Salubrinal Enhances Cancer Cell Death during Glucose Deprivation through the Upregulation of xCT and Mitochondrial Oxidative Stress
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cell Culture
2.2. Western Blot Analysis
2.3. Sulforhodamine B (SRB) Assay for Cell Growth
2.4. Clonogenic Assay
2.5. Transwell Migration Assay
2.6. Propidium Iodide (PI) Exclusion Assay
2.7. Intracellular ROS and Mitochondrial ROS Measurement
2.8. Small Interfering RNA (siRNA)-Mediated Genetic Knockdown
2.9. Intracellular Glutamate Measurement
2.10. Statistical Analysis
3. Results
3.1. Salubrinal Activated ISR and Inhibited Cell Growth and Migration
3.2. Salubrinal Increased Cell Death Rate and ROS Levels in Cancer Cells under Glucose Deprivation
3.3. ROS Were Involved in Salubrinal-Enhanced Cell Death during Glucose Deprivation
3.4. Salubrinal Promoted Cell Death during Glucose Deprivation through the Upregulation of xCT
3.5. Dm-αKG Rescued the Salubrinal-Enhanced Cell Death during Glucose Deprivation
4. Discussion
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Acknowledgments
Conflicts of Interest
References
- Hirayama, A.; Kami, K.; Sugimoto, M.; Sugawara, M.; Toki, N.; Onozuka, H.; Kinoshita, T.; Saito, N.; Ochiai, A.; Tomita, M.; et al. Quantitative metabolome profiling of colon and stomach cancer microenvironment by capillary electrophoresis time-of-flight mass spectrometry. Cancer Res. 2009, 69, 4918–4925. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Martin, J.D.; Fukumura, D.; Duda, D.G.; Boucher, Y.; Jain, R.K. Reengineering the Tumor Microenvironment to Alleviate Hypoxia and Overcome Cancer Heterogeneity. Cold Spring Harb. Perspect. Med. 2016, 6, a027094. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Grasmann, G.; Mondal, A.; Leithner, K. Flexibility and Adaptation of Cancer Cells in a Heterogenous Metabolic Microenvironment. Int. J. Mol. Sci. 2021, 22, 1476. [Google Scholar] [CrossRef]
- Balachandran, S.; Roberts, P.C.; Brown, L.E.; Truong, H.; Pattnaik, A.K.; Archer, D.R.; Barber, G.N. Essential role for the dsRNA-dependent protein kinase PKR in innate immunity to viral infection. Immunity 2000, 13, 129–141. [Google Scholar] [CrossRef] [Green Version]
- Han, A.P.; Yu, C.; Lu, L.; Fujiwara, Y.; Browne, C.; Chin, G.; Fleming, M.; Leboulch, P.; Orkin, S.H.; Chen, J.J. Heme-regulated eIF2alpha kinase (HRI) is required for translational regulation and survival of erythroid precursors in iron deficiency. EMBO J. 2001, 20, 6909–6918. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Harding, H.P.; Zhang, Y.; Bertolotti, A.; Zeng, H.; Ron, D. Perk is essential for translational regulation and cell survival during the unfolded protein response. Mol. Cell 2000, 5, 897–904. [Google Scholar] [CrossRef]
- Harding, H.P.; Zhang, Y.; Zeng, H.; Novoa, I.; Lu, P.D.; Calfon, M.; Sadri, N.; Yun, C.; Popko, B.; Paules, R.; et al. An integrated stress response regulates amino acid metabolism and resistance to oxidative stress. Mol. Cell 2003, 11, 619–633. [Google Scholar] [CrossRef]
- Pakos-Zebrucka, K.; Koryga, I.; Mnich, K.; Ljujic, M.; Samali, A.; Gorman, A.M. The integrated stress response. EMBO Rep. 2016, 17, 1374–1395. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Wang, S.F.; Chen, S.; Tseng, L.M.; Lee, H.C. Role of the mitochondrial stress response in human cancer progression. Exp. Biol. Med. (Maywood) 2020, 245, 861–878. [Google Scholar] [CrossRef] [PubMed]
- Quirós, P.M.; Prado, M.A.; Zamboni, N.; D’Amico, D.; Williams, R.W.; Finley, D.; Gygi, S.P.; Auwerx, J. Multi-omics analysis identifies ATF4 as a key regulator of the mitochondrial stress response in mammals. J. Cell Biol. 2017, 216, 2027–2045. [Google Scholar] [CrossRef]
- Pavlova, N.N.; Thompson, C.B. The Emerging Hallmarks of Cancer Metabolism. Cell Metab. 2016, 23, 27–47. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Wang, S.F.; Chen, M.S.; Chou, Y.C.; Ueng, Y.F.; Yin, P.H.; Yeh, T.S.; Lee, H.C. Mitochondrial dysfunction enhances cisplatin resistance in human gastric cancer cells via the ROS-activated GCN2-eIF2α-ATF4-xCT pathway. Oncotarget 2016, 7, 74132–74151. [Google Scholar] [CrossRef] [Green Version]
- Ye, P.; Mimura, J.; Okada, T.; Sato, H.; Liu, T.; Maruyama, A.; Ohyama, C.; Itoh, K. Nrf2- and ATF4-dependent upregulation of xCT modulates the sensitivity of T24 bladder carcinoma cells to proteasome inhibition. Mol. Cell Biol. 2014, 34, 3421–3434. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Yang, Y.; Yee, D. IGF-1 regulates redox status in breast cancer by activating the amino acid transport molecule xC-. Cancer Res. 2014, 74, 2295–2305. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- DeBerardinis, R.J.; Chandel, N.S. Fundamentals of Cancer Metabolism. Sci. Adv. 2016, 2, e1600200. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Cluntun, A.A.; Lukey, M.J.; Cerione, R.A.; Locasale, J.W. Glutamine Metabolism in Cancer: Understanding the Heterogeneity. Trends Cancer 2017, 3, 169–180. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Koppula, P.; Zhang, Y.; Shi, J.; Li, W.; Gan, B. The glutamate/cystine antiporter SLC7A11/xCT enhances cancer cell dependency on glucose by exporting glutamate. J. Biol. Chem. 2017, 292, 14240–14249. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Chen, M.C.; Hsu, L.L.; Wang, S.F.; Hsu, C.Y.; Lee, H.C.; Tseng, L.M. ROS Mediate xCT-Dependent Cell Death in Human Breast Cancer Cells under Glucose Deprivation. Cells 2020, 9, 1598. [Google Scholar] [CrossRef]
- Boyce, M.; Bryant, K.F.; Jousse, C.; Long, K.; Harding, H.P.; Scheuner, D.; Kaufman, R.J.; Ma, D.; Coen, D.M.; Ron, D.; et al. A selective inhibitor of eIF2alpha dephosphorylation protects cells from ER stress. Science 2005, 307, 935–939. [Google Scholar] [CrossRef]
- Ye, J.; Kumanova, M.; Hart, L.S.; Sloane, K.; Zhang, H.; De Panis, D.N.; Bobrovnikova-Marjon, E.; Diehl, J.A.; Ron, D.; Koumenis, C. The GCN2-ATF4 pathway is critical for tumour cell survival and proliferation in response to nutrient deprivation. EMBO J. 2010, 29, 2082–2096. [Google Scholar] [CrossRef] [Green Version]
- Bi, M.; Naczki, C.; Koritzinsky, M.; Fels, D.; Blais, J.; Hu, N.; Harding, H.; Novoa, I.; Varia, M.; Raleigh, J.; et al. ER stress-regulated translation increases tolerance to extreme hypoxia and promotes tumor growth. EMBO J. 2005, 24, 3470–3481. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Feng, Y.X.; Sokol, E.S.; Del Vecchio, C.A.; Sanduja, S.; Claessen, J.H.; Proia, T.A.; Jin, D.X.; Reinhardt, F.; Ploegh, H.L.; Wang, Q.; et al. Epithelial-to-mesenchymal transition activates PERK-eIF2α and sensitizes cells to endoplasmic reticulum stress. Cancer Discov. 2014, 4, 702–715. [Google Scholar] [CrossRef] [Green Version]
- Huber, A.L.; Lebeau, J.; Guillaumot, P.; Pétrilli, V.; Malek, M.; Chilloux, J.; Fauvet, F.; Payen, L.; Kfoury, A.; Renno, T.; et al. p58(IPK)-mediated attenuation of the proapoptotic PERK-CHOP pathway allows malignant progression upon low glucose. Mol. Cell 2013, 49, 1049–1059. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Ranganathan, A.C.; Ojha, S.; Kourtidis, A.; Conklin, D.S.; Aguirre-Ghiso, J.A. Dual function of pancreatic endoplasmic reticulum kinase in tumor cell growth arrest and survival. Cancer Res. 2008, 68, 3260–3268. [Google Scholar] [CrossRef] [Green Version]
- Stone, S.; Ho, Y.; Li, X.; Jamison, S.; Harding, H.P.; Ron, D.; Lin, W. Dual role of the integrated stress response in medulloblastoma tumorigenesis. Oncotarget 2016, 7, 64124–64135. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Chen, Y.S.; Huang, W.L.; Chang, S.H.; Chang, K.W.; Kao, S.Y.; Lo, J.F.; Su, P.F. Enhanced filopodium formation and stem-like phenotypes in a novel metastatic head and neck cancer cell model. Oncol. Rep. 2013, 30, 2829–2837. [Google Scholar] [CrossRef]
- Wang, S.F.; Wung, C.H.; Chen, M.S.; Chen, C.F.; Yin, P.H.; Yeh, T.S.; Chang, Y.L.; Chou, Y.C.; Hung, H.H.; Lee, H.C. Activated integrated stress response induced by salubrinal promotes cisplatin resistance in human gastric cancer cells via enhanced xCT expression and glutathione biosynthesis. Int. J. Mol. Sci. 2018, 19, 3389. [Google Scholar] [CrossRef] [Green Version]
- Lo, M.; Wang, Y.Z.; Gout, P.W. The x(c)- cystine/glutamate antiporter: A potential target for therapy of cancer and other diseases. J. Cell Physiol. 2008, 215, 593–602. [Google Scholar] [CrossRef] [PubMed]
- Gout, P.W.; Buckley, A.R.; Simms, C.R.; Bruchovsky, N. Sulfasalazine, a potent suppressor of lymphoma growth by inhibition of the x(c)- cystine transporter: A new action for an old drug. Leukemia 2001, 15, 1633–1640. [Google Scholar] [CrossRef] [Green Version]
- Ron, D. Translational control in the endoplasmic reticulum stress response. J. Clin. Investig. 2002, 110, 1383–1388. [Google Scholar] [CrossRef] [PubMed]
- Brewer, J.W.; Diehl, J.A. PERK mediates cell-cycle exit during the mammalian unfolded protein response. Proc. Natl. Acad. Sci. USA 2000, 97, 12625–12630. [Google Scholar] [CrossRef] [Green Version]
- Brewer, J.W.; Hendershot, L.M.; Sherr, C.J.; Diehl, J.A. Mammalian unfolded protein response inhibits cyclin D1 translation and cell-cycle progression. Proc. Natl. Acad. Sci. USA 1999, 96, 8505–8510. [Google Scholar] [CrossRef] [Green Version]
- Kami, K.; Fujimori, T.; Sato, H.; Sato, M.; Yamamoto, H.; Ohashi, Y.; Sugiyama, N.; Ishihama, Y.; Onozuka, H.; Ochiai, A.; et al. Metabolomic profiling of lung and prostate tumor tissues by capillary electrophoresis time-of-flight mass spectrometry. Metabolomics 2013, 9, 444–453. [Google Scholar] [CrossRef] [Green Version]
- Pelicano, H.; Zhang, W.; Liu, J.; Hammoudi, N.; Dai, J.; Xu, R.H.; Pusztai, L.; Huang, P. Mitochondrial dysfunction in some triple-negative breast cancer cell lines: Role of mTOR pathway and therapeutic potential. Breast Cancer Res. 2014, 16, 434. [Google Scholar] [CrossRef] [Green Version]
- Raut, G.K.; Chakrabarti, M.; Pamarthy, D.; Bhadra, M.P. Glucose starvation-induced oxidative stress causes mitochondrial dysfunction and apoptosis via Prohibitin 1 upregulation in human breast cancer cells. Free Radic. Biol. Med. 2019, 145, 428–441. [Google Scholar] [CrossRef]
- Shutt, D.C.; O’Dorisio, M.S.; Aykin-Burns, N.; Spitz, D.R. 2-deoxy-D-glucose induces oxidative stress and cell killing in human neuroblastoma cells. Cancer Biol. Ther. 2010, 9, 853–861. [Google Scholar] [CrossRef] [Green Version]
- Xi, H.; Kurtoglu, M.; Liu, H.; Wangpaichitr, M.; You, M.; Liu, X.; Savaraj, N.; Lampidis, T.J. 2-Deoxy-D-glucose activates autophagy via endoplasmic reticulum stress rather than ATP depletion. Cancer Chemother. Pharmacol. 2011, 67, 899–910. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Liou, G.Y.; Storz, P. Reactive oxygen species in cancer. Free Radic. Res. 2010, 44, 479–496. [Google Scholar] [CrossRef] [Green Version]
- Lee, J.I.; Dominy, J.E., Jr.; Sikalidis, A.K.; Hirschberger, L.L.; Wang, W.; Stipanuk, M.H. HepG2/C3A cells respond to cysteine deprivation by induction of the amino acid deprivation/integrated stress response pathway. Physiol. Genomics 2008, 33, 218–229. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Birsoy, K.; Possemato, R.; Lorbeer, F.K.; Bayraktar, E.C.; Thiru, P.; Yucel, B.; Wang, T.; Chen, W.W.; Clish, C.B.; Sabatini, D.M. Metabolic determinants of cancer cell sensitivity to glucose limitation and biguanides. Nature 2014, 508, 108–112. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- TeSlaa, T.; Bartman, C.R.; Jankowski, C.S.R.; Zhang, Z.; Xu, X.; Xing, X.; Wang, L.; Lu, W.; Hui, S.; Rabinowitz, J.D. The Source of Glycolytic Intermediates in Mammalian Tissues. Cell Metab. 2021, 33, 367–378. [Google Scholar] [CrossRef]
- Abdel-Wahab, A.F.; Mahmoud, W.; Al-Harizy, R.M. Targeting glucose metabolism to suppress cancer progression: Prospective of anti-glycolytic cancer therapy. Pharmacol. Res. 2019, 150, 104511. [Google Scholar] [CrossRef]
- Vijayaraghavan, R.; Kumar, D.; Dube, S.N.; Singh, R.; Pandey, K.S.; Bag, B.C.; Kaushik, M.P.; Sekhar, K.; Dwarakanath, B.S.; Ravindranath, T. Acute toxicity and cardio-respiratory effects of 2-deoxy-D-glucose: A promising radio sensitiser. Biomed. Environ. Sci. 2006, 19, 96–103. [Google Scholar]
- Samokhina, E.; Popova, I.; Malkov, A.; Ivanov, A.I.; Papadia, D.; Osypov, A.; Molchanov, M.; Paskevich, S.; Fisahn, A.; Zilberter, M.; et al. Chronic inhibition of brain glycolysis initiates epileptogenesis. J. Neurosci. Res. 2017, 95, 2195–2206. [Google Scholar] [CrossRef] [Green Version]
- Simons, A.L.; Ahmad, I.M.; Mattson, D.M.; Dornfeld, K.J.; Spitz, D.R. 2-Deoxy-D-glucose combined with cisplatin enhances cytotoxicity via metabolic oxidative stress in human head and neck cancer cells. Cancer Res. 2007, 67, 3364–3370. [Google Scholar] [CrossRef] [Green Version]
- Raez, L.E.; Papadopoulos, K.; Ricart, A.D.; Chiorean, E.G.; Dipaola, R.S.; Stein, M.N.; Rocha Lima, C.M.; Schlesselman, J.J.; Tolba, K.; Langmuir, V.K.; et al. A phase I dose-escalation trial of 2-deoxy-D-glucose alone or combined with docetaxel in patients with advanced solid tumors. Cancer Chemother. Pharmacol. 2013, 71, 523–530. [Google Scholar] [CrossRef]
- Venkataramanaa, N.K.; Venkatesh, P.K.; Dwarakanath, B.S.; Vani, S. Protective effect on normal brain tissue during a combinational therapy of 2-deoxy-d-glucose and hypofractionated irradiation in malignant gliomas. Asian J. Neurosurg. 2013, 8, 9–14. [Google Scholar] [CrossRef] [Green Version]
- Cheng, G.; Zielonka, J.; Dranka, B.P.; McAllister, D.; Mackinnon, A.C., Jr.; Joseph, J.; Kalyanaraman, B. Mitochondria-targeted drugs synergize with 2-deoxyglucose to trigger breast cancer cell death. Cancer Res. 2012, 72, 2634–2644. [Google Scholar] [CrossRef] [Green Version]
- Alsterda, A.; Asha, K.; Powrozek, O.; Repak, M.; Goswami, S.; Dunn, A.M.; Memmel, H.C.; Sharma-Walia, N. Salubrinal Exposes Anticancer Properties in Inflammatory Breast Cancer Cells by Manipulating the Endoplasmic Reticulum Stress Pathway. Front. Oncol. 2021, 20, 654940. [Google Scholar] [CrossRef]
- Kardos, G.R.; Gowda, R.; Dinavahi, S.S.; Kimball, S.; Robertson, G.P. Salubrinal in Combination With 4E1RCat Synergistically Impairs Melanoma Development by Disrupting the Protein Synthetic Machinery. Front. Oncol. 2020, 10, 834. [Google Scholar] [CrossRef]
- Bastola, P.; Neums, L.; Schoenen, F.J.; Chien, J. VCP inhibitors induce endoplasmic reticulum stress, cause cell cycle arrest, trigger caspase-mediated cell death and synergistically kill ovarian cancer cells in combination with Salubrinal. Mol. Oncol. 2016, 10, 1559–1574. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Drexler, H.C. Synergistic apoptosis induction in leukemic cells by the phosphatase inhibitor salubrinal and proteasome inhibitors. PLoS ONE 2009, 4, e4161. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Jeon, Y.J.; Kim, J.H.; Shin, J.I.; Jeong, M.; Cho, J.; Lee, K. Salubrinal-Mediated Upregulation of eIF2α Phosphorylation Increases Doxorubicin Sensitivity in MCF-7/ADR Cells. Mol. Cells 2016, 39, 129–135. [Google Scholar] [CrossRef] [Green Version]
- Zhao, X.; Zhang, C.; Zhou, H.; Xiao, B.; Cheng, Y.; Wang, J.; Yao, F.; Duan, C.; Chen, R.; Liu, Y.; et al. Synergistic antitumor activity of the combination of salubrinal and rapamycin against human cholangiocarcinoma cells. Oncotarget 2016, 7, 85492–85501. [Google Scholar] [CrossRef] [Green Version]
- Bunpo, P.; Dudley, A.; Cundiff, J.K.; Cavener, D.R.; Wek, R.C.; Anthony, T.G. GCN2 protein kinase is required to activate amino acid deprivation responses in mice treated with the anti-cancer agent L-asparaginase. J. Biol. Chem. 2009, 284, 32742–32749. [Google Scholar] [CrossRef] [Green Version]
- Long, Y.; Tsai, W.B.; Wangpaichitr, M.; Tsukamoto, T.; Savaraj, N.; Feun, L.G.; Kuo, M.T. Arginine deiminase resistance in melanoma cells is associated with metabolic reprogramming, glucose dependence, and glutamine addiction. Mol. Cancer Ther. 2013, 12, 2581–2590. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Sundrud, M.S.; Koralov, S.B.; Feuerer, M.; Calado, D.P.; Kozhaya, A.E.; Rhule-Smith, A.; Lefebvre, R.E.; Unutmaz, D.; Mazitschek, R.; Waldner, H.; et al. Halofuginone inhibits TH17 cell differentiation by activating the amino acid starvation response. Science 2009, 324, 1334–1338. [Google Scholar] [CrossRef] [Green Version]
- Chen, G.Q.; Gong, R.H.; Yang, D.J.; Zhang, G.; Lu, A.P.; Yan, S.C.; Lin, S.H.; Bian, Z.X. Halofuginone dually regulates autophagic flux through nutrient-sensing pathways in colorectal cancer. Cell Death Dis. 2017, 8, e2789. [Google Scholar] [CrossRef]
- De Gassart, A.; Bujisic, B.; Zaffalon, L.; Decosterd, L.A.; Di Micco, A.; Frera, G.; Tallant, R.; Martinon, F. An inhibitor of HIV-1 protease modulates constitutive eIF2α dephosphorylation to trigger a specific integrated stress response. Proc. Natl. Acad. Sci. USA 2016, 113, E117–E126. [Google Scholar] [CrossRef] [Green Version]
- Sato, A.; Asano, T.; Okubo, K.; Isono, M.; Asano, T. Nelfinavir and Ritonavir Kill Bladder Cancer Cells Synergistically by Inducing Endoplasmic Reticulum Stress. Oncol. Res. 2018, 26, 323–332. [Google Scholar] [CrossRef]
- Xia, C.; He, Z.; Liang, S.; Chen, R.; Xu, W.; Yang, J.; Xiao, G.; Jiang, S. Metformin combined with nelfinavir induces SIRT3/mROS-dependent autophagy in human cervical cancer cells and xenograft in nude mice. Eur. J. Pharmacol. 2019, 848, 62–69. [Google Scholar] [CrossRef] [PubMed]
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. |
© 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Chen, M.-C.; Hsu, L.-L.; Wang, S.-F.; Pan, Y.-L.; Lo, J.-F.; Yeh, T.-S.; Tseng, L.-M.; Lee, H.-C. Salubrinal Enhances Cancer Cell Death during Glucose Deprivation through the Upregulation of xCT and Mitochondrial Oxidative Stress. Biomedicines 2021, 9, 1101. https://doi.org/10.3390/biomedicines9091101
Chen M-C, Hsu L-L, Wang S-F, Pan Y-L, Lo J-F, Yeh T-S, Tseng L-M, Lee H-C. Salubrinal Enhances Cancer Cell Death during Glucose Deprivation through the Upregulation of xCT and Mitochondrial Oxidative Stress. Biomedicines. 2021; 9(9):1101. https://doi.org/10.3390/biomedicines9091101
Chicago/Turabian StyleChen, Mei-Chun, Li-Lin Hsu, Sheng-Fan Wang, Yi-Ling Pan, Jeng-Fan Lo, Tien-Shun Yeh, Ling-Ming Tseng, and Hsin-Chen Lee. 2021. "Salubrinal Enhances Cancer Cell Death during Glucose Deprivation through the Upregulation of xCT and Mitochondrial Oxidative Stress" Biomedicines 9, no. 9: 1101. https://doi.org/10.3390/biomedicines9091101