Tumor-specific in situ IL-12 expression combined with αPD-L1 suppresses melanoma growth by remodeling the tumor immune microenvironment.

Interleukin-12 (IL-12) potently activates antitumor immune responses and compensates for the challenge of insufficient T cell activation and infiltration faced in immune checkpoint therapy. However, its clinical application is limited by severe systemic toxicity and the upregulation of PD-L1 on tumor cells during treatment. Here, we developed iLANpTyr-IL-12, a lipid-assisted nanoparticle system loaded with tyrosinase (Tyr) promoter-driven IL-12 plasmids, designed to achieve specific and sustained IL-12 expression exclusively in melanoma cells. The secreted IL-12 promoted dendritic cell maturation, T cell activation and proliferation, and IFN-γ secretion. Notably, iLANpTyr-IL-12 treatment upregulated PD-L1 expression on tumor cells, suggesting a negative feedback loop that supports the rationale for combination with anti-PD-L1 (αPD-L1). In a subcutaneous B16-F10 melanoma model, iLANpTyr-IL-12 monotherapy significantly enhanced tumor suppression and increased intratumoral infiltration of T cells and NKT cells, along with a decreased M2/M1 macrophage ratio. When combined with αPD-L1, it produced an enhanced antitumor effect, achieved a tumor growth inhibition rate of 83% and further augmented intratumoral CD8+ T cell infiltration. Additionally, no significant systemic toxicity or organ damage was observed. Collectively, these findings demonstrate that iLANpTyr-IL-12 enables tumor-specific IL-12 expression and, when combined with αPD-L1, provides a promising cytokine-immune checkpoint combination immunotherapy with reduced off-target toxicity.