| Popis: |
Introduction: High-grade serous ovarian cancer (OvCa) remains the deadliest gynecologic cancer in the United States because it often presents at an advanced stage when the disease is incurable. Despite the impressive clinical success of cancer immunotherapy in patients with lung cancer and melanoma, it is largely ineffective in OvCa patients. Resistance to cancer immunotherapy is mediated by the apoptosis of T-cells, which can be prevented by blocking Fas ligand (FasL), a death gene. However, little is known about the expression and function of FasL in OvCa. Therefore, the aims of this study were to identify the cells in the OvCa tumor microenvironment that produce and secrete FasL, and to delineate the functional role of FasL in T-cell apoptosis within the OvCa tumor microenvironment. Methods: FasL protein localization and levels were evaluated in OvCa tumors by immunohistochemistry (IHC) using a FasL antibody on tissue microarrays (TMAs) containing primary and matching metastases from 72 OvCa patients. In-situ hybridization (ISH) using a FasL specific probe was performed on omental OvCa tumors from pre- (n=12) and post-chemotherapy (n=12) patients to localize FasL mRNA. To define the role of OvCa and endothelial cell derived FasL on T cell apoptosis, co-culture experiments were completed. First, FasL was knocked down with a FasL siRNA or neutralized with a FasL antibody in OvCa (OVCAR8 and OVCAR5) cells or endothelial (HUVEC) cells. Next, the OvCa or endothelial cells were co-cultured with primary human blood mononuclear cell-derived T cells. Subsequently, the level of FasL secreted was measured using a FasL specific enzyme-linked immunoassay, and early (4h; cleaved-caspase 3) and late (24h; TUNEL assay) T cell apoptosis were analyzed by flow cytometry. Results: FasL protein and mRNA are present in OvCa and endothelial cells in the tumor microenvironment of OvCa patients. Higher levels of FasL are detected in the endothelial cells when compared to the OvCa cells in these OvCa tumors. These results were confirmed in vitro as primary human endothelial cells secrete higher levels of FasL when compared to the OVCAR8 and OVCAR5 ovarian cancer cell lines. Functionally, both OvCa and endothelial cells stimulate early and late apoptosis in primary human blood mononuclear cell-derived T cells after a four hour co-culture. Knockdown or neutralization of FasL significantly inhibits the ability of OvCa or endothelial cells to induce early and late T cell apoptosis. In summary, FasL is localized to OvCa and endothelial cells in the OvCa tumor microenvironment, and both OvCa and endothelial cell derived FasL induce T cell apoptosis. Discussion: These findings indicate that blocking FasL may be an effective approach for reducing T-cell apoptosis and improving cancer immunotherapy response in OvCa patients. Citation Format: Grace Keegan, Hilary Kenny, Agnes Bilecz, Ricardo Lastra, Ernst Lengyel. Exploring the role of FasL in the ovarian cancer tumor microenvironment. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3655. |
