Functionalized nanoprobes for disease-associated nucleic acid detection and neurodegenerative disease therapy

Abstract

Functionalized nanoprobes have been a powerful tool for biosensing, bioimaging, and therapeutics. Förster resonance energy transfer (FRET)-based biosensors with fluorescence as the output signal have a wide range of applications in detecting viral nucleic acids and monitoring disease-associated biomarkers. To overcome the unfavorable aggregation-caused quenching (ACQ) effect of conventional organic dyes, we introduced aggregation-induced emission fluorogens (AIEgens) as donor fluorophores into the FRET sensing platform to endow fluorescent nanoprobes with optimal signal-to-noise ratio and sensitivity due to distinctive fluorescence properties of AIEgens including high brightness, large stokes shift, strong resistance to photobleaching. Furthermore, inspired by the natural photosynthesis process, we developed a hybrid upconversion nanoparticle (UCNP)-based nanoreactor for Alzheimer’s disease (AD) therapy based on the generation of photocatalytic hydrogen gas to attenuate local oxidative stress. In the first work, we developed a FRET-based biosensor in which AIEgen-labelled oligonucleotide probes as donor fluorophores were immobilized on the surface of graphene oxide (GO) nanosheets as acceptor motif (AIEgen@GO). This biosensor was highly specific to the nucleic acid sequences of Orf1ab and N genes in the genome of SARS-CoV-2 virus. The sensing mechanism is based on dual fluorescence “turn-on” process in the presence of the target sequence. Here, the first-stage fluorescence recovery is due to dissociation of the AIEgen from GO surface in the presence of target viral nucleic acid. The second-stage enhancement of AIE-based fluorescent signal is caused by the formation of a nucleic acid duplex to restrict the intramolecular rotation of the AIEgen. Our work demonstrated that this AIEgen@GO nanoprobe could identify mimic Orf1ab, N genes, and SARS-CoV-2 plasmids with rapid detection around 1 h and good sensitivity at picomolar level without amplification. AIEgen@GO nanoprobe could be a promising tool in assisting the initial rapid detection of the SARS-CoV-2 viral sequence. In the second work, we further developed an AIEgen/polymeric molybdenum disulfide (MoS2)-based FRET nanoprobe for in situ detection of characteristic microRNA-125b (miR­125b) for early diagnosis of AD. To optimize the sensing platform for in vivo applications, we extended the emission wavelength of AIEgen from blue to red range to avoid autofluorescence of living systems. Accordingly, the proposed AIEgen@MoS2 nanoprobe was highly specific to miR-125b, a promising biomarker for early AD diagnosis. In the presence of the target, AIEgen hybridized with miR-125b to form a DNA/RNA duplex, causing the donor fluorophore to detach from the surface of MoS2, which simultaneously activated the dual fluorescence enhancement processes. The sensing performance of AIEgen@MoS2 was demonstrated by detecting miR-125b in both solution and a tau-based cell model in vitro with good sensitivity and specificity. Furthermore, the fluorescent nanoprobe also successfully demonstrated the capability of in situ monitoring of the endogenous miR-125b in tau-based AD mice. Therefore, AIEgen@MoS2 could be a promising tool for in situ and real-time monitoring of the AD-related microRNA biomarkers and thus would provide mechanistic insight into the early diagnosis of AD. In the third work, we designed a UCNP-based artificial nanoreactor for near-infrared (NIR) light-triggered in situ hydrogen gas (H2) generation to scavenge reactive oxygen species (ROS) for AD therapy. This multi-component nanoreactor contained platinum nanoparticles (Pt NPs) and ascorbic acid encapsulated by cross-linking vesicles, which comprised two photosensitizers, chlorophyll a (Chla) and indoline dye (Ind), absorbing the red and green luminescence from UCNP coupled on the surface. In the nanoreactor system, the excited electrons separated from Chla and Ind were quickly transferred to Pt NPs to combine with protons from ascorbic acid, facilitating rapid activation of the photosynthesis of H2, locally providing a high therapeutic concentration. Our results successfully demonstrated that the artificial nanoreactor could efficiently photosynthesize H2 to restore ROS homeostasis, repair mitochondrial damage, and attenuate hyperphosphorylated tau in AD mice. Such an artificial nanoreactor with efficient H2 generation has great potential to provide a new window for AD treatment.