gw-4869 and Ovarian-Neoplasms

Exosomes, which are nanovesicles secreted by almost all the cells, mediate intercellular communication and are involved in various physiological and pathological processes. We aimed to investigate the effects of graphene oxide (GO) on the biogenesis and release of exosomes in human ovarian cancer (SKOV3) cells.. Exosomes were isolated using ultracentrifugation and ExoQuick and characterized by various analytical techniques. The expression levels of exosome markers were analyzed via quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay.. Graphene oxide (10-50 μg/mL), cisplatin (2-10 μg/mL), and C6-ceramide (5-25 μM) inhibited the cell viability, proliferation, and cytotoxicity in a dose-dependent manner. We observed that graphene oxide (GO), cisplatin (CIS), and C6-Ceramide (C6-Cer) stimulated acetylcholine esterase and neutral sphingomyelinase activity, total exosome protein concentration, and exosome counts associated with increased level of apoptosis, oxidative stress and endoplasmic reticulum stress. In contrast, GW4869 treatment inhibits biogenesis and release of exosomes. We observed that the human ovarian cancer cells secreted exosomes with typical cup-shaped morphology and surface protein biomarkers. The expression levels of TSG101, CD9, CD63, and CD81 were significantly higher in GO-treated cells than in control cells. Further, cytokine and chemokine levels were significantly higher in exosomes isolated from GO-treated SKOV3 cells than in those isolated from control cells. SKOV3 cells pre-treated with N-acetylcysteine or GW4869 displayed a significant reduction in GO-induced exosome biogenesis and release. Furthermore, endocytic inhibitors decrease exosome biogenesis and release by impairing endocytic pathways.. This study identifies GO as a potential tool for targeting the exosome pathway and stimulating exosome biogenesis and release. We believe that the knowledge acquired in this study can be potentially extended to other exosome-dominated pathologies and model systems. Furthermore, these nanoparticles can provide a promising means to enhance exosome production in SKOV3 cells.

Topics: Carcinoma, Ovarian Epithelial; Cisplatin; Exosomes; Female; Humans; Ovarian Neoplasms

Ovarian cancer is the most common malignancy in women. Owing to late syndromic presentation and lack of efficient early detection, most cases are diagnosed at advanced stages. Surgery and platinum-based chemotherapy are still the standard care currently. However, resistance invoked often compromises the clinical value of the latter. Expression of DNA methyltransferase 1 (DNMT1) was analysed by gene array. Protein was determined by immunoblotting. Exosome was isolated with commercial kit. Cell proliferation was measured by CCK8 method. Annexin V-PI double staining was performed for apoptosis evaluation. Xenograft model was established and administrated with exosome. Tumour growth and overall survival were monitored. We demonstrated the upregulation of DNMT1 in both tumour and derived cell line. DNMT1 transcripts were highly enriched in exosomes from conditioned medium of ovarian cells. Co-incubation with exosomes stimulated endogenous expression and rendered host cell the resistance to cytotoxicity of cisplatin. In vivo administration of DNMT1-containing exosomes exacerbated xenograft progression and reduced overall survival significantly. Moreover, treatment with exosome inhibitor GW4869 almost completely restored sensitivity in resistant cells. Our data elucidated an unappreciated mechanism of exosomal DNMT1 in cisplatin resistance in ovarian cancer, also indicating the potential of the combination of exosome inhibitor with cisplatin in resistant patients.

Topics: Aniline Compounds; Animals; Antineoplastic Agents; Benzylidene Compounds; Cell Line, Tumor; Cell Proliferation; Cisplatin; DNA (Cytosine-5-)-Methyltransferase 1; DNA (Cytosine-5-)-Methyltransferases; Drug Resistance, Neoplasm; Exosomes; Female; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Ovarian Neoplasms; RNA, Messenger; Transplantation, Heterologous; Up-Regulation