Long-term culture following ES-like gene-induced reprogramming elicits an aggressive phenotype in mutated cholangiocellular carcinoma cells.
BACKGROUND
We recently reported that gastrointestinal (GI) cancer cells can be reprogrammed to a pluripotent state by the ectopic expression of defined embryonic stem (ES)-like transcriptional factors. The induced pluripotent cancer (iPC) cells from GI cancer were sensitized to chemotherapeutic agents and differentiation-inducing treatment during a short-term culture, although a phenotype induced by long-term culture needs to be studied.


METHODS
A long-term cultured (Lc)-iPC cells were produced in GI cancer cell lines by virus-mediated introduction of four ES-like genes-c-MYC, SOX2, OCT3/4, and KLF4-followed by a culture more than three months after iPC cells induction. An acquired state was studied by expression of immature-related surface antigens, Tra-1-60, Tra-1-81, Tra-2-49, and Ssea-4; and epigenetic trimethyl modification at lysine 4 of histone H3. Sensitivity to chemotherapeutic agents and tumorigenicity were studied in Lc-iPC cells.


RESULTS
Whereas the introduction of defined factors of iPC cells once induced an immature state and sensitized cells to therapeutic reagents, the endogenous expression of the ES-like genes except for activated endogenous c-MYC was down-regulated in a long-term culture, suggesting a high magnitude of the reprogramming induction by defined factors and the requirement of therapeutic maintenance in Lc-iPC cells from cholangiocellular carcinoma HuCC-T1 cells, which harbor TP53(R175H) and KRAS(G12D). The Lc-iPC cells showed resistance to 5-fluorouracil in culture, and high tumorigenic ability with activated endogenous c-MYC in immunodeficient mice.


CONCLUSIONS
The Lc-iPC cells from HuCC-T1 might be prone to an undesirable therapeutic response because of an association with the activated endogenous c-MYC. To consider the possible therapeutic approach in GI cancer, it would be necessary to develop a predictive method for evaluating the improper reprogramming-associated aggressive phenotype of iPC cells.
Nagai
Ken-ichi
K
Department of Gastroenterological Surgery, Osaka University Graduate School of Medicine, Yamadaoka 2-2, Suita, Osaka 565-0871, Japan.
Ishii
Hideshi
H
Miyoshi
Norikatsu
N
Hoshino
Hiromitsu
H
Saito
Toshiyuki
T
Sato
Tetsuya
T
Tomimaru
Yoshito
Y
Kobayashi
Shogo
S
Nagano
Hiroaki
H
Sekimoto
Mitsugu
M
Doki
Yuichiro
Y
Mori
Masaki
M
D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D001650:Bile Duct Neoplasms / Q000235:genetics / Q000473:pathology*; D001653:Bile Ducts, Intrahepatic / Q000378:metabolism / Q000473:pathology*; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects; D065150:Cellular Reprogramming / Q000235:genetics; D018281:Cholangiocarcinoma / Q000235:genetics / Q000473:pathology*; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D053595:Embryonic Stem Cells / Q000378:metabolism; D044127:Epigenesis, Genetic; D005472:Fluorouracil / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D000090062:Kruppel-Like Factor 4; D051379:Mice; D016513:Mice, SCID; D009154:Mutation; D014411:Neoplastic Stem Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics*; D015533:Transcriptional Activation
D000964:Antimetabolites, Antineoplastic; C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D016271:Proto-Oncogene Proteins c-myc; D005472:Fluorouracil