Mn-MOF–Driven Enhanced Activation and Delivery of Pistol Ribozyme: A Strategy for Checkpoint Silencing and Cancer Immunotherapy

Checkpoint blockade targeting PD-L1 has transformed cancer immunotherapy but remains constrained by systemic immune-related toxicity and limited efficacy in poorly infiltrated tumors. Here, we report a manganese-enhanced ribozyme nanoplatform (PS473@NKMOF-101-[Mn]) that enables precise PD-L1 gene silencing and innate immune activation. PS473 is a rationally engineered variant of the natural Pistol ribozyme that exhibits high catalytic efficiency at GU-rich motifs, which is further potentiated by the use of Mn²? as a cofactor. To overcome extracellular instability and ensure intracellular delivery, PS473 was encapsulated within a pH-responsive, Mn-doped metal–organic framework (NKMOF-101-[Mn]), which confers ribozyme protection and sustained Mn²? release in the acidic tumor microenvironment. In vitro, PS473@NKMOF-101-[Mn] suppressed PD-L1 expression and macrophage activation. In vivo, the nanoplatform achieved over 90% tumor inhibition in a B16F10 melanoma model and enhanced CD8? T-cell infiltration without systemic toxicity. Transcriptomic profiling further revealed the upregulation of type I interferon signaling and cGAS–STING pathway activation, indicating a synergistic mechanism combining gene knockdown and immunostimulation. This work presents a catalytically robust and immunologically active ribozyme-based nanotherapeutic, offering a promising strategy for targeted gene silencing and tumor immune reprogramming. Overall design: Tumor tissues were obtained at 48 h after the last intratumoral injection dministration, and RNA-seq analysis was performed on the tumor tissues of the blank control roup and multiple treatment groups (FreePS473, PS473@CPs, NKMOF-101-[Mn], M5@NKMOF-101-[Mn], PS473@NKMOF-101-[Mn]).