ABSTRACT Worldwide, colorectal cancer is responsible for approximately 0.4 million deaths annually, which represent approximately 10% of all cancer deaths. The main cause of death in colorectal cancer patients is hepatic metastasis. Although regional treatment options, including hyperthermic isolated hepatic perfusion (IHP) and percutaneous IHP, offer the benefits of both aggressive local treatment and limited systemic toxicity, the management of unresectable hepatic colorectal metastases remains a major unsolved issue and more effective novel regimens are needed. During the grant period, we propose developing a novel treatment strategy for hepatic colorectal metastases. Considering our previous studies, the combined treatment of hyperthermia, TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), and ferroptotic agent synergistically induces cytotoxicity and effectively enhances the tumoricidal efficacy of subcutaneous xenografts. In this grant application, we hypothesize that a combinatorial treatment of mild hyperthermia, the biologic agent TRAIL, and the ferroptotic agent artesunate (ART) is effective in treating unresectable hepatic colorectal metastases (HCM). The specific aims of this project are to (1) elucidate the mechanism of synergistic anti-tumor efficacy caused by hyperthermia in combination with TRAIL and ART treatment (HTA: hyperthermia + TRAIL + ART) in tumoroid models, and (2) investigate the preclinical efficacy of this HTA treatment in humanized rat isolated hepatic perfusion (IHP) models. The proposed studies for the first aim will establish tumoroid models with tumor tissues from patients with HCM and employ biochemical and molecular techniques to investigate the cell death mechanism induced by synergistic effects of the HTA treatment. For the second aim, we will develop humanized rat IHP models with tumor tissues from patients with HCM and then evaluate the therapeutic advantage of the HTA treatment. We believe that the successful outcome of this study will support the application of this multimodal approach to HCM.