APE1-Responsive Adaptive DNA Nanomachines for Prolonged Intracellular Bioanalysis and Sustained Gene Silencing

Abstract Rapid cellular clearance severely restricts the effective analytical and functional windows of intracellular probes. Herein, we developed an endogenous enzyme-driven adaptive DNA nanomachine (TRZL) that synchronizes target recognition with in situ morphological transformation to overcome premature cellular clearance for prolonged intracellular theranostics. Specifically, a spatially confined hybridization chain reaction (HCR) circuit and an apurinic/apyrimidinic endonuclease 1 (APE1)-responsive module, which initially locks a DNAzyme, are integrated onto a rigid DNA prism framework. In tumor cells, APE1 recognition triggers the HCR circuit accompanied by fluorescence recovery, enabling sensitive APE1 detection with an ultralow limit of detection (LOD) of 9.60 × 10−4 U/mL. Concurrently, the continuous extension of the HCR polymers drives an in situ hydrodynamic size expansion from 37 nm monomers into 479 nm 3-dimensional networks. The resulting steric hindrance effectively mitigates rapid cellular clearance, successfully extending the signal retention window by maintaining 85.34% of the APE1-triggered signal over 24 h. Furthermore, benefiting from this aggregation-driven retention, the synchronously released DNAzyme can continuously cleave survivin mRNA to achieve sustained protein downregulation, thereby extending the functional window. Ultimately, this programmable size-adaptive strategy provides a highly versatile theranostic platform for precise bioanalysis and long-acting intervention.