Targeting Inhibition of Notch 1 Can Promote mTOR Signaling Pathway in Tumor-Related Macrophage to Regulate the Development of Ovarian Cancer.

BACKGROUND: Ovarian cancer is the leading cause of death linked to gynecological cancers. Notch1, as an important component of Notch signaling, plays an important role in a variety of cancers. This study aims to discuss the mechanisms through which Notch 1 influences the development of ovarian cancer.

METHODS: To design and establish the short hairpin (sh) RNA for targeting Notch 1, we transfected THP-1 cells (one of the human macrophagic lines). The cells were divided into shRNA negative control (NC) group and the Notch 1 shRNA group. The CoC1 cells and THP-1 cells (human mononuclear macrophages) are co-cultured, which are injected into the nude mice subcutaneously based on proposition. The sizes of tumors and their volumes are observed through HE staining. Flow cytometry is used to sort out macrophages from subcutaneous tumors of nude mice, whose protein-related expression is detected through western blot. Then the NC group and the Notch 1 shRNA group in the co-culture system are treated with PI3K/mTOR Inhibitor-13 sodium (200 nM) for 48h and then co-cultured with human endothelial cell lines HUVEC, CoC1, and THP-1 to test the tube-forming capacity of HUVEC cells in each group to detect the protein-related expression in THP-1 cells using western blot.

RESULTS: It is seen that the Notch 1 shRNA group includes a significantly larger tumor size, decreased relative expression, and the obvious increase of the relative protein expression in p-PI3K, p-mTOR, HIF1α, and VEGF compared with the NC group. Through tube-forming experiments, the Notch1 shRNA group significantly increased the number of HUVEC tubes. However, after the use of PI3K/mTOR Inhibitor-13 sodium, the number of tubes decreased in the NC and Notch1 shRNA groups, and there is no significant discrepancy in comparison to the NC group. The in vitro western blotting results indicate no obvious variation of Notch 1's relative protein expression in both the NC group and Notch 1 shRNA group after the use of PI3K/mTOR Inhibitor-13 sodium, while the relative protein expression of p-PI3K, p-mTOR, HIF1α, and VEGF was significantly reduced and there was no significant difference.

CONCLUSION: This study found that specific knockout of Notch 1 in tumor-associated macrophages will promote the activation of the PI3K/mTOR signaling pathway and the expression of HIF1α and VEGF, thus promoting angiogenesis and the development of ovarian cancer. Thus, this study provides insight into novel prognostic biomarkers and therapeutic targets for the treatment and research of ovarian cancer.