Triply activated localized DNA circuit based on dual enzyme-guaranteed and analyte-stimulated strategy for robust tumor-cell-specific microRNA imaging.

BACKGROUND

Localized catalytic hairpin assembly (LCHA) circuit has emerged as a potential tool for intracellular bioimaging, yet its application is limited by uncontrolled signal leakage, insufficient cell specificity, and unreliable readouts.

RESULTS

Herein, a triply activated LCHA (TA-LCHA) circuit was engineered by integrating an endogenous dual enzyme-guaranteed and analyte-stimulated strategy to enable robust tumor-cell-specific microRNA (miRNA) imaging. Initially, the TA-LCHA hairpin reactants were pre-blocked by incorporating the dual-enzyme recognition/cleavage site, inactivating their sensing function. Then, the miRNA sensing function of the blocked TA-LCHA reactants was specifically deblocked in tumor cells by two overexpressed enzymes, namely uracil DNA glycosylase (UDG) and apurinic/apyrimidinic endonuclease 1 (APE1). By virtue of the synergistic activation of endogenous UDG, APE1, and miRNA, this TA-LCHA circuit effectively reduced undesired signal leakage, achieving robust miRNA imaging with tumor cell specificity.

SIGNIFICANCE: The proposed TA-LCHA circuit represented a promising platform for precise biomolecules imaging in clinical diagnosis and biomedical applications.