Liqian Gao

Self-Assembling Peptide Nanofibrous Hydrogel as a Versatile Drug Delivery Platform

Molecular hydrogels have been widely explored in various biomedical applications, such as cell culture, tissue engineering and drug delivery. Peptide-based hydrogel nanoparticles represent a promising alternative to current drug delivery approaches and cell carriers for tissue engineering, due to their encapsulation stability, water solubility and biocompatibility. This review focuses on recent advances in the use of self-assembling peptide nanogels for applications in drug delivery. We firstly introduce a self-assembly mechanism for small molecules used in the peptide hydrogel, and then describe recent methods for controlling the assembly of molecular hydrogelations. A particular emphasis is placed on recent advances in the use of different types of peptide hydrogels as drug delivery carriers. Lastly, the current challenges and future perspectives for self-assembling peptide hydrogels in drug delivery applications are discussed.

Target identification of natural and traditional medicines with quantitative chemical proteomics approaches

Natural and traditional medicines, being a great source of drugs and drug leads, have regained wide interests due to the limited success of high-throughput screening of compound libraries in the past few decades and the recent technology advancement. Many drugs/bioactive compounds exert their functions through interaction with their protein targets, with more and more drugs showing their ability to target multiple proteins, thus target identification has an important role in drug discovery and biomedical research fields. Identifying drug targets not only furthers the understanding of the mechanism of action (MOA) of a drug but also reveals its potential therapeutic applications and adverse side effects. Chemical proteomics makes use of affinity chromatography approaches coupled with mass spectrometry to systematically identify small molecule-protein interactions. Although traditional affinity-based chemical proteomics approaches have made great progress in the identification of cellular targets and elucidation of MOAs of many bioactive molecules, nonspecific binding remains a major issue which may reduce the accuracy of target identification and may hamper the drug development process. Recently, quantitative proteomics approaches, namely, metabolic labeling, chemical labeling, or label-free approaches, have been implemented in target identification to overcome such limitations. In this review, we will summarize and discuss the recent advances in the application of various quantitative chemical proteomics approaches for the identification of targets of natural and traditional medicines.

A general strategy for site-directed enzyme immobilization by using NiO nanoparticle decorated mesoporous silica.

Mesoporous materials have recently gained much attention owing to their large surface area, narrow pore size distribution, and superior pore structure. These materials have been demonstrated as excellent solid supports for immobilization of a variety of proteins and enzymes for their potential applications as biocatalysts in the chemical and pharmaceutical industries. However, the lack of efficient and reproducible methods for immobilization has limited the activity and recyclability of these biocatalysts. Furthermore, the biocatalysts are usually not robust owing to their rapid denaturation in bulk solvents. To solve these problems, we designed a novel hybrid material system, mesoporous silica immobilized with NiO nanoparticles (SBA-NiO), wherein enzyme immobilization is directed to specific sites on the pore surface of the material. This yielded the biocatalytic species with higher activity than free enzyme in solution. These biocatalytic species are recyclable with minimal loss of activity after several cycles, demonstrating an advantage over free enzymes.

Activity‐based high‐throughput determination of PTPs substrate specificity using a phosphopeptide microarray

Protein tyrosine phosphatases (PTPs) constitute a large family of enzymes that play key roles in cell signaling. Malfunctions of PTP activity have been linked to major human diseases including cancer. One key aspect in PTP biology is the elucidation of roles of PTPs, as well as substrates they act on, in different cellular events. Herein, a library of 144 putative peptide substrates against different PTPs was synthesized and immobilized onto a glass slide to generate the corresponding phosphopeptide microarray. Subsequent screening of the microarray against various PTPs provided a distinctive and comparative substrate fingerprint against each PTP. Several new substrates were identified, which might aid in the future design of potent and selective PTPs inhibitors. The signal-decrease microarray assay used in our studies provided a facile and efficient way for high-throughput determination of kinetic constants for peptide/PTP interactions en masse. Finally, our microarray results were independently verified by traditional microplate-based enzymatic assays.

Targeted Delivery of Bleomycin: A Comprehensive Anticancer Review

Despite being one of the most effective broad-spectrum chemotherapeutic agents in the treatment of cancers, the clinical applications of bleomycins (BLMs) have been limited due to their poor drug delivery abilities, and the side effect of causing lung fibrosis. With the increased therapeutics and the reduction of side effects, research and development of targeted drug delivery systems (TDDS) with BLMs have become essential for the expansive clinical usage of BLM-based therapeutics. This review summarizes the recent developments of various TDDS for BLMs, including techniques such as photochemical internalization, ultrasound, and micelle, liposome, and nanoparticle formation. The advantages and disadvantages for each delivery approach are outlined, along with the specific challenges associated with each delivery system.

The rational design of a peptide-based hydrogel responsive to H2S

The development of hydrogels that are responsive to external stimuli in a well-controlled manner is important for numerous biomedical applications. Herein we reported the first example of a hydrogel responsive to hydrogen sulphide (H2S). H2S is an important gasotransmitter whose deregulation has been associated with a number of pathological conditions. Our hydrogel design is based on the functionalization of an ultrashort hydrogelating peptide sequence with an azidobenzyl moiety, which was reported to react with H2S selectively under physiological conditions. The resulting peptide was able to produce hydrogels at a concentration as low as 0.1 wt%. It could then be fully degraded in the presence of excess H2S. We envision that the novel hydrogel developed in this study may provide useful tools for biomedical research.

A capillary electrophoresis method to explore the self-assembly of a novel polypeptide ligand with quantum dots

Polyhistidine peptides are effective ligands to coat quantum dots (QDs). It is known that both the number of histidine (His) residues repeats and their structural arrangements in a peptide ligand play important roles in the assembly of the peptide onto CdSe/ZnS QDs. However, due to steric hindrance, a peptide sequence with more than six His residue tandem repeats would hardly coordinate well with Zn2+ in the QD shell to further enhance the binding affinity. To solve this problem, a His‐containing peptide ligand, ATTO 590‐E2G (NH)6 (ATTO‐NH), was specifically designed and synthesized for assembly with QDs. With sequential injection of QDs and ATTO‐NH into the capillary electrophoresis with fluorescence detection, strong Förster resonance energy transfer phenomenon between the QDs and the ATTO 590 dye was observed, indicating efficient self‐assembly of the novel peptide onto the QDs to form ATTO‐NH capped QDs inside the capillary. The binding stability of the ligand onto the QD was then systematically investigated by titrating with imidazole, His, and a his‐tag containing competitive peptide. It is believed that this new in‐capillary assay significantly reduced the sample consumption and the analysis time. By functionalizing QDs with certain metal cation‐specific group fused peptide ligand, the QD‐based probes could be even extended to the online detection of metal cations for monitoring environment in the future.

Peptide microarrays for high-throughput studies of Ser/Thr phosphatases.

Protein phosphorylation and dephosphorylation play an important role in regulation of intracellular signal transduction pathways in the biological system. A key step in the biological characterization of phosphatases and their use as drug targets is the identification of their cellular partners and suitable substrates for potential inhibitor development. Herein we describe a microarray-based protocol to map the substrate specificity of protein Ser/Thr phosphatases. This protocol uses Pro-Q dye to sensitively and quantitatively detect the amount of dephosphorylation that occurs from many putative peptide substrates in parallel, and therefore could be used to generate the so-called peptide substrate fingerprints as well as detailed kinetic information of a target phosphatase. Excluding the synthesis of the peptide substrates, the whole protocol takes a total of 11 h to complete and in future can be readily extended to the study of other classes of phosphatases, i.e., protein tyrosine phosphatases.

Developing a fluorescence coupled capillary electrophoresis based method to probe interactions between QDs and colorectal cancer‐targeting peptides

As is well known, quantum dots (QDs) have become valuable probes for cancer imaging. In particular, QD‐labeled targeting peptides are capable of identifying cancer or tumors cells. A new colorectal cancer targeting peptide, cyclo(1, 9)‐CTPSPFSHC, has strong targeting ability and also shows great potential in the identification and treatment of colon cancer. Herein, we synthesized a dual functional polypeptide, cyclo(1, 9)‐CTPSPFSHCD2G2DP9G3H6 (H6‐TCP), to investigate its interaction with QDs inside the capillary. Fluorescence‐coupled CE was adopted and applied to characterize the self‐assembly of H6‐TCP onto QDs. It was indicated that the formation of the assembly was affected by H6‐TCP/QD molar ratio and sampling time. This novel in‐capillary assay greatly reduced the sample consumption and the detection time, which was beneficial for the environment. It is expected that this kind of detection method could find more applications to provide more useful information for cancer diagnosis and detection of harm and hazardous substances/organisms in the environment in the future.

Phosphopeptide Microarrays for Comparative Proteomic Profiling of Cellular Lysates

Protein phosphorylation is one of the most important and well-studied posttranslational modifications. Aberrant phosphorylation causes a wide spectrum of diseases, including cancers. As a result, many of the proteins involved in these pathways are seen as vital drug targets and biomarkers in treatment and diagnosis. The availability of broad-based platforms that identify changes across cellular states is critical in understanding unique disease characteristics and changes at the proteomic level. To highlight how microarrays can be applied in this regard, we describe here a comparative proteomic profiling method using two-color sample labeling and application on phosphopeptide microarrays, followed by a pull-down strategy and MS-based protein identification. This strategy has been applied to uncover candidate biomarkers in breast cancer and colon cancer cell lines. Apart from the synthesis of the phosphopeptide libraries and growth/isolation of cellular lysates, the protocol takes approximately 15 days to complete, once key steps have been optimized, and can be readily extended to other similarly complex biological specimens/samples.