Phytochlorin‐Based Sonosensitizers Combined with Free‐Field Ultrasound for Immune‐Sonodynamic Cancer Therapy

Phytochlorins, a class of plant-derived tetrapyrroles, show great potential as sonosensitizers in sonodynamic therapy (SDT). The development of new phytochlorin-based sonosensitizers has significantly improved SDT, yet the absence of specialized sonodynamic systems limits their clinical translation. Herein, a dedicated ultrasound system along with a detailed step-by-step sonodynamic process from in vitro to in vivo is developed to activate phytochlorin-based sonosensitizers. Compared to standing-wave ultrasound, free-field ultrasound maintains stable acoustic pressure amplitudes and minimizes mechanical damage to cell membranes. In vitro experiments demonstrate that free-field ultrasound effectively activates naturally occurring phytochlorin, reducing the cavitation threshold for reactive oxygen species production and triggering immunogenic cell death. Furthermore, the intravenously injectable phytochlorin-based sonosensitizer (C34) enhances sonodynamic efficiency by reducing interfacial tension. Driven by in vivo free-field ultrasound, C34 effectively inhibits tumor growth in an orthotopic murine breast cancer model and elicits an immune response, preventing tumor metastasis. The reliable protocol provided by the free-field ultrasound system facilitates the activation of phytochlorin-based sonosensitizers while simultaneously stimulating the immune system, highlighting the potential of immune-sonodynamic therapy.