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RNA interference screening identifies a novel role for autocrine fibroblast growth factor signaling in neuroblastoma chemoresistance

Oncogene volume 32, pages 3944–3953 (2013)Cite this article

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Abstract

Chemotherapeutic drug resistance is one of the major causes for treatment failure in high-risk neuroblastoma (NB), the most common extra cranial solid tumor in children. Poor prognosis is typically associated with MYCN amplification. Here, we utilized a loss-of-function kinome-wide RNA interference screen to identify genes that cause cisplatin sensitization. We identified fibroblast growth factor receptor 2 (FGFR2) as an important determinant of cisplatin resistance. Pharmacological inhibition of FGFR2 confirmed the importance of this kinase in NB chemoresistance. Silencing of FGFR2 sensitized NB cells to cisplatin-induced apoptosis, which was regulated by the downregulation of the anti-apoptotic proteins BCL2 and BCLXL. Mechanistically, FGFR2 was shown to activate protein kinase C-δ to induce BCL2 expression. FGFR2, as well as the ligand fibroblast growth factor-2, were consistently expressed in primary NB and NB cell lines, indicating the presence of an autocrine loop. Expression analysis revealed that FGFR2 correlates with MYCN amplification and with advanced stage disease, demonstrating the clinical relevance of FGFR2 in NB. These findings suggest a novel role for FGFR2 in chemoresistance and provide a rational to combine pharmacological inhibitors against FGFR2 with chemotherapeutic agents for the treatment of NB.

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References

  1. Wilson TR, Longley DB, Johnston PG . Chemoresistance in solid tumours. Ann Oncol 2006; 17 (Suppl 10): x315–x324.

    Article  Google Scholar 

  2. Siddik ZH . Cisplatin: mode of cytotoxic action and molecular basis of resistance. Oncogene 2003; 22: 7265–7279.

    Article  CAS  Google Scholar 

  3. Kelland L . The resurgence of platinum-based cancer chemotherapy. Nat Rev Cancer 2007; 7: 573–584.

    Article  CAS  Google Scholar 

  4. Galanski M . Recent developments in the field of anticancer platinum complexes. Recent Pat Anticancer Drug Discov 2006; 1: 285–295.

    Article  CAS  Google Scholar 

  5. Erovic BM, Pelzmann M, Grasl M, Pammer J, Kornek G, Brannath W et al. Mcl-1, vascular endothelial growth factor-R2, and 14-3-3sigma expression might predict primary response against radiotherapy and chemotherapy in patients with locally advanced squamous cell carcinomas of the head and neck. Clin Cancer Res 2005; 11 (24 Pt 1): 8632–8636.

    Article  CAS  Google Scholar 

  6. Michaud WA, Nichols AC, Mroz EA, Faquin WC, Clark JR, Begum S et al. Bcl-2 blocks cisplatin-induced apoptosis and predicts poor outcome following chemoradiation treatment in advanced oropharyngeal squamous cell carcinoma. Clin Cancer Res 2009; 15: 1645–1654.

    Article  CAS  Google Scholar 

  7. Maris JM . Recent advances in neuroblastoma. N Engl J Med 2010; 362: 2202–2211.

    Article  CAS  Google Scholar 

  8. Brodeur GM, Seeger RC, Schwab M, Varmus HE, Bishop JM . Amplification of N-myc in untreated human neuroblastomas correlates with advanced disease stage. Science 1984; 224: 1121–1124.

    Article  CAS  Google Scholar 

  9. Goldsmith KC, Hogarty MD . Targeting programmed cell death pathways with experimental therapeutics: opportunities in high-risk neuroblastoma. Cancer Lett 2005; 228: 133–141.

    Article  CAS  Google Scholar 

  10. MacKeigan JP, Murphy LO, Blenis J . Sensitized RNAi screen of human kinases and phosphatases identifies new regulators of apoptosis and chemoresistance. Nat Cell Biol 2005; 7: 591–600.

    Article  CAS  Google Scholar 

  11. Iorns E, Turner NC, Elliott R, Syed N, Garrone O, Gasco M et al. Identification of CDK10 as an important determinant of resistance to endocrine therapy for breast cancer. Cancer Cell 2008; 13: 91–104.

    Article  CAS  Google Scholar 

  12. Holzel M, Huang S, Koster J, Ora I, Lakeman A, Caron H et al. NF1 is a tumor suppressor in neuroblastoma that determines retinoic acid response and disease outcome. Cell 2010; 142: 218–229.

    Article  Google Scholar 

  13. Diep CH, Munoz RM, Choudhary A, Von Hoff DD, Han H . Synergistic effect between erlotinib and MEK inhibitors in KRAS wild-type human pancreatic cancer cells. Clin Cancer Res 2011; 17: 2744–2756.

    Article  CAS  Google Scholar 

  14. Chou TC . Drug combination studies and their synergy quantification using the Chou-Talalay method. Cancer Res 2010; 70: 440–446.

    Article  CAS  Google Scholar 

  15. Sorenson CM, Barry MA, Eastman A . Analysis of events associated with cell cycle arrest at G2 phase and cell death induced by cisplatin. J Natl Cancer Inst 1990; 82: 749–755.

    Article  CAS  Google Scholar 

  16. Sheridan C, Brumatti G, Elgendy M, Brunet M, Martin SJ . An ERK dependent pathway to Noxa expression regulates apoptosis by platinum-based chemotherapeutic drugs. Oncogene 2010; 29: 6428–6441.

    Article  CAS  Google Scholar 

  17. Turner N, Grose R . Fibroblast growth factor signalling: from development to cancer. Nat Rev Cancer 2010; 10: 116–129.

    Article  CAS  Google Scholar 

  18. Flahaut M, Muhlethaler-Mottet A, Martinet D, Fattet S, Bourloud KB, Auderset K et al. Molecular cytogenetic characterization of doxorubicin-resistant neuroblastoma cell lines: evidence that acquired multidrug resistance results from a unique large amplification of the 7q21 region. Genes Chromosomes Cancer 2006; 45: 495–508.

    Article  CAS  Google Scholar 

  19. Muth D, Ghazaryan S, Eckerle I, Beckett E, Pohler C, Batzler J et al. Transcriptional repression of SKP2 is impaired in MYCN-amplified neuroblastoma. Cancer Res 70: 3791–3802.

    Article  CAS  Google Scholar 

  20. Terrile M, Bryan K, Vaughan L, Hallsworth A, Webber H, Chesler L et al. miRNA expression profiling of the murine TH-MYCN neuroblastoma model reveals similarities with human tumors and identifies novel candidate miRNAs. PLoS One 2011; 6: e28356.

    Article  CAS  Google Scholar 

  21. Cole KA, Huggins J, Laquaglia M, Hulderman CE, Russell MR, Bosse K et al. RNAi screen of the protein kinome identifies checkpoint kinase 1 (CHK1) as a therapeutic target in neuroblastoma. Proc Natl Acad Sci USA 2011; 108: 3336–3341.

    Article  CAS  Google Scholar 

  22. Otto T, Horn S, Brockmann M, Eilers U, Schuttrumpf L, Popov N et al. Stabilization of N-Myc is a critical function of Aurora A in human neuroblastoma. Cancer Cell 2009; 15: 67–78.

    Article  CAS  Google Scholar 

  23. Cole C, Lau S, Backen A, Clamp A, Rushton G, Dive C et al. Inhibition of FGFR2 and FGFR1 increases cisplatin sensitivity in ovarian cancer. Cancer Biol Ther 2010; 10: 495–504.

    Article  CAS  Google Scholar 

  24. Turner N, Lambros MB, Horlings HM, Pearson A, Sharpe R, Natrajan R et al. Integrative molecular profiling of triple negative breast cancers identifies amplicon drivers and potential therapeutic targets. Oncogene 2010; 29: 2013–2023.

    Article  CAS  Google Scholar 

  25. Fanale D, Amodeo V, Corsini LR, Rizzo S, Bazan V, Russo A . Breast cancer genome-wide association studies: there is strength in numbers. Oncogene 2011; 31: 2121–2128.

    Article  Google Scholar 

  26. Byron SA, Pollock PM . FGFR2 as a molecular target in endometrial cancer. Future Oncol 2009; 5: 27–32.

    Article  CAS  Google Scholar 

  27. Katoh M . Cancer genomics and genetics of FGFR2 (review). Int J Oncol 2008; 33: 233–237.

    CAS  PubMed  Google Scholar 

  28. Shukla N, Ameur N, Yilmaz I, Nafa K, Lau CY, Marchetti A et al. Oncogene mutation profiling of pediatric solid tumors reveals significant subsets of embryonal rhabdomyosarcoma and neuroblastoma with mutated genes in growth signaling pathways. Clin Cancer Res 2012; 18: 748–757.

    Article  CAS  Google Scholar 

  29. Eggert A, Ikegaki N, Kwiatkowski J, Zhao H, Brodeur GM, Himelstein BP . High-level expression of angiogenic factors is associated with advanced tumor stage in human neuroblastomas. Clin Cancer Res 2000; 6: 1900–1908.

    CAS  Google Scholar 

  30. Fang H, Harned T, Kalous O, Maldonado V, Declerck YA, Reynolds CP . Synergistic activity of fenretinide and the Bcl-2 family protein inhibitor ABT-737 against human neuroblastoma. Clin Cancer Res 2011; 17: 7093–7104.

    Article  CAS  Google Scholar 

  31. Goldsmith KC, Lestini BJ, Gross M, Ip L, Bhumbla A, Zhang X et al. BH3 response profiles from neuroblastoma mitochondria predict activity of small molecule Bcl-2 family antagonists. Cell Death Differ 2010; 17: 872–882.

    Article  CAS  Google Scholar 

  32. Pardo OE, Wellbrock C, Khanzada UK, Aubert M, Arozarena I, Davidson S et al. FGF-2 protects small cell lung cancer cells from apoptosis through a complex involving PKCepsilon, B-Raf and S6K2. EMBO J 2006; 25: 3078–3088.

    Article  CAS  Google Scholar 

  33. Pardo OE, Arcaro A, Salerno G, Raguz S, Downward J, Seckl MJ . Fibroblast growth factor-2 induces translational regulation of Bcl-XL and Bcl-2 via a MEK-dependent pathway: correlation with resistance to etoposide-induced apoptosis. J Biol Chem 2002; 277: 12040–12046.

    Article  CAS  Google Scholar 

  34. Mattson MP, Meffert MK . Roles for NF-kappaB in nerve cell survival, plasticity, and disease. Cell Death Differ 2006; 13: 852–860.

    Article  CAS  Google Scholar 

  35. Lucas PC, McAllister-Lucas LM, Nunez G . NF-kappaB signaling in lymphocytes: a new cast of characters. J Cell Sci 2004; 117 (Pt 1): 31–39.

    Article  CAS  Google Scholar 

  36. Guo B, Su TT, Rawlings DJ . Protein kinase C family functions in B-cell activation. Curr Opin Immunol 2004; 16: 367–373.

    Article  CAS  Google Scholar 

  37. Lee CH, Shieh DC, Tzeng CY, Chen CP, Wang SP, Chiu YC et al. Bradykinin-induced IL-6 expression through bradykinin B2 receptor, phospholipase C, protein kinase Cdelta and NF-kappaB pathway in human synovial fibroblasts. Mol Immunol 2008; 45: 3693–3702.

    Article  CAS  Google Scholar 

  38. Ma S, Rosen ST . Enzastaurin. Curr Opin Oncol 2007; 19: 590–595.

    Article  CAS  Google Scholar 

  39. Herbst RS, Oh Y, Wagle A, Lahn M . Enzastaurin, a protein kinase Cbeta-selective inhibitor, and its potential application as an anticancer agent in lung cancer. Clin Cancer Res 2007; 13 (15 Pt 2): s4641–s4646.

    Article  Google Scholar 

  40. Ghobrial IM, Munshi NC, Harris BN, Shi P, Porter NM, Schlossman RL et al. A phase I safety study of enzastaurin plus bortezomib in the treatment of relapsed or refractory multiple myeloma. Am J Hematol 2011; 86: 573–578.

    Article  CAS  Google Scholar 

  41. Boller D, Schramm A, Doepfner KT, Shalaby T, von Bueren AO, Eggert A et al. Targeting the phosphoinositide 3-kinase isoform p110delta impairs growth and survival in neuroblastoma cells. Clin Cancer Res 2008; 14: 1172–1181.

    Article  CAS  Google Scholar 

  42. Zhang JH, Chung TD, Oldenburg KRA . Simple statistical parameter for use in evaluation and validation of high throughput screening assays. J Biomol Screen 1999; 4: 67–73.

    Article  CAS  Google Scholar 

  43. Arcaro A, Khanzada UK, Vanhaesebroeck B, Tetley TD, Waterfield MD, Seckl MJ . Two distinct phosphoinositide 3-kinases mediate polypeptide growth factor-stimulated PKB activation. EMBO J 2002; 21: 5097–5108.

    Article  CAS  Google Scholar 

  44. Giulietti A, Overbergh L, Valckx D, Decallonne B, Bouillon R, Mathieu C . An overview of real-time quantitative PCR: applications to quantify cytokine gene expression. Methods 2001; 25: 386–401.

    Article  CAS  Google Scholar 

  45. Nikolova Z, Djonov V, Zuercher G, Andres AC, Ziemiecki A . Cell-type specific and estrogen dependent expression of the receptor tyrosine kinase EphB4 and its ligand ephrin-B2 during mammary gland morphogenesis. J Cell Sci 1998; 111 (Pt 18): 2741–2751.

    CAS  Google Scholar 

  46. Oberthuer A, Berthold F, Warnat P, Hero B, Kahlert Y, Spitz R et al. Customized oligonucleotide microarray gene expression-based classification of neuroblastoma patients outperforms current clinical risk stratification. J Clin Oncol 2006; 24: 5070–5078.

    Article  CAS  Google Scholar 

  47. Sagulenko V, Muth D, Sagulenko E, Paffhausen T, Schwab M, Westermann F . Cathepsin D protects human neuroblastoma cells from doxorubicin-induced cell death. Carcinogenesis 2008; 29: 1869–1877.

    Article  CAS  Google Scholar 

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Acknowledgements

This work was supported by grants from the Gottfried und Julia Bangerter-Rhyner-Stiftung and Jubiläumsstiftung Swiss Life. The research leading to these results has received funding from the European Union’s Seventh Framework Programme (FP7/2007–2013) under grant agreement no. 259348.

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Authors and Affiliations

  1. Department of Clinical Research, University of Bern, Bern, Switzerland

    F Salm, P Cwiek, C Wotzkow, K Höland & A Arcaro

  2. Department of Oncology, University Children’s Hospital Zurich, Zurich, Switzerland

    A Ghosal, A Lucia Buccarello, F Largey & N Bodmer

  3. Institute of Anatomy, University of Bern, Bern, Switzerland

    B Styp-Rekowska & V Djonov

  4. Institute of Pathology, University of Bern, Bern, Switzerland

    I Zlobec & S C Schäfer

  5. Department of Paediatrics, Paediatric Oncology Research, Lausanne University Hospital (CHUV), Lausanne, Switzerland

    N Gross

  6. Department Tumor Genetics, German Cancer Research Center, Heidelberg (DKFZ), Germany

    F Westermann

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Correspondence to A Arcaro.

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Salm, F., Cwiek, P., Ghosal, A. et al. RNA interference screening identifies a novel role for autocrine fibroblast growth factor signaling in neuroblastoma chemoresistance. Oncogene 32, 3944–3953 (2013). https://doi.org/10.1038/onc.2012.416

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  • DOI: https://doi.org/10.1038/onc.2012.416

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