Targeted Thermosensitive Ultrasound Contrast Agent-Enabled Ultrasound-Guided Precise Radiofrequency Ablation/Immunotherapy Synergistic Treatment for Hepatocellular Carcinoma

Radiofrequency ablation (RFA), a widely used minimally invasive therapy for primary liver cancer, induces tumor cell necrosis and immunogenic cell death, thereby activating antitumor immunity. However, its efficacy is often compromised by incomplete ablation of marginal regions and insufficiently robust immune activation, leading to a high risk of post-treatment recurrence and metastasis. Currently, integrating RFA with multifunctional nanomedicines is a promising strategy to overcome these limitations. Herein, we developed a multifunctional, targeted thermosensitive ultrasound contrast agent, termed TUCA(R837)-SP94, for synergistic RFA/immunotherapy against hepatocellular carcinoma (HCC). TUCA(R837)-SP94 integrates three major innovations: (1) tumor-targeting via the SP94 peptide enables enrichment at tumor sites; (2) under the thermal effects of RFA, the loaded perfluoropentane (PFP) undergoes a liquid–gas phase transition, enhancing the ultrasound contrast imaging effect; (3) RFA thermal effects trigger liposome release of imiquimod (R837). This study prepared TUCA(R837)-SP94 via the thin-film hydration method and characterized it, evaluated its ultrasound imaging performance and in vitro immune responses, and systematically investigated the synergistic therapeutic effects and immunomodulatory mechanisms of TUCA(R837)-SP94 combined with RFA through the establishment of mouse models of in situ and metastatic hepatocellular carcinoma. Results indicate that TUCA(R837)-SP94 combined with RFA significantly inhibits tumor growth both in situ and distally. This approach promotes DC maturation, increases the ratio of cytotoxic T cells to helper T cells in the tumor region, and reduces the proportion of regulatory T cells, while demonstrating excellent biosafety. Collectively, TUCA(R837)-SP94-based RFA/immunotherapy synergistic treatment represents a safe and effective theranostic platform for hepatocellular carcinoma, demonstrating promising potential for clinical translation and application.