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

ABSTRACT 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.