Inducing Targeted Mild Hyperthermia in Murine Tumor Models through Photothermal Conversion of Near-infrared Light by Intratumoral Gold Nanorods

Mild hyperthermia (42-48 °C) is a well-established therapeutic modality that can induce controlled tumor cell death and stimulate anti-cancer immune responses. However, delivering heat precisely to tumor tissue while sparing surrounding healthy tissue remains a significant challenge. Traditional hyperthermia methods, such as isolated limb perfusion, require complex, invasive procedures and carry substantial risk of local toxicity and patient morbidity. In contrast, photothermal therapy using gold nanoparticles activated by near-infrared (NIR) light has emerged as a promising, less invasive strategy for achieving localized hyperthermia. Gold nanorods (GNRs), in particular, exhibit tunable optical properties and high photothermal conversion efficiency, making them ideal candidates for precise thermal modulation of tumor sites. Although this technique has shown considerable promise, especially for superficial and accessible tumors, reproducible delivery, spatial confinement, and safety remain active areas of refinement. In this protocol, we present our validated and optimized method for achieving localized, mild hyperthermia using intratumoral injection of biocompatible GNRs followed by short-duration, targeted NIR laser exposure. This approach enables rapid, controllable heating within the therapeutic range, promoting immunogenic tumor cell death and stimulating innate immune responses, mechanisms particularly relevant for immunologically "cold" tumors. Real-time temperature monitoring and local delivery ensure reproducibility, safety, and minimal systemic exposure. This streamlined protocol offers a robust and accessible platform for preclinical studies, supporting broader efforts to harness mild hyperthermia in cancer immunotherapy.