The FN1-ITGB4 Axis Drives Acquired Chemoresistance in Bladder Cancer by Activating FAK Signaling.

OBJECTIVE: While cisplatin-based chemotherapy is pivotal for advanced bladder cancer, acquired resistance remains a major obstacle. This study investigates key molecular drivers of this resistance and potential reversal strategies.

METHODS: We established GC (Gemcitabine and Cisplatin)-resistant T24-R and UC3-R cell lines from T24 and UM-UC-3 (UC3) cells. Transcriptomic and proteomic analyses identified differentially expressed molecules. Apoptosis and cell viability were assessed by flow cytometry and CCK-8 (Cell Counting Kit-8) assays, while RT-qPCR (Reverse Transcription Quantitative Polymerase Chain Reaction) and Western blot analyzed gene and protein expression. Immunofluorescence evaluated FAK (Focal Adhesion Kinase) phosphorylation, and a xenograft mouse model validated the findings in vivo.

RESULTS: Integrated transcriptomic and proteomic analysis identified FN1 (fibronectin) as a consistently upregulated top candidate in resistant cells (T24-R transcript log2FC = 2.8, protein log2FC = 0.9; UC3-R transcript log2FC = 3.7; all p < 0.001). Knockdown of FN1 reduced chemoresistance (Resistance Index: 5.2 in T24-R and 2.0 in UC3-R cells, p < 0.001) and enhanced apoptosis (approximately 4.5-fold in T24-R and 7.5-fold in UC3-R, p < 0.001). ITGB4 (Integrin Subunit Beta 4) was upregulated in resistant cells (transcript log2FC: 4.2 in T24-R and 3.03 in UC3-R; protein log2FC: 0.67 in T24-R; all p < 0.01). Critically, ITGB4 knockdown abolished the chemoresistance promoted by exogenous FN1, which was associated with increased FAK (Y397) phosphorylation.

CONCLUSION: Our results demonstrate that the FN1-ITGB4 axis drives chemoresistance in bladder cancer via FAK signaling. Targeting this axis represents a promising strategy to overcome chemoresistance.