Yinghua Sun

Dual targeting folate-conjugated hyaluronic acid polymeric micelles for paclitaxel delivery

A series of novel self-assembled hyaluronic acid derivatives (HA-C(18)) grafted with hydrophobic octadecyl moiety and further dual targeting folic acid-conjugated HA-C(18) (FA-HA-C(18)) were synthesized. With the increase in the degree of substitution of octadecyl group from 12.7% to 19.3%, the critical micellar concentration of HA-C(18) copolymers decreased from 37.3 to 10.0 μg/mL. Paclitaxel (PTX) was successfully encapsulated into the hydrophobic cores of the HA-C(18) and FA-HA-C(18) micelles, with encapsulation efficiency as high as 97.3%. The physicochemical properties of the polymeric micelles were measured by DLS, TEM and DSC. Moreover, in vitro release behavior of PTX was investigated by dialysis bag method and PTX was released from micelles in a near zero-order sustained manner. In vitro antitumor activity tests suggested PTX-loaded HA-C(18) and FA-HA-C(18) micelles exhibited significantly higher cytotoxic activity against MCF-7 and A549 cells compared to Taxol at a lower PTX concentration. The cellular uptake experiments were conducted by quantitative assay of PTX cellular accumulation and confocal laser scanning microscopy imaging of coumarin-6 labeled HA-C(18) and FA-HA-C(18) micelles in folate receptor overexpressing MCF-7 cells. Folate and CD44 receptor competitive inhibition studies performed by fluorescence microscopy imaging suggested intracellular delivery of HA-C(18) and FA-HA-C(18) micelles were efficiently taken up via CD44 receptor-mediated endocytosis. The folate receptor-mediated endocytosis further enhanced internalized amounts of FA-HA-C(18) micelles in MCF-7 cells, as compared with HA-C(18) micelles. The internalization pathways of PTX-loaded HA-C(18) and FA-HA-C(18) micelles might include clathrin-mediated endocytosis, caveolae-mediated endocytosis and macropinocytosis. Therefore, the present study suggested that HA-C(18) and FA-HA-C(18) copolymers as biodegradable, biocompatible and cell-specific targetable nanostructure carriers, are promising nanosystems for cellular and intracellular targeting delivery of hydrophobic anticancer drugs.

Enhanced Oral Delivery of Paclitaxel Using Acetylcysteine Functionalized Chitosan-Vitamin E Succinate Nanomicelles Based on a Mucus Bioadhesion and Penetration Mechanism

In addition to being a physiological protective barrier, the gastrointestinal mucosal membrane is also a primary obstacle that hinders the oral absorption of many therapeutic compounds, especially drugs with a poor permeability. In order to resolve this impasse, we have designed multifunctional nanomicelles based on the acetylcysteine functionalized chitosan-vitamin E succinate copolymer (CS-VES-NAC, CVN), which exhibit marked bioadhesion, possess the ability to penetrate mucus, and enhance the oral absorption of a hydrophobic drug with a poor penetrative profile, paclitaxel. The intestinal absorption (Ka = 0.38 ± 0.04 min(-1), Papp = 0.059 cm · min(-1)) of CVN nanomicelles was greatly improved (4.5-fold) in comparison with paclitaxel solution, and CLSM (confocal laser scanning microscope) pictures also showed not only enhanced adhesion to the intestinal surface but improved accumulation within intestinal villi. The in vivo pharmacokinetics indicated that the AUC0-t (586.37 ng/mL · h) of CVN nanomicelles was markedly enhanced compared with PTX solution. In summary, the novel multifunctional CVN nanomicelles appear to be a promising nanocarrier for insoluble and poorly permeable drugs due to their high bioadhesion and permeation-enhancing capability.

Preparation and in vitro/in vivo evaluation of fenofibrate nanocrystals

The majority of the candidate drug entities exhibit solubility-limiting absorption. Nanocrystal suspensions with particle size in the nanometer scale (nanonization) can increase aqueous solubility and improve oral bioavailability. Regarding the importance of nanosuspension solidification, this study intended to study the critical parameters on redispersed particle size of dried nanocrystals as pretabletting material during spray drying process, such as supporting agents, inlet temperature and feed rate. Fenofibrate with poor water solubility and low melting point was used as a model drug. Nanocrystals of fenofibrate were prepared by a bead-milling method. Five types of hydrophilic excipients in combination with sodium dodecyl sulfate (SDS) were studied as supporting agents during spray drying. The resultant products were characterized by particle size analysis, scanning electron microscopy imaging, differential scanning calorimetry, X-ray powder diffraction and dissolution testing. Spray dried powder with a mean redispersed particle size of 699 nm was produced by using mannitol and SDS as supporting agent. Weight ratio (RF/m) of fenofibrate:mannitol and inlet temperature strongly influenced the particle size of the nanocrystals. The optimal inlet temperature and feed rate was optimized as 75 °C and 4 mL min(-1), respectively. Partially transformation of fenofibrate crystalline to the amorphous form was observed. The dissolution profiles of tablets prepared with the spray dried powder were similar to the commercial nanocrystal formulation Lipidil™ ez, and faster than that of the micronized formulation. The relative bioavailability of the spray-dried formulation was determined to be 89.6% taking Lipidil™ ez as the reference. There were no significant statistic differences of AUC0-72 and Cmax between the two formulations.

Nimodipine nanocrystals for oral bioavailability improvement: Role of mesenteric lymph transport in the oral absorption

PURPOSE We had conducted a comprehensive study on preparation, characterization and pharmacokinetics of nimodipine nanocrystals for oral administration previously, and nimodipine nanocrystals displayed lower dissolution profiles but higher bioavailability than Nimotop(®). In this study, we aimed at elucidating the reasons of unfavorable in vitro in vivo correlation for NMD nanocrystals and Nimotop(®) with a hypothesis that special oral absorption mechanism was involved in the absorption of nimodipine nanocrystals. METHODS Investigations of oral absorption mechanism of the nanocrystals were performed on everted gut sac models, lymphatically (mesenteric lymph duct) cannulated SD rats, Caco-2 cell monolayers and chylomicron flow blocking rats, respectively. RESULTS The permeability of nanocrystals in duodenum, ileum and colon was not superior to that of Nimotop(®), suggestive of special absorption mechanisms involved. Exudates of nanocrystals from enterocytes were detected in mesenteric lymphatic fluids using a transmission electron microscope, and the bioavailability was only about half of the control after the mesenteric lymph was blocked. The nanocrystals were taken up by enterocytes via macropinocytosis and caveolin-mediated endocytosis pathways. CONCLUSIONS It was impossible to establish a favorable in vitro in vivo correlation for NMD nanocrystals and Nimotop(®), because portions of the nanocrystals underwent macropinocytosis and caveolin-mediated endocytosis by enterocytes as intact nanocrystal forms, then bypassed the liver first-pass metabolism.

A drug-in-adhesive transdermal patch for S-amlodipine free base: In vitro and in vivo characterization

The objective of the present investigation was to develop and evaluate a drug-in-adhesive transdermal patch for S-amlodipine (S-AM) free base. Initial in vitro experiments were conducted to optimize the formulation parameters before transdermal delivery in rats. The effects of the type of adhesive and the content of permeation enhancers on S-AM free base transport across excised rat skin were evaluated. For in vivo studies, patches were administered transdermally to rats while orally administered S-AM in suspension and intravenously administered S-AM solution were used as controls. The plasma level of S-AM following transdermal application could be maintained for 72 h. After transdermal administration to rats, the absolute bioavailability was 88.8% for S-AM free base. After dose normalization, the areas under the plasma concentration-time curve (AUC) and mean residence times (MRT) were evidently increased and extended, respectively. This suggests that the transdermal application of S-AM in a drug-in-adhesive transdermal patch may be used for the treatment for hypertension.

Preparation and in vitro/in vivo evaluation of sustained-release venlafaxine hydrochloride pellets.

The objective of this study was to prepare different venlafaxine hydrochloride sustained-release products and to elucidate the influence of composition of the coating film on the in vitro drug release profiles and in vivo pharmacokinetics. Pellets were prepared by a standardized process of extrusion/spheronization. A selected fraction size (0.8-1.0 mm diameter) of pellets of each formulation was coated with Eudragit NE30D or ethylcellulose (10 cps). Many efforts have been made to tailor drug release rate by choosing different coating materials, different percent of pore forming components and coating weight variation to achieve a desired sustained-release effect. The dissolution studies were performed and data were analyzed in terms of cumulative release as a function of time. The influence on the release of venlafaxine from sustained-release capsules was observed in dissolution media of different pH and gradient pH. Scanning electron microscope (SEM) micrographs revealed morphological changes of the pellet coating surface which were related to in vitro drug release profiles. The relative bioavailability for Formulation 1 and Formulation 2 was evaluated in six healthy beagle dogs after oral administration in a fast state using sustained-release capsules (Effexor XR) as a reference. The results suggested that Formulation 1 and Formulation 2 both had better bioavailability compared with Effexor XR. It could be found that there existed quite difference in the in vivo release and oral absorption performances, despite the similar in vitro drug release behavior for the two formulations. It might be attributable to complex in vivo environment and then variation in the release behavior. Thus differences in the film micro-structure and surface roughness caused by aqueous dispersion and organic solvent coating techniques strongly influence the in vivo release and oral absorption performances.

Structure-Based Prediction of the Nonspecific Binding of Drugs to Hepatic Microsomes

For the accurate prediction of in vivo hepatic clearance or drug–drug interaction potential through in vitro microsomal metabolic data, it is essential to evaluate the fraction unbound in hepatic microsomal incubation media. Here, a structure-based in silico predictive model of the nonspecific binding (fumic, fraction unbound in hepatic microsomes) for 86 drugs was successfully developed based on seven selected molecular descriptors. The R2 of the predicted and observed log((1 − fumic)/fumic) for the training set (n = 64) and test set (n = 22) were 0.82 and 0.85, respectively. The average fold error (AFE, calculated by fumic rather than log((1 − fumic)/fumic)) of the in silico model was 1.33 (n = 86). The predictive capability of fumic for neutral drugs compared well to that for basic compounds (R2 = 0.82, AFE = 1.18 and fold error values were all below 2, except for felodipine and progesterone) in our model. This model appears to perform better for neutral compounds when compared to models previously published in the literature. Therefore, this in silico model may be used as an additional tool to estimate fumic and for predicting in vivo hepatic clearance and inhibition potential from in vitro hepatic microsomal studies.

A two-step strategy to design high bioavailable controlled-release nimodipine tablets: The push–pull osmotic pump in combination with the micronization/solid dispersion techniques

In order to decrease the fluctuation of blood concentration and to increase the oral bioavailability of nimodipine (NMD), a two-step strategy including the push-pull osmotic pump (PPOP) method in combination with micronization and solid dispersion techniques, was used to prepare the controlled-release high-bioavailability solid dosages. The optimization of formulation and process was conducted by comparing effects of different solubilization methods on release behavior of NMD. The in vitro dissolution studies indicated that both the two strategies were able to deliver NMD in the predetermined zero-order manner from 2 to 12h, regardless of effects of release media and agitation rates. Although the Cmax values of two PPOP tablets were lower than that of the reference formulation, both the Tmax values were prolonged, demonstrating the prominent controlled release performance. In comparison with the commercial reference tables, the relative bioavailability of the two formulations was 67.0% and 121.1%, respectively, indicating the solid dispersion technique was more efficient than the micronization technique in terms of solubilization capability and absorption enhancement. In summary, the two-step strategy, combining the push-pull osmotic pump method with the solid dispersion technique, is a very effective method to prepare high bioavailable controlled-release formulations of the poorly soluble drugs, i.e. NMD, taking into account the therapeutical efficiency and safety.

Considerations in the development of an in vitro dissolution condition for lacidipine tablets: in vivo pharmacokinetic evaluation

In this study, a new discriminative dissolution condition for lacidipine tablets was developed by the established in vitro–in vivo relationship. Series of dissolution media of phosphate buffer solution (PBS) covering the pH range of 1–7.2 and pH 6.8 PBS containing different concentrations of sodium dodecyl sulfate (SDS), were prepared and used to investigate the dissolution behavior of lacidipine tablets. There was an obvious difference in the dissolution profiles of the both brands in pH 6.8 PBS medium containing 0.1% SDS. The pharmacokinetic study of the two lacidipine tablets was carried out in the healthy beagle dogs at a single dose of 4 mg. Statistical comparison of the AUC0–24, Cmax, and Tmax showed a significant difference in the two brand tablets, coinciding with the dissolution performance with pH 6.8 PBS containing 0.1% SDS. The superiority of the proposed system, pH 6.8 PBS containing 0.1% SDS, could serve as a dissolution medium for lacidipine tablets, and more important it could discriminate the in vivo pharmacokinetic behavior for different brands of products. In summary, in vivo pharmacokinetic evaluation is essential to develop an appropriate in vitro dissolution condition for oral solid dosage forms of poorly soluble drugs.

Rapid and sensitive hydrophilic interaction chromatography/tandem mass spectrometry method for the determination of glycyl-sarcosine in cell homogenates

A rapid, selective and sensitive hydrophilic interaction chromatography/tandem mass spectrometry (HILIC/MS/MS) was developed and validated for the determination of glycyl-sarcosine (Gly-Sar) in Caco-2 cell homogenates. After a simple protein precipitation with acetonitrile, the analyte was separated on a HILIC column and detected by a triple quadrupole mass spectrometry equipped with an electrospray ionization (ESI) source. The method was linear among the concentration range of 1-2000 ng/mL for Gly-Sar and the lower limit of quantification (LLOQ) was 1 ng/mL using as l ittle as 50 microL of cell homogenates. The intra-day and inter-day relative standard deviations (RSD) were less than 15% and the relative errors (RE) were all within +/-15%. The validated method was successfully employed in the study of Gly-Sar uptake inhibition in Caco-2 cells by valcytarabine, a potential substrate of the peptide transporter 1 (PEPT1).