Establishment and Characterization of Primary Human Ovarian Cancer Stem Cell Line (CD44+ve)


  • Amoura Abouelnaga Mansoura University, Faculty of Sciences and Faculty of Medicine, Mansoura Egypt
  • Ghada A. Mutawa Mansoura University, Faculty of Sciences and Faculty of Medicine, Mansoura Egypt
  • Hassan Abdelghaffar Mansoura University, Faculty of Sciences and Faculty of Medicine, Mansoura Egypt
  • Mohamed Sobh Mansoura University, Faculty of Sciences and Faculty of Medicine, Mansoura Egypt
  • Sahar Hamed Mansoura University, Faculty of Sciences and Faculty of Medicine, Mansoura Egypt
  • Shaker A. Mousa Mansoura University, Faculty of Sciences and Faculty of Medicine, Mansoura Egypt



Ovarian cancer, Cancer stem cell, Stemness genes, CD44, Chemo-resistance.


 Ovarian cancer is ranked as the 7th most lethal cancer worldwide with 239,000 new cases annually. The mortality rate is high because most ovarian tumors are diagnosed at advanced stages and are resistant to chemotherapy and thus incurable due to the lack of effective early detection of ovarian tumors. There is a small sub-population of ovarian tumor cells capable of self-renewal and differentiation into different cancer cell types, called cancer stem cells (CSCs), which might be responsible for cancer relapse. The CD44+ phenotype in ovarian tumor cells elucidates cancer initiating cell-like properties of promoting differentiation, metastasis, and chemotherapy-resistance. Increased expression of genes previously associated with CSCs promotes regenerative capacity by promoting stem cell function that can drive cancer relapse and metastasis. In this study we present a method to isolate the primary epithelial ovarian cancer cells from human solid tumor and establish CD44+ve primary ovarian cancer stem cell (OCSCCD44+ve) line using magnetic microbeads. Also we evaluated the expression of stemness genes NanogSox2Oct4, and Nestin by real-time qPCR analysis. Thequantitative analysis by real-time qPCRshows that OCSCCD44+ve overexpressed the embryonic stem cell marker genes NanogOct4Sox2, and Nestin when compared with ovarian cancer cells OCCCD44-ve as positive control and ovarian cells as negative control. We demonstrate that CD44 in malignant ovarian tumors is a critical molecule that exhibits cancer stem cell properties that enhance tumorigenicity and cancer metastasis. Our results provide a better understanding of ovarian CSCs, which is important for future in vivo studies with subsequent target therapy for preclinical studies.


Ferlay J, Soerjomataram I, Dikshit R, Eser S, Mathers C, Rebelo M, et al. Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer 2015; 136: E359-86.

Donninger H, Bonome T, Radonovich M, Pise-Masison CA, Brady J, Shih JH, et al. Whole genome expression profiling of advance stage papillary serous ovarian cancer reveals activated pathways. Oncogene 2004; 23: 8065-77.

Zhan Q, Wang C, Ngai S. Ovarian cancer stem cells: a new target for cancer therapy. Biomed Res Int 2013; 2013: 916819.

Ebben JD, Treisman DM, Zorniak M, Kutty RG, Clark PA, Kuo JS. The cancer stem cell paradigm: a new understanding of tumor development and treatment. Expert Opin Ther Targets 2010; 14: 621-32.

Li L, Neaves WB. Normal stem cells and cancer stem cells: the niche matters. Cancer Res 2006; 66: 4553-7.

Al-Hajj M, Clarke MF. Self-renewal and solid tumor stem cells. Oncogene 2004; 23: 7274-82.

Stickeler E, Mobus VJ, Kieback DG, Kohlberger P, Runnebaum IB, Kreienberg R. Intron 9 retention in gene transcripts suggests involvement of CD44 in the tumorigenesis of ovarian cancer. Anticancer Res 1997; 17: 4395-8.

Marhaba R, Klingbeil P, Nuebel T, Nazarenko I, Buechler MW, Zoeller M. CD44 and EpCAM: cancer-initiating cell markers. Curr Mol Med 2008; 8: 784-804.

Naor D, Sionov RV, Ish-Shalom D. CD44: structure, function, and association with the malignant process. Adv Cancer Res 1997; 71: 241-319.

Meng E, Long B, Sullivan P, McClellan S, Finan MA, Reed E, et al. CD44+/CD24- ovarian cancer cells demonstrate cancer stem cell properties and correlate to survival. Clin Exp Metastasis 2012; 29: 939-48.

Liu AF, Yu XY, Liu SR. Pluripotency transcription factors and cancer stem cells: small genes make a big difference. Chinese Journal of Cancer 2013; 32: 483-7.

Clark AT, Rodriguez RT, Bodnar MS, Abeyta MJ, Cedars MI, Turek PJ, et al. Human STELLAR, NANOG, and GDF3 genes are expressed in pluripotent cells and map to chromosome 12p13, a hotspot for teratocarcinoma. Stem Cells 2004; 22: 169-79.

Iv Santaliz-Ruiz LE, Xie X, Old M, Teknos TN, Pan Q. Emerging role of nanog in tumorigenesis and cancer stem cells. Int J Cancer 2014; 135: 2741-8.

Shih J, Rahman M, Luong QT, Lomeli SH, Riss J, Prins RM, et al. Dominant B-cell epitopes from cancer/stem cell antigen SOX2 recognized by serum samples from cancer patients. Am J Clin Exp Immunol 2014; 3: 84-90.

Weina K, Utikal J. SOX2 and cancer: current research and its implications in the clinic. Clin Transl Med 2014; 3: 19.

Ren JJ, Meng XK. A relative quantitative method to detect OCT4A gene expression by exon-junction primer and locked nucleic acid-modified probe. J Zhejiang Univ Sci B 2011; 12: 149-55.

Peng S, Maihle NJ, Huang Y. Pluripotency factors Lin28 and Oct4 identify a sub-population of stem cell-like cells in ovarian cancer. Oncogene 2010; 29: 2153-9.

Matsuda Y, Yoshimura H, Naito Z, Ishiwata T. The roles and molecular mechanisms of nestin expression in cancer with a focus on pancreatic cancer. J Carcinogen Mutagen 2013; S9:002.

Qin Q, Sun Y, Fei M, Zhang J, Jia Y, Gu M, et al. Expression of putative stem marker nestin and CD133 in advanced serous ovarian cancer. Neoplasma 2012; 59: 310-5.

Weigmann B Cell isolation of spleen mononuclear cells.

Puiffe ML, Le Page C, Filali-Mouhim A, Zietarska M, Ouellet V, Tonin PN, et al. Characterization of ovarian cancer ascites on cell invasion, proliferation, spheroid formation, and gene expression in an in vitro model of epithelial ovarian cancer. Neoplasia 2007; 9: 820-9.

Casagrande F, Cocco E, Bellone S, Richter CE, Bellone M, Todeschini P, et al. Eradication of chemotherapy-resistant CD44+ human ovarian cancer stem cells in mice by intraperitoneal administration of Clostridium perfringens enterotoxin. Cancer 2011; 117: 5519-28.

Boccaccio C, Comoglio PM. Invasive growth: a MET-driven genetic programme for cancer and stem cells. Nat Rev Cancer 2006; 6: 637-45.

Yan W, Chen Y, Yao Y, Zhang H, Wang T. Increased invasion and tumorigenicity capacity of CD44+/CD24- breast cancer MCF7 cells in vitro and in nude mice. Cancer Cell International 2013; 13: 62.

Foster R, Buckanovich RJ, Rueda BR. Ovarian cancer stem cells: working towards the root of stemness. Cancer Letters 2013; 338: 147-57.

Steg AD, Bevis KS, Katre AA, Ziebarth A, Dobbin ZC, Alvarez RD, et al. Stem cell pathways contribute to clinical chemoresistance in ovarian cancer. Clin Cancer Res 2012; 18: 869-81.




How to Cite

Amoura Abouelnaga, Ghada A. Mutawa, Hassan Abdelghaffar, Mohamed Sobh, Sahar Hamed, & Shaker A. Mousa. (2016). Establishment and Characterization of Primary Human Ovarian Cancer Stem Cell Line (CD44+ve). Journal of Cancer Research Updates, 5(2),  59–66.