Drug resistance and recurrence are the major obstacles to bladder cancer chemotherapy. Our laboratory had reported that nucleophosmin1 was one of the differentially expressed proteins between bladder cancer cell lines PUMC-91 and PUMC-91/1.0ADM based on 2D-PAGE proteomics approaches. In this study, we want to explore the relationship among nucleophosmin1, drug resistance, and recurrence of bladder cancer, using normal bladder epithelia cell line SV-HUC-1, bladder cancer cell lines PUMC-91, PUMC-91 against gradient doses of adriamycin (0.3, 0.6, and 1.0 μg/ml), and bladder cancer tissue samples. The bladder cancer tissue samples were divided into two groups according to the interval of recurrence (<6 months and >2 years). The differences were detected by Western blotting and immunohistochemistry. The protein of nucleophosmin1 was differentially expressed with each other in SV-HUC-1, PUMC-91, PUMC-91/0.3ADM, and PUMC-91/1.0ADM (p < 0.05). Nucleophosmin1 was less expressed in later recurring (>2 years) bladder cancer tissue samples compared with samples that recurred <6 months (p = 0.035). The expression of nucleophosmin1 was independently associated with gradient drug resistance and recurrent frequency of bladder cancer. Nucleophosmin1 was a key regulator in either a drug-resistant bladder cancer or bladder cancer recurrence model. It may be possible to think nucleophosmin1 can provide more helpful information for clinical drug treatment of bladder cancer patients and frequently recurred ones.