xCT inhibitor sulfasalazine depletes paclitaxel‑resistant tumor cells through ferroptosis in uterine serous carcinoma

Sugiyama,Akiko; Ohta,Tsuyoshi; Obata,Miyuki; Takahashi,Kanako; Seino,Manabu; Nagase,Satoru

doi:10.3892/ol.2020.11813

xCT inhibitor sulfasalazine depletes paclitaxel‑resistant tumor cells through ferroptosis in uterine serous carcinoma

Authors:
- Akiko Sugiyama
- Tsuyoshi Ohta
- Miyuki Obata
- Kanako Takahashi
- Manabu Seino
- Satoru Nagase
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Affiliations: Department of Obstetrics and Gynecology, Faculty of Medicine, Yamagata University, Yamagata 990‑9585, Japan
Published online on: July 6, 2020 https://doi.org/10.3892/ol.2020.11813
Pages: 2689-2700
Copyright: © Sugiyama et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Glutathione (GSH) is a primary antioxidant that protects cells against reactive oxygen species (ROS), and high levels of GSH promote cancer cell survival and resistance to chemotherapy. The glutamine transporter xCT is essential for the intracellular synthesis of GSH, whereby xCT determines the intracellular redox balance. However, whether xCT inhibition can overcome GSH‑mediated resistance to chemotherapeutic agents in uterine serous carcinoma (USC) remains unclear. Thus, the present study investigated the effect of the xCT inhibitor, sulfasalazine (SAS) on cytotoxicity in paclitaxel‑sensitive and ‑resistant USC cell lines. The molecular mechanism by which SAS induces ferroptotic cell death in paclitaxel‑resistant cells was assessed. The results of the cytotoxicity assay demonstrated that SAS was more cytotoxic in paclitaxel‑resistant cells compared with in ‑sensitive cells; however, paclitaxel cytotoxicity was not enhanced in either of the USC cell lines. Immunoblotting analysis and the cell death assays performed using ferroptosis inhibitors indicated that SAS‑mediated cell death was induced through ferroptosis, and not apoptosis, in paclitaxel‑resistant cells. Furthermore, ROS production was increased in paclitaxel‑resistant but not in ‑sensitive cells, even at low SAS concentration, and JNK was activated, which is a downstream target in the Ras signaling pathway. Knockdown of JNK reversed the inhibitory effect of SAS on cell proliferation and cell death. The synthetic lethal interaction between ROS accumulation and Ras effector JNK activation may be critical for enhancing the sensitivity to ferroptotic cell death mediated by xCT inhibitor, SAS. Taken together, the results of the present study suggest that xCT inhibition may be an effective treatment for patients with recurrent paclitaxel‑resistant USC.

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1	Menderes G, Clark M and Santin AD: Novel targeted therapies in uterine serous carcinoma, an aggressive variant of endometrial cancer. Discov Med. 21:293–303. 2016.PubMed/NCBI
2	Hamilton CA, Cheung MK, Osann K, Chen L, Teng NN, Longacre TA, Powell MA, Hendrickson MR, Kapp DS and Chan JK: Uterine papillary serous and clear cell carcinomas predict for poorer survival compared to grade 3 endometrioid corpus cancers. Br J Cancer. 94:642–646. 2006. View Article : Google Scholar : PubMed/NCBI
3	Boruta DM II, Gehrig PA, Fader AN and Olawaiye AB: Management of women withuterine papillary serous cancer: A society of gynecologic oncology (SGO) review. Gynecol Oncol. 115:142–153. 2009. View Article : Google Scholar : PubMed/NCBI
4	Goff BA, Kato D, Schmidt RA, Ek M, Ferry JA, Munts HG, Cain JM, Tamimi HK, Figge DC and Greer BE: Uterine papillary serous carcinoma: Patterns of metastatic spread. Gynecol Oncol. 54:264–268. 1994. View Article : Google Scholar : PubMed/NCBI
5	Lecane PS, Karaman MW, Sirisawad M, Naumovski L, Miller RA, Hacia JG and Magda D: Motexafin gadolinium and zinc induce oxidative stress responses and apoptosis in B-cell lymphoma lines. Cancer Res. 65:11676–11688. 2005. View Article : Google Scholar : PubMed/NCBI
6	Godwin AK, Meister A, O'Dwyer PJ, Huang CS, Hamilton TC and Anderson ME: High resistance to cisplatin in human ovarian cancer cell lines is associated with marked increase of glutathione synthesis. Proc Natl Acad Sci USA. 89:3070–3074. 1992. View Article : Google Scholar : PubMed/NCBI
7	Mulcahy RT, Untawale S and Gipp JJ: Transcriptional up-regulation of gamma-glutamylcysteine synthetase gene expression in melphalan-resistant human prostate carcinoma cells. Mol Pharmacol. 46:909–914. 1994.PubMed/NCBI
8	Trachootham D, Alexandre J and Huang P: Targeting cancer cells by ROS-mediated mechanisms: A radical therapeutic approach? Nat Rev Drug Discov. 8:579–591. 2009. View Article : Google Scholar : PubMed/NCBI
9	Seino M, Ohta T, Sugiyama A, Sakaki H, Sudo T, Tsutsumi S, Shigeta S, Tokunaga H, Toyoshima M, Yaegashi N and Nagase S: Metabolomic analysis of uterine serous carcinoma with acquired resistance to paclitaxel. Oncotarget. 9:31985–31998. 2018. View Article : Google Scholar : PubMed/NCBI
10	Lo M, Wang Y and Gout PW: The x(c)- cystine/glutamate antiporter: A potential target for therapy of cancer and other diseases. J Cell Physiol. 215:593–602. 2008. View Article : Google Scholar : PubMed/NCBI
11	Aquilano K, Baldelli S and Ciriolo MR: Glutathione: New roles in redox signaling for an old antioxidant. Front Pharmacol. 5:1962014. View Article : Google Scholar : PubMed/NCBI
12	Yoshikawa M, Tsuchihashi K, Ishimoto T, Yae T, Motohara T, Sugihara E, Onishi N, Masuko T, Yoshizawa K, Kawashiri S, et al: xCT inhibition depletes CD44v-expressing tumor cells that are resistant to EGFR-targeted therapy in head and neck squamous cell carcinoma. Cancer Res. 73:1855–1866. 2013. View Article : Google Scholar : PubMed/NCBI
13	Wu WJ, Zhang Y, Zeng ZL, Li XB, Hu KS, Luo HY, Yang J, Huang P and Xu RH: β-phenylethyl isothiocyanate reverses platinum resistance by a GSH-dependent mechanism in cancer cells with epithelial-mesenchymal transition phenotype. Biochem Pharmacol. 85:486–496. 2013. View Article : Google Scholar : PubMed/NCBI
14	Ma MZ, Chen G, Wang P, Lu WH, Zhu CF, Song M, Yang J, Wen S, Xu RH, Hu Y and Huang P: Xc- inhibitor sulfasalazine sensitizes colorectal cancer to cisplatin by a GSH-dependent mechanism. Cancer Lett. 368:88–96. 2015. View Article : Google Scholar : PubMed/NCBI
15	Zenlea T and Peppercorn MA: Immunosuppressive therapies for inflammatory bowel disease. 20:3146–3152. 2014.PubMed/NCBI
16	Zhang W, Trachootham D, Liu J, Chen G, Pelicano H, Garcia-Prieto C, Lu W, Burger JA, Croce CM, Plunkett W, 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
17	Podolsky DK: Inflammatory bowel disease. N Engl J Med. 347:417–429. 2002. View Article : Google Scholar : PubMed/NCBI
18	Nielsen OH, Verspaget HW and Elmgreen J: Inhibition of intestinal macrophage chemotaxis to leukotriene B4 by sulphasalazine, olsalazine, and 5-aminosalicylic acid. Aliment Pharmacol Ther. 2:203–211. 1988. View Article : Google Scholar : PubMed/NCBI
19	Wahl C, Liptay S, Adler G and Schmid RM: Sulfasalazine: A potent and specific inhibitor of nuclear factor kappa B. J Clin Invest. 101:1163–1174. 1998. View Article : Google Scholar : PubMed/NCBI
20	Lo M, Ling V, Low C, Wang YZ and Gout PW: Potential use of the anti-inflammatory drug, sulfasalazine, for targeted therapy of pancreatic cancer. Curr Oncol. 17:9–16. 2010.PubMed/NCBI
21	Doxsee DW, Gout PW, Kurita T, Lo M, Buckley AR, Wang Y, Xue H, Karp CM, Cutz JC, Cunha GR and Wang YZ: Sulfasalazine-induced cystine starvation: Potential use for prostate cancer therapy. Prostate. 67:162–171. 2007. View Article : Google Scholar : PubMed/NCBI
22	Nnrang VS, Pauletti GM, Gout PW, Buckley DJ and Buckley AR: Sulfasalazine-induced reduction of glutathione levels in breast cancer cells: Enhancement of growth-inhibitory activity of Doxorubicin. Chemotherapy. 53:210–217. 2007. View Article : Google Scholar : PubMed/NCBI
23	Kagami T, Wang Y, Tien A, Watahiki A, Lo M, Xue H, Gout P and Wang ZY: Sulfasalazine enhances growth-inhibitory activity of doxorubicin: Potential use in combination therapy of advanced prostate cancer. Proc 98th Ann Assoc Cancer Res (Ros Angeles, CA). 2007.
24	Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, Patel DN, Bauer AJ, Cantley AM, Yang WS, et al: Ferroptosis: An iron-dependent form of nonapoptotic cell death. Cell. 149:1060–1072. 2012. View Article : Google Scholar : PubMed/NCBI
25	Cao JY and Dixon SJ: Mechanisms of ferroptosis. Cell Mol Life Sci. 73:2195–2209. 2016. View Article : Google Scholar : PubMed/NCBI
26	Santin AD, Bellone S, Gokden M, Palmieri M, Dunn D, Agha J, Roman JJ, Hutchins L, Pecorelli S, O'Brien T, et al: Overexpression of HER-2/neu in uterine serous papillary cancer. Clin Cancer Res. 8:1271–1279. 2002.PubMed/NCBI
27	Sakaki H, Okada M, Kuramoto K, Takeda H, Watarai H, Suzuki S, Seino S, Seino M, Ohta T, Nagase S, et al: GSKJ4, a selective Jumonji H3K27 demethylase inhibitor, effectively targets ovarian cancer stem cells. Anticancer Res. 35:6607–6614. 2015.PubMed/NCBI
28	Orian-Rousseau V: CD44 acts as a signaling platform controlling tumor progression and metastasis. Front Immunol. 6:1542015. View Article : Google Scholar : PubMed/NCBI
29	Tanabe KK, Nishi T and Saya H: Novel variants of CD44 arising from alternative splicing: Changes in the CD44 alternative splicing pattern of MCF-7 breast carcinoma cells treated with hyaluronidase. Mol Carcinog. 7:212–220. 1993. View Article : Google Scholar : PubMed/NCBI
30	Ishimoto T, Nagano O, Yae T, Tamada M, Motohara T, Oshima H, Oshima M, Ikeda T, Asaba R, Yagi H, et al: CD44 variant regulates redox status in cancer cells by stabilizing the xCT subunit of system xc(−) and thereby promotes tumor growth. Cancer Cell. 19:387–400. 2011. View Article : Google Scholar : PubMed/NCBI
31	Yae T, Tsuchihashi K, Ishimoto T, Motohara T, Yoshikawa M, Yoshida GJ, Wada T, Masuko T, Mogushi K, Tanaka H, et al: Alternative splicing of CD44 mRNA by ESRP1 enhances lung colonization of metastatic cancer cell. Nat Commun. 3:8832012. View Article : Google Scholar : PubMed/NCBI
32	Oliver FJ, de la Rubia G, Rolli V, Ruiz-Ruiz MC, de Murcia G and Murcia JM: Importance of poly(ADP-ribose) polymerase and its cleavage in apoptosis. Lesson from an uncleavable mutant. J Biol Chem. 273:33533–33539. 1998. View Article : Google Scholar : PubMed/NCBI
33	Shem T, Fan Z, Ghoochani A, Rauh M, Engelhorn T, Minakaki G, Dörfler A, Klucken J, Buchfelder M, Eyüpoglu IY and Savaskan N: Sulfasalazine impacts on ferroptotic cell death and alleviates the tumor microenvironment and glioma-induced brain edema. Oncotarget. 7:36021–36033. 2016. View Article : Google Scholar : PubMed/NCBI
34	Misaghi S, Korbel GA, Kessler B, Spooner E and Ploegh HL: z-VAD-fmk inhibits peptide:N-glycanase and may result in ER stress. Cell Death Differ. 13:163–165. 2006. View Article : Google Scholar : PubMed/NCBI
35	Enomoto M, Kizawa D, Ohsawa S and Igaki T: JNK signaling is converted from anti- to pro-tumor pathway by Ras-mediated switch of Warts activity. Dev Biol. 403:162–171. 2015. View Article : Google Scholar : PubMed/NCBI
36	Robe PA, Bentires-Alj M, Bonif M, Rogister B, Deprez M, Haddada H, Khac MT, Jolois O, Erkmen K, Merville MP, et al: In vitro and in vivo activity of the nuclear factor-kappaB inhibitor sulfasalazine in human glioblastomas. Clin Cancer Res. 10:5595–5603. 2004. View Article : Google Scholar : PubMed/NCBI
37	Müerköster S, Arlt A, Witt M, Gehrz A, Haye S, March C, Grohmann F, Wegehenkel K, Kalthoff H, Fölsch UR and Schäfer H: Usage of the NF-kappaB inhibitor sulfasalazine as sensitizing agent in combined chemotherapy of pancreatic cancer. Int J Cancer. 104:469–476. 2003. View Article : Google Scholar : PubMed/NCBI
38	Lay JD, Hong CC, Huang JS, Yang YY, Pao CY, Liu CH, Lai YP, Lai GM, Cheng AL, Su IJ and Chuang SE: Sulfasalazine suppresses drug resistance and invasiveness of lung adenocarcinoma cells expressing AXL. Cancer Res. 67:3878–3887. 2007. View Article : Google Scholar : PubMed/NCBI
39	Okuno S, Sato H, Kuriyama-Matsumura K, Tamba M, Wang H, Sohda S, Hamada H, Yoshikawa H, Kondo T and Bannai S: Role of cystine transport in intracellular glutathione level and cisplatin resistance in human ovarian cancer cell lines. Br J Cancer. 88:951–956. 2003. View Article : Google Scholar : PubMed/NCBI
40	Huang Y, Dai Z, Barbacioru C and Sadée W: Cystine-glutamate transporter SLC7A11 in cancer chemosensitivity and chemoresistance. Cancer Res. 65:7446–7456. 2005. View Article : Google Scholar : PubMed/NCBI
41	Kandil S, Brennan L and McBean GJ: Glutathione depletion causes a JNK and p38MAPK-mediated increase in expression of cystathionine-gamma-lyase and upregulation of the transsulfuration pathway in C6 glioma cells. Neurochem Int. 56:611–619. 2010. View Article : Google Scholar : PubMed/NCBI
42	Roh JL, Kim EH, Jang HJ, Park JY and Shin D: Induction of ferroptotic cell death for overcoming cisplatin resistance of head and neck cancer. Cancer Lett. 381:96–103. 2016. View Article : Google Scholar : PubMed/NCBI
43	Yang WS, SriRamaratnam R, Welsch ME, Shimada K, Skouta R, Viswanathan VS, Cheah JH, Clemons PA, Shamji AF, Clish CB, et al: Regulation of ferroptotic cancer cell death by GPX4. Cell. 156:317–331. 2014. View Article : Google Scholar : PubMed/NCBI
44	Dolma S, Lessnick SL, Hahn WC and Stockwell BR: Identification of genotype-selective antitumor agents using synthetic lethal chemical screening in engineered human tumor cells. Cancer Cell. 3:285–296. 2003. View Article : Google Scholar : PubMed/NCBI
45	Yagoda N, von Rechenberg M, Zaganjor E, Bauer AJ, Yang WS, Fridman DJ, Wolpaw AJ, Smukste I, Peltier JM, Boniface JJ, et al: RAS-RAF-MEK-dependent oxidative cell death involving voltage-dependent anion channels. Nature. 447:864–868. 2007. View Article : Google Scholar : PubMed/NCBI
46	Irani K, Xia Y, Zweier JL, Sollott SJ, Der CJ, Fearon ER, Sundaresan M, Finkel T and Goldschmidt-Clermont PJ: Mitogenic signaling mediated by oxidants in Ras-transformed fibroblasts. Science. 275:1649–1652. 1997. View Article : Google Scholar : PubMed/NCBI
47	Garama DJ, Harris TJ, White CL, Rossello FJ, Abdul-Hay M, Gough DJ and Levy DE: A synthetic lethal interaction between glutathione synthesis and mitochondrial reactive oxygen species provides a tumor-specific vulnerability dependent on STAT3. Mol Cell Biol. 35:3646–3656. 2015. View Article : Google Scholar : PubMed/NCBI
48	Seino M, Okada M, Sakaki H, Takeda H, Watarai H, Suzuki S, Seino S, Kuramoto K, Ohta T, Nagase S, et al: Time-staggered inhibition of JNK effectively sensitizes chemoresistant ovarian cancer cells to cisplatin and paclitaxel. Oncol Rep. 35:593–601. 2016. View Article : Google Scholar : PubMed/NCBI
49	Okada M, Kuramoto K, Takeda H, Watarai H, Sakaki H, Seino S, Seino M, Suzuki S and Kitanaka C: The novel JNK inhibitor AS602801 inhibits cancer stem cells in vitro and in vivo. Oncotarget. 7:27021–27032. 2016. View Article : Google Scholar : PubMed/NCBI
50	Jiang L, Kon N, Li T, Wang SJ, Su T, Hibshoosh H, Baer R and Gu W: Ferroptosis as a p53-mediated activity during tumour suppression. Nature. 520:57–62. 2015. View Article : Google Scholar : PubMed/NCBI
51	Gilmore TD: Introduction to NF-kappaB: Players, pathways, perspectives. Oncogene. 25:6680–6684. 2006. View Article : Google Scholar : PubMed/NCBI
52	Robe PA, Martin DH, Nguyen-Khac MT, Artesi M, Deprez M, Albert A, Vanbelle S, Califice S, Bredel M and Bours V: Early termination of ISRCTN45828668, a phase 1/2 prospective, randomized study of sulfasalazine for the treatment of progressing malignant gliomas in adults. BMC Cancer. 9:3722009. View Article : Google Scholar : PubMed/NCBI
53	Shitara K, Doi T, Nagano O, Imamura CK, Ozeki T, Ishii Y, Tsuchihashi K, Takahashi S, Nakajima TE, Hironaka S, et al: Dose-escalation study for the targeting of CD44v+ cancer stem cells by sulfasalazine in patients with advanced gastric cancer (EPOC1205). Gastric Cancer. 20:341–349. 2017. View Article : Google Scholar : PubMed/NCBI
54	Shitara K, Doi T, Nagano O, Fukutani M, Hasegawa H, Nomura S, Sato A, Kuwata T, Asai K, Einaga Y, et al: Phase 1 study of sulfasalazine and cisplatin for patients with CD44v-positive gastric cancer refractory to cisplatin (EPOC1407). Gastric Cancer. 20:1004–1009. 2017. View Article : Google Scholar : PubMed/NCBI
55	Sandborn WJ and Feagan BG: Review article: Mild to moderate Crohn's disease-defining the basis for a new treatment algorithm. Aliment Pharmacol Ther. 18:263–277. 2003. View Article : Google Scholar : PubMed/NCBI