Supramolecular DNA Di-Tetrahedral Neoadjuvants Dynamically Prime Melanoma Microenvironment for Enhancing Radio-Immunotherapy Responses

The melanoma microenvironment is hallmarked by severe hypoxia and excessive regulatory T cells (Tregs), which significantly limits anti-melanoma efficacy of radiotherapy by impairing both its tumoricidal and immunostimulatory potential. Here, we report a supramolecular aptamer-engineered DNA di-tetrahedron nanoplatform (AEN) as a neoadjuvant treatment of melanoma radiotherapy, aiming to improve the radio-immunotherapeutic responses through dynamically remodeling the melanoma microenvironment. Specifically, DNA tetrahedra are separately loaded with vascular endothelial growth factor A inhibitor Thalidomide (Thal) and Treg-inhibitory drug Epirubicin (EPI) via DNA intercalation, followed by modification with plrogrammed death-ligand 1 (PD-L1) or cytotoxic T-lymphocyte-associated protein 4 (CTLA4) vascular endothelial growth factor A (VEGF-A)-binding aptamers and further conjugation with i-motif or chain-1 motifs to generate aptPD-L1-TDN@Thal and aptCTLA-4-TDN@EPI components, which eventually self-assemble into a di-tetrahedron through complementary binding between i-motif and chain-1. AENs dissociate into aptPD-L1-TDN@Thal and aptCTLA-4-TDN@EPI in acidic melanoma microenvironment, which are then taken up by PD-L1-overexpressing melanoma cells and CTLA4-overexpressing Tregs through ligand-mediated endocytosis. Thal inhibits tumor-intrinsic vascular endothelial growth factor A (VEGFA) signaling to normalize tumor vasculature for relieving hypoxia, while EPI inhibits Treg-intrinsic Foxp3-p65 signaling to abolish their immunosuppressive activity. The AEN treatment effectively remodels the melanoma microenvironment into a radiosensitive and immunosupportive state for enhancing the radio-immunotherapy response, providing an approach for melanoma radiotherapy in the clinic.