Pengcheng Gao

Protease-Responsive Prodrug with Aggregation-Induced Emission Probe for Controlled Drug Delivery and Drug Release Tracking in Living Cells

Controlled drug delivery and real-time tracking of drug release in cancer cells are essential for cancer therapy. Herein, we report a protease-responsive prodrug (DOX-FCPPs-PyTPE, DFP) with aggregation-induced emission (AIE) characteristics for controlled drug delivery and precise tracking of drug release in living cells. DFP consists of three components: AIE-active tetraphenylethene (TPE) derivative PyTPE, functionalized cell penetrating peptides (FCPPs) containing a cell penetrating peptide (CPP) and a short protease-responsive peptide (LGLAG) that can be selectively cleaved by a cancer-related enzyme matrix metalloproteinase-2 (MMP-2), and a therapeutic unit (doxorubicin, DOX). Without MMP-2, this prodrug cannot go inside the cells easily. In the presence of MMP-2, DFP can be cleaved into two parts. One is cell penetrating peptides (CPPs) linked DOX, which can easily interact with cell membrane and then go inside the cell with the help of CPPs. Another is the PyTPE modified peptide which will self-aggregate because of the hydrophobic interaction and turn on the yellow fluorescence of PyTPE. The appearance of the yellow fluorescence indicates the release of the therapeutic unit to the cells. The selective delivery of the drug to the MMP-2 positive cells was also confirmed by using the intrinsic red fluorescence of DOX. Our result suggests a new and promising method for controlled drug delivery and real-time tracking of drug release in MMP-2 overexpression cells.

A highly sensitive and facile graphene oxide-based nucleic acid probe: Label-free detection of telomerase activity in cancer patient's urine using AIEgens

Molecular beacon (MB)-based sensing platforms that consist of a fluorogen-quencher pair play an important role in medical and biological researches. However, the synthesis of both fluorogen and quencher in the nucleic acid probes will increase the burden of organic synthesis works and induce the difficulties for precisely controlling the relative distance between fluorogen and quencher, which may lead to false-positive and false-negative results. In this work, initially we report a single labeled MB (FAM-MB, with carboxyfluorescein as fluorogen and without quencher) thus simplifies MBs with the aid of graphene oxide (GO) to detect telomerase activity. To further simplify this structure, namely label-free strategy, we design a facile, sensitive and selective platform using a label-free beacon (AIE-MB, without fluorogen and quencher), based on aggregation-induced emission fluorogen (silole-R). Upon the addition of telomerase, AIE-MB induced comb-like DNA structure leads to high aggregation of silole-R and thus exhibits strong fluorescence emission. By exploitation of this, we can detect telomerase with superior sensitivity and demonstrate their applications in bladder cancer diagnosis. Compared to single-labeled FAM-MB based telomerase activity assay, the label-free AIE-MB induced method could perform the sensitive detection with high signal-to-background ratio.

Facile Probe Design: Fluorescent Amphiphilic Nucleic Acid Probes without Quencher Providing Telomerase Activity Imaging Inside Living Cells

Nowadays, the probe with fluorophore but no quencher is promising for its simple preparation, environmental friendliness, and wide application scope. This study designs a new amphiphilic nucleic acid probe (ANAP) based on aggregation-caused quenching (ACQ) effect without any quencher. Upon binding with targets, the dispersion of hydrophobic part (conjugated fluorene, CF) in ANAP is enhanced as a signal-on model for proteins, nucleic acids, and small molecules detection or the aggregation of CF is enhanced as a signal-off model for ion detection. Meanwhile, because of the high specificity of ANAP, a one-step method is developed powerfully for monitoring the telomerase activity not only from the cell extracts but also from 50 clinic urine samples (positive results from 45 patients with bladder cancer and negative results from 5 healthy people). ANAPs can also readily enter into cells and exhibit a good performance for distinguishing natural tumor cells from the tumor cells pretreated by telomerase-related drugs or normal cells. In contrast to our previous results ( Anal. Chem. 2015 , 87 , 3890 - 3894 ), the present CF is a monomer which is just the structure unit of the previous fluorescent polymer. Since the accurate molecular structure and high DNA/CF ratio of the present CF, these advanced experiments obtain an easier preparation of probes, an improved sensitivity and specificity, and broader detectable targets.

Role of outer surface probes for regulating ion gating of nanochannels

Nanochannels with functional elements have shown promise for DNA sequencing, single-molecule sensing, and ion gating. Ionic current measurement is currently a benchmark, but is focused solely on the contribution from nanochannels’ inner-wall functional elements (NIWFE); the attributes of functional elements at nanochannels’ outer surface (NOSFE) are nearly ignored, and remain elusive. Here we show that the role of NOSFE and NIWFE for ion gating can be distinguished by constructing DNA architectures using dual-current readout. The established molecular switches have continuously tunable and reversible ion-gating ability. We find that NOSFE exhibits negligible ion-gating behavior, but it can produce a synergistic effect in alliance with NIWFE. Moreover, the high-efficiency gating systems display more noticeable synergistic effect than the low-efficiency ones. We also reveal that the probe amount of NOSFE and NIWFE is almost equally distributed in our biomimetic nanochannels, which is potentially a premise for the synergistic ion-gating phenomena.Ion gating in biological channels is commonly controlled by functional elements. Here, the authors elucidate the contribution of outer-surface functional elements on ion gating of biomimetic nanochannels, providing insight into the design of effective nanochannel-based biosensors and electronics.

A photostable AIE fluorogen for lysosome-targetable imaging of living cells

Disruptive variation in intracellular pH and its fluctuations in lysosomes have a close relationship with the more acidic lysosome lumen of cancer cells (pH 4.5-5.5). Traditional lysosome-targeted probes, such as LysoTracker Green DND-26 (LTG) and LysoTracker Red DND-99 (LTR), can fluoresce when the weak base units in the probes are removed after donating protons under the photoinduced electron-transfer (PET) effect. However they can only be used at low concentration to avoid the aggregation-caused quenching (ACQ) effect and are also easily photobleached under continuous excitation irradiation, displaying low photostability. Herein, a tetraphenylethylene (TPE)-based lysosome-targetable fluorescence probe, TPE-CA, was synthesized, which could selectively monitor the pH change in subcellular organelles and exhibited a strong blue emission under an acidic condition with pH = 4. Using crystallographic, NMR and HRMS analyses, the mechanism regarding the pH dependent fluorescent performance of TPE-CA has been illustrated at the molecular level. In addition, experimental results show that TPE-CA is cell-permeable and biocompatible with HeLa, MCF-7 and HLF cells. The punctate fluorescent spots in the co-staining experiment of TPE-CA with LTG and LTR proves that the blue fluorescence spots of TPE-CA are indeed localized in the most acidic lysosome organelles. In particular, TPE-CA also inherits the aggregation-induced emission (AIE) feature of TPE, showing better photostability under continuous UV illumination compared with the commercial dyes (LTG and LTR). These results show that TPE-CA would be beneficial for understanding the acid environment of lysosomes in related cells and organs with potential biological significance.

Nanopore-Based DNA-Probe Sequence-Evolution Method Unveiling Characteristics of Protein–DNA Binding Phenomena in a Nanoscale Confined Space

Almost all of the important functions of DNA are realized by proteins which interact with specific DNA, which actually happens in a limited space. However, most of the studies about the protein-DNA binding are in an unconfined space. Here, we propose a new method, nanopore-based DNA-probe sequence-evolution (NDPSE), which includes up to 6 different DNA-probe systems successively designed in a nanoscale confined space which unveil the more realistic characteristics of protein-DNA binding phenomena. There are several features; for example, first, the edge-hindrance and core-hindrance contribute differently for the binding events, and second, there is an equilibrium between protein-DNA binding and DNA-DNA hybridization.

Highly Robust Nanopore-Based Dual-Signal-Output Ion Detection System for Achieving Three Successive Calibration Curves

In recent years, artificial stimuli-responsive bioinspired nanopores have attracted a lot of attention due to their unique property of confined spaces and flexibility in terms of shapes and sizes. Most of the nanopore systems demonstrated their transmembrane properties and applications in target detections. However, almost all of the nanopores can be used only once due to either the irreversible reactions between targets and probes or the plugged nanopores not easily being unplugged again. In this work, we propose a dual-signal-output nanopore system that could detect the cations (Hg(2+)) inducing the plugged nanopores. The detection system is highly recoverable by the anions (S(2-)) inducing the unplugged nanopores. More importantly, as far as we know, it is seldom reported for the same nanopores to achieve successive calibration curves for three times by subsequent reversible plug-unplug processes, which strongly demonstrates the high robustness of this novel nanopore-detection system. In addition, unlike monitoring the plug-unplug phenomena by only one type of signal, we combined the ionic current signal with the fluorescence output and could directly observe that the change of ionic current does in fact correspond to the plug-unplug of the nanopores by the target stimuli.

Publisher Correction: Role of outer surface probes for regulating ion gating of nanochannels

The original version of this Article contained an error in Fig. 3. The scale bars in Figs 3c and 3d were incorrectly labelled as 50 μA. In the correct version, the scale bars are labelled as 0.5 μA. This has now been corrected in both the PDF and HTML versions of the Article.

Distinct functional elements for outer-surface anti-interference and inner-wall ion gating of nanochannels

Over the decades, widespread advances have been achieved on nanochannels, including nanochannel-based DNA sequencing, single-molecule detection, smart sensors, and energy transfer and storage. However, most interest has been focused on the contribution from the functional elements (FEs) at the inner wall (IW) of nanochannels, whereas little attention has been paid to the contribution from the FEs at the nanochannels’ outer surface (OS). Herein, we achieve explicit partition of FEOS and FEIW based on accurate regional-modification of OS and IW. The FEIW are served for ionic gating, and the chosen FEOS (hydrophobic or charged) are served for blocking interference molecules into the nanochannels, decreasing the false signals for the ionic gating in complex environments. Furthermore, we define a composite factor, areas of a radar map, to evaluate the FEOS performance for blocking interference molecules.Nanochannels are often modified with functional elements, but most studies have focused on functionalizing only the inner wall. Here, the authors design nanochannels with distinct chemical modifications on the inner and outer surfaces, providing a route to dual-function channels.

Integrated Solid-state Nanopore Electrochemistry Array for Sensitive, Specific, and Label-free Biodetection

Nanopore ionic current measurement is currently a prevailing readout and offers considerable opportunities for bioassays. Extending conventional electrochemistry to nanoscale space, albeit noteworthy, remains challenging. Here, we report a versatile electrochemistry array established on a nanofluidic platform by controllably depositing gold layers on the two outer sides of anodic aluminum oxide (AAO) nanopores, leading to form an electrochemical microdevice capable of performing amperometry in a label-free manner. Electroactive species ferricyanide ions passing through gold-decorated nanopores act as electrochemical indicator to generate electrolytic current signal. The electroactive species flux that dominates current signal response is closely related to the nanopore permeability. Such well-characteristic electrolytic current-species flux correlation lays a premise for quantitative electrochemical analysis. As a proof-of-concept demonstration, we preliminarily verify the analytical utility by detection of nucleic acid and protein at picomolar concentration levels. Universal surface modification and molecule assembly, specific target recognition and reliable signal output in nanopore enable direct electrochemical detection of biomolecules without the need of cumbersome probe labeling and signal amplification.