Cationic Pillar[6]arene Induces Cell Apoptosis by Inhibiting Protein Tyrosine Phosphorylation Via Host-Guest Recognition.

We reported for the first time that cationic pillar[6]arene (cPA6) could tightly bind to peptide polymer (MW~20-50 kDa), an artificial substrate for tyrosine (Tyr) phosphorylation, and efficiently inhibit Tyr protein phosphorylation through host-guest recognition. We synthesized a nanocomposite of black phosphorus nanosheets loaded with cPA6 (BPNS@cPA6) to explore the effect of cPA6 on cells. BPNS@cPA6 was able to enter HepG2 cells, induced apoptosis, and inhibited cell proliferation by reducing the level of Tyr phosphorylation. Furthermore, BPNS@cPA6 showed a stronger ability of inhibiting cell proliferation in tumor cells than in normal cells. Our results revealed the supramolecular modulation of enzymatic Tyr phosphorylation by the host-guest recognition of cPA6.

Therapeutic delivery of microRNA-143 by cationic lipoplexes for non-small cell lung cancer treatment in vivo.

PURPOSE: Non-small cell lung cancer (NSCLC) remains the leading cause of cancer-related deaths worldwide and new improvements are urgently needed. Several miRNA-targeted therapeutics have reached clinical development. MicroRNA-143 (miR-143) was found to significantly suppress the migration and invasion of NSCLC. It might be of great potential for NSCLC treatment. However, the therapeutic effect of miR-143 against NSCLC in vivo has not been explored until now. METHODS: The cationic liposome/pVAX-miR-143 complex (CL-pVAX-miR-143) was prepared and its biodistribution was assessed. The tumor suppression effects of CL-pVAX-miR-143 were evaluated in early-stage and advanced experimental lung cancer metastasis mice models by systemic delivery, respectively, and also in subcutaneous tumor models by intratumoral injection. The toxicity of CL-pVAX-miR-143 was assessed by H&E analysis and biochemical measurements. The preliminary mechanism of CL-pVAX-miR-143 on tumor suppression was explored by immunochemistry and western blotting. RESULTS: The assays on the stability and safety of CL-pVAX-miR-143 showed that it mainly accumulated in the lung after systemic administration. The intratumoral delivery of CL-pVAX-miR-143 effectively inhibited A549 subcutaneous tumor growth. Notably, systemic delivery of CL-pVAX-miR-143 significantly inhibited tumor metastasis and prolonged survival dose dependently in early-stage experimental lung cancer metastasis models. More importantly, same results were shown in advanced mice models with metastasis. CL-pVAX-miR-143 treatment did not induce obvious acute toxicity. The preliminary mechanism on inhibiting tumor metastasis might be induced by targeting CD44v3. CONCLUSIONS: Our results suggested that CL-pVAX-miR-143 might be a promising strategy for clinical treatment of non-small cell lung cancer, especially for advanced NSCLC with metastasis.

Nucleic acid delivery to differentiated retinal pigment epithelial cells using cell-penetrating peptide as a carrier.

Nucleic acid delivery to the eye is a promising treatment strategy for many retinal disorders. In this manuscript, retinal gene delivery with non-coated and chondroitin sulphate (CS) coated amphipathic and cationic peptides was tested. The transfection and gene knockdown efficiencies were evaluated in different retinal pigment epithelial (RPE) cell models including both dividing and differentiated cells. In addition, the mobility of peptide-based gene delivery systems was examined in porcine vitreous by particle tracking analysis. The results indicate that amphipathic and cationic peptides are safe in vitro and are capable of high transgene expression and gene knockdown in dividing cells. We further demonstrate that incorporation of CS improves the efficiency of gene delivery of peptide-based systems. Most importantly, the transgene expression mediated by both non-coated and CS coated peptides was high in differentiated as well as in human primary RPE cells which are typically difficult to transfect. Coating of peptide-based gene delivery systems with CS improved diffusion in the vitreous and enhanced the stability of the polyplexes. The results indicate that a peptide-based system can be fine-tuned as a promising approach for retinal gene delivery.

Hybrid Alginate@TiO2 Porous Microcapsules as a Reservoir of Animal Cells for Cell Therapy.

The number of patients suffering from diseases linked with hormone deficiency (e.g., type 1 diabetes mellitus) has significantly increased in recent years. As organ transplantation presents its limits, the design of novel robust devices for cell encapsulation is of great interest. The current study reports the design of a novel hybrid alginate microcapsule reinforced by titania via a biocompatible synthesis from an aqueous stable titania precursor (TiBALDH) and a cationic polyamine (PDDAC) under mild conditions. The biocompatibility of this one-pot synthesis was confirmed by evaluation of the cytotoxicity of the precursor, additive, product, and by-product. The morphology, structure, and properties of the obtained hybrid microcapsule were characterized in detail. The microcapsule displayed mesoporous, which was a key parameter to allow the diffusion of nutrients and metabolites and to avoid the entry of immune defenders. The hybrid microcapsule also showed enhanced mechanical stability compared to the pure alginate microcapsule, making it an ideal candidate as a cell reservoir. HepG2 model cells encapsulated in the hybrid microcapsules remained intact for 43 days as highlighted by fluorescent viability probes, their oxygen consumption, and their albumin secretion. The study provides a significant progress in the conception of the robust and biocompatible reservoirs of animal cells for cell therapy.

Cationic lipids as one-component vaccine adjuvants: A promising alternative to alum.

Effective vaccine formulations consist of several components: an antigen carrier, the antigen, a stimulator of cellular immunity such as a Toll-like Receptors (TLRs) ligand, and a stimulator of humoral response such as an inflammasome activator. Here, we investigated the immunostimulatory and adjuvant properties of lipopolyamines, cationic lipids used as gene carriers. We identified new lipopolyamines able to activate both TLR2 and TLR4 and showed that lipopolyamines interact with TLRs via a mechanism different from the one used by bacterial ligands, activating a strong type-I IFN response, pro-inflammatory cytokines and IL-1ß secretion. The TLR and inflammasome stimulations, together with the antigen carrier properties of lipopolyamines, resulted in both humoral and cellular immunity in mice vaccinated against OVA and make lipopolyamines promising one-component vaccine adjuvants.

Anticancer activity of the intraperitoneal-delivered DFP-10825, the cationic liposome-conjugated RNAi molecule targeting thymidylate synthase, on peritoneal disseminated ovarian cancer xenograft model.

INTRODUCTION: Peritoneal disseminated ovarian cancer is one of the most difficult cancers to treat with conventional anti-cancer drugs and the treatment options are very limited, although an intraperitoneal (ip) paclitaxel has shown some clinical benefit. Therefore, treatment of peritoneal disseminated ovarian cancer is a highly unmet medical need and it is urgent to develop a new ip delivered drug regulating the fast DNA synthesis. METHODS: We developed a unique RNAi molecule consisting of shRNA against the thymidylate synthase (TS) and a cationic liposome (DFP-10825) and tested its antitumor activity and PK profile in peritoneally disseminated human ovarian cancer ascites models by the luciferase gene-transfected SCID mice. DFP-10825 alone, paclitaxel alone or combination with DFP-10825 and paclitaxel were administered in an ip route to the tumor-bearing mice. The TS expression level was measured by conventional RT-PCR. The anti-tumor activity and host survival benefit by DFP-10825 treatment on tumor-bearing mice were observed as resulting from the specific TS mRNA knock-down in tumors. RESULTS: DFP-10825 alone significantly suppressed the growth of SKOV3-luc tumore ascites cells and further extended the survival time of these tumor-bearing mice. Combination with the ip paclitaxel augmented the antitumor efficacy of DFP-10825 and significantly prolonged the survival time in the tumor-bearing mice. Short-hairpin RNA for TS (TS shRNA) levels derived from DFP-10825 in the ascetic fluid were maintained at a nM range across 24 hours but not detected in the plasma, suggesting that TS shRNA is relatively stable in the peritoneal cavity, to be able to exert its anti-tumor activity, but not in blood stream, indicating little or no systemic effect. CONCLUSION: Collectively, the ip delivery of DFP-10825, TS shRNA conjugated with cationic liposome, shows a favorable antitumor activity without systemic adverse events via the stable localization of TS shRNA for a sufficient time and concentration in the peritoneal cavity of the peritoneally disseminated human ovarian cancer-bearing mice.

Ultrasound-mediated nanobubble destruction (UMND) facilitates the delivery of A10-3.2 aptamer targeted and siRNA-loaded cationic nanobubbles for therapy of prostate cancer.

The Forkhead box M1 (FoxM1) transcription factor is an important anti-tumor target. A novel targeted ultrasound (US)-sensitive nanobubble that is likely to make use of the physical energy of US exposure for the improvement of delivery efficacy to target tumors and specifically silence FoxM1 expression appears as among the most potential nanocarriers in respect of drug delivery. In this study, we synthesized a promising anti-tumor targeted FoxM1 siRNA-loaded cationic nanobubbles (CNBs) conjugated with an A10-3.2 aptamer (siFoxM1-Apt-CNBs), which demonstrate high specificity when binding to prostate-specific membrane antigen (PSMA) positive LNCaP cells. Uniform nanoscaled siFoxM1-Apt-CNBs were developed using a thin-film hydration sonication, carbodiimide chemistry approaches, and electrostatic adsorption methods. Fluorescence imaging as well as flow cytometry evidenced the fact that the siFoxM1-Apt-CNBs were productively developed and that they specifically bound to PSMA-positive LNCaP cells. siFoxM1-Apt-CNBs combined with ultrasound-mediated nanobubble destruction (UMND) significantly improved transfection efficiency, cell apoptosis, and cell cycle arrest in vitro while reducing FoxM1 expression. In vivo xenografts tumors in nude-mouse model results showed that siFoxM1-Apt-CNBs combined with UMND led to significant inhibition of tumor growth and prolonged the survival of the mice, with low toxicity, an obvious reduction in FoxM1 expression, and a higher apoptosis index. Our study suggests that siFoxM1-Apt-CNBs combined with UMND might be a promising targeted gene delivery strategy for therapy of prostate cancer.

Dimeric camptothecin-loaded RGD-modified targeted cationic polypeptide-based micelles with high drug loading capacity and redox-responsive drug release capability.

Camptothecin (CPT) is a broad spectrum anticancer drug, but its application is limited due to the poor water solubility, lactone ring instability, and low drug loading potential. In this study, biocompatible cationic polypeptide-based micelles were developed to deliver dimeric CPT (DCPT) with the aim of overcoming the above-mentioned obstacles and achieving favorable therapeutic effects. Cationic polypeptide poly-lysine-block-poly-leucine (PLys-b-PLeu) was fabricated via the ring-opening polymerization of N-ε-carbobenzoxy-l-lysine (ε-Lys(Z)) and l-leucine (Leu) and further grafted with polyethylene glycol (PEG) and an arginine-glycine-aspartic acid (RGD) peptide. DCPT was synthesized by reacting CPT and 2-hydroxyethyl disulfide, and micelles were prepared using a dialysis method. The obtained DCPT-loaded RGD-PEG-g-poly-l-lysine-b-poly-l-leucine (DRPPP) micelles showed a high encapsulation efficiency of 89.7% and a high drug loading capacity of 46.1%. In addition, the DRPPP micelles remained stable under physiological conditions (PBS at a pH of 7.4) but showed rapid release when triggered by a reductive environment (PBS at a pH of 7.4 with 10 mM dithiothreitol). Compared to micelles without RGD decoration, the DRPPP micelles exhibited an increased cellular uptake through RGD targeting and were internalized into cells via caveolae-mediated endocytosis and macropinocytosis. Furthermore, the DRPPP micelles exerted an enhanced cytotoxicity against MDA-MB-231 cells compared to MCF-7 cells, which expressed less αvß3 receptors. Besides, the DRPPP micelles induced cell apoptosis and caused a decrease of mitochondrial membrane potential. These results indicate that dimeric camptothecin-loaded cationic polypeptide-based micelle is a promising strategy for cancer therapy.

pH-sensitive polymer-modified liposome-based immunity-inducing system: Effects of inclusion of cationic lipid and CpG-DNA.

Efficient vaccine carriers for cancer immunotherapy require two functions: antigen delivery to dendritic cells (DCs) and the activation of DCs, a so-called adjuvant effect. We previously reported antigen delivery system using liposomes modified with pH-sensitive polymers, such as 3-methylglutarylated hyperbranched poly(glycidol) (MGlu-HPG), for the induction of antigen-specific immune responses. We reported that inclusion of cationic lipids to MGlu-HPG-modified liposomes activates DCs and enhances antitumor effects. In this study, CpG-DNA, a ligand to Toll-like receptor 9 (TLR9) expressing in endosomes of DCs, was introduced to MGlu-HPG-modified liposomes containing cationic lipids using two complexation methods (Pre-mix and Post-mix) for additional activation of antigen-specific immunity. For Pre-mix, thin membrane of lipids and polymers were dispersed by a mixture of antigen/CpG-DNA. For Post-mix, CpG-DNA was added to pre-formed liposomes. Both Pre-mix and Post-mix delivered CpG-DNA to DC endosomes, where TLR9 is expressing, more efficiently than free CpG-DNA solution did. These liposomes promoted cytokine production from DCs and the expression of co-stimulatory molecules in vitro and induced antigen-specific immune responses in vivo. Both Pre-mix and Post-mix exhibited strong antitumor effects compared with conventional pH-sensitive polymer-modified liposomes. Results show that inclusion of multiple adjuvant molecules into pH-sensitive polymer-modified liposomes and suitable CpG-DNA complexation methods are important to design potent vaccine carriers.

PEGylated Cationic Vectors Containing a Protease-Sensitive Peptide as a miRNA Delivery System for Treating Breast Cancer.

Several targeted drug delivery systems have recently been developed to increase the bioavailability of a drug at its site of action, allowing simultaneous reduction of the total necessary drug dose as well as side effects. Here, we designed a cationic gene vector containing matrix metalloproteinase-2 (MMP2)-cleavable substrate peptides that specifically target tumor sites where MMP2 levels are high. The targeted delivery system is fabricated by linking enzyme-cleavable polyethylene glycol (PEG) derivatives to cationic ß-cyclodextrin-polyethylenimine conjugates, which reduce the toxicity of polyethylenimine and condense the therapeutic cargo. In the present study, tumor suppressor microRNA miR-34a, which suppresses onset and progression of many types of cancers, was investigated for its therapeutic potential for treating breast cancer. The PEG coating markedly reduces nonspecific interaction between cationic particles and serum proteins, permitting accumulation at the target site; subsequent peptide cleavage by MMP2 facilitates miR-34a delivery into tumor cells. The nanopreparation shows excellent stability, and its internalization, tumor targeting, and antitumor efficacy in vitro and in vivo are better than those of a nanopreparation containing MMP2-uncleavable peptide.

Multifunctional polycationic photosensitizer conjugates with rich hydroxyl groups for versatile water-soluble photodynamic therapy nanoplatforms.

Photodynamic therapy (PDT) has already shown immense potential in antitumor fields due to its low systemic toxicity and negligible drug resistance. However, the clinical application of current photosensitizers is still restricted by the low singlet oxygen yield or insolubility. Herein, series of star-like hydroxyl-rich polycations (Pc-PGEA/Pc) with flanking phthalocyanine (Pc) were proposed for effective water-soluble photosensitizers. The designed Pc-PGEA/Pc polymers consist of one Pc core and four ethanolamine and Pc-difunctionalized poly(glycidyl methacrylate) arms. The strong π-π stacking and hydrophobicity of introduced Pc units drive the amphipathic Pc-PGEA/Pc polymers to self-assemble into well-defined cationic nanoparticles. Such Pc-PGEA/Pc nanoparticles present impressive photodynamic therapy effects under moderate irradiation and remarkable photoacoustic imaging (PAI) ability. These kinds of nanoparticles also exhibit good performance as gene vectors. The PAI ability given by the proper wavelength absorbance of Pc units provides one promising method for PAI-guided combined antitumor therapy. The present work would contribute valuable information for the development of new strategies of visible antitumor therapy.

Cell-Targeting Cationic Gene Delivery System Based on a Modular Design Rationale.

En route to target cells, a gene carrier faces multiple extra- and intracellular hurdles that would affect delivery efficacy. Although diverse strategies have been proposed to functionalize gene carriers for individually overcoming these barriers, it is challenging to generate a single multifunctional gene carrier capable of surmounting all these barriers. Aiming at this challenge, we have developed a supramolecular modular approach to fabricate a multifunctional cationic gene delivery system. It consists of two prefunctionalized modules: (1) a host module: a polymer (PCD-SS-PDMAEMA) composed of poly(ß-cyclodextrin) backbone and disulfide-linked PDMAEMA arms, expectedly acting to compact DNA and release DNA upon cleavage of disulfide linkers in reductive microenvironment; and (2) a guest module: adamantyl and folate terminated PEG (Ad-PEG-FA), expectedly functioning to reduce nonspecific interactions, improve biocompatibility, and provide folate-mediated cellular targeting specificity. Through the host-guest interaction between ß-cyclodextrin units of the "host" module and adamantyl groups of the "guest" module, the PCD-SS-PDMAEMA-1 (host) and Ad-PEG-FA (guest) self-assemble forming a supramolecular pseudocopolymer (PCD-SS-PDMAEMA-1/PEG-FA). Our comprehensive analyses demonstrate that the functions preassigned to the two building modules are well realized. The gene carrier effectively compacts DNA into stable nanosized polyplexes resistant to enzymatic digestion, triggers DNA release in reducing environment, possesses significantly improved hemocompatibility, and specifically targets folate-receptor positive cells. Most importantly, endowed with these predesigned functions, the PCD-SS-PDMAEMA-1/PEG-FA supramolecular gene carrier exhibits excellent transfection efficacy for both pDNA and siRNA. Thus, this work represents a proof-of-concept example showing the efficiency and convenience of an adaptable, modular approach for conferring multiple functions to a single supramolecular gene carrier toward effective in vivo delivery of therapeutic nucleic acids.

Tissue Permeability Effects Associated with the Use of Mucoadhesive Cationic Nanoformulations of Docetaxel in the Bladder.

PURPOSE: Recently, efficacy studies in mice have shown that amine-terminated cationic (CNP) nanoparticulate carriers of DTX offer an improved formulation of the drug for intravesical delivery. It is hypothesized that this improved efficacy may arise from a carrier mediated bladder exfoliation process that removes the urothelial barrier allowing for increased drug uptake into bladder tissue. The objective of this study was to investigate exfoliation processes in fresh pig's bladders (ex vivo) exposed to three cationic polyglycerols with increasing degrees of amination (denoted 350, 580 and 780). The study also compared the tissue depth profile of DTX uptake into these tissues using these different carriers. MATERIALS AND METHODS: Aminated polyglycerols were synthesized and characterized in the laboratory with low (CNP-360), medium (CNP-580) and high (CNP-780) levels of amine content. CNP-based DTX solutions and commercial DTX solutions in polysorbate 80 (Taxotere®) were doped with (3)H-radiolabeled DTX and prepared by solvent evaporation from acetonitrile, followed by drying and reconstitution in pH 6.4 buffer. Sections of fresh pig's bladder tissue were clamped into Franz diffusion cells and the urothelial side was exposed to the DTX solutions for 2 h. Tissue sections were then frozen for sectioning by cryotome sectioning and subsequently processed for drug analysis by liquid scintillation counting. Alternatively tissue sections were fixed in 2% glutaraldehyde and 2% paraformaldehyde in 0.1 M sodium cacodylate buffer for the purposes of scanning electron microscopy (SEM). RESULTS: Exposure of the urothelial surface to the amine-terminated polyglycerol solutions resulted in the exfoliation of bladder tissues in a time- and concentration-dependent manner. Exfoliation was significantly more pronounced when using CNPs with a medium or high levels of amination whereas only minor levels of exfoliation were seen with low levels. Following incubation of tissues in Tween-based commercial formulations (Taxotere) of DTX (0.5 mg/mL) the drug was detectable at low levels (10-40 µg/g tissue) in all depths of tissue. Similar drug uptake was observed using the CNP-360 formulation. However drug uptake levels were increased to 60-100 µg/g tissue when samples were incubated with either the CNP-580 or CNP-780 formulations. CONCLUSION: The use of cationic polyglycerols with higher levels of amine termination allows for an enhanced uptake of DTX into bladder tissues as compared to commercial (Taxotere) formulations. These increased drug levels probably arise from exfoliation processes resulting in a temporary elimination of the urothelial permeability barrier and increased drug penetration into the tissue.

Synthesis and evaluation of novel lipopeptide as a vehicle for efficient gene delivery and gene silencing.

Nucleic acid-based therapeutics have recently emerged as a new class of next generation agents for treatment and prevention of viral infection, cancer, and genetic disorders, but their wide use is limited by their relatively weak delivery into target cells. Usage of synthetic cationic amphiphiles with peptide hydrophilic domain as agents for non-viral gene delivery is an attractive approach. We developed the schemes for the synthesis of aliphatic peptides with different length of the hydrocarbon chains in hydrophobic domains and different amino acids in polar head. For the obtained derivatives we determined transfection efficiency, critical vesicle concentration, particle size, ζ-potential and aggregates stability. We have found that the transfection efficiency is increased if the ornithine is a part of polar head in an amphiphile. The most promising amphiphile for liposomal formation OrnOrnGlu(C16H33)2 was examined more carefully. It has been shown that the lipopeptide possesses low toxicity (in vitro and in vivo) and high transfection efficiency with pDNA and siRNA in different cell lines. In addition, the production of liposomes based on this lipopeptide is simple, quick and cheap. Thus OrnOrnGlu(C16H33)2 is a promising vehicle for gene delivery and gene silencing.

Pharmacokinetics, Tissue Distribution and Therapeutic Effect of Cationic Thermosensitive Liposomal Doxorubicin Upon Mild Hyperthermia.

PURPOSE: To evaluate pharmacokinetic profile, biodistribution and therapeutic effect of cationic thermosensitive liposomes (CTSL) encapsulating doxorubicin (Dox) upon mild hyperthermia (HT). METHODS: Non-targeted thermosensitive liposomes (TSL) and CTSL were developed, loaded with Dox and characterized. Blood kinetics and biodistribution of Dox-TSL and Dox-CTSL were followed in B16BL6 tumor bearing mice upon normothermia (NT) or initial hyperthermia conditions. Efficacy study in B16BL6 tumor bearing mice was followed with Dox-TSL or Dox-CTSL upon NT or HT. Efficacy study in LLC tumor bearing mice was performed upon two HT conditions. Intravital microscopy was performed on B16BL6 tumors implanted in dorsal-skin fold window-bearing mice. RESULTS: Targeting did not cause faster blood clearance of CTSL compared to TSL. Highest uptake of liposomes was observed in spleen, kidneys and liver. Applying HT prior to CTSL administration increased drug delivery to the tumor and CTSL delivered ~1.7 fold higher Dox concentration compared to TSL. Efficacy in B16BL6 murine melanoma showed that HT had a significant effect on CTSL in tumor suppression and prolonged survival. Efficacy in LLC Lewis lung carcinoma tumor model demonstrates that two HT treatments hold promises for a successful treatment option. CONCLUSION: CTSL have potency to increase drug efficacy in tumors due to their targeted and drug release functions.

Stapled endosome disrupting alginate particles for cytosolic delivery of cations.

Divalent cations, the most prevalent minerals in the body, are responsible for a wide variety of cellular functions including signaling, proliferation, differentiation and cell death, and therefore their transmembrane transportation is tightly regulated. Despite the importance of divalent cations in cell activity, there are currently no intracellular delivery methods for divalent cations or modulation of intracellular levels of minerals. Here, we describe endosome disrupting alginate nanoparticles termed Alginoketals, which can deliver divalent cations to the cytosol of the cells. Alginoketals are generated by crosslinking alginic acid with endosome disrupting ketals, and using divalent cations as the stapling or binding agent. We show that Alginoketals were able to deliver copper (II) in the cytosol of the cancer cells thereby disrupting copper homeostasis and inducing cell death via accumulation of hydrogen peroxide. Alginoketal-copper (II)-based particles act as superoxide dismutase mimics and are the first class of divalent cation delivery vehicles, with potential application in cancer therapy, regenerative medicine and drug delivery.

The impact of cationic solid lipid nanoparticles on human neutrophil activation and formation of neutrophil extracellular traps (NETs).

Cationic solid lipid nanoparticles (cSLNs) are extensively employed as the nanocarriers for drug/gene targeting to tumors and the brain. Investigation into the possible immune response of cSLNs is still lacking. The aim of this study was to evaluate the impact of cSLNs upon the activation of human polymorphonuclear neutrophil cells (PMNs). The cytotoxicity, pro-inflammatory mediators, Ca(2+) mobilization, mitogen-activated protein kinases (MAPKs), and neutrophil extracellular traps (NETs) as the indicators of PMN stimulation were examined in this work. The cSLNs presented a diameter of 195 nm with a zeta potential of 44 mV. The cSLNs could interact with the cell membrane to produce a direct membrane lysis and the subsequent cytotoxicity according to lactate dehydrogenase (LDH) elevation. The interaction of cSLNs with the membrane also triggered a Ca(2+) influx, followed by the induction of oxidative stress and degranulation. The cationic nanoparticles elevated the levels of superoxide anion and elastase by 24- and 9-fold, respectively. The PMN activation by cSLNs promoted the phosphorylation of p38 and Jun-N-terminal kinases (JNK) but not extracellular signal-regulated kinases (ERK). The imaging of scanning electron microscopy (SEM) and immunofluorescence demonstrated the production of NETs by cSLNs. This phenomenon was not significant for the neutral SLNs (nSLNs), although histones in NETs also increased after treatment of nSLNs. Our results suggest an important role of cSLNs in governing the activation of human neutrophils.

siRNA delivery to lung-metastasized tumor by systemic injection with cationic liposomes.

CONTEXT: Cationic liposomes can efficiently deliver siRNA to the lung by intravenous injection of cationic liposome/siRNA complexes (lipoplexes). OBJECTIVE: The aim of this study was to examine a formulation of cationic liposomes for siRNA delivery to lung metastasis of breast tumor. MATERIALS AND METHODS: For the preparation of cationic liposomes, 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) or dimethyldioctadecylammonium bromide (DDAB) as a cationic lipid and cholesterol (Chol) or 1,2-dioleoyl-L-α-glycero-3-phosphatidylethanolamine (DOPE) as a neutral lipid were used. In vitro and in vivo gene silencing effects by cationic lipoplexes were evaluated after transfection into stably luciferase-expressing human breast tumor MCF-7-Luc cells and after intravenous injection into mice with lung MCF-7-Luc metastasis, respectively. Intracellular localization of siRNA after transfection into MCF-7 cells by cationic lipoplexes and biodistribution of siRNA after intravenous injection of cationic lipoplexes into the mice with lung metastasis were examined by confocal and fluorescent microscopy analyses, respectively. RESULTS: In in vitro transfection, DOTAP/DOPE and DDAB/DOPE lipoplexes of luciferase siRNA strongly suppressed luciferase activity in MCF-7-Luc cells, but DOTAP/Chol and DDAB/Chol lipoplexes did not, although DOTAP/Chol and DDAB/Chol lipoplexes exhibited higher cellular uptake than DOTAP/DOPE and DDAB/DOPE lipoplexes. When their cationic lipoplexes were intravenously injected into mice with lung MCF-7-Luc metastasis, siRNAs were mainly accumulated in the lungs; however, the reduced luciferase activities in the lung-metastasized tumors were observed only by injections of DOTAP/Chol and DOTAP/DOPE lipoplexes, but not by DDAB/Chol and DDAB/DOPE lipoplexes. CONCLUSIONS: DOTAP-based liposomes might be useful as an in vivo siRNA delivery carrier that can induce gene silencing in lung-metastasized tumors.

Synergistic effect of pendant hydroxypropyl and pyrrolidine moieties randomly distributed along polymethacrylamide backbones on in vitro DNA-transfection.

In this work, the cationic monomer N-ethyl pyrrolidine methacrylamide (EPA) was copolymerized with the neutral monomer N-hydroxypropyl methacrylamide (HPMA) at different molar ratios obtaining linear random copolymers that were characterized and evaluated in vitro as non-viral gene carriers using murine Swiss 3T3 fibroblasts. The copolymers with excess or equimolar amount of EPA were able to complex DNA forming stable polyplexes with an average size between 50 and 200 nm, while the copolymers with an excess of HPMA do not. Cell viability was shown to depend on the EPA/HPMA molar ratio, exhibiting the equimolar copolymer poly (EPA-co-HPMA) 50:50 (EPA50) a full cytocompatibility, similar to the HPMA-rich systems. This copolymer EPA50 has also shown significantly higher transfection levels than the systems with other compositions and the positive controls poly L-lysine (PLL) and poly EPA (pEPA). This statistical equimolar copolymer EPA50 has unique properties related to its composition and microstructure, which allows it to complex DNA, showing an excellent biocompatibility and high transfection efficiency.

Reversal of multidrug resistance in vitro by co-delivery of MDR1 targeting siRNA and doxorubicin using a novel cationic poly(lactide-co-glycolide) nanoformulation.

Over expression of drug efflux transporters such as P-glycoprotein (P-gp) cumulatively leading to multidrug resistance (MDR) embodies a major hindrance for successful cancer therapy. A paradigm nanomedicinal approach involving an anticancer drug and modulators of drug resistance within one multifunctional nanocarrier-based delivery system represent an ideal modality for the treatment of MDR. In this regards, we have developed a cationic polymeric nanoparticulate system loaded with MDR1-siRNA and doxorubicin. Results indicated augmented synergistic effect of combinational nanoformulation in overcoming MDR in MCF-7/ADR cells. Therefore, the above regime could be a promising co-delivery system for effective therapy of drug resistant breast cancer.