Dawei Chen

Esterase-catalyzed dePEGylation of pH-sensitive vesicles modified with cleavable PEG-lipid derivatives

To make vesicles for the better controlled content release, we modified cholesteryl hemisuccinate (CHEMS)-derived vesicles with PEG-lipid derivatives. Two cholesterol analogs, cholesteryl hemisuccinate (CHEMS) and cholesteryl chloroformate (CHM), have been conjugated to the polyethylene glycol (PEG) via their ester bonds for the vesicle modifications, so the PEGs can be cleaved by esterases. The effects of PEG-lipid proportions, serum concentrations and the lipid types on the esterase-catalyzed cleavage of PEG polymer off the vesicles surface were determined. We observed that PEG cleavages decreased as the molar ratios of the PEG-lipids in vesicles increased; on the other hand, PEG cleavages gradually increased with the increased serum concentrations. In contrast to conventional long circulation materials mPEG-DSPE and mPEG-CHOL, the two new conjugates enabled higher degrees of PEG cleavages from modified vesicles. After incubation in the serum at acidic conditions, esterase-catalyzed dePEGylation destabilized these vesicles, increasing the mean particle sizes and promoting content releases. These results suggested that ester linkages between the PEG and lipid anchors allow faster content releases in the suitable conditions. Together, the two esterase cleavable PEG-lipid conjugations may be applied not only to stabilize vesicles and prolong their circulation time, but also to provide more efficient content releases by the esterase controlled dePEGylation.

Effect of honey bee venom on proliferation of K1735M2 mouse melanoma cells in-vitro and growth of murine B16 melanomas in-vivo

Bee venom has been reported to exhibit antitumour activity in‐vitro and in‐vivo. Apoptosis, necrosis and lysis of tumour cells were suggested as possible mechanisms by which bee venom inhibited tumour growth. The aim of this study was to investigate potential mechanisms by which bee venom inhibits K1735M2 mouse melanoma cells in‐vitro and B16 melanoma, a transplantable solid melanoma in C57BL/6 mice, in‐vivo. The proliferation of K1735M2 cells in‐vitro was inhibited by bee venom in a concentration‐ and time‐dependent manner. The inhibition was indicated by the arrest of the cell cycle at the G1 stage, as detected by flow cytometric measurements. The bee venom induced apoptosis‐like cell death as identified by histological observations and by DNA fragmentation. In the in‐vivo experiments, the bee venom (1.0, 3.0, 9.0 mg kg−1 of body weight, on days 1–12) was injected intraperitoneally into mice 24 h after the mice were inoculated with B16 cells. Inhibition of the solid tumour was observed. Apoptosis of the K1735M2 cells was suggested as the possible mechanism by which bee venom inhibited cell proliferation and induced K1735M2 cell differentiation in‐vitro. The in‐vivo experiment indicated that bee venom could be used as a chemotherapeutic agent against malignant tumours.

Characterization of water-in-oil microemulsion for oral delivery of earthworm fibrinolytic enzyme

Earthworm fibrinolytic enzyme (EFE-d, Mr 24177), a water-soluble protein, is clinically used for the management of cardiovascular diseases. However, this protein drug has a very low oral bioavailability because of its low oil/water partitioning, low membrane permeability and unstable nature in harsh gastric juice. This study explored the possibility of absorption and efficacy enhancement for EFE-d through the delivery of the water-in-oil (w/o) microemulsions. The w/o microemulsion consisting of Labrafac CC, Labrasol, Plurol Oleique CC 497 and saline (54/18/18/10, % w/w) was developed and characterized, including conductivity, viscosity, particle size and in vitro membrane permeability. The w/o microemulsion and the control solution of EFE-d were administered intraduodenally (or orally) to rats. The w/o microemulsion possessed a higher intestinal membrane permeability in vitro as well as a higher absorption and efficacy in vivo, when compared to control solution. The intraduodenal bioavailability of EFE-d for microemulsions was 208-fold higher than that of control solution and the absolute bioavailability was 17.55%. Meanwhile, there was no tissue damage of the intestinal mucosa found after oral multiple-dose administration of the EFE-d microemulsion to rats. These findings indicated that the w/o microemulsion may represent a safe and effective oral delivery system for hydrophilic bioactivity macromolecules.

Effect of polypeptides in bee venom on growth inhibition and apoptosis induction of the human hepatoma cell line SMMC-7721 in-vitro and Balb/c nude mice in-vivo

Polypeptides in bee venom (PBV) produced a significant growth inhibition against SMMC‐7721 human hepatoma cell line. Analysis of the mechanisms of cell death indicated that PBV induced an apoptotic cell death. SMMC‐7721 cells exposed to PBV (10.0 μg mL−1) produced an insignificant morphological change. Analysis of the cytotoxicity with the MTT (3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium) assay confirmed that the cytotoxic effects of PBV were dose‐ and time‐dependent. The result of Ki67 immunohistochemistry demonstrated that the proliferation of SMMC‐7721 cells treated with PBV (10.0 μg mL−1) was inhibited. The apoptotic cell death was then confirmed by annexin V, propidium iodide staining and DNA fragmentation analysis. In in‐vivo experiments, treatment with PBV (1.5 or 3 mg kg−1) resulted in a significant retardation of SMMC‐7721 cell growth in Balb/c nude mice. These findings suggested that PBV could be used as a chemotherapeutic agent against tumours.

Synchronic release of two hormonal contraceptives for about one month from the PLGA microspheres: In vitro and in vivo studies

A controlled drug release system based on the injectable PLGA microspheres loaded with gestodene and ethinyl estradiol was prepared and evaluated for the feasibility of monthly synchronic delivery of the two hormonal contraceptives. The scanning electron microscopy, light-scattering analyzer and gel permeation chromatography were used to study the morphology, particle size and molecular weight of the polymer microspheres, respectively. HPLC was utilized to determine the drug loading and the drug released, while a LC-MS-MS system was employed to analyze the plasma drug concentration. Result indicated that the PLGA particles obtained were spherical and appropriate in size. The formulation was stable during the test period. In vitro drug release from the microspheres for both drugs was sustained for about 30 days mostly by the diffusion mechanism. The plasma drug concentration-time profiles of the drug-loaded microspheres were relatively smooth after subcutaneous injection to rats for about 1-month, compared with that for drug suspension. In vitro and in vivo correlation was established. One of the most important facts is the synchronicity of the two contraceptives both in the release kinetics in vitro and the pharmacokinetic behaviors in vivo. Therefore, the synchronic delivery of two contraceptives is achieved for about 1 month by using the injectable PLGA-based microspheres.

Nanoparticles of 5‐fluorouracil (5‐FU) loaded N‐succinyl‐chitosan (Suc‐Chi) for cancer chemotherapy: preparation, characterization — in‐vitro drug release and anti‐tumour activity

N‐Succinyl‐chitosan has favourable properties as a drug carrier, such as biocompatibility, low toxicity and long‐term retention in the body. It is a good candidate for cancer chemotherapy as a polymeric drug carrier. This study describes the preparation and characterization of 5‐fluorouracil‐loaded N‐succinyl‐chitosan nanoparticles (5‐FU‐Suc‐Chi/NP) by an emulsification solvent diffusion method. The influence of the initial 5‐FU concentration on the nanoparticle characteristics and release behaviour in phosphate‐buffered saline solution (PBS) was evaluated. The Suc‐Chi nanoparticles had a particle diameter (Z‐average) in the range 202∼273 nm and a negative zeta‐potential (approx. −27 to −18 mV). The formulation with an initial 5‐FU concentration of 1000 μg mL−1 provided the highest loading capacity (19%) and the highest extent of release (61% at 24 h). The 5‐FU‐Suc‐Chi/NP showed good anti‐tumour activity against Sarcoma 180 solid tumour and mild toxicity. According to the data obtained, this Suc‐Chi‐based nanotechnology opens new and interesting perspectives for cancer chemotherapy.