The combination of temsirolimus and chloroquine increases radiosensitivity in colorectal cancer cells

  • Authors:
    • Hiroshi Shiratori
    • Kazushige Kawai
    • Keisuke Hata
    • Toshiaki Tanaka
    • Takeshi Nishikawa
    • Kensuke Otani
    • Kazuhito Sasaki
    • Manabu Kaneko
    • Koji Murono
    • Shigenobu Emoto
    • Hirofumi Sonoda
    • Hiroaki Nozawa
  • View Affiliations

  • Published online on: April 23, 2019     https://doi.org/10.3892/or.2019.7134
  • Pages: 377-385
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Abstract

The PI3K/AKT/mTOR pathway and autophagy are known to play important roles in cancer radioresistance. The aim of the present study was to investigate whether the combination of temsirolimus (TEM), an mTOR inhibitor, and chloroquine (CQ), an autophagy inhibitor, can increase radiosensitivity in colorectal cancer (CRC) cells. The efficacies of TEM and/or CQ as radiosensitizers were examined using clonogenic assays in CRC cell lines SW480 and HT‑29. The expression levels of the phosphorylated isoforms of S6 and 4E‑BP1, downstream proteins of mTOR, as well as the expression levels of p62 and LC3, autophagy‑related proteins, were assessed by western blot analysis. The formation of acidic organelles was detected in acridine orange‑stained cells. Apoptosis and caspase activity were assessed using flow cytometry. The results revealed that ionizing radiation (IR) activated the downstream proteins of mTOR and induced autophagy. In the clonogenic assays, neither TEM nor CQ influenced the efficacy of IR, whereas their combination significantly increased the dose‑dependent efficacy of IR. TEM inhibited phosphorylation of the downstream proteins of mTOR and induced autophagy. CQ inhibited autophagy in the late phase and did not influence the downstream proteins of mTOR. TEM and CQ inhibited both the phosphorylation of downstream proteins of mTOR and autophagy. Cell death analysis revealed that the combination of TEM and CQ strongly induced apoptosis in cells exposed to IR. In conclusion, the combination of TEM and CQ increased radiosensitivity in CRC cells through co‑inhibition of mTOR and autophagy.

References

1 

Martin ST, Heneghan HM and Winter DC: Systematic review and meta-analysis of outcomes following pathological complete response to neoadjuvant chemoradiotherapy for rectal cancer. Br J Surg. 99:918–928. 2012. View Article : Google Scholar : PubMed/NCBI

2 

Rödel C, Liersch T, Becker H, Fietkau R, Hohenberger W, Hothorn T, Graeven U, Arnold D, Lang-Welzenbach M, Raab HR, et al: Preoperative chemoradiotherapy and postoperative chemotherapy with fluorouracil and oxaliplatin versus fluorouracil alone in locally advanced rectal cancer: Initial results of the German CAO/ARO/AIO-04 randomised phase 3 trial trial. Lancet Oncol. 13:679–687. 2012. View Article : Google Scholar : PubMed/NCBI

3 

Albert JM, Kim KW, Cao C and Lu B: Targeting the Akt/mammalian target of rapamycin pathway for radiosensitization of breast cancer. Mol Cancer Ther. 5:1183–1189. 2006. View Article : Google Scholar : PubMed/NCBI

4 

Chen H, Ma Z, Vanderwaal RP, Feng Z, Gonzalez-Suarez I, Wang S and Zhang J, Roti Roti JL, Gonzalo S and Zhang J: The mTOR inhibitor rapamycin suppresses DNA double-strand break repair. Radiat Res. 175:214–224. 2011. View Article : Google Scholar : PubMed/NCBI

5 

Buijsen J, van den Bogaard J, Jutten B, Belgers E, Sosef M, Leijtens JW, Beets GL, Jansen RL, Riedl RG, Clarijs R, et al: A phase I–II study on the combination of rapamycin and short course radiotherapy in rectal cancer. Radiother Oncol. 116:214–220. 2015. View Article : Google Scholar : PubMed/NCBI

6 

Chaurasia M, Bhatt AN, Das A, Dwarakanath BS and Sharma K: Radiation-induced autophagy: Mechanisms and consequences. Free Radic Res. 50:273–290. 2016. View Article : Google Scholar : PubMed/NCBI

7 

Hu L, Wang H, Huang L, Zhao Y and Wang J: Crosstalk between autophagy and intracellular radiation response (Review). Int J Oncol. 49:2217–2226. 2016. View Article : Google Scholar : PubMed/NCBI

8 

Rosich L, Xargay-Torrent S, López-Guerra M, Campo E, Colomer D and Roué G: Counteracting autophagy overcomes resistance to everolimus in mantle cell lymphoma. Clin Cancer Res. 18:5278–5289. 2012. View Article : Google Scholar : PubMed/NCBI

9 

Kaneko M, Nozawa H, Hiyoshi M, Tada N, Murono K, Nirei T, Emoto S, Kishikawa J, Iida Y, Sunami E, et al: Temsirolimus and chloroquine cooperatively exhibit a potent antitumor effect against colorectal cancer cells. J Cancer Res Clin Onco. 140:769–781. 2014. View Article : Google Scholar

10 

Mo N, Lu YK, Xie WM, Liu Y, Zhou WX, Wang HX, Nong L, Jia YX, Tan AH, Chen Y, et al: Inhibition of autophagy enhances the radiosensitivity of nasopharyngeal carcinoma by reducing Rad51 expression. Oncol Rep. 32:1905–1912. 2014. View Article : Google Scholar : PubMed/NCBI

11 

Sun Q, Liu T, Yuan Y, Guo Z, Xie G, Du S, Lin X, Xu Z, Liu M, Wang W, et al: MiR-200c inhibits autophagy and enhances radiosensitivity in breast cancer cells by targeting UBQLN1. Int J Cancer. 136:1003–1012. 2015. View Article : Google Scholar : PubMed/NCBI

12 

Apel A, Herr I, Schwarz H, Rodemann HP and Mayer A: Blocked autophagy sensitizes resistant carcinoma cells to radiation therapy. Cancer Res. 68:1485–1494. 2008. View Article : Google Scholar : PubMed/NCBI

13 

Okuno T, Kawai K, Hata K, Murono K, Emoto S, Kaneko M, Sasaki K, Nishikawa T, Tanaka T and Nozawa H: SN-38 Acts as a radiosensitizer for colorectal cancer by inhibiting the radiation-induced Up-regulation of HIF-1α. Anticancer Res. 38:3323–3331. 2018. View Article : Google Scholar : PubMed/NCBI

14 

Saxton RA and Sabatini DM: mTOR signaling in growth, metabolism, and disease. Cell. 168:960–976. 2017. View Article : Google Scholar : PubMed/NCBI

15 

Fingar DC, Richardson CJ, Tee AR, Cheatham L, Tsou C and Blenis J: mTOR controls cell cycle progression through its cell growth effectors S6K1 and 4E-BP1/eukaryotic translation initiation factor 4E. Mol Cell Biol. 24:200–216. 2004. View Article : Google Scholar : PubMed/NCBI

16 

Miyasaka A, Oda K, Ikeda Y, Sone K, Fukuda T, Inaba K, Makii C, Enomoto A, Hosoya N, Tanikawa M, et al: PI3K/mTOR pathway inhibition overcomes radioresistance via suppression of the HIF1-α/VEGF pathway in endometrial cancer. Gynecol Oncol. 138:174–180. 2015. View Article : Google Scholar : PubMed/NCBI

17 

Edwards E, Geng L, Tan J, Onishko H, Donnelly E and Hallahan DE: Phosphatidylinositol 3-kinase/Akt signaling in the response of vascular endothelium to ionizing radiation. Cancer Res. 62:4671–4677. 2002.PubMed/NCBI

18 

Nishikawa T, Tsuno NH, Okaji Y, Shuno Y, Sasaki K, Hongo K, Sunami E, Kitayama J, Takahashi K and Nagawa H: Inhibition of autophagy potentiates sulforaphane-induced apoptosis in human colon cancer cells. Ann Surg Oncol. 17:592–602. 2010. View Article : Google Scholar : PubMed/NCBI

19 

Sasaki K, Tsuno NH, Sunami E, Tsurita G, Kawai K, Okaji Y, Nishikawa T, Shuno Y, Hongo K, Hiyoshi M, et al: Chloroquine potentiates the anti-cancer effect of 5-fluorouracil on colon cancer cells. BMC Cancer. 10:3702010. View Article : Google Scholar : PubMed/NCBI

20 

Al-Bari MA: Chloroquine analogues in drug discovery: New directions of uses, mechanisms of actions and toxic manifestations from malaria to multifarious diseases. J Antimicrob Chemother. 70:1608–1621. 2015.PubMed/NCBI

21 

Cao C, Subhawong T, Albert JM, Kim KW, Geng L, Sekhar KR, Gi YJ and Lu B: Inhibition of mammalian target of rapamycin or apoptotic pathway induces autophagy and radiosensitizes PTEN nulll prostate cancer cells. Cancer Res. 66:10040–10047. 2006. View Article : Google Scholar : PubMed/NCBI

22 

Manegold PC, Paringer C, Kulka U, Krimmel K, Eichhorn ME, Wilkowski R, Jauch KW, Guba M and Bruns CJ: Antiangiogenic therapy with mammalian target of rapamycin inhibitor RAD001 (Everolimus) increases radiosensitivity in solid cancer. Clin Cancer Res. 14:892–900. 2008. View Article : Google Scholar : PubMed/NCBI

23 

Lomonaco SL, Finniss S, Xiang C, Decarvalho A, Umansky F, Kalkanis SN, Mikkelsen T and Brodie C: The induction of autophagy by gamma-radiation contributes to the radioresistance of glioma stem cells. Int J Cancer. 125:717–722. 2009. View Article : Google Scholar : PubMed/NCBI

24 

Yan C, Luo L, Goto S, Urata Y, Guo CY, Doi H, Kitazato K and Li TS: Enhanced autophagy in colorectal cancer stem cells does not contribute to radio-resistance. Oncotarget. 7:45112–45121. 2016. View Article : Google Scholar : PubMed/NCBI

25 

Bristol ML, Emery SM, Maycotte P, Thorburn A, Chakradeo S and Gewirtz DA: Autophagy inhibition for chemosensitization and radiosensitization in cancer: Do the preclinical data support this therapeutic strategy? J Pharmacol Exp Ther. 344:544–552. 2013. View Article : Google Scholar : PubMed/NCBI

26 

Enzenmuller S, Gonzalez P, Debatin KM and Fulda S: Chloroquine overcomes resistance of lung carcinoma cells to the dual PI3K/mTOR inhibitor PI103 by lysosome-mediated apoptosis. Anticancer Drugs. 24:14–19. 2013. View Article : Google Scholar : PubMed/NCBI

27 

Seitz C, Hugle M, Cristofanon S, Tchoghandjian A and Fulda S: The dual PI3K/mTOR inhibitor NVP-BEZ235 and chloroquine synergize to trigger apoptosis via mitochondrial-lysosomal cross-talk. Int J Cancer. 132:2682–2693. 2013. View Article : Google Scholar : PubMed/NCBI

28 

Choo AY, Yoon SO, Kim SG, Roux PP and Blenis J: Rapamycin differentially inhibits S6Ks and 4E-BP1 to mediate cell-type-specific repression of mRNA translation. Proc Natl Acad Sci USA. 105:17414–17419. 2008. View Article : Google Scholar : PubMed/NCBI

29 

Shi TT, Yu XX, Yan LJ and Xiao HT: Research progress of hydroxychloroquine and autophagy inhibitors on cancer. Cancer Chemother Pharmacol. 79:287–294. 2017. View Article : Google Scholar : PubMed/NCBI

30 

Boya P, Gonzalez-Polo RA, Poncet D, Andreau K, Vieira HL, Roumier T, Perfettini JL and Kroemer G: Mitochondrial membrane permeabilization is a critical step of lysosome-initiated apoptosis induced by hydroxychloroquine. Oncogene. 22:3927–3936. 2003. View Article : Google Scholar : PubMed/NCBI

31 

Lakhter AJ, Sahu RP, Sun Y, Kaufmann WK, Androphy EJ, Travers JB and Naidu SR: Chloroquine promotes apoptosis in melanoma cells by inhibiting BH3 domain-mediated PUMA degradation. J Invest Dermatol. 133:2247–2254. 2013. View Article : Google Scholar : PubMed/NCBI

32 

Rangwala R, Chang YC, Hu J, Algazy KM, Evans TL, Fecher LA, Schuchter LM, Torigian DA, Panosian JT, Troxel AB, et al: Combined MTOR and autophagy inhibition: Phase I trial of hydroxychloroquine and temsirolimus in patients with advanced solid tumors and melanoma. Autophagy. 10:1391–1402. 2014. View Article : Google Scholar : PubMed/NCBI

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Copy and paste a formatted citation
APA
Shiratori, H., Kawai, K., Hata, K., Tanaka, T., Nishikawa, T., Otani, K. ... Nozawa, H. (2019). The combination of temsirolimus and chloroquine increases radiosensitivity in colorectal cancer cells. Oncology Reports, 42, 377-385. https://doi.org/10.3892/or.2019.7134
MLA
Shiratori, H., Kawai, K., Hata, K., Tanaka, T., Nishikawa, T., Otani, K., Sasaki, K., Kaneko, M., Murono, K., Emoto, S., Sonoda, H., Nozawa, H."The combination of temsirolimus and chloroquine increases radiosensitivity in colorectal cancer cells". Oncology Reports 42.1 (2019): 377-385.
Chicago
Shiratori, H., Kawai, K., Hata, K., Tanaka, T., Nishikawa, T., Otani, K., Sasaki, K., Kaneko, M., Murono, K., Emoto, S., Sonoda, H., Nozawa, H."The combination of temsirolimus and chloroquine increases radiosensitivity in colorectal cancer cells". Oncology Reports 42, no. 1 (2019): 377-385. https://doi.org/10.3892/or.2019.7134