Simvastatin Enhanced Anti-tumor Effects of Bevacizumab against Lung Adenocarcinoma A549 Cells via Abating HIF-1α-Wnt/β-Catenin Signaling Pathway


Cite item

Full Text

Abstract

Background: Bevacizumab increased hypoxia-inducible factor (HIF-1α) expression attenuates its antitumor effect. Simvastatin can reduce the expression of HIF-1α to exert a tumor-suppressive effect in many in vitro experiments. Therefore, this study aimed to determine whether simvastatin could strengthen the anti-tumor activity of bevacizumab in lung adenocarcinoma.

Objective: To determine whether simvastatin could strengthen the anti-tumor activity of bevacizumab in lung adenocarcinoma.

Methods: The changes in the biological behavior of A549 cells treated with different drugs were determined through colony forming assay, Cell Counting Assay-8 (CCK-8), transwell assay, wound healing assay, and flow cytometry. The expressions of pathway-related factors HIF-1α and β-Catenin were determined via qRT-PCR and western blotting. The expressions of proliferation-related proteins, invasion-related proteins, and apoptosis-related proteins were detected by western blotting. In addition, a xenograft non-small cell lung cancer model in nude mice was used to explore in vivo tumor growth.

Results: We found that simvastatin combined with bevacizumab synergistically suppressed the proliferation, migration, and invasion of A549 cells while promoting their apoptosis. As demonstrated by qRT-PCR and western blotting experiments, the bevacizumab group displayed a higher expression of pathway-related factors HIF-1α and β-Catenin than the control groups, however simvastatin group showed the opposite trend. Its combination with bevacizumab induced elevation of HIF-1α and β-catenin expressions. During in vivo experiments, simvastatin inhibited tumor growth, and in comparison, the inhibitory effects of its combination with bevacizumab were stronger.

Conclusion: Based on our findings, simvastatin may affect the biological responses of bevacizumab on A549 cells by restraining the HIF-1α-Wnt/β-catenin signaling pathway, thus representing a novel and effective combination therapy that can be potentially applied in a clinical therapy for lung adenocarcinoma.

About the authors

Xin Tu

Department of Respiratory and Critical Care Medicine, The Third Affiliated Hospital of Chengdu Medical College, Chengdu Pidu District People's Hospital

Email: info@benthamscience.net

Jian Zhang

Department of Gastroenterology, The Second People's Hospital of Yibin

Email: info@benthamscience.net

Wei Yuan

Department of Neurology, The Third Affiliated Hospital of Chengdu Medical College, Chengdu Pidu District People's Hospital

Email: info@benthamscience.net

Xia Wu

Department of Respiratory and Critical Care Medicine, The Third Affiliated Hospital of Chengdu Medical College, Chengdu Pidu District People's Hospital

Email: info@benthamscience.net

Zhi Xu

Department of Respiratory and Critical Care Medicine, The Third Affiliated Hospital of Chengdu Medical College, Chengdu Pidu District People's Hospital

Email: info@benthamscience.net

Cuo Qing

Department of Respiratory and Critical Care Medicine, The Third Affiliated Hospital of Chengdu Medical College, Chengdu Pidu District People's Hospital

Author for correspondence.
Email: info@benthamscience.net

References

  1. Chen, W.; Zheng, R.; Baade, P.D.; Zhang, S.; Zeng, H.; Bray, F.; Jemal, A.; Yu, X.Q.; He, J. Cancer statistics in China, 2015. CA Cancer J. Clin., 2016, 66(2), 115-132. doi: 10.3322/caac.21338 PMID: 26808342
  2. Chen, Z.; Fillmore, C.M.; Hammerman, P.S.; Kim, C.F.; Wong, K.K. Non-small-cell lung cancers: A heterogeneous set of diseases. Nat. Rev. Cancer, 2014, 14(8), 535-546. doi: 10.1038/nrc3775 PMID: 25056707
  3. Schwartzberg, L.; Korytowsky, B.; Penrod, J.R.; Zhang, Y.; Le, T.K.; Batenchuk, C.; Krug, L. Real-world clinical impact of immune checkpoint inhibitors in patients with advanced/metastatic non–small cell lung cancer after platinum chemotherapy. Clin. Lung Cancer, 2019, 20(4), 287-296.e4. doi: 10.1016/j.cllc.2019.04.004 PMID: 31130450
  4. Chen, J.H.; Yang, J.L.; Chou, C.Y.; Wang, J.Y.; Hung, C.C. Indirect comparison of efficacy and safety between immune checkpoint inhibitors and antiangiogenic therapy in advanced non–small-cell lung cancer. Sci. Rep., 2018, 8(1), 9686. doi: 10.1038/s41598-018-27994-x PMID: 29946182
  5. Reck, M.; Garassino, M.C.; Imbimbo, M.; Shepherd, F.A.; Socinski, M.A.; Shih, J.Y.; Tsao, A.; Lee, P.; Winfree, K.B.; Sashegyi, A.; Cheng, R.; Varea, R.; Levy, B.; Garon, E. Antiangiogenic therapy for patients with aggressive or refractory advanced non-small cell lung cancer in the second-line setting. Lung Cancer, 2018, 120, 62-69. doi: 10.1016/j.lungcan.2018.03.025 PMID: 29748017
  6. Alevizakos, M.; Kaltsas, S.; Syrigos, K.N. The VEGF pathway in lung cancer. Cancer Chemother. Pharmacol., 2013, 72(6), 1169-1181. doi: 10.1007/s00280-013-2298-3 PMID: 24085262
  7. Blagosklonny, M.V. Antiangiogenic therapy and tumor progression. Cancer Cell, 2004, 5(1), 13-17. doi: 10.1016/S1535-6108(03)00336-2 PMID: 14749122
  8. De Francesco, E.M.; Sims, A.H.; Maggiolini, M.; Sotgia, F.; Lisanti, M.P.; Clarke, R.B. GPER mediates the angiocrine actions induced by IGF1 through the HIF-1α/VEGF pathway in the breast tumor microenvironment. Breast Cancer Res., 2017, 19(1), 129. doi: 10.1186/s13058-017-0923-5 PMID: 29212519
  9. Rapisarda, A.; Melillo, G. Overcoming disappointing results with antiangiogenic therapy by targeting hypoxia. Nat. Rev. Clin. Oncol., 2012, 9(7), 378-390. doi: 10.1038/nrclinonc.2012.64 PMID: 22525710
  10. Mazumdar, J.; O'Brien, W.T.; Johnson, R.S.; LaManna, J.C.; Chavez, J.C.; Klein, P.S.; Simon, M.C. O2 regulates stem cells through Wnt/β-catenin signalling. Nat. Cell Biol., 2010, 12(10), 1007-1013. doi: 10.1038/ncb2102 PMID: 20852629
  11. Lv, Z.; Liu, R.D.; Chen, X.Q.; Wang, B.; Li, L.F.; Guo, Y.S.; Chen, X.J.; Ren, X.Q. HIF 1α promotes the stemness of oesophageal squamous cell carcinoma by activating the Wnt/β catenin pathway. Oncol. Rep., 2019, 42(2), 726-734. doi: 10.3892/or.2019.7203 PMID: 31233197
  12. Wang, X.; Yu, Z.; Wang, C.; Cheng, W.; Tian, X.; Huo, X.; Wang, Y.; Sun, C.; Feng, L.; Xing, J.; Lan, Y.; Sun, D.; Hou, Q.; Zhang, B.; Ma, X.; Zhang, B. Alantolactone, a natural sesquiterpene lactone, has potent antitumor activity against glioblastoma by targeting IKKβ kinase activity and interrupting NF-κB/COX-2-mediated signaling cascades. J. Exp. Clin. Cancer Res., 2017, 36(1), 93. doi: 10.1186/s13046-017-0563-8 PMID: 28701209
  13. McIntyre, A.; Harris, A.L. Metabolic and hypoxic adaptation to anti‐angiogenic therapy: A target for induced essentiality. EMBO Mol. Med., 2015, 7(4), 368-379. doi: 10.15252/emmm.201404271 PMID: 25700172
  14. Vasudev, N.S.; Goh, V.; Juttla, J.K.; Thompson, V.L.; Larkin, J.M.G.; Gore, M.; Nathan, P.D.; Reynolds, A.R. Changes in tumour vessel density upon treatment with anti-angiogenic agents: Relationship with response and resistance to therapy. Br. J. Cancer, 2013, 109(5), 1230-1242. doi: 10.1038/bjc.2013.429 PMID: 23922108
  15. Huang, W.; Zhang, C.; Cui, M.; Niu, J.; Ding, W. Inhibition of bevacizumab-induced epithelial-mesenchymal transition by BATF2 overexpression involves the suppression of Wnt/β-catenin signaling in glioblastoma cells. Anticancer Res., 2017, 37(8), 4285-4294. doi: 10.21873/anticanres.11821 PMID: 28739720
  16. Xie, W.; Zhao, H.; Wang, F.; Wang, Y.; He, Y.; Wang, T.; Zhang, K.; Yang, H.; Zhou, Z.; Shi, H.; Wang, J.; Huang, G. A novel humanized Frizzled-7-targeting antibody enhances antitumor effects of Bevacizumab against triple-negative breast cancer via blocking Wnt/β-catenin signaling pathway. J. Exp. Clin. Cancer Res., 2021, 40(1), 30. doi: 10.1186/s13046-020-01800-x PMID: 33436039
  17. Shepherd, J.; Cobbe, S.M.; Ford, I.; Isles, C.G.; Lorimer, A.R.; Macfarlane, P.W.; McKillop, J.H.; Packard, C.J. Prevention of coronary heart disease with pravastatin in men with hypercholesterolemia. N. Engl. J. Med., 1995, 333(20), 1301-1308. doi: 10.1056/NEJM199511163332001 PMID: 7566020
  18. Fujikake, K.; Kajiyama, H.; Yoshihara, M.; Nishino, K.; Yoshikawa, N.; Utsumi, F.; Suzuki, S.; Niimi, K.; Sakata, J.; Mitsui, H.; Shibata, K.; Senga, T.; Kikkawa, F. A novel mechanism of neovascularization in peritoneal dissemination via cancer-associated mesothelial cells affected by TGF-β derived from ovarian cancer. Oncol. Rep., 2017, 39(1), 193-200. doi: 10.3892/or.2017.6104 PMID: 29192324
  19. Murai, T. Cholesterol lowering: Role in cancer prevention and treatment. Biol. Chem., 2015, 396(1), 1-11. doi: 10.1515/hsz-2014-0194 PMID: 25205720
  20. Lee, Y.; Lee, K.H.; Lee, G.K.; Lee, S.H.; Lim, K.Y.; Joo, J.; Go, Y.J.; Lee, J.S.; Han, J.Y. Randomized phase ii study of afatinib plus simvastatin versus afatinib alone in previously treated patients with advanced nonadenocarcinomatous non-small cell lung cancer. Cancer Res. Treat., 2017, 49(4), 1001-1011. doi: 10.4143/crt.2016.546 PMID: 28111428
  21. Feng, J.; Dai, W.; Mao, Y.; Wu, L.; Li, J.; Chen, K.; Yu, Q.; Kong, R.; Li, S.; Zhang, J.; Ji, J.; Wu, J.; Mo, W.; Xu, X.; Guo, C. Simvastatin re-sensitizes hepatocellular carcinoma cells to sorafenib by inhibiting HIF-1α/PPAR-γ/PKM2-mediated glycolysis. J. Exp. Clin. Cancer Res., 2020, 39(1), 24. doi: 10.1186/s13046-020-1528-x PMID: 32000827
  22. Chou, T.C.; Martin, N. CompuSyn software. CompuSyn for drug combinations: PC software and user's guide: A computer program for quantitation of synergism and antagonism in drug combinations, and the determination of IC50 and ED50 and LD50 values. ComboSyn Inc., Paramus, NJ. 2005. Available from: https://www.combosyn.com
  23. Chou, T.C.; Talalay, P. Generalized equations for the analysis of inhibitions of Michaelis-Menten and higher-order kinetic systems with two or more mutually exclusive and nonexclusive inhibitors. Eur. J. Biochem., 1981, 115(1), 207-216. doi: 10.1111/j.1432-1033.1981.tb06218.x PMID: 7227366
  24. Welti, J.; Loges, S.; Dimmeler, S.; Carmeliet, P. Recent molecular discoveries in angiogenesis and antiangiogenic therapies in cancer. J. Clin. Invest., 2013, 123(8), 3190-3200. doi: 10.1172/JCI70212 PMID: 23908119
  25. Kerbel, R.; Folkman, J. Clinical translation of angiogenesis inhibitors. Nat. Rev. Cancer, 2002, 2(10), 727-739. doi: 10.1038/nrc905 PMID: 12360276
  26. Lu, X.; Kang, Y. Hypoxia and hypoxia-inducible factors: Master regulators of metastasis. Clin. Cancer Res., 2010, 16(24), 5928-5935. doi: 10.1158/1078-0432.CCR-10-1360 PMID: 20962028
  27. Ai, Z.; Lu, Y.; Qiu, S.; Fan, Z. Overcoming cisplatin resistance of ovarian cancer cells by targeting HIF-1-regulated cancer metabolism. Cancer Lett., 2016, 373(1), 36-44. doi: 10.1016/j.canlet.2016.01.009 PMID: 26801746
  28. Tong, D.; Liu, Q.; Liu, G.; Yuan, W.; Wang, L.; Guo, Y.; Lan, W.; Zhang, D.; Dong, S.; Wang, Y.; Xiao, H.; Mu, J.; Mao, C.; Wong, J.; Jiang, J. The HIF/PHF8/AR axis promotes prostate cancer progression. Oncogenesis, 2016, 5(12), e283. doi: 10.1038/oncsis.2016.74 PMID: 27991916
  29. Zhang, Y.; Bian, Y.; Wang, Y.; Wang, Y.; Duan, X.; Han, Y.; Zhang, L.; Wang, F.; Gu, Z.; Qin, Z. HIF‐1α is necessary for activation and tumour‐promotion effect of cancer‐associated fibroblasts in lung cancer. J. Cell. Mol. Med., 2021, 25(12), 5457-5469. doi: 10.1111/jcmm.16556 PMID: 33943003
  30. Valenta, T.; Hausmann, G.; Basler, K. The many faces and functions of β-catenin. EMBO J., 2012, 31(12), 2714-2736. doi: 10.1038/emboj.2012.150 PMID: 22617422
  31. Pastushenko, I.; Brisebarre, A.; Sifrim, A.; Fioramonti, M.; Revenco, T.; Boumahdi, S.; Van Keymeulen, A.; Brown, D.; Moers, V.; Lemaire, S.; De Clercq, S.; Minguijón, E.; Balsat, C.; Sokolow, Y.; Dubois, C.; De Cock, F.; Scozzaro, S.; Sopena, F.; Lanas, A.; D'Haene, N.; Salmon, I.; Marine, J.C.; Voet, T.; Sotiropoulou, P.A.; Blanpain, C. Identification of the tumour transition states occurring during EMT. Nature, 2018, 556(7702), 463-468. doi: 10.1038/s41586-018-0040-3 PMID: 29670281
  32. Brabletz, T.; Kalluri, R.; Nieto, M.A.; Weinberg, R.A. EMT in cancer. Nat. Rev. Cancer, 2018, 18(2), 128-134. doi: 10.1038/nrc.2017.118 PMID: 29326430
  33. Nusse, R.; Clevers, H. Wnt/β-catenin signaling, disease, and emerging therapeutic modalities. Cell, 2017, 169(6), 985-999. doi: 10.1016/j.cell.2017.05.016 PMID: 28575679
  34. Han, P.; Li, J.; Zhang, B.; Lv, J.; Li, Y.; Gu, X.; Yu, Z.; Jia, Y.; Bai, X.; Li, L.; Liu, Y.; Cui, B. The lncRNA CRNDE promotes colorectal cancer cell proliferation and chemoresistance via miR-181a-5p-mediated regulation of Wnt/β-catenin signaling. Mol. Cancer, 2017, 16(1), 9. doi: 10.1186/s12943-017-0583-1 PMID: 28086904
  35. Teng, Y.; Wang, X.; Wang, Y.; Ma, D. Wnt/β-catenin signaling regulates cancer stem cells in lung cancer A549 cells. Biochem. Biophys. Res. Commun., 2010, 392(3), 373-379. doi: 10.1016/j.bbrc.2010.01.028 PMID: 20074550
  36. Niehrs, C. The complex world of WNT receptor signalling. Nat. Rev. Mol. Cell Biol., 2012, 13(12), 767-779. doi: 10.1038/nrm3470 PMID: 23151663
  37. Zhou, Y.Y.; Zhu, G.Q.; Wang, Y.; Zheng, J.N.; Ruan, L.Y.; Cheng, Z.; Hu, B.; Fu, S.W.; Zheng, M.H. Systematic review with network meta-analysis: Statins and risk of hepatocellular carcinoma. Oncotarget, 2016, 7(16), 21753-21762. doi: 10.18632/oncotarget.7832 PMID: 26943041
  38. Ishikawa, S.; Hayashi, H.; Kinoshita, K.; Abe, M.; Kuroki, H.; Tokunaga, R.; Tomiyasu, S.; Tanaka, H.; Sugita, H.; Arita, T.; Yagi, Y.; Watanabe, M.; Hirota, M.; Baba, H. Statins inhibit tumor progression via an enhancer of zeste homolog 2-mediated epigenetic alteration in colorectal cancer. Int. J. Cancer, 2014, 135(11), 2528-2536. doi: 10.1002/ijc.28672 PMID: 24346863
  39. Cardwell, C.R.; Mc Menamin, Ú.; Hughes, C.M.; Murray, L.J. Statin use and survival from lung cancer: A population-based cohort study. Cancer Epidemiol. Biomarkers Prev., 2015, 24(5), 833-841. doi: 10.1158/1055-9965.EPI-15-0052 PMID: 25934831
  40. Yu, X.; Pan, Y.; Ma, H.; Li, W. Simvastatin inhibits proliferation and induces apoptosis in human lung cancer cells. Oncol. Res., 2013, 20(8), 351-357. doi: 10.3727/096504013X13657689382897 PMID: 23924855
  41. Han, J.Y.; Lee, S.H.; Yoo, N.J.; Hyung, L.S.; Moon, Y.J.; Yun, T.; Kim, H.T.; Lee, J.S. A randomized phase II study of gefitinib plus simvastatin Versus gefitinib alone in previously treated patients with advanced non-small cell lung cancer. Clin. Cancer Res., 2011, 17(6), 1553-1560. doi: 10.1158/1078-0432.CCR-10-2525 PMID: 21411446
  42. Rauca, V.F.; Licarete, E.; Luput, L.; Sesarman, A.; Patras, L.; Bulzu, P.; Rakosy-Tican, E.; Banciu, M. Combination therapy of simvastatin and 5, 6-dimethylxanthenone-4-acetic acid synergistically suppresses the aggressiveness of B16.F10 melanoma cells. PLoS One, 2018, 13(8), e0202827. doi: 10.1371/journal.pone.0202827 PMID: 30138430
  43. Wang, J.C.; Li, X.X.; Sun, X.; Li, G.Y.; Sun, J.L.; Ye, Y.P.; Cong, L.L.; Li, W.M.; Lu, S.Y.; Feng, J.; Liu, P.J. Activation of AMPK by simvastatin inhibited breast tumor angiogenesis via impeding HIF ‐1α‐induced pro‐angiogenic factor. Cancer Sci., 2018, 109(5), 1627-1637. doi: 10.1111/cas.13570 PMID: 29532562

Supplementary files

Supplementary Files
Action
1. JATS XML
Download

Copyright (c) 2023 Bentham Science Publishers