Myxoma virus-mediated oncolysis of ascites-derived human ovarian cancer cells and spheroids is impacted by differential AKT activity.
OBJECTIVE
We propose that metastatic epithelial ovarian cancer (EOC) is a potential therapeutic target for the oncolytic agent, Myxoma virus (MYXV).


METHODS
Primary EOC cells were isolated from patient ascites and cultured as adherent cells or in suspension using Ultra Low-Attachment dishes. MYXV expressing green fluorescent protein was used to infect cells and spheroids. Infection was monitored by fluorescence microscopy, viral titering and immunoblotting for M-T7 and M130 virus protein expression, and cell viability by alamarBlue assay. Akti-1/2 (5 μM) and rapamycin (20 nM) were used to assay the role of PI3K-AKT signaling in mediating MYXV infection.


RESULTS
Ascites-derived EOC cells grown in adherent culture are effectively killed by MYXV infection. EOC cells grown in suspension to form three-dimensional EOC spheroids readily permit MYXV entry into cells, yet are protected from the cytopathic effects of late MYXV infection. Upon reattachment (to model secondary metastasis), EOC spheroids are re-sensitized to MYXV-mediated oncolysis. The critical determinant that facilitates efficient MYXV infection is the presence of an activated PI3K-AKT signaling pathway. Treatment with the specific AKT inhibitor Akti-1/2 reduces infection of monolayer EOC cells and spheroids. Direct infection of freshly-collected ascites demonstrated that 54.5% of patient samples were sensitive to MYXV-mediated oncolytic cell killing. We also demonstrate that factor(s) present in ascites may negatively impact MYXV infection and oncolysis of EOC cells, which may be due to a down-regulation in endogenous AKT activity.


CONCLUSIONS
Differential activity of AKT serves as the mechanistic basis for regulating MYXV-mediated oncolysis of EOC spheroids during key steps of the metastatic program. In addition, we provide the first evidence that MYXV oncolytic therapy may be efficacious for a significant proportion of ovarian cancer patients with metastatic disease.
Translational Ovarian Cancer Research Program, London Regional Cancer Program, London, Ontario, Canada.
Rohann J M
RJ
Correa
Monica
M
Komar
Jessica G K
JG
Tong
Milani
M
Sivapragasam
Masmudur M
MM
Rahman
Grant
G
McFadden
Gabriel E
GE
Dimattia
Trevor G
TG
Shepherd
D001201:Ascites / Q000473:pathology; D000077216:Carcinoma, Ovarian Epithelial; D005260:Female; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods; D009233:Myxoma virus / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D009375:Neoplasms, Glandular and Epithelial / Q000201:enzymology / Q000628:therapy* / Q000821:virology; D051058:Oncogene Protein v-akt / Q000378:metabolism*; D050130:Oncolytic Virotherapy / Q000379:methods*; D010051:Ovarian Neoplasms / Q000201:enzymology / Q000628:therapy* / Q000821:virology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D018874:Spheroids, Cellular / Q000473:pathology; D014407:Tumor Cells, Cultured
D049452:Green Fluorescent Proteins; D019869:Phosphatidylinositol 3-Kinases; D051058:Oncogene Protein v-akt