Chemosensitization of endometrial cancer cells through AKT inhibition involves FOXO1.
OBJECTIVE
Endometrial cancer is the most common type of gynecologic cancer in the United States. In this study, we propose that inhibition of the AKT pathway sensitizes cells to chemotherapeutic agents by increasing FOXO1 expression.


METHODS
Ishikawa and RL95 cells were treated with the AKT inhibitor (API-59CJ-OMe) alone and in combination with carboplatin or paclitaxel. Cells were counted using a hemocytometer and cell cycle analysis done with flow cytometry. Apoptosis was measured with TUNEL and Annexin V/DAPI staining. FOXO1 protein expression and localization was done using immunofluorescent staining of cells. Finally, the adenovirus containing triple mutant FOXO1 was used to overexpress the constitutively active FOXO1 in Ishikawa cells and its effects on cell viability were studied.


RESULTS
Treatment with 6 microM API-59CJ-OME resulted in preferential cell death in Ishikawa and RL95 cells compared to another endometrial cancer cell line, ECC1 after 48 h of treatment. API-59CJ-OME treatment of Ishikawa cells resulted in cell cycle arrest in the G2/M phase. The addition of API-59CJ-OME to carboplatin resulted in a synergistic increase in cell death by apoptosis compared to the responses to each agent separately. Treatment with API-59CJ-OME, carboplatin, paclitaxel or the combinations for 24 h increased nuclear expression of FOXO1 in Ishikawa cells. Overexpression of FOXO1 caused 37% of the cells to die within 24 h. Addition of carboplatin to the AD-FOXO1 expressing cells further increased cell death to 71%.


CONCLUSIONS
Inhibition of AKT signaling potentiates cell death in Ishikawa and RL95 cells when combined with carboplatin through mechanisms involving FOXO1 activation.
Hoekstra
Anna V
AV
Department of Obstetrics and Gynecology, Division of Gynecologic Oncology, Northwestern University, Chicago, IL 60611, USA.
Ward
Erin C
EC
Hardt
Jennifer L
JL
Lurain
John R
JR
Singh
Diljeet K
DK
Buttin
Barbara M
BM
Schink
Julian C
JC
Kim
J Julie
JJ
D000971:Antineoplastic Combined Chemotherapy Protocols / Q000008:administration & dosage / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D016190:Carboplatin / Q000008:administration & dosage / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor / Q000187:drug effects / Q000378:metabolism; D004611:Ellipticines / Q000008:administration & dosage / Q000494:pharmacology*; D016889:Endometrial Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D005260:Female; D005434:Flow Cytometry; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors / Q000378:metabolism*; D006801:Humans; D017239:Paclitaxel / Q000008:administration & dosage / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt; D015398:Signal Transduction / Q000187:drug effects
D004611:Ellipticines; C085126:FOXO1 protein, human; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; D016190:Carboplatin; C494918:AKT1 protein, human; D051057:Proto-Oncogene Proteins c-akt; D017239:Paclitaxel