Self-assembled micelles of biodegradable triblock copolymers based on poly(ethyl ethylene phosphate) and poly(-caprolactone) as drug carriers

A series of novel amphiphilic triblock copolymers of poly(ethyl ethylene phosphate) and poly(-caprolactone) (PEEP-PCL-PEEP) with various PEEP and PCL block lengths were synthesized and characterized. These triblock copolymers formed micelles composed of a hydrophobic core of poly(-caprolactone) (PCL) and a hydrophilic shell of poly(ethyl ethylene phosphate) (PEEP) in aqueous solution. The micelle morphology was spherical, determined by transmission electron microscopy. It was found that the size and critical micelle concentration values of the micelles depended on both hydrophobic PCL block length and PEEP hydrophilic block length. The in vitro degradation characteristics of the triblock copolymers were investigated in micellar form, showing that these copolymers were completely biodegradable under enzymatic catalysis of Pseudomonas lipase and phosphodiesterase I. These triblock copolymers were used for paclitaxel (PTX) encapsulation to demonstrate the potential in drug delivery. PTX was successfully loaded into the micelles, and the in vitro release profile was found to be correlative to the polymer composition. These biodegradable triblock copolymer micelles are potential as novel carriers for hydrophobic drug delivery.     

Enhanced stability of core-surface cross-linked micelles fabricated from amphiphilic brush copolymers

"Stealth" nanoparticles made from polymer micelles have been widely explored as drug carriers for targeted drug delivery. High stability (i.e., low critical micelle concentration (CMC)) is required for their intravenous applications. Herein, we present a "core-surface cross-linking" concept to greatly enhance nanoparticle's stability: amphiphilic brush copolymers form core-surface cross-linked micelles (nanoparticles) (SCNs). The amphiphilic brush copolymers consisted of hydrophobic poly(epsilon-caprolactone) (PCL) and hydrophilic poly(ethylene glycol) (PEG) or poly(2-(N,N-dimethylamino)ethyl methacrylate) (PDMA) chains were synthesized by macromonomer copolymerization method and used to demonstrate this concept. The resulting SCNs were about 100 times more stable than micelles from corresponding amphiphilic block copolymers. The size and surface properties of the SCNs could be easily tailored by the copolymer's compositions.     

Influence of LA and GA sequence in the PLGA block on the properties of thermogelling PLGA-PEG-PLGA block copolymers

This paper reports the influence of sequence structures of block copolymers composed of poly(lactic acid-co-glycolic acid) (PLGA) and poly(ethylene glycol) (PEG) on their thermogelling aqueous behaviors. A series of thermogelling PLGA-PEG-PLGA triblock copolymers with similar chemical compositions and block lengths but different sequences of D,L-lactide (LA) and glycolide (GA) in the PLGA block were synthesized. The difference of sequence structures arises from the different reactivities of LA and GA during the copolymerization and the transesterification after polymerization. The sol-gel transition temperature and height of gel window were found to be regulated by the sequence structure. Our study reveals that the macromolecular sequence structure influences the hydrophobic/hydrophilic balance of this kind of amphiphilic copolymers and thus alters mesoscopic micellization and the forthcoming macroscopic physical gelation in water. This finding might be helpful to guide the molecular design of the underlying thermogelling systems as injectable hydrogels.     

Osteogenic differentiation of human placenta-derived mesenchymal stem cells (PMSCs) on electrospun nanofiber meshes

Numerous challenges remain in the successful clinical translation of cell-based therapeutic studies for skeletal tissue repair, including appropriate cell sources and viable cell delivery systems. Poly(ethylene glycol)-poly(ε-caprolactone) (PEG-PCL) amphiphilic block copolymers have been extensively explored in microspheres preparation. Due to the introduction of hydrophilic PEG segments into PCL backbones, these copolymers have shown much more potentials in carrying protein, lipophilic drugs or genes than commonly used poly (ε-caprolactone) (PCL) and poly (lactic acid). The aim of this study is to investigate the attachment and osteogenic differentiation of human placenta derived mesenchymal stem cells (PMSCs) on PEG-PCL triblock copolymers nanofiber scaffolds. Here we demonstrated that PMSCs proliferate robustly and can be effectively differentiated into osteogenic-like cells on nanofiber scaffolds. This study provides evidence for the use of nanofiber scaffolds as an ideal supporting material for in vitro PMSCs culture and an in vivo cell delivery vehicle for bone repair.     

Lipase-catalyzed transformation of poly(butylene adipate) and poly(butylene succinate) into repolymerizable cyclic oligomers

The enzymatic degradation and repolymerization were carried out with the objectives of developing the chemical recycling of aliphatic polyester-type plastics, such as the poly(butylene adipate) (PBA), poly(butylene succinate), and poly(butylene adipate-co-succinate) copolymers which are typical biodegradable plastics. They were degraded by lipase in an organic solvent solution containing a small amount of water to produce cyclic oligomers mainly consisting of the cyclic diester. The produced cyclic oligomer was readily repolymerized by lipase to produce a polyester having an equal or higher molecular weight compared to the parent polymer. As an example, PBA having an Mw of 22,000 was almost quantitatively transformed by lipase CA (Novozym 435) in water-containing toluene at 50 degrees C into the corresponding cyclic oligomers mainly consisting of dimers. Thus, the obtained oligomers were readily polymerized by lipase CA to produce the PBA with an Mw of 52,000.     

Synthesis and characterization of star poly(epsilon-caprolactone)-b-poly(ethylene glycol) and poly(L-lactide)-b-poly(ethylene glycol) copolymers: evaluation as drug delivery carriers

Two types of 32 arm star polymers incorporating amphiphilic block copolymer arms have been synthesized and characterized. The first type, stPCL-PEG 32, is composed of a polyamidoamine (PAMAM) dendrimer as the core with radiating arms having poly(epsilon-caprolactone) (PCL) as an inner lipophilic block in the arm and poly(ethylene glycol) (PEG) as an outer hydrophilic block. The second type, stPLA-PEG 32, is similar but with poly(L-lactide) (PLA) as the inner lipophilic block. Characterization with SEC, (1)H NMR, FTIR, and DSC confirmed the structure of the polymers. Micelle formation by both star copolymers was studied by fluorescence spectroscopy. The stPCL-PEG 32 polymer exhibited unimolecular micelle behavior. It was capable of solubilizing hydrophobic molecules, such as pyrene, in aqueous solution, while not displaying a critical micelle concentration. In contrast, the association behavior of stPLA-PEG 32 in aqueous solution was characterized by an apparent critical micelle concentration of ca. 0.01 mg/mL. The hydrophobic anticancer drug etoposide can be encapsulated in the micelles formed from both polymers. Overall, the stPCL-PEG 32 polymer exhibited a higher etoposide loading capacity (up to 7.8 w/w % versus 4.3 w/w % for stPLA-PEG 32) as well as facile release kinetics and is more suitable as a potential drug delivery carrier.     

Polymeric micelles with water-insoluble drug as hydrophobic moiety for drug delivery

The hydrophobic block of polymeric micelles formed by amphiphilic copolymers has no direct therapeutical effect, and the metabolites of these hydrophobic segments might lead to some unexpected side effects. Here the hydrophobic core of polymeric micelles is replaced by highly water-insoluble drugs themselves, forming a new micellar drug delivery system. By grafting hydrophobic drugs of paclitaxel (PTX) onto the surface of hydrophilic hyperbranched poly(ether-ester) (HPEE), we constructed an amphiphilic copolymer (HPEE-PTX). HPEE-PTX could self-assemble into micellar nanoparticles in aqueous solution with tunable drug contents from 4.1 to 10.7%. Moreover, the hydrolysis of HPEE-PTX in serum resulted in the cumulative release of PTX. In vivo evaluation indicated that the dosage toleration of PTX in mice had been improved greatly and HPEE-PTX micellar nanoparticles could be used as an efficient prodrug with satisfactory therapeutical effect. We believe that most of the lipophilic drugs could improve their characters through this strategy.     

Synthesis and self-assembly of chitosan-based copolymer with a pair of hydrophobic/hydrophilic grafts of polycaprolactone and poly(ethylene glycol)

Chitosan-based copolymers with binary grafts of hydrophobic polycaprolactone and hydrophilic poly(ethylene glycol) (CS-g-PCL&PEG) were prepared by a homogeneous coupling reaction of phthaloyl-protected chitosan with functional PCL-COOH and PEG-COOH, following deprotection to regenerate free amino groups back to chitosan backbone. They were characterized by ¹H NMR, Fourier transform infrared and X-ray diffraction analysis. These CS-g-PCL&PEG copolymers could form nano-size self-aggregates in acidic aqueous solution without a specific processing technique, which were investigated using dynamic light scattering and transmission electron microscopy. The formed self-aggregates become smaller with weakened stability upon pH increasing. Moreover, the aggregates of copolymer with higher content of PEG and PCL grafts could remain stable for over 30 days in both acid and neutral condition. A possible mechanism was proposed for the formation of self-aggregates from CS-g-PCL&PEG and their structural changes as pH. It is warranted to find promising application of these self-aggregates based on chitosan as drug carriers.     

Synthesis and evaluation of a star amphiphilic block copolymer from poly(epsilon-caprolactone) and poly(ethylene glycol) as a potential drug delivery carrier

A star polymer composed of amphiphilic block copolymer arms has been synthesized and characterized. The core of the star polymer is polyamidoamine (PAMAM) dendrimer, the inner block in the arm is lipophilic poly(epsilon-caprolactone) (PCL), and the outer block in the arm is hydrophilic poly(ethylene glycol) (PEG). The star-PCL polymer was synthesized first by ring-opening polymerization of epsilon-caprolactone with a PAMAM-OH dendrimer as initiator. The PEG polymer was then attached to the PCL terminus by an ester-forming reaction. Characterization with SEC, (1)H NMR, FTIR, TGA, and DSC confirmed the star structure of the polymers. The micelle formation of the star copolymer (star-PCL-PEG) was studied by fluorescence spectroscopy. Hydrophobic dyes and drugs can be encapsulated in the micelles. A loading capacity of up to 22% (w/w) was achieved with etoposide, a hydrophobic anticancer drug. A cytotoxicity assay demonstrated that the star-PCL-PEG copolymer is nontoxic in cell culture. This type of block copolymer can be used as a drug delivery carrier.     

A coarse-grain MD (molecular dynamic) simulation of PCL–PEG and PLA–PEG aggregation as a computational model for prediction of the drug-loading efficacy of doxorubicin

Abstract

Formulating a hydrophobic drug that is water-soluble is a pharmaceutical challenge. One way is to incorporate the drug in an amphiphilic micelle composed from an aggregation of block copolymers. Design of a good nano-micelle requires many trial-and-error attempts. In this article, we developed a computational model based on a coarse-grained molecular dynamic (MD) simulation and correlated outputs with previous studies. A good correlation shows that this model reliably simulates poly-lactic acid–poly-ethylene glycol (PLA–PEG) and poly-caprolactone (PCL)–PEG aggregation in water with and without the presence of doxorubicin.

Communicated by Ramaswamy H. Sarma     

Hydrophobic drug delivery by self-assembling triblock copolymer-derived nanospheres

We describe the synthesis and characterization of a family of biocompatible ABA-triblock copolymers that comprised of hydrophilic A-blocks of poly(ethylene glycol) and hydrophobic B-blocks of oligomers of suberic acid and desaminotyrosyl-tyrosine esters. The triblock copolymers spontaneously self-assemble in aqueous solution into nanospheres, with hydrodynamic diameters between 40 and 70 nm, that do not dissociate under chromatographic and ultracentrifugation conditions. These nanospheres form strong complexes with hydrophobic molecules, including the fluorescent dye 5-dodecanoylaminofluorescein (DAF) and the antitumor drug, paclitaxel, but not with hydrophilic molecules such as fluorescein and Oregon Green. The nanosphere-paclitaxel complexes retain in vitro the high antiproliferative activity of paclitaxel, demonstrating that these nanospheres may be useful for delivery of the hydrophobic drugs.     

Organocatalytic approach to amphiphilic comb-block copolymers capable of stereocomplexation and self-assembly

Biocompatible amphiphilic block copolymers comprised of poly(ethylene glycol) (PEG) as the hydrophilic component and a poly(methylcarboxytrimethylene carbonate) (PMTC) as a hydrophobic backbone having either poly(L-lactide) (L-PLA) or poly(D-lactide) (D-PLA) branches were prepared by organocatalytic ring-opening polymerization (ROP). The polycarbonate backbone was prepared by copolymerization of two different MTC-type monomers (MTCs) including a tetrahydropyranyloxy protected hydroxyl group, a masked initiator for a subsequent ROP step. Interestingly, the organic catalyst used in the ROP of MTCs was also effective for acetylation of the hydroxyl end-groups by the addition of acetic anhydride added after polymerization. Acidic deprotection of the tetrahydropyranyloxy (THP) protecting group on the carbonate chain generated hydroxyl functional groups that served as initiators for the ROP of either D- or L-lactide. Comb-shaped block copolymers of predictable molecular weights and narrow polydispersities (approximately 1.3) were prepared with up to 8-PLA branches. Mixtures of the D- and L-lactide based copolymers were studied to understand the effect of noncovalent interactions or stereocomplexation on the properties.     

Cocrystallization model for synthetic biodegradable poly(butylene adipate-co-butylene terephthalate)

Synthetic biodegradable poly(butylene adipate-co-butylene terephthalate), P(BA-co-BT), with 56 mol % butylene adipate, BA, was characterized by solid-state NMR spectroscopy, thermal analysis, X-ray diffraction, computer modeling, and polarization microscopy. The NMR study showed the presence of BA and butylene terephthalate, BT. T(1C) NMR measurements proved that some BA and BT units were in crystalline regions. Thermal analysis showed one glass-transition temperature and a single diffuse melting endotherm corresponding to a large melting-point depression of about 100 degrees C compared with poly(butylene terephthalate), PBT. These results suggest that there is only one crystalline phase. An X-ray fiber diagram of a stretched film could be indexed with the same unit cell as that for PBT. Computer modeling showed that the adipate unit fits into the crystal structure of PBT by adopting a TTGTG dihedral angle sequence in the crystalline conformation proposed for the cocrystallization model. The predicted fiber diagram from the proposed model qualitatively agrees with the experimental one. Polarization microscopy revealed that the spherulite growth rate of P(BA-co-BT) was similar to that for poly(butylene adipate), PBA.     

Synthesis and in vitro characterization of an ABC triblock copolymer for siRNA delivery

The ability to specifically down-regulate gene expression using the RNAi pathway in mammalian cells has tremendous potential in therapy and in basic science. However, delivery systems capable of efficient and biocompatible delivery of siRNA to target cells are not yet satisfactory. Here, we report the synthesis and in vitro characterization of ABC triblock copolymers that self-assemble with siRNA based on electrostatics and with each other by hydrophobic interactions. The ABC triblock copolymer is based on poly(ethylene glycol) (PEG), poly(propylene sulfide) (PPS), and a positively charged peptide (PEG-PPS-peptide). The diblock copolymer PEG(45)-PPS(5,10) was synthesized using anionic polymerization of propylene sulfide upon a PEG macroinitiator, and the peptide domain was coupled to the PPS terminus using a disulfide exchange reaction with an N-terminal cysteine residue on the peptide. The peptides were designed to interact electrostatically with siRNA, selecting the TAT peptide domain of HIV (RKKRRQRRR) and an oligolysine (Lys(9)). The resulting triblock copolymers were able to self-assemble with siRNA as demonstrated by dynamic light scattering and gel electrophoresis. Complex size was found to be dependent on the amount of polymer used (charge ratio) and the length of the hydrophobic PPS block, achieving sizes ranging from 171 nm to 601 nm. Cell internalization and gene expression down-regulation studies showed that the triblock copolymers are able to transport siRNA inside the cell and mediate gene expression down-regulation, with the amount of internalization and gene transfer affected by charge ratio, PPS length, and the presence of serum. The proposed triblock was able to mediate gene expression down-regulation of GAPDH, achieving up to 90.5% +/- 0.02% down-regulation.     

Effect of tyrosine-derived triblock copolymer compositions on nanosphere self-assembly and drug delivery

We have obtained structure-activity relations for nanosphere drug delivery as a function of the chemical properties of a tunable family of self-assembling triblock copolymers. These block copolymers are synthesized with hydrophobic oligomers of a desaminotyrosyl tyrosine ester and diacid and hydrophilic poly(ethylene glycol). We have calculated the thermodynamic interaction parameters for the copolymers with anti-tumor drugs to provide an understanding of the drug binding by the nanospheres. We find that there is an optimum ester chain length, C8, for nanospheres in terms of their drug loading capacities. The nanospheres release the drugs under dialysis conditions, with release rates strongly influenced by solution pH. The nanospheres, which are themselves non-cytotoxic, deliver the hydrophobic drugs very effectively to tumor cells as measured by cell killing activity in vitro and thus offer the potential for effective parentarel in vivo delivery of many hydrophobic therapeutic agents.     

Oxime linkage: a robust tool for the design of pH-sensitive polymeric drug carriers

Oxime bonds dispersed in the backbones of the synthetic polymers, while young in the current spectrum of the biomedical application, are rapidly extending into their own niche. In the present work, oxime linkages were confirmed to be a robust tool for the design of pH-sensitive polymeric drug delivery systems. The triblock copolymer (PEG-OPCL-PEG) consisting of hydrophilic poly(ethylene glycol) (PEG) and hydrophobic oxime-tethered polycaprolactone (OPCL) was successfully prepared by aminooxy terminals of OPCL ligating with aldehyde-terminated PEG (PEG-CHO). Owing to its amphiphilic architecture, PEG-OPCL-PEG self-assembled into the micelles in aqueous media, validated by the measurement of critical micelle concentration (CMC). The MTT assay showed that PEG-OPCL-PEG exhibited low cytotoxicity against NIH/3T3 normal cells. Doxorubicin (DOX) as a model drug was encapsulated into the PEG-OPCL-PEG micelles. Drug release study revealed that the DOX release from micelles was significantly accelerated at mildly acid pH of 5.0 compared to physiological pH of 7.4, suggesting the pH-responsive feature of the drug delivery systems with oxime linkages. Flow cytometry and confocal laser scanning microscopy (CLSM) measurements indicated that these DOX-loaded micelles were easily internalized by living cells. MTT assay against HeLa cancer cells showed DOX-loaded PEG-OPCL-PEG micelles had a high anticancer efficacy. All of these results demonstrate that these polymeric micelles self-assembled from oxime-tethered block copolymers are promising carriers for the pH-triggered intracellular delivery of hydrophobic anticancer drugs.     

Synthesis, characterization, and morphology studies of biodegradable amphiphilic poly[(R)-3-hydroxybutyrate]-alt-poly(ethylene glycol) multiblock copolymers

Novel biodegradable amphiphilic alternating block copolymers based on poly[(R)-3-hydroxybutyrate] (PHB) as biodegradable and hydrophobic block and poly(ethylene glycol) (PEG) as hydrophilic block (PHB-alt-PEG) were successfully synthesized through coupling reaction. Their chemical structures have been characterized by using gel permeation chromatography, (1)H nuclear magnetic resonance, and Fourier transform infrared spectroscopy. Differential scanning calorimetry (DSC) analysis revealed that both PHB and PEG blocks in PHB-alt-PEG block copolymers can crystallize to form separate crystalline phase except in those with a short PEG block (M(n) 600) only PHB crystalline phase has been observed. However, due to the mutual interference from each other, the melting transition of both PHB and PEG crystalline phases shifted to lower temperature with lower crystallinity in comparison with corresponding pure PHB and PEG. The crystallization behavior of PHB block and PEG block has also been studied by X-ray diffraction, and the results were in good agreement with those deduced from DSC study. The surface morphologies of PHB-alt-PEG block copolymer thin films spin-coated on mica have been visualized by atomic force microscopy with tapping mode, indicating formation of laterally regular lamellar surface patterns. Static water contact angle measurement revealed that surface hydrophilicity of these spin-coated thin films increases with increasing PEG block content.     

Functionalized amphiphilic hyperbranched polymers for targeted drug delivery

Amphiphilic hyperbranched core-shell polymers with folate moieties as the targeting groups were synthesized and characterized. The core of the amphiphilic polymers was hyperbranched aliphatic polyester Boltorn H40. The inner part and the outer shell of the amphiphilic polymers were composed of hydrophobic poly(epsilon-caprolactone) segments and hydrophilic poly(ethylene glycol) (PEG) segments, respectively. To achieve tumor cell targeting property, folic acid was further incorporated to the surface of the amphiphilic polymers via a coupling reaction between the hydroxyl group of the PEG segment and the carboxyl group of folic acid. The polymers were characterized by (1)H NMR, (13)C NMR, and combined size-exclusion chromatography and multiangle laser light scattering analysis. The nanoparticles of the amphiphilic polymers prepared by dialysis method were characterized by transmission electron microscopy and particle size analysis. Two antineoplastic drugs, 5-fluorouracil and paclitaxel, were encapsulated into the nanoparticles. The drug release property and the targeting of the drug-loaded nanoparticles to different cells were evaluated in vitro. The results showed the drug-loaded nanoparticles exhibited enhanced cell inhibition because folate targeting increased the cytotoxicity of drug-loaded nanoparticles against folate receptor expressing tumor cells.     

Core-cross-linked polymeric micelles as paclitaxel carriers

Cross-linkable di- and triblock copolymers of poly(epsilon-caprolactone) (PCL) and monomethoxyl poly(ethylene glycol) (MPEG) were synthesized. These amphiphilic copolymers self-assembled into nanoscale micelles capable of encapsulating hydrophobic paclitaxel in their hydrophobic cores in aqueous solutions. To further enhance their thermodynamic stability, the micelles were cross-linked by radical polymerization of the double bonds introduced into the PCL blocks. Reaction conditions were found to significantly affect both the cross-linking efficiency and the micelle size. The encapsulation of paclitaxel into the micelles was confirmed by the proton nuclear magnetic resonance (1H NMR) spectroscopy. Encouragingly, paclitaxel-loading efficiency of micelles was enhanced significantly upon micelle core-cross-linking. Both the micelle size and the drug loading efficiency increased markedly with increasing the PCL block lengths, no matter if the micelles were core-cross-linked or not. However, paclitaxel-loading did not obviously affect the micelle size or size distribution. The cross-linked micelles exhibited a significantly enhanced thermodynamic stability against dilution with aqueous solvents. The efficient cellular uptake of paclitaxel loaded in the nanomicelles was demonstrated by confocal laser scanning microscopy (CLSM) imaging. This new biodegradable nanoscale carrier system merits further investigations for parenteral drug delivery.     

Synthesis and characterization of biocompatible thermo-responsive gelators based on ABA triblock copolymers

The synthesis of biocompatible, thermo-responsive ABA triblock copolymers in which the outer A blocks comprise poly(N-isopropylacrylamide) and the central B block is poly(2-methacryloyloxyethyl phosphorylcholine) is achieved using atom transfer radical polymerization with a commercially available bifunctional initiator. These novel triblock copolymers are water-soluble in dilute aqueous solution at 20 degrees C and pH 7.4 but form free-standing physical gels at 37 degrees C due to hydrophobic interactions between the poly(N-isopropylacrylamide) blocks. This gelation is reversible, and the gels are believed to contain nanosized micellar domains; this suggests possible applications in drug delivery and tissue engineering.