Abstract Background: Peritoneal metastases (PM) are the leading cause of cancer-related death in gastric cancer (GC), with typical survival of 9 months and no effective targeted or immune therapies available. T-cell immunoglobulin and mucin domain-3 (TIM3) was initially identified on the surface of T helper 1 (Th1) cells and cytotoxic (CTL) lymphocytes as an immunosuppressive molecule that interacts with its ligands (GAL-9, CEACAM1, HMGB1, and PtdSer), and its expression is associated with advanced cancer stage. However, its functional role in promoting GC-associated PM (GCPM) and its clinical relevance in advanced GC remain unclear. Methods: multiple profiling and functional assays including scRNA-seq, CyTOF, qRT-PCR, cytokine arrays, kinase arrays, ELISA, and Western blotting were used to evaluate gene expression and secretion. TIM3 was genetically ablated using CRISPR/Cas9 in in macrophages to determine its functional role in tumor cells and T cells by co-culture system. In vivo, genetic and pharmacologic inhibition of TIM3 in macrophages co-innovation with tumor cells were tested in TIM3high vs TIM3KO macrophages on tumor growth in patient-derived xenograft (PDX) and patient-derived orthotopic (PDO) models and in the KP-luc2 syngeneic GC model. Results: We demonstrate that TIM3 and its ligands increased along the GC continuum and are associated with poor survival. Integrated omics analyses, multi-flow and co-immunofluorescent staining revealed that TIM3 is highly enriched in CD163+ tumor associated M2 immunosuppressive macrophages, which significantly increased tumor cell invasion in vitro, and promoted tumor growth in vivo. TIM3 depletion in macrophages reduced tumor cell malignant phenotypes and enhanced T-cell immunity from PBMCs or CD45+ immune cells isolated from malignant ascites in co-culture assays. Cytokine and kinase array analyses showed that TIM3 depletion in macrophages reduced production of key cytokines and chemokines from M2 macrophages. We further identified the RSK/CCL20/CCR6 axis as essential for the pro-tumor activity of TIM3+ macrophages. Finally, we show that TIM3 blockade or genetic knockout exhibits strong antitumor effects, especially when combined with anti-PD-1/PD-L1 immunotherapy or mitomycin C (MMC) chemotherapy. Together, these findings reveal a critical role for TIM3 in tumor associated macrophages and highlight the potential of TIM3 targeted immune therapy in GC patients with PM. Conclusion: TIM3 is highly enriched in tumor associated macrophages in GC with PM and plays a pivotal role in promoting GC progression and suppressing T-cell function. Thus, targeting TIM3 especially in combination with anti-PD1/PD-L1 or chemotherapy MMC may provide a novel therapeutic strategy for advanced GC patients with PM. Citation Format: Xiaodan Yao, Junsong Zhao, Yibo Fan, Yanting Zhang, Dipti Athavale, Curt Balch, Mikel Ghelfi, Anthony Pompetti, Jiankang Jin, Yong Ki Hong, Jamin Morrison, Madeline Torres, Francis Spitz, Generosa Grana, Shilpa S. Dhar, Linghua Wang, Jaffer A. Ajani, Shumei Song. TIM3+tumor associated M2 macrophages impair antitumor T cell immunity and promote gastric cancer progression and peritoneal metastasis abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 6124.
Yao et al. (Fri,) studied this question.