimoto M, Kitamura T, Saito H et al. Tandem-duplicated Flt3 constitutively activates STAT5 and MAP kinase and introduces autonomous cell growth in IL-3-dependent cell lines. Oncogene 2000; 19: 624–631.

    Article  CAS  Google Scholar 

  30. Lundin C, Schultz N, Arnaudeau C, Mohindra A, Hansen LT, Helleday T . RAD51 is involved in repair of damage associated with DNA replication in mammalian cells. J Mol Biol 2003; 328: 521–535.

    Article  CAS  Google Scholar 

  31. Birkenkamp KU, Dokter WH, Esselink MT, Jonk LJ, Kruijer W, Vellenga E . A dual function for p38 MAP kinase in hematopoietic cells: involvement in apoptosis and cell activation. Leukemia 1999; 13: 1037–1045.

    Article  CAS  Google Scholar 

  32. Ghoda L, Lin X, Greene WC . The 90-kDa ribosomal S6 kinase (pp90rsk) phosphorylates the N-terminal regulatory domain of IkappaBalpha and stimulates its degradation in vitro. J Biol Chem 1997; 272: 21281–21288.

    Article  CAS  Google Scholar 

  33. Coffer PJ, Jin J, Woodgett JR . Protein kinase B (c-Akt): a multifunctional mediator of phosphatidylinositol 3-kinase activation. Biochem J 1998; 335 (Part 1): 1–13.

    Article  CAS  Google Scholar 

  34. Ozes ON, Mayo LD, Gustin JA, Pfeffer SR, Pfeffer LM, Donner DB . NF-kappaB activation by tumour necrosis factor requires the Akt serine–threonine kinase. Nature 1999; 401: 82–85.

    Article  CAS  Google Scholar 

  35. Romashkova JA, Makarov SS . NF-kappaB is a target of AKT in anti-apoptotic PDGF signalling. Nature 1999; 401: 86–90.

    Article  CAS  Google Scholar 

  36. Alessi DR, Andjelkovic M, Caudwell B, Cron P, Morrice N, Cohen P et al. Mechanism of activation of protein kinase B by insulin and IGF-1. EMBO J 1996; 15: 6541–6551.

    Article  CAS  Google Scholar 

  37. Finco TS, Westwick JK, Norris JL, Beg AA, Der CJ, Baldwin Jr AS . Oncogenic Ha-Ras-induced signaling activates NF-kappaB transcriptional activity, which is required for cellular transformation. J Biol Chem 1997; 272: 24113–24116.

    Article  CAS  Google Scholar 

  38. Van Antwerp DJ, Martin SJ, Kafri T, Green DR, Verma IM . Suppression of TNF-alpha-induced apoptosis by NF-kappaB. Science 1996; 274: 787–789.

    Article  CAS  Google Scholar 

  39. Beg AA, Baltimore D . An essential role for NF-kappaB in preventing TNF-alpha-induced cell death. Science 1996; 274: 782–784.

    Article  CAS  Google Scholar 

  40. Reuther GW, Lambert QT, Rebhun JF, Caligiuri MA, Quilliam LA, Der CJ . RasGRP4 is a novel Ras activator isolated from acute myeloid leukemia. J Biol Chem 2002; 277: 30508–30514.

    Article  CAS  Google Scholar 

  41. Milella M, Kornblau SM, Estrov Z, Carter BZ, Lapillonne H, Harris D et al. Therapeutic targeting of the MEK/MAPK signal transduction module in acute myeloid leukemia. J Clin Invest 2001; 108: 851–859.

    Article  CAS  Google Scholar 

  42. Dokter WH, Tuyt L, Sierdsema SJ, Esselink MT, Vellenga E . The spontaneous expression of interleukin-1 beta and interleukin-6 is associated with spontaneous expression of AP-1 and NF-kappa B transcription factor in acute myeloblastic leukemia cells. Leukemia 1995; 9: 425–432.

    CAS  PubMed  Google Scholar 

  43. Schuringa JJ, Wierenga AT, Kruijer W, Vellenga E . Constitutive Stat3, Tyr705, and Ser727 phosphorylation in acute myeloid leukemia cells caused by the autocrine secretion of interleukin-6. Blood 2000; 95: 3765–3770.

    CAS  PubMed  Google Scholar 

  44. Schepers H, Geugien M, Eggen BJL, Vellenga E . Constitutive cytoplasmic localization of p21Waf1/Cip1 affects the apoptotic process in monocytic leukemia. Leukemia 2003; 17: 2113–2121.

    Article  CAS  Google Scholar 

  45. Pennington KN, Taylor JA, Bren GD, Paya CV . IkappaB kinase-dependent chronic activation of NF-kappaB is necessary for p21(WAF1/Cip1) inhibition of differentiation-induced apoptosis of monocytes. Mol Cell Biol 2001; 21: 1930–1941.

    Article  CAS  Google Scholar 

  46. Asada M, Yamada T, Ichijo H, Delia D, Miyazono K, Fukumuro K et al. Apoptosis inhibitory activity of cytoplasmic p21(Cip1/WAF1) in monocytic differentiation. EMBO J 1999; 18: 1223–1234.

    Article  CAS  Google Scholar 

  47. Asada M, Yamada T, Fukumuro K, Mizutani S . p21Cip1/WAF1 is important for differentiation and survival of U937 cells. Leukemia 1998; 12: 1944–1950.

    Article  CAS  Google Scholar 

  48. Mayo MW, Wang CY, Cogswell PC, Rogers-Graham KS, Lowe SW, Der CJ et al. Requirement of NF-kappaB activation to suppress p53-independent apoptosis induced by oncogenic Ras. Science 1997; 278: 1812–1815.

    Article  CAS  Google Scholar 

  49. Vellenga E, Ostapovicz D, O'Rourke B, Griffin JD . Effects of recombinant IL-3, GM-CSF, and G-CSF on proliferation of leukemic clonogenic cells in short-term and long-term cultures. Leukemia 1987; 1: 584–589.

    CAS  PubMed  Google Scholar 

  50. Birkenkamp KU, Esselink MT, Kruijer W, Vellenga E . Differential effects of interleukin-3 and interleukin-1 on the proliferation and interleukin-6 protein secretion of acute myeloid leukemic cells; the involvement of ERK, p38 and STAT5. Eur Cytokine Netw 1999; 10: 479–490.

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Dr Robert Hofstra (Department of Medical Genetics, University of Groningen, Groningen, The Netherlands) for his cooperation in screening for mutations in the K-Ras gene by DGGE analysis.

This study was supported by grant RUG 99-1944 from the Dutch Cancer Foundation.

Author information

Author notes

  1. Contributed equally to this work

Authors and Affiliations

  1. Division of Hematology, Department of Medicine, University of Groningen, The Netherlands

    K U Birkenkamp, M Geugien, H Schepers & E Vellenga

  2. Department of Medical Genetics, University of Groningen, The Netherlands

    J Westra & H H Lemmink

Authors
  1. K U Birkenkamp
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. M Geugien
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. H Schepers
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. J Westra
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. H H Lemmink
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. E Vellenga
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Birkenkamp, K., Geugien, M., Schepers, H. et al. Constitutive NF-κB DNA-binding activity in AML is frequently mediated by a Ras/PI3-K/PKB-dependent pathway. Leukemia 18, 103–112 (2004). https://doi.org/10.1038/sj.leu.2403145

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.leu.2403145

Keywords

This article is cited by

Search

Advanced search

Quick links