A Cascaded DNA Nanocircuit for Multi‐Signal‐Responsive Precision siRNA Delivery in Cancer Therapy
ABSTRACT Precision control over nucleic acid delivery remains a critical challenge in cancer therapy, particularly for siRNA‐based gene silencing, where off‐target effects limit clinical translation. Herein, we report a programmably engineered DNA nanocircuit with cascaded dual‐AND logic gates, which enables the development of a spatiotemporally controlled siRNA delivery strategy for precision cancer therapy. The DNA nanocircuit is engineered to respond to three tumor‐specific signals in a sequential manner: extracellular acidic pH, membrane‐overexpressed nucleolin (NCL), and intracellular glutathione (GSH). The first AND gate is activated by the co‐occurrence of acidic pH and NCL, triggering a conformational rearrangement that generates a molecular output. This integrated output, combined with intracellular GSH, serves as the dual input to co‐activate the second AND gate, initiating siRNA release via a cascade reaction inherent to the DNA circuit. As a proof‐of‐concept, when harnessing this DNA circuit in a temozolomide (TMZ)‐resistant glioblastoma (GBM) mouse model, we demonstrate that this design ensures highly selective release of siPARP1 in GBM cells, achieving efficient PARP1 silencing, reversed TMZ resistance, and minimized off‐target toxicity. Collectively, the cascaded dual‐AND logic, enabled by precise DNA sequence programming, represents a generalizable strategy for multi‐signal‐responsive delivery systems, highlighting the potential of DNA circuits in precision cancer therapy.