ω‐3PUFAs Inhibit Gallbladder Cancer Through Enhancing STK31 Methylation

Gallbladder cancer (GBC) is an aggressive malignancy with limited treatment options. ω-3PUFAs have known anticancer activity, but their role in GBC remains unclear. The anti-GBC effects of ω-3PUFAs (DHA/EPA) were evaluated using in vitro assays and mouse xenograft models. Epigenetic changes were analyzed by RRBS, MSP, and MethyLight PCR. Downstream mechanisms were explored via proteomics, Co-IP, and Western blot. This study demonstrates that ω-3PUFAs suppress GBC progression by inducing promoter hypermethylation of STK31, which is an oncogene found in GBC by our previous study. MethyLight PCR revealed that ω-3PUFAs treatment significantly increased STK31 promoter methylation in GBC-SD cells from 34.1% ± 20.91% to 89.1% ± 13.11%. Moreover, STK31(DNMT1)/GSK3β pathway and COL4A1 (ITGB1) were particularly associated with ω-3PUFAs' inhibitory mechanisms to GBC. In vivo experimental results demonstrated that ω-3PUFAs significantly inhibited the growth of gallbladder cancer xenografts, with tumor growth inhibition rates of 38.6% for GBC-SD and 50.2% for NOZ at day 33, and effectively suppressed tumor cell metastasis in an intraperitoneal mouse model. This study provides first evidence that ω-3PUFAs target STK31 via epigenetic reprogramming, opening new avenues for dietary intervention and targeted therapy of GBC.