Poly( -histidine)–PEG block copolymer micelles and pH-induced destabilization

Poly(L-histidine)-poly(ethylene glycol) diblock copolymers (polyHis-b-PEG) were prepared and used for the construction of polymeric micelles responding to local pH changes in the body. PolyHis was synthesized by ring opening polymerization of L-histidine N-carboxyanhydride, the imidazole amine group of which was protected by the dinitrophenyl group. The resulting polymer (M(n): 5,000 g/mole) was coupled to poly(ethylene glycol) (M(n): 2,000 g/mole) via an amide linkage using the dicyclohexyl carbodiimide and N-hydroxysuccinimide-mediated reaction. The block copolymer in dimethyl sulfoxide formed polymeric micelles on diafiltration against a borate buffer at pH 8. Dynamic light scattering and atomic force microscopy showed the micelles were spherical, diameter approximately 114 nm, with a unimodal distribution. The critical micelle concentration (CMC) at pH 8.0 was 2.3 mg/l. The CMC increased markedly on decreasing the pH of the diafiltration medium below 7.2. Micelles prepared at pH 8.0 were gradually destabilized below pH 7.4, as evidenced by a slight increase in light transmittance, an alteration in size distribution, and a decrease in the pyrene fluorescence intensity. It was concluded that the ionization of the polyHis block forming the micelle core determined the pH-dependent CMC and stability. After further optimization of the pH-sensitivity, pH-sensitive micelles are expected to have application for solid tumor treatment, exploiting the fact that most solid tumors have an acidic extracellular pH.     

Mucoadhesive platforms for targeted delivery to the colon

A novel platform system, comprising a mucoadhesive core and a rapid release carrier, was designed for targeted drug delivery to the colon. Prednisolone pellets containing different carbomers, including Carbopol 971P, Carbopol 974P and Polycarbophil AA-1, with or without organic acids, were produced by extrusion-spheronization. Mucoadhesive pellets were coated with a new enteric double-coating system, which dissolves at pH 7. This system comprises an inner layer of partially neutralized Eudragit^® S and buffer salt and an outer coating of standard Eudragit^® S. A single layer of standard Eudragit S was also applied for comparison purposes. Dissolution of the coated pellets was assessed in USP II apparatus in 0.1 N HCl followed by Krebs bicarbonate buffer pH 7.4. Visualization of the coating dissolution process was performed by confocal laser scanning microscopy using fluorescent markers in both layers. The mucoadhesive properties of uncoated, single-coated and-double coated pellets were evaluated ex vivo on porcine colonic mucosa. Mucoadhesive pellets coated with a single layer of Eudragit^® S release its cargo after a lag time of 120 min in Krebs buffer. In contrast, drug release from the double-coated mucoadhesive pellets was significantly accelerated, starting at 75 min. In addition, the mucoadhesive properties of the core of the double coated pellets were higher than those from single-coated pellets after the core had been exposed to the buffer medium. This novel platform technology has the potential to target the colon and overcome the variability in transit and harmonize drug release and bioavailability.     

N-辛基-N’-(2-羧基环己甲酰基)-壳聚糖的制备、表征与pH敏感性

目的以pH敏感的酰胺键为连接臂,制备两亲性壳聚糖衍生物。方法通过在壳聚糖2-NH2上引入疏水辛基和pH敏感的酰胺键,制备两亲性壳聚糖接枝共聚物,用FTIR、1H-NMR和13C-NMR对其结构进行表征,使用XRD、DSC对其物理性质进行分析,采用紫外-可见分光光度计和粒径仪评价其pH敏感性。结果合成了7种取代度的N-辛基-N’-(2-羧基环己甲酰基)-壳聚糖,在pH 5.0~6.0内具有pH敏感性。结论合成的pH敏感衍生物有望用于智能型药物释放系统,在体内特定部位(肿瘤、梗塞)或细胞内(内涵体、细胞质)释放包载的难溶性药物。     

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.     

New amphiphilic and pH-sensitive hydrogel for controlled release of a model poorly water-soluble drug

This paper presents the development of new pH-sensitive, amphiphilic and biocompatible hydrogels based on alginate-g-PCL, cross-linked with calcium ions to form beads, prepared for controlled delivery of poorly water-soluble drug. We have focused our study on the effect of the length of PCL chains (530 and 1250 g mol⁻¹). Swelling profiles obtained clearly indicated that these hydrogels swell slightly (10-14%) in a simulated gastric fluid (pH 1.2), and strongly (700-1300% before disintegration) in a simulated intestinal fluid (pH 6.8). In both media, rates of swelling were lower for beads based on amphiphilic derivatives than for alginate/Ca²⁺ ones due to the hydrophobic PCL grafts, and decreased when hydrophobic character increased. A model drug, theophylline, was entrapped into these hydrogels and release studies were carried out. The drug was protected in acidic fluid (only 14-20% of release for alginate-g-PCL hydrogel against 35% of release for alginate hydrogel during 350 min). The drug is released completely in neutral fluid due to ion exchanges and disintegration of the hydrogel. PCL leads to decrease in the release kinetics in SIF (2 h for alginate-g-PCL/Ca²⁺ beads against 1 h for alginate/Ca²⁺ beads). It was demonstrated that the establishment of clusters inside beads by intramolecular interactions between PCL grafts of 530 g mol⁻¹ in salt media allowed to retain the drug and to slow down its release considerably.     

Chemical vectors for gene delivery: a current review on polymers, peptides and lipids containing histidine or imidazole as nucleic acids carriers

DNA/cationic lipid (lipoplexes), DNA/cationic polymer (polyplexes) and DNA/cationic polymer/cationic lipid (lipopolyplexes) electrostatic complexes are proposed as non-viral nucleic acids delivery systems. These DNA-nanoparticles are taken up by the cells through endocytosis processes, but the low capacity of DNA to escape from endosomes is regarded as the major limitations of their transfection efficiency. Here, we present a current report on a particular class of carriers including the polymers, peptides and lipids, which is based on the exploitation of the imidazole ring as an endosome destabilization device to favour the nucleic acids delivery in the cytosol. The imidazole ring of histidine is a weak base that has the ability to acquire a cationic charge when the pH of the environment drops bellow 6. As it has been demonstrated for poly(histidine), this phenomena can induce membrane fusion and/or membrane permeation in an acidic medium. Moreover, the accumulation of histidine residues inside acidic vesicles can induce a proton sponge effect, which increases their osmolarity and their swelling. The proof of concept has been shown with polylysine partially substituted with histidine residues that has caused a dramatic increase by 3–4.5 orders of magnitude of the transfection efficiency of DNA/polylysine polyplexes. Then, several histidine-rich polymers and peptides as well as lipids with imidazole, imidazolinium or imidazolium polar head have been reported to be efficient carriers to deliver nucleic acids including genes, mRNA or SiRNA in vitro and in vivo. More remarkable, histidylated carriers are often weakly cytotoxic, making them promising chemical vectors for nucleic acids delivery.This article is part of a themed section on Vector Design and Drug Delivery. For a list of all articles in this section see the end of this paper, or visit: http://www3.interscience.wiley.com/journal/121548564/issueyear?year=2009     

Poly(Ethylene Oxide)-Modified Poly(β-Amino Ester) Nanoparticles as a pH-Sensitive System for Tumor-Targeted Delivery of Hydrophobic Drugs: Part 2. <Emphasis Type="BoldItalic">In Vivo</Emphasis> Distribution and Tumor Localization Studies

Purpose This study was carried out to determine the biodistribution profiles and tumor localization potential of poly(ethylene oxide) (PEO)-modified poly(β-amino ester) (PbAE) as a novel, pH-sensitive biodegradable polymeric nanoparticulate system for tumor-targeted drug delivery. Methods The biodistribution studies of PEO-modified PbAE and PEO-modified poly(ɛ-caprolactone) (PCL), a non-pH-sensitive polymer, nanoparticle systems were carried out in normal mice using ¹¹¹indium-oxine [¹¹¹In] as a lipophilic radiolabel encapsulated within the polymeric matrix, and the distribution of the nanoparticles was studied in plasma and all the vital organs following intravenous administration. Solid tumors were developed on nude mice using human ovarian carcinoma xenograft (SKOV-3) and the change in concentrations of tritium [³H]-labeled paclitaxel encapsulated in polymeric nanoparticles was examined in blood, tumor mass, and liver. Results Study in normal mice with a gamma-emitting isotope [¹¹¹In] provided a thorough biodistribution analysis of the PEO-modified nanoparticulate carrier systems, whereas ³H-paclitaxel was useful to understand the change in concentration and tumor localization of anticancer compound directly in major sites of distribution. Both PEO-PbAE and PEO-PCL nanoparticles showed long systemic circulating properties by virtue of surface modification with PEO-containing triblock block copolymer (Pluronic?) stabilizer. Although the PCL nanoparticles showed higher uptake by the reticuloendothelial system, the PbAE nanoparticles effectively delivered the encapsulated payload into the tumor mass. Conclusions PEO-modified PbAE nanoparticles showed considerable passive tumor targeting potential in early stages of biodistribution via the enhanced permeation and retention (EPR) mechanism. This prompts a detailed biodistribution profiling of the nanocarrier for prolonged periods to provide conclusive evidence for superiority of the delivery system.     

Stimulated release of photosensitizers from graft and diblock micelles for photodynamic therapy

To understand the effect of photosensitizer (PS) release from graft copolymer based micelles in photodynamic therapy (PDT), the two pH-sensitive and non-pH-sensitive graft copolymers, (poly(N-vinyly caprolactam)-g-poly(d,l-lactide) and poly(N-vinyly caprolactam-co-N-vinyl imidazole)-g-poly(d,l-lactide)), were synthesized and utilized for the encapsulation of protoporphyrin IX (PPIX) for in vitro and in vivo PDT studies. Photochemical internalization (PCI) was utilized to study the localization of pH- and non-pH-sensitive micelles uptake in the lysosome. After non-toxic light treatment, PPIX was found in the nucleus with pH-sensitive micelles, while PPIX was still localized in the lysosomal organism with the non-pH-sensitive micelles, as observed by confocal microscopy. Because the formation of singlet oxygen was observed for the block and graft micelles, dramatic differences in the cell viability could be ascribed to the damage occurring at the region where the PPIX was located. An in vivo study revealed that PPIX-loaded graft and diblock micelles presented prolonged blood circulation and enhanced tumor targeting ability. The PPIX released from g-CIM micelles on tumor site was further proved by ex vivo confocal image. In addition, non-pH-sensitive micelle-treated mice showed a better repression of tumor growth than PPIX-treated mice, which was likely due to the larger amount of PS localized in the tumor region still exhibiting therapeutic effects. Finally, effective PDT-induced inhibition of tumor growth was found in pH-sensitive micelle-treated mice. This work provides insight into PS-loaded graft and diblock micelles for the PDT of tumors.     

The degradation and biocompatibility of pH-sensitive biodegradable polyurethanes for intracellular multifunctional antitumor drug delivery

To obtain controllable stepwise biodegradable polymer for multifunctional antitumor drug carriers, pH-sensitive biodegradable polyurethanes were firstly synthesized using poly(ε-caprolactone) (PCL) and pH-sensitive poly(ε-caprolactone)-hydrazone-poly(ethylene glycol)-hydrazone-poly(ε-caprolactone) macrodiol (PCLH) as soft segment; l-lysine ethyl ester diisocyanate (LDI), l-lysine derivative tripeptide and 1,4-butandiol (BDO) as hard segment; and hydrazone-linked methoxyl-poly(ethylene glycol)(m-PEG-Hyd) as end-capper. Then, an extensive degradation process of the prepared pH-sensitive polyurethanes was investigated in vitro with proton nuclear magnetic resonance spectra (¹H NMR), gel permeation chromatograph (GPC), scanning electron microscopy (SEM), and weight loss. It was found that the degradation of these polyurethanes occurred via the random hydrolytic ester cleavage along the PCL segments close to PEG segments in enzymatic solutions while the hydrazone bond in the polymer chain was more easily cleaved in acidic media, which was accelerated with decreasing pH value. Furthermore, the biocompatibility in vivo was evaluated in an intramuscular implantation model on Sprague-Dawley rats, using SEM and light microscopy. The result showed that the prepared polyurethanes can be easily degraded and the degradation products do not induce any adverse response from surrounding muscle tissues. Our work suggests that the prepared pH-sensitive polyurethanes could be promising materials as controllable biodegradable and non-cyctotoxic multifunctional carriers for active intracellular drug delivery.     

环孢素A-Eudragit S100纳米粒冻干粉制剂在犬体内的药动学

目的:研究环孢素A-Eudragit S100纳米粒冻干粉制剂(freeze-dried cyclosporine A-Eudragit S100 nanoparticles,CyA-S100-NP)在家犬体内药动学及相对生物利用度。方法:以双周期交叉随机试验设计法,高效液相色谱法测定6只家犬口服给予环孢素A-Eudragit S100纳米粒冻干粉硬胶囊和新山地明微乳软胶囊(Neoral)后环孢素A的血药浓度,采用3P97软件计算药动学参数。结果:经3P97软件拟合,环孢素A的药动学过程符合二室模型。与Neoral相比,CyA-S100-NP的AUC显著增大(P<0.05),CL显著降低(P<0.05),相对生物利用度为135.9%。结论:CyA-S100-NP可促进药物口服吸收,显著提高环孢素A的生物利用度,有望开发成为一种新型口服环孢素A纳米粒固体制剂。