Rapamycin inhibits growth of cholangiocarcinoma cells.
OBJECTIVE
The immunosuppressive agent rapamycin is currently being evaluated for its antineoplastic effect. In the present study, the antineoplastic effect of rapamycin against cholangiocarcinoma was studied in vitro.


METHODS
To explore the therapeutic potential of rapamycin, expression of mTOR in four cholangiocarcinoma cell lines--TFK1, HuCCT1, NOZW, and OZ--was evaluated by real-time PCR. The cell lines were then cultured with rapamycin (200 nM), and changes in the expression of Akt, phosphorylated PTEN (pPTEN), and phosphorylated S6 (pS6) were evaluated by western blotting. Finally, the cell lines were cultured with rapamycin (0, 25, 50, 100, 200 nM), gemcitabine (0, 0.5, 1, 2 microM), or both, and the antiproliferative effect was evaluated by MTT assay.


RESULTS
All four cholangiocarcinoma cell lines expressed endogenous mTOR-mRNA, the of expression being highest in HuCCT1 (65.8) and lowest in TFK1 (17.6). Western blotting revealed that rapamycin treatment decreased Akt expression significantly in all four cell lines (TFK1; 15.5%, HuCCT1; 6.3%, NOZW; 9.8%, OZ; 19.5%), and also decreased the expression of p-PTEN (TFK1; 10.6%, HuCCT1; 5.4%, NOZ-W; 12.2%, OZ; 12.2%) and pS6 (TFK1; 64.0%, HuCCT1; 73.9%, NOZW; 78.6%, OZ; 47.6%) in all four cell lines. Finally, rapamycin significantly inhibited the growth of all four cell lines in a dose-dependent manner. Gemcitabine inhibited the growth of NOZW and HuCCT1, but its effect was less marked on TFK1 and OZ. Furthermore, a synergistic anti-proliferative effect of rapamycin and gemcitabine was observed in TFK1, NOZW, and OZ, but not in HuCCT1.


CONCLUSIONS
Rapamycin effectively inhibited the growth of the four cholangiocarcinoma cell lines tested, and a synergistic effect with gemcitabine was observed in three of them. Rapamycin offers a new therapeutic strategy to inhibit the growth of cholangiocarcinoma.
Okada
Toshie
T
Second Department of Surgery, Dokkyo University School of Medicine, Kitakobayashi 880, Mibu, Shimotsuga, Tochigi 321-0293, Japan.
Sawada
Tokihiko
T
Kubota
Keiichi
K
D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D015153:Blotting, Western; D045744:Cell Line, Tumor / Q000187:drug effects / Q000378:metabolism; D018281:Cholangiocarcinoma / Q000188:drug therapy*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D066298:In Vitro Techniques; D051059:PTEN Phosphohydrolase / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism; D020123:Sirolimus / Q000494:pharmacology*; D058570:TOR Serine-Threonine Kinases
D000903:Antibiotics, Antineoplastic; D011494:Protein Kinases; C546842:MTOR protein, human; D051057:Proto-Oncogene Proteins c-akt; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human; D020123:Sirolimus