DNA nanorobot for mitochondria-targeted microRNAs detection and tailored regulation

<b>Rationale:</b> Mitochondrial miRNAs (mitomiRs) are crucial regulators of mitochondrial functions and play pivotal roles in tumorigenesis and cancer progression. Nevertheless, direct monitoring mitomiRs and regulating mitochondrial function at the subcellular level remains challenging. <b>Methods:</b> In this study, we present a versatile DNA framework-based nanorobot for synchronous ultrasensitive detection of mitochondrial miRNAs (mitomiRs) and modulation of the mitochondria-associated apoptosis process. The DNA nanorobot features a tetrahedral nucleic acid framework as its structural body, two DNA hairpins (H1 and H2) as functional arms, and a mitochondria-targeting triphenylphosphine (TPP) group as the command center. The DNA nanorobot was comprehensively characterized for its morphological properties, mitochondria-targeting capacity, mitomiRs detection performance, DOX-loading and release behaviors, and antineoplastic effects both <i>in vitro and in vivo</i>. <b>Results:</b> Upon recognizing mitomiRs, the arms of the DNA nanorobot activate and trigger spatially restricted catalytic hairpin assembly (CHA) reactions with accelerated kinetics to generate amplified fluorescence signals. Additionally, the lipophilic anticancer drug doxorubicin (DOX) encapsulated within the DNA nanorobot induces reactive oxygen species (ROS) production, leading to mitochondria damage and promoting mitochondria-associated apoptosis in tumor cells. <b>Conclusion:</b> This newly developed DNA nanorobot provides a multifunctional platform for precise mitochondria-targeted diagnosis and enhanced therapeutic efficacy, advancing innovative strategies for mitochondria-focused tumor diagnosis and treatment.