<p>We investigated the role of the intraperitoneal immune microenvironment in the therapeutic response of gastric cancer with peritoneal metastasis to intraperitoneal paclitaxel. Gastric cancer with peritoneal metastasis has a poor prognosis, even with systemic chemotherapy. Although intraperitoneal paclitaxel administration has demonstrated clinical benefit, treatment resistance remains a major hurdle. Between 2017 and 2023, 30 patients undergoing staging laparoscopy were prospectively enrolled. Immune cell subsets and checkpoint marker expression in peritoneal lavage fluid were analyzed using flow cytometry and immunohistochemistry. Their associations with clinical outcomes were assessed using univariable and multivariable analyses, including Kaplan–Meier, logistic regression, and Ridge regression analyses. Elevated CD4⁺ T and CD19⁺ B cell levels were associated with inferior progression-free survival, whereas increased natural killer T-like cell levels correlated with improved survival. High FOXP3 and CTLA-4 expression were linked to worse outcomes. Ridge regression identified CD4⁺ T cell levels and CTLA-4 expression as unfavorable factors, while the presence of natural killer cells emerged as a favorable factor. An immunoregulatory-dominant immune landscape may drive resistance to intraperitoneal paclitaxel. Immune profiling of ascitic fluid could identify patients likely to benefit from immune-modulating strategies, including those targeting FOXP3⁺ and CTLA-4⁺ regulatory cells.</p>