Sonodynamic therapy (SDT) has emerged as a promising noninvasive approach for tumor therapy. However, the effectiveness of traditional inorganic semiconductor sonosensitizers is hindered by rapid electron (e) and hole (h) recombination under ultrasonic (US) stimulation, as well as the hypoxic and reductive conditions of tumor microenvironment (TME), which limit the generation of reactive oxygen species (ROS). Herein, a ruthenium (Ru) single-atom nanozyme-driven superimposition-enhanced titanium dioxide-based sonosensitizer (Ru/TiO SAE) is presented that features sufficient oxygen vacancies and high e/h separation efficiency. Through synchrotron radiation-based X-ray absorption spectroscopy and extended X-ray absorption fine structure analysis it is confirmed that oxygen vacancies in TiO nanoparticles promote the immobilization of single-atomic Ru, forming Ru-O₄ active sites. Density functional theory calculations demonstrate that oxygen vacancies alter the electronic structure of nanosensitizer, enhanced e/h separation, increasing oxygen adsorption, and accelerating reaction kinetics under US stimulation, ultimately improving ROS production. Moreover, Ru/TiO SAE boosts sonodynamic efficacy by mitigating the hypoxic and reductive TME. This is attributed to its catalase- and glutathione peroxidase 4-like activities, which facilitate the generation of ROS and trigger lipid peroxidation-mediated ferroptosis. These findings highlight the innovative role of single-atom Ru in optimizing sonosensitizers for SDT-induced ferroptosis, demonstrating its potential for advancing cancer therapy.