Cancer remains one of the most important threats to human health, with its high incidence and mortality rates consistently being a focus of global medical research. In recent years, nanomedicines have attracted explosive growth in cancer therapy research due to their enhanced permeability and targeting capabilities. However, the biosafety of anticancer nanoparticles cannot be overlooked, as their potential toxicity may not only compromise therapeutic efficacy but also cause additional harm to the organisms. Thus, we explored the underlying toxicity mechanisms of anticancer nanoparticles from three aspects: payload, nanomaterials, and bio-corona in this article. To address these challenges, we proposed targeted strategies including enhancing nanoparticles targeting design, using the biodegradable natural polymers, and implementing advanced antifouling strategy. We hope that our work can provide a reference for more researchers to pay attention to the biosafety of anticancer nanoparticles and develop safer and more effective methods to fight cancer.