Huanhuan Fan

Activatable fluorescence/MRI bimodal platform for tumor cell imaging via MnO2 nanosheet-aptamer nanoprobe.

A novel dual-activatable fluorescence/MRI bimodal platform is designed for tumor cell imaging by using a redoxable manganese dioxide (MnO2) nanosheet-aptamer nanoprobe. The redoxable MnO2 nanosheet acts as a DNA nanocarrier, fluorescence quencher, and intracellular glutathione (GSH)-activated MRI contrast agent. In the absence of target cells, neither fluorescence signaling nor MRI contrast of the nanoprobe is activated. In the presence of target cells, the binding of aptamers to their targets weakens the adsorption of aptamers on the MnO2 nanosheets, causing partial fluorescence recovery, illuminating the target cells, and also facilitating the endocytosis of nanoprobes into target cells. After endocytosis, the reduction of MnO2 nanosheets by GSH further activates the fluorescence signals and generates large amounts of Mn(2+) ions suitable for MRI. This platform should facilitate the development of various dual-activatable fluorescence/MRI bimodalities for use in cells or in vivo.

A Smart Photosensitizer–Manganese Dioxide Nanosystem for Enhanced Photodynamic Therapy by Reducing Glutathione Levels in Cancer Cells

Photodynamic therapy (PDT) has been applied in cancer treatment by utilizing reactive oxygen species to kill cancer cells. However, a high concentration of glutathione (GSH) is present in cancer cells and can consume reactive oxygen species. To address this problem, we report the development of a photosensitizer-MnO2 nanosystem for highly efficient PDT. In our design, MnO2 nanosheets adsorb photosensitizer chlorin e6 (Ce6), protect it from self-destruction upon light irradiation, and efficiently deliver it into cells. The nanosystem also inhibits extracellular singlet oxygen generation by Ce6, leading to fewer side effects. Once endocytosed, the MnO2 nanosheets are reduced by intracellular GSH. As a result, the nanosystem is disintegrated, simultaneously releasing Ce6 and decreasing the level of GSH for highly efficient PDT. Moreover, fluorescence recovery, accompanied by the dissolution of MnO2 nanosheets, can provide a fluorescence signal for monitoring the efficacy of delivery.

A Smart DNAzyme–MnO2 Nanosystem for Efficient Gene Silencing†

DNAzymes hold promise for gene-silencing therapy, but the lack of sufficient cofactors in the cell cytoplasm, poor membrane permeability, and poor biostability have limited the use of DNAzymes in therapeutics. We report a DNAzyme-MnO2 nanosystem for gene-silencing therapy. MnO2 nanosheets adsorb chlorin e6-labelled DNAzymes (Ce6), protect them from enzymatic digestion, and efficiently deliver them into cells. The nanosystem can also inhibit (1)O2 generation by Ce6 in the circulatory system. In the presence of intracellular glutathione (GSH), MnO2 is reduced to Mn(2+) ions, which serve as cofactors of 10-23 DNAzyme for gene silencing. The release of Ce6 generates (1)O2 for more efficient photodynamic therapy. The Mn(2+) ions also enhance magnetic resonance contrast, providing GSH-activated magnetic resonance imaging (MRI) of tumor cells. The integration of fluorescence recovery and MRI activation provides fluorescence/MRI bimodality for monitoring the delivery of DNAzymes.

DNA Dendrimer: An Efficient Nanocarrier of Functional Nucleic Acids for Intracellular Molecular Sensing

Functional nucleic acid (FNA)-based sensing systems have been developed for efficient detection of a wide range of biorelated analytes by employing DNAzymes or aptamers as recognition units. However, their intracellular delivery has always been a concern, mainly in delivery efficiency, kinetics, and the amount of delivered FNAs. Here we report a DNA dendrimer scaffold as an efficient nanocarrier to deliver FNAs and to conduct in situ monitoring of biological molecules in living cells. A histidine-dependent DNAzyme and an anti-ATP aptamer were chosen separately as the model FNAs to make the FNA dendrimer. The FNA-embedded DNA dendrimers maintained the catalytic activity of the DNAzyme or the aptamer recognition function toward ATP in the cellular environment, with no change in sensitivity or specificity. Moreover, these DNA dendrimeric nanocarriers show excellent biocompatibility, high intracellular delivery efficiency, and sufficient stability in a cellular environment. This FNA dendrimeric nanocarrier may find a broad spectrum of applications in biomedical diagnosis and therapy.

Gold nanorod-photosensitizer conjugate with extracellular pH-driven tumor targeting ability for photothermal/photodynamic therapy

AbstractChlorin e6-pHLIPss-AuNRs, a gold nanorod-photosensitizer conjugate containing a pH (low) insertion peptide (pHLIP) with a disulfide bond which imparts extracellular pH (pHe)-driven tumor targeting ability, has been successfully developed for bimodal photodynamic and photothermal therapy. In this bimodal therapy, chlorin e6 (Ce6), a second-generation photosensitizer (PS), is used for photodynamic therapy (PDT). Gold nanorods (AuNRs) are used as a hyperthermia agent for photothermal therapy (PTT) and also as a nanocarrier and quencher of Ce6. pHLIPss is designed as a pHe-driven targeting probe to enhance accumulation of Ce6 and AuNRs in cancer cells at low pH. In Ce6-pHLIPss-AuNRs, Ce6 is close to and quenched by AuNRs, causing little PDT effect. When exposed to normal physiological pH 7.4, Ce6-pHLIPss-AuNRs loosely associate with the cell membrane. However, once exposed to acidic pH 6.2, pHLIP actively inserts into the cell membrane, and the conjugates are translocated into cells. When this occurs, Ce6 separates from the AuNRs as a result of disulfide bond cleavage caused by intracellular glutathione (GSH), and singlet oxygen is produced for PDT upon light irradiation. In addition, as individual PTT agent, AuNRs can enhance the accumulation of PSs in the tumor by the enhanced permeation and retention (EPR) effect. Therefore, as indicated by our data, when exposed to acidic pH, Ce6-pHLIPss-AuNRs can achieve synergistic PTT/PDT bimodality for cancer treatment.

Recent advances in DNAzyme-based gene silencing

DNAzymes, generated through in vitro selection processes, are single-stranded DNA catalysts that can catalyze a wide variety of reactions, such as RNA or DNA cleavage and ligation or DNA phosphorylation. Based on specific cofactor dependence and potent catalytic ability, DNAzymes have been extensively used to develop highly sensitive and specific sensing platforms for metal ions, small molecules, and biomacromolecules. However, in spite of their multiple strong enzymatic turnover properties, few reports have addressed the potential application of RNA-cleaving DNAzymes as therapeutic gene-silencing agents. The main challenges are being met with low efficiency of cellular uptake, instability and the lack of sufficient cofactors for cellular or in vivo study, which have limited the development of DNAzymes for clinical application. In recent years, substantial progress has been made to enhance the delivery efficiency and stability of DNAzymes by developing variety of methods. Smart metal oxide nanomaterials have also been used to meet the requirement of cofactors in situ. This review focuses on the gene silencing application of DNAzymes as well as their physicochemical properties. Methods of increasing the efficacy of DNAzymes in gene therapy are also discussed: delivery systems to enhance the cellular uptake, modifications to enhance the stability and smart systems to generate sufficient cofactors in situ. Finally, some future trends and perspectives in these research areas are outlined.

Imaging Endogenous Metal Ions in Living Cells Using a DNAzyme-Catalytic Hairpin Assembly Probe

DNAzymes are a promising platform for metal ion detection, and a few DNAzyme-based sensors have been reported to detect metal ions inside cells. However, these methods required an influx of metal ions to increase their concentrations for detection. To address this major issue, the design of a catalytic hairpin assembly (CHA) reaction to amplify the signal from photocaged Na+ -specific DNAzyme to detect endogenous Na+ inside cells is reported. Upon light activation and in the presence of Na+ , the NaA43 DNAzyme cleaves its substrate strand and releases a product strand, which becomes an initiator that trigger the subsequent CHA amplification reaction. This strategy allows detection of endogenous Na+ inside cells, which has been demonstrated by both fluorescent imaging of individual cells and flow cytometry of the whole cell population. This method can be generally applied to detect other endogenous metal ions and thus contribute to deeper understanding of the role of metal ions in biological systems.