The influence of the WWOX gene on the regulation of biological processes during endometrial carcinogenesis

Płuciennik,E.; Nowakowska,M.; Gałdyszyńska,M.; Popęda,M.; Bednarek,A. K.

doi:10.3892/ijmm.2016.2469

The influence of the WWOX gene on the regulation of biological processes during endometrial carcinogenesis

Authors:
- E. Płuciennik
- M. Nowakowska
- M. Gałdyszyńska
- M. Popęda
- A. K. Bednarek
View Affiliations

Affiliations: Department of Molecular Carcinogenesis, Medical University of Lodz, 90-752 Lodz, Poland, Department of Comparative Endocrinology, Medical University of Lodz, 90-752 Lodz, Poland, Faculty of Biomedical Sciences and Postgraduate Education, Medical University of Lodz, 90-752 Lodz, Poland
Published online on: January 27, 2016 https://doi.org/10.3892/ijmm.2016.2469
Pages: 807-815

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

The purpose of the present study was to investigate the role of WW domain containing oxidoreductase (WWOX) downregulation in biological cancer-related processes in normal (non-malignant) and cancer endometrial cell lines. We created an in vitro model using the normal endometrial cell line, THESC, and 2 endometrial cancer cell lines with varying degrees of differentiation, the Ishikawa (well-differentiated) and the MFE296 (moderately differentiated) cells, in which the WWOX tumor suppressor gene was silenced using Gipz lentiviral shRNA. In this model, we examined the changes in invasiveness via biological assays, such as zymography, migration through a basement membrane, the adhesion of cells to extracellular matrix proteins, anchorage-independent growth and colony formation assay. We also evaluated the correlation between the mRNA expression of the WWOX gene and genes involved in the processes of carcinogenesis, namely catenin beta-1 (CTNNB1) and zinc finger E-box binding homeobox 1 (ZEB1) (gene transcription), cadherin 1 (CDH1) and ezrin (EZR) (cell adhesion), vimentin (VIM) (structural proteins), as well as phosphatase and tensin homolog (PTEN) (tumor suppression) and secreted protein, acidic, cysteine-rich (osteonectin) (SPARC) (SPARC) (cell growth regulation) by RT-qPCR. Downregulation of the WWOX gene in the moderately differentiated MFE296 cell line caused decreased migratory capacity, and a reduction of matrix metalloproteinase-2 (MMP-2) activity. However, these cells grew in semisolid medium and exhibited higher expression of CDH1 and EZR (cell adhesion) and secreted protein, acidic, cysteine-rich (osteonectin) (SPARC) (cell growth regulation). Moreover, in the well-differentiated endometrial cancer (Ishikawa) cell line, WWOX gene silencing resulted in an increased ability of the cells to proliferate indefinitely. Additionally, WWOX regulated changes in adhesion potential in both the normal and cancer cell lines. Our results suggest that the WWOX tumor suppressor gene modulated the processes of cell motility, cell adhesion, gene expression and remodeling in endometrial cell lines.

View Figures

View References

1	Ministry of Health: National Cancer Registry of Poland: Reports based on data of National Cancer Registry of Poland. (updated 19-11-2010). http://epid.coi.waw.pl/krn/english/index.asp. Accessed June 30, 2012.
2	Bokhman JV: Two pathogenetic types of endometrial carcinoma. Gynecol Oncol. 15:10–17. 1983. View Article : Google Scholar : PubMed/NCBI
3	Doll A, Abal M, Rigau M, Monge M, Gonzalez M, Demajo S, Colás E, Llauradó M, Alazzouzi H, Planagumá J, et al: Novel molecular profiles of endometrial cancer-new light through old windows. J Steroid Biochem Mol Biol. 108:221–229. 2008. View Article : Google Scholar
4	Ryan AJ, Susil B, Jobling TW and Oehler MK: Endometrial cancer. Cell Tissue Res. 322:53–61. 2005. View Article : Google Scholar : PubMed/NCBI
5	O'Hara AJ and Bell DW: The genomics and genetics of endometrial cancer. Adv Genomics Genet. 2012:33–47. 2012.PubMed/NCBI
6	Okuda T, Sekizawa A, Purwosunu Y, Nagatsuka M, Morioka M, Hayashi M and Okai T: Genetics of endometrial cancers. Obstet Gynecol Int. 2010:9840132010. View Article : Google Scholar : PubMed/NCBI
7	Merritt MA and Cramer DW: Molecular pathogenesis of endometrial and ovarian cancer. Cancer Biomark. 9:287–305. 2010.
8	Ferguson BW, Gao X, Zelazowski MJ, Lee J, Jeter CR, Abba MC and Aldaz CM: The cancer gene WWOX behaves as an inhibitor of SMAD3 transcriptional activity via direct binding. BMC Cancer. 13:5932013. View Article : Google Scholar : PubMed/NCBI
9	Kurek KC, Del Mare S, Salah Z, Abdeen S, Sadiq H, Lee SH, Gaudio E, Zanesi N, Jones KB, DeYoung B, et al: Frequent attenuation of the WWOX tumor suppressor in osteosarcoma is associated with increased tumorigenicity and aberrant RUNX2 expression. Cancer Res. 70:5577–5586. 2010. View Article : Google Scholar : PubMed/NCBI
10	Gaudio E, Palamarchuk A, Palumbo T, Trapasso F, Pekarsky Y, Croce CM and Aqeilan RI: Physical association with WWOX suppresses c-Jun transcriptional activity. Cancer Res. 66:11585–11589. 2006. View Article : Google Scholar : PubMed/NCBI
11	Aqeilan RI, Pekarsky Y, Herrero JJ, Palamarchuk A, Letofsky J, Druck T, Trapasso F, Han SY, Melino G, Huebner K, et al: Functional association between Wwox tumor suppressor protein and p73, a p53 homolog. Proc Natl Acad Sci USA. 101:4401–4406. 2004. View Article : Google Scholar : PubMed/NCBI
12	Aqeilan RI, Palamarchuk A, Weigel RJ, Herrero JJ, Pekarsky Y and Croce CM: Physical and functional interactions between the Wwox tumor suppressor protein and the AP-2gamma transcription factor. Cancer Res. 64:8256–8261. 2004. View Article : Google Scholar : PubMed/NCBI
13	Aqeilan RI, Donati V, Palamarchuk A, Trapasso F, Kaou M, Pekarsky Y, Sudol M and Croce CM: WW domain-containing proteins, WWOX and YAP, compete for interaction with ErbB-4 and modulate its transcriptional function. Cancer Res. 65:6764–6772. 2005. View Article : Google Scholar : PubMed/NCBI
14	Aqeilan RI, Donati V, Gaudio E, Nicoloso MS, Sundvall M, Korhonen A, Lundin J, Isola J, Sudol M, Joensuu H, et al: Association of Wwox with ErbB4 in breast cancer. Cancer Res. 67:9330–9336. 2007. View Article : Google Scholar : PubMed/NCBI
15	Abu-Remaileh M and Aqeilan RI: Tumor suppressor WWOX regulates glucose metabolism via HIF1α modulation. Cell Death Differ. 21:1805–1814. 2014. View Article : Google Scholar : PubMed/NCBI
16	Aqeilan RI and Croce CM: WWOX in biological control and tumorigenesis. J Cell Physiol. 212:307–310. 2007. View Article : Google Scholar : PubMed/NCBI
17	Aqeilan RI, Hagan JP, de Bruin A, Rawahneh M, Salah Z, Gaudio E, Siddiqui H, Volinia S, Alder H, Lian JB, et al: Targeted ablation of the WW domain-containing oxidoreductase tumor suppressor leads to impaired steroidogenesis. Endocrinology. 150:1530–1535. 2009. View Article : Google Scholar :
18	Bednarek AK, Laflin KJ, Daniel RL, Liao Q, Hawkins KA and Aldaz CM: WWOX, a novel WW domain-containing protein mapping to human chromosome 16q23.3-24.1, a region frequently affected in breast cancer. Cancer Res. 60:2140–2145. 2000.PubMed/NCBI
19	Lan C, Chenggang W, Yulan B, Xiaohui D, Junhui Z and Xiao W: Aberrant expression of WWOX protein in epithelial ovarian cancer: A clinicopathologic and immunohistochemical study. Int J Gynecol Pathol. 31:125–132. 2012. View Article : Google Scholar : PubMed/NCBI
20	Qin HR, Iliopoulos D, Nakamura T, Costinean S, Volinia S, Druck T, Sun J, Okumura H and Huebner K: Wwox suppresses prostate cancer cell growth through modulation of ErbB2-mediated androgen receptor signaling. Mol Cancer Res. 5:957–965. 2007. View Article : Google Scholar : PubMed/NCBI
21	Płuciennik E, Kusińska R, Potemski P, Kubiak R, Kordek R and Bednarek AK: WWOX--the FRA16D cancer gene: Expression correlation with breast cancer progression and prognosis. Eur J Surg Oncol. 32:153–157. 2006. View Article : Google Scholar
22	Płuciennik E, Kośla K, Wójcik-Krowiranda K, Bieńkiewicz A and Bednarek AK: The WWOX tumor suppressor gene in endometrial adenocarcinoma. Int J Mol Med. 32:1458–1464. 2013.
23	Cui Z, Lin D, Cheng F, Luo L, Kong L, Xu J, Hu J and Lan F: The role of the WWOX gene in leukemia and its mechanisms of action. Oncol Rep. 29:2154–2162. 2013.PubMed/NCBI
24	Ishii H, Vecchione A, Furukawa Y, Sutheesophon K, Han SY, Druck T, Kuroki T, Trapasso F, Nishimura M, Saito Y, et al: Expression of FRA16D/WWOX and FRA3B/FHIT genes in hematopoietic malignancies. Mol Cancer Res. 1:940–947. 2003.PubMed/NCBI
25	Liu SY, Chiang MF and Chen YJ: Role of WW domain proteins WWOX in development, prognosis, and treatment response of glioma. Exp Biol Med (Maywood). 240:315–323. 2015. View Article : Google Scholar
26	Nowakowska M, Pospiech K, Lewandowska U, Piastowska-Ciesielska AW and Bednarek AK: Diverse effect of WWOX overexpression in HT29 and SW480 colon cancer cell lines. Tumour Biol. 35:9291–9301. 2014. View Article : Google Scholar : PubMed/NCBI
27	Xie B, Zen Q, Wang X, He X, Xie Y, Zhang Z and Li H: ACK1 promotes hepatocellular carcinoma progression via downregulating WWOX and activating AKT signaling. Int J Oncol. 46:2057–2066. 2015.PubMed/NCBI
28	Yan H and Sun Y: Evaluation of the mechanism of epithelial-mesenchymal transition in human ovarian cancer stem cells transfected with a WW domain-containing oxidoreductase gene. Oncol Lett. 8:426–430. 2014.PubMed/NCBI
29	Płuciennik E, Nowakowska M, Pospiech K, Stępień A, Wołkowicz M, Gałdyszyńska M, Popęda M, Wójcik-Krowiranda K, Bieńkiewicz A and Bednarek AK: The role of WWOX tumor suppressor gene in the regulation of EMT process via regulation of CDH1-ZEB1-VIM expression in endometrial cancer. Int J Oncol. 46:2639–2648. 2015.
30	Pfaffl MW, Horgan GW and Dempfle L: Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res. 30:e362002. View Article : Google Scholar : PubMed/NCBI
31	Aqeilan RI, Kuroki T, Pekarsky Y, Albagha O, Trapasso F, Baffa R, Huebner K, Edmonds P and Croce CM: Loss of WWOX expression in gastric carcinoma. Clin Cancer Res. 10:3053–3058. 2004. View Article : Google Scholar : PubMed/NCBI
32	Nunez MI, Rosen DG, Ludes-Meyers JH, Abba MC, Kil H, Page R, Klein-Szanto AJ, Godwin AK, Liu J, Mills GB, et al: WWOX protein expression varies among ovarian carcinoma histotypes and correlates with less favorable outcome. BMC Cancer. 5:642005. View Article : Google Scholar : PubMed/NCBI
33	Qu J, Lu W, Li B, Lu C and Wan X: WWOX induces apoptosis and inhibits proliferation in cervical cancer and cell lines. Int J Mol Med. 31:1139–1147. 2013.PubMed/NCBI
34	Bednarek AK, Keck-Waggoner CL, Daniel RL, Laflin KJ, Bergsagel PL, Kiguchi K, Brenner AJ and Aldaz CM: WWOX, the FRA16D gene, behaves as a suppressor of tumor growth. Cancer Res. 61:8068–8073. 2001.PubMed/NCBI
35	Nunez MI, Ludes-Meyers J, Abba MC, Kil H, Abbey NW, Page RE, Sahin A, Klein-Szanto AJ and Aldaz CM: Frequent loss of WWOX expression in breast cancer: Correlation with estrogen receptor status. Breast Cancer Res Treat. 89:99–105. 2005. View Article : Google Scholar : PubMed/NCBI
36	Göthlin Eremo A, Wegman P, Stål O, Nordenskjöld B, Fornander T and Wingren S: Wwox expression may predict benefit from adjuvant tamoxifen in randomized breast cancer patients. Oncol Rep. 29:1467–1474. 2013.PubMed/NCBI
37	Schwenke M, Knöfler M, Velicky P, Weimar CH, Kruse M, Samalecos A, Wolf A, Macklon NS, Bamberger AM and Gellersen B: Control of human endometrial stromal cell motility by PDGF-BB, HB-EGF and trophoblast-secreted factors. PLoS One. 8:e543362013. View Article : Google Scholar : PubMed/NCBI
38	Desgrosellier JS and Cheresh DA: Integrins in cancer: biological implications and therapeutic opportunities. Nat Rev Cancer. 10:9–22. 2010. View Article : Google Scholar
39	Li X, Regezi J, Ross FP, Blystone S, Ilić D, Leong SP and Ramos DM: Integrin alphavbeta3 mediates K1735 murine melanoma cell motility in vivo and in vitro. J Cell Sci. 114:2665–2672. 2001.PubMed/NCBI
40	Abdeen SK, Salah Z, Khawaled S and Aqeilan RI: Characterization of WWOX inactivation in murine mammary gland development. J Cell Physiol. 228:1391–1396. 2013. View Article : Google Scholar
41	Gourley C, Paige AJ, Taylor KJ, Ward C, Kuske B, Zhang J, Sun M, Janczar S, Harrison DJ, Muir M, et al: WWOX gene expression abolishes ovarian cancer tumorigenicity in vivo and decreases attachment to fibronectin via integrin α3. Cancer Res. 69:4835–4842. 2009. View Article : Google Scholar : PubMed/NCBI
42	Zhang JQ, Li L, Song HL, Paige A and Gabra H: Effect of WWOX gene on the attachment and adhesion of ovarian cancer cells. Zhonghua Fu Chan Ke Za Zhi. 44:529–532. 2009.In Chinese. PubMed/NCBI
43	Palumbo JS and Degen JL: Fibrinogen and tumor cell metastasis. Haemostasis. 31(Suppl 1): S11–S15. 2001.
44	Lessey BA, Castelbaum AJ, Buck CA, Lei Y, Yowell CW and Sun J: Further characterization of endometrial integrins during the menstrual cycle and in pregnancy. Fertil Steril. 62:497–506. 1994.PubMed/NCBI
45	Rebhun RB, Cheng H, Gershenwald JE, Fan D, Fidler IJ and Langley RR: Constitutive expression of the alpha4 integrin correlates with tumorigenicity and lymph node metastasis of the B16 murine melanoma. Neoplasia. 12:173–182. 2010. View Article : Google Scholar : PubMed/NCBI
46	Holzmann B1, Gosslar U and Bittner M: Alpha 4 integrins and tumor metastasis. Curr Top Microbiol Immunol. 231:125–141. 1998.PubMed/NCBI
47	Jin H, Aiyer A, Su J, Borgstrom P, Stupack D, Friedlander M and Varner J: A homing mechanism for bone marrow-derived progenitor cell recruitment to the neovasculature. J Clin Invest. 116:652–662. 2006. View Article : Google Scholar : PubMed/NCBI
48	Garmy-Susini B, Jin H, Zhu Y, Sung RJ, Hwang R and Varner J: Integrin α4β1-VCAM-1-mediated adhesion between endothelial and mural cells is required for blood vessel maturation. J Clin Invest. 115:1542–1551. 2005. View Article : Google Scholar : PubMed/NCBI
49	Prifti S, Zourab Y, Koumouridis A, Bohlmann M, Strowitzki T and Rabe T: Role of integrins in invasion of endometrial cancer cell lines. Gynecol Oncol. 84:12–20. 2002. View Article : Google Scholar
50	Lessey BA, Albelda S, Buck CA, Castelbaum AJ, Yeh I, Kohler M and Berchuck A: Distribution of integrin cell adhesion molecules in endometrial cancer. Am J Pathol. 146:717–726. 1995.PubMed/NCBI
51	Lewandowska U, Żelazowski M, Seta K, Byczewska M, Płuciennik E and Bednarek AK: WWOX, the tumour suppressor gene affected in multiple cancer. J Physiol Pharmacol. 60(Suppl 1): 47–56. 2009.
52	Martin TA and Jiang WG: Loss of tight junction barrier function and its role in cancer metastasis. Biochim Biophys Acta. 1788:872–891. 2009. View Article : Google Scholar
53	Neuzillet C, Tijeras-Raballand A, Cros J, Faivre S, Hammel P and Raymond E: Stromal expression of SPARC in pancreatic adenocarcinoma. Cancer Metastasis Rev. 32:585–602. 2013. View Article : Google Scholar : PubMed/NCBI
54	Yusuf N, Inagaki T, Kusunoki S, Okabe H, Yamada I, Matsumoto A, Terao Y, Takeda S and Kato K: SPARC was overexpressed in human endometrial cancer stem-like cells and promoted migration activity. Gynecol Oncol. 134:356–363. 2014. View Article : Google Scholar : PubMed/NCBI
55	Köbel M, Langhammer T, Hüttelmaier S, Schmitt WD, Kriese K, Dittmer J, Strauss HG, Thomssen C and Hauptmann S: Ezrin expression is related to poor prognosis in FIGO stage I endometrioid carcinomas. Mod Pathol. 19:581–587. 2006. View Article : Google Scholar : PubMed/NCBI
56	Ohtani K, Sakamoto H, Rutherford T, Chen Z, Kikuchi A, Yamamoto T, Satoh K and Naftolin F: Ezrin, a membrane-cytoskeletal linking protein, is highly expressed in atypical endometrial hyperplasia and uterine endometrioid adenocarcinoma. Cancer Lett. 179:79–86. 2002. View Article : Google Scholar : PubMed/NCBI