Nucleus, which plays a central role in the storage of genetic information and cell proliferation, growth and differentiation, is recognized as the optimal treatment site for cancer therapy. The nucleus is not only reactive oxygen species (ROS)-sensitive for photodynamic therapy (PDT), but also sensitive to hyperthermia for photothermal therapy (PTT). However, owing to the limited size of nuclear pores, it is extremely difficult for the phototherapeutic agent to pass through nuclear membrane for nucleus-targeting PDT/PTT synergistic therapy. Additionally, specific cancer cell or subcellular organelle-targeting capability of phototherapeutic agent is also very importance for improving therapeutic efficacy. Here, we rationally designed and synthesized a phototherapeutic agent BTPB-Py with two positive charges to enhance its electrostatic interaction with the nucleus, meanwhile, the lipophilic cation equipped BTPB-Py with potential mitochondria-targeting capability. In order to increase the specificity upon cancer cells of BTPB-Py, folic acid-modified amphiphilic polymer F127 (FA-F127) was synthesized to encapsulate BTPB-Py to obtain FA-BTPB-Py NPs. FA-BTPB-Py NPs with cancer cells, nucleus and mitochondria targeting ability display good near-infrared absorbance, excellent ROS generation for PDT, as well as satisfying photothermal efficiency up to 60.3 % for PTT and photothermal imaging. Notably, FA-BTPB-Py NPs can completely ablate tumor cells in 4T1 tumor-bearing mice via PDT/PTT synergistic therapy in vivo.