Yuehong Xu

Hepatocellular carcinoma targeting effect of PEGylated liposomes modified with lactoferrin

A hepatocellular carcinoma targeting lactoferrin (Lf) modified PEGylated liposome system was developed for improving drug efficacies to hepatic cancer cells. In this present work, PEGylated liposomes (PLS) were successfully prepared by the thin film hydration method combined with peglipid post insertion. Lf was covalently conjugated to the distal end of DSPE-PEG2000-COOH lipid by amide bound and loaded onto PEGylated liposomes surface as the targeting ligand. To confirm the targeting efficacies to hepatic cancer, coumarin-6 and DiR were encapsulated as fluorescent probes. The confocal microscopy and flow cytometry demonstrated that Lf conjugated PEGylated liposomes (Lf-PLS) were efficiently associated by HepG2 cells, while limited interaction was found for liposomes modified with a negative control protein. A similar pharmacokinetic behavior was observed in pharmacokinetics study of the liposomal formulations. Meanwhile, the in vivo imaging of liposomes in HepG2 tumor bearing mice indicated that Lf-PLS achieved more accumulation in tumor compared with PLS without Lf conjugated. The significant in vitro and in vivo results suggested that Lf-PLS might be a promising drug delivery system for hepatocellular carcinoma therapy with low toxicity.

Percutaneous delivery of econazole using microemulsion as vehicle: Formulation, evaluation and vesicle-skin interaction

This project was carried out to exploit the feasibility of using microemulsion (ME) as an alternative carrier for percutaneous delivery econazole nitrate (ECN) and elucidate the underlying mechanism of permeation enhancement. The ME was developed based on Labrafil M 1944 Cs as oil phase, Solutol HS15 and Span 80 as surfactants, Transcutol P as cosurfactant and water as aqueous phase. The solubility of ECN was firstly determined for screening the ingredients of the system. Pseudo-ternary phase diagrams were constructed to formulate ME and select surfactant and cosurfactant. Central composite design-response surface methodology (CCD-RSM) was utilized to optimize the formulation of ME. The ECN loaded ME was characterized in terms of morphology, particle size and size distribution, pH value, refractive index, viscosity and conductivity, and storage stability of the ECN loaded ME was assayed. Percutaneous permeation of ECN from ME in vitro through rat skin was investigated in comparison with PBS aqueous suspension (1%, w/w), and results showed that ME enhanced drug retention in the skin and permeation through the skin, the enhancement of ME on skin deposition was further visualized through fluorescent-labeled ME by confocal laser scanning microscope (CLSM). The action mechanism of ME on improving percutaneous delivery was studied by performing a pretreatment test. It can speculate that ME does not simply behave as enhancer but it also acts as drug carrier. Furthermore, ME-skin interaction was elucidated through transmission electron microscope (TEM), and attenuated total reflectance fourier-transform infrared (ATR-FTIR). TEM was performed to visualize the micro morphological change of skin. ATR-FTIR was carried out to investigate the molecular vibrations of the components of stratum corneum (SC). The results indicate that the ME system may be a promising vehicle for percutaneous delivery of ECN.

Proniosome-derived niosomes for tacrolimus topical ocular delivery: In vitro cornea permeation, ocular irritation, and in vivo anti-allograft rejection

The objective of this study was to develop proniosome-derived niosomes for topical ophthalmic delivery of Tacrolimus (FK506). The FK506 loaded proniosomes containing poloxamer 188 and lecithin as surfactants, cholesterol as a stabilizer, and minimal amount of ethanol and trace water reconstituted to niosomes prior to use. The stability of FK506 loaded proniosomes was assessed, and the morphology, size, zeta potential, surface tension, and entrapment efficiency of the derived niosomes were characterized, indicating they were feasible for instillation in the eyes. The in vitro permeation of FK506 through the freshly excised rabbit cornea, the cumulative permeation amount of FK506 from niosomes, and the drug retention in the cornea all exhibited significant increase as compared to 0.1% FK506 commercial ointments. The in vivo ocular irritation test of 0.1% FK506 loaded niosomes instilled 4 times per day in rat eyes for 21 consecutive days showed no irritation and good biocompatibility with cornea. The in vivo anti-allograft rejection assessment was performed in a Sprague-Dawley (SD) rat corneal xenotransplantation model. The results showed treatment with 0.1% FK506 loaded niosomes delayed the occurrence of corneal allograft rejection and significantly prolonged the median survival time of corneal allografts to13.86±0.80days as compared with those treated with 1% Cyclosporine (CsA) eye drops, drug-free niosomes, or untreated. In conclusion, the proniosome-derived niosomes may be a promising vehicle for effective ocular drug delivery of FK506.

Phytantriol-based inverted type bicontinuous cubic phase for vascular embolization and drug sustained release.

The potential feasibility of using phytantriol-based inverted type bicontinuous cubic phase as an embolization agent and sustained release system was evaluated in this study. In the ternary phytantriol-water-ethanol system, when water content was less than 30% (w/w), the injectable isotropic solution was formed and could transform into the bicontinuous cubic phase upon contacting the dissolution/body fluids. The transformation of the isotropic solution was confirmed by polarized light microscopy (PLM), small angle X-ray scattering (SAXS), resonance Raman spectroscopy, and rheological measurements. The in vitro dissolution results showed that the release was sustained for up to 30 days and was affected by drug loading and the initial compositions of isotropic solutions. In vivo, the embolization study was performed with normal rabbits using transcatheter arterial embolization technique and was monitored under digital subtraction angiography (DSA). The angiographical results showed that the hepatic artery was successfully embolized with phytantriol cubic phase. Therefore, with the vascular embolization and sustained release characteristics, the phytantriol-based inverted type bicontinuous cubic phase could be used for arterial transcatheter chemoembolization on hepatocellular carcinoma.

In vitro and in vivo evaluation of ordered mesoporous silica as a novel adsorbent in liquisolid formulation

Background A liquisolid technique has been reported to be a new approach to improve the release of poorly water-soluble drugs for oral administration. However, an apparent limitation of this technique is the formulation of a high dose because a large amount of liquid vehicle is needed, which finally results in a low-dose liquisolid formulation. Silica as an absorbent has been used extensively in liquisolid formulations. Although nanoparticle silica can be prepared and used to improve liquid adsorption capacity, loading a high dose of drug into a liquisolid is still a challenge. With the aim of improving adsorption capacity and accordingly achieving high drug loading, ordered mesoporous silica with a high surface area and narrow pore size distribution was synthesized and used in a liquisolid formulation. Methods Ordered mesoporous silica was synthesized and its particle size and morphology were tailored by controlling the concentration of cetyltrimethyl ammonium bromide. The ordered mesoporous silica synthesized was characterized by transmission electron microscopy, scanning electron microscopy, Fourier transform infrared spectroscopy, small-angle x-ray diffraction, wide angle x-ray diffraction, and nitrogen adsorption-desorption measurements. The liquid adsorption capacity of ordered mesoporous silica was subsequently compared with that of conventional silica materials using PEG400 as the model liquid. Carbamazepine was chosen as a model drug to prepare the liquisolid formulation, with ordered mesoporous silica as the adsorbent material. The preparation was evaluated and compared with commercially available fast-release carbamazepine tablets in vitro and in vivo. Results Characterization of the ordered mesoporous silica synthesized in this study indicated a huge Brunauer–Emmett–Teller surface area (1030 m2/g), an ordered mesoporous structure with a pore size of 2.8 nm, and high adsorption capacity for liquid compared with conventional silica. Compared with fast-release commercial carbamazepine tablets, drug release from the liquisolid capsules was greatly improved, and the bioavailability of the liquisolid preparation was enhanced by 182.7%. Conclusion Ordered mesoporous silica is a potentially attractive adsorbent which may lead to a new approach for development of liquisolid products.

Lactoferrin-modified PEGylated liposomes loaded with doxorubicin for targeting delivery to hepatocellular carcinoma.

Lactoferrin (Lf) is a potential-targeting ligand for hepatocellular carcinoma (HCC) cells because of its specific binding with asialoglycoprotein receptor (ASGPR). In this present work, a doxorubicin (DOX)-loaded, Lf-modified, polyethylene glycol (PEG)ylated liposome (Lf-PLS) system was developed, and its targeting effect and antitumor efficacy to HCC was also explored. The DOX-loaded Lf-PLS system had spherical or oval vesicles, with mean particle size approximately 100 nm, and had an encapsulation efficiency of 97%. The confocal microscopy and flow cytometry indicated that the cellular uptake of Lf-PLS was significantly higher than that of PEGylated liposome (PLS) in ASGPR-positive cells (P<0.05) but not in ASGPR-negative cells (P>0.05). Cytotoxicity assay by MTT demonstrated that DOX-loaded Lf-PLS showed significantly stronger antiproliferative effects on ASGPR-positive HCC cells than did PLS without the Lf modification (P<0.05). The in vivo antitumor studies on male BALB/c nude mice bearing HepG2 xenografts demonstrated that DOX-loaded Lf-PLS had significantly stronger antitumor efficacy compared with PLS (P<0.05) and free DOX (P<0.05). All these results demonstrated that a DOX-loaded Lf-PLS might have great potential application for HCC-targeting therapy.

A novel bottom-up process to produce nanoparticles containing protein and peptide for suspension in hydrofluoroalkane propellants

To overcome the disadvantages of microemulsion and nanoprecipitation methods to produce protein-containing nanoparticles, a novel bottom-up process was developed to produce nanoparticles containing the model protein lysozyme. The nanoparticles were generated by freeze-drying a solution of lysozyme, lecithin and lactose in tert-butyl alcohol (TBA)/water co-solvent system and washing off excess lecithin in lyophilizate by centrifugation. Formulation parameters such as lecithin concentration in organic phase, water content in TBA/water co-solvent, and lactose concentration in water were optimized so as to obtain desired nanoparticles with retention of the bioactivity of lysozyme. Based on the results, 24.0% (w/v) of lecithin, 37.5% (v/v) of water content, and 0.56% (w/v) of lactose concentration were selected to generate spherical nanoparticles with approximately 200 nm in mean size, 0.1 in polydispersity index (PI), and 99% retained bioactivity of lysozyme. These nanoparticles rinsed with ethanol containing dipalmitoylphosphatidylcholine (DPPC), Span 85 or oleic acid (3%, w/v) could readily be dispersed in HFA 134a to form a stable suspension with good redispersibility and 98% retained bioactivity of lysozyme. The study indicates there is a potential to produce pressed metered dose inhaler (pMDI) formulations containing therapeutic protein and peptide nanoparticles.

Optimization of the preparation process for an oral phytantriol-based amphotericin B cubosomes

In order to develop an oral formation of Amphotericin B (AmB) using phytantriol- (PYT-) based cubosomes with desirable properties, homogenization conditions were firstly investigated to determine their effects on the morphological and dimensional characteristics of cubosomes. Under the optimized homogenization conditions of 1200 bar for 9 cycles, cubosomes with reproducible, narrow particle size distribution and a mean particle size of 256.9 nm ± 4.9 nm were obtained. The structure of the dispersed cubosomes was revealed by SAXS (small-angle X-ray scattering) and Cryo-TEM (cryogenic transmission electron microscopy) as a bicontinuous cubic liquid crystalline phase with Pn3m geometry. To overcome the poor drug solubility and increase the drug-loading rate, a solubilization method was firstly used to develop cubosomes containing AmB. The encapsulation efficiency determined by HPLC assay was 87.8% ± 3.4%, and UV spectroscopy and stability studies in simulated gastric fluids further confirmed that AmB was successfully encapsulated in cubosomes.

Microemulsion based gel for topical dermal delivery of pseudolaric acid B: In vitro and in vivo evaluation

Pseudolaric acid B (PAB) possesses comparable fungicidal activity against Candida albicans to amphotericin B and antifungal activity against azole-resistant Candida species. However, its poor water solubility makes the formulation a considerable challenge for dermal permeation of PAB. The aim of this project was to improve the solubility and eventually the dermal permeability and bioavailability by developing a microemulsion based gel for PAB (PAB ME-gel). PAB ME-gel was formulated using isopropyl myristate as oil phase, cremphor EL as surfactant, transcutol P as cosurfactant, and carbopol as gel matrix, and characterized by droplet size, morphology, pH, and rheology. The 3 month storage test showed that PAB ME-gel possessed good physicochemical stability. In vitro permeation of PAB through rat skin from ME-gel was investigated in comparison with PAB microemulsion (PAB ME) and PAB hydrogel (PAB gel), and results showed that ME significantly enhanced PAB retention in the skin and permeation through the skin whether it was incorporated into the gel or not. In vivo dermatopharmacokinetics study using microdialysis further confirmed that ME-gel significantly increased PAB dermal bioavailability compared with the gel (41.95 ± 8.89 μg/ml vs. 13.90 ± 2.22 μg/ml). In vitro sensitivity against C. albicans test indicated that the antifungal activity of PAB ME-gel increased with the increase of PAB loading, and 8 mg/g of PAB ME-gel exhibited a higher antifungal activity than that of 20mg/g miconazole nitrate cream. In vivo antifungal activity evaluation in C. albicans infected guinea pigs showed that 8 mg/g of PAB ME-gel exhibited a higher efficacy than that of 20mg/g miconazole nitrate cream after 7 day treatment. Overall, the results indicated that the ME enhanced in vitro permeability, in vivo dermal bioavailability, and antifungal activity of PAB. Therefore, ME-gel may be a promising approach for topical dermal delivery of PAB to treat skin fungal infection.

Hyaluronic acid-coated niosomes facilitate tacrolimus ocular delivery: Mucoadhesion, precorneal retention, aqueous humor pharmacokinetics, and transcorneal permeability

Tacrolimus (FK506) was used to prevent corneal allograft rejection in patients who were resistant to steroids and cyclosporine. However, the formulation for FK506 ocular delivery remained a challenge due to the drugs high hydrophobicity, high molecular weight, and eyes physiological and anatomical constraints. The aim of this project is to develop an ocular delivery system for FK506 based on a combined strategy of niosomes and mucoadhesive hyaluronic acid (HA), i.e., FK506HA-coated niosomes, which exploits virtues of both niosomes and HA to synergistically improve ophthalmic bioavailability. The FK506HA-coated niosomes were characterized with particle size, zeta potential, and rheology behavior. Mucoadhesion of FK506HA-coated niosomes to mucin was investigated through surface plasmon resonance in comparison with non-coated niosomes and HA solution. The results showed that niosomes possessed adhesion to mucin, and HA coating enhanced the adhesion. The in vivo precorneal retention was evaluated in rabbit, and the results showed that HA-coated niosomes prolonged the residence of FK506 significantly in comparison with non-coated niosomes or suspension. Aqueous humor pharmacokinetics test showed that area under curve of HA-coated niosomes was 2.3-fold and 1.2-fold as that of suspension and non-coated niosomes, respectively. Moreover, the synergetic corneal permeability enhancement of the hybrid delivery system on FK506 was visualized and confirmed by confocal laser scanning microscope. Overall, the results indicated that the hybrid system facilitated FK506 ocular delivery on mucoadhesion, precorneal retention, aqueous humor pharmacokinetics and transcorneal permeability. Therefore, HA-coated niosomes may be a promising approach for ocular targeting delivery of FK506.