The tumor microenvironment is a complex system that involves the interaction between malignant and neighbor stromal cells embedded in a mesh of extracellular matrix (ECM) components. Stromal cells (fibroblasts, endothelial, and inflammatory cells) are co-opted at different stages to help malignant cells invade the surrounding ECM and disseminate. Malignant cells have developed adaptive mechanisms to survive under the extreme conditions of the tumor microenvironment such as restricted oxygen supply (hypoxia), nutrient deprivation, and a prooxidant state among others. These conditions could be eventually used to target drugs that will be activated specifically in this microenvironment. Preclinical studies have shown that modulating cellular/tissue redox state by different gene therapy (GT) approaches was able to control tumor growth. In this review, we describe the most relevant features of the tumor microenvironment, addressing reactive oxygen species-generating sources that promote a prooxidative microenvironment inside the tumor mass. We describe different GT approaches that promote either a decreased or exacerbated prooxidative microenvironment, and those that make use of the differential levels of ROS between cancer and normal cells to achieve tumor growth inhibition.