Photodynamic therapy (PDT) is an emerging effective treatment for cancer. However, the great promise of PDT for bladder cancer therapy has not yet been realized because of tumor hypoxia. To address this challenge, we fabricated O-generating HSA-MnO-Ce6 NPs (HSA for human serum albumin, Ce6 for chlorin e6, and NPs for nanoparticles) to overcome tumor hypoxia and thus enhance the photodynamic effect for bladder cancer therapy. The HSA-MnO-Ce6 NPs were prepared. We investigated the O generation of NPs and . The orthotopic bladder cancer model in C57BL/6 mice was established for study, and dual-modal imaging of NPs were demonstrated. Therapeutic efficacy of NPs for bladder cancer was evaluated. HSA-MnO-Ce6 NPs had an excellent performance in generating O upon reaction with HO at endogenous levels. Moreover, O generation was increased two-fold by using HSA-MnO-Ce6 NPs instead of HSA-Ce6 NPs in the presence of HO under 660 nm laser irradiation. cell viability assays showed that HSA-MnO-Ce6 NPs themselves were non-toxic but greatly enhanced PDT effects on bladder cancer cells under laser irradiation. near-infrared (NIR) fluorescence and magnetic resonance (MR) imaging suggested the excellent bladder tumor-targeting property of HSA-MnO-Ce6 NPs. O content in orthotopic bladder cancer was increased 3.5-fold after injection of HSA-MnO-Ce6 NPs as compared with pre-injection. Given the excellent tumor-targeting ability and negligible toxicity, HSA-MnO-Ce6 NPs were then used to treat orthotopic bladder cancer by PDT. The PDT with HSA-MnO-Ce6 NPs showed remarkably improved therapeutic efficacy and significantly prolonged lifetime of mice as compared with controls. This study not only demonstrated the great potential of HSA-MnO-Ce6 NPs for bladder cancer photodynamic ablation but also provided a new therapeutic strategy to overcoming tumor hypoxia.