KEY POINTS
- This translational preclinical study combined TCGA analysis, bladder cancer tissue cohorts, six human bladder cancer cell lines, subcutaneous and orthotopic mouse models, and low-USP15 patient-derived xenografts.
- USP15 expression was lower in bladder tumors than adjacent tissue across 16 paired immunohistochemistry samples, 22 paired immunofluorescence samples, and eight paired Western blot samples. Higher USP15 expression was associated with better survival in a 63-patient tissue microarray cohort and in TCGA bladder cancer data (p = 0.016).
- In clonogenic experiments using 0, 2, 4, and 8 Gy, USP15 overexpression reduced colony formation, increased radiation-induced apoptosis, and increased γ-H2AX foci and comet-assay DNA damage. USP15 knockdown produced the opposite phenotype, supporting a functional association with radioresistance.
- In subcutaneous xenografts, mice received 8 Gy on days 7, 17, and 22. USP15 loss reduced radiation response, whereas an orthotopic bladder model treated with 8 Gy on days 7, 12, 17, and 22 showed the greatest tumor suppression and survival benefit when USP15 overexpression was combined with irradiation.
- Mechanistically, mitochondrial USP15 directly interacted with FIS1 and removed ubiquitin at the K25 residue, stabilizing the FIS1 protein. This promoted mitochondrial fragmentation, reduced oxygen consumption and oxidative phosphorylation, and increased radiation-induced DNA damage; FIS1 knockdown reversed these effects.
- In USP15-silenced BIU87 xenografts, metformin 200 mg/kg/day plus four 8 Gy fractions produced significantly smaller tumors than either treatment alone (n = 4 per group). The finding was reproduced in low-USP15 patient-derived xenografts using metformin 300 mg/kg/day, where combined treatment again produced the lowest tumor volumes and weights (n = 3 per group).
- The study did not evaluate metformin-radiotherapy treatment in patients, define a clinically transferable radiation schedule, or provide human safety data. The very small animal groups also make the apparent treatment synergy exploratory rather than confirmatory.
CLINICAL TAKEAWAY
USP15 may identify a mitochondrial radioresistance pathway in bladder cancer, with metformin offering a potential way to restore radiosensitivity in USP15-low tumors. This is mechanistically strong but entirely preclinical evidence and should not alter bladder-preservation treatment outside a clinical trial.