Background: Gemcitabine/Cisplatin (GemCis) in combination with durvalumab have been established as a global new standard of care in first-line (1L) advanced biliary tract cancer (BTC) in the TOPAZ-1 trial (Oh et al. NEJM Evid, 2022). In our phase 2 study (NCT03046862, Oh et al. Lancet Gastroenterol Hepatol, 2022) exploring GemCis/durvalumab +/- tremelimumab, dynamics of PD-L1 expression after one cycle of treatment has been suggested as a predictor for clinical outcomes. Tumor metabolic activity assessed with ¹⁸F-FDG PET is usually associated with prognosis in solid tumors. However, the role of metabolic activity in patients treated with the combination of chemotherapy and immunotherapy has not yet been much studied, in terms of correlation with tumor immune landscape and prognosis. Methods: This study recruited 1L, advanced BTC patients in 3 different cohorts. ¹⁸F-FDG PET was conducted at baseline (pre-treatment) and at the time of first tumor response evaluation (post-treatment). Metabolic indices of the main tumor lesion were measured, including maximum standardized uptake values (SUVmax), metabolic tumor volume (MTV), and total lesion glycolysis (TLG). Metabolic response evaluation was assessed according to EORTC criteria. Tumor biopsies were done at baseline and after 1 cycle of treatment. Results: Among total 124 enrolled patients, 118 patients were included in this PET analysis. The early metabolic response rate was 54.3% (mCR 10.2%, mPR 44.1%, mSD 36.4%, mPD 9.3%). Pre-treatment metabolic parameters (SUVmax, MTV, TLG) were associated with disease status, previous operation history, and CRP level. High pre-treatment SUVmax was associated with poor progression-free survival (PFS) and overall survival (OS). Low post-treatment metabolism and large reduction in metabolism between pre- and post-treatment were associated with responder (vs non-responder), long PFS and long OS. In terms of PD-L1 expression and tumor metabolism, Tumor cell (TC) PD-L1 at pre-treatment was not associated with metabolic parameters; however, high immune cell (IC) PD-L1 expression showed significant association with lower MTV and TLG. In terms of dynamics of PD-L1 and tumor metabolism, post-treatment SUVmax was higher in patients with TC PD-L1 decrease. In patients with decrease of PD-L1 on both TC and IC (worst prognostic group), smaller reduction in metabolism between pre- and post-treatment was observed than the other patients. Conclusions: In BTC patients treated with 1L GemCis plus durvalumab +/- tremelimumab, tumor metabolic activity and its dynamics conferred prognostic impact. Furthermore, tumor metabolic activity has a potential to be associated with tumor immune landscape. Clinical trial information: NCT03046862.