A Layered-Responsive DNA Tetrahedral Nanomachine for Precise Cancer Cell Imaging and Selective cGAS-STING Signaling Activation

The dual challenges of accurately imaging cancer cells and selectively activating immune signaling pathways require innovative DNA nanomachines with targeted recognition and multistimulus response capabilities. In this study, we developed a dual tetrahedral DNA nanomachine (DTDN) with layered responsiveness, which integrated a cascaded AND logic gate driven by sequential activation of reconfigurable DNA modules for precise cancer cell imaging and selective activation of the cGAS-STING pathway. Through AS1411 aptamer modification, DTDN achieved selective targeting and efficient cancer cell internalization. The overexpression of apurinic/apyrimidinic endonuclease 1 (APE1) in cancer cells first triggered the release of functional hairpins H1 and H2, after which miR-21 initiated a hybridization chain reaction (HCR) to generate long fluorescent nicked double-stranded DNA (dsDNA). The dsDNA product was recognized by cGAS, thereby activating the cGAS-STING pathway. The design of DNA tetrahedra gate prevented signal leakage by blocking the HCR toehold sequences. Moreover, the dual-locked cascade strategy exhibited high specificity and anti-interference capability, ensuring the specificity of cancer cell recognition and the activation of downstream events. Furthermore, the generated long nicked dsDNA not only provided an amplified fluorescence signal for cancer cell imaging but also acted as potent cGAS activators, thereby triggering the cGAS-STING pathway. Hence, this work provides a programmable, safe, and reliable nucleic acid nanoplatform for cancer diagnosis and cGAS-STING pathway-based regulatory therapy.