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Effects of Raf kinase inhibitor protein expression on pancreatic cancer cell growth and motility: an in vivo and in vitro study

Journal of Cancer Research and Clinical Oncology volume 142, pages 2107–2117 (2016)Cite this article

Abstract

Purpose

Raf kinase inhibitor protein (RKIP) is a tumor suppressor that inhibits cell growth and metastasis of malignant tumors. Pancreatic cancer is a leading cause of cancer death with a low survival rate. RKIP expression and its role in tumorigenesis and metastasis in pancreatic cancer are poorly understood. The aims of our study were to assess the effects of RKIP on pancreatic carcinoma cells in vitro and in tumor tissues in vivo.

Methods

This study included 84 patients with histologically confirmed pancreatic adenocarcinoma. The expression levels of RKIP were measured in pancreatic cancer tissues and adjacent normal tissues using real-time PCR and immunohistochemistry. Overexpression plasmid of RKIP was transfected into SW1990 and AsPC-1 cell lines, and the effects on cell proliferation were studied using a Cell Counting Kit-8 assay. MEK1/2 and ERK1/2 were detected by Western blot and immunofluorescence assay.

Results

Results showed a reduced expression of RKIP in pancreatic carcinoma tissues compared with adjacent normal tissues, which closely correlated with patient outcomes. Overexpression of RKIP suppressed cell proliferation and promoted apoptosis in cultured SW1990 and AsPC-1 cell lines. Transwell assay showed RKIP can inhibit cell migration and invasion, and in vivo RKIP can suppress tumorigenesis by diminishing the volume of the tumors.

Conclusions

In conclusion, expression of RKIP is closely correlated with the survival of pancreatic cancer patients. RKIP can inhibit pancreatic adenocarcinoma cells proliferation, activities of migration and invasion, through downregulating Raf-1-MEK1/2-ERK1/2 signaling pathway.

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References

  • Arlt A, Gehrz A, Müerköster S, Vorndamm J, Kruse M-L, Fölsch UR, Schäfer H (2003) Role of NF-κB and Akt/PI3 K in the resistance of pancreatic carcinoma cell lines against gemcitabine-induced cell death. Oncogene 22:3243–3251

    CAS  Article  PubMed  Google Scholar 

  • Beshir AB, Ren G, Magpusao AN, Barone LM, Yeung KC, Fenteany G (2010) Raf kinase inhibitor protein suppresses nuclear factor-κB-dependent cancer cell invasion through negative regulation of matrix metalloproteinase expression. Cancer Lett 299:137–149

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Chaffer CL, Weinberg RA (2011) A perspective on cancer cell metastasis. Science 331:1559–1564

    CAS  Article  PubMed  Google Scholar 

  • Chambers AF, Groom AC, MacDonald IC (2002) Metastasis: dissemination and growth of cancer cells in metastatic sites. Nat Rev Cancer 2:563–572

    CAS  Article  PubMed  Google Scholar 

  • Correia DV, d’Orey F, Cardoso BA, Lanca T, Grosso AR, deBarros A, Martins LR, Barata JT, Silva-Santos B (2009) Highly active microbial phosphoantigen induces rapid yet sustained MEK/Erk- and PI-3K/Akt-mediated signal transduction in anti-tumor human gammadelta T-cells. PLoS One 4:e5657

    Article  PubMed  PubMed Central  Google Scholar 

  • Edin ML, Juliano RL (2005) Raf-1 serine 338 phosphorylation plays a key role in adhesion-dependent activation of extracellular signal-regulated kinase by epidermal growth factor. Mol Cell Biol 25:4466–4475

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Ehrenreiter K, Piazzolla D, Velamoor V, Sobczak I, Small JV, Takeda J, Leung T, Baccarini M (2005) Raf-1 regulates Rho signaling and cell migration. J Cell Biol 168:955–964

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Escara-Wilke J, Yeung K, Keller ET (2012) Raf kinase inhibitor protein (RKIP) in cancer. Cancer Metastasis Rev 31:615–620

    CAS  Article  PubMed  Google Scholar 

  • Fatima I, Chandra V, Saxena R, Manohar M, Sanghani Y, Hajela K, Negi MP, Sankhwar PL, Jain SK, Dwivedi A (2012) 2,3-Diaryl-2H-1-benzopyran derivatives interfere with classical and non-classical estrogen receptor signaling pathways, inhibit Akt activation and induce apoptosis in human endometrial cancer cells. Mol Cell Endocrinol 348:198–210

    CAS  Article  PubMed  Google Scholar 

  • Fu Z, Smith PC, Zhang L, Rubin MA, Dunn RL, Yao Z, Keller ET (2003) Effects of raf kinase inhibitor protein expression on suppression of prostate cancer metastasis. J Natl Cancer Inst 95:878–889

    CAS  Article  PubMed  Google Scholar 

  • Fu Z, Kitagawa Y, Shen R, Shah R, Mehra R, Rhodes D, Keller PJ, Mizokami A, Dunn RL, Chinnaiyan AM (2006) Metastasis suppressor gene raf kinase inhibitor protein (RKIP) is a novel prognostic marker in prostate cancer. Prostate 66:248–256

    CAS  Article  PubMed  Google Scholar 

  • Fujioka S, Sclabas GM, Schmidt C, Frederick WA, Dong QG, Abbruzzese JL, Evans DB, Baker C, Chiao PJ (2003) Function of nuclear factor κB in pancreatic cancer metastasis. Clin Cancer Res 9:346–354

    CAS  PubMed  Google Scholar 

  • Gollob JA, Wilhelm S, Carter C, Kelley SL (2006) Role of Raf kinase in cancer: therapeutic potential of targeting the Raf/MEK/ERK signal transduction pathway. Semin Oncol 33:392–406

    CAS  Article  PubMed  Google Scholar 

  • Gray-Schopfer VC, da Rocha Dias S, Marais R (2005) The role of B-RAF in melanoma. Cancer Metastasis Rev 24:165–183

    CAS  Article  PubMed  Google Scholar 

  • Hagan S, Al-Mulla F, Mallon E, Oien K, Ferrier R, Gusterson B, García JJC, Kolch W (2005) Reduction of Raf-1 kinase inhibitor protein expression correlates with breast cancer metastasis. Clin Cancer Res 11:7392–7397

    CAS  Article  PubMed  Google Scholar 

  • He Y, Zheng R, Li D, Zeng H, Zhang S, Chen W (2015) Pancreatic cancer incidence and mortality patterns in China, 2011. Chin J Cancer Res 27:29–37  

    PubMed  PubMed Central  Google Scholar 

  • Hidalgo M (2010) Pancreatic cancer. N Engl J Med 362:1605–1617

    CAS  Article  PubMed  Google Scholar 

  • Holcomb B, Yip-Schneider M, Schmidt CM (2008) The role of nuclear factor κB in pancreatic cancer and the clinical applications of targeted therapy. Pancreas 36:225–235

    CAS  Article  PubMed  Google Scholar 

  • Hu C, Zhou L, Zhang J, Huang C, Zhang G (2011) Immunohistochemical detection of Raf kinase inhibitor protein in normal cervical tissue and cervical cancer tissue. J Int Med Res 39:229–237

    CAS  Article  PubMed  Google Scholar 

  • Karamitopoulou E, Zlobec I, Panayiotides I, Patsouris ES, Peros G, Rallis G, Lapas C, Karakitsos P, Terracciano LM, Lugli A (2011) Systematic analysis of proteins from different signaling pathways in the tumor center and the invasive front of colorectal cancer. Hum Pathol 42:1888–1896

    CAS  Article  PubMed  Google Scholar 

  • Kim SO, Kim MR (2013) (-)-Epigallocatechin 3-gallate inhibits invasion by inducing the expression of Raf kinase inhibitor protein in AsPC-1 human pancreatic adenocarcinoma cells through the modulation of histone deacetylase activity. Int J Oncol 42:349–358

    PubMed  Google Scholar 

  • Kulemann B, Hoeppner J, Wittel U, Glatz T, Keck T, Wellner UF, Bronsert P, Sick O, Hopt UT, Makowiec F (2015) Perioperative and long-term outcome after standard pancreaticoduodenectomy, additional portal vein and multivisceral resection for pancreatic head cancer. J Gastrointest Surg 19:438–444

    Article  PubMed  Google Scholar 

  • Liu H, Li P, Li B, Sun P, Zhang J, Wang B, Jia B (2014) RKIP inhibits gastric cancer cell survival and invasion by regulating the expression of HMGA2 and OPN. Tumor Biol 35:11949–11958

    CAS  Article  Google Scholar 

  • Lorenz K, Lohse MJ, Quitterer U (2003) Protein kinase C switches the Raf kinase inhibitor from Raf-1 to GRK-2. Nature 426:574–579

    CAS  Article  PubMed  Google Scholar 

  • Ma S, Tang KH, Chan YP, Lee TK, Kwan PS, Castilho A, Ng I, Man K, Wong N, To KF, Zheng BJ, Lai PB, Lo CM, Chan KW, Guan XY (2010) miR-130b Promotes CD133(+) liver tumor-initiating cell growth and self-renewal via tumor protein 53-induced nuclear protein 1. Cell Stem Cell 7:694–707

    CAS  Article  PubMed  Google Scholar 

  • Matsuoka H, Tsubaki M, Yamazoe Y, Ogaki M, Satou T, Itoh T, Kusunoki T, Nishida S (2009) Tamoxifen inhibits tumor cell invasion and metastasis in mouse melanoma through suppression of PKC/MEK/ERK and PKC/PI3K/Akt pathways. Exp Cell Res 315:2022–2032

    CAS  Article  PubMed  Google Scholar 

  • Mavros MN, Xu L, Maqsood H, Gani F, Ejaz A, Spolverato G, Al-Refaie WB, Frank SM, Pawlik TM (2015) Perioperative blood transfusion and the prognosis of pancreatic cancer surgery: systematic review and meta-analysis. Ann Surg Oncol 22:4382–4391

    Article  PubMed  Google Scholar 

  • Melle C, Ernst G, Escher N, Hartmann D, Schimmel B, Bleul A, Thieme H, Kaufmann R, Felix K, Friess HM (2007) Protein profiling of microdissected pancreas carcinoma and identification of HSP27 as a potential serum marker. Clin Chem 53:629–635

    CAS  Article  PubMed  Google Scholar 

  • Michalski C, Kleeff J, Wente M, Diener M, Büchler M, Friess H (2007) Systematic review and meta-analysis of standard and extended lymphadenectomy in pancreaticoduodenectomy for pancreatic cancer. Br J Surg 94:265–273

    CAS  Article  PubMed  Google Scholar 

  • Minn AJ, Bevilacqua E, Yun J, Rosner MR (2012) Identification of novel metastasis suppressor signaling pathways for breast cancer. Cell Cycle 11:2452–2457

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Prasad R, Vaid M, Katiyar SK (2012) Grape proanthocyanidin inhibit pancreatic cancer cell growth in vitro and in vivo through induction of apoptosis and by targeting the PI3K/Akt pathway. PLoS One 7:e43064

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Ren G, Baritaki S, Marathe H, Feng J, Park S, Beach S, Bazeley PS, Beshir AB, Fenteany G, Mehra R (2012) Polycomb protein EZH2 regulates tumor invasion via the transcriptional repression of the metastasis suppressor RKIP in breast and prostate cancer. Cancer Res 72:3091–3104

    CAS  Article  PubMed  Google Scholar 

  • Sharpe SM, In H, Winchester DJ, Talamonti MS, Baker MS (2015) Surgical Resection Provides an Overall Survival Benefit for Patients with Small Pancreatic Neuroendocrine Tumors. J Gastrointest Surg 19:117–123

    Article  PubMed  Google Scholar 

  • Siegel RL, Miller KD, Jemal A (2015) Cancer statistics, 2015. CA: Cancer J Clin 65:5–29

    Google Scholar 

  • Stocken D, Büchler M, Dervenis C, Bassi C, Jeekel H, Klinkenbijl J, Bakkevold K, Takada T, Amano H, Neoptolemos J (2005) Meta-analysis of randomised adjuvant therapy trials for pancreatic cancer. Br J Cancer 92:1372–1381

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Torre LA, Bray F, Siegel RL, Ferlay J, Lortet-Tieulent J, Jemal A (2015) Global cancer statistics, 2012. CA: Cancer J Clin 65:87–108

    Google Scholar 

  • Vincent A, Herman J, Schulick R, Hruban RH, Goggins M (2011) Pancreatic cancer. Lancet 378:607–620

    Article  PubMed  PubMed Central  Google Scholar 

  • Wang W, Abbruzzese JL, Evans DB, Larry L, Cleary KR, Chiao PJ (1999) The nuclear factor-κB RelA transcription factor is constitutively activated in human pancreatic adenocarcinoma cells. Clin Cancer Res 5:119–127

    CAS  PubMed  Google Scholar 

  • Wolfgang CL, Herman JM, Laheru DA, Klein AP, Erdek MA, Fishman EK, Hruban RH (2013) Recent progress in pancreatic cancer. CA: Cancer J Clin 63:318–348

    Google Scholar 

  • Yeung K, Seitz T, Li S, Janosch P, McFerran B, Kaiser C, Fee F, Katsanakis KD, Rose DW, Mischak H, Sedivy JM, Kolch W (1999) Suppression of Raf-1 kinase activity and MAP kinase signalling by RKIP. Nature 401:173–177

    CAS  Article  PubMed  Google Scholar 

  • Zebisch A, Wölfler A, Fried I, Wolf O, Lind K, Bodner C, Haller M, Drasche A, Pirkebner D, Matallanas D (2012) Frequent loss of RAF kinase inhibitor protein expression in acute myeloid leukemia. Leukemia 26:1842–1849

    CAS  Article  PubMed  Google Scholar 

  • Zeng L, Imamoto A, Rosner M (2008) Raf kinase inhibitory protein (RKIP): a physiological regulator and future therapeutic target. Expert Opin Ther Targets 12:1275

    CAS  Article  PubMed  Google Scholar 

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Funding

This work was supported by grants from National Natural Science Foundation of China (No. 81472288).

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    Affiliations

    1. Department of Hepatobiliary Surgery, Southwest Hospital, Third Military Medical University, Chongqing, 404100, People’s Republic of China

      Haisu Dai, Haowei Chen, Wei Liu, Yu You, Jiaxin Tan, Aigang Yang, Xiangdong Lai & Ping Bie

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    1. Haisu Dai
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    2. Haowei Chen
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    3. Wei Liu
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    4. Yu You
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    5. Jiaxin Tan
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    7. Xiangdong Lai
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    8. Ping Bie
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    Correspondence to Ping Bie.

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    Human and animal rights

    All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Declaration of Helsinki and its later amendments or comparable ethical standards. All animal experiments were carried out strictly in accordance with international ethical guidelines and the National Institutes of Health Guide concerning the Care and Use of Laboratory Animals. The experiments were approved by Medical Ethics Committee of the Southwest Hospital and the First Affiliated Hospital of the Third Military Medical University.

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    Informed consent was obtained from all individual participants included in the study.

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    Haisu Dai and Haowei Chen contributed equally to this work.

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    Cite this article

    Dai, H., Chen, H., Liu, W. et al. Effects of Raf kinase inhibitor protein expression on pancreatic cancer cell growth and motility: an in vivo and in vitro study. J Cancer Res Clin Oncol 142, 2107–2117 (2016). https://doi.org/10.1007/s00432-016-2206-4

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    • DOI: https://doi.org/10.1007/s00432-016-2206-4

    Keywords

    • Raf kinase inhibitor protein
    • Pancreatic adenocarcinoma
    • Pancreatic cancer
    • Metastasis
    • Tumorigenesis