Effects of hydrophobic and hydrophilic modifications on gene delivery of amphiphilic chitosan based nanocarriers

The structure-activity relationships between hydrophobic and hydrophilic modification on chitosan and resultant physicochemical properties along with performances in dealing with critical gene delivery barriers were investigated through amphiphilic linoleic acid(LA) and poly (β-malic acid) (PMLA) double grafted chitosan (LMC)/plasmid DNA (pDNA) nanocomplexes. LMC polymers with various LA and PMLA substitution degrees were synthesized and their hydrophilicity/hydrophobicity was characterized. Compared to chitosan, LMC nanoparticles retained the pDNA binding ability at pH 5.5 when they formed nanocomplexes with pDNA encoding enhanced green fluorescence protein (pEGFP) and the resultant complexes showed diameters below 300 nm. Hydrophobic LA and hydrophilic PMLA substitution contributed to suppressed non-specific adsorption, reduced interactions inside LMC/pDNA nanocomplexes, and enhanced pDNA dissociation. However, enzymatic degradation resistance, cell adsorption, and cellular uptake through clathrin-mediated pathway were promoted by hydrophobic LA grafting while being inhibited by hydrophilic PMLA substitution. In vitro transfection assay suggested the optimal LMC/pEGFP nanocomplexes mediated an 8.0-fold improved transfection compared to chitosan/pEGFP nanocomplexes. The 4.2-fold and 2.2-fold higher intramuscular gene expression in mice compared to chitosan/pEGFP and polyethyleneimine (PEI)/pEGFP nanocomplexes further demonstrated the superiority of LMC/pDNA nanocomplexes. Therefore, amphiphilic chitosan derivates with appropriate combination of hydrophobic and hydrophilic modification would be promising gene delivery nanocarriers.     

Oral delivery of shRNA and siRNA via multifunctional polymeric nanoparticles for synergistic cancer therapy

Galactose modified trimethyl chitosan-cysteine (GTC) conjugates with various galactose grafting densities were developed for oral delivery of Survivin shRNA-expression pDNA (iSur-pDNA) and vascular endothelial growth factor (VEGF) siRNA (siVEGF) in the synergistic and targeted treatment of hepatoma. iSur-pDNA and siVEGF loaded GTC nanoparticles (NPs) were prepared via electrostatic complexation and showed desirable stability in physiological fluids and improved intestinal permeation compared to naked genes. Galactose grafting density of GTC NPs significantly affected their in vitro and in vivo antitumor activities. GTC NPs with moderate galactose grafting density, termed GTC2 NPs, were superior in facilitating cellular uptake, promoting nuclear distribution, and silencing target genes, leading to notable inhibition of cell growth. In tumor-bearing mice, orally delivered GTC2 NPs could effectively accumulate in the tumor tissues and silence the expression of Survivin and VEGF, evoking increased apoptosis, inhibited angiogenesis, and thus the most efficient tumor regression. Moreover, compared with single gene delivery, co-delivery of iSur-pDNA and siVEGF showed synergistic effects on inhibiting in vitro cell proliferation and in vivo tumor growth. This study could serve as an effective approach for synergistic cancer therapy via oral gene delivery, and highlighted the importance of ligand grafting density in the rational design of targeted nanocarriers.     

Co-delivery of doxorubicin and paclitaxel by PEG-polypeptide nanovehicle for the treatment of non-small cell lung cancer

Despite progress, combination therapy of different functional drugs to increase the efficiency of anticancer treatment still remains challenges. An amphiphilic methoxy poly(ethylene glycol)-b-poly(l-glutamic acid)-b-poly(l-lysine) triblock copolymer decorated with deoxycholate (mPEsG-b-PLG-b-PLL/DOCA) was synthesized and developed as a nanovehicle for the co-delivery of anticancer drugs: doxorubicin (DOX) and paclitaxel (PTX). The amphiphilic copolymer spontaneously self-assembled into micellar-type nanoparticles in aqueous solutions and the blank nanoparticles possessed excellent stability. Three different domains of the copolymer performed distinct functions: PEG outer corona provided prolonged circulation, middle biodegradable and hydrophilic PLG shell was designed for DOX loading through electrostatic interactions, and hydrophobic deoxycholate modified PLL served as the container for PTX. In vitro cytotoxicity assays against A549 human lung adenocarcinoma cell line demonstrated that the DOX + PTX co-delivered nanoparticles (Co-NPs) exhibited synergistic effect in inducing cancer cell apoptosis. Ex vivo DOX fluorescence imaging revealed that Co-NPs had highly efficient targeting and accumulation at the implanted site of A549 xenograft tumor in vivo. Co-NPs exhibited significantly higher antitumor efficiency in reducing tumor size compared to free drug combination or single drug-loaded nanoparticles, while no obvious side effects were observed during the treatment, indicating this co-delivery system with different functional antitumor drugs provides the clinical potential in cancer therapy.     

Supramolecular self-assembly forming a multifunctional synergistic system for targeted co-delivery of gene and drug

For developing a multifunctional bioreducible targeted and synergistic co-delivery system for anticancer drug paclitaxel (PTX) and p53 gene for potential cancer therapy, supramolecular self-assembled inclusion complex was prepared from PTX and star-shaped cationic polymer containing γ-cyclodextrin (γ-CD) and multiple oligoethylenimine (OEI) arms with folic acid (FA) conjugated via a disulfide linker. The inclusion complex, termed as γ-CD-OEI-SS-FA/PTX, was formed between PTX and the hydrophobic cavity of γ-CD core of the star polymer. The γ-CD-OEI-SS-FA/PTX complex further formed polyplexes with pDNA to give positively charged nanoparticles, becoming multifunctional supramolecular self-assembled co-delivery system for PTX and pDNA targeting to cancer cells that overexpress folate receptors (FRs). The results showed that the FA-targeted function induced higher gene transfection efficiency in the FR-positive KB cells. The redox-sensitive disulfide linker in the self-assembly system led to the detachment of the FA groups from the carrier after the FR-mediated endocytosis, which resulted in the release of the bound FRs followed by the recycling of the FRs from the cytosol onto the cell membrane surface, facilitating continuous FR-mediated endocytosis to achieve enhanced gene transfection. In addition, the complexed PTX was co-delivered to the cells with pDNA, which further enhanced the gene transfection even at low N/P ratios in the FR-positive KB cells. Further, the efficient delivery of wild-type p53 gene resulted in large cell population at sub G1 and G2/M phases, inducing significant cell apoptosis. Therefore, the multifunctional γ-CD-OEI-SS-FA/PTX self-assembly system with the synergistic effects of redox-sensitive FA-targeted and PTX-enhanced p53 gene delivery may be promising for cancer therapeutic application.     

Co-delivery of hydrophobic paclitaxel and hydrophilic AURKA specific siRNA by redox-sensitive micelles for effective treatment of breast cancer

In this study, a novel redox-sensitive micellar system constructed from a hyaluronic acid-based amphiphilic conjugate (HA-ss-(OA-g-bPEI), HSOP) was successfully developed for tumor-targeted co-delivery of paclitaxel (PTX) and AURKA specific siRNA (si-AURKA). HSOP exhibited excellent loading capacities for both PTX and siRNA with adjustable dosing ratios and desirable redox-sensitivity independently verified by morphological changes of micelles alongside in vitro release of both drugs in different reducing environments. Moreover, flow cytometry and confocal microscopy analysis confirmed that HSOP micelles were capable of simultaneously delivering PTX and siRNA into MDA-MB-231 breast cancer cells via HA-receptor mediated endocytosis followed by rapid transport of cargoes into the cytosol. Successful delivery and transport amplified the synergistic effects between the drugs while leading to substantially greater antitumor efficacy when compared with single drug-loaded micelles and non-sensitive co-loaded micelles. In vivo investigation demonstrated that HSOP micelles could effectively accumulate in tumor sites and possessed the greatest antitumor efficacy over non-sensitive co-delivery control and redox-sensitive single-drug controls. These findings indicated that redox-sensitive HSOP co-delivery system holds great promise for combined drug/gene treatment for targeted cancer therapy.     

Amphiphilic block copolymer NPs obtained by coupling ricinoleic acid/sebacic acids and mPEG: Synthesis,characterization, and controlled release of paclitaxel

Abstract

Currently, nanoparticles (NPs) made of amphiphilic block copolymer are still an essential part of drug delivery system. Here, we report a novel amphiphilic block copolymer and paclitaxel (PTX)-loaded copolymer NPs for the controlled delivery of PTX. The block copolymer was synthesized via melt polycondensation method of methoxy poly(ethylene glycol) (mPEG), sebacic acid (SA) and ricinoleic acid (RA). A series of characterization approaches such as Fourier Transform Infrared Spectroscopy (FTIR), ¹Hydrogen-Nuclear Magnetic Resonance (¹H-NMR), Differential Scanning Calorimetry (DSC), X-Ray Diffraction (XRD) and Gel Permeation Chromatography (GPC) applied have shown that the amphiphilic block copolymer was prepared as designed. NPs prepared by nanoprecipitation method consist of mPEG segments as the hydrophilic shell and RA-SA segments as the hydrophobic core, hydrophobic PTX was encapsulated as model drug. Subsequently, Transmission Electron Microscopy (TEM) analysis indicated that the spherical NPs have effective mean diameters ranging from 100 to 400?nm. Dynamic Light Scattering (DLS) analysis also revealed the controllable NPs diameter by modulating the mass ratio of RA to SA and drug loading amount (DLA). Besides, biphasic profile with zero order drug release was observed in general in vitro release behaviors of PTX from NPs. Further investigation confirmed that the release behaviors depend on the crystallinity of hydrophobic RA-SA segments. Results above suggest that NPs with amphiphlic block copolymer mPEG-b-P(RA-SA)-b-mPEG have a remarkable potential as a carrier for hydrophobic drug delivery in cancer therapy.     

Co-delivery of SOX9 genes and anti-Cbfa-1 siRNA coated onto PLGA nanoparticles for chondrogenesis of human MSCs

Some genes expressed in stem cells interrupt and/or enhance differentiation. Therefore, the aim of this study was to inhibit the expression of unnecessary genes and enhance the expression of specific genes involved in stem cell differentiation by using small interfering RNA (siRNA) and plasmid DNA (pDNA) incorporated into cationic polymers as co-delivery factors. To achieve co-delivery of siRNA and pDNA to human mesenchymal stem cells (hMSCs), two different genes were complexed with poly(ethyleneimine) (PEI) and then coated onto poly(lactide-co-glycolic acid) (PLGA) nanoparticles (NP). To evaluate co-delivery of siRNA and pDNA into hMSCs, cells were transfected with green fluorescence protein (GFP) pDNA (GFP pDNA) and GFP siRNA (GFP siRNA). The percentage of GFP-expressing hMSCs decreased from 25.35 to 3.7% after transfection with GFP-DNA/PLGA NP (NPs) or GFP siRNA/PLGA NPs, whereas GFP-DNA/PLGA NPs and scramble siRNA (MOCK)/PLGA NPs had no effect on GFP expression. hMSCs cotransfected with coSOX9-pDNA/NPs and Cbfa-1-siRNA/NPs were tested both in vitro and in vivo using gel retardation, dynamic light scattering (DLS), and scanning electron microscope (SEM). The expression of genes and proteins associated with chondrogenesis was evaluated by FACS, RT-PCR, real time-qPCR, Western blotting, immunohistochemistry, and immunofluorescence imaging.     

Self-aggregated nanoparticles of linoleic acid-modified glycol chitosan conjugate as delivery vehicles for paclitaxel: preparation,characterization and evaluation

A series of linoleic acid-modified glycol chitosan (LAGC) conjugates were synthesized and characterized by FTIR and ¹H NMR. The effect of the amount of linoleic acid (LA) on the physicochemical properties of LAGC conjugates was investigated. The mean diameters of three LAGC nanoparticles determined by dynamic light scattering ranged from 204 to 289 nm. The critical aggregation concentration values of LAGC conjugates in aqueous solution were 0.0148, 0.0348, and 0.0807 mg/ml, respectively. Paclitaxel (PTX) was physically loaded into the LAGC nanoparticles by a dialysis method. The drug loading content and encapsulation efficiency of PTX-loaded LAGC (PTX-LAGC) nanoparticles increased with an increasing ratio of the hydrophobic LA to hydrophilic glycol chitosan in the conjugates. PTX-LAGC nanoparticles were almost spherical in shape observed by transmission electron microscopy. In vitro release revealed that PTX release from the nanoparticles was reduced as the LA substitution degree of LAGC conjugates increased. Compared with the commercial formulation Taxol, PTX-LAGC-1 nanoparticles exhibited comparable cellular uptake and cytotoxicity against HepG2 cells in vitro. Importantly, PTX-LAGC-1 nanoparticles demonstrated the stronger antitumor efficacy against hepatic H22 tumor-bearing mice than Taxol (p < 0.05). Therefore, glycolipid-like LAGC nanoparticles had a potential as delivery vehicles for tumor therapy.     

Enhanced anti-tumor efficacy by co-delivery of doxorubicin and paclitaxel with amphiphilic methoxy PEG-PLGA copolymer nanoparticles

The use of single chemotherapeutic drug has shown some limitations in anti-tumor treatment, such as development of drug resistance, high toxicity and limited regime of clinical uses. The combination of two or more therapeutic drugs is feasible means to overcome the limitations. Co-delivery strategy has been proposed to minimize the amount of each drug and to achieve the synergistic effect for cancer therapies. Attempts have been made to deliver chemotherapeutic drugs simultaneously using drug carriers, such as micelles, liposomes, and inorganic nanoparticles (NPs). Here we reported core-shell NPs that were doubly emulsified from an amphiphilic copolymer methoxy poly(ethylene glycol)-poly(lactide-co-glycolide) (mPEG-PLGA). These NPs offered advantages over other nanocarriers, as they were easy to fabricate by improved double emulsion method, biocompatible, and showed high loading efficacy. More importantly, these NPs could co-deliver hydrophilic doxorubicin (DOX) and hydrophobic paclitaxel (TAX). The drug-loaded NPs possessed a better polydispersity, indicating that they are more readily subject to controlled size distribution. Studies on drug release and cellular uptake of the co-delivery system demonstrated that both drugs were effectively taken up by the cells and released simultaneously. Furthermore, the co-delivery nanocarrier suppressed tumor cells growth more efficiently than the delivery of either DOX or TAX at the same concentrations, indicating a synergistic effect. Moreover, the NPs loading drugs with a DOX/TAX concentration ratio of 2:1 showed the highest anti-tumor activity to three different types of tumor cells. This nanocarrier might have important potential in clinical implications for co-delivery of multiple anti-tumor drugs with different properties.     

Synergistic inhibition of breast cancer by co-delivery of VEGF siRNA and paclitaxel via vapreotide-modified core–shell nanoparticles

A somatostatin analog, vapreotide (VAP), can be used as a ligand for targeting drug delivery based on its high affinity to somatostatin receptors (SSTRs), which is overexpressed in many tumor cells. RNA interference plays an important role on downregulation of vascular endothelial growth factor (VEGF), which is important for tumor growth, progression and metastasis. To improve tumor therapy efficacy, the vapreotide-modified core–shell type nanoparticles co-encapsulating VEGF targeted siRNA (siVEGF) and paclitaxel (PTX), termed as VAP-PLPC/siRNA NPs, were developed in this study. When targeted via somatostatin receptors to tumor cells, the VAP-PLPC/siRNA NPs could simultaneously delivery siVEGF and PTX into cells and achieve a synergistic inhibition of tumor growth. Interestingly, in vitro cell uptake and gene silencing experiments demonstrated that the targeted VAP-PLPC/siRNA NPs exhibited significant higher intracellular siRNA accumulation and VEGF downregulation in human breast cancer MCF-7 cells, compared to those of the non-targeted PEG-PLPC/siRNA NPs. More importantly, in vivo results further demonstrated that the targeted VAP-PLPC/siRNA NPs had significant stronger drug distribution in tumor tissues and tumor growth inhibition efficacy via receptor-mediated targeting delivery, accompany with an obvious inhibition of neovascularization induced by siVEGF silencing. These results suggested that the co-delivery of siRNA and paclitaxel via vapreotide-modified core–shell nanoparticles would be a promising approach for tumor targeted therapy.     

Contribution of hydrophobic/hydrophilic modification on cationic chains of poly(ε-caprolactone)-graft-poly(dimethylamino ethylmethacrylate) amphiphilic co-polymer in gene delivery

Nanoparticles (NPs) assembled from amphiphilic polycations have been certified as potential carriers for gene delivery. Structural modification of polycation moieties may be an efficient route to further enhance gene delivery efficiency. In this study two electroneutral monomers with different hydrophobicities, 2-hydroxyethyl methacrylate (HEMA) and 2-hydroxyethyl acrylate (HEA), were incorporated into the cationic poly(dimethylamino ethyl methacrylate) (PDMAEMA) side-chains of amphiphilic poly(ε-caprolactone)-graft-poly(dimethylamino ethylmethacrylate) (PCD) by random co-polymerization, to obtain poly(ε-caprolactone)-graft-poly(dimethylamino ethyl methacrylate-co-2-hydroxyethyl methacrylate) (PCD-HEMA) and poly(ε-caprolactone)-graft-poly(dimethylamino ethyl methacrylate-co-2-hydroxyethyl acrylate) (PCD-HEA). Minimal HEA or HEMA moieties in PDMAEMA do not lead to statistically significant changes in particle size, zeta potential, DNA condensation properties and buffering capacity of the naked NPs. However, the incorporation of HEMA and HEA lead to reductions and increases, respectively, in the surface hydrophilicity of the naked NPs and NPs/DNA complexes, which was confirmed by water contact angle assay. These simple modifications of PDMAEMA with HEA and HEMA moieties significantly affect the gene transfection efficiency on HeLa cells in vitro: PCD-HEMA NP/DNA complexes show a much higher transfection efficiency than PCD NPs/DNA complexes, while PCD-HEA NPs/DNA complexes show a lower transfection efficiency than PCD NP/DNA complexes. Fluorescence activated cell sorter and confocal laser scanning microscope results indicate that the incorporation of hydrophobic HEMA moieties facilitates an enhancement in both cellular uptake and endosomal/lysosomal escape, leading to a higher transfection efficiency. Moreover, the process of endosomal/lysosomal escape confirmed in our research that PCD and its derivatives do not just rely on the proton sponge mechanism, but also on membrane damage due to the polycation chains, especially hydrophobic modified ones. Hence, it is proved that hydrophobic modification of cationic side-chains is a crucial route to improve gene transfection mediated by polycation NPs.     

Gold nanoparticles with a monolayer of doxorubicin-conjugated amphiphilic block copolymer for tumor-targeted drug delivery

Gold (Au) nanoparticles (NPs) stabilized with a monolayer of folate-conjugated poly(l-aspartate-doxorubicin)-b-poly(ethylene glycol) copolymer (Au-P(LA-DOX)-b-PEG-OH/FA) was synthesized as a tumor-targeted drug delivery carrier. The Au-P(LA-DOX)-b-PEG-OH/FA NPs consist of an Au core, a hydrophobic poly(l-aspartate-doxorubicin) (P(LA-DOX)) inner shell, and a hydrophilic poly(ethylene glycol) and folate-conjugated poly(ethylene glycol) outer shell (PEG-OH/FA). The anticancer drug, doxorubicin (DOX), was covalently conjugated onto the hydrophobic inner shell by acid-cleavable hydrazone linkage. The DOX loading level was determined to be 17 wt%. The Au-P(LA-DOX)-b-PEG-OH/FA NPs formed stable unimolecular micelles in aqueous solution. The size of the Au-P(LA-DOX)-b-PEG-OH/FA micelles were determined as 24–52 and 10–25 nm by dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. The conjugated DOX was released from the Au-P(LA-DOX)-b-PEG-OH/FA micelles much more rapidly at pH 5.3 and 6.6 than at pH 7.4, which is a desirable characteristic for tumor-targeted drug delivery. Cellular uptake of the Au-P(LA-DOX)-b-PEG-OH/FA micelles facilitated by the folate-receptor-mediated endocytosis process was higher than that of the micelles without folate. This was consistent with the higher cytotoxicity observed with the Au-P(LA-DOX)-b-PEG-OH/FA micelles against the 4T1 mouse mammary carcinoma cell line. These results suggest that Au-P(LA-DOX)-b-PEG-OH/FA NPs could be used as a carrier with pH-triggered drug releasing properties for tumor-targeted drug delivery.     

Gelatin coating to stabilize the transfection ability of nucleic acid polyplexes

Amphiphilic polymers are effective in complexing and delivering therapeutic nucleic acids, such as plasmid DNA (pDNA) and short interfering RNA (siRNA). However, long-term stability of the complexes is not desirable, as it may have an impact on the transfection efficiency in vivo. To develop a method to preserve complex stability we first showed that pDNA complexes formed with the amphiphilic polymer linoleic acid-substituted polyethylenimine (PEI–LA) and incubated at 37 °C lost ～90% of their transfection efficiency after only 24 h of complex formation. Polyethyleneglycol modification of complexes to control the increase in complex size and incubation in scaffolds used for implantation did not preserve the transfection ability of the complexes. Among a variety of approaches explored, gelatin coating of complexes was found to be the best at maintaining the original transfection efficiency. Mechanistic studies suggested that improved complex uptake, not size stability, was responsible for retention of the transfection efficiency. Similarly to the results with pDNA, gelatin coating also prevented the decreases in uptake and silencing efficiency of siRNA complexes observed following incubation at 37 °C. Gelatin-stabilized complexes were, furthermore, effective in vivo and led to subcutaneous transgene expression with a low pDNA dose that was otherwise ineffective. We conclude that a simple gelatin coating approach offers an efficient means to preserve the transfection efficiency of polyplexes.     

The role of hydrophobicity in the disruption of erythrocyte membrane by nanoparticles composed of hydrophobically modified poly(γ-glutamic acid)

Polymeric nanoparticles (NPs) prepared from biocompatible polymers have been studied extensively as carriers for the targeted and controlled delivery of antigens to develop safe and effective vaccines. Especially, the endosomal escape of antigens is essential for the induction of antigen-specific potent immune responses, and the NPs which can control the endosomal escape are urgently required. It has been reported that the hydrophobicity of polymers affected the interactions between the polymer and the membranes, but there have no reports about investigating the effect of the hydrophobicity of the NPs on the membrane disruptive property. In this study, we evaluated the effect of hydrophobicity of NPs on the membrane disruptive property for the first time. We prepared NPs composed of amphiphilic poly(γ-glutamic acid) (γ-PGA) with various grafting degrees of hydrophobic backbone (43–71%), and evaluated the membrane disruptive property. These NPs showed membrane disruptive activity only at the endosomal pH range, and this activity was dependent on the hydrophobicity of γ-PGA. The dependency of the membrane disruptive property on the hydrophobicity of NPs was due to the surface hydrophobicity of them. Our results could provide a guideline for the rational design of amphiphilic polymers as nanoparticle-based vaccine carriers.     

Star-branched amphiphilic PLA-b-PDMAEMA copolymers for co-delivery of miR-21 inhibitor and doxorubicin to treat glioma

The combined treatment of chemotherapeutant and microRNA (miR) has been proven to be a viable strategy for enhancing chemosensitivity due to its synergistic effect for tumor therapy. However, the co-delivery of drugs and genes remains a major challenge as they lack efficient co-delivery carriers. In this study, three amphiphilic star-branched copolymers comprising polylactic acid (PLA) and polydimethylaminoethyl methacrylate (PDMAEMA) with AB₃, (AB₃)₂,and (AB₃)₃ molecular architectures were synthesized respectively by a combination of ring-opening polymerization, atom transfer radical polymerization, and click chemistry via an “arm-first” approach. The star copolymers possessed a low critical micelle concentration (CMC) and formed nano-sized micelles with positive surface charges in water as well as exhibiting a much lower cytotoxicity than PEI 25 kDa. Nevertheless, their gene transfection efficiency and tumor inhibition ability showed a remarkable dependence on their molecular architecture. The (AB₃)₃ architecture micelle copolymer exhibited the highest transfection efficiency, about 2.5 times higher than PEI. In addition, after co-delivering DOX and miR-21 inhibitor (miR-21i) into LN229 glioma cells, the micelles could mediate escaping miR-21i from lysosome degradation and the release of DOX to the nucleus, which significantly decreased the miR-21 expression. Moreover, co-delivery of DOX and miR-21i surprisingly exhibited an anti-proliferative efficiency compared with DOX or the miR-21i treatment alone. These results demonstrated that amphiphilic star-branched copolymers are highly promising for their combinatorial delivery of genes and hydrophobic therapeutants.     

A self-assembling nanoparticle for paclitaxel delivery in ovarian cancer

Paclitaxel (PTX) is one of the most effective chemotherapeutic drugs for the treatment of a variety of cancers. However, it is associated with serious side effects caused by PTX itself and the Cremophor EL emulsifier. In the present study, we report the development of a well-defined amphiphilic linear-dendritic copolymer (named as telodendrimer) composed of polyethylene glycol (PEG), cholic acid (CA, a facial amphiphilic molecule) and lysine, which can form drug-loaded core/shell micelles when mixed with hydrophobic drug, such as PTX, under aqueous condition. We have used PEG^5k-CA₈, a representive telodendrimer, to prepare paclitaxel-loaded nanoparticles (PTX-PEG^5k-CA₈ NPs) with high loading capacity (7.3 mg PTX/mL) and a size of 20–60 nm. This novel nanoformulation of PTX was found to exhibit similar in vitro cytotoxic activity against ovarian cancer cells as the free drug (Taxol^®) or paclitaxel/human serum albumin nanoaggregate (Abraxane^®). The maximum tolerated doses (MTDs) of PTX-PEG^5k-CA₈ NPs after single dose and five consecutive daily doses in mice were approximately 75 and 45 mg PTX/kg, respectively, which were 2.5-fold higher than those of Taxol^®. In both subcutaneous and orthotopic intraperitoneal murine models of ovarian cancer, PTX-PEG^5k-CA₈ NPs achieved superior toxicity profiles and anti-tumor effects compared to Taxol^® and Abraxane^® at equivalent PTX doses, which were attributed to their preferential tumor accumulation, and deep penetration into tumor tissue, as confirmed by near infrared fluorescence (NIRF) imaging.     

Co-delivery of pEGFP-hTRAIL and paclitaxel to brain glioma mediated by an angiopep-conjugated liposome

In this study, we report an angiopep-2 modified cationic liposome (ANG-CLP) for the efficient co-delivery of a therapeutic gene encoding the human tumour necrosis factor-related apoptosis-inducing ligand (pEGFP-hTRAIL) and paclitaxel (PTX) to glioma. The dual targeting co-delivery system (ANG-CLP/PTX/pEGFP-hTRAIL) improved uptake and gene expression not only in U87 MG cells and BCECs, but also in the glioma bed and infiltrating margin of intracranial U87 MG glioma-bearing models. The system selectively induces apoptosis in U87 MG cells while reducing toxicity to BCECs. The results of the pharmacodynamics studies showed that the apoptosis of glioma cells in in vitro BBB models and in U87 MG glioma-bearing mice induced by ANG-CLP/PTX/pEGFP-hTRAIL was more apparent and widespread than that induced by single medication systems and unmodified co-delivery system. More importantly, the median survival time of brain tumour-bearing mice treated with ANG-CLP/PTX/pEGFP-hTRAIL was 69.5 days, significantly longer than that of other groups, even longer than the commercial temozolomide group (47 days). Therefore, the dual targeting co-delivery system is a promising drug delivery strategy against glioma.     

紫杉醇聚合物纳米囊泡的制备和体外释放研究

目的 以聚己内酯-b-聚乙二醇-6-聚己内酯（PCEP）两亲性三嵌段共聚物为载体研制紫杉醇聚合物纳米囊泡.方法 以不同分子量的聚乙二醇（PEG）引发合成不同亲水段、疏水段链长的PCEP并进行FT-IR、1H NMR和GPC表征,以合成的嵌段聚合物PCEP为载体,通过薄膜-超声分散法制备紫杉醇聚合物纳米囊泡,用透射电子显微镜（TEM）表征其形态和构造,用粒度分析仪测定其粒径及分布,用高效液相色谱（HPLC）法测定其载药量及包封率,用透析袋法研究药物体外释放;同时,研究不同亲水链长、疏水链长对紫杉醇聚合物囊泡载药量、包封率、粒径及体外释放紫杉醇药物的影响.结果 研制的紫杉醇聚合物囊泡呈核-壳结构球形,粒径为纳米级,随着PCEP共聚物相对分子质量的增加而增大;紫杉醇聚合物囊泡体外释放无突释现象,能稳定缓慢释放紫杉醇,且释放速率随共聚物中亲水段PEG含量增加而增大,随疏水段PCL含量增大而减小.结论 以PCEP两亲性三嵌段共聚物为载体制备的紫杉醇聚合物纳米囊泡,其粒径小且分布均匀,包封率较高,有望成为一种用于提高紫杉醇的药效且降低不良反应的新的紫杉醇缓控释剂型.     

Constructing doxorubicin-loaded polymeric micelles through amphiphilic graft polyphosphazenes containing ethyl tryptophan and PEG segments

By changing the molar ratio of hydrophilic and hydrophobic segments, a series of novel amphiphilic graft polyphosphazenes (PEG/EtTrp-PPPs) was synthesized via thermal ring-opening polymerization and a subsequent two-step substitution reaction of hydrophilic methoxyl polyethylene glycol (MPEG) and hydrophobic ethyl tryptophan (EtTrp). ¹H-Nuclear magnetic resonance and Fourier transform infrared studies validated the expected synthesis of copolymers. The copolymer composition was also confirmed by UV–visible spectrophotometry. The molar ratio of the segment PEG to group EtTrp was 1.33:0.67, 1.01:0.99 and 0.78:1.22, respectively. Micellization behavior of PEG/EtTrp-PPPs in an aqueous phase was characterized by fluorescence technique, dynamic light scattering and transmission electron microscopy. The critical micelle concentration (CMC) of the graft copolymer in aqueous solution was 0.158, 0.033 and 0.020 g l^?1, which decreased as the hydrophobic content in amphiphilic copolymers increased. Doxorubicin (DOX) was physically loaded into micelles prepared by an O/W emulsion method with a drug loading content increasing with DOX feeding. In vitro release of DOX from micelles can be accelerated in weak acidic solution. The results of cytotoxicity study using an MTT assay method with HeLa cell showed that amphiphilic graft polyphosphazenes were biocompatible while DOX-loaded micelles achieved comparable cytotoxicity with that of free DOX. In summary, these novel amphiphilic copolymers exhibited potential to be used as injectable drug carriers for tumor treatment.     

Thermosensitive poly(organophosphazene)–paclitaxel conjugate gels for antitumor applications

A poly(organophosphazene)–PTX conjugate was synthesized by a covalent ester linkage between PTX and carboxylic acid-terminated poly(organophosphazene), which can be readily modified by various hydrophobic, hydrophilic, and other functional substitutes. The physicochemical properties, hydrolytic degradation and PTX release behaviors of the polymer–PTX conjugate were characterized, in addition to the in vitro and in vivo antitumor activities. The aqueous solutions of these conjugates showed a sol–gel transition behavior that depended on temperature changes. The in vitro antitumor activity of the polymer–PTX conjugate was investigated by an MTT assay against human tumor cell lines. From the in vivo antitumor activity studies with tumor-induced (xenografted) nude mice, the polymer–paclitaxel conjugate hydrogels after local injection at the tumor site were shown to inhibit tumor growth more effectively and longer than paclitaxel and saline alone, indicating that the tumor-active paclitaxel from the polymer–PTX conjugate hydrogel is released slowly over a longer period of time and effectively accumulated locally in the tumor sites. These combined observations suggest that this poly(organophosphazene)–PTX conjugate holds promise for use in clinical studies as single and/or combination therapies.