Abstract 851: Reversion of epithelial to mesenchymal transition under androgen deprived conditions promotes chemoresistance in prostate cancer

Abstract

Abstract Introduction: Metastatic prostate cancer (PC) is one of the leading causes of cancer related male deaths in Australia. Radical prostatectomy followed by androgen deprivation therapy (ADT) upon recurrence of disease is the conventional form of treatment. Upon relapse, administration of androgen targeted therapy (ATT) and subsequent chemotherapy improves quality of life and overall survival of patients. However, it is not curative and the disease ultimately progresses. Epithelial to mesenchymal transition (EMT) is a process that can facilitate tumour growth and progression to metastasis and has also been associated with chemoresistance. We therefore aim to determine how EMT and resultant reversal of this state (MET) influences the chemoresistance profile of PC cells and further determine if and how specific genetic regulators are involved in this process. Methods: Doxycycline hyclate (Dox) inducible Snai1 and Zeb1, LNCaP EMT models were used to determine the effect of the EMT status in response to a clinically relevant panel of drugs using cellular and molecular assays. All assays were performed in both fetal bovine serum (FBS) and charcoal stripped serum (CSS) supplemented culture media. CSS media was used to mimic an ADT-like environment. Using 2D monolayer and 3D spheroid [Happy Cell©] formats. Cells were induced for 5 days of EMT in the presence of Dox and were allowed to revert for 14 days of MET (absence of Dox) before the drug assays were carried out over a period of 72 hours. Live/dead cell staining was used to determine viability percentage in both models. Flow cytometry was used for 2D assays and 3D assays were analysed using either a GE IN Cell Analyzer live cell imaging system or confocal microscopy. Results: Induction of Snai1 and Zeb1 induced an elongated cell shape and increased mRNA and protein expression of mesenchymal markers such as vimentin during EMT. This was associated with a decrease in expression of epithelial markers E-cadherin and EpCam,. An eventual drop in the expression levels of vimentin at around 14 days of dox removal along with an increase in epithelial markers was observed. These changes were also associated with a gradual reversal in the phenotype of these cells. However, this reversal in phenotype and expression of EMT markers was not as striking in cells grown in CSS compared to cells in FBS media (both 2D and 3D). Mitoxantrone concentration-response assays in a 2D format showed that cells that undergo a subsequent MET are more chemoresistant than uninduced or EMT cells when grown in CSS media. This was however, not observed in cells undergoing similar transitions in the FBS media. Conclusions: The chemoresistance profile of prostate cancer cells is altered by their EMT status. This effect is enhanced in the context of ADT. Cells that revert back to a more epithelial phenotype after undergoing an EMT exhibit higher resistance towards chemotherapy drugs. Citation Format: Akanksha Upadhyaya, Nataly Stylianou, Anthony M. Davies, Sarah-Louise Ryan, Brett G. Hollier, Elizabeth D. Williams. Reversion of epithelial to mesenchymal transition under androgen deprived conditions promotes chemoresistance in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 851. doi:10.1158/1538-7445.AM2017-851