Endogenously Activated Multiple DNAzyme-Encoded Nanoflowers for High-Contrast Imaging of Long Noncoding RNA and...

Long noncoding RNAs (lncRNAs) are implicated in various physiological and pathological processes with the potential as diagnostic biomarkers and therapeutic targets. Watson-Crick base pairing-based DNA nanomaterials have been developed previously for diagnosis-guided therapy, but they are limited by undesired signal leakage and uncontrollable drug release due to nonspecific activation and nuclease susceptibility. Rolling circle amplification (RCA) products can noncanonically self-assemble into compact DNA nanoflowers with high loading performance and excellent nuclease resistance, but they are scarcely explored for intracellular analysis due to inefficient integration/release/activation of probes. Herein, we design endogenous acid-activatable ZnO-encapsulated RCA nanoflowers encoded by DNA-cleaving DNAzyme (D-DNAzyme) and RNA-cleaving DNAzyme (R-DNAzyme) for high-contrast imaging of lncRNA and controlled cancer therapy in living cells and mice. Upon the endocytosis of ZnO-RCA nanoflowers into the cells, the acidic microenvironment of tumor cells stimulates the decomposition of ZnO into Zn²⁺ that serves as DNAzyme cofactor and therapeutic reactive oxygen species producer. Zn²⁺-motivated D-DNAzyme-catalyzed detachment of RCA nanoflowers releases the deactivated R-DNAzyme. In the presence of lncRNA, the activity of R-DNAzyme is restored to cleave Cy5-labeled substrate probes on the AuNP surface with high turnover rate and specifically knocks down survivin gene, resulting in the generation of an enhanced fluorescence signal and R-DNAzyme-mediated gene silencing. Notably, the intrinsic resistance to nucleases and acid-stimulated detachment of RCA nanoflowers dramatically reduce the background signal leakage and improve the imaging contrast. This nanoplatform can accurately measure HOTAIR in living cells, real-time monitor HOTAIR in mice, and distinguish HOTAIR levels in healthy and cancerous breast tissues.