Recently, diverse functional materials that take subcellular structures as therapeutic targets are playing increasingly important roles in cancer therapy. Here, particular emphasis is placed on four kinds of therapies, including chemotherapy, gene therapy, photodynamic therapy (PDT), and hyperthermal therapy, which are the most widely used approaches for killing cancer cells by the specific destruction of subcellular organelles. Moreover, some non-drug-loaded nanoformulations (i.e., metal nanoparticles and molecular self-assemblies) with a fatal effect on cells by influencing the subcellular functions without the use of any drug molecules are also included. According to the basic principles and unique performances of each treatment, appropriate strategies are developed to meet task-specific applications by integrating specific materials, ligands, as well as methods. In addition, the combination of two or more therapies based on multifunctional nanostructures, which either directly target specific subcellular organelles or release organelle-targeted therapeutics, is also introduced with the intent of superadditive therapeutic effects. Finally, the related challenges of critical re-evaluation of this emerging field are presented.