Skip to main content

Advertisement

SpringerLink
Log in

Expression of PFKFB3 and Ki67 in lung adenocarcinomas and targeting PFKFB3 as a therapeutic strategy

  • Published:
Molecular and Cellular Biochemistry Aims and scope Submit manuscript

Abstract

Phosphofructokinase-2/fructose-2, 6-bisphosphatase 3 (PFKFB3) catalyzes the synthesis of F2,6BP, which is an allosteric activator of 6-phosphofructo-1-kinase (PFK-1): the rate-limiting enzyme of glycolysis. During tumorigenesis, PFKFB3 increases glycolysis, angiogenesis, and tumor progression. In this study, our aim was to investigate the significance of PFKFB3 and Ki67 in human lung adenocarcinomas and to target PFKFB3 as a therapeutic strategy. In this study, we determined the expression levels of PFKFB3 mRNA and proteins in cancerous and normal lung adenocarcinomas by quantitative reverse transcription PCR (qRT-PCR), Western blot analysis, and tissue microarray immunohistochemistry analysis, respectively. In human adenocarcinoma tissues, PFKFB3 and Ki67 protein levels were related to the clinical characteristics and overall survival. Both PFKFB3 mRNA and protein were significantly higher in lung adenocarcinoma cells (all P < 0.05). A high expression of PFKFB3 and Ki67 were associated with the degree of differentiation, TNM staging, lymph node metastasis, and survival. A high expression of PFKFB3 protein was an independent prognostic marker in lung adenocarcinoma. Subsequently, 1-(4-pyridinyl)-3-(2-quinolinyl)-2-propen-1-one (PFK15) was used as a selective antagonist of PFKFB3. Glycolytic flux was determined by measuring glucose uptake, F2,6BP, and lactate production. Cell viability, cell cycle, cell apoptosis, cell migration, and invasion were analyzed by MTT, flow cytometry, Western blot analysis, wound healing assay, and transwell chamber assay. By targeting PFKFB3, it inhibited cell viability and glycolytic activity. It also caused apoptosis and induced cell cycle arrest. Furthermore, the migration and invasion of A549 cells was inhibited. We conclude that PFKFB3 bears an oncogene-like regulatory element in lung adenocarcinoma progression. In the treatment of lung adenocarcinoma, targeting PFKFB3 would be a promising therapeutic strategy.

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
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  1. Siegel R, Naishadham D, Jemal A (2013) Cancer statistics, 2013. CA Cancer J Clin 63:11–30. https://doi.org/10.3322/caac.21166

    Article  PubMed  Google Scholar 

  2. Saito M, Shiraishi K, Kunitoh H, Takenoshita S, Yokota J, Kohno T (2016) Gene aberrations for precision medicine against lung adenocarcinoma. Cancer Sci 107:713–720. https://doi.org/10.1111/cas.12941

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  3. Lee YS, Bae SC (2016) How do K-RAS-activated cells evade cellular defense mechanisms? Oncogene 35:827–832. https://doi.org/10.1038/onc.2015.153

    Article  PubMed  CAS  Google Scholar 

  4. Pasche B, Grant SC (2014) Non-small cell lung cancer and precision medicine: a model for the incorporation of genomic features into clinical trial design. JAMA 311:1975–1976. https://doi.org/10.1001/jama.2014.3742

    Article  PubMed  CAS  Google Scholar 

  5. Kohno T, Tsuta K, Tsuchihara K, Nakaoku T, Yoh K, Goto K (2013) RET fusion gene: translation to personalized lung cancer therapy. Cancer Sci 104:1396–1400. https://doi.org/10.1111/cas.12275

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  6. Bando H, Atsumi T, Nishio T, Niwa H, Mishima S, Shimizu C, Yoshioka N, Bucala R, Koike T (2005) Phosphorylation of the 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase/PFKFB3 family of glycolytic regulators in human cancer. Clin Cancer Res 11:5784–5792. https://doi.org/10.1158/1078-0432.CCR-05-0149

    Article  PubMed  CAS  Google Scholar 

  7. Clem B, Telang S, Clem A, Yalcin A, Meier J, Simmons A, Rasku MA, Arumugam S, Dean WL, Eaton J, Lane A, Trent JO, Chesney J (2008) Small-molecule inhibition of 6-phosphofructo-2-kinase activity suppresses glycolytic flux and tumor growth. Mol Cancer Ther 7:110–120. https://doi.org/10.1158/1535-7163.MCT-07-0482

    Article  PubMed  CAS  Google Scholar 

  8. Chesney J, Telang S, Yalcin A, Clem A, Wallis N, Bucala R (2005) Targeted disruption of inducible 6-phosphofructo-2-kinase results in embryonic lethality. Biochem Biophys Res Commun 331:139–146. https://doi.org/10.1016/j.bbrc.2005.02.193

    Article  PubMed  CAS  Google Scholar 

  9. Li HM, Yang JG, Liu ZJ, Wang WM, Yu ZL, Ren JG, Chen G, Zhang W, Jia J (2017) Blockage of glycolysis by targeting PFKFB3 suppresses tumor growth and metastasis in head and neck squamous cell carcinoma. J Exp Clin Cancer Res 36:7. https://doi.org/10.1186/s13046-016-0481-1

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  10. Minchenko OH, Tsuchihara K, Minchenko DO, Bikfalvi A, Esumi H (2014) Mechanisms of regulation of PFKFB expression in pancreatic and gastric cancer cells. World J Gastroenterol 20:13705–13717. https://doi.org/10.3748/wjg.v20.i38.13705

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  11. Wang JX, Zhang YY, Yu XM, Jin T, Pan XL (2012) Role of centromere protein H and Ki67 in relapse-free survival of patients after primary surgery for hypopharyngeal cancer. Asian Pac J Cancer Prev 13:821–825

    Article  PubMed  Google Scholar 

  12. Liu HB, Gao XX, Zhang Q, Liu J, Cui Y, Zhu Y, Liu YF (2015) Expression and prognostic implications of FOXO3a and Ki67 in lung adenocarcinomas. Asian Pac J Cancer Prev 16:1443–1448

    Article  PubMed  Google Scholar 

  13. Han J, Meng Q, Xi Q, Wang H, Wu G (2017) PFKFB3 was overexpressed in gastric cancer patients and promoted the proliferation and migration of gastric cancer cells. Cancer Biomark 18:249–256. https://doi.org/10.3233/CBM-160143

    Article  PubMed  CAS  Google Scholar 

  14. Clem BF, O’Neal J, Tapolsky G, Clem AL, Imbert-Fernandez Y, Kerr DA, 2nd, Klarer AC, Redman R, Miller DM, Trent JO, Telang S, Chesney J (2013) Targeting 6-phosphofructo-2-kinase (PFKFB3) as a therapeutic strategy against cancer. Mol Cancer Ther 12:1461–1470. https://doi.org/10.1158/1535-7163.MCT-13-0097

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  15. Liu Y, Lv L, Xue Q, Wan C, Ni T, Chen B, Liu Y, Zhou Y, Ni R, Mao G (2013) Vacuolar protein sorting 4B, an ATPase protein positively regulates the progression of NSCLC via promoting cell division. Mol Cell Biochem 381:163–171. https://doi.org/10.1007/s11010-013-1699-2

    Article  PubMed  CAS  Google Scholar 

  16. Zhu W, Ye L, Zhang J, Yu P, Wang H, Ye Z, Tian J (2016) PFK15, a small molecule inhibitor of PFKFB3, induces cell cycle arrest, apoptosis and inhibits invasion in gastric cancer. PLoS ONE 11:e0163768. https://doi.org/10.1371/journal.pone.0163768

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  17. Yalcin A, Clem BF, Imbert-Fernandez Y, Ozcan SC, Peker S, O’Neal J, Klarer AC, Clem AL, Telang S, Chesney J (2014) 6-Phosphofructo-2-kinase (PFKFB3) promotes cell cycle progression and suppresses apoptosis via Cdk1-mediated phosphorylation of p27. Cell Death Dis 5:e1337. https://doi.org/10.1038/cddis.2014.292

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  18. Yalcin A, Clem BF, Simmons A, Lane A, Nelson K, Clem AL, Brock E, Siow D, Wattenberg B, Telang S, Chesney J (2009) Nuclear targeting of 6-phosphofructo-2-kinase (PFKFB3) increases proliferation via cyclin-dependent kinases. J Biol Chem 284:24223–24232. https://doi.org/10.1074/jbc.M109.016816

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  19. Xu Y, An X, Guo X, Habtetsion TG, Wang Y, Xu X, Kandala S, Li Q, Li H, Zhang C, Caldwell RB, Fulton DJ, Su Y, Hoda MN, Zhou G, Wu C, Huo Y (2014) Endothelial PFKFB3 plays a critical role in angiogenesis. Arterioscler Thromb Vasc Biol 34:1231–1239. https://doi.org/10.1161/ATVBAHA.113.303041

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  20. Venning FA, Wullkopf L, Erler JT (2015) Targeting ECM disrupts cancer progression. Front Oncol 5:224. https://doi.org/10.3389/fonc.2015.00224

    Article  PubMed  PubMed Central  Google Scholar 

  21. Bian T, Jiang D, Liu J, Yuan X, Feng J, Li Q, Zhang Q, Li X, Liu Y, Zhang J (2017) miR-1236-3p suppresses the migration and invasion by targeting KLF8 in lung adenocarcinoma A549 cells. Biochem Biophys Res Commun 492:461–467. https://doi.org/10.1016/j.bbrc.2017.08.074

    Article  PubMed  CAS  Google Scholar 

  22. Jemal A, Siegel R, Xu J, Ward E (2010) Cancer statistics, 2010. CA Cancer J Clin 60:277–300. https://doi.org/10.3322/caac.20073

    Article  PubMed  Google Scholar 

  23. Travis WD, Brambilla E, Noguchi M, Nicholson AG, Geisinger K, Yatabe Y, Powell CA, Beer D, Riely G, Garg K, Austin JH, Rusch VW, Hirsch FR, Jett J, Yang PC, Gould M, American Thoracic Society (2011) International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society: international multidisciplinary classification of lung adenocarcinoma: executive summary. Proc Am Thorac Soc 8:381–385. https://doi.org/10.1513/pats.201107-042ST

    Article  PubMed  Google Scholar 

  24. Atsumi T, Chesney J, Metz C, Leng L, Donnelly S, Makita Z, Mitchell R, Bucala R (2002) High expression of inducible 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (iPFK-2; PFKFB3) in human cancers. Cancer Res 62:5881–5887

    PubMed  CAS  Google Scholar 

  25. Ge X, Lyu P, Cao Z, Li J, Guo G, Xia W, Gu Y (2015) Overexpression of miR-206 suppresses glycolysis, proliferation and migration in breast cancer cells via PFKFB3 targeting. Biochem Biophys Res Commun 463:1115–1121. https://doi.org/10.1016/j.bbrc.2015.06.068

    Article  PubMed  CAS  Google Scholar 

  26. Sun CM, Xiong DB, Yan Y, Geng J, Liu M, Yao XD (2016) Genetic alteration in phosphofructokinase family promotes growth of muscle-invasive bladder cancer. Int J Biol Markers 31:e286–e293. https://doi.org/10.5301/jbm.5000189

    Article  PubMed  CAS  Google Scholar 

  27. Minchenko A, Leshchinsky I, Opentanova I, Sang N, Srinivas V, Armstead V, Caro J (2002) Hypoxia-inducible factor-1-mediated expression of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (PFKFB3) gene. Its possible role in the Warburg effect. J Biol Chem 277:6183–6187. https://doi.org/10.1074/jbc.M110978200

    Article  PubMed  CAS  Google Scholar 

  28. Obach M, Navarro-Sabate A, Caro J, Kong X, Duran J, Gomez M, Perales JC, Ventura F, Rosa JL, Bartrons R (2004) 6-Phosphofructo-2-kinase (pfkfb3) gene promoter contains hypoxia-inducible factor-1 binding sites necessary for transactivation in response to hypoxia. J Biol Chem 279:53562–53570. https://doi.org/10.1074/jbc.M406096200

    Article  PubMed  CAS  Google Scholar 

  29. Klarer AC, O’Neal J, Imbert-Fernandez Y, Clem A, Ellis SR, Clark J, Clem B, Chesney J, Telang S (2014) Inhibition of 6-phosphofructo-2-kinase (PFKFB3) induces autophagy as a survival mechanism. Cancer Metab 2:2. https://doi.org/10.1186/2049-3002-2-2

    Article  PubMed  PubMed Central  Google Scholar 

  30. Zou Y, Zeng S, Huang M, Qiu Q, Xiao Y, Shi M, Zhan Z, Liang L, Yang X, Xu H (2017) Inhibition of 6-phosphofructo-2-kinase suppresses fibroblast-like synoviocytes-mediated synovial inflammation and joint destruction in rheumatoid arthritis. Br J Pharmacol. https://doi.org/10.1111/bph.13762

    Article  PubMed  PubMed Central  Google Scholar 

  31. Feng Y, Wu L (2017) mTOR up-regulation of PFKFB3 is essential for acute myeloid leukemia cell survival. Biochem Biophys Res Commun 483:897–903. https://doi.org/10.1016/j.bbrc.2017.01.031

    Article  PubMed  CAS  Google Scholar 

  32. Schoors S, De Bock K, Cantelmo AR, Georgiadou M, Ghesquiere B, Cauwenberghs S, Kuchnio A, Wong BW, Quaegebeur A, Goveia J, Bifari F, Wang X, Blanco R, Tembuyser B, Cornelissen I, Bouche A, Vinckier S, Diaz-Moralli S, Gerhardt H, Telang S, Cascante M, Chesney J, Dewerchin M, Carmeliet P (2014) Partial and transient reduction of glycolysis by PFKFB3 blockade reduces pathological angiogenesis. Cell Metab 19:37–48. https://doi.org/10.1016/j.cmet.2013.11.008

    Article  PubMed  CAS  Google Scholar 

  33. De Bock K, Georgiadou M, Schoors S, Kuchnio A, Wong BW, Cantelmo AR, Quaegebeur A, Ghesquiere B, Cauwenberghs S, Eelen G, Phng LK, Betz I, Tembuyser B, Brepoels K, Welti J, Geudens I, Segura I, Cruys B, Bifari F, Decimo I, Blanco R, Wyns S, Vangindertael J, Rocha S, Collins RT, Munck S, Daelemans D, Imamura H, Devlieger R, Rider M, Van Veldhoven PP, Schuit F, Bartrons R, Hofkens J, Fraisl P, Telang S, Deberardinis RJ, Schoonjans L, Vinckier S, Chesney J, Gerhardt H, Dewerchin M, Carmeliet P (2013) Role of PFKFB3-driven glycolysis in vessel sprouting. Cell 154:651–663. https://doi.org/10.1016/j.cell.2013.06.037

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgements

This study was funded by grants from Six talent peaks project in Jiangsu Province, China (No. WSN-059), the Science Foundation of Nantong City, Jiangsu, China (No. MS12015007), Scientific research topic of Jiangsu provincial health and Family Planning Commission, China (No. H201626), and Key talents of Medical Science in Jiangsu Province, China (No. QNRC2016682).

Author information

Author notes

  1. Xiaoli Li and Jian Liu have contributed equally.

Authors and Affiliations

  1. Department of Pathology, Affiliated Hospital of Nantong University, No. 20 Xisi Road, Nantong, 226001, Jiangsu, People’s Republic of China

    Xiaoli Li, Li Qian, Chan Yao, Wei Tian, Yifei Liu & Jianguo Zhang

  2. Department of Chemotherapy, Affiliated Hospital of Nantong University, Nantong, Jiangsu, People’s Republic of China

    Jian Liu

  3. Department of Respiratory Medicine, Haian County People’s Hospital, Affiliated to Nantong University, Nantong, Jiangsu, People’s Republic of China

    Honggang Ke

Authors
  1. Xiaoli Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jian Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Li Qian
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Honggang Ke
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Chan Yao
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Wei Tian
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Yifei Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Jianguo Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Yifei Liu or Jianguo Zhang.

Ethics declarations

Conflict of interest

The authors declared that they have no competing interests.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOC 229 KB)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Li, X., Liu, J., Qian, L. et al. Expression of PFKFB3 and Ki67 in lung adenocarcinomas and targeting PFKFB3 as a therapeutic strategy. Mol Cell Biochem 445, 123–134 (2018). https://doi.org/10.1007/s11010-017-3258-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11010-017-3258-8

Keywords

Search

Navigation