Abstract 4379: Exosomes isolated from ovarian cancer cells transfer oncogenic features to the target cells promoting epithelial to mesenchymal transition

Abstract

Abstract Introduction: Recent studies establish the role of exosomes in cell-cell communication in cancer. As adaptation to hypoxia is a critical step in tumor progression, the aim of this study was to test the hypotheses that hypoxia promotes epithelial to mesenchymal transition (EMT) and that exosomes isolated from hypoxic cancer cells transfer oncogenic properties to target cells. Methods: CaOV-3 cell line was used as model of ovarian cancer. Cells were cultured under 8% O2 (normoxia) and 1% O2 (hypoxia) for 48 hours. Exosomes were isolated from cell-conditioned media by differential and buoyant density centrifugation. Exosomes were characterised by the presence of TSG101 using Western Blot, size distribution (Nanosight™) and morphology by electron microscopy. Exosomal protein and miRNA content were determined using liquid chromatography mass spectrometry (5600 Triple TOF, AB Sciex,) and an Illumina NextSeq 500 Platform, respectively. The effects of exosomes released from CaOV-3 incubated under 1% O2 on EMT induction in CaOV-3 cells cultured under 8% O2 were assessed by the measuring the ratio of E-cadherin (epithelial marker) to N-cadherin (mesenchymal marker) by Wester Blot and the expression of 84 key genes involve in the EMT (RT2 Profiler™ PCR Array, QIAGEN). Results: Exosomes were identified as spherical vesicles with a typical cup-shape, diameters ranging from 50 to 100 nm with the expression of TSG101. Exosome release from ovarian cancer cells was ∼4-fold higher under hypoxic than normoxic conditions (p <0.001). Hypoxia-specific exosomal proteins and miRNAs were identified. Hypoxia induced EMT (increased N-cadherin/E-cadherin ratio) on CaOV-3 cells compared to cells cultured under 8% O2. Interestingly, exosomes isolated from hypoxic conditions mimic the effect of hypoxia on cells cultured under 8% O2, involving the unregulated of a set of transcription factors associated with EMT such as SNAIL1/SNAIL2, bHLH (E47, E2-2, and TWIST1/TWIST2), and ZEB1/ZEB2. Conclusion: The data obtained is consistent with the hypothesis that exosomes released from cancer cells modify the phenotype of target cells by transferring pro-oncogenic molecules inducing cancerous phenotype of receipt cells, contributing to tumour growth and metastasis. Citation Format: Miharu Kobayashi, Laura Cohelo, Dominic Guanzon, Katherin Scholz-Romero, Melissa Brown, Richard Kline, Li Li, Gregory Rice, Carlos Salomon. Exosomes isolated from ovarian cancer cells transfer oncogenic features to the target cells promoting epithelial to mesenchymal transition. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4379.