Fuqiang Hu

Synthesis and antitumor activity of doxorubicin conjugated stearic acid-g-chitosan oligosaccharide polymeric micelles

Doxorubicin conjugated stearic acid-g-chitosan oligosaccharide polymeric micelles (DOX-CSO-SA) was synthesized via cis-aconityl bond between the anticancer drug doxorubicin (DOX) and stearic acid grafted chitosan oligosaccharide (CSO-SA) in this paper. The CSO-SA micelles had been demonstrated faster internalization ability into tumor cells. Here, the CSO-SA with 6.47% amino substituted degree (SD%) was used to synthesize DOX-CSO-SA. The critical micelle concentration (CMC) was about 0.14 mg mL(-1). The micelles with 1 mg mL(-1) CSO-SA concentration had 32.7 nm number average diameter with a narrow size distribution and 51.5 mV surface potential. After conjugating with doxorubicin, CMC of DOX-CSO-SA descended; the micellar size increased; and the zeta potential decreased. The DOX-CSO-SA micelles indicated pH-dependent DOX release behavior. The release rate of DOX from DOX-CSO-SA micelles increased significantly with the reductions of the pH for release medium from 7.2 to 5.0. In vitro antitumor activity tests of DOX-CSO-SA micelles against human breast carcinoma (MCF-7) cells and their multi-drug resistant (MCF-7/Adr) cells presented the reversal activity against DOX resistance MCF-7 cells (MCF-7/Adr). The in vivo antitumor activity results showed that DOX-CSO-SA micelles treatments effectively suppressed the tumor growth and reduced the toxicity against animal body than commercial doxorubicin hydrochloride injection.

Enhanced bioavailability of the poorly water-soluble drug fenofibrate by using liposomes containing a bile salt.

The main purpose of this study was to evaluate oral bioavailability of the poorly water-soluble drug fenofibrate when liposomes containing a bile salt were used as oral drug delivery systems. Liposomes composed of soybean phosphotidylcholine (SPC) and sodium deoxycholate (SDC) were prepared by a dry-film dispersing method coupled with sonication and homogenization. Several properties of the liposomes, including particle size, entrapment efficiency and membrane fluidity, were extensively characterized. In vitro release experiments indicated that no more than 20% of total fenofibrate was released from SPC/cholesterol (CL) and SPC/SDC liposomes at 2 h, in contrast with near complete release for micronized fenofibrate capsules. Strikingly, in vivo measurements of pharmacokinetics and bioavailability demonstrated higher rates of fenofibrate absorption from both SPC/SDC and SPC/CL liposomes than micronized fenofibrate. The bioavailability of SPC/SDC and SPC/CL liposomes was 5.13- and 3.28-fold higher, respectively, than that of the micronized fenofibrate. The disparity between oral bioavailability and in vitro release for liposomes strongly suggests alternative absorption mechanisms rather than enhanced release. Importantly, SPC/SDC liposomes exhibited a 1.57-fold increase in bioavailability relative to SPC/CL liposomes, indicating that liposomes containing bile salts may be used to enhance oral bioavailability of poorly water-soluble drugs.

Food protein-stabilized nanoemulsions as potential delivery systems for poorly water-soluble drugs: preparation, in vitro characterization, and pharmacokinetics in rats

Nanoemulsions stabilized by traditional emulsifiers raise toxicological concerns for long-term treatment. The present work investigates the potential of food proteins as safer stabilizers for nanoemulsions to deliver hydrophobic drugs. Nanoemulsions stabilized by food proteins (soybean protein isolate, whey protein isolate, β-lactoglobulin) were prepared by high-pressure homogenization. The toxicity of the nanoemulsions was tested in Caco-2 cells using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide viability assay. In vivo absorption in rats was also evaluated. Food protein-stabilized nanoemulsions, with small particle size and good size distribution, exhibited better stability and biocompatibility compared with nanoemulsions stabilized by traditional emulsifiers. Moreover, β-lactoglobulin had a better emulsifying capacity and biocompatibility than the other two food proteins. The pancreatic degradation of the proteins accelerated drug release. It is concluded that an oil/water nanoemulsion system with good biocompatibility can be prepared by using food proteins as emulsifiers, allowing better and more rapid absorption of lipophilic drugs.

Reversal activity of nanostructured lipid carriers loading cytotoxic drug in multi-drug resistant cancer cells.

To overcome multi-drug resistance (MDR) of cancer cells, paclitaxel (PTX) and doxorubicin (DOX)-loaded nanostructured lipid carriers (NLC) were prepared by solvent diffusion method using monostearin as solid lipid and oleic acid as liquid lipid matrix. The cytotoxicities and reversal activity of drug-loaded NLC were tested against human breast cancer (MCF-7) cells, human ovarian cancer (SKOV3) cells and their multi-drug resistant (MCF-7/ADR and SKOV3-TR30) cells. The chemical conjugant of folic acid and stearic acid (FA-SA) was further synthesized to prepare folated NLC. Comparing with taxol and doxorubicin solution, the NLC loading PTX exhibited high cytotoxicities in MCF-7 and MCF-7/ADR cells, while the NLC loading DOX only indicated high cytotoxicity in MCF-7/ADR cells. The reversal powers of the NLC loading PTX and DOX were 34.3 and 6.4 folds, respectively. The NLC loading PTX and DOX showed the same trends of enhanced cytotoxicity against SKOV3 and SKOV3-TR30 cells. The reversal powers were 31.3 and 2.2 folds for the NLC loading PTX and DOX, respectively. The modification of NLC with FA-SA could further enhance the cytotoxicities of drug in drug sensitive and drug resistant cells.

Preparation and characteristics of linoleic acid-grafted chitosan oligosaccharide micelles as a carrier for doxorubicin

The linoleic acid (LA)-grafted chitosan oligosaccharide (CSO) (CSO-LA) was synthesized in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), and the effects of molecular weight of CSO and the charged amount of LA on the physicochemical properties of CSO-LA were investigated, such as CMC, graft ratio, size, zeta potential. The results showed that these chitosan derivatives were able to self-assemble and form spherical shape polymeric micelles with the size range of 150.7-213.9nm and the zeta potential range of 57.9-79.9mV, depending on molecular weight of CSO and the charged amount of LA. Using doxorubicin (DOX) as a model drug, the DOX-loaded CSO-LA micelles were prepared by dialysis method. The drug encapsulation efficiencies (EE) of DOX-loaded CSO-LA micelles were as high as about 75%. The sizes of DOX-loaded CSO-LA micelles with 20% charged DOX (relating the mass of CSO-LA) were near 200nm, and the drug loading (DL) capacity could reach up to 15%. The in vitro release studies indicated that the drug release from the DOX-loaded CSO-LA micelles was reduced with increasing the graft ratio of CSO-LA, due to the enhanced hydrophobic interaction between hydrophobic drug and hydrophobic segments of CSO-LA. Moreover, the drug release rate from CSO-LA micelles was faster with the drug loading. These data suggested the possible utilization of the amphiphilic micellar chitosan derivatives as carriers for hydrophobic drugs for improving their delivery and release properties.

Improved Transport and Absorption through Gastrointestinal Tract by PEGylated Solid Lipid Nanoparticles

The aim of the present study was to evaluate the potential of PEGylated solid lipid nanoparticle (pSLN) as mucus penetrating particles (MPP) for oral delivery across gastrointestinal mucus. The SLN was prepared by an aqueous solvent diffusion method, subsequently modified with PEG2000-stearic acid (PEG2000-SA) as hydrophilic groups. Surface properties, cytotoxicity, cellular uptake, and transport across Caco-2/HT29 coculture cell monolayers, intestinal absorption, and pharmacokinetics of pSLN were studied compared with that of SLN. Quantitative cellular uptake showed that the internalization of SLN and pSLN was an active transfer process, which would be restrained by several inhibitors of cell activity. Compared with SLN, the permeation ability of pSLN decreased through Caco-2 cell monolayer while it increased through a mucus-secreting Caco-2/HT29 coculture cell monolayer, which indicated that the mucus layer has a significant impact on determining the efficiency of oral nanoformulations. In addition to increasing permeation ability, the stability of the nanoparticles in simulated intestinal fluids was also increased by the PEGylation. Moreover, in vitro everted gut sac technique and the ligated intestinal loops model in vivo also demonstrated that pSLN can rapidly penetrate mucus secretions, whereas the SLN were strongly trapped by highly viscoelastic mucus barriers. The pharmacokinetic studies presented that pSLN exhibited improved absorption efficiency and prolonged blood circulation times with a 1.99-fold higher relative bioavailability compared with SLN. In conclusion, PEGylated solid lipid nanoparticles had advantages in enhancing the bioavailability of oral administration.

PEGylated chitosan-based polymer micelle as an intracellular delivery carrier for anti-tumor targeting therapy.

Stearic acid-grafted chitosan oligosaccharide (CSO-SA) micelles presented a potential candidate for intracellular drug delivery carrier due to its special spatial structure. In this article, CSO-SA was further modified by polyethylene glycol (PEG). The physicochemical properties of PEGylated CSO-SA (PEG-CSO-SA) micelles were characterized. After PEGylation, the critical micelle concentration (CMC) of PEG-CSO-SA had no significant change; the micelle size increased; and the zeta potential decreased. The cellular uptake of CSO-SA micelles before and after PEGylation in macrophage RAW264.7, immortalized rat liver cells BRL-3A and human liver tumor cells HepG2 was studied. About 58.4+/-0.63% of CSO-SA micelles were uptaked by RAW264.7 in 24h, however, only 17.7+/-0.94% of PEG-CSO-SA micelles were internalized into RAW264.7 after the CSO-SA was modified with PEG in five molar times. Meanwhile, there were no changes in the uptake after PEGylation of CSO-SA in BRL-3A and HepG2. Using mitomycin C as a model drug, the in vitro anti-tumor activities of the drug loaded in the micelles were investigated. The 50% cellular growth inhibition (IC(50)) of the drug decreased from 1.97+/-0.2 to 0.13+/-0.02mug/mL after mitomycin C was loaded into CSO-SA micelles, and the IC(50) value of the drug had no obvious change when the CSO-SA was modified by PEG.

Pharmacokinetics and enhanced oral bioavailability in beagle dogs of cyclosporine A encapsulated in glyceryl monooleate/poloxamer 407 cubic nanoparticles

Efforts to improve the oral bioavailability of cyclosporine A (CyA) remains a challenge in the field of drug delivery. In this study, glyceryl monooleate (GMO)/poloxamer 407 cubic nanoparticles were evaluated as potential vehicles to improve the oral bioavailability of CyA. Cubic nanoparticles were prepared via the fragmentation of a bulk GMO/poloxamer 407 cubic phase gel by sonication and homogenization. The cubic inner structure formed was verified using Cryo-TEM. The mean diameters of the nanoparticles were about 180 nm, and the entrapment efficiency of these particles for CyA was over 85%. The in vitro release of CyA from these nanoparticles was less than 5% at 12 h. The results of a pharmacokinetic study in beagle dogs showed improved absorption of CyA from cubic nanoparticles as compared to microemulsion-based Neoral®; higher Cmax (1371.18 ± 37.34 vs 969.68 ± 176.3 ng mL−1), higher AUC0–t (7757.21 ± 1093.64 vs 4739.52 ± 806.30 ng h mL−1) and AUC0–∞ (9004.77 ± 1090.38 vs 5462.31 ± 930.76 ng h mL−1). The relative oral bioavailability of CyA cubic nanoparticles calculated on the basis of AUC0–∞ was about 178% as compared to Neoral®. The enhanced bioavailability of CyA is likely due to facilitated absorption by cubic nanoparticles rather than improved release.

Studies on PEG-modified SLNs loading vinorelbine bitartrate (I): Preparation and evaluation in vitro

In this study, the conjugate of PEG2000-stearic acid (PEG2000-SA) was used to prepare PEGylated solid lipid nanoparticles loading vinorelbine bitartrate (VB-pSLNs) by cold homogenization technique. The particle size and zeta potential of resulted VB-pSLNs ranged 180-250nm and 0-10mV, which were determined using a Zetasizer, respectively. Although the drug entrapment efficiency (EE) slightly decreased after the PEG modification of VB-SLNs, above 60 % EE could be reached. The drug release tests in vitro indicated the faster drug release from VB-pSLNs than that from VB-SLNs without PEG modification. To investigate the cellular uptake of VB-pSLNs, the chemical conjugate of octadecylamine-fluorescein isothiocynate (FITC-ODA) was synthesized, and was used as a fluorescence marker to incorporate into nanoparticles. The results from cellular uptake indicated that the phagocytosis of VB-pSLNs by RAW264.7 cells was inhibited effectively by the PEG modification of SLNs, while the uptake by cancer cells (MCF-7 and A549) could be improved significantly. The assay of anticancer activity in vitro demonstrated that the anticancer activity of VB was significantly enhanced by the encapsulation of SLNs and pSLNs due to the increased cellular internalization of drug. The results suggested that SLNs and pSLNs could be excellent carrier candidates to entrap VB for tumor chemotherapeutics.

Stearic acid-g-chitosan polymeric micelle for oral drug delivery: in vitro transport and in vivo absorption.

Stearic acid-g-chitosan (low molecular weight chitosan CS-SA) with different amino-substituted degrees was synthesized and evaluated as an oral delivery vehicle in this paper. Synthesized CS-SA with 4.47%, 24.36% and 40.36% amino-substituted degree (SD) could form micelles by self-aggregation in aqueous medium. The critical micelle concentration (CMC) ranged from about 0.16 to 0.25 mg/mL, which decreased with the increased SD of CS-SA. The CS-SA micelles had 33.4-130.9 nm size and 22.9- 48.4 mV zeta potential. CS-SA with higher SD had the smaller size and the higher zeta potential. The permeability and possible transport route of CS-SA micelles across the gastrointestinal tract was investigated by in vitro model Caco-2 cells. The results exhibited that the CS-SA micelles had good permeability, and the permeability enhanced with increasing SD of the CS-SA. The transport of the micelles showed energy, pH and concentration dependent transcytosis process, mainly through macropinocytosis and partly via fluid-phase transcytosis and caveolar route. The reversible decrease in transepithelial electrical resistance (TEER) by treatment of micelles suggested that paracellular transport pathway was another route of the micelles crossing the gastrointestinal tract. Using doxorubicin (DOX) as a model drug, the permeation results further demonstrated that the DOX transport mediated by CS-SA micelles could avoid efflux via P-glycoprotein. In vivo study demonstrated that the micelles could significantly improve the bioavailability of encapsulated drug. The results presented that the CS-SA with higher SD was a promising vehicle for oral drugs.