RKLAHE-Mn metallopeptide nanoassembly amplifies mtDNA-driven STING signaling, boosting anti-PD-L1 efficacy in breast cancer.

The cGAS-STING pathway is a critical component of innate immunity, recognized as a powerful strategy for antitumor immunity. However, its therapeutic potential is often limited by insufficient cytosolic DNA cues, which are necessary to fully activate the pathway. Current immunotherapies, such as PD-L1 checkpoint blockade, face challenges from the highly immunosuppressive tumor microenvironment (TME). There is a pressing need for novel approaches that can effectively enhance STING activation and remodel the TME to improve therapeutic outcomes in cancer, particularly by leveraging endogenous danger signals like mitochondrial DNA (mtDNA).

Researchers engineered RKLAHE-Mn, a bioinspired metallopeptide nanoassembly, leveraging dynamic N,O-bidentate coordination between Mn(II) and a programmable hexapeptide scaffold. They used DFT calculations to confirm the thermodynamically favorable Mn-His/Glu N,O-coordination mode, orchestrating the self-assembly of uniform, chain-like spherical nanoparticles. In vitro, the study investigated the nanoassembly's impact on intracellular redox homeostasis, mitochondrial membrane potential, and mtDNA leakage, alongside its effects on STING signaling and immunogenic cell death. They also assessed RKLAHE-Mn's ability to potentiate innate immune signaling in dendritic cells, measuring markers like CD80, CD86, MHC class II, and cytokine secretion (IL-6, TNF-α, CXCL10). Finally, its therapeutic efficacy was evaluated in a murine breast cancer model, focusing on TME remodeling, CD8+ T cell infiltration, and synergy with anti-PD-L1 checkpoint blockade.

Upon internalization, RKLAHE-Mn triggered a rapid ROS burst and perturbed intracellular redox homeostasis. These cationic assemblies preferentially accumulated in mitochondria, leading to a loss of mitochondrial membrane potential and promoting mitochondrial DNA (mtDNA) leakage into the cytosol. The released mtDNA, combined with Mn(II)-dependent potentiation, robustly amplified STING signaling and elicited immunogenic cell death-associated responses. Furthermore, RKLAHE-Mn potentiated STING-related innate immune signaling in dendritic cells, driving their maturation with upregulated CD80, CD86, and MHC class II expression. This also led to increased secretion of IL-6, TNF-α, and CXCL10, alongside enhanced antigen presentation and cross-presentation. In a murine breast cancer model, RKLAHE-Mn effectively remodeled the immunosuppressive tumor microenvironment by promoting dendritic cell maturation and CD8+ T cell infiltration.

Notably, RKLAHE-Mn significantly enhanced the therapeutic efficacy of anti-PD-L1 checkpoint blockade.