The oxygen puzzle in FLASH radiotherapy: A comprehensive review and experimental outlook.

FLASH radiotherapy is attracting increasing interest because it maintains tumor control while inflicting less damage to normal tissues compared to conventional radiotherapy. This sparing effect, the so-called FLASH effect, is achieved when radiation is delivered at ultra-high dose rates (≥40 Gy/s). Although the FLASH effect has already been demonstrated in several preclinical models, a complete mechanistic description explaining why tumors and normal tissues respond differently is still missing. None of the current hypotheses fully explains the experimental evidence. A common point between many of these is the role of oxygen, which is described as a major factor, either through transient hypoxia in the form of dissolved molecules, or reactive oxygen species (ROS). Therefore, this review focuses on both forms of this molecule, retracing old and more recent theories, while proposing new mechanisms that could provide a complete description of the FLASH effect based on preclinical and experimental evidence. In addition, this manuscript describes a set of experiments designed to provide the FLASH community with new tools for exploring the post-irradiation fate of ROS and their potential biological implications.