This work presents an integrated technology for assessing in vivo anti-cancer treatments in mice, based on various heating conditions. Bioluminescence imaging (BLI) was used to assess the physiological response of tumor and tumor microenvironment (TME) to heat treatment induced by magnetic resonance-guided high-intensity focused ultrasound (MRgHIFU). Transgenic tumor cells and mouse models with either constitutive or thermoinduced luciferase expressions were combined to monitor cell viability and heat-induced processes in the tumor and TME. BLI performed after MRgHIFU heating shows that a moderate increase in temperature (45°C) over 5 min can be exploited to promote heat-activable treatments in the tumor and its TME, without inducing direct cell death. A higher temperature rise over a shorter exposure time can induce cell death in the tumor, as revealed by a reduction in the BLI signal after treatment. Under these conditions, BLI also revealed that the TME can be stimulated by heat without inducing necrosis. These integrated technologies and models are useful to assess, in vivo in mice, the efficacy of various anticancer strategies exploiting local heat deposition by noninvasive MRgHIFU, including those combining tumor ablation with local drug administration using thermosensitive nanovehicles.