Radiosensitizing effect of oleanolic acid on tumor cells through the inhibition of GSH synthesis in vitro

Wang,Juan; Yu,Maohu; Xiao,Linlin; Xu,Shiguo; Yi,Qiyi; Jin,Wensen

doi:10.3892/or.2013.2510

Radiosensitizing effect of oleanolic acid on tumor cells through the inhibition of GSH synthesis in vitro

Authors:
- Juan Wang
- Maohu Yu
- Linlin Xiao
- Shiguo Xu
- Qiyi Yi
- Wensen Jin
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Affiliations: Teaching and Research Section of Nuclear Medicine, Anhui Medical University, Hefei, Anhui 230032, P.R. China
Published online on: May 31, 2013 https://doi.org/10.3892/or.2013.2510
Pages: 917-924

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Abstract

Oleanolic acid (OA) is a natural pentacyclic triterpenoid that has been used in traditional medicine as an anticancer and anti-inflammatory agent. The aim of our study was to investigate whether or not OA increases the radiosensivity of tumor cells, and the relative mechanism was also investigated. Clonogenic assay was used to observe the radiosensitivity of C6 and A549 cells following different treatments. The alteration of intracellular DNA damage was determined using a micronucleus (MN) assay. In order to identify the mechanism of OA-mediated radiosensitization of tumor cells, the levels of glutathione (GSH) in irradiated cells following various pretreatments were determined using glutathione reductase/5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) recycling assay. Under the same condition, the activities of γ-glutamylcysteine synthetase (γ-GCS) and GSH synthase (GSS), both key enzymes for GSH synthesis, were detected using appropriate methods. In order to confirm the radiosensitizing effect of OA on cancer cells by attenuating GSH, N-acetylcysteine (NAC) was added to cells in culture for 12 h before irradiation. The results showed that the combined treatment of radiation with OA significantly decreased the clonogenic growth of tumor cells and enhanced the numbers of intracellular MN compared to irradiation alone. Furthermore, it was found that the synthesis of cellular GSH was inhibited concomitantly with the downregulation of γ-GCS activity. Therefore, the utilization of OA as a radiosensitizing agent for irradiation-inducing cell death offers a potential therapeutic approach to treat cancer.

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1	Shah C, Grills IS, Kestin LL, McGrath S, Ye H, Martin SK and Yan D: Intrafraction variation of mean tumor position during image-guided hypofractionated stereotactic body radiotherapy for lung cancer. Int J Radiat Oncol Biol Phys. 82:1636–1641. 2012. View Article : Google Scholar : PubMed/NCBI
2	Lagadec C, Vlashi E, Della Donna L, Meng Y, Dekmezian C, Kim K and Pajonk F: Survival and self-renewing capacity of breast cancer initiating cells during fractionated radiation treatment. Breast Cancer Res. 12:R132010. View Article : Google Scholar : PubMed/NCBI
3	Jamal M, Rath BH, Williams ES, Camphausen K and Tofilon PJ: Microenvironmental regulation of glioblastoma radioresponse. Clin Cancer Res. 16:6049–6059. 2010. View Article : Google Scholar : PubMed/NCBI
4	Ayouaz A, Raynaud C, Heride C, Revaud D and Sabatier L: Telomeres: hallmarks of radiosensitivity. Biochimie. 90:60–72. 2008. View Article : Google Scholar : PubMed/NCBI
5	Dumont F, Altmeyer A and Bischoff P: Radiosensitising agents for the radiotherapy of cancer: novel molecularly targeted approaches. Expert Opin Ther Pat. 19:775–799. 2009. View Article : Google Scholar : PubMed/NCBI
6	Bondza-Kibangou P, Millot C, El Khoury V and Millot JM: Antioxidants and doxorubicin supplementation to modulate CD14 expression and oxidative stress induced by vitamin D₃ and seocalcitol in HL60 cells. Oncol Rep. 18:1513–1519. 2007.PubMed/NCBI
7	Pani G, Galeotti T and Chiarugi P: Metastasis: cancer cell’s escape from oxidative stress. Cancer Metastasis Rev. 29:351–378. 2010.
8	Forkink M, Smeitink JA, Brock R, Willems PH and Koopman WJ: Detection and manipulation of mitochondrial reactive oxygen species in mammalian cells. Biochim Biophys Acta. 1797:1034–1044. 2010. View Article : Google Scholar : PubMed/NCBI
9	Forman HJ, Zhang H and Rinna A: Glutathione: overview of its protective roles, measurement, and biosynthesis. Mol Aspects Med. 30:1–12. 2009. View Article : Google Scholar : PubMed/NCBI
10	Circu ML and Aw TY: Glutathione and apoptosis. Free Radic Res. 42:689–706. 2008. View Article : Google Scholar
11	Wu G, Fang YZ, Yang S, Lupton JR and Turner ND: Glutathione metabolism and its implications for health. J Nutr. 134:489–492. 2004.PubMed/NCBI
12	Botta D, White CC, Vliet-Gregg P, et al: Modulating GSH synthesis using glutamate cysteine ligase transgenic and gene-targeted mice. Drug Metab Rev. 40:465–477. 2008. View Article : Google Scholar : PubMed/NCBI
13	Zhang W, Trachootham D, Liu J, et al: Stromal control of cystine metabolism promotes cancer cell survival in chronic lymphocytic leukaemia. Nat Cell Biol. 14:276–286. 2012. View Article : Google Scholar : PubMed/NCBI
14	Lewis-Wambi JS, Kim HR, Wambi C, Patel R, Pyle JR, Klein-Szanto AJ and Jordan VC: Buthionine sulfoximine sensitizes antihormone-resistant human breast cancer cells to estrogen-induced apoptosis. Breast Cancer Res. 10:R1042008. View Article : Google Scholar
15	Ogunrinu TA and Sontheimer H: Hypoxia increases the dependence of glioma cells on glutathione. J Biol Chem. 285:37716–37724. 2010. View Article : Google Scholar : PubMed/NCBI
16	Ruiz-Gómez MJ, Souviron A, Martínez-Morillo M and Gil L: P-glycoprotein, glutathione and glutathione S-transferase increase in a colon carcinoma cell line by colchicine. J Physiol Biochem. 56:307–312. 2000.PubMed/NCBI
17	Inci E, Civelek S, Seven A, Inci F, Korkut N and Burçax G: Laryngeal cancer: in relation to oxidative stress. Tohoku J Exp Med. 200:17–23. 2003. View Article : Google Scholar : PubMed/NCBI
18	Honda T, Coppola S, Ghibelli L, et al: GSH depletion enhances adenoviral bax-induced apoptosis in lung cancer cells. Cancer Gene Ther. 11:249–255. 2004. View Article : Google Scholar : PubMed/NCBI
19	Simons AL, Parsons AD, Foster KA, Orcutt KP, Fath MA and Spitz DR: Inhibition of glutathione and thioredoxin metabolism enhances sensitivity to perifosine in head and neck cancer cells. J Oncol. 2009:5195632009. View Article : Google Scholar : PubMed/NCBI
20	Boivin A, Hanot M, Malesys C, Maalouf M, Rousson R, Rodriguez-Lafrasse C and Ardail D: Transient alteration of cellular redox buffering before irradiation triggers apoptosis in head and neck carcinoma stem and non-stem cells. PLoS One. 6:e145582011. View Article : Google Scholar : PubMed/NCBI
21	Pollier J and Goossens A: Oleanolic acid. Phytochemistry. 77:10–15. 2012. View Article : Google Scholar
22	Yang EJ, Lee W, Ku SK, Song KS and Bae JS: Anti-inflammatory activities of oleanolic acid on HMGB1 activated HUVECs. Food Chem Toxicol. 50:1288–1294. 2012. View Article : Google Scholar : PubMed/NCBI
23	Reisman SA, Aleksunes LM and Klaassen CD: Oleanolic acid activates Nrf2 and protects from acetaminophen hepatotoxicity via Nrf2-dependent and Nrf2-independent processes. Biochem Pharmacol. 77:1273–1282. 2009. View Article : Google Scholar : PubMed/NCBI
24	Yim TK, Wu WK, Pak WF and Ko KM: Hepatoprotective action of an oleanolic acid-enriched extract of Ligustrum lucidum fruits is mediated through an enhancement on hepatic glutathione regeneration capacity in mice. Phytother Res. 15:589–592. 2001.PubMed/NCBI
25	Hsu HY, Yang JJ and Lin CC: Effects of oleanolic acid and ursolic acid on inhibiting tumor growth and enhancing the recovery of hematopoietic system postirradiation in mice. Cancer Lett. 111:7–13. 1997. View Article : Google Scholar : PubMed/NCBI
26	Fujiwara Y, Komohara Y, Kudo R, Tsurushima K, Ohnishi K, Ikeda T and Takeya M: Oleanolic acid inhibits macrophage differentiation into the M2 phenotype and glioblastoma cell proliferation by suppressing the activation of STAT3. Oncol Rep. 26:1533–1537. 2011.PubMed/NCBI
27	Wei J, Liu H, Liu M, et al: Oleanolic acid potentiates the antitumor activity of 5-fluorouracil in pancreatic cancer cells. Oncol Rep. 28:1339–1345. 2012.PubMed/NCBI
28	Bishayee A, Ahmed S, Brankov N and Perloff M: Triterpenoids as potential agents for the chemoprevention and therapy of breast cancer. Front Biosci. 16:980–996. 2011. View Article : Google Scholar : PubMed/NCBI
29	Yamai H, Sawada N, Yoshida T, et al: Triterpenes augment the inhibitory effects of anticancer drugs on growth of human esophageal carcinoma cells in vitro and suppress experimental metastasis in vivo. Int J Cancer. 125:952–960. 2009. View Article : Google Scholar
30	White CC, Viernes H, Krejsa CM, Botta D and Kavanagh TJ: Fluorescence-based microtiter plate assay for glutamate-cysteine ligase activity. Anal Biochem. 318:175–180. 2003. View Article : Google Scholar : PubMed/NCBI
31	Frosina G: DNA repair and resistance of gliomas to chemotherapy and radiotherapy. Mol Cancer Res. 7:989–999. 2009. View Article : Google Scholar : PubMed/NCBI
32	Chang JY and Cox JD: Improving radiation conformality in the treatment of non-small cell lung cancer. Semin Radiat Oncol. 20:171–177. 2010. View Article : Google Scholar : PubMed/NCBI
33	Kim M, Lee J, Ha B, Lee R, Lee KJ and Suh HS: Factors predicting radiation pneumonitis in locally advanced non-small cell lung cancer. Radiat Oncol J. 29:181–190. 2011. View Article : Google Scholar : PubMed/NCBI
34	Zerp SF, Stoter R, Kuipers G, et al: AT-101, a small molecule inhibitor of anti-apoptotic Bcl-2 family members, activates the SAPK/JNK pathway and enhances radiation-induced apoptosis. Radiat Oncol. 4:472009. View Article : Google Scholar : PubMed/NCBI
35	Rao SK, Rao PS and Rao BN: Preliminary investigation of the radiosensitizing activity of guduchi (Tinospora cordifolia) in tumor-bearing mice. Phytother Res. 22:1482–1489. 2008. View Article : Google Scholar : PubMed/NCBI
36	Dai Y, DeSano JT, Meng Y, Ji Q, Ljungman M, Lawrence TS and Xu L: Celastrol potentiates radiotherapy by impairment of DNA damage processing in human prostate cancer. Int J Radiat Oncol Biol Phys. 74:1217–1225. 2009. View Article : Google Scholar : PubMed/NCBI
37	Tsai SJ and Yin MC: Anti-oxidative, anti-glycative and anti-apoptotic effects of oleanolic acid in brain of mice treated by D-galactose. Eur J Pharmacol. 689:81–88. 2012. View Article : Google Scholar : PubMed/NCBI
38	Koh SJ, Tak JK, Kim ST, Nam WS, Kim SY, Park KM and Park JW: Sensitization of ionizing radiation-induced apoptosis by ursolic acid. Free Radic Res. 46:339–345. 2012. View Article : Google Scholar : PubMed/NCBI
39	Eder-Czembirek C, Erovic BM, Czembirek C, Brunner M, Selzer E, Pötter R and Thurnher D: Betulinic acid a radiosensitizer in head and neck squamous cell carcinoma cell lines. Strahlenther Onkol. 186:143–148. 2010. View Article : Google Scholar : PubMed/NCBI
40	Selzer E, Pimentel E, Wacheck V, Schlegel W, Pehamberger H, Jansen B and Kodym R: Effects of betulinic acid alone and in combination with irradiation in human melanoma cells. J Invest Dermatol. 114:935–940. 2000. View Article : Google Scholar : PubMed/NCBI
41	Yang JC, Lu MC, Lee CL, Chen GY, Lin YY, Chang FR and Wu YC: Selective targeting of breast cancer cells through ROS-mediated mechanisms potentiates the lethality of paclitaxel by a novel diterpene, gelomulide K. Free Radic Biol Med. 51:641–657. 2011. View Article : Google Scholar : PubMed/NCBI
42	Karthikeyan S, Kanimozhi G, Prasad NR and Mahalakshmi R: Radiosensitizing effect of ferulic acid on human cervical carcinoma cells in vitro. Toxicol In Vitro. 25:1366–1375. 2011. View Article : Google Scholar : PubMed/NCBI
43	Benassi B, Zupi G and Biroccio A: Gamma-glutamylcysteine synthetase mediates the c-Myc-dependent response to antineoplastic agents in melanoma cells. Mol Pharmacol. 72:1015–1023. 2007. View Article : Google Scholar : PubMed/NCBI
44	Oliveira NG, Castro M, Rodrigues AS, et al: Wortmannin enhances the induction of micronuclei by low and high LET radiation. Mutagenesis. 18:37–44. 2003. View Article : Google Scholar
45	Sedelnikova OA, Nakamura A, Kovalchuk O, et al: DNA double-strand breaks form in bystander cells after microbeam irradiation of three-dimensional human tissue models. Cancer Res. 67:4295–4302. 2007. View Article : Google Scholar
46	Kashino G, Prise KM, Suzuki K, et al: Effective suppression of bystander effects by DMSO treatment of irradiated CHO cells. J Radiat Res. 48:327–333. 2007. View Article : Google Scholar : PubMed/NCBI
47	Shao C, Furusawa Y, Kobayashi Y, Funayama T and Wada S: Bystander effect induced by counted high-LET particles in confluent human fibroblasts: a mechanistic study. FASEB J. 17:1422–1427. 2003. View Article : Google Scholar : PubMed/NCBI
48	Thierens H and Vral A: The micronucleus assay in radiation accidents. Ann Ist Super Sanita. 45:260–264. 2009.PubMed/NCBI
49	McConnachie LA, Mohar I, Hudson FN, et al: Glutamate cysteine ligase modifier subunit deficiency and gender as determinants of acetaminophen-induced hepatotoxicity in mice. Toxicol Sci. 99:628–636. 2007. View Article : Google Scholar : PubMed/NCBI
50	Bai X, Qiu A, Guan J and Shi Z: Antioxidant and protective effect of an oleanolic acid-enriched extract of A. deliciosa root on carbon tetrachloride induced rat liver injury. Asia Pac J Clin Nutr. 16(Suppl 1): 169–173. 2007.PubMed/NCBI
51	Abdel-Zaher AO, Abdel-Rahman MM, Hafez MM and Omran FM: Role of nitric oxide and reduced glutathione in the protective effects of aminoguanidine, gadolinium chloride and oleanolic acid against acetaminophen-induced hepatic and renal damage. Toxicology. 234:124–134. 2007. View Article : Google Scholar
52	Petronelli A, Saulle E, Pasquini L, et al: High sensitivity of ovarian cancer cells to the synthetic triterpenoid CDDO-imidazolide. Cancer Lett. 282:214–228. 2009. View Article : Google Scholar : PubMed/NCBI
53	Ikeda T, Sporn M, Honda T, Gribble GW and Kufe D: The novel triterpenoid CDDO and its derivatives induce apoptosis by disruption of intracellular redox balance. Cancer Res. 63:5551–5558. 2003.PubMed/NCBI
54	Haddad JJ, Olver RE and Land SC: Antioxidant/pro-oxidant equilibrium regulates HIF-1alpha and NF-kappa B redox sensitivity. Evidence for inhibition by glutathione oxidation in alveolar epithelial cells. J Biol Chem. 275:21130–21139. 2000. View Article : Google Scholar