Resibufogenin suppresses tumor growth and inhibits glycolysis in ovarian cancer by modulating PIM1

  • Qian Li
  • Chuanwu Jiang
  • Yan Wang
  • Minghua Wei
  • Huijin Zheng
  • Yanqi Xu
  • Xuegang Xu
  • Fengyu Jia
  • Kai Liu
  • Gang Sun
  • Jianhua ZangEmail author
  • Ping MoEmail author
Original Article


Ovarian cancer is a common human malignancy of the female reproductive system. However, chemotherapy has been proven to have limited effectiveness in a majority of patients. Resibufogenin (RB) is a major active ingredient in cinobufacini, which has been used in the treatment of human malignancies as adjunct agents. This study was designed to examine the anti-cancer effect of RB and the underlying mechanisms in ovarian cancer. Our results showed that RB treatment resulted in cell death, cell cycle arrest, and apoptosis in ovarian cancer cells. The anti-growth and pro-apoptotic effects of RB were also validated in xenograft mice models. Proteomics analysis indicated that RB was able to alter the expressions of several genes, which were involved in the regulation of glycolysis. The suppression effect of RB in the glycolysis pathway of ovarian cancer cells was validated by decreased glucose consumption, lactate production, and extracellular acidification rate (ECAR). We proposed that PIM1 functioned as the key target that mediated the anti-cancer effect of RB against ovarian cancer cells. Our results have revealed that RB downregulated PIM1 in ovarian cancer cells and its downstream genes involved in glycolysis. Moreover, our results indicated that the anti-growth activities and suppressing effect of RB on glycolysis were enhanced significantly by PIM1 knockdown but was attenuated by ectopic PIM1 expression. This provided evidence to support the role of PIM1 in the anti-cancer activities of RB.


Resibufogenin Ovarian cancer PIM1 Apoptosis Glycolysis 


Author contributions

Ping Mo and Jianhua Zang designed the research. Qian Li, Chuanwu Jiang, Yan Wang, Minghua Wei, Huijin Zheng, Yanqi Xu, and Xuegang Xu conducted the experiments. Fengyu Jia, Kai Liu, and Gang Sun analyzed the data. Qian Li and Chuanwu Jiang wrote the paper under the guidance of Ping Mo and Jianhua Zang. All authors read and approved the manuscript.

Compliance with ethical standards

The Institutional Animal Care and Use Committee at Qingdao Hospital of Traditional Chinese Medicine approved the protocol for animal experiments.

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Ahmad S et al (2013) First cancer statistics report from Hazara division. J Ayub Med Coll Abbottabad 25(1–2):71–73Google Scholar
  2. Cao L et al (2018) PIM1 kinase promotes cell proliferation, metastasis and tumor growth of lung adenocarcinoma by potentiating the c-MET signaling pathway. Cancer Lett 444:116–126Google Scholar
  3. Chatterjee, S., et al., Targeting PIM kinase with PD1 inhibition improves immunotherapeutic antitumor T-cell response. Clin Cancer Res, 2018Google Scholar
  4. Deng D, Wang L, Chen Y, Li B, Xue L, Shao N, Wang Q, Xia X, Yang Y, Zhi F (2016) MicroRNA-124-3p regulates cell proliferation, invasion, apoptosis, and bioenergetics by targeting PIM1 in astrocytoma. Cancer Sci 107(7):899–907Google Scholar
  5. Dhanasekaran SM, Barrette TR, Ghosh D, Shah R, Varambally S, Kurachi K, Pienta KJ, Rubin MA, Chinnaiyan AM (2001) Delineation of prognostic biomarkers in prostate cancer. Nature 412(6849):822–826Google Scholar
  6. Dong J et al (2016) Treatment of huge hepatocellular carcinoma using cinobufacini injection in transarterial chemoembolization: a retrospective study. Evid Based Complement Alternat Med 2016:2754542Google Scholar
  7. Gojda J, Waldauf P, Hrušková N, Blahutová B, Krajčová A, Urban T, Tůma P, Řasová K, Duška F (2019) Lactate production without hypoxia in skeletal muscle during electrical cycling: crossover study of femoral venous-arterial differences in healthy volunteers. PLoS One 14(3):e0200228Google Scholar
  8. Han Q, Ma Y, Wang H, Dai Y, Chen C, Liu Y, Jing L, Sun X (2018) Resibufogenin suppresses colorectal cancer growth and metastasis through RIP3-mediated necroptosis. J Transl Med 16(1):201Google Scholar
  9. Ichikawa M, Sowa Y, Iizumi Y, Aono Y, Sakai T (2015) Resibufogenin induces G1-phase arrest through the proteasomal degradation of cyclin D1 in human malignant tumor cells. PLoS One 10(6):e0129851Google Scholar
  10. Jiang Y, Zhang Y, Luan J, Duan H, Zhang F, Yagasaki K, Zhang G (2010) Effects of bufalin on the proliferation of human lung cancer cells and its molecular mechanisms of action. Cytotechnology 62(6):573–583Google Scholar
  11. Leung CO et al (2015) PIM1 regulates glycolysis and promotes tumor progression in hepatocellular carcinoma. Oncotarget 6(13):10880–10892Google Scholar
  12. Li X, Liu Y, Shen A, Wang C, Yan J, Zhao W, Liang X (2014) Efficient purification of active bufadienolides by a class separation method based on hydrophilic solid-phase extraction and reversed-phase high performance liquid chromatography. J Pharm Biomed Anal 97:54–64Google Scholar
  13. Liberti MV, Locasale JW (2016) The Warburg effect: how does it benefit cancer cells? Trends Biochem Sci 41(3):211–218Google Scholar
  14. Lin F, Luo X, Tsun A, Li Z, Li D, Li B (2015) Kaempferol enhances the suppressive function of Treg cells by inhibiting FOXP3 phosphorylation. Int Immunopharmacol 28(2):859–865Google Scholar
  15. Liu C et al (2014) Comparison of toad skins Bufo bufo gargarizans Cantor from different regions for their active constituents content and cytotoxic activity on lung carcinoma cell lines. Pharmacogn Mag 10(39):207–212Google Scholar
  16. Liu K, Gao H, Wang Q, Wang L, Zhang B, Han Z, Chen X, Han M, Gao M (2018a) Hispidulin suppresses cell growth and metastasis by targeting PIM1 through JAK2/STAT3 signaling in colorectal cancer. Cancer Sci 109(5):1369–1381Google Scholar
  17. Liu L, Liu Y, Liu X, Zhang N, Mao G, Zeng Q, Yin M, Song D, Deng H (2018b) Resibufogenin suppresses transforming growth factor-beta-activated kinase 1-mediated nuclear factor-kappaB activity through protein kinase C-dependent inhibition of glycogen synthase kinase 3. Cancer Sci 109(11):3611–3622Google Scholar
  18. Ma XC, Zhang BJ, Xin XL, Huang SS, Deng S, Zhang HL, Shu XH, Diao YP, Cui J (2009) Simultaneous quantification of seven major bufadienolides in three traditional Chinese medicinal preparations of chansu by HPLC-DAD. Nat Prod Commun 4(2):179–184Google Scholar
  19. Magnuson NS et al (2010) Why target PIM1 for cancer diagnosis and treatment. Future Oncol 6(9):1461–1478Google Scholar
  20. Narlik-Grassow M, Blanco-Aparicio C, Carnero A (2014) The PIM family of serine/threonine kinases in cancer. Med Res Rev 34(1):136–159Google Scholar
  21. Nawijn MC, Alendar A, Berns A (2011) For better or for worse: the role of Pim oncogenes in tumorigenesis. Nat Rev Cancer 11(1):23–34Google Scholar
  22. Ozols RF (2005) Treatment goals in ovarian cancer. Int J Gynecol Cancer 15(Suppl 1):3–11Google Scholar
  23. Qi F, Li A, Inagaki Y, Kokudo N, Tamura S, Nakata M, Tang W (2011) Antitumor activity of extracts and compounds from the skin of the toad Bufo bufo gargarizans Cantor. Int Immunopharmacol 11(3):342–349Google Scholar
  24. Qi F, Li A, Inagaki Y, Xu H, Wang D, Cui X, Zhang L, Kokudo N, du G, Tang W (2012) Induction of apoptosis by cinobufacini preparation through mitochondria- and Fas-mediated caspase-dependent pathways in human hepatocellular carcinoma cells. Food Chem Toxicol 50(2):295–302Google Scholar
  25. Qian KC, Wang L, Hickey ER, Studts J, Barringer K, Peng C, Kronkaitis A, Li J, White A, Mische S, Farmer B (2005) Structural basis of constitutive activity and a unique nucleotide binding mode of human Pim-1 kinase. J Biol Chem 280(7):6130–6137Google Scholar
  26. Qin TJ, Zhao XH, Yun J, Zhang LX, Ruan ZP, Pan BR (2008) Efficacy and safety of gemcitabine-oxaliplatin combined with huachansu in patients with advanced gallbladder carcinoma. World J Gastroenterol 14(33):5210–5216Google Scholar
  27. Reiser-Erkan C, Erkan M, Pan Z, Bekasi S, Giese NA, Streit S, Michalski CW, Friess H, Kleeff J (2008) Hypoxia-inducible proto-oncogene Pim-1 is a prognostic marker in pancreatic ductal adenocarcinoma. Cancer Biol Ther 7(9):1352–1359Google Scholar
  28. Shen Z et al (2018) miR494BAG1 axis is involved in cinobufaciniinduced cell proliferation and apoptosis in gastric cancer. Mol Med Rep 17(5):7435–7441Google Scholar
  29. Siegel RL, Miller KD, Jemal A (2017) Cancer statistics, 2017. CA Cancer J Clin 67(1):7–30Google Scholar
  30. Siu MK et al (2010) p21-activated kinase 4 regulates ovarian cancer cell proliferation, migration, and invasion and contributes to poor prognosis in patients. Proc Natl Acad Sci U S A 107(43):18622–18627Google Scholar
  31. Wang DL et al (2010) Apoptosis-inducing activity of compounds screened and characterized from cinobufacini by bioassay-guided isolation. Mol Med Rep 3(4):717–722Google Scholar
  32. Wang JY et al (2012) Cinobufocini inhibits NF-kappaB and COX-2 activation induced by TNF-alpha in lung adenocarcinoma cells. Oncol Rep 27(5):1619–1624Google Scholar
  33. Wang Y, Ren F, Chen P, Liu S, Song Z, Ma X (2019) Effects of CytoReductive surgery plus hyperthermic IntraPEritoneal chemotherapy (HIPEC) versus CytoReductive surgery for ovarian cancer patients: a systematic review and meta-analysis. Eur J Surg Oncol 45(3):301–309Google Scholar
  34. Warnecke-Eberz U et al (2008) Frequent down-regulation of pim-1 mRNA expression in non-small cell lung cancer is associated with lymph node metastases. Oncol Rep 20(3):619–624Google Scholar
  35. Wei X, Si N, Zhang Y, Zhao H, Yang J, Wang H, Wang L, Han L, Bian B (2017) Evaluation of bufadienolides as the main antitumor components in cinobufacin injection for liver and gastric Cancer therapy. PLoS One 12(1):e0169141Google Scholar
  36. Wu Y, Deng Y, Zhu J, Duan Y, Weng WW, Wu X (2018) Pim1 promotes cell proliferation and regulates glycolysis via interaction with MYC in ovarian cancer. Onco Targets Ther 11:6647–6656Google Scholar
  37. Xie RF, Li ZC, Gao B, Shi ZN, Zhou X (2012) Bufothionine, a possible effective component in cinobufocini injection for hepatocellular carcinoma. J Ethnopharmacol 141(2):692–700Google Scholar
  38. Xie X, Huang X, Li J, Lv X, Huang J, Tang S, Sun Y (2013) Efficacy and safety of Huachansu combined with chemotherapy in advanced gastric cancer: a meta-analysis. Med Hypotheses 81(2):243–250Google Scholar
  39. Xu W, Luo H, Zhang Y, Shan L, Li H, Yang M, Liu R, Zhang W (2007) Simultaneous determination of five main active bufadienolides of Chan Su in rat plasma by liquid chromatography tandem mass spectrometry. J Chromatogr B Anal Technol Biomed Life Sci 859(2):157–163Google Scholar
  40. Xu S et al (2019) 1,25-(OH)2D3 protects Schwann cells against advanced glycation end products-induced apoptosis through PKA-NF-kappaB pathway. Life Sci 225:107–116Google Scholar
  41. Xue C, He Y, Hu Q, Yu Y, Chen X, Chen J, Ren F, Li J, Ren Z, Cui G, Sun R (2018) Downregulation of PIM1 regulates glycolysis and suppresses tumor progression in gallbladder cancer. Cancer Manag Res 10:5101–5112Google Scholar
  42. Yang Y, Cao Y, Chen L, Liu F, Qi Z, Cheng X, Wang Z (2018) Cryptotanshinone suppresses cell proliferation and glucose metabolism via STAT3/SIRT3 signaling pathway in ovarian cancer cells. Cancer Med 7(9):4610–4618Google Scholar
  43. Zhang X et al (2018) Cinobufacini injection improves the efficacy of chemotherapy on advanced stage gastric cancer: a systemic review and meta-analysis. Evid Based Complement Alternat Med 2018:7362340Google Scholar
  44. Zhou RP, Chen G, Shen ZL, Pan LQ (2014) Cinobufacin suppresses cell proliferation via miR-494 in BGC- 823 gastric cancer cells. Asian Pac J Cancer Prev 15(3):1241–1245Google Scholar
  45. Zou J, Wang Y, Liu M, Huang X, Zheng W, Gao Q, Wang H (2018) Euxanthone inhibits glycolysis and triggers mitochondria-mediated apoptosis by targeting hexokinase 2 in epithelial ovarian cancer. Cell Biochem Funct 36:303–311Google Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Qian Li
    • 1
  • Chuanwu Jiang
    • 2
    • 3
  • Yan Wang
    • 2
  • Minghua Wei
    • 2
  • Huijin Zheng
    • 2
  • Yanqi Xu
    • 2
  • Xuegang Xu
    • 4
  • Fengyu Jia
    • 3
  • Kai Liu
    • 3
  • Gang Sun
    • 3
  • Jianhua Zang
    • 5
    Email author
  • Ping Mo
    • 6
    Email author
  1. 1.Department of GynecologyJining No. 1 People’s HospitalJiningChina
  2. 2.Department of Interventional RadiologyQingdao Hospital of Traditional Chinese MedicineQingdaoChina
  3. 3.Department of Interventional RadiologyJinan Military General HospitalJinanChina
  4. 4.Department of Interventional RadiologyJining No. 2 People’s HospitalJiningChina
  5. 5.Department of OncologyQingdao Hospital of Traditional Chinese MedicineQingdaoChina
  6. 6.Department of GynecologyQingdao Hospital of Traditional Chinese MedicineQingdaoChina

Personalised recommendations