Background <p>Gastric cancers are classified into four molecular subtypes according to The Cancer Genome Atlas (TCGA) classification: Epstein-Barr virus-positive (EBV-positive), microsatellite instability-high (MSI-H), chromosomal instability (CIN), and genomically stable (GS). Unlike MSI-H gastric cancer, GS and CIN subtypes exhibit immunologically inert microenvironments and demonstrate minimal response to immune checkpoint blockade (ICB), necessitating novel strategies to overcome immunotherapy resistance.</p> Methods <p>Through weighted gene co-expression network analysis (WGCNA), we identified the E3 ubiquitin ligase TRIM6 as inversely associated with MSI-H status. <i>TRIM6</i>-knockout murine models and subcutaneous tumors were subjected to flow cytometry, RNA sequencing, immunoblotting, and ubiquitination assays to characterize tumor-infiltrating lymphocytes (TILs), pathway activation, and TRIM6-mediated regulation of the cGAS-STING axis.</p> Results <p>Hypermethylation-mediated <i>TRIM6</i> downregulation distinguished MSI-H from microsatellite stable (MSS) gastric cancers. Clinically, <i>TRIM6</i> expression inversely correlated with cytotoxic T lymphocyte (CTL) infiltration and anti-PD-1/PD-L1 therapeutic efficacy. Mechanistically, TRIM6 catalyzed K27-linked polyubiquitination of cGAS, triggering its proteasomal degradation and consequent suppression of the cGAS-STING pathway. <i>TRIM6</i> ablation enhanced CD8⁺ T lymphocytes infiltration via cGAS-mediated innate immune response and synergized with anti-PD-L1 therapy in MSS gastric tumors.</p> Conclusions <p>Our results elucidate TRIM6-mediated suppression of antitumor immunity as a novel mechanism underlying ICB resistance in MSS gastric cancer, positioning TRIM6 as both a predictive biomarker and therapeutic target for immunologically cold subtypes.</p>