title: Induction of artificial cancer stem cells from tongue cancer cells by defined reprogramming factors.
abstract: The cancer stem cells (CSCs), a small subpopulation of cells in tumor are responsible for the tumor initiation, growth, recurrence and metastasis of cancer, as well as resistance of cancers to drugs or radiotherapy. CSCs are an important target for the development of novel strategies in cancer treatment. However, CSCs-targeted new anti-cancer drug discovery is currently hindered by the lack of easy and reliable methods for isolating, collecting and maintaining sufficient number of CSCs. Here, we examined whether introduction of defined reprogramming factors (Oct4, shp53, Sox2, Klf4, l-Myc and Lin28) into HSC2 tongue cancer cells could transform the HSC2 into HSC2 with CSCs properties.
  
  
  
  We introduced the defined reprogramming factors into HSC2 tongue cancer cells via episomal vectors by electroporation method to generate transfectant cells. We investigated the malignant properties of the transfectant cells by cell proliferation assay, migration assay, wound healing assay, sphere formation assay, chemosensitivity and radiosensitivity assay in vitro; and also examined the tumorigenic potential of the transfectants in vivo.
  
  
  
  The transfectant cells (HSC2/hOCT3/4-shp53-F, HSC2/hSK, HSC2/hUL, HSC2/hOCT3/4-shp53-F + hSK, HSC2/hOCT3/4-shp53-F + hUL, HSC2/hSK + hUL, HSC2/hOCT3/4-shp53-F + hSK + hUL) displayed a malignant phenotype in culture and form tumors on the back of nude mice more efficiently than parental HSC2 and control HSC2/EGFP transfectant cells. They exhibited increased resistance to chemotherapeutic agents; 5-fluorouracil, cisplatin, docetaxel, trifluorothymidine, zoledronic acid, cetuximab, bortezomib and radiation when compared with HSC2 and HSC2/EGFP. Among all the transfected cells, HSC2/hOCT3/4-shp53-F + hSK + hUL cell containing all of the reprogramming factors showed the most aggressive and malignant properties and presented the highest number of spheres in the culture medium containing human recombinant fibroblast Growth Factor-2 (FGF-2) and epidermal Growth Factor (EGF).
  
  
  
  These findings suggest that artificial cancer stem cells obtained by the induction of cellular reprogramming may be useful for investigating the acquisition of potential malignancy as well as screening the CSCs-targeting drugs.
authors.affiliation: Department of Oral and Maxillofacial Surgery, Yamaguchi University Graduate School of Medicine, 1-1-1, Minamikogushi, Ube, 755-8505, Japan. harako@yamaguchi-u.ac.jp.
authors.forename: Koji
authors.initials: K
authors.lastname: Harada
authors.affiliation: Department of Oral and Maxillofacial Surgery, Yamaguchi University Graduate School of Medicine, 1-1-1, Minamikogushi, Ube, 755-8505, Japan.
authors.forename: Tarannum
authors.initials: T
authors.lastname: Ferdous
authors.affiliation: Department of Pathology, Yamaguchi University Graduate School of Medicine, 1-1-1, Minamikogushi, Ube, 755-8505, Japan.
authors.forename: Dan
authors.initials: D
authors.lastname: Cui
authors.affiliation: Department of Biochemistry and Functional Proteomics, Yamaguchi University Graduate School of Medicine, 1-1-1, Minamikogushi, Ube, 755-8505, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kuramitsu
authors.affiliation: Department of Physiology, Keio University, School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Matsumoto
authors.affiliation: Department of Pathology, Yamaguchi University Graduate School of Medicine, 1-1-1, Minamikogushi, Ube, 755-8505, Japan.
authors.forename: Eiji
authors.initials: E
authors.lastname: Ikeda
authors.affiliation: Department of Physiology, Keio University, School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Okano
authors.affiliation: Department of Oral and Maxillofacial Surgery, Yamaguchi University Graduate School of Medicine, 1-1-1, Minamikogushi, Ube, 755-8505, Japan.
authors.forename: Yoshiya
authors.initials: Y
authors.lastname: Ueyama
mesh_terms: D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D065150:Cellular Reprogramming; D018274:Electroporation; D004815:Epidermal Growth Factor / Q000494:pharmacology; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D006801:Humans; D000090062:Kruppel-Like Factor 4; D014411:Neoplastic Stem Cells / Q000378:metabolism / Q000473:pathology*; D018874:Spheroids, Cellular / Q000378:metabolism; D014062:Tongue Neoplasms / Q000378:metabolism / Q000473:pathology*; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured
chemical_list: C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor
keywords: Artificial CSC; Episomal vector; Reprogramming factor; Tongue cancer