Isolation and Identification of Cancer Stem Cell in Nasopharyngeal Carcinoma

• Main Authors: Yao-An Shen, 沈耀安
• Other Authors: Yann-Jang Chen
• Format: Others
• Language: en_US
• Published: 2009
• Online Access: http://ndltd.ncl.edu.tw/handle/tcjhy7

碩士 === 國立陽明大學 === 生命科學暨基因體科學研究所 === 97 === Cancer stem cell (CSC) describes the rare subset stem-like cancer cells that exist in tumors. These cancer cells are able to self-renew and cause clinical treatment obstacles, such as therapy resistance and metastasis. In order to investigate CSC, we isolate CSC through side population, cell surface markers, or other available methods. However, the surface marker of CSC in nasopharyngeal carcinoma (NPC) remains unknown. Hence we took advantage of CSC’s natural properties, such as drug resistance, radioresistance, and the ability to form tumor spheres, to isolate CSC from five NPC cell lines (Tw01, Tw06, HONE-1, CNE-1, and CNE-2); the three isolation methods, when used together as one combination method, increased the precision in identifying CSC. Selected from this isolation method, the CSC-candidates demonstrated significant radioresistance, tumor sphere formation, side population percentage, clonogenic growth in soft agar, and metastatic potential. These candidates also expressed high-level stemness genes, p63 and Cytokeratin 14. Interestingly, these ectoderm NPC cancer stem cell candidates were able to differentiate into endoderm liver cells, and they even possessed liver function. We also found the surface marker changed into another surface antigen during differentiation. We then initiated surface marker screen on these CSC-candidates and located cell surface markers by identifying that CSCs have high level of CD44 and CD24 surface antigen expression. We inferred that CSC enriched subpopulation in NPC appeared as CD44+/CD24+. CD44+/CD24+ cells expressed significant CSC properties compared with differentiated cell-like CD44-/CD24- subpopulations. 500 CD44+/CD24+ cells formed tumor in NOD/SCID mice, while the same amount of CD44-/CD24- cells were not able to form tumor. Furthermore, we generated NPC-induced cancer stem cell (iCSC) via transfecting Twist (EMT factor) as well as Oct4, Sox2, Klf4 and c-Myc (Yamanaka factors). iCSC acquired CSC properties along with CD44+/CD24+ antigen phenotype, which confirmed the CSC surface marker in NPC as we had predicted. Taken together, the combination isolation method can serve as a model isolation method in CSC research. CD44+/CD24+ will help to identify CSC and aid in the subsequent drug design to target CSC in NPC. p63 and CK14 have prognosis and diagnosis marker potential. Looking ahead, we plan to investigate the mechanism of CSC in NPC even more thoroughly.