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Medical Oncology

, 34:25 | Cite as

Inhibiting autophagy with chloroquine enhances the anti-tumor effect of high-LET carbon ions via ER stress-related apoptosis

  • Xiaogang Zheng
  • Xiaodong Jin
  • Feifei Li
  • Xiongxiong Liu
  • Yan Liu
  • Fei Ye
  • Ping Li
  • Ting Zhao
  • Qiang LiEmail author
Original Paper

Abstract

Energetic carbon ions (CI) offer great advantages over conventional radiations such as X- or γ-rays in cancer radiotherapy. High linear energy transfer (LET) CI can induce both endoplasmic reticulum (ER) stress and autophagy in tumor cells under certain circumstances. The molecular connection between ER stress and autophagy in tumor exposed to high-LET radiation and how these two pathways influence the therapeutic effect against tumor remain poorly understood. In this work, we studied the impact of autophagy and apoptosis induced by ER stress following high-LET CI radiation on the radiosensitivity of S180 cells both in vitro and in vivo. In the in vitro experiment, X-rays were also used as a reference radiation. Our results documented that the combination of CI radiation with chloroquine (CQ), a special autophagy inhibitor, produced more pronounced proliferation suppression in S180 cells and xenograft tumors. Co-treatment with CI radiation and CQ could block autophagy through the IRE1/JNK/Beclin-1 axis and enhance apoptotic cell death via the activation of C/EBP homologous protein (CHOP) by the IRE1 pathway rather than PERK in vitro and in vivo. Thus, our study indicates that inhibiting autophagy might be a promising therapeutic strategy in CI radiotherapy via aggravating the ER stress-related apoptosis.

Keywords

High-LET radiation Autophagy Chloroquine ER stress Apoptosis CHOP 

Notes

Acknowledgements

This work was jointly supported by the Key Project of the National Natural Science Foundation of China (Grant No. U1232207), the National Key Technology Support Program of the Ministry of Science and Technology of China (Grant No. 2015BAI01B11), the National Key Research and Development Program of the Ministry of Science and Technology of China (Grant No. 2016YFC0904602) and the National Natural Science Foundation of China (Grant No. 10905080, 11075191 and 11205217).

Author’s contribution

QL designed and supervised the study. XZ, XJ and FL performed the experiments. XZ wrote the manuscript. XL and YL analyzed the data. FY, PL and TZ contributed to the data analysis and discussion. XZ and FL performed the irradiation. All authors read and approved the final manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical standards

All procedures performed in studies involving animals were in accordance with the ethical standards of the Animal Care Committee and with the approval of the Academic Committee at Institute of Modern Physics.

Supplementary material

12032_2017_883_MOESM1_ESM.pdf (1.9 mb)
Supplementary material 1 (PDF 1905 kb)

References

  1. 1.
    Hamada N, Imaoka T, Masunaga S, Ogata T, Okayasu R, Takahashi A, et al. Recent advances in the biology of heavy-ion cancer therapy. J Radiat Res. 2010;51(4):365–83.CrossRefPubMedGoogle Scholar
  2. 2.
    Zhang B, Wang Y, Pang X, Su Y, Ai G, Wang T. ER stress induced by ionising radiation in IEC-6 cells. Int J Radiat Biol. 2010;86(6):429–35. doi: 10.3109/09553001003668014.CrossRefPubMedGoogle Scholar
  3. 3.
    Anscher MS, Vujaskovic Z. Mechanisms and potential targets for prevention and treatment of normal tissue injury after radiation therapy. Semin Oncol. 2005;32(2 Suppl 3):S86–91.CrossRefPubMedGoogle Scholar
  4. 4.
    Verfaillie T, Salazar M, Velasco G, Agostinis P. Linking ER stress to autophagy: potential implications for cancer therapy. Int J Cell Biol. 2010;2010:930509. doi: 10.1155/2010/930509.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Senft D, Ronai ZA. UPR, autophagy, and mitochondria crosstalk underlies the ER stress response. Trends Biochem Sci. 2015;40(3):141–8. doi: 10.1016/j.tibs.2015.01.002.CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Hetz C. The unfolded protein response: controlling cell fate decisions under ER stress and beyond. Nat Rev Mol Cell Biol. 2012;13(2):89–102. doi: 10.1038/nrm3270.PubMedGoogle Scholar
  7. 7.
    Wang WA, Groenendyk J, Michalak M. Endoplasmic reticulum stress associated responses in cancer. Biochim Biophys Acta. 2014;1843(10):2143–9. doi: 10.1016/j.bbamcr.2014.01.012.CrossRefPubMedGoogle Scholar
  8. 8.
    Zhang G, Liu K, Ling X, Wang Z, Zou P, Wang X, et al. DBP-induced endoplasmic reticulum stress in male germ cells causes autophagy, which has a cytoprotective role against apoptosis in vitro and in vivo. Toxicol Lett. 2016;245:86–98. doi: 10.1016/j.toxlet.2016.01.016.CrossRefPubMedGoogle Scholar
  9. 9.
    Bohnert KR, Gallot YS, Sato S, Xiong G, Hindi SM, Kumar A. Inhibition of ER stress and unfolding protein response pathways causes skeletal muscle wasting during cancer cachexia. Faseb J. 2016. doi: 10.1096/fj.201600250RR.PubMedGoogle Scholar
  10. 10.
    Levine B, Kroemer G. SnapShot: macroautophagy. Cell. 2008;132(1):162–U14.CrossRefGoogle Scholar
  11. 11.
    Klionsky DJ, Abdelmohsen K, Abe A, Abedin MJ, Abeliovich H, Acevedo Arozena A, et al. Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition). Autophagy. 2016;12(1):1–222. doi: 10.1080/15548627.2015.1100356.CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Roy S, Debnath J. Autophagy and tumorigenesis. Semin Immunopathol. 2010;32(4):383–96. doi: 10.1007/s00281-010-0213-0.CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Zois CE, Koukourakis MI. Radiation-induced autophagy in normal and cancer cells: towards novel cytoprotection and radio-sensitization policies? Autophagy. 2009;5(4):442–50.CrossRefPubMedGoogle Scholar
  14. 14.
    Bristol ML, Di X, Beckman MJ, Wilson EN, Henderson SC, Maiti A, et al. Dual functions of autophagy in the response of breast tumor cells to radiation: cytoprotective autophagy with radiation alone and cytotoxic autophagy in radiosensitization by vitamin D 3. Autophagy. 2012;8(5):739–53. doi: 10.4161/auto.19313.CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Moretti L, Cha YI, Niermann KJ, Lu B. Switch between apoptosis and autophagy: radiation-induced endoplasmic reticulum stress? Cell Cycle. 2007;6(7):793–8.CrossRefPubMedGoogle Scholar
  16. 16.
    Choi JY, Hong WG, Cho JH, Kim EM, Kim J, Jung CH, et al. Podophyllotoxin acetate triggers anticancer effects against non-small cell lung cancer cells by promoting cell death via cell cycle arrest, ER stress and autophagy. Int J Oncol. 2015;47(4):1257–65. doi: 10.3892/ijo.2015.3123.PubMedPubMedCentralGoogle Scholar
  17. 17.
    Rouschop KM, van den Beucken T, Dubois L, Niessen H, Bussink J, Savelkouls K, et al. The unfolded protein response protects human tumor cells during hypoxia through regulation of the autophagy genes MAP1LC3B and ATG5. J Clin Investig. 2010;120(1):127–41. doi: 10.1172/JCI40027.CrossRefPubMedGoogle Scholar
  18. 18.
    Jin X, Li F, Zheng X, Liu Y, Hirayama R, Liu X, et al. Carbon ions induce autophagy effectively through stimulating the unfolded protein response and subsequent inhibiting Akt phosphorylation in tumor cells. Sci Rep. 2015;5:13815. doi: 10.1038/srep13815.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Lv S, Sun E-C, Xu Q-Y, Zhang J-K, Wu D-L. Endoplasmic reticulum stress-mediated autophagy contributes to bluetongue virus infection via the PERK-eIF2α pathway. Biochem Biophys Res Commun. 2015;466(3):406–12. doi: 10.1016/j.bbrc.2015.09.039.CrossRefPubMedGoogle Scholar
  20. 20.
    Shimodaira Y, Takahashi S, Kinouchi Y, Endo K, Shiga H, Kakuta Y, et al. Modulation of endoplasmic reticulum (ER) stress-induced autophagy by C/EBP homologous protein (CHOP) and inositol-requiring enzyme 1α (IRE1α) in human colon cancer cells. Biochem Biophys Res Commun. 2014;445(2):524–33. doi: 10.1016/j.bbrc.2014.02.054.CrossRefPubMedGoogle Scholar
  21. 21.
    Panganiban RAM, Mungunsukh O, Day RM. X-irradiation induces ER stress, apoptosis, and senescence in pulmonary artery endothelial cells. Int J Radiat Biol. 2013;89(8):656–67. doi: 10.3109/09553002.2012.711502.CrossRefPubMedGoogle Scholar
  22. 22.
    Chiu HW, Fang WH, Chen YL, Wu MD, Yuan GF, Ho SY, et al. Monascuspiloin enhances the radiation sensitivity of human prostate cancer cells by stimulating endoplasmic reticulum stress and inducing autophagy. PLoS ONE. 2012;7(7):e40462. doi: 10.1371/journal.pone.0040462.CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Lee ES, Lee HJ, Lee YJ, Jeong JH, Kang S, Lim YB. Chemical chaperones reduce ionizing radiation-induced endoplasmic reticulum stress and cell death in IEC-6 cells. Biochem Biophys Res Commun. 2014;450(2):1005–9. doi: 10.1016/j.bbrc.2014.06.091.CrossRefPubMedGoogle Scholar
  24. 24.
    Wang ZC, Wang JF, Li YB, Guo CX, Liu Y, Fang F, et al. Involvement of endoplasmic reticulum stress in apoptosis of testicular cells induced by low-dose radiation. J Huazhong Univ Sci Technol Med Sci = Hua zhong ke ji da xue xue bao Yi xue Ying De wen ban = Huazhong keji daxue xuebao Yixue Yingdewen ban. 2013;33(4):551–8. doi: 10.1007/s11596-013-1157-0.CrossRefPubMedGoogle Scholar
  25. 25.
    Pang XL, He G, Liu YB, Wang Y, Zhang B. Endoplasmic reticulum stress sensitizes human esophageal cancer cell to radiation. World J Gastroenterol WJG. 2013;19(11):1736–48. doi: 10.3748/wjg.v19.i11.1736.CrossRefPubMedGoogle Scholar
  26. 26.
    Li F, Zheng X, Liu Y, Li P, Liu X, Ye F, et al. Different roles of CHOP and JNK in mediating radiation-induced autophagy and apoptosis in breast cancer cells. Radiat Res. 2016;185(5):539–48. doi: 10.1667/RR14344.1.CrossRefPubMedGoogle Scholar
  27. 27.
    Kim KW, Moretti L, Mitchell LR, Jung DK, Lu B. Endoplasmic reticulum stress mediates radiation-induced autophagy by perk-eIF2alpha in caspase-3/7-deficient cells. Oncogene. 2010;29(22):3241–51. doi: 10.1038/onc.2010.74.CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Harhaji-Trajkovic L, Arsikin K, Kravic-Stevovic T, Petricevic S, Tovilovic G, Pantovic A, et al. Chloroquine-mediated lysosomal dysfunction enhances the anticancer effect of nutrient deprivation. Pharm Res. 2012;29(8):2249–63. doi: 10.1007/s11095-012-0753-1.CrossRefPubMedGoogle Scholar
  29. 29.
    Kazuhito S, Tsuno NH, Sunami E. Resistance of colon cancer to 5-fluorouracil may be overcome by combination with chloroquine, an in vivo study. Anti-Cancer Drugs. 2012. doi: 10.1097/CAD.0b013e328353f8c7.Google Scholar
  30. 30.
    Jin X, Li Q, Wu Q, Li P, Matsumoto Y, Furusawa Y, et al. Radiosensitization by inhibiting survivin in human hepatoma HepG2 cells to high-LET radiation. J Radiat Res. 2011;52(3):335–41. doi: 10.1269/jrr.10134.CrossRefPubMedGoogle Scholar
  31. 31.
    Jiang P, Mizushima N. LC3- and p62-based biochemical methods for the analysis of autophagy progression in mammalian cells. Methods. 2015;75:13–8. doi: 10.1016/j.ymeth.2014.11.021.CrossRefPubMedGoogle Scholar
  32. 32.
    Salazar M, Carracedo A, Salanueva IJ, Hernandez-Tiedra S, Lorente M, Egia A, et al. Cannabinoid action induces autophagy-mediated cell death through stimulation of ER stress in human glioma cells. J Clin Investig. 2009;119(5):1359–72. doi: 10.1172/JCI37948.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Park MA, Yacoub A, Sarkar D, Emdad L, Rahmani M, Spiegel S, et al. PERK-dependent regulation of MDA-7/IL-24-induced autophagy in primary human glioma cells. Autophagy. 2008;4(4):513–5.CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Luo B, Lee AS. The critical roles of endoplasmic reticulum chaperones and unfolded protein response in tumorigenesis and anticancer therapies. Oncogene. 2013;32(7):805–18. doi: 10.1038/onc.2012.130.CrossRefPubMedGoogle Scholar
  35. 35.
    Kouroku Y, Fujita E, Tanida I, Ueno T, Isoai A, Kumagai H, et al. ER stress (PERK/eIF2alpha phosphorylation) mediates the polyglutamine-induced LC3 conversion, an essential step for autophagy formation. Cell Death Differ. 2007;14(2):230–9. doi: 10.1038/sj.cdd.4401984.CrossRefPubMedGoogle Scholar
  36. 36.
    Sano R, Reed JC. ER stress-induced cell death mechanisms. BBA-Mol Cell Res. 2013;1833(12):3460–70. doi: 10.1016/j.bbamcr.2013.06.028.Google Scholar
  37. 37.
    Shi YH, Ding ZB, Zhou J, Hui B, Shi GM, Ke AW, et al. Targeting autophagy enhances sorafenib lethality for hepatocellular carcinoma via ER stress-related apoptosis. Autophagy. 2011;7(10):1159–72. doi: 10.4161/auto.7.10.16818.CrossRefPubMedGoogle Scholar
  38. 38.
    Agostinis P. Endoplasmic reticulum stress in health and disease. 1st ed. New York: Springer; 2012.CrossRefGoogle Scholar
  39. 39.
    Liu F, Shang Y, Chen SZ. Chloroquine potentiates the anti-cancer effect of lidamycin on non-small cell lung cancer cells in vitro. Acta Pharmacol Sin. 2014;35(5):645–52. doi: 10.1038/aps.2014.3.CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Gulow K, Bienert D, Haas IG. BiP is feed-back regulated by control of protein translation efficiency. J Cell Sci. 2002;115(11):2443–52.PubMedGoogle Scholar
  41. 41.
    Pattingre S, Tassa A, Qu X, Garuti R, Liang XH, Mizushima N, et al. Bcl-2 antiapoptotic proteins inhibit Beclin 1-dependent autophagy. Cell. 2005;122(6):927–39. doi: 10.1016/j.cell.2005.07.002.CrossRefPubMedGoogle Scholar
  42. 42.
    Xu Z, Bu Y, Chitnis N, Koumenis C, Fuchs SY, Diehl JA. miR-216b regulation of c-Jun mediates GADD153/CHOP-dependent apoptosis. Nat Commun. 2016;7:11422. doi: 10.1038/ncomms11422.CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Mori E, Takahashi A, Yamakawa N, Kirita T, Ohnishi T. High LET heavy ion radiation induces p53-independent apoptosis. J Radiat Res. 2009;50(1):37–42.CrossRefPubMedGoogle Scholar
  44. 44.
    Amornwichet N, Oike T, Shibata A, Ogiwara H, Tsuchiya N, Yamauchi M, et al. Carbon-ion beam irradiation kills X-ray-resistant p53-null cancer cells by inducing mitotic catastrophe. PLoS ONE. 2014;9(12):e115121. doi: 10.1371/journal.pone.0115121.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Xiaogang Zheng
    • 1
    • 2
    • 3
    • 4
  • Xiaodong Jin
    • 1
    • 2
    • 3
  • Feifei Li
    • 1
    • 2
    • 3
    • 4
  • Xiongxiong Liu
    • 1
    • 2
    • 3
  • Yan Liu
    • 1
    • 2
    • 3
    • 4
  • Fei Ye
    • 1
    • 2
    • 3
  • Ping Li
    • 1
    • 2
    • 3
  • Ting Zhao
    • 1
    • 2
    • 3
  • Qiang Li
    • 1
    • 2
    • 3
    Email author
  1. 1.Institute of Modern PhysicsChinese Academy of SciencesLanzhouChina
  2. 2.Key Laboratory of Heavy Ion Radiation Biology and MedicineChinese Academy of SciencesLanzhouChina
  3. 3.Key Laboratory of Basic Research on Heavy Ion Radiation Application in MedicineLanzhouChina
  4. 4.University of Chinese Academy of SciencesBeijingChina

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