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However, predictive biomarkers for therapeutic response remain insufficiently defined. Although some genomic alterations have been implicated in modulating tumor immunogenicity and ICI sensitivity in other cancers, evidence in mUC remains limited and warrants further investigation.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>We retrospectively analyzed 67 patients with mUC treated with ICIs and performed targeted next-generation sequencing using a 440-gene cancer panel. Tumor mutational burden (TMB), genomic alterations, and clinical outcomes were evaluated to identify biomarkers associated with ICI response. Functional studies were conducted using urothelial carcinoma cell lines co-cultured with peripheral blood mononuclear cells (PBMCs) with or without anti-programmed death-ligand 1 (PD-L1) treatment to evaluate tumor viability, cytokine responses, and antigen presentation. Gene knockdown and overexpression experiments, quantitative PCR, immunofluorescence, and western blotting were used to assess cytokine production, antigen presentation machinery components, and major histocompatibility complex (MHC) class I expression. In vivo validation was performed using CRISPR\/Cas9-mediated knockout syngeneic murine models treated with anti-PD-L1 to assess tumor growth, immune infiltration, and therapeutic response through immunohistochemistry and flow cytometry.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      Responders to ICI monotherapy exhibited significantly higher TMB and enriched mutations in\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      ,\n                      <jats:italic>ADAMTSL1<\/jats:italic>\n                      ,\n                      <jats:italic>NSD1<\/jats:italic>\n                      , and\n                      <jats:italic>PRKDC<\/jats:italic>\n                      . Hotspot\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      mutations correlated with elevated TMB across multiple TCGA pan-cancer, breast and colorectal cohorts. Functionally,\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      -mutant tumor cells demonstrated enhanced cytotoxicity when co-cultured with PBMCs and anti-PD-L1, whereas\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      knockdown abolished this effect. Silencing\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      reduced expression of pro-inflammatory cytokines (interferon-\u03b3, tumor necrosis factor-\u03b1, interleukin (IL)-2, IL-6) and CD8\n                      <jats:sup>+<\/jats:sup>\n                      effector molecules (granzyme B, perforin A), while\n                      <jats:italic>PIK3CA E545K<\/jats:italic>\n                      overexpression restored these immune mediators. Mechanistically,\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      mutations enhanced antigen presentation via the IRF1-NLRC5-MHC-I axis, increasing HLA-A and B2M expression. In vivo,\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      -deficient tumors displayed diminished response to anti-PD-L1 therapy, reduced CD8\n                      <jats:sup>+<\/jats:sup>\n                      T-cell infiltration, and attenuated MHC-I expression.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>\n                      This study is the first to identify\n                      <jats:italic>PIK3CA<\/jats:italic>\n                      mutation as an immune-modulating driver that enhances tumor immunogenicity through activating IRF1-NLRC5-MHC-I pathway in urothelial carcinoma.\n                    <\/jats:p>\n                  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