Amphipathic Polyethylene Glycol Research Articles

Preliminary study on fabrication, characterization and synergistic anti-lung cancer effects of self-assembled micelles of covalently conjugated celastrol–polyethylene glycol–ginsenoside Rh2

The aim of this study was to develop an amphipathic polyethylene glycol (PEG) derivative that was bi-terminally modified with celastrol and ginsenoside Rh2 (Celastrol-PEG-G Rh2). Such derivative was capable of forming novel, celastrol-loaded polymeric micelles (CG-M) for endo/lysosomal delivery and thereby synergistic treatment of lung cancer. Celastrol-PEG-G Rh2 with a yield of 55.6% was first synthesized and characterized. Its critical micellar concentration was 1 × 10−5 M, determined by pyrene entrapment method. CG-M had a small particle size of 121.53 ± 2.35 nm, a narrow polydispersity index of 0.214 ± 0.001 and a moderately negative zeta potential of –23.14 ± 3.15 mV. Celastrol and G Rh2 were rapidly released from CG-M under acidic and enzymatic conditions, but slowly released in normal physiological environments. In cellular studies, the internalization of celastrol and G Rh2 by human non-small cell lung cancer (A549) cells treated with CG-M was 5.8-fold and 1.8-fold higher than that of non-micelle control. Combinational therapy of celastrol and G Rh2 using CG-M exhibited synergistic anticancer activities in cell apoptosis and proliferation assays via rapid drug release within endo/lysosomes. Most importantly, the celastrol in CG-M exhibited a long elimination half-life of 445.3 ± 43.5 min and an improved area under the curve of 645060.8 ± 63640.7 ng/mL/h, that were 1.03-fold and 2.44-fold greater than those of non-micelle control, respectively. These findings suggest that CG-M is a promising vector for precisely releasing anticancer drugs within the tumor cells, and thereby exerts an improved synergistic anti-lung cancer effect.

A submicron lipid emulsion coated with amphipathic polyethylene glycol for parenteral administration of paclitaxel (Taxol).

Paclitaxel is a promising anticancer agent with poor solubility in water and requires a suitable formulation for intravenous administration. Presently paclitaxel is formulated for clinical use in ethanol and Cremophor EL (Diluent 12), a solvent system associated with severe adverse effects. In this study paclitaxel was entrapped in lipid emulsion droplets with triolein as oil core and dipalmitoyl phosphatidylcholine as the principal emulsifier. The emulsion was further stabilized with polysorbate 80 and polyethylene glycol-dipalmitoyl phosphatidylethanolamine. The drug-emulsion droplets (diameter about 40 nm) were physically and chemically stable during several months at 4 degrees C. Lyophilized preparations in 5% glucose were completely restored by distilled water. Studies of the integrity of the drug-emulsion showed a release of the drug from emulsion globules and surface transfer was found to be the major mechanism for cellular uptake. The in-vitro antiproliferative activity of paclitaxel against T-47D cells was retained by the drug-emulsion with an ID50 value of 7 nM compared to 10 and 35 nM for paclitaxel in liposomes and Diluent 12, respectively. Long-circulating submicron lipid emulsions may prove useful, not only for replacement of the more toxic Cremophor EL vehicle, but also by improving the distribution of the drug to the tumour.

膜の構造と機能設計 ステルスリポソームを用いたデリバリーシステム

Newly developed liposomes, containing amphipathic polyethylene glycol (PEG) derivatives, are not readily taken up by the macrophages in the RES and hence stay in the circulation for a relatively long period of time. PEG is useful because of its ease of preparation, relatively low cost, controllability of molecular weight and linkability to lipids or protein including the antibody by a variety of methods. The presence of PEG reduces binding of serum protein, i.e. opsonins marking the liposome for clearance by MPS. PEG-liposome can take advantage of the EPR effect (enhanced permeability and retention effect) for efficient targeting binding in the tumor.The blood clearance of immunoliposomes must be minimized in comparison with rate of extravasation in the tumor. As described above, PEG-liposomes offer the development of immunoliposomes with both long survival times in circulation and target recognition being retained in vivo. For the active targeting following the passive targeting to the solid tumor tissue, Fab' fragment of 21B2 antibody which is anti-human CEA or transferrin (TF) was conjugated to prepared the pendant type immunoliposome (Fab'-PEG-LP or TF-PEG-ILP). TF-PEG-LP could internalized into tumor cells with receptor mediated endocytosis following extravasation into tumor tissue. Such liposomal formulations should be useful for endocytotic internalization of plasmid DNA and other bioactive materials.

Targeting chemotherapy to solid tumors with long-circulating thermosensitive liposomes and local hyperthermia.

The effectiveness of the combination of long‐circulating, thermosensitive liposomes and hyperthermia is described. Small‐sized, thermosensitive liposomes that encapsulate doxorubicin (DXR‐PEGTSL (SUV)) have a prolonged circulation time and are extravasated to targeted solid tumors in vivo, where they preferentially release the agent in an anatomical site subjected to local hyperthermia. Liposomes were prepared by the incorporation of amphipathic polyethyleneglycol (PEG) to prolong their circulation time. DXR‐PEG‐TSL (SUV) was retained longest and was accumulated most efficiently in solid tumors in Balb/c mice. The combination of DXR‐PEG‐TSL (SUV) and hyperthermia at the tumor sites 3 h after injection, gave high concentrations of doxorubicin in tumor tissue and resulted in more effective tumor retardation and increased survival time. A large amount of DXR‐PEG‐TSL (SUV) was extravasated into the tumors during circulation for 3 h after injection, suggesting that the encapsulated drug was released into the interstitial spaces of the lesions by local hyperthermia. This system is expected to be clinically valuable for the delivery of a wide range of chemotherapeutic agents in the treatment of solid tumors.

リポソームによるパッシブターゲティング

Passive targeting with liposomal drug carrier systems was reviewed. The current status of the passive targeting by the pnlyethyleneglycol coated liposome (PEG-liposome) were described in this review. Newly developed liposomes, containing either monosialoganglioside GM1 or amphipathic polyethylene glycol (PEG) derivatives, are not readily taken up by the macrophages in the RES and hence stay in the circulation for a relatively long period of time. Particularly, PEG is useful because of its ease of preparation, relatively low cast, controllability of molecular weight and linkability to lipids or protein including the antibody by a variety of methods as compared with GM1 molecules. So many recent studies have focused on the use of liposomes with surface associated PEG. The presence of PEG reduces binding of serum protein, i.e. opsonins marking the lipnsome for clearance by MPS. Pharmacokinetic analysis and therapeutic studies with tumor bearing mice revealed that PEG-liposomes with an average diameter of 100-200 nm were accumulated efficiently in tumor tissue. Due to the capillary permeability of the endothelial barrier in newly vascularized tumors is significantly greater than that of normal tissues, PEG-liposomes could extravasate from blood circulation to tumor tissue. Results from clinical studies with doxorubicin encapsulated into PEG-liposomes (DOXIL®) in AIDS-related Kaposi's sarcoma revealed an increased therapeutic efficacy compared to free-drug. Solid tumors generally possess the following pathophysiological characteristics : (a) hypervasculature, (b) incomplete vascular architecture, (c) secretion of vascular permeability factors that stimulate extravasation within the cancer, (d) little drainage (lack of lymph vessel) of macromolecules and particles, which results in their long-term retention in tumor tissue. These characteristics of solid tumors are the basis of the so-called EPR effect (enhanced permeability and retention effect). Thus, the permeability of the endothelial barrier in newly vascularized tumors is increased compared with that of healthy tissues. PEG-liposome can take advantage of the EPR effect for efficient targeting binding in the tumor. The localization of PEG-liposomes into the interstitial space between tumor cells by a process of extravasatiun from tumor vessels (EPR effect) was revealed by the electron microscopic observations. Immunoliposomes for the treatment of solid tumor should satisfy a number of requirements aimed at maximum targeting effect of immunoliposome administered systemically in the bloodstream. Antigen binding site of the liposome-cunjugated antibody must be accessible for unperturbed interaction with antigen on the surface of target cells. The blood clearance of immunoliposomes must be minimized in comparison with rate of extravasation in the tumor. As described above, PEG-liposomes offer the development of immunoliposomes with both long survival times in circulation and target recongnition being retained in vivo. A new type of long-circulating immunoliposome, which was PEG-immunoliposome attached antibody at the distal end of PEG chain, so called the pendant type immunoliposome, was designed. To assist extravasation, the liposomes were of uniform, small size (100-130 nm). Elimination of immunogenic effect of Fc portion and of the increased RES clearance through specific recognition by MPS cells carrying Fc receptor, was achieved by using Fab' fragment instead of the whole antibody.

Stabilization and Regulated Fusion of Liposomes Containing a Cationic Lipid Using Amphipathic Polyethyleneglycol Derivatives

Current fusogenic liposomal delivery systems have limited applicability in vivo due to poor stability in the blood and rapid clearance from the circulation. This is particularly true for liposomes composed of dioleoylphosphatidylethanolamine (DOPE) and cationic lipids, currently being developed for systemic delivery of gene-based drugs. This paper describes a potential strategy to overcome these problems, involving the incorporation of exchangeable amphipathic polyethyleneglycol (PEG) derivatives to transiently stabilize fusogenic liposomes while in the circulation, but where the PEG coating dissipates to reveal fusogenic character at later times after arrival at target sites. It is shown here that large unilamellar vesicles (LUVs) containing Dope and the cationic lipid, N,N-dioleoyl-N, N-dimethylammonium chloride (DODAC) can be stabilized against serum-induced aggregation and fusion by inclusion of at least 2 mol% of PEG coupled to phosphatidylethanolamine (PEG-PE) or ceramide (PEG-Cer). However, low in vitro recovery of fusogenic activity was obtained for the PEG-PE-containing system, presumably due to electrostatic interactions between the negatively charged PEG-PE and the cationic lipid which prevent PEG-PE dissociation from the LUV. Improved recovery of fusogenic activity was achieved for LUVs stabilized by the neutral PEG-Cer derivatives, with shorter chain ceramides exhibiting more rapid recovery rates. Biodistribution studies showed that DOPE/DODAC (85:15, mol/mol) LUVs were rapidly cleared from the circulation, whereas inclusion of 10 mol% PEG-Cer(C20) resulted in significantly prolonged circulation time. Inclusion of shorter ceramide chain lengths resulted in decreased circulation times, consistent with increased exchangeability. These findings demonstrate the feasibility of developing a cationic liposome that is stable in the circulation, but retains its ability to fuse with membranes. This work represents the first step toward the rationale design of fusogenic cationic liposomes for the systemic delivery of gene-based drugs to target tissues, such as tumors.

Preparation of Thermosensitive Liposomes Containing Amphipathic Polyethyleneglycol for Macromolecule Delivery

Preparation of Thermosensitive Liposomes Containing Amphipathic Polyethyleneglycol for Macromolecule Delivery

Submicron lipid emulsions containing amphipathic polyethylene glycol for use as drug-carriers with prolonged circulation time

Submicron-sized lipid emulsions possess many favourable properties as drug-carriers like biocompatibility, physical stability and ease of preparation. The use of colloidal carriers is, however, hampered by their rapid clearance from the circulation. In the present study, this problem was tackled by coating the emulsion droplets with polyethylene glycol modified phosphatidylethanolamine (PEG-PE). Physical stability was obtained by using a mixture of phosphatidylcholine and polysorbate 80 as emulsifying agent. Sonicated preparations with the standard composition; triolein (TO): dipalmitoyl phosphatidylcholine (DPPC): polysorbate 80: PEG-PE at mass ratios of 2: 1: 0.4: 0.1 exhibited a mean particle size of 44 nm (by quasi-elastic light scattering) and an excellent physical stability. In vivo plasma clearance data were obtained by intravenous injection of emulsions into mice. Coating of the emulsion droplets with PEG-PE gave a considerable increase in circulation lifetime. A further notable effect was obtained when the cosurfactant polysorbate 80 was introduced into the system, apparently as a result of decreased particle size. Lipid emulsions with the standard composition showed first order kinetics during 6 h with a circulation half-life of about 3 h. Phospholipid transition temperature and emulsion particle size were found to be important factors while the clearance rate was essentially independent of lipid dose.

Amphipathic polyethylene glycol stabilized emulsions (o/w): Physical characterization and in vivo distribution

Injectable submicron oil-in-water emulsions have important potential as a carrier for hydrophobic drugs. Conventional formulations, however, using phosphatidylcholine (PC) either with or without co-emulsifiers cannot generally be used because of their fast elimination from the blood by the reticuloendothelial system. To overcome this obstacle, we have included the amphipathic molecules of polyethylene glycol derivatives of phosphatidylethanolamine (PEG-PE) as a co-emulsifier into emulsions containing castor oil and phosphatidylcholine. Compared to the emulsions with phosphatidylcholine as the sole emulsifier or those with Tween 80 as the co-emulsifier, emulsions containing PEG-PE as a co-emulsifier of PC show a decreased liver uptake and increased blood circulation time in mice. Furthermore, they are much more stable than the conventional formulations as tested under various conditions. With superior stability and a prolonged circulation time in blood, emulsions containing amphipathic molecules of PEG may be useful for sustained release and targeted delivery of lipophilic drugs.

Long-circulating emulsions (oil-in-water) as carriers for lipophilic drugs.

Rapid clearance of parenterally administered oil-in-water emulsions from blood by the reticuloendothelial system (RES), mainly macrophages of the liver and spleen, has been one of the major obstacles for delivering lipophilic drugs to cells other than those in the RES. The purpose of this study therefore is to overcome this problem and develop emulsions that will have prolonged blood circulation time. A series of amphipathic polyethylene-glycol (PEG) derivatives have been included as co-emulsifier into emulsions composed of Castor oil and phosphatidylcholine. The effect of amphipathic PEG on reducing the RES uptake and prolonging the blood circulation of the emulsion particles has been tested in vivo using mice as an animal model. Inclusion of PEG derivatives such as Tween-80 or dioleoyl N-(monomethoxy-polyethyleneglycol succinyl)phosphotidylethanolamine (PEG-PE) into emulsions composed of Castor oil and phosphatidylcholine decreases the RES uptake and increases blood residence time of the emulsions. The activity of PEG derivatives in prolonging the circulation time of emulsions depends on the PEG chain length (PEG2000 > PEG5000 > PEG1000, Tween-80) and the PEG density on emulsion surface. Inclusion of amphipathic PEG as emulsifier into oil-in-water emulsions is a very effective method to prolong the blood half life of the emulsions. Emulsions with long circulating half life in blood should be very useful as a delivery vehicle for lipophilic drugs.

Biodistribution and Antitumor Effect of Adriamycin Encapsulated in Long-Circulating Liposomes Containing Amphipathic Polyethylene Glycol or Ganglioside GM1

AbstractBiodistribution and antitumor effect of adriamycin (ADM) encapsulated in liposomes with reduced uptake by reticuloendothelial system (RES) and prolonged circulation time were investigated in mice. Two different types of long-circulating liposomes, ganglioside GMI (GMl)/distearoyl phosphatidyl choline (DSPC) /cholesterol (CH) (0.13:1:1, m/m) and amphipathic polyethylene glycol (PEG)/DSPC/CH (0.13:1:1, m/m) were used. They were sized to 180-200 nm in mean diameter. In the case of amphipathic PEG, distearoyl phosphatidylethanolamine (DSPE) derivatives of PEG with various molecular weight (1000-12000 in mean molecular weight) were used. ADM was encapsulated by transmembrane pH gradient method. GM~DSPC /CH

Enhanced delivery and antitumor activity of doxorubicin using long-circulating thermosensitive liposomes containing amphipathic polyethylene glycol in combination with local hyperthermia.

Enhanced delivery of doxorubicin (DXR) to a solid tumor subjected to local hyperthermia was achieved by using long-circulating, thermosensitive liposomes (TSL) composed of dipalmitoyl phosphatidylcholine (DPPC)/distearoyl phosphatidylcholine (DSPC) (9:1, m/m) and 3 mol% amphipathic polyethylene glycol (PEG) in colon 26-bearing mice. Inclusion of 3 mol% of distearoyl phosphatidylethanolamine derivatives of PEG (DSPE-PEG, amphipathic PEG) with a mean molecular weight of 1000 or 5000 in DPPC/DSPC liposomes resulted in decreased reticuloendothelial system (RES) uptake and a concomitant prolongation of circulation time, affording sustained increased blood levels of the liposomes. Concomitantly, DXR levels in blood were also kept high over a long period. The presence of amphipathic PEG did not interfere with the encapsulation of DXR by the pH gradient method (> 90% trapping efficiency) or with the temperature-dependent drug release from the liposomes. The optimal size of these liposomes was 180-200 nm in mean diameter for thermosensitive drug release and prolonged circulation time. The DXR levels in the tumor after injection of long-circulating TSL (DXR-PEG1000TSL or DXR-PEG5000TSL, at a dose of 5 mg DXR/kg) with local hyperthermia were much higher than after treatment with DXR-TSL lacking PEG or with free DXR, reaching 7.0-8.5 DXR micrograms/g tumor (approximately 2 times or 6 times higher than that of DXR-TSL or free DXR, respectively). Furthermore, the combination of DXR-PEGTSL and hyperthermia effectively retarded tumor growth and increased survival time.(ABSTRACT TRUNCATED AT 250 WORDS)

Enhanced Delivery and Antitumor Effect of Doxorubicin Encapsulated in Long-Circulating Liposomes

AbstractLong-circulating liposomes containing amphipathic polyethyleneglycol (PEG) or ganglioside GM1 (GM1) have been tested for their utility as enhanced delivery system of doxorubicin (DXR) in vivo. DXR was entrapped into liposomes by pH gradient method.The long-circulating LUV (200 nm in size) composed of DSPC/CH (1:1, m/m) and either 6 mol% of DSPE-PEG1000 or GM1 entrapped DXR with >95% in trapping efficiency. DXR-long-circulating LUVs were administered to leukemic (LI210) mice via the tail vein at a dose of 5mg DXR/kg. The high blood concentration was kept for long time, and significantly increased survival time was observed as compared with free DXR and DXR-LUV. The data indicated that DXR was slowly released from long-circulating LUV during that stayed in bloodstream for long time. Administration of DXR-long-circulating SUV (100 nm) to the colon 26 bearing mice produced the increased DXR level in tumor compared with bear SUV or free drug did, respectively, and resulted in effective tumor growth ret...

ポリエチレングリコール付与リポソームによるドキソルビシンの固形癌への移行量増加と抗腫よう効果

Long-circulating liposomes containing amphipathic polyethyleneglycol (PEG) have been tested for their utility as enhanced delivery system of doxorubicin(DXR) to the solid tumor. Inclusion of PEG to the small unilamellar liposomes(SUV, DSPC/CH (1 : 1, m/m), 120 nm in mean diameter) encapsulating DXR showed the increased blood level and the decreased RES uptake of SUV, and resulted in high accumulation of SUV into the colon 26 solid tumor site. Reflecting to these behavior of PEG-SUV, DXR concentration in tumor tissue was significantly increased, and effective tumor-growth retardation and increased survival time were observed. These data indicate that since the capillary permeability of the endothelial barrier in newly vascularized tumors is significantly greater than that of normal organs, SUV with prolonged circulation time could predominantly pass through leaky endothelium into tumor area by passive convective transport. DXR encapsulated in long-circulating liposomes will be much more effective for cancer chemotherapy than either the usual liposomal DXR or free DXR.

Doxorubicin Encapsulated in Long-Circulating Thermosensitive Liposomes

AbstractDoxorubicin (DXR) was encapsulated in long-circulating, thermosensitive liposomes (TSL, 180-200 nm in mean diameter), prepared from dipalmitoyl phosphatidyl choline (DPPC)/distearoyl phosphatidyl choline (DSPC) (9:1, m/m) and either 3 mol% of amphipathic polyethylene glycol (PEG) with 1000 in average molecular weight or 6 mol% of ganglioside GMI (GMI), with 95-98% entrapping efficiency by the pH gradient method. 57% or 45% of the entrapped DXR was released from PEG/DPPC/DSPC or GM1/DPPC/DSPC liposomes, respectively, by incubation with 20% serum at 42°C for 5 min. Inclusion of PEG or GM1 endowed TSL with prolonged circulation ability, resulting in increased blood levels of liposomes and decreased reticuloendothelial system (RES) uptake over 6 hours after injection. Concomitantly, high DXR level in blood was kept for long time.Accumulation of DXR into tumor tissue of tumor-bearing mice (mouse colon carcinoma 26) by local hyperthermia after injection of DXR-long-circulating TSL was 2 times or 7 times...

Effect of molecular weight in amphipathic polyethyleneglycol on prolonging the circulation time of large unilamellar liposomes.

The effect in mice of the molecular weight of polyethyleneglycol on prolonging the circulation time of large unilamellar liposomes (LUVs) was examined using four different distearoyl N-(monomethoxy polyethyleneglycol succinyl) phosphatidylethanolamines (DSPE-PEGs). The molecular weights tested were 1000, 2000, 5000 and 12000. Incorporation of 6 mol% of DSPE-PEG in LUV composed of distearoylphosphatidylcholine (DSPC) / cholesterol (CH) (1:1 in molar ratio) increased the blood circulation half-life significantly more than those without DSPE-PEG derivatives. DSPE-PEGs with molecular weights of 1000 and 2000 prolonged the circulation time of liposomes more than other DSPE-PEGs with higher molecular weights, such as 5000 and 12000. Their effects are also higher than ganglioside GM1, a well described glycolipid with this effect. DSPC/CH LUV-incorporated DSPE-PEG with a molecular weight of 2000 displayed a high concentration in the blood, approximately 40% of the dose, 6 h after the injection.