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Anti-Cancer Agents in Medicinal Chemistry

Editor-in-Chief

ISSN (Print): 1871-5206
ISSN (Online): 1875-5992

Research Article

Establishment and Characterization of Pemetrexed-resistant NCI-H460/PMT Cells

Author(s): Yu-Lian Xu, Xiao-Ming Jiang, Le-Le Zhang, Xiuping Chen, Zhang-Jian Huang and Jin-Jian Lu*

Volume 19, Issue 6, 2019

Page: [731 - 739] Pages: 9

DOI: 10.2174/1871520619666190307120441

Price: $65

Abstract

Background: Pemetrexed (PMT) is a multitargeted antifolate agent that is used for treating patients with Non-Small Cell Lung Cancer (NSCLC). However, patients have presented clinical responses of drug resistance to PMT.

Objective: This study aimed to explore the underlying mechanisms of PMT resistance in NSCLC cells.

Methods: PMT-resistant NCI-H460/PMT cells were established by treating with PMT in a concentrationescalation manner. MTT assay and colony formation were performed to detect cell proliferation. Immunofluorescence was used to detect the expression of Ki-67. Transwell assay was performed to measure cell migration ability. qPCR and Western blot were used to detect the mRNA and protein expression levels of indicated genes. Small interfering RNAs (siRNA) were used to knockdown ATP binding cassette subfamily B member 1 (ABCB1) and Thymidylate Synthase (TYMS).

Results: This study showed that compared with the parental cells, the NCI-H460/PMT cells displayed weakened proliferation and enhanced cell mobility. In addition, the NCI-H460/PMT cells demonstrated cellular senescence, which might result in PMT resistance. The NCI-H460/PMT cells exhibited cross-resistance to other chemotherapeutics, including fluorouracil, paclitaxel, doxorubicin, etoposide and gemcitabine, possibly because of the upregulated expression of ABCB1. However, the ABCB1 knockdown by siRNA failed to eradicate PMT resistance. Moreover, TYMS, a target of PMT, was obviously upregulated in the resistant cells. The genetic silence of TYMS partially abrogated PMT resistance, suggesting that the overexpression of TYMS was a key resistant mechanism of PMT.

Conclusion: The overexpression of TYMS was an important resistance mechanism of PMT for KRAS-mutated NCI-H460 cells. Cross-resistance to other chemotherapeutics should be considered in addressing PMT resistance.

Keywords: NSCLC, pemetrexed, resistance, TYMS, ABCB1, chemotherapy.

Graphical Abstract

[1]
Ferlay, J.; Soerjomataram, I.; Dikshit, R.; Eser, S.; Mathers, C.; Rebelo, M.; Parkin, D.M.; Forman, D.; Bray, F. Cancer incidence and mortality worldwide: Sources, methods and major patterns in GLOBOCAN 2012. Int. J. Cancer, 2015, 136(5), E359-E386.
[2]
Swanton, C.; Govindan, R. Clinical implications of genomic discoveries in lung cancer. N. Engl. J. Med., 2016, 374(19), 1864-1873.
[3]
Hirsch, F.R.; Scagliotti, G.V.; Mulshine, J.L.; Kwon, R.; Curran, W.J., Jr; Wu, Y.L.; Paz-Ares, L. Lung cancer: Current therapies and new targeted treatments. Lancet, 2017, 389(10066), 299-311.
[4]
Mok, T.S.; Wu, Y.L.; Ahn, M.J.; Garassino, M.C.; Kim, H.R.; Ramalingam, S.S.; Shepherd, F.A.; He, Y.; Akamatsu, H.; Theelen, W.S.; Lee, C.K.; Sebastian, M.; Templeton, A.; Mann, H.; Marotti, M.; Ghiorghiu, S.; Papadimitrakopoulou, V.A.; Investigators, A. Osimertinib or platinum-pemetrexed in EGFR T790M-positive lung cancer. N. Engl. J. Med., 2017, 376(7), 629-640.
[5]
Tan, W.L.; Jain, A.; Takano, A.; Newell, E.W.; Iyer, N.G.; Lim, W.T.; Tan, E.H.; Zhai, W.; Hillmer, A.M.; Tam, W.L.; Tan, D.S.W. Novel therapeutic targets on the horizon for lung cancer. Lancet Oncol., 2016, 17(8), e347-e362.
[6]
Ettinger, D.S.; Wood, D.E.; Aisner, D.L.; Akerley, W.; Bauman, J.; Chirieac, L.R.; D’Amico, T.A.; DeCamp, M.M.; Dilling, T.J.; Dobelbower, M.; Doebele, R.C.; Govindan, R.; Gubens, M.A.; Hennon, M.; Horn, L.; Komaki, R.; Lackner, R.P.; Lanuti, M.; Leal, T.A.; Leisch, L.J.; Lilenbaum, R.; Lin, J.; Loo, B.W., Jr; Martins, R.; Otterson, G.A.; Reckamp, K.; Riely, G.J.; Schild, S.E.; Shapiro, T.A.; Stevenson, J.; Swanson, S.J.; Tauer, K.; Yang, S.C.; Gregory, K.; Hughes, M. Non-small cell lung cancer, version 5.2017, NCCN clinical practice guidelines in oncology. J. Natl. Compr. Canc. Netw., 2017, 15(4), 504-535.
[7]
Hirsch, F.R.; Suda, K.; Wiens, J.; Bunn, P.A., Jr New and emerging targeted treatments in advanced non-small-cell lung cancer. Lancet, 2016, 388(10048), 1012-1024.
[8]
Tomasini, P.; Barlesi, F.; Mascaux, C.; Greillier, L. Pemetrexed for advanced stage nonsquamous non-small cell lung cancer: Latest evidence about its extended use and outcomes. Ther. Adv. Med. Oncol., 2016, 8(3), 198-208.
[9]
Hazarika, M.; White, R.M.; Johnson, J.R.; Pazdur, R. FDA drug approval summaries: Pemetrexed (Alimta). Oncologist, 2004, 9(5), 482-488.
[10]
Gibbs, D.; Jackman, A. Pemetrexed disodium. Nat. Rev. Drug Discov., 2005. Suppl, S16-17.
[11]
Hanna, N.; Shepherd, F.A.; Fossella, F.V.; Pereira, J.R.; De Marinis, F.; von Pawel, J.; Gatzemeier, U.; Tsao, T.C.; Pless, M.; Muller, T.; Lim, H.L.; Desch, C.; Szondy, K.; Gervais, R. Shaharyar; Manegold, C.; Paul, S.; Paoletti, P.; Einhorn, L.; Bunn, P.A., Jr. Randomized phase III trial of pemetrexed versus docetaxel in patients with non-small-cell lung cancer previously treated with chemotherapy. J. Clin. Oncol., 2004, 22(9), 1589-1597.
[12]
Scagliotti, G.V.; Parikh, P.; von Pawel, J.; Biesma, B.; Vansteenkiste, J.; Manegold, C.; Serwatowski, P.; Gatzemeier, U.; Digumarti, R.; Zukin, M.; Lee, J.S.; Mellemgaard, A.; Park, K.; Patil, S.; Rolski, J.; Goksel, T.; de Marinis, F.; Simms, L.; Sugarman, K.P.; Gandara, D. Phase III study comparing cisplatin plus gemcitabine with cisplatin plus pemetrexed in chemotherapy-naive patients with advanced-stage non-small-cell lung cancer. J. Clin. Oncol., 2008, 26(21), 3543-3551.
[13]
Cohen, M.H.; Justice, R.; Pazdur, R. Approval summary: Pemetrexed in the initial treatment of advanced/metastatic non-small cell lung cancer. Oncologist, 2009, 14(9), 930-935.
[14]
Cohen, M.H.; Cortazar, P.; Justice, R.; Pazdur, R. Approval summary: Pemetrexed maintenance therapy of advanced/metastatic nonsquamous, Non-Small Cell Lung Cancer (NSCLC). Oncologist, 2010, 15(12), 1352-1358.
[15]
Schuette, W.H.; Groschel, A.; Sebastian, M.; Andreas, S.; Muller, T.; Schneller, F.; Guetz, S.; Eschbach, C.; Bohnet, S.; Leschinger, M.I.; Reck, M. A randomized phase II study of pemetrexed in combination with cisplatin or carboplatin as first-line therapy for patients with locally advanced or metastatic non-small-cell lung cancer. Clin. Lung Cancer, 2013, 14(3), 215-223.
[16]
Pujol, J.L.; Paul, S.; Chouaki, N.; Peterson, P.; Moore, P.; Berry, D.A.; Salzberg, M. Survival without common toxicity criteria grade 3/4 toxicity for pemetrexed compared with docetaxel in previously treated patients with advanced Non-Small Cell Lung Cancer (NSCLC): A risk-benefit analysis. J. Thorac. Oncol., 2007, 2(5), 397-401.
[17]
Lee, S.H.; Noh, K.B.; Lee, J.S.; Lee, E.J.; Min, K.H.; Hur, G.Y.; Lee, S.H.; Lee, S.Y.; Kim, J.H.; Lee, S.Y.; Shin, C.; Shim, J.J.; Kim, C.H.; Kang, K.H. In, K.H. Thymidylate synthase and ERCC1 as predictive markers in patients with pulmonary adenocarcinoma treated with pemetrexed and cisplatin. Lung Cancer, 2013, 81(1), 102-108.
[18]
Wang, T.; Chuan, P.C.; Rui, Y.J.; Long, Y.; Hong, C.X.; De, Y.X.; Qiong, H.L.; Li, L.L. Association between TYMS expression and efficacy of pemetrexed-based chemotherapy in advanced non-small cell lung cancer: A meta-analysis. PLoS One, 2013, 8(9), e74284.
[19]
Chamizo, C.; Zazo, S.; Domine, M.; Cristobal, I.; Garcia-Foncillas, J.; Rojo, F.; Madoz-Gurpide, J. Thymidylate synthase expression as a predictive biomarker of pemetrexed sensitivity in advanced non-small cell lung cancer. BMC Pulm. Med., 2015, 15, 132.
[20]
Ozasa, H.; Oguri, T.; Uemura, T.; Miyazaki, M.; Maeno, K.; Sato, S.; Ueda, R. Significance of thymidylate synthase for resistance to pemetrexed in lung cancer. Cancer Sci., 2010, 101(1), 161-166.
[21]
Liu, Y.; Yin, T.J.; Zhou, R.; Zhou, S.; Fan, L.; Zhang, R.G. Expression of thymidylate synthase predicts clinical outcomes of pemetrexed-containing chemotherapy for non-small-cell lung cancer: a systemic review and meta-analysis. Cancer Chemother. Pharmacol., 2013, 72(5), 1125-1132.
[22]
Zhang, D.; Ochi, N.; Takigawa, N.; Tanimoto, Y.; Chen, Y.; Ichihara, E.; Hotta, K.; Tabata, M.; Tanimoto, M.; Kiura, K. Establishment of pemetrexed-resistant non-small cell lung cancer cell lines. Cancer Lett., 2011, 309(2), 228-235.
[23]
Wood, K.; Hensing, T.; Malik, R.; Salgia, R. Prognostic and predictive value in KRAS in non-small-cell lung cancer: A review. JAMA Oncol., 2016, 2(6), 805-812.
[24]
Moran, D.M.; Trusk, P.B.; Pry, K.; Paz, K.; Sidransky, D.; Bacus, S.S. KRAS mutation status is associated with enhanced dependency on folate metabolism pathways in non-small cell lung cancer cells. Mol. Cancer Ther., 2014, 13(6), 1611-1624.
[25]
Tang, Z.H.; Jiang, X.M.; Guo, X.; Fong, C.M.; Chen, X.; Lu, J.J. Characterization of osimertinib (AZD9291)-resistant non-small cell lung cancer NCI-H1975/OSIR cell line. Oncotarget, 2016, 7(49), 81598-81610.
[26]
Jiang, X.M.; Xu, Y.L.; Huang, M.Y.; Zhang, L.L.; Su, M.X.; Chen, X.; Lu, J.J. Osimertinib (AZD9291) decreases programmed death ligand-1 in EGFR-mutated non-small cell lung cancer cells. Acta Pharmacol. Sin., 2017, 38(11), 1512-1520.
[27]
Zhang, L.L.; Xu, Y.L.; Tang, Z.H.; Xu, X.H.; Chen, X.; Li, T.; Ding, C.Y.; Huang, M.Q.; Chen, X.P.; Wang, Y.T.; Yuan, X.F.; Lu, J.J. Effects of alisol B 23-acetate on ovarian cancer cells: G1 phase cell cycle arrest, apoptosis, migration and invasion inhibition. Phytomedicine, 2016, 23(8), 800-809.
[28]
Scholzen, T.; Gerdes, J. The Ki-67 protein: From the known and the unknown. J. Cell. Physiol., 2000, 182(3), 311-322.
[29]
Munoz-Espin, D.; Serrano, M. Cellular senescence: From physiology to pathology. Nat. Rev. Mol. Cell Biol., 2014, 15(7), 482-496.
[30]
Chiu, L.Y.; Hsin, I.L.; Yang, T.Y.; Sung, W.W.; Chi, J.Y.; Chang, J.T.; Ko, J.L.; Sheu, G.T. The ERK-ZEB1 pathway mediates epithelial-mesenchymal transition in pemetrexed resistant lung cancer cells with suppression by vinca alkaloids. Oncogene, 2017, 36(2), 242-253.
[31]
Longley, D.B.; Harkin, D.P.; Johnston, P.G. 5-fluorouracil: Mechanisms of action and clinical strategies. Nat. Rev. Cancer, 2003, 3(5), 330-338.
[32]
Horwitz, S.B. Taxol (paclitaxel): Mechanisms of action. Ann. Oncol., 1994, 5(Suppl. 6), S3-S6.
[33]
Cavalcante, S.L.; Monteiro, G. Gemcitabine: Metabolism and molecular mechanisms of action, sensitivity and chemoresistance in pancreatic cancer. Eur. J. Pharmacol., 2014, 741, 8-16.
[34]
Pommier, Y.; Leo, E.; Zhang, H.; Marchand, C. DNA topoisomerases and their poisoning by anticancer and antibacterial drugs. Chem. Biol., 2010, 17(5), 421-433.
[35]
Yang, M.; Fan, W.F.; Pu, X.L.; Liu, F.Y.; Meng, L.J.; Wang, J. Significance of thymidylate synthase expression for resistance to pemetrexed in pulmonary adenocarcinoma. Oncol. Lett., 2014, 7(1), 227-232.
[36]
Del Bufalo, D.; Desideri, M.; De Luca, T.; Di Martile, M.; Gabellini, C.; Monica, V.; Busso, S.; Eramo, A.; De Maria, R.; Milella, M.; Trisciuoglio, D. Histone deacetylase inhibition synergistically enhances pemetrexed cytotoxicity through induction of apoptosis and autophagy in non-small cell lung cancer. Mol. Cancer, 2014, 13, 230.
[37]
Gordon, R.R.; Nelson, P.S. Cellular senescence and cancer chemotherapy resistance. Drug Resist. Updat., 2012, 15(1-2), 123-131.
[38]
Rebbaa, A. Targeting senescence pathways to reverse drug resistance in cancer. Cancer Lett., 2005, 219(1), 1-13.
[39]
Wang, L.; Wang, R.; Pan, Y.; Sun, Y.; Zhang, J.; Chen, H. The pemetrexed-containing treatments in the non-small cell lung cancer is -/low thymidylate synthase expression better than +/high thymidylate synthase expression: A meta-analysis. BMC Cancer, 2014, 14, 205.
[40]
Shimizu, T.; Nakagawa, Y.; Takahashi, N.; Hashimoto, S. Thymidylate synthase gene amplification predicts pemetrexed resistance in patients with advanced non-small cell lung cancer. Clin. Transl. Oncol., 2016, 18(1), 107-112.
[41]
Yoshida, T.; Okamoto, T.; Yano, T.; Takada, K.; Kohno, M.; Suda, K.; Takenoyama, M.; Oda, Y.; Maehara, Y. Molecular factors associated with pemetrexed sensitivity according to histological type in non-small cell lung cancer. Anticancer Res., 2016, 36(12), 6319-6326.
[42]
Racanelli, A.C.; Rothbart, S.B.; Heyer, C.L.; Moran, R.G. Therapeutics by cytotoxic metabolite accumulation: pemetrexed causes ZMP accumulation, AMPK activation, and mammalian target of rapamycin inhibition. Cancer Res., 2009, 69(13), 5467-5474.
[43]
Desmoulin, S.K.; Hou, Z.; Gangjee, A.; Matherly, L.H. The human proton-coupled folate transporter: Biology and therapeutic applications to cancer. Cancer Biol. Ther., 2012, 13(14), 1355-1373.
[44]
Giovannetti, E.; Zucali, P.A.; Assaraf, Y.G.; Funel, N.; Gemelli, M.; Stark, M.; Thunnissen, E.; Hou, Z.; Muller, I.B.; Struys, E.A.; Perrino, M.; Jansen, G.; Matherly, L.H.; Peters, G.J. Role of proton-coupled folate transporter in pemetrexed resistance of mesothelioma: Clinical evidence and new pharmacological tools. Ann. Oncol., 2017, 28(11), 2725-2732.
[45]
Uemura, T.; Oguri, T.; Ozasa, H.; Takakuwa, O.; Miyazaki, M.; Maeno, K.; Sato, S.; Ueda, R. ABCC11/MRP8 confers pemetrexed resistance in lung cancer. Cancer Sci., 2010, 101(11), 2404-2410.
[46]
Wang, D.S.; Patel, A.; Shukla, S.; Zhang, Y.K.; Wang, Y.J.; Kathawala, R.J.; Robey, R.W.; Zhang, L.; Yang, D.H.; Talele, T.T.; Bates, S.E.; Ambudkar, S.V.; Xu, R.H.; Chen, Z.S. Icotinib antagonizes ABCG2-mediated multidrug resistance, but not the pemetrexed resistance mediated by thymidylate synthase and ABCG2. Oncotarget, 2014, 5(12), 4529-4542.

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