Introduction: There is still a lack of alternative therapeutic drugs to cope with the dilemma of BCG treatment failure in non-muscle-invasive bladder cancer (NMIBC). This study aimed to expand the panel of druggable targets and drug candidates for BCG incurable NMIBC.
Methods: A detailed analysis of 345 BCG-treated NMIBC patients was conducted on transcriptomics and proteomics data. LASSO and stepwise regression were performed to establish a prognostic signature and validated its robustness in multiple datasets. The Pearson coefficients between CERES score and prognostic score were calculated to evaluate the dependence of bladder cancer cell lines on 2249 genes. Bladder cancer cell line expression profiles and drug sensitivity data (AUC values) for 1929 drug agents archived in the CTRP and PRISM databases were used as the training dataset, expression profiles of NMIBC patients were used as the test dataset to calculate its estimated AUC values by running a ridge regression model with 10-fold cross-validation. Spearman correlations between recurrence-risk scores and estimated AUC values were calculated and drugs with coefficients < -0.30 and more sensitive to the high-risk group were retained and validated in the CMap database. BCG was co-cultured with T24 at 10 CFU/cell until 80% of the cells died, and BCG was removed to restore cell growth to more than 50%, and the cycle operation exceeded 6 months to obtain the BCG resistant T24 cell line (BCG-R-T24). CCK8 and Flow Cytometry Assay on BCG-R-T24 were performed to test the therapeutic effect.
Results: A 14-gene prognostic signature was established and it outperformed the published signatures in predicting BCG treatment outcome (AUC > 0.75 for recurrence, and > 0.80 for progression, respectively). CERES score correlation analysis revealed that the proliferation of bladder cancer cell lines was highly dependent on TOP1, EED, HLA-A, PDE6D, PSMB1, PSMB4 and RRM2. After qPCR and Western blot confirmed their over-expression in BCG-R-T24, they were identified as potential therapeutic targets. Fourteen drugs were screened from the CTRP and PRISM databases, and CMap initially identified the therapeutic potential of clofarabine (targeting RRM2), irinotecan and SN-38 (targeting TOP1). CCK8 assay demonstrated the inhibition effect of clofarabine, irinotecan, and SN-38 on BCG-R-T24. Flow cytometry showed that the apoptotic ratio of BCG-R-T24 cells increased significantly under clofarabine, irinotecan, and SN-38 treatment.
Conclusions: Clofarabine, irinotecan, and SN-38, especially clofarabine that targets RRM2, may have therapeutic value for NMIBC that underwent BCG treatment failure.
Source of Funding: This presented study was supported by the National Natural Science Foundation of China, Youth Science Fund Project (Grant Number: 81700615) and the Chen Xiao-ping Foundation for The Development of Science and Technology of Hubei Province (No.202094). The funders have no role in designing and completing this study.
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