Abstract: Immune checkpoint inhibitors (ICIs) have shown limited clinical benefits for advanced gastric cancer (GC) because of a highly immunosuppressive tumour microenvironment (TME) that suppresses the therapeutic effect. Hua Chan Su (HCS) is a bioactive fraction of toad venom from traditional Chinese medicine that has shown anti-tumour effects in many preclinical and clinical studies. The particular function of changing tumour immunometabolism in this way is unknown. Here, we investigated whether HCS could enhance the therapeutic effect of PD-1 blockade in GC by targeting the aryl hydrocarbon receptor (AhR)–indoleamine 2,3-dioxygenase 1 (IDO1)–kynurenine (Kyn) signalling axis. First, we studied the anti-tumour effects of HCS on patient-derived organoids (PDOs) and HGC-27 cells. Integrated proteomics and untargeted metabolomics were then employed to identify key altered metabolic and signalling signatures under HCS treatment. A PDO/HGC-27-T cell co-culture system and a syngeneic xenograft mouse model were also used to study the therapeutic effect of HCS in combination with αPD-1 antibody. HCS inhibited the proliferation of cancer cells and induced apoptosis. In conjunction with HCS, αPD-1 can enhance the proliferation and cytotoxic effects of CD8 ⁺ T cells more effectively than either one alone. AhR overexpression and the addition of exogenous kynurenine both lost the immunomodulatory effect mechanistically. Collectively, our data show that HCS strengthens the cytotoxic activity of CD8 ⁺ T cells by targeting the AhR/IDO1 signaling axis to suppress the development of gastric cancer, and in conjunction with PD-1 inhibitors, offers robust mechanistic support for its application as an immunometabolic adjuvant in gastric cancer therapy.