Rapamycin enhances the anti-tumor effect of gemcitabine in pancreatic cancer cells.
OBJECTIVE
Rapamycin is a potent inhibitor of PI3K/Akt pathway activation and its chemotherapeutic effect against pancreatic cancer has been demonstrated. In the present study, the combined effect of rapamycin with gemcitabine was examined and a screening method for detecting sensitivity to combined effects was investigated.


METHODS
Four pancreatic cancer cell lines, MIA, PK-1, PANK-1, and PK-45H, were cultured with or without rapamycin (200, 100, 50, 25nM), or gemcitabine (2, 1, 0.5, 0.25 microM), or with rapamycin and gemcitabine in combination. Growth inhibition of each cell line in response to the agents was examined using the MTT assay. Data were expressed as percentage inhibition at each concentration. Sensitivity to the combined effect of rapamycin and gemcitabine was investigated with real-time PCR (Akt1, 2, and 3, PTEN, mTOR) and western blotting (Akt, Akt/ Ser473, Akt/ Thr308, PTEN).


RESULTS
Rapamycin inhibited the growth of MIA (% inhibition: 36.2+/-9.0, 19.5+/-9.0, 10.8+/-6.8, 8.8+/-5.9), PK-1 (41.3+/-0.1, 33.5+/-0.1, 25.2+/-0.1, 22.6+/-0.1), PANK-1 (27.3+/-0.1, 21.7+/-0.1, 15.9+/-0.1, 14.9+/-0.1), and PK-45H (30.0+/-0.1, 24.4+/-0.1, 23.5+/-0.1, 20.5+/-0.1), whereas gemcitabine inhibited the growth in MIA (37.5+/-11.1, 11.5+/-7.8, 3.5+/-1.7, 0.1+/-0.1) and PK-1 (29.2+/-2.8, 26.7+/-0.8, 26.2+/-1.0, 25.1+/-1.0), but only weakly in PANK-1 (7.5+/-5.2, 1.2+/-0.9, 4.2+/-0.8, 2.7+/-0.6) and PK-45H (16.1+/-5.8, 11.8+/-10.0, 7.9+/-1.8, 10.1+/-1.8). However, gemcitabine administered with rapamycin had an enhanced inhibitory effect in PK-45H (61.3+/-12.0%), and no change in PANK-1, when compared to the inhibition by rapamycin or gemcitabine alone.


CONCLUSIONS
As a screening method for assessing the combined effects of rapamycin and gemcitabine, the decrease in expression of Akt/Ser473 and Akt/Thr 308 with rapamycin treatment was promising. Rapamycin enhances the anti-tumor effect of gemcitabine. Detecting a decrease in Akt/ Ser473 and Akt/Thr308 after rapamycin treatment, by western blotting, may be an effective way for assessing the combined effect of rapamycin and gemcitabine.
Okada
Toshie
T
Second Department of Surgery, Dokkyo University School of Medicine, Tochigi, Japan.
Sawada
Tokihiko
T
Kubota
Keiichi
K
D000903:Antibiotics, Antineoplastic / Q000494:pharmacology* / Q000627:therapeutic use; D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D004354:Drug Screening Assays, Antitumor; D004357:Drug Synergism; D006801:Humans; D066298:In Vitro Techniques; D051059:PTEN Phosphohydrolase / Q000378:metabolism; D010190:Pancreatic Neoplasms / Q000188:drug therapy*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020123:Sirolimus / Q000494:pharmacology* / Q000627:therapeutic use; D058570:TOR Serine-Threonine Kinases; D000093542:Gemcitabine
D000903:Antibiotics, Antineoplastic; D003841:Deoxycytidine; D011494:Protein Kinases; C546842:MTOR protein, human; C494918:AKT1 protein, human; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human; D020123:Sirolimus; D000093542:Gemcitabine