Di Li

Optical Analysis of Hg2+ Ions by Oligonucleotide–Gold‐Nanoparticle Hybrids and DNA‐Based Machines

Zwei verschiedene optische Methoden fur die Analyse von Hg2+-Ionen, die auf der Bildung von Thymin-Hg2+-Komplexen beruhen, werden vorgestellt. Bei diesen Methoden werden die Hg2+-Ionen mithilfe aggregierter Goldnanopartikel und einer auf der DNA-Chemie basierenden Maschine (siehe Schema) analysiert.

Target-Responsive Structural Switching for Nucleic Acid-Based Sensors

Interest in the development of sensitive, selective, rapid, and cost-effective biosensors for biomedical analysis, environmental monitoring, and the detection of bioterrorism agents is rapidly increasing. A classic biosensor directly transduces ligand-target binding events into a measurable physical readout. More recently, researchers have proposed novel biosensing strategies that couple ligand-induced structural switching of biomolecules with advanced optical and electronic transducers. This approach has proven to be a highly general platform for the development of new biosensors. In this Account, we describe a series of electrochemical and optical nucleic acid sensors that use target-responsive DNA structures. By employing surface-confined DNA structures with appropriate redox labels, we can monitor target-induced structural switching of DNA or aptamer-specific small molecule probes by measuring electrochemical currents that are directly associated with the distance between the redox label and the electrode surface. We have also demonstrated significant improvements in sensing performance through optimization of the DNA self-assembly process at electrode surfaces or the introduction of nanomaterial-based signal amplification. Alternatively, gold nanoparticles interact differently with folded and unfolded DNA structures, which provides a visual method for detecting target-induced structural switching based on the plasmonic change of gold nanoparticles. This novel method using gold nanoparticles has proven particularly suitable for the detection of a range of small-molecule targets (e.g., cocaine) and environmentally toxic metal ions (e.g., Hg(2+)). Rational sequence design of DNA aptamers improves the sensitivity and increases the reaction kinetics. Recently, we have also designed microfluidic devices that allow rapid and portable mercury detection with the naked eye. This Account focuses on the use of bulk and nanoscale gold and DNA/aptamer molecules. We expect that researchers will further expand the analyte spectrum and improve the sensitivity and selectivity of nucleic acid sensors using functional biomolecules, such as DNAzymes, peptide aptamers and engineered proteins, and nanomaterials of different sizes, dimensions and compositions, such as carbon nanotubes, graphene, silicon nanowires, and metal nanoparticles or nanorods.

Highly Sensitive Electrochemical Sensor for Mercury(II) Ions by Using a Mercury-Specific Oligonucleotide Probe and Gold Nanoparticle-Based Amplification

We report a highly sensitive electrochemical sensor for the detection of Hg(2+) ions in aqueous solution by using a thymine (T)-rich, mercury-specific oligonucleotide (MSO) probe and gold nanoparticles (Au NPs)-based signal amplification. The MSO probe contains seven thymine bases at both ends and a mute spacer in the middle, which, in the presence of Hg(2+), forms a hairpin structure via the Hg(2+)-mediated coordination of T-Hg(2+)-T base pairs. The thiolated MSO probe is immobilized on Au electrodes to capture free Hg(2+) in aqueous media, and the MSO-bound Hg(2+) can be electrochemically reduced to Hg(+), which provides a readout signal for quantitative detection of Hg(2+). This direct immobilization strategy leads to a detection limit of 1 microM. In order to improve the sensitivity, MSO probe-modified Au NPs are employed to amplify the electrochemical signals. Au NPs are comodified with the MSO probe and a linking probe that is complementary to a capture DNA probe immobilized on gold electrodes. We demonstrated that this Au NPs-based sensing strategy brings about an amplification factor of more than 3 orders of magnitude, leading to a limit of detection of 0.5 nM (100 ppt), which satisfactorily meets the sensitivity requirement of U.S. Environmental Protection Agency (EPA). This Au NPs-based Hg(2+) sensor also exhibits excellent selectivity over a spectrum of interference metal ions. Considering the high sensitivity and selectivity of this sensor, as well as the cost-effective and portable features of electrochemical techniques, we expect this Au NPs amplified electrochemical sensor will be a promising candidate for field detection of environmentally toxic mercury.

Self-Catalyzed, Self-Limiting Growth of Glucose Oxidase-Mimicking Gold Nanoparticles

Size and shape of nanoparticles are generally controlled by external influence factors such as reaction temperature, time, precursor, and/or surfactant concentration. Lack of external influence may eventually lead to unregulated growth of nanoparticles and possibly loss of their nanoscale properties. Here we report a gold nanoparticle (AuNPs)-based self-catalyzed and self-limiting system that exploits the glucose oxidase-like catalytic activity of AuNPs. We find that the AuNP-catalyzed glucose oxidation in situ produces hydrogen peroxide (H(2)O(2)) that induces the AuNPs seeded growth in the presence of chloroauric acid (HAuCl(4)). This crystal growth of AuNPs is internally regulated via two negative feedback factors, size-dependent activity decrease of AuNPs and product (gluconic acid)-induced surface passivation, leading to a rapidly self-limiting system. Interestingly, the size, shape, and catalytic activities of AuNPs are simultaneously controlled in this system. We expect that it provides a new method for controlled synthesis of novel nanomaterials, design of smart self-limiting nanomedicine, as well as in-depth understanding of self-limiting systems in nature.

Parallel analysis of two analytes in solutions or on surfaces by using a bifunctional aptamer: applications for biosensing and logic gate operations.

A bifunctional aptamer that includes two aptamer units for cocaine and adenosine 5′‐monophosphate (AMP) is blocked by a nucleic acid to form a hybrid structure with two duplex regions. The blocked bifunctional aptamer assembly is used as a functional structure for the simultaneous sensing of cocaine or AMP. The blocked bifunctional aptamer is dissociated by either of the two analytes, and the readout of the separation of the sensing structure is accomplished by a colorimetric detection, by a released DNAzyme, or by electronic means that use Faradaic impedance spectroscopy or field‐effect transistors. In one configuration, the blocked bifunctional aptamer structure is separated by the substrates cocaine or AMP, and the displaced blocker units act as a horseradish peroxidase‐mimicking DNAzyme that permits the colorimetric detection of the analytes. In the second system, the blocked bifunctional aptamer hybrid is associated with a Au electrode. The displacement of the aptamer by any of the substrates alters the interfacial electron transfer resistance at the electrode surface, thus providing an electronic signal for the sensing process. In the third configuration, the blocked aptamer hybrid is linked to the gate of a field‐effect transistor device. The separation of the complex by means of any of the analytes, cocaine, or AMP alters the gate potential, and this allows the electronic transduction of the sensing process by following the changes in the gate‐to‐source potentials. The different systems enable not only the simultaneous detection of the two analytes, but they provide a functional assembly that performs a logic gate “OR” operation.

Inhibition of the In Vitro Replication of DNA by an Aptamer–Protein Complex in an Autonomous DNA Machine

DNA replication plays a central role in living organisms. Unregulated or uncontrollable DNA replication is well known to result in many pathological states, such as cancer, autoimmune diseases, and viral/bacterial infections. We report that an aptamer-protein complex could indirectly inhibit in vitro replication of DNA. An isothermal DNA machine based on the strand-displacement amplification is employed to support our assumption. An antithrombin aptamer sequence is rationally encoded into the DNA replication template. Once thrombin binds to the template, the as-formed aptamer-protein complexes can, in turn, become a barrier to the polymerase and inhibit the DNA replication activities in both static and dynamic modes. The inhibition is successfully confirmed by both fluorescence and gel-electrophoresis experiments. Considering the availability of a broad library of aptamers and the existence of various DNA/protein interactions, our results imply the possibility for the rational regulation of DNA replication in vivo.

Synthesis of Amphiphilic Alternating Polyesters with Oligo(ethylene glycol) Side Chains and Potential Use for Sustained Release Drug Delivery

Novel amphiphilic alternating polyesters, poly((N-phthaloyl-l-glutamic anhydride)-co-(2-(2-(2-methoxyethoxy)ethoxy)methyl)oxirane) (P(PGA-co-ME(2)MO)), were synthesized by alternating copolymerization of PGA and ME(2)MO. The structures of the synthesized polyesters were characterized by (1)H NMR, (13)C NMR, FT-IR, and GPC analyses. Because of the presence of oligo(ethylene glycol) (OEG) side chains, the polyesters could self-assemble into thermosensitive micelles. Dynamic light scattering (DLS) showed that these micelles underwent thermoinduced size decrease without intermicellar aggregation. In vitro methyl thiazolyl tetrazolium (MTT) assay demonstrated that the polyesters were biocompatible to Henrietta Lacks (HeLa) cells, rendering their potential for drug delivery applications. Two hydrophobic drugs, rifampin and doxorubicin (DOX), were loaded into the polyester micelles and observed to be released in a zero-order sustained manner. The sustained release could be accelerated in lower pH or in the presence of proteinase K, due to the degradation of the polyester under these conditions. Remarkably, in vitro cell experiments showed that the polyester micelles accomplished fast release of DOX inside cells and higher anticancer efficacy as compared with the free DOX. With enhanced stability during circulation condition and accelerated drug release at the target sites (e.g., low pH or enzyme presence), these novel polyesters with amphiphilic structures are promising to be used in sustained release drug delivery systems.

Amplified electrochemical detection of DNA through the aggregation of Au nanoparticles on electrodes and the incorporation of methylene blue into the DNA-crosslinked structure

The amplified electrochemical sensing of DNA is accomplished by the analyte-induced aggregation of nucleic acid-functionalized Au nanoparticles, deposition of the aggregates on a thiolated monolayer-functionalized electrode, and the voltammetric analysis of the redox-active methylene blue intercalated in the nucleic acid duplexes associated with the aggregates.

Targeted hydroxyethyl starch prodrug for inhibiting the growth and metastasis of prostate cancer.

Prostate cancer is one of the most prevalent malignancies among men. Although chemotherapy has been an effective therapeutic approach for treating metastatic prostate cancer, serious undesired side effects have hampered its wide application clinically. In this work, a pH-responsive LHRH-conjugated hydroxyethyl starch-doxorubicin (HES-DOX/LHRH) prodrug was facilely synthesized by conjugating oxidized HES (HES-CHO) with DOX and LHRH through an acid-sensitive Schiff base bond. The resulting prodrug spontaneously self-assembled into nanoscopic micelle with a radius of about 55xa0nm in an aqueous environment. HES-DOX/LHRH significantly improved the inxa0vivo tissue distribution of the drug. Compared to its non-targeted counterpart, targeted HES-DOX/LHRH demonstrated a greater inxa0vitro anti-proliferative capability toward mouse RM-1 prostate cells. More importantly, targeted HES-DOX/LHRH exhibited higher levels of anti-tumor and anti-metastasis activities against an RM-1-xenografted mouse model, with lower systemic toxicity compared to free DOX·HCl and non-targeted HES-DOX. Hence, these results revealed that targeted HES-DOX/LHRH possesses great potential application in clinical chemotherapy of metastatic prostate cancer.

In vitro evaluation of anticancer nanomedicines based on doxorubicin and amphiphilic Y-shaped copolymers

Four monomethoxy poly(ethylene glycol)-poly(L-lactide-co-glycolide)2 (mPEG-P( LA-co-GA)2) copolymers were synthesized by ring-opening polymerization of L-lactide and glycolide with double hydroxyl functionalized mPEG (mPEG-(OH)2) as macroinitiator and stannous octoate as catalyst. The copolymers self-assembled into nanoscale micellar/vesicular aggregations in phosphate buffer at pH 7.4. Doxorubicin (DOX), an anthracycline anticancer drug, was loaded into the micellar/vesicular nanoparticles, yielding micellar/vesicular nanomedicines. The in vitro release behaviors could be adjusted by content of hydrophobic polyester and pH of the release medium. In vitro cell experiments showed that the intracellular DOX release could be adjusted by content of P(LA-co-GA), and the nanomedicines displayed effective proliferation inhibition against Henrietta Lacks’s cells with different culture times. Hemolysis tests indicated that the copolymers were hemocompatible, and the presence of copolymers could reduce the hemolysis ratio of DOX significantly. These results suggested that the novel anticancer nanomedicines based on DOX and amphiphilic Y-shaped copolymers were attractive candidates as tumor tissular and intracellular targeting drug delivery systems in vivo, with enhanced stability during circulation and accelerated drug release at the target sites.