Simple Summary Cyclooxygenase-2 (COX-2) expression causes several changes in the tumor microenvironment (TME) of ovarian cancer, which are associated with a low immunotherapy response in patients and immunosuppression in the TME, mainly due to cytotoxic T-lymphocyte-associated-protein-4 (CTLA-4) expression. However, its effect on the immune component of the TME has not been fully elucidated. In this study, using several integrated bioinformatic tools, we analyzed public transcriptomic data from four groups of ovarian cancer patients. We found that high expression of COX-2 is linked to poor survival, changes in the immune ecosystem, cell dysfunction, and lower effector activity of natural killer (NK) cells. Afterwards, we validated these results using flow cytometry and cytotoxicity assays on circulating NK cells from HGSOC patients. These cells were co-cultured with HGSOC cell lines while undergoing COX-2 and CTLA-4 blockade. Our results suggest that targeting COX-2 prior to the anti-CTLA-4 immunotherapy scheme, which takes advantage of NK cells cytotoxic capacity, may be a promising strategy to improve the effectiveness of immunotherapy for ovarian cancer patients.Abstract Chronic inflammation influences the tumor immune microenvironment (TIME) in high-grade serous ovarian cancer (HGSOC). Specifically, cyclooxygenase-2 (COX-2) overexpression promotes cytotoxic T-lymphocyte-associated protein-4 (CTLA-4) expression. Notably, elevated COX-2 levels in the TIME have been associated with reduced response to anti-CTLA-4 immunotherapy. However, the precise impact of COX-2, encoded by PTGS2, on the immune profile remains unknown. To address this, we performed an integrated bioinformatics analysis using data from the HGSOC cohorts (TCGA-OV, n = 368; Australian cohort AOCS, n = 80; GSE26193, n = 62; and GSE30161, n = 45). Employing Gene Set Variation Analysis (GSVA), MIXTURE and Ecotyper cell deconvolution algorithms, we concluded that COX-2 was linked to immune cell ecosystems associated with shorter survival, cell dysfunction and lower NK cell effector cytotoxicity capacity. Next, we validated these results by characterizing circulating NK cells from HGSOC patients through flow cytometry and cytotoxic assays while undergoing COX-2 and CTLA-4 blockade. The blockade of COX-2 improved the cytotoxic capacity of NK cells against HGSOC cell lines. Our findings underscore the relevance of COX-2 in shaping the TIME and suggest its potential as a prognostic indicator and therapeutic target. Increased COX-2 expression may hamper the effectivity of immunotherapies that require NK cell effector function. These results provide a foundation for experimental validation and clinical trials investigating combined therapies targeting COX-2 and CTLA-4 in HGSOC.