ISSN 2410-7751 (Print)
ISSN 2410-776X (Online)
Biotechnologia Acta Т. 16, No. 5 , 2023
P. 34-44, Bibliography 36, Engl.
UDC::616.091:576.31
DOI: https://doi.org/10.15407/biotech16.05.034
Full text: (PDF, in English)
Deniz Özdemir1, Seher Saruhan2, Can Ali Agca3
Department of Molecular Biology and Genetics, Bingol University, Bingol, TÜRKİYE
Aim.FKFB3 is a glycolytic activator that is overexpressed in human lung cancer and plays a crucial role in multiple cellular functions, including programmed cell death. Despite the many small molecules described as PFKFB3 inhibitors, some of them have shown disappointing results in vitro and in vivo. On the other hand, KAN0438757, a selective and potent small molecule inhibitor, has been developed. However, the effects of KAN0438757 in non-small cell lung carcinoma cells remain unknown. Herein, we sought to decipher the impact of KAN0438757 on proliferation, migration, DNA damage, and programmed cell death in non-small cell lung carcinoma cells.
Methods. The effects of KAN0438757 on cell viability, proliferation, DNA damage, migration, apoptosis, and autophagy in non-small cell lung carcinoma cells were tested by WST-1, real-time cell analysis, comet assay, wound-healing migration test, and MMP/JC-1 and AO/ER dual staining assays as well as western blot analysis.
Results. Our results revealed that KAN0438757 significantly suppressed the viability and proliferation of A549 and H1299 cells and inhibited migration of A549 cells. More importantly, KAN0438757 caused DNA damage and triggered apoptosis and this was accompanied by the up-regulation of cleaved PARP in A549 cells. Furthermore, treatment with KAN0438757 resulted in increased LC3 II and Beclin1, which indicated that KAN0438757 stimulated autophagy.
Conclusions. Overall, targeting PFKFB3 with KAN0438757 may be a promising, practical treatment approach, requiring further in vitro and in vivo evaluation of KAN0438757 as a therapy in non-small cell lung carcinoma cells.
Keywords: PFKFB3, KAN0438757, Apoptosis, Autophagy, Lung Cancer.
© Palladin Institute of Biochemistry of National Academy of Sciences of Ukraine, 2023
{
References
1. Warburg O. On the origin of cancer cells. Science. New York, 1956, 123(3191), 309–314. https://doi.org/10.1126/science.123.3191.309
2. Koppenol W. H., Bounds P. L, Dang C. V. Otto Warburg’s contributions to current concepts of cancer metabolism. Nature Reviews Cancer. 2011, 11(5), 325–337. https://doi.org/10.1038/nrc3038
3. Boroughs L. K., Deberardinis R. J. Nisan 30. Metabolic pathways promoting cancer cell survival and growth. Nature Cell Biology Nature Publishing Group. 2015. https://doi.org/10.1038/ncb3124
4. Lu L., Chen Y., Zhu Y., Lu L., Chen Y., Zhu Y. The molecular basis of targeting PFKFB3 as a therapeutic strategy against cancer. Oncotarget 2017, 8(37), 62793–62802. https://doi.org/10.18632/oncotarget.19513
5. Chesney J., Telang S., Yalcin A., Clem A., Wallis N., Bucala R. Targeted disruption of inducible 6-phosphofructo-2-kinase results in embryonic lethality. Biochemical and Biophysical Research Communications. 2005, 331(1), 139–146. https://doi.org/10.1016/j.bbrc.2005.02.193
6. Yalcin Abdullah, Telang S., Clem B., Chesney J. Regulation of glucose metabolism by 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatases in cancer. Experimental and Molecular Pathology. 2009, 86(3), 174–179. https://doi.org/10.1016/j.yexmp.2009.01.003
7. Cordero-Espinoza L., Hagen T. Increased Concentrations of Fructose 2,6-Bisphosphate Contribute to the Warburg Effect in Phosphatase and Tensin Homolog (PTEN)-deficient Cells. Journal of Biological Chemistry. 2013, 288(50), 36020–36028. https://doi.org/10.1074/jbc.M113.510289
8. Bolaños J. P. Adapting glycolysis to cancer cell proliferation: the MAPK pathway focuses on PFKFB3. Biochemical Journal. 2013, 452(3), e7–e9. https://doi.org/10.1042/BJ20130560
9. N. L, S.M. D., L. L., J. C., Y. I.F. PFKFB3 Inhibition Impairs Erlotinib-Induced Autophagy in NSCLCs. Cells. 2021, 10(7). https://doi.org/10.3390/cells10071679
10.Obach M., Navarro-Sabaté A., Caro J., Kong X., Duran J., Gómez M., Bartrons R. 6-Phosphofructo-2-kinase (pfkfb3) Gene Promoter Contains Hypoxia-inducible Factor-1 Binding Sites Necessary for Transactivation in Response to Hypoxia. Journal of Biological Chemistry. 2004, 279(51), 53562–53570. https://doi.org/10.1074/jbc.M406096200
11. Clem B., Telang S., Clem A., Yalcin A., Meier J., Simmons A., Chesney J. Small-molecule inhibition of 6-phosphofructo-2-kinase activity suppresses glycolytic flux and tumor growth. Molecular cancer therapeutics. 2008, 7(1), 110–120. https://doi.org/10.1158/1535-7163.MCT-07-0482
12. Horváthová J., Moravčík R., Boháč A., Zeman M. Synergic effects of inhibition of glycolysis and multikinase receptor signalling on proliferation and migration of endothelial cells. General physiology and biophysics. 2019, 38(2), 157–163. https://doi.org/10.4149/gpb_2018047
13. Clem B.F., O’Neal J., Tapolsky G., Clem A. L., Imbert-Fernandez Y., Kerr D. A., Chesney J. Targeting 6-phosphofructo-2-kinase (PFKFB3) as a therapeutic strategy against cancer. Molecular Cancer Therapeutics. 2013, 12(8), 1461–1470. https://doi.org/10.1158/1535-7163.MCT-13-0097
14. Li H. M., Yang J. G., Liu Z. J., Wang W. M., Yu Z. L., Ren J. G., Jia J. Blockage of glycolysis by targeting PFKFB3 suppresses tumor growth and metastasis in head and neck squamous cell carcinoma. Journal of Experimental and Clinical Cancer Research. 2017, 36(1), 1–12. https://doi.org/10.1186/s13046-016-0481-1
15. Gustafsson N. M. S., Färnegårdh K., Bonagas N., Ninou A. H., Groth P., Wiita E., Helleday T. Targeting PFKFB3 radiosensitizes cancer cells and suppresses homologous recombination. Nature Communications. 2018, 9(1). https://doi.org/10.1038/s41467-018-06287-x
16. Li X., Liu J., Qian L., Ke H., Yao C., Tian W., Zhang J. Expression of PFKFB3 and Ki67 in lung adenocarcinomas and targeting PFKFB3 as a therapeutic strategy. Molecular and Cellular Biochemistry. 2018, 445(1–2), 123–134. https://doi.org/10.1007/s11010-017-3258-8
17. Cory G. Scratch-wound assay. Methods in Molecular Biology (Clifton, N.J.). 2011, 769, 25–30. https://doi.org/10.1007/978-1-61779-207-6_2
18. Liu K., Liu Pcheng, Liu R., Wu X. Dual AO/EB staining to detect apoptosis in osteosarcoma cells compared with flow cytometry. Medical science monitor basic research. 2015, 21, 15–20. https://doi.org/10.12659/MSMBR.893327
19. Yalcin A,. Clem BF., Imbert-Fernandez Y., Ozcan S. C., Peker S., O’Neal J., Chesney J. 6-Phosphofructo-2-kinase (PFKFB3) promotes cell cycle progression and suppresses apoptosis via Cdk1-mediated phosphorylation of p27. Cell Death & Disease, 2014, 5(7), e1337–e1337. https://doi.org/10.1038/cddis.2014.292
20. Thirusangu P., Ray U., Sarkar Bhattacharya S., Oien D. B., Jin L., Staub J., Shridhar V. PFKFB3 regulates cancer stemness through the hippo pathway in small cell lung carcinoma. Oncogene. 2022, 41(33), 4003–4017. https://doi.org/10.1038/s41388-022-02391-x
21. De Oliveira T., Goldhardt T., Edelmann M., Rogge T.., Rauch K, Kyuchukov N. D., Conradi L. C. Effects of the Novel PFKFB3 Inhibitor KAN0438757 on Colorectal Cancer Cells and Its Systemic Toxicity Evaluation In Vivo. Cancers. 2021, 13(5), 1–24. https://doi.org/10.3390/cancers13051011
22. Shi W. K., Zhu X. D., Wang C. H., Zhang Y. Y., Cai H., Li X. L., Sun H. C. PFKFB3 blockade inhibits hepatocellular carcinoma growth by impairing DNA repair through AKT. Cell Death & Disease. 2018, 9(4), 1–12. https://doi.org/10.1038/s41419-018-0435-y
23. Ninou A. H.., Lehto J, Chioureas D., Stigsdotter H., Schelzig K., Åkerlund E., Gustafsson N. M. S. PFKFB3 Inhibition Sensitizes DNA Crosslinking Chemotherapies by Suppressing Fanconi Anemia Repair. Cancers. 2021, 13(14). https://doi.org/10.3390/cancers13143604
24. Agca C. A., Kırıcı M., Nedzvetsky V. S., Gundogdu R., Tykhomyrov A. A. The Effect of TIGAR Knockdown on Apoptotic and Epithelial-Mesenchymal Markers Expression in Doxorubicin-Resistant Non-Small Cell Lung Cancer A549 Cell Lines. Chemistry and Biodiversity. 2020, 17(9). https://doi.org/10.1002/cbdv.202000441
25. Ly J. D., Grubb D. R., Lawen A. The mitochondrial membrane potential (δψm) in apoptosis; an update. Apoptosis. 2003, 8(2), 115–128. https://doi.org/10.1023/a:1022945107762
26. Herceg Z., Wang Z. Q. Functions of poly(ADP-ribose) polymerase (PARP) in DNA repair, genomic integrity and cell death. Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis. 2001, 477(1–2), 97–110. https://doi.org/10.1016/s0027-5107(01)00111-7
27. Zhu W., Ye L., Zhang J., Yu P., Wang H., Ye Z., Tian J. PFK15, a Small Molecule Inhibitor of PFKFB3, Induces Cell Cycle Arrest, Apoptosis and Inhibits Invasion in Gastric Cancer. 2016, 11(9), e0163768. https://doi.org/10.1371/journal.pone.0163768
28. Peng F., Li Q., Sun J. Y., Luo Y., Chen M., Bao Y. PFKFB3 is involved in breast cancer proliferation, migration, invasion and angiogenesis. International journal of oncology. 2018, 52(3), 945–954. https://doi.org/10.3892/ijo.2018.4257
29. Gu M., Li L., Zhang Z., Chen J., Zhang W., Zhang J., You Y. PFKFB3 promotes proliferation, migration and angiogenesis in nasopharyngeal carcinoma. Journal of Cancer. 2017, 8(18), 3887–3896. https://doi.org/10.7150/jca.19112
30. Han J., Meng Q., Xi Q., Wang H., Wu G. PFKFB3 was overexpressed in gastric cancer patients and promoted the proliferation and migration of gastric cancer cells. Cancer biomarkers : section A of Disease markers. 2017, 18(3), 249–256. https://doi.org/10.3233/CBM-160143
31. Zheng W. D., Zhou F. L., Lin N. MicroRNA-26b inhibits osteosarcoma cell migration and invasion by down-regulating PFKFB3 expression. Genetics and molecular research : GMR. 2015, 14(4), 16872–16879. https://doi.org/10.4238/2015.Aralık.14.14
32. Zhang D. M., Zhang T., Wang M. M., Wang X. X., Qin Y. Y., Wu J., Qin Z. H. TIGAR alleviates ischemia/reperfusion-induced autophagy and ischemic brain injury. Free Radical Biology and Medicine. 2019, 137, 13–23. https://doi.org/10.1016/j.freeradbiomed.2019.04.002
33. Yun C. W., Lee S. H. The Roles of Autophagy in Cancer. International Journal of Molecular Sciences. 2018, 19(11). https://doi.org/10.3390/ijms19113466
34. Mondal S., Roy D., Sarkar Bhattacharya S., Jin L., Jung D., Zhang S., Shridhar V. Therapeutic targeting of PFKFB3 with a novel glycolytic inhibitor PFK158 promotes lipophagy and chemosensitivity in gynecologic cancers. International Journal of Cancer. 2019, 144(1), 178. https://doi.org/10.1002/ijc.31868
35. Klarer A. C., O’neal J., Imbert-Fernandez Y., Clem A., Ellis S. R., Clark J., Telang S. Inhibition of 6-phosphofructo-2-kinase (PFKFB3) induces autophagy as a survival mechanism. 2014, 2, 1–14. https://doi.org/10.1186/2049-3002-2-2
36. Xi H., Wang S., Wang B., Hong X., Liu X., Li M., Dong Q. The role of interaction between autophagy and apoptosis in tumorigenesis (Review). Oncology reports, 2022, 48(6), 1–16. https://doi.org/10.3892/or.2022.8423