Liquid Core Nanoparticle with High Deformability Enables Efficient Penetration across Biological Barriers.

Biological barriers significantly limit the delivery efficiency of drug delivery systems, resulting in undesired therapeutic effects. When designing a delivery system with optimized penetration behavior across the biological barriers, mechanical properties, such as deformability, are emerging as important parameters that need to be considered, although they are usually neglected in current research. Herein, a liquid core nanoparticle (LCN) composed of a polymer-encapsulated edible oil droplet is demonstrated. Owing to the unique structure in which the liquid oil core is encapsulated by a layer of highly hydrophilic and cross-linked polymer, the LCN exhibits high mechanical softness, making it deformable under external forces. With high deformability, LCNs can effectively penetrate through several important biological barriers including deep tumor tissue, blood-brain barriers, mucus layers, and bacterial biofilms. Moreover, the potential of the LCN as a drug delivery system is also demonstrated by the loading and release of several clinical drugs. With the capability of penetrating biological barriers and delivering drugs, LCN provides a potential platform for disease treatments, particularly for those suffering from inadequate drug penetration.

Transferrin receptor targeting segment T7 containing peptide gene delivery vectors for efficient transfection of brain tumor cells.

Successful gene therapy for brain tumors are often limited by two important factors, the existence of blood brain barrier (BBB) and inefficient transfection of brain tumor cells. In this study, we designed a series of peptide-based gene delivery vectors decorated with T7 segment for binding the transferrin (Tf) receptors which were highly expressed on brain tumor cells, and evaluated their ability of gene delivery. The physicochemical properties of peptide vectors or peptide/DNA complexes were studied as well. The in vitro transfection efficiency was investigated in normal and glioma cell lines. Among these complexes, PT-02/DNA complexes showed the highest transfection efficiency in glioma cells and low cytotoxicity in normal cell lines, and it could transport DNA across the BBB model in vitro. Furthermore, PT-02/DNA could deliver pIRES2-EGFP into the brain site of zebrafish in vivo. The designed peptide vectors offered a promising way for glioma gene therapy.

Arginine-Rich Polymers with Pore-Forming Capability Enable Efficient Intracellular Delivery via Direct Translocation Across Cell Membrane.

Efficient delivery of biomacromolecules or drugs across the cell membrane via endocytosis usually encounters inevitable entrapment in endosomes and subsequent degradation in lyso-endosomes. To address this issue, a series of arginine-rich cell penetrating polymers is designed and synthesized, which internalize into cells by inducing the formation of pores on the cell membrane, thereby crossing the cell membrane via direct translocation that fundamentally avoids endo/lysosomal entrapment. The structure-activity relationship studies show that PTn-R2-C6, which is a type of polymer that has two arginine residues and a flexible hexanoic acid linker in each side chain, exhibits excellent pore-formation ability on the cell membrane. Further investigations indicate that PTn-R2-C6 rapidly transports plasmid DNAs into cytosol through a similar endocytosis-independent pathway, thereby achieving significantly higher transfection efficiency and lower cytotoxicity than the gold-standard transfection reagent PEI 25K. These results suggest the great potential of PTn-R2-C6 as a safe and efficient gene transfection reagent for wide applications including disease treatments, vaccine development, and biomedical research purposes.

Spatial Distribution Control of Antimicrobial Peptides through a Novel Polymeric Carrier for Safe and Efficient Cancer Treatment.

Antimicrobial peptides (AMPs) hold great potential for use in tumor treatment. However, developing AMP-based antitumor therapies is challenging due to circulatory instability, hemolytic toxicity, low selectivity, and poor cell permeability of AMPs. In this study, a polymeric carrier for AMPs (denoted as PAMPm -co-PPBEn /PCA) is presented that effectively enhances their anticancer efficacy while minimizing their potential side effects. By integrating multiple responsive structures at the molecular level, the carrier finely controls the spatial distribution of AMPs in different biological microenvironments, thereby effectively modulating their membranolytic ability. Upon employing KLA as the model AMP, the polymeric carrier's hemolytic toxicity during blood circulation is suppressed, its cellular internalization when reaching tumor tissues facilitated, and its membranolytic toxicity toward the mitochondria upon entering cancer cells restored and further enhanced. Animal studies indicate that this approach significantly improves the antitumor efficacy of KLA and reduces its toxicity. Considering that the loading method for most AMPs is identical to that of KLA, the polymeric carrier reported in this study may provide a feasible approach for the development of AMP-based cancer treatments.

A near-infrared light-excitable immunomodulating nano-photosensitizer for effective photoimmunotherapy.

Photodynamic therapy has great potential for tumor ablation and the activation of antitumor immune responses. However, its overall therapeutic efficiency is often limited by the immunosuppressive tumor microenvironment. We developed a near-infrared light-excitable immunomodulating nano-photosensitizer (NeINP) that can improve reactive oxygen species production and regulate the immunosuppressive TME to improve photoimmunotherapy. The NeINP is composed of a photosensitive core and a pH-responsive polymer shell, which allows for NeINP loading and delivery of small-molecular immunomodulators to tumor sites for regulation of the immunosuppressive TME and effective photoimmunotherapy. Through the co-delivery of celecoxib and the NIR-triggered photodynamic core to tumors, the NeINP was shown to regulate the immunosuppressive TME and enhance antitumor immunity, leading to the elimination of residual tumor and reduction of metastasis and recurrence. The NeINP can be optimized to co-deliver other immunomodulators, and thus has potential as a universal platform for efficient, precise photoimmunotherapy.

Calixarene-Embedded Nanoparticles for Interference-Free Gene-Drug Combination Cancer Therapy.

Combination therapy based on molecular drugs and therapeutic genes provides an effective strategy for malignant tumor treatment. However, effective gene and drug combinations for cancer treatment are limited by the widespread antagonism between therapeutic genes and molecular drugs. Herein, a calixarene-embedded nanoparticle (CENP) is developed to co-deliver molecular drugs and therapeutic genes without compromising their biological functions, thereby achieving interference-free gene-drug combination cancer therapy. CENP is composed of a cationic polyplex core and an acid-responsive polymer shell, allowing CENP loading and delivering therapeutic genes with improved circulation stability and enhanced tumor accumulation. Moreover, the introduction of carboxylated azocalix[4]arene, which is a hypoxia-responsive calixarene derivatives, in the polyplex core endows CENP with the capability to load molecular drugs through the host-guest complexation as well as inhibit the interference between the drugs and genes by encapsulating the drugs into its cavity. By loading doxorubicin and a plasmid DNA-based CRISPR interference system that targets miR-21, CENP exhibits the significantly enhanced anti-tumor effects in mice. Considering the wide variety of calixarene derivatives, CENP can be adapted to deliver almost any combination of drugs and genes, providing the potential as a universal platform for the development of interference-free gene-drug combination cancer therapy.

Peptide Gene Delivery Vectors for Specific Transfection of Glioma Cells.

Gene therapy offers an alternative approach to malignant glioma; however, glioma cells are difficult to transfect. Peptides, as nonviral vectors, can achieve efficient gene transfection in glioma cells due to their good biocompatibility and easy functionalization. In this article, we reported a series of peptide vectors, which were composed of amphiphilic α-helical segments, cationic cell-penetrating segments, and cysteine and glycine residues. The physicochemical properties of peptide vectors or peptide/pGL3 complexes, including conformation, DNA-loading capacity, size, zeta potential, and morphology, were characterized. Their gene delivery abilities were evaluated in U373, U87, and C6 glioma cell lines and a normal cell line 293 T. Compared with Lipo 2000 and other peptide vectors, the efficiency of P-03 (CLLHHLLHHLLHHGGRKKRRQRRR) to transfect glioma cells was higher. While in 293 T cells, the transfection efficiency of P-03 was much lower than that of Lipo 2000 and another positive control P-07. Furthermore, P-03 could facilitate the pGL3 plasmids crossing a blood-brain barrier model in vitro and achieved the expression of EGFP gene in the brain sites of zebrafish.

The rational design of cell-penetrating peptides for application in delivery systems.

Cell penetrating peptides (CPPs) play a crucial role in the transportation of bioactive molecules. Although CPPs have been used widely in various delivery systems, further applications of CPPs are hampered by several drawbacks, such as high toxicity, low delivery efficiency, proteolytic instability and poor specificity. To design CPPs with great cell-penetrating ability, physicochemical properties and safety, researchers have tried to develop new methods to overcome the defects of CPPs. Briefly, (1) the side chain of arginine containing the guanidinium group is essential for the facilitation of cellular uptake; (2) the hydrophobic counterion complex around the guanidinium-rich backbone can "coat" the highly cationic structure with lipophilic moieties and act as an activator; (3) the conformation-constrained strategy was pursued to shield the peptide, thereby impeding access of the proteolytic enzyme; (4) targeting strategies can increase cell-type specificity of CPPs. In this review, the above four aspects were discussed in detail.

Peptide-based gene delivery vectors.

Gene therapy as a strategy for disease treatment requires safe and efficient gene delivery systems that encapsulate nucleic acids and deliver them to effective sites in the cell. Due to the insecurity of viral vectors, non-viral vectors have gained great attention recently. Additional advantages of non-viral vectors include their safety, flexibility in packaging nucleic acids, and ease of production. To construct an ideal gene carrier, peptides can be incorporated into non-viral gene delivery systems as functional motifs to overcome the current barriers in gene delivery. In this review, we summarize recent developments in peptide-based gene delivery vector research.

Enhanced gene transfection efficiency by use of peptide vectors containing laminin receptor-targeting sequence YIGSR.

This study presents the design and evaluation of a series of multifunctional peptides and their gene delivery abilities. The peptide sequences contained a cell-penetrating segment, six continuous histidine residues, a stearyl moiety and a laminin receptor-targeting segment. The YIGSR segment promoted cellular uptake through the interaction with laminin receptors on the surface of cells, which resulted in a great improvement in gene transfection efficiency. The conformation, particle size and zeta potential of peptide/DNA complexes were characterized via circular dichroism and dynamic light scattering. Their gene transfection efficiency was investigated by fluorescence-activated cell sorting and confocal microscopy. The transfection efficiency of the designed peptide vectors was higher than that of Lipo 2000. The peptide TAT-H6-K(C18)-YIGSR displayed transfection efficiencies at N/P ratios of 6, which was 3.5 and 7 times higher than that of Lipo 2000 in B16F10 and 293T cells, respectively. All peptides exhibited lower cytotoxicity than Lipo 2000 in B16F10 and 293T cells. In summary, the designed YIGSR-containing multifunctional peptide gene vectors promoted cellular uptake and gene transfection. Their in vivo transfection ability was investigated in zebrafish, and the transfection efficiency was determined by confocal microscopy and bioluminescence imaging. The peptide vectors, owing to their relatively short sequences and ease of functionalization, offer a promising approach for gene delivery because of their low cytotoxicity and high transfection efficiency.

The structure and configuration changes of multifunctional peptide vectors enhance gene delivery efficiency.

We designed a series of peptide vectors that contain functional fragments with the goal of enhancing cellular internalization and gene transfection efficiency. The functional fragments included a cell-penetrating peptide (R9), a cationic amphiphilic α-helical peptide [(LLKK)3-H6 or (LLHH)3], a stearyl moiety, and cysteine residues. Vectors were also synthesized with D-type amino acids to improve their proteolytic stability. The conformations, particle sizes, and zeta potentials for complexes of these peptides with pGL3 plasmid DNA were characterized by circular dichroism and dynamic light scattering. In addition, cellular uptake of the peptide/DNA complexes and gene transfection efficiency were investigated with fluorescence-activated cell sorting and confocal laser-scanning microscopy. Greater transfection efficiency was achieved with the vectors containing the R9 segment, and the efficiency was greater than Lipo2000. In addition, the D-type C18-c(llkk)3ch6-r9 had about 7 times and 5.5 times the transfection efficiency of Lipo2000 in 293T cells and NIH-3T3 cells at the N/P ratio of 6, respectively. Overall, the multifunctional peptide gene vectors containing the R9 segment exhibited enhanced cellular internalization, a high gene transfection efficiency, and low cytotoxicity.

A pH responsive complexation-based drug delivery system for oxaliplatin.

A responsive drug delivery system (DDS) for oxaliplatin (OX) has been designed with a view to overcoming several drawbacks associated with this anticancer agent, including fast degradation/deactivation in the blood stream, lack of tumor selectivity, and low bioavailability. The present approach is based on the direct host-guest encapsulation of OX by a pH-responsive receptor, carboxylatopillar[6]arene (CP6A). The binding affinities of CP6A for OX were found to be pH-sensitive at biologically relevant pH. For example, the association constant (Ka) at pH 7.4 [Ka = (1.02 ± 0.05) × 104 M-1] is 24 times larger than that at pH 5.4 [Ka = (4.21 ± 0.06) × 102 M-1]. Encapsulation of OX within the CP6A cavity did not affect its in vitro cytotoxicity as inferred from comparison studies carried out in several cancer cells (e.g., the HepG-2, MCF-7, and A549 cell lines). On the other hand, complexation by CP6A serves to increase the inherent stability of OX in plasma by 2.8-fold over a 24 h incubation period. The formation of a CP6A⊃OX host-guest complex served to enhance in a statistically significant way the ability of OX to inhibit the regrowth of sarcoma 180 (S180) tumors in Kunming (KM) mice xenografts. The improved anticancer activity observed in vivo for CP6A⊃OX is attributed to the combined effects of enhanced stability of the host-guest complex and the pH-responsive release of OX. Specifically, it is proposed that OX is protected as the result of complex formation and then released effectively in the acidic tumor environment.

Histidine-enriched multifunctional peptide vectors with enhanced cellular uptake and endosomal escape for gene delivery.

Peptide vectors offer a promising gene delivery approach because of their biocompatibility and ease of functionalization. This article describes the design and evaluation of a series of multifunctional peptides and their gene delivery abilities. The peptides were composed of a cell-penetrating segment, stearyl moiety, cationic amphiphilic α-helical segment, and cysteine and histidine residues. The proton sponge effect of histidine residues at low pH and the α-helical conformation should improve endosomal escape. Inclusion of d-type amino acids should improve proteolytic stability. The conformation, particle size and zeta potential of peptide/DNA complexes were characterized by circular dichroism and dynamic light scattering. Gene transfection efficiency was investigated by fluorescence-activated cell sorting and confocal microscopy. Transfection efficiencies of the designed peptide vectors were better than those of C18-C(LLKK)3C-TAT and Lipo2000. d-Type peptide C18-c(llhh)3c-tat showed three times higher transfection efficiency at N/P ratios of 6 and 8 than Lipo2000 in NIH-3T3 and 293T cells. All peptides showed lower cytotoxicity than Lipo2000 in NIH-3T3 and 293T cells. In the presence of trypsin or serum in vitro, d-type peptides showed better stability than l-type peptides. Overall, the designed histidine-enriched multifunctional peptide gene vectors promoted cellular uptake, endosomal escape and gene transfection.