WWOX promotes apoptosis and inhibits autophagy in paclitaxel‑treated ovarian carcinoma cells

Zhao,Yan; Wang,Wen; Pan,Wenying; Yu,Yunhai; Huang,Wenqian; Gao,Jiangang; Zhang,Ying; Zhang,Shiqian

doi:10.3892/mmr.2020.11754

WWOX promotes apoptosis and inhibits autophagy in paclitaxel‑treated ovarian carcinoma cells

Authors:
- Yan Zhao
- Wen Wang
- Wenying Pan
- Yunhai Yu
- Wenqian Huang
- Jiangang Gao
- Ying Zhang
- Shiqian Zhang
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Affiliations: School of Medicine, Shandong University, Jinan, Shandong 250012, P.R. China, Department of Gynecology and Obstetrics, Tengzhou Central People's Hospital, Zaozhuang, Shandong 277500, P.R. China, Department of Gynecology and Obstetrics, The Affiliated Hospital of Binzhou Medical College, Binzhou, Shandong 264003, P.R. China, Department of Gynecology and Obstetrics, The Second Hospital of Shandong University, Jinan, Shandong 250032, P.R. China, School of Life Sciences, Shandong University, Jinan, Shandong 250100, P.R. China, Department of Gynecology and Obstetrics, The First People's Hospital of Jining, Jining, Shandong 272000, P.R. China, Department of Gynecology and Obstetrics, Qilu Hospital, Shandong University, Jinan, Shandong 250012, P.R. China
Published online on: December 3, 2020 https://doi.org/10.3892/mmr.2020.11754
Article Number: 115

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Abstract

Although paclitaxel (PTX) is a first‑line chemotherapeutic agent for the treatment of epithelial ovarian cancer (EOC), its clinical use is restricted by chemoresistance. Autophagy is believed to promote drug resistance, and WW domain‑containing oxidoreductase (WWOX) has been predicted to serve an essential role in apoptosis induction and to suppress autophagy in various tumor cell types. In the present study, the role of WWOX was demonstrated using PTX‑treated EOC cells. Cell viability and apoptosis were detected using Cell Counting Kit‑8, morphological and flow cytometric analyses. WWOX and phosphorylated (p)‑WWOX were highly expressed in PTX‑treated sensitive EOC cells (A2780), which was accompanied by activation of the apoptosis‑related proteins caspase‑3 and poly (ADP‑ribose) polymerase (PARP). Conversely, PTX‑resistant EOC cells (A2780/T) were characterized by reduced WWOX expression and constant phosphorylation levels, as well as undetectable levels of activated caspase‑3 and PARP when cells were treated with PTX. The altered expression of WWOX between the two cell types was further validated by reverse transcription‑quantitative PCR. The apoptosis‑inducing role of WWOX was also confirmed by flow cytometry after WWOX overexpression was induced in PTX‑treated A2780 cells. These findings indicated that WWOX activation may inhibit chemoresistance and trigger cancer cell death. The upregulated expression levels of the autophagy‑related protein 12‑5 complex, Beclin‑1 and LC3, as well as the downregulation of P62, were also detected following PTX treatment, suggesting that PTX induced autophagic flux in both types of EOC cells. This conclusion was further supported by visualizing the accumulation of autophagosome and autolysosome vesicles, using confocal microscopy and transmission electron microscopy. PTX was also shown to inhibit mTOR signaling, indicated by a decreased level of p‑mTOR and increased expression of eukaryotic translation initiation factor 4E‑binding protein 1. Finally, the interaction between WWOX, mTOR and autophagy was investigated via WWOX transfection experimentation, and indicated that WWOX activated mTOR whilst inhibiting autophagy. These data indicated that WWOX may serve a critical role in PTX‑induced apoptosis and could suppress autophagy by downregulating essential autophagic effectors in EOC cells via mTOR signaling.

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1	Ferlay J, Soerjomataram I, Dikshit R, Eser S, Mathers C, Rebelo M, Parkin DM, Forman D and Bray F: Cancer incidence and mortality worldwide: Sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer. 136:E359–E386. 2015. View Article : Google Scholar : PubMed/NCBI
2	Mozzetti S, Ferlini C, Concolino P, Filippetti F, Raspaglio G, Prislei S, Gallo D, Martinelli E, Ranelletti FO, Ferrandina G, et al: Class III beta-tubulin overexpression is a prominent mechanism of paclitaxel resistance in ovarian cancer patients. Clin Cancer Res. 11:298–305. 2005.PubMed/NCBI
3	Veldhoen RA, Banman SL, Hemmerling DR, Odsen R, Simmen T, Simmonds AJ, Underhill DA and Goping IS: The chemotherapeutic agent paclitaxel inhibits autophagy through two distinct mechanisms that regulate apoptosis. Oncogene. 32:736–746. 2013. View Article : Google Scholar : PubMed/NCBI
4	Lee Y, Na J, Lee MS, Cha EY, Sul JY, Park JB and Lee JS: Combination of pristimerin and paclitaxel additively induces autophagy in human breast cancer cells via ERK1/2 regulation. Mol Med Rep. 18:4281–4288. 2018.PubMed/NCBI
5	Zhan L, Zhang Y, Wang W, Song E, Fan Y, Li J and Wei B: Autophagy as an emerging therapy target for ovarian carcinoma. Oncotarget. 7:83476–83487. 2016. View Article : Google Scholar : PubMed/NCBI
6	Hale AN, Ledbetter DJ, Gawriluk TR and Rucker EB III: Autophagy: Regulation and role in development. Autophagy. 9:951–972. 2013. View Article : Google Scholar : PubMed/NCBI
7	Wang C, Yang Y, Sun L, Wang J, Jiang Z, Li Y, Liu D, Sun H and Pan Z: Baicalin reverses radioresistance in nasopharyngeal carcinoma by downregulating autophagy. Cancer Cell Int. 20:352020. View Article : Google Scholar : PubMed/NCBI
8	Tyutyunyk-Massey L and Gewirtz DA: Roles of autophagy in breast cancer treatment: Target, bystander or benefactor. Semin Cancer Biol. 66:155–162. 2020. View Article : Google Scholar : PubMed/NCBI
9	Gao K, Yin J and Dong J: Deregulated WWOX is involved in a negative feedback loop with microRNA-214-3p in osteosarcoma. Int J Mol Med. 38:1850–1856. 2016. View Article : Google Scholar : PubMed/NCBI
10	Yan H, Yu N and Tong J: Effects of 5-Aza-2′-deoxycytidine on the methylation state and function of the WWOX gene in the HO-8910 ovarian cancer cell line. Oncol Lett. 6:845–849. 2013. View Article : Google Scholar : PubMed/NCBI
11	Guo W, Wang G, Dong Y, Guo Y, Kuang G and Dong Z: Decreased expression of WWOX in the development of esophageal squamous cell carcinoma. Mol Carcinog. 52:265–274. 2013. View Article : Google Scholar : PubMed/NCBI
12	Li J, Liu J, Ren Y and Liu P: Roles of the WWOX in pathogenesis and endocrine therapy of breast cancer. Exp Biol Med (Maywood). 240:324–328. 2015. View Article : Google Scholar : PubMed/NCBI
13	Baykara O, Demirkaya A, Kaynak K, Tanju S, Toker A and Buyru N: WWOX gene may contribute to progression of non-small-cell lung cancer (NSCLC). Tumour Biol. 31:315–320. 2010. View Article : Google Scholar : PubMed/NCBI
14	Del MS, Husanie H, Iancu O, Abu-Odeh M, Evangelou K, Lovat F, Volinia S, Gordon J, Amir G, Stein J, et al: WWOX and p53 dysregulation synergize to drive the development of osteosarcoma. Cancer Res. 76:6107–6117. 2016. View Article : Google Scholar : PubMed/NCBI
15	Aderca I, Moser CD, Veerasamy M, Bani-Hani AH, Bonilla-Guerrero R, Ahmed K, Shire A, Cazanave SC, Montoya DP, Mettler TA, et al: The JNK inhibitor SP600129 enhances apoptosis of HCC cells induced by the tumor suppressor WWOX. J Hepatol. 49:373–383. 2008. View Article : Google Scholar : PubMed/NCBI
16	Chang NS, Doherty J, Ensign A, Lewis J, Heath J, Schultz L, Chen ST and Oppermann U: Molecular mechanisms underlying WOX1 activation during apoptotic and stress responses. Biochem Pharmacol. 66:1347–1354. 2003. View Article : Google Scholar : PubMed/NCBI
17	Tsai CW, Lai FJ, Sheu HM, Lin YS, Chang TH, Jan MS, Chen SM, Hsu PC, Huang TT, Huang TC, et al: WWOX suppresses autophagy for inducing apoptosis in methotrexate-treated human squamous cell carcinoma. Cell Death Dis. 4:e7922013. View Article : Google Scholar : PubMed/NCBI
18	Kong W, Mao J, Yang Y, Yuan J, Chen J, Luo Y, Lai T and Zuo L: Mechanisms of mTOR and autophagy in human endothelial cell infected with dengue Virus-2. Viral Immunol. 33:61–70. 2020. View Article : Google Scholar : PubMed/NCBI
19	Jia J, Abudu YP, Claude-Taupin A, Gu Y, Kumar S, Choi SW, Peters R, Mudd MH, Allers L, Salemi M, et al: Galectins control MTOR and AMPK in response to lysosomal damage to induce autophagy. Autophagy. 15:169–171. 2019. View Article : Google Scholar : PubMed/NCBI
20	Munson MJ and Ganley IG: MTOR, PIK3C3, and autophagy: Signaling the beginning from the end. Autophagy. 11:2375–2376. 2015. View Article : Google Scholar : PubMed/NCBI
21	simplehttps://www.atcc.org/products/all/HTB-77.aspx#characteristics
22	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using Real-Time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
23	Hu BS, Tan JW, Zhu GH, Wang DF, Zhou X and Sun ZQ: WWOX induces apoptosis and inhibits proliferation of human hepatoma cell line SMMC-7721. World J Gastroenterol. 18:3020–3026. 2012. View Article : Google Scholar : PubMed/NCBI
24	Lo JY, Chou YT, Lai FJ and Hsu LJ: Regulation of cell signaling and apoptosis by tumor suppressor WWOX. Exp Biol Med (Maywood). 240:383–391. 2015. View Article : Google Scholar : PubMed/NCBI
25	Ekizoglu S, Bulut P, Karaman E, Kilic E and Buyru N: Epigenetic and genetic alterations affect the WWOX gene in head and neck squamous cell carcinoma. PLoS One. 10:e01153532015. View Article : Google Scholar : PubMed/NCBI
26	Chen X, Li P, Yang Z and Mo WN: Expression of fragile histidine triad (FHIT) and WW-domain oxidoreductase gene (WWOX) in nasopharyngeal carcinoma. Asian Pac J Cancer Prev. 14:165–171. 2013. View Article : Google Scholar : PubMed/NCBI
27	Barth S, Glick D and Macleod KF: Autophagy: Assays and artifacts. J Pathol. 221:117–124. 2010. View Article : Google Scholar : PubMed/NCBI
28	Schmitz KJ, Ademi C, Bertram S, Schmid KW and Baba HA: Prognostic relevance of autophagy-related markers LC3, p62/sequestosome 1, Beclin-1 and ULK1 in colorectal cancer patients with respect to KRAS mutational status. World J Surg Oncol. 14:1892016. View Article : Google Scholar : PubMed/NCBI
29	Li RF, Chen G, Ren JG, Zhang W, Wu ZX, Liu B, Zhao Y and Zhao YF: The adaptor protein p62 is involved in RANKL-induced autophagy and osteoclastogenesis. J Histochem Cytochem. 62:879–888. 2014. View Article : Google Scholar : PubMed/NCBI
30	Kim YC and Guan KL: mTOR: A pharmacologic target for autophagy regulation. J Clin Invest. 125:25–32. 2015. View Article : Google Scholar : PubMed/NCBI
31	Noureldein MH and Eid AA: Gut microbiota and mTOR signaling: Insight on a new pathophysiological interaction. Microb Pathog. 118:98–104. 2018. View Article : Google Scholar : PubMed/NCBI
32	Yan H, Tong J, Lin X, Han Q and Huang H: Effect of the WWOX gene on the regulation of the cell cycle and apoptosis in human ovarian cancer stem cells. Mol Med Rep. 12:1783–1788. 2015. View Article : Google Scholar : PubMed/NCBI
33	Lin JT, Li HY, Chang NS, Lin CH, Chen YC and Lu PJ: WWOX suppresses prostate cancer cell progression through cyclin D1-mediated cell cycle arrest in the G1 phase. Cell Cycle. 14:408–416. 2015. View Article : Google Scholar : PubMed/NCBI
34	Chiang MF, Chou PY, Wang WJ, Sze CI and Chang NS: Tumor suppressor WWOX and p53 alterations and drug resistance in glioblastomas. Front Oncol. 3:432013. View Article : Google Scholar : PubMed/NCBI
35	Płuciennik E, Nowakowska M, Gałdyszyńska M, Popęda M and Bednarek AK: The influence of the WWOX gene on the regulation of biological processes during endometrial carcinogenesis. Int J Mol Med. 37:807–815. 2016. View Article : Google Scholar : PubMed/NCBI
36	Baryła I, Styczeń-Binkowska E and Bednarek AK: Alteration of WWOX in human cancer: A clinical view. Exp Biol Med (Maywood). 240:305–314. 2015. View Article : Google Scholar : PubMed/NCBI
37	Hung PS, Chuang FJ, Chen CY, Chou CH, Tu HF and Lo SS: miR-187* enhances SiHa cervical cancer cell oncogenicity via suppression of WWOX. Anticancer Res. 40:1427–1436. 2020. View Article : Google Scholar : PubMed/NCBI
38	Nakayama S, Semba S, Maeda N, Aqeilan RI, Huebner K and Yokozaki H: Role of the WWOX gene, encompassing fragile region FRA16D, in suppression of pancreatic carcinoma cells. Cancer Sci. 99:1370–1376. 2008. View Article : Google Scholar : PubMed/NCBI
39	Khawaled S, Suh SS, Abdeen SK, Monin J, Distefano R, Nigita G, Croce CM and Aqeilan RI: WWOX inhibits metastasis of triple-negative breast cancer cells via modulation of miRNAs. Cancer Res. 79:1784–1798. 2019. View Article : Google Scholar : PubMed/NCBI
40	Kimmelman AC and White E: Autophagy and tumor metabolism. Cell Metab. 25:1037–1043. 2017. View Article : Google Scholar : PubMed/NCBI
41	O'Donovan TR, O'Sullivan GC and McKenna SL: Induction of autophagy by drug-resistant esophageal cancer cells promotes their survival and recovery following treatment with chemotherapeutics. Autophagy. 7:509–524. 2011. View Article : Google Scholar : PubMed/NCBI
42	Zhang SF, Wang XY, Fu ZQ, Peng QH, Zhang JY, Ye F, Fu YF, Zhou CY, Lu WG, Cheng XD and Xie X: TXNDC17 promotes paclitaxel resistance via inducing autophagy in ovarian cancer. Autophagy. 11:225–238. 2015. View Article : Google Scholar : PubMed/NCBI
43	Li LQ, Xie WJ, Pan D, Chen H and Zhang L: Inhibition of autophagy by bafilomycin A1 promotes chemosensitivity of gastric cancer cells. Tumour Biol. 37:653–659. 2016. View Article : Google Scholar : PubMed/NCBI
44	Polishchuk EV, Merolla A, Lichtmannegger J, Romano A, Indrieri A, Ilyechova EY, Concilli M, De Cegli R, Crispino R, Mariniello M, et al: Activation of autophagy, observed in liver tissues from patients with Wilson disease and from ATP7B-deficient animals, protects hepatocytes from copper-induced apoptosis. Gastroenterology. 156:1173–1189. 2019. View Article : Google Scholar : PubMed/NCBI