Endogenous miRNA-Activated DNA Nanomachine for Intracellular miRNA Imaging and Gene Silencing

Abstract

The development of multifunctional nanoplatforms that integrate both diagnostic and therapeutic functions has always been extremely desirable and challenging in the cancer combat. Here, we report an endogenous miRNA-activated DNA nanomachine (EMDN) in living cells for concurrent sensitive miRNA imaging and activatable gene silencing. EMDN is constructed by interval hybridization of two functional DNA monomers (R/HP and F) to a DNA nanowire generated by hybridization chain reaction. After the target cell-specific transportation of EMDN, intracellular let-7a miRNA initiates the DNA nanomachine by DNA strand displacement cascades, resulting in an amplified fluorescence resonance energy-transfer signal and the release of many free HP sequences. The restoration of HP hairpin structures further activates the split-DNAzyme to identify and cleave the EGR-1 mRNA to realize gene silencing therapy. The proposed EMDN shows efficient cell internalization, good biological stability, rapid reaction kinetics, and the ability to avoid false-positive signals, thus ensuring reliable miRNA imaging in living cells. Meanwhile, the controlled activation of the split-DNAzyme activity regulated by the intracellular specific miRNA may be promising in the precise treatment of cancer. Collectively, this strategy provides a valuable nanoplatform for early clinical diagnosis and activatable gene therapy of tumors.

Self-Protected DNAzyme Walker with a Circular Bulging DNA Shield for Amplified Imaging of miRNAs in Living Cells and Mice

“…The detection limit was determined to be 10 pM by using 3σ method. Compared to the previous studies (SI Tables S4 and S5), ,− the proposed ES-AuNP system shows a relatively lower detection limit, demonstrating tremendous potential for application in the ultrasensitive detection of miR-21.…”

“…After incubation at 37 °C for 30 min, 200 mM Mg 2+ (the final concentration, 10 mM) was added to initiate the catalytic cleavage of substrates. At designated time points (5,10,15,20,40,60,90, and 120 min), the enzymatic cleavage was quenched by addition of 50 mM EDTA to chelate the cofactor Mg 2+ , and the resulting reaction solutions were analyzed by 10% denaturing polyacrylamide gel electrophoresis (dPAGE). Electrophoresis was carried out at 300 V in 0.5 × TBE buffer (45 mM Tris, 45 mM boric acid, 1 mM EDTA, pH 7.9) for 15 min on a gel electrophoresis instrument (BIO-RAD, U.S.).…”

Size-Discriminative DNA Nanocage Framework Enables Sensitive and High-Fidelity Imaging of Mature MicroRNA in Living Cells

“…Mature microRNAs (miRNAs) are small, noncoding, and conserved RNAs, which are processed from the large precursor microRNAs (pre-miRNAs) with stem-loop structures. − MiRNAs can bind messenger RNA (mRNA) to regulate the transcription and translation of target genes and thus have regulatory effects on cell proliferation and differentiation, stress response, and carcinogenesis. , Recent lines of evidence have confirmed the use of miRNAs as biomarkers for the diagnosis, treatment, and cell-level research of a disease. − Therefore, accurately monitoring intracellular tumor-related and low-abundance miRNAs is crucial for understanding their biological functions and the molecular mechanisms of cancer progression. Currently, prevalent sensing techniques such as colorimetry, electrochemiluminescence, and electrochemistry were developed to sensitively detect miRNAs.…”

Fuel‐Powered DNA Nanomachines for Biosensing and Cancer Therapy

“…Here, our groups proposed an endogenous miRNAtriggered DNA nanomachine based on entropy-driven catalytic reaction for simultaneous miRNA imaging and gene silencing. [67] A DNA nanowire serves as a carrier to deliver the diagnostic and therapeutic agents into live cells. Upon in the presence of let-7a miRNA, it can initiate DNA nanomachines to generate multiplexed FRET signals and free hairpin strands, in which FRET signal is used for miRNA imaging in real-time and hairpin strands further activate split-DNAzyme to cleave the EGR-1 mRNA for gene silencing-mediated cancer therapy.…”