Skip to main content

Advertisement

SpringerLink
Log in

Hypoxia-Induced PLOD2 is a Key Regulator in Epithelial-Mesenchymal Transition and Chemoresistance in Biliary Tract Cancer

  • Translational Research and Biomarkers
  • Published:
Annals of Surgical Oncology Aims and scope Submit manuscript

Abstract

Background

The prognosis of biliary tract cancer (BTC) is unfavorable due to its chemoresistance. Hypoxia triggers epithelial-to-mesenchymal transition (EMT), which is closely related to drug resistance. In this study, we focused on the functional roles of procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2), a hypoxia-induced gene, in BTC, and assessed the clinical significance of PLOD2.

Methods

The expression of PLOD2 under hypoxia was assessed in BTC cell lines. Gemcitabine-resistant (GR) BTC cell lines were transfected with small interfering RNA (siRNA) against PLOD2, and EMT markers and chemoresistance were evaluated. PLOD2 expression was also characterized using immunohistochemistry in BTC clinical specimens following resection. Patient survival was analyzed and the role of PLOD2 expression was examined.

Results

The expression of PLOD2 was induced by hypoxia in vitro and was upregulated in BTC-GR cell lines, which had low expression of epithelial markers and high expression of mesenchymal markers. Downregulation of PLOD2 by siRNA resulted in improved chemoresistance, recovery of epithelial markers, and reduction of mesenchymal markers. In the resected BTC samples, PLOD2 expression was significantly correlated with lymph node metastasis (p = 0.037) and stage (p = 0.001). Recurrence-free survival (p = 0.011) and overall survival (p < 0.001) rates were significantly lower in patients with high expression of PLOD2. PLOD2 expression was an independent prognostic factor for overall survival (p = 0.019).

Conclusions

The expression of PLOD2 influenced chemoresistance through EMT, and high expression of PLOD2 was a significant unfavorable prognostic factor in BTC patients. PLOD2 might be a potential therapeutic target for overcoming chemoresistance.

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

Similar content being viewed by others

References

  1. Zhu AX, Hezel AF. Development of molecularly targeted therapies in biliary tract cancers: reassessing the challenges and opportunities. Hepatology. 2011;53(2):695–704.

    Article  CAS  PubMed  Google Scholar 

  2. Kim HJ, Kim CY, Hur YH, et al. Prognostic factors for survival after curative resection of distal cholangiocarcinoma: perineural invasion and lymphovascular invasion. Surg Today. 2014;44(10):1879–86.

    Article  PubMed  Google Scholar 

  3. Murakami S, Ajiki T, Okazaki T, et al. Factors affecting survival after resection of intrahepatic cholangiocarcinoma. Surg Today. 2014;44(10):1847–54.

    Article  PubMed  Google Scholar 

  4. Kobayashi S, Nagano H, Marubashi S, et al. Treatment of borderline cases for curative resection of biliary tract cancer. J Surg Oncol. 2011;104(5):499–503.

    Article  PubMed  Google Scholar 

  5. Colvin H, Mizushima T, Eguchi H, Takiguchi S, Doki Y, Mori M. Gastroenterological surgery in Japan: The past, the present and the future. Ann Gastroenterol Surg. 2017;1(1):5–10.

    Article  PubMed  PubMed Central  Google Scholar 

  6. Mizuno T, Ebata T, Yokoyama Y, et al. Adjuvant gemcitabine monotherapy for resectable perihilar cholangiocarcinoma with lymph node involvement: a propensity score matching analysis. Surg Today. 2017;47(2):182–92.

    Article  CAS  PubMed  Google Scholar 

  7. Noda T, Shimoda M, Ortiz V, Sirica AE, Wands JR. Immunization with aspartate-beta-hydroxylase-loaded dendritic cells produces antitumor effects in a rat model of intrahepatic cholangiocarcinoma. Hepatology. 2012;55(1):86–97.

    Article  CAS  PubMed  Google Scholar 

  8. Valle JW, Wasan H, Johnson P, et al. Gemcitabine alone or in combination with cisplatin in patients with advanced or metastatic cholangiocarcinomas or other biliary tract tumours: a multicentre randomised phase II study—the UK ABC-01 Study. Br J Cancer. 2009;101(4):621–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Okusaka T, Nakachi K, Fukutomi A, et al. Gemcitabine alone or in combination with cisplatin in patients with biliary tract cancer: a comparative multicentre study in Japan. Br J Cancer. 2010;103(4):469–474.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Valle JW, Furuse J, Jitlal M, et al. Cisplatin and gemcitabine for advanced biliary tract cancer: a meta-analysis of two randomised trials. Ann Oncol. 2014;25(2):391–8.

    Article  CAS  PubMed  Google Scholar 

  11. Ji RC. Hypoxia and lymphangiogenesis in tumor microenvironment and metastasis. Cancer Lett. 2014;346(1):6–16.

    Article  CAS  PubMed  Google Scholar 

  12. Liang D, Ma Y, Liu J, et al. The hypoxic microenvironment upgrades stem-like properties of ovarian cancer cells. BMC Cancer. 2012;12:201.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Zhang L, Huang G, Li X, et al. Hypoxia induces epithelial-mesenchymal transition via activation of SNAI1 by hypoxia-inducible factor-1α in hepatocellular carcinoma. BMC Cancer. 2013;13:108.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Unwith S, Zhao H, Hennah L, Ma D. The potential role of HIF on tumour progression and dissemination. Int J Cancer. 2015;136(11):2491–503.

    Article  CAS  PubMed  Google Scholar 

  15. Vanichapol T, Leelawat K, Hongeng S. Hypoxia enhances cholangiocarcinoma invasion through activation of hepatocyte growth factor receptor and the extracellular signalregulated kinase signaling pathway. Mol Med Rep. 2015;12(3):3265–72.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Zhan M, Wang H, Chen T, et al. NOX1 mediates chemoresistance via HIF1alpha/MDR1 pathway in gallbladder cancer. Biochem Biophys Res Commun. 2015;468(1–2):79–85.

    Article  CAS  PubMed  Google Scholar 

  17. Ruotsalainen H, Vanhatupa S, Tampio M, Sipila L, Valtavaara M, Myllyla R. Complete genomic structure of mouse lysyl hydroxylase 2 and lysyl hydroxylase 3/collagen glucosyltransferase. Matrix Biol. 2001;20(2):137–46.

    Article  CAS  PubMed  Google Scholar 

  18. Gilkes DM, Bajpai S, Wong CC, et al. Procollagen lysyl hydroxylase 2 is essential for hypoxia-induced breast cancer metastasis. Mol Cancer Res. 2013;11(5):456–66.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Eisinger-Mathason TS, Zhang M, Qiu Q, et al. Hypoxia-dependent modification of collagen networks promotes sarcoma metastasis. Cancer Discov. 2013;3(10):1190–205.

    Article  CAS  PubMed  Google Scholar 

  20. van der Slot AJ, Zuurmond AM, Bardoel AF, et al. Identification of PLOD2 as telopeptide lysyl hydroxylase, an important enzyme in fibrosis. J Biol Chem. 2003;278(42):40967–72.

    Article  CAS  PubMed  Google Scholar 

  21. Wu J, Reinhardt DP, Batmunkh C, et al. Functional diversity of lysyl hydroxylase 2 in collagen synthesis of human dermal fibroblasts. Exp Cell Res. 2006;312(18):3485–94.

    Article  CAS  PubMed  Google Scholar 

  22. Bergman AM, Pinedo HM, Peters GJ. Determinants of resistance to 2′,2′-difluorodeoxycytidine (gemcitabine). Drug Resist Updat. 2002;5(1):19–33.

    Article  CAS  PubMed  Google Scholar 

  23. Iwagami Y, Eguchi H, Nagano H, et al. miR-320c regulates gemcitabine-resistance in pancreatic cancer via SMARCC1. Br J Cancer. 2013;109(2):502–11.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Yamada D, Kobayashi S, Wada H, et al. Role of crosstalk between interleukin-6 and transforming growth factor-beta 1 in epithelial-mesenchymal transition and chemoresistance in biliary tract cancer. Eur J Cancer. 2013;49(7):1725–40.

    Article  CAS  PubMed  Google Scholar 

  25. Hasegawa S, Eguchi H, Nagano H, et al. MicroRNA-1246 expression associated with CCNG2-mediated chemoresistance and stemness in pancreatic cancer. Br J Cancer. 2014;111(8):1572–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Mikamori M, Yamada D, Eguchi H, et al. MicroRNA-155 Controls Exosome Synthesis and Promotes Gemcitabine Resistance in Pancreatic Ductal Adenocarcinoma. Sci Rep. 2017;7:42339.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Harris AL. Hypoxia: a key regulatory factor in tumour growth. Nat Rev Cancer. 2002;2(1):38–47.

    Article  CAS  PubMed  Google Scholar 

  28. Semenza GL. Targeting HIF-1 for cancer therapy. Nat Rev Cancer. 2003;3(10):721–32.

    Article  CAS  PubMed  Google Scholar 

  29. Erkan M, Kleeff J, Esposito I, et al. Loss of BNIP3 expression is a late event in pancreatic cancer contributing to chemoresistance and worsened prognosis. Oncogene. 2005;24(27):4421–32.

    Article  CAS  PubMed  Google Scholar 

  30. Song X, Liu X, Chi W, et al. Hypoxia-induced resistance to cisplatin and doxorubicin in non-small cell lung cancer is inhibited by silencing of HIF-1alpha gene. Cancer Chemother Pharmacol. 2006;58(6):776–84.

    Article  CAS  PubMed  Google Scholar 

  31. Samanta D, Gilkes DM, Chaturvedi P, Xiang L, Semenza GL. Hypoxia-inducible factors are required for chemotherapy resistance of breast cancer stem cells. Proc Natl Acad Sci U S A. 2014;111(50):E5429–38.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Shukla SK, Purohit V, Mehla K, et al. MUC1 and HIF-1alpha signaling crosstalk induces anabolic glucose metabolism to impart gemcitabine resistance to pancreatic cancer. Cancer Cell. 2017;32(1):71–87.e7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Gilkes DM, Bajpai S, Chaturvedi P, Wirtz D, Semenza GL. Hypoxia-inducible factor 1 (HIF-1) promotes extracellular matrix remodeling under hypoxic conditions by inducing P4HA1, P4HA2, and PLOD2 expression in fibroblasts. J Biol Chem. 2013;288(15):10819-10829.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Chen Y, Terajima M, Yang Y, et al. Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma. J Clin Invest. 2015;125(3):1147–62.

    Article  PubMed  PubMed Central  Google Scholar 

  35. Miyamoto K, Seki N, Matsushita R, et al. Tumour-suppressive miRNA-26a-5p and miR-26b-5p inhibit cell aggressiveness by regulating PLOD2 in bladder cancer. Br J Cancer. 2016;115(3):354–63.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Arao T, Yanagihara K, Takigahira M, et al. ZD6474 inhibits tumor growth and intraperitoneal dissemination in a highly metastatic orthotopic gastric cancer model. Int J Cancer. 2006;118(2):483–9.

    Article  CAS  PubMed  Google Scholar 

  37. Dong S, Nutt CL, Betensky RA, et al. Histology-based expression profiling yields novel prognostic markers in human glioblastoma. J Neuropathol Exp Neurol. 2005;64(11):948–55.

    Article  PubMed  Google Scholar 

  38. Noda T, Yamamoto H, Takemasa I, et al. PLOD2 induced under hypoxia is a novel prognostic factor for hepatocellular carcinoma after curative resection. Liver Int. 2012;32(1):110–8.

    Article  CAS  PubMed  Google Scholar 

  39. Song Y, Zheng S, Wang J, et al. Hypoxia-induced PLOD2 promotes proliferation, migration and invasion via PI3K/Akt signaling in glioma. Oncotarget. 2017;8(26):41947–62.

    PubMed  PubMed Central  Google Scholar 

  40. Xu F, Zhang J, Hu G, Liu L, Liang W. Hypoxia and TGF-beta1 induced PLOD2 expression improve the migration and invasion of cervical cancer cells by promoting epithelial-to-mesenchymal transition (EMT) and focal adhesion formation. Cancer Cell Int. 2017;17:54.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Xu Y, Zhang L, Wei Y, et al. Procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 promotes hypoxia-induced glioma migration and invasion. Oncotarget. 2017;8(14):23401–13.

    PubMed  PubMed Central  Google Scholar 

  42. Thiery JP. Epithelial-mesenchymal transitions in tumour progression. Nat Rev Cancer. 2002;2(6):442–54.

    Article  CAS  PubMed  Google Scholar 

  43. Shah AN, Summy JM, Zhang J, Park SI, Parikh NU, Gallick GE. Development and characterization of gemcitabine-resistant pancreatic tumor cells. Ann Surg Oncol. 2007;14(12):3629–37.

    Article  PubMed  Google Scholar 

  44. Cannito S, Novo E, Compagnone A, et al. Redox mechanisms switch on hypoxia-dependent epithelial-mesenchymal transition in cancer cells. Carcinogenesis. 2008;29(12):2267–78.

    Article  CAS  PubMed  Google Scholar 

  45. Arumugam T, Ramachandran V, Fournier KF, et al. Epithelial to mesenchymal transition contributes to drug resistance in pancreatic cancer. Cancer Res. 2009;69(14):5820–8.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Sakamoto T, Kobayashi S, Yamada D, et al. A histone deacetylase inhibitor suppresses epithelial-mesenchymal transition and attenuates chemoresistance in biliary tract cancer. PLoS One. 2016;11(1):e0145985.

    Article  PubMed  PubMed Central  Google Scholar 

  47. Thiery JP, Sleeman JP. Complex networks orchestrate epithelial-mesenchymal transitions. Nat Rev Mol Cell Biol. 2006;7(2):131–42.

    Article  CAS  PubMed  Google Scholar 

  48. Hotz B, Arndt M, Dullat S, Bhargava S, Buhr HJ, Hotz HG. Epithelial to mesenchymal transition: expression of the regulators snail, slug, and twist in pancreatic cancer. Clin Cancer Res. 2007;13(16):4769–76.

    Article  CAS  PubMed  Google Scholar 

  49. Peinado H, Olmeda D, Cano A. Snail, Zeb and bHLH factors in tumour progression: an alliance against the epithelial phenotype? Nat Rev Cancer. 2007;7(6):415–28.

    Article  CAS  PubMed  Google Scholar 

  50. Lluis JM, Buricchi F, Chiarugi P, Morales A, Fernandez-Checa JC. Dual role of mitochondrial reactive oxygen species in hypoxia signaling: activation of nuclear factor-{kappa}B via c-SRC and oxidant-dependent cell death. Cancer Res. 2007;67(15):7368–77.

    Article  CAS  PubMed  Google Scholar 

  51. Fischer KR, Durrans A, Lee S, et al. Epithelial-to-mesenchymal transition is not required for lung metastasis but contributes to chemoresistance. Nature. 2015;527(7579):472–6.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgment

This work was supported in part by grants from Grant-in-Aid for Scientific Research of the Japan Society for the Promotion of Science (JSPS) [(C) 16K10572], Grant-in-Aid for Young Scientists of the JSPS [(B) 17K16543], and the Suzuken Memorial Foundation. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The authors thank the members of their laboratory for technical advice and helpful discussions.

Author information

Authors and Affiliations

  1. Department of Gastroenterological Surgery, Graduate School of Medicine, Osaka University, Osaka, Japan

    Yuichiro Okumura MD, Takehiro Noda MD, PhD, Hidetoshi Eguchi MD, PhD, Takuya Sakamoto MD, PhD, Yoshifumi Iwagami MD, PhD, Daisaku Yamada MD, PhD, Tadafumi Asaoka MD, PhD, Hiroshi Wada MD, PhD, Koichi Kawamoto MD, PhD, Kunihito Gotoh MD, PhD, Shogo Kobayashi MD, PhD, Yutaka Takeda MD, PhD, Masahiro Tanemura MD, PhD, Yuichiro Doki MD, PhD & Masaki Mori MD, PhD, FACS

  2. Department of Surgery, Kansai Rosai Hospital, Hyogo, Japan

    Yutaka Takeda MD, PhD

  3. Department of Surgery, Osaka Police Hospital, Osaka, Japan

    Masahiro Tanemura MD, PhD

  4. Division of Health Science, Graduate School of Medicine, Osaka University, Osaka, Japan

    Koji Umeshita MD, PhD

Authors
  1. Yuichiro Okumura MD
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Takehiro Noda MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hidetoshi Eguchi MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Takuya Sakamoto MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yoshifumi Iwagami MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Daisaku Yamada MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Tadafumi Asaoka MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Hiroshi Wada MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Koichi Kawamoto MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Kunihito Gotoh MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Shogo Kobayashi MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Yutaka Takeda MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Masahiro Tanemura MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. Koji Umeshita MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Yuichiro Doki MD, PhD
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Masaki Mori MD, PhD, FACS
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Hidetoshi Eguchi MD, PhD.

Ethics declarations

Conflicts of interest

The authors declare no conflicts of interest associated with this manuscript.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 50 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Okumura, Y., Noda, T., Eguchi, H. et al. Hypoxia-Induced PLOD2 is a Key Regulator in Epithelial-Mesenchymal Transition and Chemoresistance in Biliary Tract Cancer. Ann Surg Oncol 25, 3728–3737 (2018). https://doi.org/10.1245/s10434-018-6670-8

Download citation

  • Received:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1245/s10434-018-6670-8

Keywords

Search

Navigation