Biliary tract cancer (BTC) exhibits a poor prognosis and limited responses to current therapeutic strategies. While surgical resection followed by adjuvant S-1 therapy is the standard curative treatment for BTC, long-term postoperative remission is hardly achieved. Therefore, more effective perioperative strategies are urgently needed. Here, we show that an immune-cold tumor microenvironment (TME) in KRAS-mutated BTC correlated with resistance to postoperative adjuvant S-1 therapy. Surgically resected tumor specimens were collected from 31 BTC patients who received adjuvant S-1 therapy after surgery and were subjected to integrated immunogenomic analysis, including multiplexed immunohistochemistry staining and whole-exome sequencing. The analysis revealed a strong correlation between limited CD8⁺ T cell and antigen-presenting cell (APC) infiltration into the TME and KRAS mutations in BTC. In addition, the distances between tumor cells and APCs, as well as between APCs and CD8⁺ T cells, were significantly greater in the TME of KRAS-mutated BTC than in that of KRAS wild-type BTC. These findings indicate that interactions between effector T cells and APCs were impaired in the TME of KRAS-mutated BTC, thereby disrupting antitumor immune responses. Furthermore, wild-type KRAS and abundant CD8⁺ T cells and APCs correlated with a favorable prognosis following adjuvant S-1 therapy for BTC. Altogether, we propose a novel immunogenomic-based biomarker for optimizing perioperative chemotherapy for BTC.