Targeted Oncology

, Volume 11, Issue 2, pp 183–195 | Cite as

Inhibition of Survival Pathways MAPK and NF-kB Triggers Apoptosis in Pancreatic Ductal Adenocarcinoma Cells via Suppression of Autophagy

  • Daniela Laura Papademetrio
  • Silvina Laura Lompardía
  • Tania Simunovich
  • Susana Costantino
  • Cintia Yamila Mihalez
  • Victoria Cavaliere
  • Élida Álvarez
Original Research Article



Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease with a survival rate of 4–6 months from diagnosis. PDAC is the fourth leading cause of cancer-related death in the Western world, with a mortality rate of 10 cases per 100,000 population. Chemotherapy constitutes only a palliative strategy, with limited effects on life expectancy.


To investigate the biological response of PDAC to mitogen-activated protein kinase (MAPK) and NF-kappaB (NF-kB) inhibitors and the role of autophagy in the modulation of these signaling pathways in order to address the challenge of developing improved medical protocols for patients with PDAC.


Two ATCC cell lines, MIAPaCa-2 and PANC-1, were used as PDAC models. Cells were exposed to inhibitors of MAPK or NF-kB survival pathways alone or after autophagy inhibition. Several aspects were analyzed, as follows: cell proliferation, by [3H]TdR incorporation; cell death, by TUNEL assay, regulation of autophagy by LC3-II expression level and modulation of pro-and anti-apoptotic proteins by Western blot.


We demonstrated that the inhibition of the MAPK and NF-kB survival pathways with U0126 and caffeic acid phenethyl ester (CAPE), respectively, produced strong inhibition of pancreatic tumor cell growth without inducing apoptotic death. Interestingly, U0126 and CAPE induced apoptosis after autophagy inhibition in a caspase-dependent manner in MIA PaCa-2 cells and in a caspase-independent manner in PANC-1 cells.


Here we present evidence that allows us to consider a combined therapy regimen comprising an autophagy inhibitor and a MAPK or NF-kB pathway inhibitor as a possible treatment strategy for pancreatic cancer.


Gemcitabine Pancreatic Ductal Adenocarcinoma Propolis Caffeic Acid Phenethyl Ester PDAC Cell 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



The authors thank Dr. Daniela Ureta (Servicio de Citometría de flujo, Departamento de Microbiología, Inmunología y Biotecnología, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Argentina) for technical assistance and Martín Levermann for language assistance during the edition of the manuscript.

Compliance with Ethical Standards


This study was funded by Universidad de Buenos Aires and Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET).

Conflict of Interest

The authors (DLP, SLL, TS, SC, CYM, VC, EA) declared no conflict of interest.


  1. 1.
    Hidalgo M (2010) Pancreatic cancer. N Engl J Med 362(17):1605–17CrossRefPubMedGoogle Scholar
  2. 2.
    Hidalgo M (2012) New insights into pancreatic cancer biology. Ann Oncol 23(Suppl 10):135–8CrossRefGoogle Scholar
  3. 3.
    Siegel R, Naishadham D, Jemal A (2012) Cancer statistics. CA Cancer J Clin 62:10–29CrossRefPubMedGoogle Scholar
  4. 4.
    O'Reilly KE, Rojo F, She QB et al (2006) mTOR inhibition induces upstream receptor tyrosine kinase signaling and activates Akt. Cancer Res 66:1500–8CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Garcia MG, Alaniz LD, Cordo Russo RI et al (2009) PI3K/Akt inhibition modulates multidrug resistance and activates NFkB in murine Lymphoma cell lines. Leuk Res 33:288–96CrossRefPubMedGoogle Scholar
  6. 6.
    Cox AD, Der CJ (1997) Farnesyltransferase inhibitors and cancer treatment: targeting simply Ras? Biochim Biophys Acta 1333(1):F51–71PubMedGoogle Scholar
  7. 7.
    Muerkoster S, Arlt A, Sipos B et al (2005) Increased expression of the E3-ubiquitin ligase receptor subunit betaTRCP1 relates to constitutive nuclear factor-kappaB activation and chemoresistance in pancreatic carcinoma cells. Cancer Res 65(4):1316–24CrossRefPubMedGoogle Scholar
  8. 8.
    Aksamitiene E, Kiyatkin A, Kholodenko BN (2012) Cross-talk between mitogenic Ras/MAPK and survival PI3K/Akt pathways: a fine balance. Biochem Soc Trans 40:139–46CrossRefPubMedGoogle Scholar
  9. 9.
    De Luca A, Maiello MR, D'Alessio A et al (2012) The RAS/RAF/MEK/ERK and the PI3K/AKT signalling pathways: role in cancer pathogenesis and implications for therapeutic approaches. Expert Opin Ther Targets 16:S17–27CrossRefPubMedGoogle Scholar
  10. 10.
    Shimizu T, Tolcher AW, Papadopoulos KP et al (2012) The clinical effect of the dual-targeting strategy involving PI3K/AKT/mTOR and RAS/MEK/ERK pathways in patients with advanced cancer. Clin Cancer Res 18:2316–25CrossRefPubMedGoogle Scholar
  11. 11.
    Wang LH (2014) LiY, Yang SN, et al. Gambogic acid synergistically potentiates cisplatin-induced apoptosis in non-small-cell lung cancer through suppressing NF-κB and MAPK/HO-1 signalling. Br J Cancer 110(2):34–52CrossRefGoogle Scholar
  12. 12.
    Sylvester RJ, van der Meijden AP, Oosterlinck W et al (2006) Predicting recurrence and progression in individual patients with stage Ta T1 bladder cancer using EORTC risk tables: a combined analysis of 2596 patients from seven EORTC trials. Eur Urol 49:466–75CrossRefPubMedGoogle Scholar
  13. 13.
    Vaux DL, Silke J (2003) Mammalian mitochondrial IAP binding proteins. Biochem Biophys Res Commun 304:499–504CrossRefPubMedGoogle Scholar
  14. 14.
    Wei Y, Fan T, Yu M (2008) Inhibitor of apoptosis proteins and apoptosis. Acta Biochim Biophys Sin (Shanghai) 40:278–88CrossRefGoogle Scholar
  15. 15.
    Srinivasula SM, Ashwell JD (2008) IAPs: what's in a name? Mol Cell 30:123–35CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Dubrez-Daloz L, Dupoux A, Cartier J (2008) IAPs: more than just inhibitors of apoptosis proteins. Cell Cycle 7:1036–46CrossRefPubMedGoogle Scholar
  17. 17.
    LaCasse EC, Mahoney DJ, Cheung HH et al (2008) IAP-targeted therapies for cancer. Oncogene 27:6252–75CrossRefPubMedGoogle Scholar
  18. 18.
    Vucic D, Fairbrother WJ (2007) The inhibitor of apoptosis proteins as therapeutic targets in cancer. Clin Cancer Res 13:5995–6000CrossRefPubMedGoogle Scholar
  19. 19.
    Reggiori F, Klionsky DJ (2002) Autophagy in the eukaryotic cell. Eukaryot Cell 1(1):11–21CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Shintani T, Klionsky DJ (2004) Autophagy in health and disease: a double-edged sword. Science 306(5698):990–5CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Kirkegaard K, Taylor MP, Jackson WT (2004) Cellular autophagy: surrender, avoidance and subversion by microorganisms. Nat Rev Microbiol 2(4):301–14CrossRefPubMedGoogle Scholar
  22. 22.
    Ogawa M, Yoshimori T, Suzuki T et al (2005) Escape of intracellular Shigella from autophagy. Science 307(5710):727–31CrossRefPubMedGoogle Scholar
  23. 23.
    Hara T, Nakamura K, Matsui M et al (2006) Suppression of basal autophagy in neural cells causes neurodegenerative disease in mice. Nature 441(7095):885–9CrossRefPubMedGoogle Scholar
  24. 24.
    Komatsu M, Waguri S, Chiba T et al (2006) Loss of autophagy in the central nervous system causes neurodegeneration in mice. Nature 441(7095):880–4CrossRefPubMedGoogle Scholar
  25. 25.
    Liang XH, Jackson S, Seaman M et al (1999) Induction of autophagy and inhibition of tumorigenesis by beclin 1. Nature 402(6762):672–6CrossRefPubMedGoogle Scholar
  26. 26.
    Liang XH, Yu J, Brown K et al (2001) Beclin 1 contains a leucine-rich nuclear export signal that is required for its autophagy and tumor suppressor function. Cancer Res 61(8):3443–9PubMedGoogle Scholar
  27. 27.
    Ravikumar B, Berger Z, Vacher C et al (2006) Rapamycin pre-treatment protects against apoptosis. Hum Mol Genet 15(7):1209–16CrossRefPubMedGoogle Scholar
  28. 28.
    Papademetrio DL, Cavaliere V, Simunovich T et al (2014) Interplay between autophagy and apoptosis in pancreatic tumors in response to gemcitabine. Target Oncol 9(2):123–34CrossRefPubMedGoogle Scholar
  29. 29.
    Kabeya Y, Mizushima N, Ueno T et al (2000) LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing. EMBO J 19(21):5720–8CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Rubinsztein DC, Cuervo AM, Ravikumar B et al (2009) In search of an "autophagomometer". Autophagy 5(5):585–9CrossRefPubMedGoogle Scholar
  31. 31.
    Conroy T, Desseigne F, Ychou M et al (2001) FOLFIRINOX versus gemcitabine for metastatic pancreatic cancer. N Engl J Med 364:1817–25CrossRefGoogle Scholar
  32. 32.
    Hill R, Rabb M, Madureira PA et al (2013) Gemcitabine-mediated tumour regression and p53-dependent gene expression: implications for colon and pancreatic cancer therapy. Cell Death Dis 4:e791CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Jones S, Zhang X, Parsons DW et al (2008) Core signaling pathways in human pancreatic cancers revealed by global genomic analyses. Science 321:1801–6CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Biankin AV, Waddell N, Kassahn KS et al (2012) Pancreatic cancer genomes reveal aberrations in axon guidance pathway genes. Nature 491:399–405CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Baines AT, Xu D, Der CJ (2011) Inhibition of Ras for cancer treatment: the search continues. Future Med Chem 3:1787–808CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Chen Z, Cheng K, Walton Z et al (2012) A murine lung cancer coclinical trial identifies genetic modifiers of therapeutic response. Nature 483(7391):613–7CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Jänne PA, Shaw AT, Pereira JR et al (2013) Selumetinib plus docetaxel for KRAS-mutant advanced non-small-cell lung cancer: a randomised, multicentre, placebo-controlled, phase 2 study. Lancet Oncol 14:38–47CrossRefPubMedGoogle Scholar
  38. 38.
    McCubrey JA, Steelman LS, Chappell WH et al (2012) Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR cascade inhibitors: how mutations can result in therapy resistance and how to overcome resistance. Oncotarget 3:1068–111CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Fujioka S, Sclabas GM, Schmidt C et al (2003) Function of nuclear factor kappaB in pancreatic cancer metastasis. Clin Cancer Res 9:346–54PubMedGoogle Scholar
  40. 40.
    Hu L, Shi Y, Hsu JH et al (2003) Downstream effectors of oncogenic ras in multiple myeloma cells. Blood 101:3126–35CrossRefPubMedGoogle Scholar
  41. 41.
    Mayo MW, Wang CY, Cogswell PC et al (1997) Requirement of NF-kappaB activation to suppress p53-independent apoptosis induced by oncogenic Ras. Science 278:1812–5CrossRefPubMedGoogle Scholar
  42. 42.
    Li L, Aggarwal BB, Shishodia S et al (2004) Nuclear factor-kappaB and IkappaB kinase are constitutively active in human pancreatic cells, and their down-regulation by curcumin (diferuloylmethane) is associated with the suppression of proliferation and the induction of apoptosis. Cancer 101:2351–62CrossRefPubMedGoogle Scholar
  43. 43.
    Aggarwal BB (2004) Nuclear factor-kappaB: the enemy within. Cancer Cell 6:203–8CrossRefPubMedGoogle Scholar
  44. 44.
    Yamamoto Y, Gaynor RB (2001) Role of the NF-kappaB pathway in the pathogenesis of human disease states. Curr Mol Med 1:287–96CrossRefPubMedGoogle Scholar
  45. 45.
    Aggarwal BB, Takada Y, Shishodia S et al (2004) Nuclear transcription factor NF-kappa B: role in biology and medicine. Indian J Exp Biol 42:341–53PubMedGoogle Scholar
  46. 46.
    Karin M, Cao Y, Greten FR et al (2002) NF-kappaB in cancer: from innocent bystander to major culprit. Nat Rev Cancer 2:301–10CrossRefPubMedGoogle Scholar
  47. 47.
    Garg A, Aggarwal BB (2002) Nuclear transcription factor-kappaB as a target for cancer drug development. Leukemia 16:1053–68CrossRefPubMedGoogle Scholar
  48. 48.
    Lin Y, Shi R, Wang X et al (2008) Luteolin, a flavonoid with potential for cancer prevention and therapy. Curr Cancer Drug Targets 8:634–46CrossRefPubMedPubMedCentralGoogle Scholar
  49. 49.
    Cai X, Lu W, Yang Y et al (2013) Digitoflavone inhibits IκBα kinase and enhances apoptosis induced by TNFα through downregulation of expression of nuclear factor κB-regulated gene products in human pancreatic cancer cells. PLoS One 8(10):e77126CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Cavaliere V, Papademetrio DL, Lorenzetti M et al (2009) Caffeic Acid Phenylethyl Ester and MG-132 have apoptotic and antiproliferative effects on Leukemic cells but not on normal mononuclear cells. Transl Oncol 2(1):46–58CrossRefPubMedPubMedCentralGoogle Scholar
  51. 51.
    Velculescu VE (1999) Essay: Amersham Pharmacia Biotech & Science prize. Tantalizing transcriptomes SAGE and its use in global gene expression analysis. Science 286(5444):1491–2CrossRefPubMedGoogle Scholar
  52. 52.
    Altieri DC (2008) Survivin, cancer networks and pathway-directed drug discovery. Nat Rev Cancer 8(1):61–70CrossRefPubMedGoogle Scholar
  53. 53.
    Sarela AI, Macadam RC, Farmery SM et al (2000) Expression of the antiapoptosis gene, survivin, predicts death from recurrent colorectal carcinoma. Gut 46(5):645–50CrossRefPubMedPubMedCentralGoogle Scholar
  54. 54.
    Monzo M, Rosell R, Felip E et al (1999) A novel anti-apoptosis gene: Re-expression of survivin messenger RNA as a prognosis marker in non-small-cell lung cancers. J Clin Oncol 17(7):2100–4PubMedGoogle Scholar
  55. 55.
    Shariat SF, Lotan Y, Saboorian H et al (2004) Survivin expression is associated with features of biologically aggressive prostate carcinoma. Cancer 100(4):751–7CrossRefPubMedGoogle Scholar
  56. 56.
    Tanaka K, Iwamoto S, Gon G et al (2000) Expression of survivin and its relationship to loss of apoptosis in breast carcinomas. Clin Can 6(1):127–34Google Scholar
  57. 57.
    Jourdan M, Reme T, Goldschmidt H et al (2009) Gene expression of anti- and pro-apoptotic proteins in malignant and normal plasma cells. Br J Haematol 145(1):45–58CrossRefPubMedPubMedCentralGoogle Scholar
  58. 58.
    Cheng SM, Chang YC, Liu CY et al (2015) YM155 down-regulates survivin and XIAP, modulates autophagy and induces autophagy-dependent DNA damage in breast cancer cells. Br J Pharmacol 172(1):214–34CrossRefPubMedGoogle Scholar
  59. 59.
    Wang J, Whiteman MW, Lian H et al (2009) A Non-canonical MEK/ERK Signaling Pathway Regulates Autophagy via Regulating Beclin 1. J Biol Chem 284(32):21412–24CrossRefPubMedPubMedCentralGoogle Scholar
  60. 60.
    Pattingre S, Bauvy C, Codogno PZ (2003) Amino acids interfere with the ERK1⁄ 2-dependent control of macroautophagy by controlling the activation of Raf-1 in human colon cancer HT-29 cells. J Biol Chem 278:16667–74CrossRefPubMedGoogle Scholar
  61. 61.
    Ellington AA, Berhow MA, Singletary KW (2006) Inhibition of Akt signaling and enhanced ERK1⁄ 2 activity are involved in induction of macroautophagy by triterpenoid B-group soyasaponins in colon cancer cells. Carcinogenesis 27:298–306CrossRefPubMedGoogle Scholar
  62. 62.
    Copetti T, Demarchi F, Schneider C (2009) p65/RelA binds and activates the beclin 1 promoter. Autophagy 5(6):858–9CrossRefPubMedGoogle Scholar
  63. 63.
    Vadlamudi RK, Shin J (1998) Genomic structure and promoter analysis of the p62 gene encoding a nonproteasomal multiubiquitin chain binding protein. FEBS Lett 435:138–42CrossRefPubMedGoogle Scholar
  64. 64.
    David A (2014) An autophagic switch in the response of tumor cells to radiation and chemotherapy. Biochem Pharmacol 90:208–11CrossRefGoogle Scholar
  65. 65.
    Yang S, Wang X, Contino G et al (2011) Pancreatic cancers require autophagy for tumor growth. Genes Dev 25(7):717–29CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer International Publishing Switzerland 2015

Authors and Affiliations

  • Daniela Laura Papademetrio
    • 1
    • 2
  • Silvina Laura Lompardía
    • 1
    • 2
  • Tania Simunovich
    • 1
  • Susana Costantino
    • 1
    • 2
  • Cintia Yamila Mihalez
    • 1
    • 2
  • Victoria Cavaliere
    • 1
  • Élida Álvarez
    • 1
    • 2
  1. 1.Cátedra de Inmunología, Facultad de Farmacia y BioquímicaUniversidad de Buenos AiresCiudad Autónoma de Buenos AiresArgentina
  2. 2.IDEHUCONICETCiudad Autónoma de Buenos AiresArgentina

Personalised recommendations