Common enriched genera include Escherichia coli, its subtype can produce Shiga toxin (Stx): toxin-producing strains (e.g., STEC) can cleave the 28S RNA of the ribosome of the host cell through the Stx2 subtype, inhibit protein synthesis and induce cellular growth. The Stx2 subtype inhibits protein synthesis and induces apoptotic mechanisms, leading to the development of renal tubular epithelial damage. Adhesion factor modality: the intimin protein encoded by LEE pathogenicity islands mediates tight adhesion between bacteria and host cells, thereby promoting inflammatory responses (12). Enterococcus faecalis: it produces reactive oxygen species (ROS) via glycerol phosphate oxidase (GPO), which activates the NF-κB pathway and upregulates IL-8 and TNF-α expression, which can induce the onset of oxidative stress. which can further promote biofilm formation: the Esp protein promotes biofilm formation and enhances its antibiotic resistance and persistent infections (13). Klebsiella pneumoniae (K. pneumoniae): releases pro-inflammatory factors, lipopolysaccharide (LPS) activates macrophages via TLR4, secretes IL-6 and IL-1β, and inhibits NLRP3 inflammatory vesicles which in turn leads to chronic inflammation (14). Disruption of the epithelial barrier: its secretion of proteases to degrade tight junction proteins (e.g., occludin), which increases transcellular permeability, can cause some disruption of the epithelial barrier (15).

The absence of certain flora of the environment may lead to the development of UTUC. For example, the reduced abundance of the symbiotic bacterium Lactobacillus crispatus showed a correlation with impaired barrier function. Its deficiency leads to a decrease in H2O2 and lactate secretion, allowing local pH to increase and promoting the proliferation of pathogenic bacteria (e.g., E. coli), and its S-layer protein (SlpB) deficiency impairs adhesion and reduces the protective effect on epithelial cells (16–18).

Increase in pro-tumor-associated genera: Acinetobacter, Anaerococcus and Sphingobacterium are significantly more abundant in the urinary microbiota of bladder cancer patients. The genus Streptococcus is significantly enriched in some patients with uroepithelial carcinoma and may be associated with tumorigenesis. Fusobacterium spp. (e.g. Clostridium nucleatum) have been postulated as potential pro-tumor pathogens that may promote cancer progression by modulating the tumor immune microenvironment (19).

Decrease in protective bacterial genera: The genera Serratia, Proteus and Roseomonas are reduced in abundance in patients with bladder cancer, which may suggest a potential role in tumor suppression (19).

Colibactin-producing Escherichia coli (pks+ E. coli) carrying the pks gene island can directly attack host DNA via its genotoxin colibactin. colibactin contains two cyclopropane slug structures that form alkylation adducts with DNA to form alkylating adducts that lead to DNA interstrand crosslinks (ICLs) (20). This cross-linking triggers double-strand breaks (DSBs) in DNA during its replication, activating a p53-dependent DNA damage response (DDR), and this sustained damage leads to p53 mutations and genomic instability (21). Meanwhile, some animal experiments have shown that specific adenine-colibactin adducts were detected in the colonic epithelium of mice infected with pks+ E. coli, and the level of cross-linking showed a positive correlation with tumor progression (20). In addition, colibactin can make repair defects more severe and lead to the accumulation of mutations by inhibiting the activity of the DNA repair protein FANCD2 (21).

Nitrosamines (e.g., NNK, NNAL) are metabolically activated by cytochrome P450 to generate alkyl diazene ions, which covalently bind to DNA bases (e.g., guanine) to form adducts, such as O^6-methylguanine (22). Such damage can lead to failure of mismatch repair as well as generation of point mutations in uroepithelial cells. In particular, they cause activation of oncogenes such as RAS (23). At the same time, elevated levels of nitrosamines in the urine of patients with chronic urinary tract infections and their metabolites lead to oxidative stress and mitochondrial dysfunction through preferential alkylation of mitochondrial DNA, which further contributes to the malignant transformation of the urinary tract epithelium (22).

Overexpression of B7-H4 (B7 homolog 4) is an important immunosuppressive molecule of the B7 family, which is highly expressed in UTUC, and promotes tumor immune escape by inhibiting effector T cell function. It has been found that B7-H4 enhances immunosuppressive signaling through glycolytic metabolic reprogramming in the UTUC tumor microenvironment, which in turn leads to the depletion of CD8+ T cell function (24). In addition, B7-H4 expression was significantly associated with clinical stage and poor prognosis of UTUC, also suggesting its value as a potential therapeutic target. Bidirectional regulation of the PD-1/PD-L1 axis Microbial metabolites such as short-chain fatty acids (SCFAs) and secondary bile acids can indirectly affect PD-L1 expression by regulating dendritic cell (DC) function. For example, it has been shown that Bifidobacterium bifidum reduced the expression of PD-L1 on the surface of tumor cells by lowering cholesterol levels in the tumor microenvironment, thereby enhancing the antitumor activity of CD8+ T cells (25). However, some microbial metabolites (e.g., tryptophan catabolic products) may also upregulate PD-L1 through activation of AhR receptors, creating an immunosuppressive environment (26). The urinary microbiota can modulate immune checkpoint expression via metabolites (e.g. indole-3-propionic acid, IPA).IPA up-regulates the Tcf7 gene and enhances CD8+ T-cell infiltration, which improves the efficacy of PD-1/PD-L1 inhibitors (27). In addition, tumors with high FGFR3 expression in UTUC often exhibit an ‘immune-cold’ microenvironment and are less responsive to immune checkpoint inhibitors (ICIs), whereas patients with low FGFR3 expression but abundant CD8+ T cells are more sensitive to ICIs (28), this may be related to microbiota-driven immune activation.

Metabolism and AhR pathway activation: Gut microbes metabolize tryptophan to products such as kynurenine (kynurenine), which inhibits CD8+ T cell proliferation and promotes differentiation of regulatory T cells (Tregs) through the activation of the aryl hydrocarbon receptor (AhR).AhR signaling also induces the expression of IDO1 (indoleamine 2,3-di oxygenase 1) expression, further exacerbating tryptophan depletion and creating an immunosuppressive microenvironment (26). And in UTUC, high expression of IDO1 presents a correlation with increased Foxp3+ Treg infiltration and poor patient prognosis (29). Bile acids and NKT cell regulation: intestinal microorganisms convert primary bile acids into secondary bile acids (e.g., deoxycholic acid), which diminishes the immune surveillance function of the liver and local tumors by inhibiting the expression of CXCL16 in hepatic sinusoidal endothelial cells and reducing the recruitment of CXCR6+ natural killer T (NKT) cells (30). In contrast, abnormal bile acid metabolism may be associated with an elevated risk of tumor liver metastasis in UTUC patients. Dual role of short-chain fatty acids (SCFAs): SCFAs (e.g., butyric acid, propionic acid) regulate T-cell differentiation through inhibition of histone deacetylase (HDAC): on the one hand, it promotes Treg differentiation to maintain immune tolerance, and on the other hand, it enhances the memory function of CD8+ T cells. In UTUC, the concentration gradient of SCFAs may have determined the direction of polarization of the immune microenvironment (31).

On the one hand CD8+ T-cell depletion: microbial metabolites (e.g., kynurenine) can induce CD8+ T-cell depletion via the AhR signaling pathway, while upregulating the expression of inhibitory receptors such as TIM-3, LAG-3, etc (26). On the other hand reg expansion: SCFAs and AhR ligands promote Treg differentiation and inhibit anti-tumor immune response. elevated Treg ratio in tumor tissues of UTUC patients correlates with PD-1 inhibitor resistance (29). regulation of T cell subsets in turn has a dynamic impact on the tumor microenvironment. Macrophage polarization and NK cell function: microbial-derived lipopolysaccharide (LPS) induces M2-type macrophage polarization through the TLR4/NF-κB pathway, secretes IL-10 and TGF-β, and promotes tumor angiogenesis and immunosuppression. Bifidobacteria also promote NK cell infiltration into tumor tissues by increasing intestinal permeability, which in turn enhances cytotoxicity (32). However, high expression of B7-H4 in UTUC may also inhibit NK cell activation (24). Dynamics of tertiary lymphoid structures (TLS): increased CD3+ and CD8+ T cell infiltration and TLS formation in metastases correlate with response to immunotherapy. The microbiome may promote TLS formation by regulating chemokines (e.g., CXCL13), but the exact mechanism still needs to be further verified (33).

Glycolysis and lactate metabolism are enhanced, and tumor cells generally suffer from the “Warburg effect”, i.e., they are preferentially supplied with energy through glycolysis rather than oxidative phosphorylation. The microbiome can exacerbate this metabolic profile through the following pathways. In terms of lactate production and utilization, certain commensal bacteria (e.g., Lactobacillus spp. in the intestinal or urinary tract) and pathogenic bacteria (e.g., Clostridium nucleatum Fusobacterium nucleatum) can produce lactate via glycolysis, which directly provides an energy substrate for tumor cells (34). In addition, microbial metabolites (e.g., butyrate) can induce the expression of lactate dehydrogenase (LDH) and α-hydroxybutyrate dehydrogenase (α-HBDH) in tumor cells, which promotes lactate production and metabolic cycling (35). Also the regulation of α-HBDH plays a role in the reprogramming of metabolic pathways. α-HBDH is one of the LDH isoenzymes, and its elevated levels are significantly associated with poor prognosis in UTUC patients (36). This enzyme may promote the adaptive metabolic process of tumor cells in a hypoxic environment by catalyzing the conversion of α-ketobutyric acid to α-hydroxybutyric acid, while releasing lactic acid to further acidify the microenvironment, which promotes invasion and immune escape (37).

Microbial metabolites show a bidirectional role in tumor progression, depending on the concentration, composition of the bacterial flora and the host environment. Specifically, short-chain fatty acids (SCFAs) such as butyrate can inhibit histone deacetylases (HDACs), exerting anti-inflammatory and anti-tumor effects in many contexts. However, at high concentrations or under specific conditions, butyrate may also activate pro-inflammatory pathways such as NF-κB, potentially promoting tumor progression (38). Polyamines: polyamines (e.g., spermidine) produced by certain bacteria (e.g., Pseudomonas fragilis) may also activate the mTOR pathway and promote the tumor cell proliferation process. Hydrogen sulfide (H₂S): H₂S produced by sulfate-reducing bacteria can promote genomic instability by inducing DNA damage and inhibiting the mitochondrial respiratory chain (39), which can further have an effect on metabolic reprogramming.

Some clinical studies found that elevated serum α-HBDH levels in UTUC patients were significantly correlated with tumor stage (TNM), pathological grade and lymph node metastasis (P<0.05), and were independent prognostic factors (HR = 1.36-2.04) suggesting that the α-HBDH levels may have a definite Correlation. The mechanisms may include there are microbe-host metabolic interactions: intestinal or urinary tract pathogenic bacteria (e.g., E. coli) activate the STAT3 pathway by inducing local inflammatory responses, and promote lactate accumulation and tumor cell invasion by up-regulating the expression of LDH/α-HBDH (35, 37). Metabolite feedback regulation process: high lactate environment further enriches acid-resistant flora (e.g., Lactobacillus spp.), forming a positive feedback loop that continuously drives glycolysis and α-HBDH activity, which in turn exerts a certain influence on metabolic reprogramming process, tumor staging (TNM), pathological grading and lymph node metastasis.