A dual-locked and self-feedback CHA-DNAzyme nanomachine for tumor-specific molecular imaging in living cells

Background The detection and imaging of intracellular biomarkers can provide important information for the early diagnosis of cancer. Leveraging the inherent advantages of DNA such as biocompatibility, ease of synthesis, programmability, and sequence-specific recognition, DNA-based probes have been widely employed for biomarker analysis. Currently, a variety of DNA probes combined with signal amplification methods have been widely used to visualize biomarkers in living cells. However, conventional single-locked DNA probes cannot ensured the accurate diagnosis. Furthermore, the lack of efficient cascade signal amplification strategies makes it challenging to detect and image low-abundance biomarkers. Results In this work, we report a dual-locked, self-feedback catalytic hairpin assembly (CHA)-DNAzyme cascade amplification platform, termed hMNS@SCD. This platform utilizes MnO 2 nanoflowers as nucleic acid carriers and DNAzyme cofactor supplement, employing apurinic/apyrimidinic endonuclease 1 (APE1) and miR-21 as two molecular keys. When the APE1 and miR-21 existence, hairpin probe carry out the CHA cycle, leading to the assembly of a Y-shaped DNAzyme complex. When Mn 2+ is present, the Y-shaped DNAzyme can undergo a hybridization-cleavage-release cycle on the MB, leading to a gradual restoration of Cy5 fluorescence. In addition, the released target analogue from MB can trigger a new CHA cycle, generating more Y-shaped DNAzyme complexs and cleaving more MB, further amplifying the fluorescence signal. Through the self-feedback CHA-DNAzyme cascade amplification mechanism, hMNS@SCD effectively enhances the detection sensitivity, with a detection limit for miR-21 reaching 34.6 fM. Experimental data show that hMNS@SCD has high sensitivity, selectivity, and biocompatibility. Significance we constructed a dual-locked, self-feedback CHA-DNAzyme cascade nanomachine, which enables tumor-specific molecular imaging and effectively distinguishes between normal cells and cancer cells. In addition, due to the programmability of nucleic acid probes, this strategy can be readily adapted for detection and imaging of other low-abundance biomarkers, offering a promising way forward for tumor-specific molecular imaging and early cancer diagnosis.