Immunotherapy resistance presents a formidable challenge in tumor biology. While fibroblast growth factor receptor 3 (FGFR3) serves as a pivotal oncogenic driver in a multitude of cancers, the exploration of its role in immune checkpoint inhibitor (ICI) resistance remains scarce, thus impeding a deeper understanding of the tumor immune microenvironment (TIME) in the era of immunotherapy. Employing patient-derived urothelial carcinoma (UC) organoids and co-cultured systems, along with single-cell RNA sequencing (scRNA-seq), whole-exome sequencing (WES), bulk RNA-seq, and CUT CUT investigation; writing—original draft; software; methodology; validation; visualization; writing—review and editing; resources; supervision; data curation; formal analysis; project administration. Yuxuan Song: Validation; visualization; writing—review and editing; project administration; formal analysis; software; data curation; supervision; resources. Yun Peng: Conceptualization; investigation; writing—original draft; methodology; software; formal analysis; project administration; writing—review and editing; resources; data curation. Ran Yan: Conceptualization; investigation; visualization; validation; methodology; formal analysis; project administration. Yunze Niu: Conceptualization; investigation; visualization; validation; methodology; software; project administration; formal analysis; resources; data curation. Baoqiang Chen: Methodology; validation; software; formal analysis; project administration; resources; supervision; data curation. Jiaxing Lin: Methodology; validation; visualization; software; formal analysis; project administration; data curation; supervision. Jilin Wu: Methodology; validation; writing—review and editing; visualization; investigation; conceptualization. Shixiang Wang: Methodology; data curation. Yiqing Du: Conceptualization; investigation; funding acquisition; writing—original draft; project administration; formal analysis; software. Caipeng Qin: Conceptualization; investigation; resources; supervision; data curation; formal analysis; software. Yihan Lin: Conceptualization; investigation; writing—original draft; methodology; validation; visualization; writing—review and editing; project administration; formal analysis; software; data curation; resources; supervision. Tao Xu: Conceptualization; investigation; funding acquisition; writing—review and editing; visualization; validation; methodology; software; formal analysis; project administration; resources; supervision; data curation. All authors have read the final manuscript and approved it for publication. This study was supported by Noncommunicable Chronic Diseases-National Science and Technology Major Project (No. 2024ZD0525700), National Key Research and Development Program of China (No. 2023YFC2507000), National Natural Science Foundation of China (Nos. 82471866, 82271877, 82472912, and 82371840), Natural Science Foundation of Beijing, China (Nos. 7242150, 7264343, and QY25150), Beijing Municipal Science & Technology Commission (No. Z221100007422097), Capital's Funds for Health Improvement and Research of China (No. 2022-4-4087), Peking University People's Hospital Scientific Research Development Funds (Nos. RDGS2022-02, RDX2024-01, and RDEB2025-02). We acknowledge Sogen Biotechnology, Zhengzhou, Henan Province, Novogene Co. Ltd. , and the SolvingLab team for the advice and assistance they provided in data processing. We apologize for not being able to cite additional work owing to space limitations. The authors declare no conflicts of interest. All UC tissues were obtained with written informed consent from all patients prior to participation in the study. The Ethical Review Committee of Peking University People's Hospital reviewed and approved this study and the use of UC tissues (2026PHB128-001). All animal experiments were reviewed and approved by the Ethics Committee for Laboratory Animal Research at Peking University Health Science Center (2025PHB139-001, 2019PHB133-01). All aspects of this study complied with the Declaration of Helsinki. The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions. Due to ethical and legal restrictions, de-identified individual participant data and the accompanying data dictionary cannot be made publicly available. All data are available upon request from the corresponding author, subject to local rules and regulations. The data and scripts used are saved in GitHub (https: //github. com/Yunuuuu/UCImmunoErda). Public data used in the present study are available from the R package IMvigor210CoreBiologies (http: //research-pub. gene. com/IMvigor210CoreBiologies/). Supplementary materials (methods, figures, tables, graphical abstract, slides, videos, Chinese translated version, and updated materials) can be found in the online DOI or iMeta Science http: //www. imeta. science/. Figure S1. FGFR3 mutation status, expression profiles, and T cell cytotoxicity and exhaustion scores. Figure S2. Gene expression levels and multicolor immunofluorescence images. Figure S3. Generation of tumor-reactive T cells via co-culture system. Figure S4. Flow cytometric and immunofluorescence analysis of T-cell phenotype, FGFR3 expression, and organoid cytotoxicity in FGFR3-mutant UC models. Figure S5. FGFR3 inhibition increases NK cell proportion/function in co-culture via IRF2-dependent IFN-mediated anti-tumor immunity. Figure S6. STAT5 inhibition suppresses IFN-stimulated gene expression, which is partially rescued by combined erdafitinib treatment in co-culture system. Figure S7. FGFR3 inhibiting promotes chemokine secretion and recruits NK cells into TME via STAT5-IRF2 axis. Figure S8. Erdafitinib combined with anti-PD-1 immunotherapy reduces T-cell exhaustion and enhances cytotoxic function in co-culture system. Figure S9. Combination of FGFR3 inhibition (erdafitinib) and anti-PD1 immunotherapy is synergistic in FGFR3-mutant UC. Table S1. Clinical and molecular characteristics of UC patient cohort. Table S2. Summary of UCOs-PBMCs co-culture outcomes across 6 UC patients. Table S3. Homer known Motif enrichment results (homerSTAT5IRF2). Table S4. Differentially expressed genes (DEGs) identified in bulk RNA-seq analysis. Table S5. Research resource identifiers (RRIDs) for key reagents and resources. 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Jiang et al. (2026) studied this question.