A biomimetic nanointegrator synergizing “dual-throttle” immune checkpoint blockade and enhanced DAMPs-mediated immune activation for potent anti-tumor T‑cell immunity

Living cell-derived bioactive nanotherapeutics have advanced the development of highly effective cancer treatment strategies. In this study, we elaborately develop a multifunctional nanointegrator (DOX@DPNVs) from bioengineered nanovesicles (NVs) that act as a potent immunoactivator to overcome immunosuppression for enhanced tumor therapy. By genetically modifying tumor cells with PD-L1 knockout as well as PD-1 overexpression, these cells were subsequently treated with immunogenic cell death (ICD)-inducer (i.e., doxorubicin (DOX)) to trigger the release of damage-associated molecular patterns (DAMPs) in the extracellular environment. The resultant cell-involved mixture undergoes a one-step sonication and extrusion process, resulting in the formation of DOX@DPNVs, which are co-loaded with DAMPs and DOX and display PD-1 molecules on their surface. Consequently, the bioactive DOX@DPNVs enable the delivered DAMPs to combine with the DOX-induced ICD effect to accelerate DAMPs-mediated immune activation and effectively blockage the PD-1/PD-L1 axis for restoring cytotoxic function of T cells, eliciting an robust anti-tumor response in an aggressive squamous cell carcinoma mouse model. This naturally occurring NVs-based dual-throttle strategy provides an effective immunoactivation strategy to overcome immunosuppressive tumor environment, offering a promising platform for combating aggressive cancers. Nanovesicles derived from genetic engineered cancer cells (PD1-overexpressing and PD-L1 knockout) were modified with surface-enriched ICD-derived DAMPs and loaded with DOX. DOX@DPNVs efficiently induced ICD in cancer cells, whereas DAMPs and tumor-associated antigen presentation promoted dendritic cell maturation and antigen presentation. Synergistic immune checkpoint blockade with DOX@DPNVs enhances cytotoxic T-cell infiltration, decreased T-cell exhaustion and establishes a robust and long-lasting anticancer immune memory.