Advertisement

Tumor Biology

, Volume 37, Issue 6, pp 7277–7286 | Cite as

LB-100 sensitizes hepatocellular carcinoma cells to the effects of sorafenib during hypoxia by activation of Smad3 phosphorylation

  • Qi-Han Fu
  • Qi Zhang
  • Jing-Ying Zhang
  • Xu Sun
  • Yu Lou
  • Guo-Gang Li
  • Zhi-Liang Chen
  • Xue-Li Bai
  • Ting-Bo Liang
Original Article

Abstract

Hepatocellular carcinoma (HCC) is a common cancer with poor prognosis. The multikinase inhibitor sorafenib is the only clinically proved systematic treatment for HCC. However, few patients respond to sorafenib. Hypoxic microenvironments contribute to sorafenib resistance. LB-100, a serine/threonine protein phosphatase 2A (PP2A) inhibitor was previously found to be a chemosensitizer in HCC. Here, we tested whether LB-100 could sensitize HCC to the effects of sorafenib. Intriguingly, LB-100 enhanced the effects of sorafenib in HCC cells only during hypoxic environments. LB-100 dramatically increased intracellular p-Smad3 level, which was responsible for the effect of LB-100 as a sensitizer. LB-100 downregulated Bcl-2 expression and enhanced sorafenib-induced apoptosis in HCC cells. We further proved that PP2A mediated LB-100-induced p-Smad3 overexpression. In addition, p38 mitogen-activated protein kinase pathway was activated in hypoxic conditions, and enhanced p-Smad3-dependent Bcl-2 inhibition and consequent apoptosis. In conclusion, LB-100 sensitized HCC cells to sorafenib in hypoxic environments. This effect was mediated by inactivation of PP2A, resulting in enhanced level of p-Smad3. Increased p-Smad3 downregulated Bcl-2, causing increased apoptosis of HCC cells.

Keywords

p-Smad3 Apoptosis Drug resistance PP2A p38 MAPK 

Notes

Acknowledgments

We thank Lixte Biotechnology Holdings, Inc. (East Setauket, NY, USA) for the gift of LB-100. This work was financially supported by the National Natural Science Foundation of China (81401954), Science and Technology Program of Traditional Medicine of Zhejiang Province (2014ZZ007), and Medical Science and Technology Program of Zhejiang Province, China (2015KYA114 and 2013KYB264). We appreciate Mr. Wang Yi and Mr. Qin Hao (The Second Affiliated Hospital, Zhejiang University School of Medicine, China) for their help in certain experiments.

Author contributions

Liang TB, Bai XL, and Zhang Q conceived the idea. Fu QH, Zhang Q, Zhang JY, Sun X, Lou Y, Li GG, and Chen ZL performed the experiments. Fu QH and Zhang Q analyzed the data. Fu QH and Zhang Q wrote the manuscript. All authors approved the manuscript.

Compliance with ethical standards

Conflicts of interest

None

Statement on the welfare of animals

  • All applicable international, national, and institutional guidelines for the care and use of animals have been followed.

  • All procedures performed in studies involving animals were in accordance with the ethical standards of the second affiliated hospital, Zhejiang University School of Medicine.

References

  1. 1.
    Torre LA, Bray F, Siegel RL, et al. Global cancer statistics, 2012. CA Cancer J Clin. 2015;65:87–108.CrossRefPubMedGoogle Scholar
  2. 2.
    Llovet JM, Ricci S, Mazzaferro V, et al. Sorafenib in advanced hepatocellular carcinoma. N Engl J Med. 2008;359:378–90.CrossRefPubMedGoogle Scholar
  3. 3.
    Cheng AL, Kang YK, Chen Z, et al. Efficacy and safety of Sorafenib in patients in the Asia-Pacific region with advanced hepatocellular carcinoma: a phase III randomised, double-blind, placebo-controlled trial. Lancet Oncol. 2009;10:25–34.CrossRefPubMedGoogle Scholar
  4. 4.
    Liang Y, Zheng T, Song R, et al. Hypoxia-mediated sorafenib resistance can be overcome by EF24 through Von Hippel-Lindau tumor suppressor-dependent HIF-1alpha inhibition in hepatocellular carcinoma. Hepatology. 2013;57:1847–57.CrossRefPubMedGoogle Scholar
  5. 5.
    Xu H, Zhao L, Fang Q, et al. MiR-338-3p inhibits hepatocarcinoma cells and sensitizes these cells to sorafenib by targeting hypoxia-induced factor 1alpha. PLoS One. 2014;9:e115565.CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Zhang Q, Bai XL, Chen W, et al. Wnt/beta-catenin signaling enhances hypoxia-induced epithelial-mesenchymal transition in hepatocellular carcinoma via crosstalk with hif-1alpha signaling. Carcinogenesis. 2013;34:962–73.CrossRefPubMedGoogle Scholar
  7. 7.
    Yang YA, Zhang GM, Feigenbaum L, et al. Smad3 reduces susceptibility to hepatocarcinoma by sensitizing hepatocytes to apoptosis through downregulation of Bcl-2. Cancer Cell. 2006;9:445–57.CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Ali A, Zhang P, Liangfang Y, et al. KLF17 empowers TGF-beta/Smad signaling by targeting Smad3-dependent pathway to suppress tumor growth and metastasis during cancer progression. Cell Death Dis. 2015;6:e1681.CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Chen CL, Tsukamoto H, Liu JC, Kashiwabara C, Feldman D, Sher L, et al. Reciprocal regulation by TLR4 and TGF-beta in tumor-initiating stem-like cells. J Clin Invest. 2013;123:2832–49.CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Heikkinen PT, Nummela M, Leivonen SK, et al. Hypoxia-activated Smad3-specific dephosphorylation by PP2A. J Biol Chem. 2010;285:3740–9.CrossRefPubMedGoogle Scholar
  11. 11.
    Chen YL, Lv J, Ye XL, et al. Sorafenib inhibits transforming growth factor beta1-mediated epithelial-mesenchymal transition and apoptosis in mouse hepatocytes. Hepatology. 2011;53:1708–18.CrossRefPubMedGoogle Scholar
  12. 12.
    Bai XL, Zhang Q, Ye LY, et al. Inhibition of protein phosphatase 2A enhances cytotoxicity and accessibility of chemotherapeutic drugs to hepatocellular carcinomas. Mol Cancer Ther. 2014;13:2062–72.CrossRefPubMedGoogle Scholar
  13. 13.
    Bai XL, Zhi X, Zhang Q, et al. Inhibition of protein phosphatase 2A sensitizes pancreatic cancer to chemotherapy by increasing drug perfusion via HIF-1alpha-VEGF mediated angiogenesis. Cancer Lett. 2014;355:281–7.CrossRefPubMedGoogle Scholar
  14. 14.
    Kim BC, van Gelder H, Kim TA, et al. Activin receptor-like kinase-7 induces apoptosis through activation of MAPKs in a Smad3-dependent mechanism in hepatoma cells. J Biol Chem. 2004;279:28458–65.CrossRefPubMedGoogle Scholar
  15. 15.
    Feng X, Xu R, Du X, et al. Combination therapy with Sorafenib and radiofrequency ablation for BCLC stage 0-B1 hepatocellular carcinoma: a multicenter retrospective cohort study. Am J Gastroenterol. 2014;109:1891–9.CrossRefPubMedGoogle Scholar
  16. 16.
    Cainap C, Qin S, Huang WT, et al. Linifanib versus sorafenib in patients with advanced hepatocellular carcinoma: results of a randomized phase III trial. J Clin Oncol. 2015;33:172–9.CrossRefPubMedGoogle Scholar
  17. 17.
    Zhu AX, Rosmorduc O, Evans TR, et al. SEARCH: a phase III, randomized, double-blind, placebo-controlled trial of Sorafenib plus Erlotinib in patients with advanced hepatocellular carcinoma. J Clin Oncol. 2015;33:559–66.CrossRefPubMedGoogle Scholar
  18. 18.
    Fu QH, Zhang Q, Bai XL, et al. Sorafenib enhances effects of transarterial chemoembolization for hepatocellular carcinoma: a systematic review and meta-analysis. J Cancer Res Clin Oncol. 2014;140:1429–40.CrossRefPubMedGoogle Scholar
  19. 19.
    Jia L, Ma X, Gui B, et al. Sorafenib ameliorates renal fibrosis through inhibition of TGF-beta-induced epithelial-mesenchymal transition. PLoS One. 2015;10:e0117757.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Zhou T, Lv X, Guo X, et al. RACK1 modulates apoptosis induced by sorafenib in HCC cells by interfering with the IRE1/XBP1 axis. Oncol Rep. 2015;33:3006–14.PubMedGoogle Scholar
  21. 21.
    Serova M, Tijeras-Raballand A, Dos Santos C, et al. Effects of TGF-beta signalling inhibition with galunisertib (LY2157299) in hepatocellular carcinoma models and in ex vivo whole tumor tissue samples from patients. Oncotarget. 2015;6:21614–27.CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Lin YT, Lu HP, Chao CC. Oncogenic c-Myc and prothymosin-alpha protect hepatocellular carcinoma cells against sorafenib-induced apoptosis. Biochem Pharmacol. 2015;93:110–24.CrossRefPubMedGoogle Scholar
  23. 23.
    Jiao M, Nan KJ. Activation of PI3 kinase/Akt/HIF-1alpha pathway contributes to hypoxia-induced epithelial-mesenchymal transition and chemoresistance in hepatocellular carcinoma. Int J Oncol. 2012;40:461–8.PubMedGoogle Scholar
  24. 24.
    Negri FV, Dal Bello B, Porta C, et al. Expression of pERK and VEGFR-2 in advanced hepatocellular carcinoma and resistance to sorafenib treatment. Liver Int. 2015;35:2001–8.CrossRefPubMedGoogle Scholar
  25. 25.
    Liao JH, Chen JS, Chai MQ, et al. The involvement of p38 MAPK in transforming growth factor beta1-induced apoptosis in murine hepatocytes. Cell Res. 2001;11:89–94.CrossRefPubMedGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2015

Authors and Affiliations

  • Qi-Han Fu
    • 1
  • Qi Zhang
    • 1
    • 2
  • Jing-Ying Zhang
    • 1
  • Xu Sun
    • 3
  • Yu Lou
    • 1
  • Guo-Gang Li
    • 1
  • Zhi-Liang Chen
    • 4
  • Xue-Li Bai
    • 1
    • 2
  • Ting-Bo Liang
    • 1
    • 5
  1. 1.Department of Hepatobiliary and Pancreatic Surgery, the Second Affiliated Hospital, School of MedicineZhejiang UniversityHangzhouChina
  2. 2.Key Laboratory of Cancer Prevention and Intervention, the Second Affiliated Hospital, School of MedicineZhejiang UniversityHangzhouChina
  3. 3.Department of General SurgeryHuzhou Central HospitalHuzhouChina
  4. 4.Department of Hepatobiliary and Pancreatic SurgeryShaoxing People’s HospitalShaoxingChina
  5. 5.Collaborative Innovation Center for Cancer MedicineZhejiang UniversityGuangzhouChina

Personalised recommendations