Abstract 1579: Exosomes promote ovarian cancer invasion through CD-44 transfer to mesothelial cells

Abstract

Abstract Purpose: The peritoneum and organs in the peritoneal cavity are covered by a single layer of mesothelial cells. Therefore, ovarian cancer cells, which metastasize within the peritoneal cavity, directly encounter mesothelial cells as the initial step of metastasis. This contact has been found to involve cell-cell communication that affects cancer progression. Possible actors in this cell-cell communication are exosomes, 30-100 nm membrane vesicles of endocytic origin, through the cell-cell transfer of proteins, mRNAs, and microRNAs. Here, we aim to identify the functional roles of ovarian cancer-derived exosomes in this metastatic process. Methods: Exosomes were isolated from two ovarian cancer cell lines (HeyA8 and TYK-NU) and immortalized normal ovarian epithelial cell line (IOSE) using differential centrifugation. Human peritoneal mesothelial cells (HPMCs) were isolated from normal omentum of patients undergoing gynecologic surgery. The isolation of exosomes was confirmed by electron microscope, nanoparticle tracking analysis, Western blotting and electrophoresis of RNA. The transfer of exosomes into HPMCs was confirmed by fluorescent-labeled exosomes. The effects of exosome transfer from ovarian cancer cells to mesothelial cells in cancer invasion were analyzed in vitro 3D culture model, morphological assessment, Western blotting and gelatin zymography. CD-44 was enriched in cancer derived exosome. Thus, gain or loss of function of CD-44 was analyzed. CD-44 expression in ovarian cancer omental metastasis and surrounding organs was assessed by immunohistochemistry using clinical samples. Results: Fluorescent-labeled exosomes were evidently transferred into HPMCs. Exosome-treated HPMCs changed in cellular morphology to spindle phenotype. Ovarian cancer invasion was significantly promoted in the presence of exosome-treated HPMCs. In exosome-treated HPMCs, MMP-9 secretion was up-regulated and E-cadherin expression down-regulated. The clearance of mesothelial barrier was increased in exosome-treated HPMC monolayer. CD-44 was enriched in cancer-derived exosomes and exosome-treated HPMCs display high-level of CD-44. When CD-44 expression was knocked down by siRNA in ovarian cancer cells, these effects to HPMCs were significantly attenuated. In contrast, the enforced expression of CD-44 into HPMCs promoted cancer invasion. In human omentum with microscopic metastasis of ovarian cancer, positive CD-44 expression was confirmed in a mesothelial cell layer when cancer cells are attaching onto it, while CD-44 expression was generally negative in normal mesothelial cells. Conclusion: Ovarian cancer-derived exosomes transfer CD-44 to HPMCs, which can facilitate ovarian cancer invasion by up-regulating of MMP-9 secretion and increasing mesothelial clearance. Citation Format: Koji Nakamura, Kenjiro Sawada, Yasuto Kinose, Akihiko Yoshimura, Erika Nakatsuka, Seiji Mabuchi, Tadashi Kimura. Exosomes promote ovarian cancer invasion through CD-44 transfer to mesothelial cells. [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 1579.