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Three-Dimensional Cellular Arrangement in Epithelial Ovarian Cancer Cell Lines TOV-21G and SKOV-3 is Associated with Apoptosis-Related miRNA Expression Modulation

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Cancer Microenvironment

Abstract

Epithelial ovarian cancer (EOC) is the most lethal gynecological malignancy, and the lack of chemoresistance biomarkers contributes to the poor prognosis. Cancer stem cells (CSC) have been investigated in EOC to understand its relationship with chemoresistance and recurrence. In this context, in vitro cultivation-models are important tools for CSC studies. MicroRNAs (miRNAs) play key roles in cancer, CSC regulation and apoptosis. Thus, this study aims to evaluate the tumorsphere model as CSC-enrichment method in EOC studies and investigate apoptosis-related miRNAs in tumorspheres-derived EOC cell lines. TOV-21G and SKOV-3 were cultured in monolayer and tumorspheres. Genetic profiles of cell lines were obtained using COSMIC database. CD24/CD44/CD146/CD177 and ALDH1 markers were evaluated in cell lines and tumorspheres-derived by flow cytometry. Eleven miRNAs were selected by in silico analysis for qPCR analysis. According to COSMIC, TOV-21G and SKOV-3 have eight and nine cancer-related mutations, respectively. TOV-21G showed a CD44+/high/CD24−/low/CD117−/low/CD146−/low/ALDH1low profile in both culture models; thus, no significant difference between cultivation models was identified. SKOV-3 showed a CD44+/high/CD24+/high/ CD117−/low/CD146−/low/ALDH1low profile in both culture models, although the tumorsphere model showed a significant increase in CD24+/high subpopulation (ovarian CSC-like). Among eleven miRNAs, we observed differences in miRNA expression between culture models. MiR-26a was overexpressed in TOV-21G tumorspheres, albeit downregulated in SKOV-3 tumorspheres. MiR-125b-5p, miR-17-5p and miR-221 was downregulated in tumorsphere model in both cell lines. Given that tumorsphere-derived SKOV-3 had a higher ratio of CD24+/high cells, we suggest that miR-26a, miR-125b-5p, miR-17-5p and miR-221 downregulation could be related to poor EOC prognosis.

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References

  1. Garson K, Vanderhyden BC (2015) Epithelial ovarian cancer stem cells: underlying complexity of a simple paradigm. Reproduction 149:R59–R70. https://doi.org/10.1530/REP-14-0234

    Article  PubMed  CAS  Google Scholar 

  2. KK A, Josahkian JA, Francis J-A et al (2015) Current state of biomarkers in ovarian cancer prognosis. Future Oncol 11:3187–3195. https://doi.org/10.2217/fon.15.251

    Article  CAS  Google Scholar 

  3. Kwon MJ, Shin YK (2013) Regulation of ovarian cancer stem cells or tumor-initiating cells. Int J Mol Sci 14:6624–6648. https://doi.org/10.3390/ijms14046624

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  4. Olivos DJ, Mayo LD (2016) Emerging non-canonical functions and regulation by p53: p53 and stemness. Int J Mol Sci 17:1–30. https://doi.org/10.3390/ijms17121982

    Article  Google Scholar 

  5. Ponti D, Costa A, Zaffaroni N et al (2005) Isolation and in vitro propagation of tumorigenic breast cancer cells with stem / progenitor cell properties. Cancer Res 65:5506–5512. https://doi.org/10.1158/0008-5472.CAN-05-0626

    Article  PubMed  CAS  Google Scholar 

  6. Weiswald LB, Bellet D, Dangles-Marie V (2015) Spherical cancer models in tumor biology. Neoplasia 17:1–15. https://doi.org/10.1016/j.neo.2014.12.004

    Article  PubMed  PubMed Central  Google Scholar 

  7. Chen D, Zhang Y, Wang J et al (2013) MicroRNA-200c overexpression inhibits tumorigenicity and metastasis of CD117+CD44+ ovarian cancer stem cells by regulating epithelial-mesenchymal transition. J Ovarian Res 6:50. https://doi.org/10.1186/1757-2215-6-50

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  8. Takahashi R-U, Miyazaki H, Ochiya T (2014) The role of microRNAs in the regulation of cancer stem cells. Front Genet 4:295. https://doi.org/10.3389/fgene.2013.00295

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  9. Laios A, O’Toole S, Flavin R et al (2008) Potential role of miR-9 and miR-223 in recurrent ovarian cancer. Mol Cancer 7:35. https://doi.org/10.1186/1476-4598-7-35

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  10. Vilming Elgaaen B, Olstad OK, Haug KBF et al (2014) Global miRNA expression analysis of serous and clear cell ovarian carcinomas identifies differentially expressed miRNAs including miR-200c-3p as a prognostic marker. BMC Cancer 14:80. https://doi.org/10.1186/1471-2407-14-80

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  11. Braga LC, Ramos APAS, Traiman P et al (2012) TRAIL-R3-related apoptosis: epigenetic and expression analyses in women with ovarian neoplasia. Gynecol Oncol 126:268–273. https://doi.org/10.1016/j.ygyno.2012.04.038

    Article  CAS  Google Scholar 

  12. Braga LC, Silva LM, Piedade JB et al (2014) Epigenetic and expression analysis of TRAIL-R2 and BCL2: on the TRAIL to knowledge of apoptosis in ovarian tumors. Arch Gynecol Obstet 289:1061–1069. https://doi.org/10.1007/s00404-013-3060-0

    Article  CAS  Google Scholar 

  13. Braga LC, Silva LM, Ramos APÁ d S et al (2014) Single CpG island methylation is not sufficient to maintain the silenced expression of CASPASE-8 apoptosis-related gene among women with epithelial ovarian cancer. Biomed Pharmacother 68:87–91. https://doi.org/10.1016/j.biopha.2013.12.004

    Article  CAS  Google Scholar 

  14. Iorio MV, Croce CM (2012) MicroRNA dysregulation in cancer: diagnostics, monitoring and therapeutics. A comprehensive review. EMBO Mol Med 4:143–159. https://doi.org/10.1002/emmm.201100209

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  15. House CD, Hernandez L, Annunziata CM (2015) In vitro enrichment of ovarian cancer tumor-initiating cells. J Vis Exp:1–8. https://doi.org/10.3791/52446

  16. Forbes SA, Beare D, Gunasekaran P et al (2015) COSMIC: exploring the world’s knowledge of somatic mutations in human cancer. Nucleic Acids Res 43:D805–D811. https://doi.org/10.1093/nar/gku1075

    Article  PubMed  CAS  Google Scholar 

  17. Griffiths-Jones S, Grocock RJ, van Dongen S et al (2006) miRBase: microRNA sequences, targets and gene nomenclature. Nucleic Acids Res 34:D140–D144. https://doi.org/10.1093/nar/gkj112

    Article  PubMed  CAS  Google Scholar 

  18. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and. Methods 25:402–408. https://doi.org/10.1006/meth.2001.1262

    Article  PubMed  CAS  Google Scholar 

  19. Coleman RL, Monk BJ, Sood AK, Herzog TJ (2013) Latest research and clinical treatment of advanced-stage epithelial ovarian cancer. Nat Rev Clin Oncol 10:211–224. https://doi.org/10.1038/nrclinonc.2013.5.Latest

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  20. Toss A, Tomasello C, Razzaboni E et al (2015) Hereditary ovarian cancer: not only BRCA 1 and 2 genes. Biomed Res Int 2015:341723. https://doi.org/10.1155/2015/341723

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  21. Kurman R, Shih I (2010) The origin and pathogenesis of epithelial ovarian cancer-a proposed unifying theory. Am J Surg Pathol 34:433–443. https://doi.org/10.1097/PAS.0b013e3181cf3d79.The

    Article  PubMed  PubMed Central  Google Scholar 

  22. Calvet CY, André FM, Mir LM (2014) The culture of cancer cell lines as tumorspheres does not systematically result in cancer stem cell enrichment. PLoS One 9:e89644. https://doi.org/10.1371/journal.pone.0089644

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  23. Zhao J (2016) Cancer stem cells and chemoresistance: the smartest survives the raid. Pharmacol Ther 160:145–158. https://doi.org/10.1016/j.pharmthera.2016.02.008

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  24. Luna JI, Grossenbacher SK, Murphy WJ, Canter RJ (2016) Targeting cancer stem cells with natural killer cell immunotherapy. Expert Opin Biol Ther 17:313–324. https://doi.org/10.1080/14712598.2017.1271874

  25. Jaggupilli A, Elkord E (2012) Significance of CD44 and CD24 as cancer stem cell markers: an enduring ambiguity. Clin Dev Immunol 2012:708036. https://doi.org/10.1155/2012/708036

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  26. Huang L, Lv W, Zhao X (2016) CD24 as a molecular marker in ovarian cancer: a literature review. Cancer Transl Med 2:29. https://doi.org/10.4103/2395-3977.177563

    Article  Google Scholar 

  27. Iglesias JM, Beloqui I, Garcia-Garcia F et al (2013) Mammosphere formation in breast carcinoma cell lines depends upon expression of E-cadherin. PLoS One 8:1–12. https://doi.org/10.1371/journal.pone.0077281

    Article  CAS  Google Scholar 

  28. Gao MQ, Choi YP, Kang S et al (2010) CD24+ cells from hierarchically organized ovarian cancer are enriched in cancer stem cells. Oncogene 29:2672–2680. https://doi.org/10.1038/onc.2010.35

    Article  PubMed  CAS  Google Scholar 

  29. Nakamura K, Terai Y, Tanabe A et al (2017) CD24 expression is a marker for predicting clinical outcome and regulates the epithelial-mesenchymal transition in ovarian cancer via both the Akt and ERK pathways. Oncol Rep 37:3189–3200. https://doi.org/10.3892/or.2017.5583

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  30. Ishiguro K, Zhu Y-L, Lin ZP et al (2016) Cataloging antineoplastic agents according to their effectiveness against platinum-resistant and platinum-sensitive ovarian carcinoma cell lines. J Transl Sci 2:117–124. https://doi.org/10.15761/JTS.1000127

  31. Mulrane L, McGee SF, Gallagher WM, O’Connor DP (2013) miRNA dysregulation in breast cancer. Cancer Res 73:6554–6562. https://doi.org/10.1158/0008-5472.CAN-13-1841

    Article  PubMed  CAS  Google Scholar 

  32. Sethi S, Ali S, Sethi S, Sarkar FH (2014) MicroRNAs in personalized cancer therapy. Clin Genet 86:68–73. https://doi.org/10.1111/cge.12362

    Article  PubMed  CAS  Google Scholar 

  33. Subramanian S, Steer CJ (2010) MicroRNAs as gatekeepers of apoptosis. J Cell Physiol 223:289–298. https://doi.org/10.1002/jcp.22066

    Article  PubMed  CAS  Google Scholar 

  34. Li C, Hashimi SM, Good DA et al (2012) Apoptosis and microRNA aberrations in cancer. Clin Exp Pharmacol Physiol 39:739–746. https://doi.org/10.1111/j.1440-1681.2012.05700.x

    Article  PubMed  CAS  Google Scholar 

  35. Jovanovic M, Hengartner MO (2006) miRNAs and apoptosis: RNAs to die for. Oncogene 25:6176–6187. https://doi.org/10.1038/sj.onc.1209912

    Article  PubMed  CAS  Google Scholar 

  36. Ouyang L, Shi Z, Zhao S et al (2012) Programmed cell death pathways in cancer: a review of apoptosis, autophagy and programmed necrosis. Cell Prolif 45:487–498. https://doi.org/10.1111/j.1365-2184.2012.00845.x

    Article  PubMed  CAS  Google Scholar 

  37. Ayala-Ortega E, Arzate-Mejía R, Pérez-Molina R et al (2016) Epigenetic silencing of miR-181c by DNA methylation in glioblastoma cell lines. BMC Cancer 16:226. https://doi.org/10.1186/s12885-016-2273-6

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  38. Yang D, Zhan M, Chen T et al (2017) miR-125b-5p enhances chemotherapy sensitivity to cisplatin by down-regulating Bcl2 in gallbladder cancer. Sci Rep 7:43109. https://doi.org/10.1038/srep43109

    Article  PubMed  PubMed Central  Google Scholar 

  39. Zhang Y, Johansson E, Miller ML et al (2011) Identification of a conserved anti-apoptotic protein that modulates the mitochondrial apoptosis pathway. PLoS One 6:e25284. https://doi.org/10.1371/journal.pone.0025284

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  40. Li H, Zhou H, Luo J, Huang J (2017) MicroRNA-17-5p inhibits proliferation and triggers apoptosis in non-small cell lung cancer by targeting transforming growth factor β receptor 2. Exp Ther Med 13:2715–2722. https://doi.org/10.3892/etm.2017.4347

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  41. Li J, Lai Y, Ma J et al (2017) miR-17-5p suppresses cell proliferation and invasion by targeting ETV1 in triple-negative breast cancer. BMC Cancer 17:745. https://doi.org/10.1186/s12885-017-3674-x

    Article  PubMed  PubMed Central  Google Scholar 

  42. Li J, Li Q, Huang H et al (2017) Overexpression of miRNA-221 promotes cell proliferation by targeting the apoptotic protease activating factor-1 and indicates a poor prognosis in ovarian cancer. Int J Oncol 50:1087–1096. https://doi.org/10.3892/ijo.2017.3898

    Article  PubMed Central  CAS  Google Scholar 

  43. Gao J, Liu QG (2011) The role of miR-26 in tumors and normal tissues (review). Oncol Lett 2:1019–1023. https://doi.org/10.3892/ol.2011.413

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  44. Deng M, lin TH, X hong L et al (2013) miR-26a suppresses tumor growth and metastasis by targeting FGF9 in gastric cancer. PLoS One 8:1–10. https://doi.org/10.1371/journal.pone.0072662

    Article  CAS  Google Scholar 

  45. Jin F, Wang Y, Li M et al (2017) MiR-26 enhances chemosensitivity and promotes apoptosis of hepatocellular carcinoma cells through inhibiting autophagy. Cell Death Dis 8:e2540. https://doi.org/10.1038/cddis.2016.461

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  46. Sun TY, Xie HJ, He H et al (2016) miR-26a inhibits the proliferation of ovarian cancer cells via regulating CDC6 expression. Am J Transl Res 8:1037–1046 eCollection 2016

    PubMed  PubMed Central  CAS  Google Scholar 

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Acknowledgements

The authors thank Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG) for CDS - PPM-00383-14 and CDS - BPV-00277-16 grants and Pró-Reitoria de Pesquisa from Universidade Federal de Minas Gerais (PRPq –UFMG) for support.

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Authors and Affiliations

  1. Serviço de Biologia Celular, Fundação Ezequiel Dias, Belo Horizonte, Minas Gerais, Brazil

    Aline Brito de Lima, Luciana Maria Silva, Nikole Gontijo Gonçales & Letícia da Conceição Braga

  2. Laboratório de Genética Humana e Médica, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil

    Aline Brito de Lima & Maria Raquel Santos Carvalho

  3. Departamento de Ginecologia e Obstetrícia, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil

    Agnaldo Lopes da Silva Filho & Letícia da Conceição Braga

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  1. Aline Brito de Lima
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  2. Luciana Maria Silva
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  3. Nikole Gontijo Gonçales
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Correspondence to Agnaldo Lopes da Silva Filho.

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de Lima, A.B., Silva, L.M., Gonçales, N.G. et al. Three-Dimensional Cellular Arrangement in Epithelial Ovarian Cancer Cell Lines TOV-21G and SKOV-3 is Associated with Apoptosis-Related miRNA Expression Modulation. Cancer Microenvironment 11, 85–92 (2018). https://doi.org/10.1007/s12307-017-0203-z

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