Induction of programmed cell death by inhibition of AKT with the alkylphosphocholine perifosine in in vitro models of platinum sensitive and resistant ovarian cancers.
OBJECTIVE
We analyzed the anti-tumor effect and the mechanism of action of perifosine, an orally active alkylphospholipid AKT inhibitor using in vitro models of human ovarian cancer.


METHODS
Ovarian cancer cells OAW42, PA-1, SKOV3, and A2780 as well as platinum resistant A2780cis cells were incubated with increasing concentrations of perifosine, with and without multi-caspase inhibitor zVAD-FMK. The effect of a combined treatment with cisplatin and perifosine was investigated in OAW42, SKOV3, A2780 and A2780cis cells. Cytotoxic effects of perifosine were analyzed using crystal violet staining, FACS analysis of DNA content as well as Annexin V/propidium iodide-double staining. The effect of perifosine on AKT phosphorylation was determined by Western blotting.


RESULTS
Perifosine displayed anti-tumor activity in all five cell lines, which increased time-dependently. While IC(50) values at 24 h were >40 μM, IC(50) values after 72 h decreased to 10 μM in OAW42 and 25 μM in PA-1 and 30 μm in SKOV3 cells. In platinum resistant A2780cis cells perifosine showed good antiproliferative activity (IC(50) = 3 μm). At adequate doses, perifosine increased cytotoxic effects of cisplatin in OAW42, A2780 and A2780cis cell. Anti-tumor activity of perifosine was not confined to a specific phase of the cell cycle and could not be decreased by the pan-caspase inhibitor zVAD-FMK. AnnexinV/propidium iodide-double staining after treatment with perifosine was not indicative of classical apoptosis. AKT phosphorylation was dose-dependently inhibited by perifosine.


CONCLUSIONS
Perifosine showed substantial cytotoxic effects in various in vitro models of ovarian cancer. Since anti-tumor effects were not confined to platinum-sensitive cells perifosine seems to be a good candidate for clinical studies in patients especially with platinum resistant ovarian cancer.
Engel
Jörg B
JB
Universitätsfrauenklinik Würzburg, Josef-Schneider-Str. 4, 97080 Würzburg, Germany. joergbengel@hotmail.com
Schönhals
Tanja
T
Häusler
Sebastian
S
Krockenberger
Mathias
M
Schmidt
Melanie
M
Horn
Evi
E
Köster
Frank
F
Dietl
Johannes
J
Wischhusen
Jörg
J
Honig
Arnd
A
D000590:Amino Acid Chloromethyl Ketones / Q000627:therapeutic use; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D000971:Antineoplastic Combined Chemotherapy Protocols; D017209:Apoptosis / Q000187:drug effects*; D002277:Carcinoma / Q000188:drug therapy*; D045744:Cell Line, Tumor; D002945:Cisplatin / Q000627:therapeutic use; D015853:Cysteine Proteinase Inhibitors / Q000627:therapeutic use; D019008:Drug Resistance, Neoplasm; D005260:Female; D006801:Humans; D010051:Ovarian Neoplasms / Q000188:drug therapy*; D010767:Phosphorylcholine / Q000031:analogs & derivatives* / Q000627:therapeutic use; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors*
D000590:Amino Acid Chloromethyl Ketones; D000970:Antineoplastic Agents; D015853:Cysteine Proteinase Inhibitors; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D010767:Phosphorylcholine; C105905:perifosine; D051057:Proto-Oncogene Proteins c-akt; D002945:Cisplatin