Extracellular vesicles (EVs) from immune cells represent a novel drug delivery system with inherent anti-tumor properties. To remodel the immunosuppressive tumor microenvironment and mediate multimodal therapy, we produced neutrophil nanovesicles at high yield and loaded them with drug (doxorubicin, DOX), near-infrared region II fluorescent dye (FD1080), and PD-1 inhibitor (TFA) through a simple extrusion method and further modified them with DSPE-PEG2000-cRGD to enhance their tumor-targeting ability. The RGD-NNV@FD1080&DOX/TFA exhibited excellent photothermal effect and efficiently suppressed tumor progression in mouse xenograft tumor, PDX, and lung metastasis models. Upon laser irradiation, the RGD-NNV@FD1080&DOX/TFA elicited the activation of anti-tumor immunity with increased infiltration of mature dendritic cells (DCs), CD8+ T cells, and decreased infiltration of regulatory T cells (Tregs) in tumors. Single-cell RNA sequencing revealed that the combination therapy increased the subtype of cytotoxic and memory T cells while decreasing that of exhausted T cells, and re-polarized M2 TAMs and N2 TANs. The re-challenge model further confirmed that RGD-NNV@FD1080&DOX/TFA exerted effective long-term immune memory in mouse models and displayed excellent biosafety in vivo. Overall, we designed a simple and potentially clinically applicable nanovesicle-based nanomedicine delivery system that exhibits a highly efficient anti-tumor effect by remodeling the tumor immune microenvironment and activating anti-tumor immunity.
