PharmaGateway

Pharmaceutical Research: Molecular Weight Effects on the Miscibility Behavior of Dextran and Maltodextrin with Poly(vinylpyrrolidone)

Van Eerdenbrugh; Taylor — 2012-02-02

Type: Original Paper
Purpose

To characterize and interpret the miscibility of dextran and maltodextrin with poly(vinylpyrrolidone) (DEX-PVP) as a function of polymer molecular weights.

Methods

Blend miscibility was studied using 4 different molecular weight (MW) grades of DEX combined with 5 MW grades of PVP, over a broad compositional range. Miscibility was evaluated by inspection of glass transition events measured by differential scanning calorimetry (DSC). Fourier transform mid-infrared spectroscopy (FTIR), combined with curve fitting, was performed to characterize the extent of hydrogen bonding. The observed miscibility behavior was further interpreted in terms of mixing thermodynamics.

Results

Miscibility of the blends ranged from fully miscible to completely immiscible with multiple partially miscible systems observed. Increasing polymer molecular weight decreased miscibility. For the lowest DEX grade, hydrogen bonding was independent of PVP MW, as expected since all systems were completely miscible. Higher molecular weights of DEX resulted in reduced intermolecular hydrogen bonding and decreased miscibility, increasingly so for higher MW PVP grades. Evaluation of the mixing thermodynamics supported these findings.

Conclusions

With higher combined molecular weights of DEX-PVP blends, phase behavior evolves from completely miscible to virtually immiscible. Concurrently, DEX-PVP hydrogen bonding decreases. From a thermodynamic perspective, the combinatorial mixing entropy was observed to decrease as the molecular weight of the polymers increased, providing a reduced counterbalance to the unfavorable mixing enthalpy thought to accompany this polymer combination.

Pharmaceutical Research: Physicochemical and Biological Evaluation of siRNA Polyplexes Based on PEGylated Poly(amido amine)s

Vader; Aa; Engbersen; Storm; Schiffelers — 2012-02-01

Type: Original Paper
Purpose

Use of RNA interference as novel therapeutic strategy is hampered by inefficient delivery of its mediator, siRNA, to target cells. Cationic polymers have been thoroughly investigated for this purpose but often display unfavorable characteristics for systemic administration, such as interactions with serum and/or toxicity.

Methods

We report the synthesis of a new PEGylated polymer based on biodegradable poly(amido amine)s with disulfide linkages in the backbone. Various amounts of PEGylated polymers were mixed with their unPEGylated counterparts prior to polyplex formation to alter PEG content in the final complex.

Results

PEGylation effectively decreased polyplex surface charge, salt- or serum-induced aggregation and interaction with erythrocytes. Increasing amount of PEG in formulation also reduced its stability against heparin displacement, cellular uptake and subsequent silencing efficiency. Yet, for polyplexes with high PEG content, significant gene silencing efficacy was found, which was combined with almost no toxicity.

Conclusions

PEGylated poly(amido amine)s are promising carriers for systemic siRNA delivery in vivo.

Pharmaceutical Research: Development of a Performance Verification Test for USP Apparatus 4

Eaton; Tran; Hauck; Stippler — 2012-02-01

Type: Original Paper
Purpose

To evaluate salicylic acid tablets as a candidate reference material in a Performance Verification Test (PVT) when a USP performance test for dissolution (General Chapter <711>) relies on USP Apparatus 4 (flow-through cell).

Methods

We developed a dissolution procedure relying on Apparatus 4 and salicylic acid tablets. Thereafter, a designed experiment was conducted to identify operational variables that significantly affect salicylic acid dissolution in this apparatus.

Results

Four variables (size of glass beads, cell temperature, flow rate, level of deaeration) and one combination effect (deaeration/bead size) were significant for mean percent dissolved. Two variables (tablet orientation, level of deaeration) were significant for standard deviation results, but these effects were less pronounced than those for mean percent dissolved results. Three variables (analyst, tester manufacturer, amount of glass beads) had no statistically significant effects on either the mean or standard deviation of the responses.

Conclusions

The proposed PVT is capable of probing effects of changes in several critical operational parameters of Apparatus 4. Salicylic acid tablets were shown to be a suitable reference material for the PVT. The PVT using salicylic acid tablets satisfies important aspects of a PVT.

Pharmaceutical Research: Cationic Lipid-Coated Gold Nanoparticles as Efficient and Non-Cytotoxic Intracellular siRNA Delivery Vehicles

Kong; Bae; Jo; Kim; Park — 2012-02-01

Type: Original Paper
Purpose

Cationic lipid-coated gold nanoparticles were developed for efficient intracellular delivery of therapeutic siRNA.

Methods

Particle formation was characterized by UV-visible spectroscopy, atomic force microscopy, and dynamic light scattering analysis. Cellular uptake, gene silencing effect, and cytotoxicity were investigated in multiple human cancer cell lines.

Results

Nanoparticles had a spherical nanostructure with highly cationic surface charge and could form stable nanosized polyelectrolyte complexes with siRNA via electrostatic interactions; complexes exhibited efficient intracellular uptake and significant gene silencing effect with markedly low cytotoxicity compared to the widely used polycationic carrier, linear polyethyleneimine.

Conclusions

We demonstrated that cationic lipid-coated gold nanoparticles could be widely utilized as efficient and safe siRNA nanocarriers for diverse therapeutic and diagnostic applications.

Pharmaceutical Research: Palmitoyl Ascorbate Liposomes and Free Ascorbic Acid: Comparison of Anticancer Therapeutic Effects Upon Parenteral Administration

Sawant; Vaze; Wang; D’Souza; Rockwell; Gada; Khaw; Torchilin — 2012-02-01

Type: Original Paper
Purpose

To evaluate and compare anticancer therapeutic effect of palmitoyl ascorbate liposomes (PAL) and free ascorbic acid (AA).

Methods

Liposomes incorporating palmitoyl ascorbate (PA) were prepared and evaluated for PA content by HPLC. To elucidate mechanism of action of cell death in vitro, effect of various H₂O₂ scavengers and metal chelators on PA-mediated cytotoxicity was studied. Effect of various combinations of PAL and free AA on in vitro cytotoxicity was evaluated on 4T1 cells. In vivo, PAL formulation was modified with polyethylene glycol; effect of PEGylation on in vitro cytotoxicity was evaluated. Biodistribution of PEG-PAL formulation was investigated in female Balb/c mice bearing murine mammary carcinoma (4T1 cells). In vivo anticancer activity of PEG-PAL (PEG-PAL equivalent to 20 mg/kg of PA injected intravenously on alternate days) was compared with free AA therapy in same model.

Results

PEG-PAL treatment was significantly more effective than free AA treatment in slowing tumor growth.

Conclusions

Nanoparticle formulations incorporating PA can kill cancer cells in vitro. The mechanism of PA cytotoxicity is based on production of extracellular reactive oxygen species and involves intracellular transition metals.

Pharmaceutical Research: Smart DNA Vectors Based on Cyclodextrin Polymers: Compaction and Endosomal Release

2012-02-01

Type: Original Paper
Purpose

Neutral β-cyclodextrin polymers (polyβCD) associated with cationic adamantyl derivatives (Ada) can be used to deliver plasmid DNA into cells. In absence of an endosomolytic agent, transfection efficiency remains low because most complexes are trapped in the endosomal compartment. We asked whether addition of an imidazole-modified Ada can increase efficiency of polyβCD/cationic Ada-based delivery system.

Methods

We synthesized two adamantyl derivatives: Ada5, which has a spacer arm between the Ada moiety and a bi-cationic polar head group, and Ada6, which presents an imidazole group. Strength of association between polyβCD and Ada derivatives was evaluated by fluorimetric titration.

Results

Gel mobility shift assay, zeta potential, and dark field transmission electron microscopy experiments demonstrated the system allowed for efficient DNA compaction. In vitro transfection experiments performed on HepG2 and HEK293 cells revealed the quaternary system polyβCD/Ada5/Ada6/DNA has efficiency comparable to cationic lipid DOTAP.

Conclusion

We successfully designed fine-tuned DNA vectors based on cyclodextrin polymers combined with two new adamantyl derivatives, leading to significant transfection associated with low toxicity.

Pharmaceutical Research: Behavior of Monoclonal Antibodies: Relation Between the Second Virial Coefficient (B2) at Low Concentrations and Aggregation Propensity and Viscosity at High Concentrations

Saito; Hasegawa; Kobayashi; Kishi; Uchiyama; Fukui — 2012-02-01

Type: Original Paper
Purpose

To investigate relationship between second virial coefficient B₂ and viscosity and aggregation propensity of highly concentrated monoclonal antibody (MAbs) solutions.

Methods

Intermolecular interactions of 3 MAbs solutions with varying pH were characterized according to B₂ estimated by analytical ultracentrifugation sedimentation equilibrium with initial loading concentrations <10 mg/mL. Viscosity measurements and stability assessments of MAb solutions at concentrations higher than 100 mg/mL were conducted.

Results

B₂ of all MAb solutions depended on solution pH and have qualitative correlation with viscosity and aggregation propensity. The more negative the B₂ values, the more viscous the solution, acquiring increased propensity to aggregate. Solutions with B₂ values of ~2 × 10⁻⁵ mL·mol/g² acquire similar viscosity and aggregation propensity regardless of amino acid sequences; for solutions with negative B₂ values, viscosity and aggregation propensity differed depending on sequences. Results suggest attractive intermolecular interactions represented by negative B₂ values are influenced by surface properties of individual MAbs.

Conclusions

B₂ can be used, within certain limitations, as an effective indicator of viscosity and aggregation propensity of highly concentrated MAb solutions.

Pharmaceutical Research: In Vitro and In Silico Strategies to Identify OATP1B1 Inhibitors and Predict Clinical Drug–Drug Interactions

Karlgren; Ahlin; Bergström; Svensson; Palm; Artursson — 2012-02-01

Type: Original Paper
Purpose

To establish in vitro and in silico models that predict clinical drug–drug interactions (DDIs) with the OATP1B1 (SLCO1B1) transporter.

Methods

The inhibitory effect of 146 drugs and drug-like compounds on OATP1B1-mediated transport was studied in HEK293 cells. A computational model was developed to predict OATP1B1 inhibition. Concentration-dependent effects were investigated for six compounds; clinical DDIs were predicted by calculating change in exposure (i.e. R-values) in eight different ways.

Results

Sixty-five compounds were identified as OATP1B1 inhibitors at 20 μM. The computational model predicted the test set with 80% accuracy for inhibitors and 91% for non-inhibitors. In vitro–in vivo comparisons underscored the importance of using drugs with known clinical effects as references. Thus, reference drugs, cyclosporin A, gemfibrozil, and fenofibrate, provided an inhibition interval to which three antiviral drugs, atazanavir, lopinavir, and amprenavir, could be compared and their clinical DDIs with OATP1B1 classified.

Conclusions

Twenty-two new OATP1B1 inhibitors were identified, a predictive OATP1B1 inhibition in silico model was developed, and successful predictions of clinical DDIs were obtained with OATP1B1.

Pharmaceutical Research: Polymeric Nanoparticles for Increased Oral Bioavailability and Rapid Absorption Using Celecoxib as a Model of a Low-Solubility, High-Permeability Drug

2012-02-01

Type: Original Paper
Purpose

To demonstrate drug/polymer nanoparticles can increase the rate and extent of oral absorption of a low-solubility, high-permeability drug.

Methods

Amorphous drug/polymer nanoparticles containing celecoxib were prepared using ethyl cellulose and either sodium caseinate or bile salt. Nanoparticles were characterized using dynamic light scattering, transmission and scanning electron microscopy, and differential scanning calorimetry. Drug release and resuspension studies were performed using high-performance liquid chromatography. Pharmacokinetic studies were performed in dogs and humans.

Results

A physical model is presented describing the nanoparticle state of matter and release performance. Nanoparticles dosed orally in aqueous suspensions provided higher systemic exposure and faster attainment of peak plasma concentrations than commercial capsules, with median time to maximum drug concentration (Tmax) of 0.75 h in humans for nanoparticles vs. 3 h for commercial capsules. Nanoparticles released celecoxib rapidly and provided higher dissolved-drug concentrations than micronized crystalline drug. Nanoparticle suspensions are stable for several days and can be spray-dried to form dry powders that resuspend in water.

Conclusions

Drug/polymer nanoparticles are well suited for providing rapid oral absorption and increased bioavailability of BCS Class II drugs.

Pharmaceutical Research: Comparative Investigations on In Vitro Serum Stability of Polymeric Micelle Formulations

Miller; Rachel; Besheer; Uezguen; Weigandt; Goepferich — 2012-02-01

Type: Original Paper
Purpose

Stability of polymeric micelles upon injection is essential for a drug delivery system but is not fully understood. We optimized an analytical test allowing quantification of micellar stability in biofluids and applied it to a variety of block copolymer micelles with different hydrophobic block architechtures.

Methods

Polymeric micelles were prepared from four different polymers and investigated via encapsulation of two fluorescent dyes. Samples were incubated in human serum; changes in Foerster Resonance Energy Transfer (FRET) were recorded as a function of time. This fluorescence-based approach was supported semi-quantitatively by results from Asymmetrical Flow Field-Flow-Fractionation (AF4).

Results

After incubation experiments, micellar stability was determined by calculation of two stability-indicating parameters: residual micellar fractions (RMFs) and in vitro serum half-lives. Both parameters showed that PEG-PVPy micelles rapidly destabilized after 3 h (RMF < 45%), whereas PEG-PLA, PEG-PLGA and PEG-PCL micelles were far more stable (RMFs 65 to 98%).

Conclusion

This FRET-based assay is a valuable tool in evaluating and screening serum stability of polymeric micelles and revealed low serum stability of PEG-PVPy micelles compared to polyester-based micelles.

Pharmaceutical Research: Delivery of Brain-Derived Neurotrophic Factor via Nose-to-Brain Pathway

Vaka; Murthy; Balaji; Repka — 2012-02-01

Type: Original Paper
Purpose

To investigate the plausibility of delivering brain-derived neurotrophic factor (BDNF) to brain via nose-to-brain pathway using chitosan as barrier-modulating agent.

Methods

Effect of different viscosity grades chitosan at different concentrations on permeation of fluorescein isothio-cyanate dextran (FD 40 K) across bovine olfactory mucosa was studied using Franz diffusion cells. Medium viscosity chitosan was used to carry out permeation studies of BDNF. Pharmacokinetic and pharmacodynamic studies were carried out in Sprague dawley rats upon intranasal/i.v administration of different formulations.

Results

Medium viscosity chitosan more efficiently enhanced permeation of FD 40 K across olfactory mucosa compared to other grades. In case of BDNF, medium viscosity chitosan (0.25% w/v) enhanced permeation ~14-fold over control (18.78 ± 16.69 ng/cm²). Brain bioavailability of rats administered intranasally with BDNF solution containing chitosan was significantly enhanced ~13-fold compared to rats administered with same concentration of BDNF solution without chitosan. In rats subjected to immobilization stress, BDNF solution containing chitosan significantly decreased immobility time.

Conclusions

Intranasal formulations containing chitosan as barrier-modulating agent significantly enhanced brain bioavailability of BDNF. Delivery of BDNF was found to counteract stress-induced depression in rats.

Pharmaceutical Research: Polymorph Formation and Nucleation Mechanism of Tolfenamic Acid in Solution: An Investigation of Pre-nucleation Solute Association

Mattei; Li — 2012-02-01

Type: Original Paper
Purpose

Crystallization from solution involves nucleation and growth; growth conditions greatly influence self-association behaviors of solute molecules in these steps, affecting crystal packing of organic molecules. We examined the role of pre-nucleation association to provide insights into the mutual influence between molecular conformation in solution and packing in the solid state.

Methods

Crystallization experiments of tolfenamic acid were conducted in ethanol under different supersaturation conditions. UV spectroscopy was performed to study self-association of solute molecules in ethanol as a function of concentration. Intermolecular interaction energies of tolfenamic acid dimers were calculated with quantum mechanical methods.

Results

As supersaturation increased, growth of the most stable polymorph outpaced the metastable one, contradicting Ostwald’s Rule of Stages. UV spectroscopy measurement suggests solute molecules exist as hydrogen-bonded dimers and more dimers form as total concentration increases. Hydrogen bonding in the most stable form is significantly stronger than that in the metastable form.

Conclusions

With the fact that molecular conformation is different in the two polymorphs, as concentration increases, solute molecules rearrange their conformations to form stronger hydrogen-bonded dimers in solution, resulting in nucleation of the most stable form.

Pharmaceutical Research: Promoted Transfection Efficiency of pDNA Polyplexes-Loaded Biodegradable Microparticles Containing Acid-Labile Segments and Galactose Grafts

Chen; Cai; Yang; Wu; Liu; Chen; Li — 2012-02-01

Type: Original Paper
Purpose

Targeting to antigen-presenting cells and efficient intracellular delivery of pDNA are essential for development of microsphere formulations of DNA vaccine.

Methods

Biodegradable polymers containing acid-labile segments and galactose grafts were developed to entrap pDNA polyplexes into microspheres, which were proposed to promote transfection efficiency of pDNA.

Results

Acid-labile characteristics were approved by the hemolysis capabilities of red blood cells and degradation behaviors of matrix polymers; release of pDNA polyplexes from microspheres was significantly accelerated after incubation in acid buffers. Presence of galactose moieties enhanced cellular uptake of microspheres and increased acid-lability due to hydrophilic grafts on acid-labile segments. There was no apparent cytotoxicity of blank microspheres; cytotoxicity of pDNA polyplexes was significantly decreased after encapsulation into and sustained release from microspheres. High transfection efficiency and a dose-dependent transfection were indicated for pDNA polyplex-loaded acid-labile microspheres when balancing with cytotoxicity.

Conclusions

Integration of acid-lability, targeting effect into full biodegradable backbone represents an exciting approach to promote transfection efficiency through modulating release of pDNA polyplexes, targeting to antigen-presenting cells and intracellular delivery of pDNA.

Pharmaceutical Research: Development of Effective Cancer Vaccine Using Targeting System of Antigen Protein to APCs

2012-02-01

Type: Original Paper
Purpose

To develop a novel cancer vaccine using the targeting system of antigen protein to antigen-presenting cells (APCs) for efficient and safe cancer therapy.

Methods

The novel delivery system was constructed with antigen protein, benzalkonium chloride (BK), and γ-polyglutamic acid (γ-PGA), using ovalbumin (OVA) as a model antigen protein and evaluating its immune induction effects and utilities for cancer vaccine.

Results

BK and γ-PGA enabled encapsulation of OVA and formed stable anionic particles at nanoscale, OVA/BK/γ-PGA complex. Complex was taken up by dendritic cell line DC2.4 cells efficiently. We subcutaneously administered the complex to mice and examined induction of IgGs. The complex induced not only Th2-type immunoglobulins but also Th1-type immunoglobulins. OVA/BK/γ-PGA complex inhibited tumor growth of E.G7 cells expressing OVA regularly; administered OVA/BK/γ-PGA complex completely rejected tumor cells.

Conclusion

The novel vaccine could be platform technology for a cancer vaccine.

Pharmaceutical Research: Subcutaneous Absorption of Monoclonal Antibodies: Role of Dose, Site of Injection, and Injection Volume on Rituximab Pharmacokinetics in Rats

Kagan; Turner; Balu-Iyer; Mager — 2012-02-01

Type: Original Paper
Purpose

To determine the effect of dose, the anatomical site of injection, and the injection volume on subcutaneous absorption of rituximab in rats and to explore absorption mechanisms using pharmacokinetic modeling.

Methods

Rituximab serum concentrations were measured following intravenous and subcutaneous administration at the back, abdomen, and foot of rats. Several pharmacokinetic models were developed that included linear and saturable absorption, and degradation and/or protective binding at the injection site.

Results

Rituximab exhibited linear kinetics following intravenous administration; however, bioavailability following subcutaneous injection was inversely related to the dose level. For the 1 mg/kg dose, bioavailability was approximately 70% at all tested injection sites, with faster absorption from the foot (T_max = 12 h for foot vs. 4.6 days for back). Bioavailability for the 10 mg/kg dose was 44 and 31% for the abdomen and back sites and 18% for 40 mg/kg injected at the back. A pharmacokinetic model that included binding as part of the absorption mechanism successfully captured the nonlinearities in rituximab absorption.

Conclusion

The anatomical site of subcutaneous injection influences the rate of absorption and bioavailability of rituximab in rats. Saturable binding may be a major determinant of the nonlinear absorptive transport of monoclonal antibodies.

Pharmaceutical Research: The Effects of PVP(Fe(III)) Catalyst on Polymer Molecular Weight and Gene Delivery Via Biodegradable Cross-Linked Polyethylenimine

Shum; Gabrielson; Forrest; Pack — 2012-02-01

Type: Original Paper
Purpose

Crosslinked, degradable derivatives of low-molecular-weight polyethylenimine (PEI) are relatively efficient and non-cytotoxic gene delivery agents. To further investigate these promising materials, a new synthetic approach was developed using a poly(4-vinylpyridine)-supported Fe(III) catalyst (PVP(Fe(III))) that provides more facile synthesis and enhanced control of polymer molecular weight.

Methods

Biodegradable polymers (D.PEI) comprising 800-Da PEI crosslinked with 1,6-hexanediol diacrylate and exhibiting molecular weights of 1.2, 6.2, and 48 kDa were synthesized utilizing the PVP(Fe(III)) catalyst. D.PEI/DNA polyplexes were characterized using gel retardation, ethidium bromide exclusion, heparan sulfate displacement, and dynamic light scattering. In vitro transfection, cellular uptake, and cytotoxicity of the polyplexes were tested in human cervical cancer cells (HeLa) and human breast cancer cells (MDA-MB-231).

Results

D.PEIs tightly complexed plasmid DNA and formed 320- to 440-nm diameter polyplexes, similar to those comprising non-degradable, 25-kDa, branched PEI. D.PEI polyplexes mediated 2- to 5-fold increased gene delivery efficacy compared to 25-kDa PEI and exhibited 20% lower cytotoxicity in HeLa and no toxicity in MDA-MB-231. In addition, 2- to 7-fold improved cellular uptake of DNA was achieved with D.PEI polyplexes.

Conclusions

PVP(Fe(III)) catalyst provided a more controlled synthesis of D.PEIs, and these materials demonstrated improved in vitro transfection efficacy and reduced cytotoxicity .

Pharmaceutical Research: The Measurement of the β/α Anomer Composition Within Amorphous Lactose Prepared by Spray and Freeze Drying Using a Simple 1H-NMR Method

Jawad; Elleman; Vermeer; Drake; Woodhead; Martin; Royall — 2012-02-01

Type: Original Paper
Purpose

Reports of the anomeric composition of amorphous lactose are rare and state a highly variable range of composition (between 0% and 60% w/w β content). We aimed to develop a quantitative measurement by ¹H-NMR of α and β anomer content in amorphous lactose produced by different production methods.

Methods

Amorphous lactose was prepared by spray and freeze drying 10% w/v aqueous solutions of lactose. NMR analysis was performed in DMSO; peak areas of partially resolved doublets at 6.3 and 6.6 ppm were used to calculate % of α and β lactose present. Polarimetery was used to determine optical rotation of lactose solutions.

Results

Observed specific rotation for supplied crystalline alpha lactose monohydrate of 88° recorded in DMSO was constant for the length of a typical NMR experiment (max. 10 min). β/α anomer contents of amorphous lactose measured by ¹H-NMR had standard deviations as low as 0.1% w/w (n = 6). Drying a lactose solution 4 h after its preparation led to almost 35% w/w difference in anomer composition within solid amorphous material compared to samples dried after only 30 min, e.g. for freeze dried samples, β content was 60 ± 0.1% w/w (4 h) and 25 ± 1.0% w/w (30 min). Mutarotation leads to this increase in β anomer concentration in aqueous solution and within the solid amorphous lactose stored at 25°C. e.g. after 56 d storage the β content of freeze dried lactose (30 min solution) increased from 25±1.0% to 50±0.5% w/w.

Conclusion

A simple solution-based ¹H-NMR method for measurement of anomeric composition of lactose has been established. The solution β/α ratio at the time of drying is mirrored in the composition of the resulting solid amorphous material. In order to produce a consistent anomer composition within spray and freeze dried amorphous lactose, the standing time for the feed solution should be greater than 4 h, such that the most dynamic region of the mutarotation profile has been exceeded. If the amorphous material has been formed from a solution that has not been allowed to equilibrate for 4 h, the resulting solid will continue to undergo mutarotation if trace amounts of moisture are present, until the anomeric β/α ratio slowly approaches 1.7.

Pharmaceutical Research: Influence of Fed-Fasted State on Intestinal PEPT1 Expression and In Vivo Pharmacokinetics of Glycylsarcosine in Wild-Type and Pept1 Knockout Mice

Ma; Hu; Smith — 2012-02-01

Type: Original Paper
Purpose

To determine if fasting would affect the intestinal expression and in vivo functional activity of PEPT1 as determined after oral dosing of the dipeptide glycylsarcosine (GlySar).

Methods

Systemic exposure and tissue distribution studies were performed in wild-type and Pept1 knockout mice, under fed and fasted conditions, following both intravenous and oral doses of [¹⁴C]GlySar at 5 nmol/g body weight. Intestinal PEPT1 expression was evaluated by real-time PCR and immunoblot analyses.

Results

We found that expression of PEPT1 protein in the small intestine was increased ~2-fold in wild-type mice during fasted as compared to fed conditions. In agreement, systemic exposure and peak plasma concentrations of orally administered GlySar were 40 and 65% greater, respectively, in wild-type mice during fasted vs. fed state. No significant differences were observed between fed and fasted animals during PEPT1 ablation. Tissue distribution of GlySar was unchanged after oral dosing for all four treatment groups.

Conclusions

As little as 16 h of fasting can cause significant upregulation of PEPT1 protein expression in the small intestine, which then translates into a significant increase in in vivo oral absorption of GlySar.

Pharmaceutical Research: Low-Molecular-Weight Methylcellulose-Based Thermo-reversible Gel/Pluronic Micelle Combination System for Local and Sustained Docetaxel Delivery

Kim; Won; Lim; Kim — 2012-02-01

Type: Original Paper
Purpose

To develop low-molecular-weight methylcellulose (LMw MC)-based gel/Pluronic F127 micelle combination system for local and sustained delivery of docetaxel (DTX).

Methods

LMw MC and Pluronic F127 were used to formulate an injectable thermo-reversible gel/micelle combination system containing DTX. The DTX-loaded combination system was characterized and its therapeutic efficacy evaluated in a subcutaneous tumor model.

Results

Mixtures of LMw MC, AS, and Pluronic F127 formed gel at ~15–40°C depending on AS concentration. The combination system released DTX for >30 days with a biphasic and sustained release pattern, and DTX stability was maintained during release. The combination system significantly enhanced anti-cancer effects of DTX and prolonged survival of the model mouse in comparison with free DTX.

Conclusions

The LMw MC gel/Pluronic F127 micelle combination system constitutes a promising tool for reducing tumor size and eradicating remaining tumor cells before and after surgery.

Pharmaceutical Research: Polyamide Nanocapsules and Nano-emulsions Containing Parsol® MCX and Parsol® 1789: In Vitro Release, Ex Vivo Skin Penetration and Photo-Stability Studies

Hanno; Anselmi; Bouchemal — 2012-02-01

Type: Original Paper
Purpose

To prepare polyamide nanocapsules for skin photo-protection, encapsulating α-tocopherol, Parsol®MCX (ethylhexyl methoxycinnamate) and/or Parsol®1789 (butyl methoxydibenzoylmethane).

Methods

Nanocapsules were obtained by combining spontaneous emulsification and interfacial polycondensation reaction between sebacoyl chloride and diethylenetriamine. Nano-emulsions used as control were obtained by the same process without monomers. The influence of carrier on release rate was studied in vitro with a membrane-free model. Epidermal penetration of encapsulated sunscreens was ex vivo evaluated using Franz diffusion cells. Ability of encapsulated sunscreens to improve photo-stability was verified by comparing percentage of degradation after UV radiation exposure.

Results

Sunscreen-containing nanocapsules (260–400 nm) were successfully prepared; yield of encapsulation was >98%. Parsol®MCX and Parsol®1789 encapsulation led to decreased release rate by up to 60% in comparison with nano-emulsion and allowed minimum penetration through pig ear epidermis. Presence of polyamide shell protected encapsulated sunscreen filters from photo-degradation without affecting their activity.

Conclusions

Encapsulation of Parsol®MCX and Parsol®1789 into oil-core of polyamide nanocapsules allowed protection from photo-degradation, controlled release from nanocapsules, and limited penetration through pig ear epidermis.

Pharmaceutical Research: A Semi-mechanistic Modeling Strategy for Characterization of Regional Absorption Properties and Prospective Prediction of Plasma Concentrations Following Administration of New Modified Release Formulations

Bergstrand; Söderlind; Eriksson; Weitschies; Karlsson — 2012-02-01

Type: Original Paper
Purpose

To outline and test a new modeling approach for prospective predictions of absorption from newly developed modified release formulations based on in vivo studies of gastro intestinal (GI) transit, drug release and regional absorption for the investigational drug AZD0837.

Methods

This work was a natural extension to the companion article “A semi-mechanistic model to link in vitro and in vivo drug release for modified release formulations”. The drug release model governed the amount of substance released in distinct GI regions over time. GI distribution of released drug substance, region specific rate and extent of absorption and the influence of food intake were estimated. The model was informed by magnetic marker monitoring data and data from an intubation study with local administration in colon.

Results

Distinctly different absorption properties were characterized for different GI regions. Bioavailability over the gut-wall was estimated to be high in duodenum (70%) compared to the small intestine (25%). Colon was primarily characterized by a very slow rate of absorption.

Conclusions

The established model was largely successful in predicting plasma concentration following administration of three newly developed formulations for which no clinical data had been applied during model building.

Pharmaceutical Research: Preclinical Pharmacokinetics and Biodistribution of Human Papillomavirus DNA Vaccine Delivered in Human Endogenous Retrovirus Envelope-Coated Baculovirus Vector

Cho; Lee; Im; Kim; Lee; Lee; Lee; Kim; Kim; Oh — 2012-02-01

Type: Original Paper
Purpose

Test pharmacokinetics and biodistribution of a human papillomavirus(HPV)16L1 DNA vaccine delivered in human endogenous retrovirus envelope protein (HERV)-expressing baculovirus (AcHERV) and those of naked plasmid vaccine.

Method

HPV16L1 gene was administrated as a naked plasmid or in AcHERV to mice via intravenous and intramuscular routes. HPV16L1 gene was extracted and assayed by quantitative real-time polymerase chain reaction, which was determined to have a detection limit of 50 copies/µg genomic DNA..

Results

Mean residence times of HPV16L1 in AcHERV were 4.8- and 272.2-fold higher than naked HPV16L1 DNA vaccines after intramuscular and intravenous administration, respectively. Naked HPV16L1 DNA levels 1 month after injection were >3 orders of magnitude lower in each tissue tested than AcHERV-delivered HPV16L1, which was retained in most tissues without specific tissue tropism. AcHERV-delivered HPV16L1 administered intramuscularly persisted at the injection sites. However, the levels of copy numbers in muscle were low (1,800/μg genomic DNA) after 1 month, and undetectable after 6 months.

Conclusions

HPV16L1 delivered via AcHERV resides longer in the body than HPV16L1 in naked form. The lack of tissue tropism ensures the safety of AcHERV vectors for further development.

Pharmaceutical Research: Discrimination Between Silicone Oil Droplets and Protein Aggregates in Biopharmaceuticals: A Novel Multiparametric Image Filter for Sub-visible Particles in Microflow Imaging Analysis

Strehl; Rombach-Riegraf; Diez; Egodage; Bluemel; Jeschke; Koulov — 2012-02-01

Type: Original Paper
Purpose

Accurate monitoring of the sub-visible particle load in protein biopharmaceuticals is increasingly important to drug development. Manufacturers are expected to characterize and control sub-visible protein particles in their products due to their potential immunogenicity. Light obscuration, the most commonly used analytical tool to count microscopic particles, does not allow discrimination between potentially harmful protein aggregates and harmless pharmaceutical components, e.g. silicone oil, commonly present in drug products. Microscopic image analysis in flow-microscopy techniques allows not only counting, but also classification of sub-visible particles based on morphology. We present a novel approach to define software filters for analysis of particle morphology in flow-microscopic images enhancing the capabilities of flow-microscopy.

Methods

Image morphology analysis was applied to analyze flow-microscopy data from experimental test sets of protein aggregates and silicone oil suspensions.

Results

A combination of four image morphology parameters was found to provide a reliable basis for automatic distinction between silicone oil droplets and protein aggregates in protein biopharmaceuticals resulting in low misclassification errors.

Conclusions

A novel, custom-made software filter for discrimination between proteinaceous particles and silicone oil droplets in flow-microscopy imaging analysis was successfully developed.

Pharmaceutical Research: Novel In Vitro-In Vivo Extrapolation (IVIVE) Method to Predict Hepatic Organ Clearance in Rat

Umehara; Camenisch — 2012-02-01

Type: Original Paper
Purpose

Drug elimination in the liver consists of uptake, metabolism, biliary excretion, and sinusoidal efflux from the hepatocytes to the blood. We aimed to establish an accurate prediction method for liver clearance in rats, considering these four elimination processes. In vitro assays were combined to achieve improved predictions.

Methods

In vitro clearances for uptake, metabolism, biliary excretion and sinusoidal efflux were determined for 13 selected compounds with various physicochemical and pharmacokinetic properties. Suspended hepatocytes, liver microsomes and sandwich-cultured hepatocytes were evaluated as in vitro models. Based on the individual processes, in vivo hepatic clearance was calculated. Subsequently, the predicted clearances were compared with the corresponding in vivo values from literature.

Results

Using this in vitro-in vivo extrapolation method good linear correlation was observed between predicted and reported clearances. Linear regression analysis revealed much improved prediction for the novel method (r² = 0.928) as compared to parameter analysis using hepatocyte uptake only (r² = 0.600), microsomal metabolism only (r² = 0.687) or overall hepatobiliary excretion in sandwich-cultured hepatocytes (r² = 0.321).

Conclusions

In this new attempt to predict hepatic elimination under consideration of multiple clearance processes, in vivo hepatic clearances of 13 compounds in rats were well predicted using an IVIVE analysis method based on in vitro assays.

Pharmaceutical Research: Nose-to-Brain Transport Pathways of Wheat Germ Agglutinin Conjugated PEG-PLA Nanoparticles

Liu; Shen; Chen; Gao; Feng; Wang; Zhang; Jiang — 2012-02-01

Type: Original Paper
Purpose

To investigate the possible pathways for transport of wheat germ agglutinin conjugated PEG-PLA nanoparticles (WGA-NP) into the brain after nasal administration.

Methods

The nose-to-brain pathways were investigated using WGA-NP containing 6-coumarin (as a fluorescent marker) and ¹²⁵I-labeled WGA-NP. Ex vivo imaging analysis was also employed to visualize the transport process.

Results

Nasal administration of WGA-NP to rats resulted in transcellular absorption across the olfactory epithelium and transfer to the olfactory bulb within 5 min. After entering the lamina propria, a proportion of WGA-NP were transferred from the olfactory nerve bundles and their surrounding connective tissue to the olfactory bulb. The trigeminal nerves also contributed to WGA-NP brain transfer, especially to WGA-NP distribution in the caudal brain areas. However, cerebrospinal fluid pathway may have little contribution to the process of transferring WGA-NP into the central nervous system (CNS) after intranasal administration.

Conclusions

These results demonstrated that intranasally administered WGA-NP reach the CNS via olfactory pathway and trigeminal nerve pathway, and extracellular transport along these nerves is the most possible mechanism.

Pharmaceutical Research: AAPS Connection

2012-02-01

Type: News
No abstract available.

Pharmaceutical Research: Comparing MDI and DPI Aerosol Deposition Using In Vitro Experiments and a New Stochastic Individual Path (SIP) Model of the Conducting Airways

Longest; Tian; Walenga; Hindle — 2012-01-31

Type: Original Paper
Purpose

Deposition characteristics of MDI and DPI aerosols were compared throughout the conducting airways for the first time using a combination of in vitro experiments and a newly developed stochastic individual path (SIP) model for different inhalation profiles.

Methods

In vitro experiments were used to determine initial particle distribution profiles and to validate computational fluid dynamics (CFD) model results for a MDI and DPI delivering the same dose of drug in a geometry of the mouth-throat and tracheobronchial airways. The validated CFD model was then used to predict the transport and deposition of the drug using correct and incorrect inhalation profiles for each inhaler.

Results

The MDI delivered approximately two times more drug to the tracheobronchial region compared with the DPI for both correct and incorrect inhalation profiles. Errors in inhalation reduced the deposited tracheobronchial dose by approximately 30% for both inhalers. The DPI delivered the largest dose to the mouth-throat (~70%) and the MDI delivered the largest dose to the alveolar airways (~50%).

Conclusions

The developed in silico model provides new insights into the lung delivery of pharmaceutical aerosols and can be applied in future studies in combination with pharmacokinetic analysis to establish bioequivalence between devices.

Pharmaceutical Research: A Bodyweight-Dependent Allometric Exponent for Scaling Clearance Across the Human Life-Span

2012-01-28

Type: Original Paper
Purpose

To explore different allometric equations for scaling clearance across the human life-span using propofol as a model drug.

Methods

Data from seven previously published propofol studies ((pre)term neonates, infants, toddlers, children, adolescents and adults) were analysed using NONMEM VI. To scale clearance, a bodyweight-based exponential equation with four different structures for the exponent was used: (I) 3/4 allometric scaling model; (II) mixture model; (III) bodyweight-cut-point separated model; (IV) bodyweight-dependent exponent model.

Results

Model I adequately described clearance in adults and older children, but overestimated clearance of neonates and underestimated clearance of infants. Use of two different exponents in Model II and Model III showed significantly improved performance, but yielded ambiguities on the boundaries of the two subpopulations. This discontinuity was overcome in Model IV, in which the exponent changed sigmoidally from 1.35 at a hypothetical bodyweight of 0 kg to a value of 0.56 from 10 kg onwards, thereby describing clearance of all individuals best.

Conclusions

A model was developed for scaling clearance over the entire human life-span with a single continuous equation, in which the exponent of the bodyweight-based exponential equation varied with bodyweight.

The AAPS Journal: A mechanism-Based Approach for Absorption Modeling: The Gastro-Intestinal Transit Time (GITT) Model

Hénin; Bergstrand; Standing; Karlsson — 2012-01-28

Type: Original Paper

Absorption models used in the estimation of pharmacokinetic drug characteristics from plasma concentration data are generally empirical and simple, utilizing no prior information on gastro-intestinal (GI) transit patterns. Our aim was to develop and evaluate an estimation strategy based on a mechanism-based model for drug absorption, which takes into account the tablet movement through the GI transit. This work is an extension of a previous model utilizing tablet movement characteristics derived from magnetic marker monitoring (MMM) and pharmacokinetic data. The new approach, which replaces MMM data with a GI transit model, was evaluated in data sets where MMM data were available (felodipine) or not available (diclofenac). Pharmacokinetic profiles in both datasets were well described by the model according to goodness-of-fit plots. Visual predictive checks showed the model to give superior simulation properties compared with a standard empirical approach (first-order absorption rate + lag-time). This model represents a step towards an integrated mechanism-based NLME model, where the use of physiological knowledge and in vitro–in vivo correlation helps fully characterize PK and generate hypotheses for new formulations or specific populations.

The AAPS Journal: Structure-Based Virtual Screening for Drug Discovery: a Problem-Centric Review

Cheng; Li; Zhou; Wang; Bryant — 2012-01-27

Type: Review Paper

Structure-based virtual screening (SBVS) has been widely applied in early-stage drug discovery. From a problem-centric perspective, we reviewed the recent advances and applications in SBVS with a special focus on docking-based virtual screening. We emphasized the researchers’ practical efforts in real projects by understanding the ligand-target binding interactions as a premise. We also highlighted the recent progress in developing target-biased scoring functions by optimizing current generic scoring functions toward certain target classes, as well as in developing novel ones by means of machine learning techniques.

AAPS PharmSciTech: Cavamax W7 Composite Ethosomal Gel of Clotrimazole for Improved Topical Delivery: Development and Comparison with Ethosomal Gel

Akhtar; Pathak — 2012-01-27

Type: Original Paper

The present research work was aimed to formulate clotrimazole encapsulated Cavamax W7 composite ethosomes by injection method for improved delivery across epidermis. 3² factorial design was used to design nine formulations (F1-F9) and compared with ethosomal formulations (F10-F12). F9 with vesicle size of 202.8 ± 4.8 nm, highest zeta potential (−83.6 ± 0.96 mV) and %EE of 98.42 ± 0.15 was selected as optimized composite ethosome and F12 as reference ethosomal formulation. As revealed by transmission electron microscopy F9 vesicles were more condensed, uniformly spherical in shape than F12 vesicles. Vesicular stability studies indicated F9 to be more stable as compared to F12. Both F9 and F12 were incorporated in carbopol 934 gel base to get G1–G8 gel formulations and evaluated for in vitro skin permeability. Cavamax W7 composite ethosomal optimized gel (G5) showed higher in vitro percent cumulative drug permeation (88.53 ± 2.10%) in 8 h and steady state flux (J_ss) of 3.39 ± 1.45 μg/cm²/min against the J_ss of 1.57 ± 0.23 μg/cm²/min for ethosomal gel (G1) and 1.13 ± 0.06 μg/cm²/min for marketed formulation. The J_ss flux of G5 was independent of amount of drug applied/unit area of skin. In vivo confocal laser scanning microscopic study of G5 depicted uniform and deeper penetration of rhodamine B (marker) in epidermis from Cavamax W7 composite ethosomal gel in comparison to G1. Finally, G5 demonstrated better (p < 0.05) antifungal activity against Candida albicans and Aspergillus niger than G1 thus, signifying that Cavamax W7 composite ethosomes present a superior stable and efficacious vesicular system than ethosomal formulation for topical delivery of clotrimazole.

Pharmaceutical Research: Tanshinones from Chinese Medicinal Herb Danshen (Salvia miltiorrhiza Bunge) Suppress Prostate Cancer Growth and Androgen Receptor Signaling

Zhang; Won; Jiang; Lee; Jeong; Lee; Zhang; Ye; Kim; Lü — 2012-01-27

Type: Original Paper
Purpose

To test whether tanshinones inhibit prostate cancer (PCa) growth at least in part through inhibiting androgen receptor (AR) signaling.

Methods

We evaluated cell growth, survival and AR signaling parameters of PCa cells after exposure to tanshinones in in vitro models. We also tested the in vivo inhibitory efficacy of tanshinone IIA (TIIA) against LNCaP xenograft model in athymic nude mice.

Results

For androgen-dependent LNCaP cells, a colony growth assay showed strong inhibitory potency following the order of TIIA≈cryptotanshinone>tanshinone I, being 10–30 folds higher than Casodex (racemic). TIIA inhibited growth of LNCaP cells more than several androgen-independent PCa cell lines. All 3 tested tanshinones were devoid of AR agonist activity under castrate condition. Mechanistically, tanshinones inhibited AR nuclear translocation within 2 h, decreased protein and mRNA abundance of AR and its target prostate-specific antigen within 12 h, and stimulated proteosomal degradation of AR. Oral administration of TIIA (25 mg/kg, once daily) retarded LNCaP xenograft growth and down-regulated tumor AR abundance in athymic nude mice.

Conclusion

AR targeting action of tanshinones was distinct from Casodex and contributed to prostate cancer growth suppression in vitro and in vivo.

Pharmaceutical Research: Multi-Compartmental Nanoparticles-in-Emulsion Formulation for Macrophage-Specific Anti-Inflammatory Gene Delivery

Attarwala; Amiji — 2012-01-27

Type: Original Paper
Purpose

To develop a safe and effective non-viral vector for gene delivery and transfection in macrophages for potential anti-inflammatory therapy.

Methods

Solid nanoparticles-in-emulsion (NiE) multi-compartmental delivery system was designed using plasmid DNA-encapsulated type B gelatin nanoparticles suspended in the inner aqueous phase of safflower oil-containing water-in-oil-in-water (W/O/W) multiple emulsion. Control and NiE formulations were evaluated for DNA delivery and transfection efficiency in J774A.1 adherent murine macrophages.

Results

Using green fluorescent protein (GFP) and murine interleukin-10 (mIL-10) expressing plasmid DNA constructs, the NiE formulation was found superior in enhancing intracellular delivery and gene transfection efficiency in cells. Anti-inflammatory effects of transfected mIL-10 were examined by suppression of tumor necrosis factor-alpha (TNFα) and interleukin 1-beta (IL-1β) production in lipopolysaccharide (LPS)-stimulated cells.

Conclusions

Overall, the results were very encouraging towards development of a macrophage-specific NiE-based multi-compartmental gene delivery strategy that can potentially affect a number of acute and chronic inflammatory diseases.

Pharmaceutical Research: Use of Partial AUC to Demonstrate Bioequivalence of Zolpidem Tartrate Extended Release Formulations

Lionberger; Raw; Kim; Zhang; Yu — 2012-01-26

Type: Original Paper
Purpose

FDA’s bioequivalence recommendation for Zolpidem Tartrate Extended Release Tablets is the first to use partial AUC (pAUC) metrics for determining bioequivalence of modified-release dosage forms. Modeling and simulation studies were performed to aid in understanding the need for pAUC measures and also the proper pAUC truncation times.

Methods

Deconvolution techniques, In Vitro/In Vivo Correlations, and the CAT (Compartmental Absorption and Transit) model were used to predict the PK profiles for zolpidem. Models were validated using in-house data submitted to the FDA. Using dissolution profiles expressed by the Weibull model as input for the CAT model, dissolution spaces were derived for simulated test formulations.

Results

The AUC_0–1.5 parameter was indicative of IR characteristics of early exposure and effectively distinguished among formulations that produced different pharmacodynamic effects. The AUC_1.5-t parameter ensured equivalence with respect to the sustained release phase of Ambien CR. The variability of AUC_0–1.5 is higher than other PK parameters, but is reasonable for use in an equivalence test.

Conclusions

In addition to the traditional PK parameters of AUCinf and Cmax, AUC_0-1.5 and AUC_1.5-t are recommended to provide bioequivalence measures with respect to label indications for Ambien CR: onset of sleep and sleep maintenance.

AAPS PharmSciTech: Polymeric Micelles of PEG-PE as Carriers of All-Trans Retinoic Acid for Stability Improvement

Wichit; Tangsumranjit; Pitaksuteepong; Waranuch — 2012-01-25

Type: Original Paper

The topical application of all-trans retinoic acid (ATRA) is an effective treatment for several skin disorders, including photo-aging. Unfortunately, ATRA is susceptible to light, heat, and oxidizing agents. Thus, this study aimed to investigate the ability of polymeric micelles prepared from polyethylene glycol conjugated phosphatidylethanolamine (PEG-PE) to stabilize ATRA under various storage conditions. ATRA entrapped in polymeric micelles with various PEG and PE structures was prepared. The critical micelle concentrations were 97–243 μM, depending on the structures of the PEG and PE molecules. All of the micelles had particle diameters of 6–20 nm and neutral charges. The highest entrapment efficiency (82.7%) of the tested micelles was exhibited by ATRA in PEG with a molecular weight of 750 Da conjugated to dipalmitoyl phosphatidylethanolamine (PEG₇₅₀-DPPE) micelles. The PEG₇₅₀-DPPE micelle could significantly retard ATRA oxidation compared to ATRA in 75% methanol/HBS solution. Up to 87% of ATRA remained in the PEG₇₅₀-DPPE micelle solution after storage in ambient air for 28 days. This result suggests that PEG₇₅₀-DPPE micelle can improve ATRA stability. Therefore, ATRA in PEG₇₅₀-DPPE micelle is an interesting alternative structure for the development of cosmeceutical formulations.

Pharmaceutical Research: Targeted Endothelial Gene Delivery by Ultrasonic Destruction of Magnetic Microbubbles Carrying Lentiviral Vectors

2012-01-25

Type: Original Paper
Purpose

Site specific vascular gene delivery is a promising tool for treatment of cardiovascular diseases. By combining ultrasound mediated microbubble destruction with site specific magnetic targeting of lentiviruses, we aimed to develop a technique suitable for systemic application.

Methods

The magnetic nanoparticle coupling to lipid microbubbles was confirmed by absorbance measurements. Association of fluorescent lentivirus to magnetic microbubbles (MMB) was determined by microscopy and flow cytometry. Functionality and efficiency of GFP-encoding lentiviral MMB transduction was evaluated by endothelial (HMEC) GFP expression and cytotoxicity was measured by MTT reduction.

Results

Microbubbles with a mean diameter of 4.3 ± 0.04 μm were stable for 2 days, readily magnetizable and magnetically steerable in vitro and efficiently associated with lentivirus. Exposure of eGFP-encoding lentiviral MMB to human endothelial cells followed by application of an external static magnetic field (30 min) and ultrasonic destruction of the microbubbles did not markedly affect cellular viability. Finally, this combination led to a 30-fold increase in transduction efficiency compared to application of naked virus alone.

Conclusions

By associating microbubbles with magnetic iron nanoparticles, these function as carriers for lentiviruses achieving tissue specific deposition at the site of interest.

Pharmaceutical Research: Magnetic Nanoparticles for Cancer Diagnosis and Therapy

Yigit; Moore; Medarova — 2012-01-25

Type: Original Paper

Nanotechnology is evolving as a new field that has a potentially high research and clinical impact. Medicine, in particular, could benefit from nanotechnology, due to emerging applications for noninvasive imaging and therapy. One important nanotechnological platform that has shown promise includes the so-called iron oxide nanoparticles. With specific relevance to cancer therapy, iron oxide nanoparticle-based therapy represents an important alternative to conventional chemotherapy, radiation, or surgery. Iron oxide nanoparticles are usually composed of three main components: an iron core, a polymer coating, and functional moieties. The biodegradable iron core can be designed to be superparamagnetic. This is particularly important, if the nanoparticles are to be used as a contrast agent for noninvasive magnetic resonance imaging (MRI). Surrounding the iron core is generally a polymer coating, which not only serves as a protective layer but also is a very important component for transforming nanoparticles into biomedical nanotools for in vivo applications. Finally, different moieties attached to the coating serve as targeting macromolecules, therapeutics payloads, or additional imaging tags. Despite the development of several nanoparticles for biomedical applications, we believe that iron oxide nanoparticles are still the most promising platform that can transform nanotechnology into a conventional medical discipline.

Pharmaceutical Research: Click Chemistry with Polymers, Dendrimers, and Hydrogels for Drug Delivery

Lallana; Fernandez-Trillo; Sousa-Herves; Riguera; Fernandez-Megia — 2012-01-25

Type: Review Paper

During the last decades, great efforts have been devoted to design polymers for reducing the toxicity, increasing the absorption, and improving the release profile of drugs. Advantage has been also taken from the inherent multivalency of polymers and dendrimers for the incorporation of diverse functional molecules of interest in targeting and diagnosis. In addition, polymeric hydrogels with the ability to encapsulate drugs and cells have been developed for drug delivery and tissue engineering applications. In the long road to this successful story, pharmaceutical sciences have been accompanied by parallel advances in synthetic methodologies allowing the preparation of precise polymeric materials with enhanced properties. In this context, the introduction of the click concept by Sharpless and coworkers in 2001 focusing the attention on modularity and orthogonality has greatly benefited polymer synthesis, an area where reaction efficiency and product purity are significantly challenged. The purpose of this Expert Review is to discuss the impact of click chemistry in the preparation and functionalization of polymers, dendrimers, and hydrogels of interest in drug delivery.

The AAPS Journal: Covariate Pharmacokinetic Model Building in Oncology and its Potential Clinical Relevance

Joerger — 2012-01-25

Type: Review Paper

When modeling pharmacokinetic (PK) data, identifying covariates is important in explaining interindividual variability, and thus increasing the predictive value of the model. Nonlinear mixed-effects modeling with stepwise covariate modeling is frequently used to build structural covariate models, and the most commonly used software—NONMEM—provides estimations for the fixed-effect parameters (e.g., drug clearance), interindividual and residual unidentified random effects. The aim of covariate modeling is not only to find covariates that significantly influence the population PK parameters, but also to provide dosing recommendations for a certain drug under different conditions, e.g., organ dysfunction, combination chemotherapy. A true covariate is usually seen as one that carries unique information on a structural model parameter. Covariate models have improved our understanding of the pharmacology of many anticancer drugs, including busulfan or melphalan that are part of high-dose pretransplant treatments, the antifolate methotrexate whose elimination is strongly dependent on GFR and comedication, the taxanes and tyrosine kinase inhibitors, the latter being subject of cytochrome p450 3A4 (CYP3A4) associated metabolism. The purpose of this review article is to provide a tool to help understand population covariate analysis and their potential implications for the clinic. Accordingly, several population covariate models are listed, and their clinical relevance is discussed. The target audience of this article are clinical oncologists with a special interest in clinical and mathematical pharmacology.

Pharmaceutical Research: Optimization of Magnetic Nanoparticle-Assisted Lentiviral Gene Transfer

2012-01-25

Type: Original Paper
Purpose

Targeting of specific cells and tissues is of great interest for clinical relevant gene- and cell-based therapies. We use magnetic nanoparticles (MNPs) with a ferrimagnetic core (Fe₃O₄) with different coatings to optimize MNP-assisted lentiviral gene transfer with focus on different endothelial cell lines.

Methods

Lentiviral vector (LV)/MNP binding was characterized for various MNPs by different methods (e.g. magnetic responsiveness measurement). Transduced cells were analyzed by flow cytometry, fluorescence microscopy and iron recovery. Cell transduction and cell positioning under physiological flow conditions were performed using different in vitro and ex vivo systems.

Results

Analysis of diverse MNPs with different coatings resulted in identification of nanoparticles with improved LV association and enhanced transduction properties of complexes in several endothelial cell lines. The magnetic moments of LV/MNP complexes are high enough to achieve local gene targeting of perfused endothelial cells. Perfusion of a mouse aorta with LV/MNP transduced cells under clinically relevant flow conditions led to local cell attachment at the intima of the vessel.

Conclusion

MNP-guided lentiviral transduction of endothelial cells can be significantly enhanced and localized by using optimized MNPs.

Pharmaceutical Research: Formulation and In Vitro Characterization of Composite Biodegradable Magnetic Nanoparticles for Magnetically Guided Cell Delivery

Chorny; Alferiev; Fishbein; Tengood; Folchman-Wagner; Forbes; Levy — 2012-01-25

Type: Original Paper
Purpose

Cells modified with magnetically responsive nanoparticles (MNP) can provide the basis for novel targeted therapeutic strategies. However, improvements are required in the MNP design and cell treatment protocols to provide adequate magnetic properties in balance with acceptable cell viability and function. This study focused on select variables controlling the uptake and cell compatibility of biodegradable polymer-based MNP in cultured endothelial cells.

Methods

Fluorescent-labeled MNP were formed using magnetite and polylactide as structural components. Their magnetically driven sedimentation and uptake were studied fluorimetrically relative to cell viability in comparison to non-magnetic control conditions. The utility of surface-activated MNP forming affinity complexes with replication-deficient adenovirus (Ad) for transduction achieved concomitantly with magnetic cell loading was examined using the green fluorescent protein reporter.

Results

A high-gradient magnetic field was essential for sedimentation and cell binding of albumin-stabilized MNP, the latter being rate-limiting in the MNP loading process. Cell loading up to 160 pg iron oxide per cell was achievable with cell viability >90%. Magnetically driven uptake of MNP-Ad complexes can provide high levels of transgene expression potentially useful for a combined cell/gene therapy.

Conclusions

Magnetically responsive endothelial cells for targeted delivery applications can be obtained rapidly and efficiently using composite biodegradable MNP.

Pharmaceutical Research: Efficient siRNA Delivery Using a Polyamidoamine Dendrimer with a Modified Pentaerythritol Core

Zhang; Zhou; Kwak; Wang; Yung; Lee; Wang; Wang; Lee — 2012-01-25

Type: Original Paper
Purpose

Delivery of siRNA into cells remains a critical challenge. Our lab has shown a novel polyamidoamine (PAMAM) dendrimer with modified pentaerythritol derivative core (PD dendrimer) to exhibit high plasmid DNA transfection efficiency and low cytotoxicity. Here, we evaluate PD dendrimer as a siRNA carrier.

Methods

Agarose gel electrophoresis and AFM were used to confirm formation of generation 5 (G5)-PD dendrimer/siRNA nanoparticles (NPs). G5 PD dendrimer/anti-luciferase siRNA NPs were used to transfect SK Hep-1 cells with stable luciferase expression. Effects of various endocytic pathway inhibitors on uptake of G5 PD dendrimer/siRNA NPs in SK Hep-1 cells were also investigated.

Results

Agarose gel electrophoresis indicated that G5 PD dendrimer and siRNA formed NPs at weight ratios >0.5:1. G5 PD dendrimer showed effective luciferase gene silencing when weight ratio was 3.0:1 and above. Treatment with endocytosis inhibitors showed that clathrin-mediated endocytosis was the main endocytic pathway by which G5-PD dendrimer/siRNA NPs enter the cell.

Conclusions

These results show that the novel G5 PD dendrimer has high siRNA delivery activity and is promising as a delivery agent for its therapeutic application.

Pharmaceutical Research: PEG-Stabilized Micellar System with Positively Charged Polyester Core for Fast pH-Responsive Drug Release

Wang; Jia; Cheng; Feng; Zhang; Zhuo — 2012-01-21

Type: Original Paper
Purpose

To design functional drug carriers for fast pH-responsive drug release.

Methods

Functional diblock terpolymers of monomethoxy poly(ethylene glycol)-block- copoly(6,14-dimethyl-1,3,9,11-tetraoxa-6,14-diaza-cyclohexadecane-2,10-dione-co-ε-caprolactone) [mPEG-b-poly(ADMC-co-CL)] were fabricated via biosynthetic pathway. The self-assembled nanosphere and drug-loaded micelles of the copolymers were further prepared by dialysis method. The pH-tunable morphology variation and drug release pattern were observed at different pH.

Results

A collection of three PEGylated terpolymers with varied compositions in poly(ADMC-co-CL) block was designed with high cell-biocompatibility. The copolymers could readily self-assemble into nanoscale micelles (~ 100 nm) in aqueous medium and exhibit high stability over 80-h incubation in different mediums including deionized water, neutral NaCl solution, and heparin sodium solution. Due to the protonation-deprotonation of tertiary amine groups in ADMC units, acid-induced structural deformation of micelles was disclosed in terms of the variation in CAC value and hydrodynamic size at different pH. Drug loading efficiency was comparable to that of reported PEG-polyester micelles with specifically designed structures purposed for drug-loading improvement. Remarkably accelerated drug release triggered by acidity was distinctly detected for ibuprofen-loaded mPEG-b-poly(ADMC-co-CL) micelle system, suggesting a fast pH-responsive characteristic.

Conclusion

Functional PEG-stabilized micellar carriers with positively charged polyester core were successfully developed for fast pH-responsive drug release.

Pharmaceutical Research: Cellular Transfer of Magnetic Nanoparticles Via Cell Microvesicles: Impact on Cell Tracking by Magnetic Resonance Imaging

Silva; Wilhelm; Kolosnjaj-Tabi; Luciani; Gazeau — 2012-01-21

Type: Original Paper
Purpose

Cell labeling with magnetic nanoparticles can be used to monitor the fate of transplanted cells in vivo by magnetic resonance imaging. However, nanoparticles initially internalized in administered cells might end up in other cells of the host organism. We investigated a mechanism of intercellular cross-transfer of magnetic nanoparticles to different types of recipient cells via cell microvesicles released under cellular stress.

Methods

Three cell types (mesenchymal stem cells, endothelial cells and macrophages) were labeled with 8-nm iron oxide nanoparticles. Then cells underwent starvation stress, during which they produced microvesicles that were subsequently transferred to unlabeled recipient cells.

Results

The analysis of the magnetophoretic mobility of donor cells indicated that magnetic load was partially lost under cell stress. Microvesicles shed by stressed cells participated in the release of magnetic label. Moreover, such microvesicles were uptaken by naïve cells, resulting in cellular redistribution of nanoparticles. Iron load of recipient cells allowed their detection by MRI.

Conclusions

Cell microvesicles released under stress may be disseminated throughout the organism, where they can be uptaken by host cells. The transferred cargo may be sufficient to allow MRI detection of these secondarily labeled cells, leading to misinterpretations of the effectiveness of transplanted cells.

Pharmaceutical Research: Magnetized Aerosols Comprising Superparamagnetic Iron Oxide Nanoparticles Improve Targeted Drug and Gene Delivery to the Lung

2012-01-21

Type: Original Paper
Purpose

Targeted delivery of aerosols could not only improve efficacy of inhaled drugs but also reduce side effects resulting from their accumulation in healthy tissue. Here we investigated the impact of magnetized aerosols on model drug accumulation and transgene expression in magnetically targeted lung regions of unanesthetized mice.

Methods

Solutions containing superparamagnetic iron oxide nanoparticles (SPIONs) and model drugs (fluorescein or complexed plasmid DNA) were nebulized to unanesthetized mice under the influence of an external magnetic gradient directed to the lungs. Drug accumulation and transgene expression was subsequently measured at different time points.

Results

We could demonstrate 2–3 fold higher accumulation of the model drug fluorescein and specific transgene expression in lung regions of mice which had been exposed to an external magnetic gradient during nebulization compared to the control mice without any exposure to magnetic gradient.

Conclusions

Magnetized aerosols present themselves as an efficient approach for targeted pulmonary delivery of drugs and gene therapeutic agents in order to treat localized diseases of the deeper airways.

Pharmaceutical Research: Erythropoietin-Coated ZP-Microneedle Transdermal System: Preclinical Formulation, Stability, and Delivery

Peters; Ameri; Wang; Maa; Daddona — 2012-01-19

Type: Original Paper
Purpose

To evaluate the feasibility of coating formulated recombinant human erythropoietin alfa (EPO) on a titanium microneedle transdermal delivery system, ZP-EPO, and assess preclinical patch delivery performance.

Methods

Formulation rheology and surface activity were assessed by viscometry and contact angle measurement. EPO liquid formulation was coated onto titanium microneedles by dip-coating and drying. Stability of coated EPO was assessed by SEC-HPLC, CZE and potency assay. Preclinical in vivo delivery and pharmacokinetic studies were conducted in rats with EPO-coated microneedle patches and compared to subcutaneous EPO injection.

Results

Studies demonstrated successful EPO formulation development and coating on microneedle arrays. ZP-EPO patch was stable at 25°C for at least 3 months with no significant change in % aggregates, isoforms, or potency. Preclinical studies in rats showed the ZP-EPO microneedle patches, coated with 750 IU to 22,000 IU, delivered with high efficiency (75–90%) with a linear dose response. PK profile was similar to subcutaneous injection of commercial EPO.

Conclusions

Results suggest transdermal microneedle patch delivery of EPO is feasible and may offer an efficient, dose-adjustable, patient-friendly alternative to current intravenous or subcutaneous routes of administration.

Pharmaceutical Research: Outstanding Reviewers of 2011

2012-01-19

Type: News
No abstract available.

The AAPS Journal: Ligand Binding Assays in the 21st Century Laboratory: Platforms

2012-01-19

Type: Book Review
No abstract available.

AAPS PharmSciTech: Application of a Pull on a Disk Method to Measure Surface Tension of Liquids

Carvalho; Horng; McConville — 2012-01-19

Type: Original Paper

The intrinsic property of liquids is a vital indicator of formulation performance and stability. Therefore, investigation of the interfacial phenomenon of surface tension is a routine procedure in the development of products in a wide variety of areas including foods, pharmaceuticals, cosmetics, and painting technologies. We hypothesize that studies related to the maximum pull on a rod can be extrapolated to disk geometry and applied to measure surface tension using a texture analyzer. A glass disk probe was attached to the arm of a texture analyzer and pulled from the liquid surface. The maximum force of detachment was used to calculate surface tension extrapolating from the theory of maximum pull on a rod. The surface tension of water, ethanol, and a hydroalcoholic solution was measured and compared with literature values to validate this hypothesis. The calculated values of surface tension for the liquids studied were within 5% of the reported values. Probe diameter appears to have an important role on surface tension accuracy compared with literature values. Slight discrepancies can be attributed to temperature control and leveling of liquid surface, although still in accordance with the reported values of surface tension measured using different methods. This study presents a simple, precise, and quick method to determine the surface tension of liquids from the maximum pull on a disk. Further studies are warranted to determine the optimum glass disk probe diameter for better accuracy.

AAPS PharmSciTech: Controlled Release of Modified Insulin Glargine from Novel Biodegradable Injectable Gels

Anand; Almoazen; Mehrotra; Johnson; Shukla — 2012-01-19

Type: Original Paper

The objective of this study was to investigate the duration of biological effects of modified insulin glargine released from a novel biodegradable injectable gel in type II diabetic Zucker diabetic fatty (ZDF) rats. Modified insulin glargine was purified from the marketed formulation by process of dialysis followed by freeze-drying, and the purity was confirmed by the single peak, corresponding to insulin glargine in the HPLC chromatogram. To determine and to compare the biological activity of purified insulin glargine with marketed formulation, it was suspended in isotonic saline solutions and administered subcutaneously to ZDF rats at a dose of 10 IU/kg of insulin and the blood glucose levels were measured. The blood glucose levels of ZDF rats after a subcutaneous injection of a suspension of purified insulin glargine decreased below 200 mg/dL within 2 h and remained at this level up to 6 h, then steadily raised above 400 mg/dL in 12 h. Insulin glargine particles were loaded into a novel biodegradable injectable gel formulation prepared from a blend of polylactic-co-glycolic acid (PLGA) and biocompatible plasticizers. Approximately 0.1 mL of insulin glargine-loaded gel prepared with PLGA was administered subcutaneously to the ZDF rats, and blood glucose levels were measured. The PLGA gel formulations prepared with insulin glargine particles had duration of action of 10 days following a single subcutaneous injection. The addition of zinc sulfate to the formulations prepared with purified insulin glargine particles further slowed down the drop in blood glucose concentrations.

AAPS PharmSciTech: Transdermal Absorption Enhancement of Rice Bran Bioactive Compounds Entrapped in Niosomes

Manosroi; Chutoprapat; Abe; Manosroi; Manosroi — 2012-01-19

Type: Original Paper

Niosomes composed of Tween 61 and cholesterol at 1:1 molar ratio were entrapped with the mixture of the three semi-purified rice (Oryza sativa L., Family Gramineae) bran bioactive compounds [ferulic acid (F), γ-oryzanol (O), and phytic acid (P)] at 0.5%, 1.5%, and 1.5%, respectively, by the supercritical CO₂ technique. The transdermal absorption by vertical Franz diffusion cells of the compounds entrapped in niosomes (Nio FOP), the unentrapped compounds (Mixed FOP), the compounds incorporated in gel and cream (Gel FOP and Cream FOP), and the compounds entrapped in niosomes and incorporated in gel and cream (Gel nio and Cream nio) was investigated. At 6 h, F and P from Nio FOP gave lower cumulative amount in viable epidermis and dermis (VED) than from Mixed FOP of 1.1 and 1.6 times, respectively, while O from Nio FOP exhibited higher cumulative amount in VED than from Mixed FOP of 2.4 times. The highest cumulative amount in VED of F, O, and P were from Gel nio, Cream nio, and Mixed FOP at 1.564 ± 0.052, 15.972 ± 0.273, and 25.857 ± 0.025 ng/cm², respectively. Niosomes enhanced the transdermal absorption of the hydrophobic compound O, while retarded the hydrophilic compounds F and P indicating the less systemic risk of F and P than O when entrapped in niosomes. Thus, transdermal absorption of F, O, and P appeared to depend on niosomal size, lipophilicity of the bioactive compounds, and types of formulations. These preclinical results can be applied for the design of the clinical study of the developed rice bran niosomal topical products.

The AAPS Journal: Ligand-Binding Assays in the 21st Century Laboratory: Recommendations for an Automated Data Interchange Process

Leung; Usansky; Lynde; Thway; Hendricks; Rusnak — 2012-01-14

Type: Editorial Notes
No abstract available.

AAPS PharmSciTech: Biotherapeutic Formulation Factors Affecting Metal Leachables from Stainless Steel Studied by Design of Experiments

Zhou; Evans; Schöneich; Singh — 2012-01-13

Type: Original Paper

Trace amounts of metals are inevitably present in biotherapeutic products. They can arise from various sources. The impact of common formulation factors such as protein concentration, antioxidant, metal chelator concentration and type, surfactant, pH, and contact time with stainless steel on metal leachables was investigated by a design of experiments approach. Three major metal leachables, iron, chromium, and nickel were monitored by inductively coupled plasma-mass spectrometry. It was observed that among all the tested factors, contact time, metal chelator concentration, and protein concentration were statistically significant factors with higher temperature resulting in higher levels of leached metals. Within a pH range of 5.5–6.5, solution pH played a minor role for chromium leaching at 25°C. No statistically significant difference was observed due to type of chelator, presence of antioxidant, or surfactant. In order to optimize a biotherapeutic formulation to achieve a target drug product shelf life with acceptable quality, each formulation component must be evaluated for its impact.

AAPS PharmSciTech: Fabrication of Carvedilol Nanosuspensions Through the Anti-Solvent Precipitation–Ultrasonication Method for the Improvement of Dissolution Rate and Oral Bioavailability

Liu; Xu; Tian; Yuan; Pan; Ma; Yang; Pan — 2012-01-13

Type: Original Paper

The present study aims to prepare carvedilol (CAR) nanosuspensions using the anti-solvent precipitation–ultrasonication technique to improve its dissolution rate and oral bioavailability. Alpha-tocopherol succinate (VES) was first used as a co-stabilizer to enhance the stability of the nanosuspensions. The effects of the process parameters on particle size of the nanosuspensions were investigated. The optimal values of the precipitation temperature, power inputs, and the time length of ultrasonication were selected as 10°C, 400 W, and 15 min, respectively. Response surface methodology based on central composite design was utilized to evaluate the formulation factors that affect the size of nanosuspensions, i.e., the concentration of CAR and VES in the organic solution, and the level of sodium dodecyl sulfate in the anti-solvent phase, respectively. The optimized formulation showed a mean size of 212 ± 12 nm and a zeta potential of −42 ± 3 mV. Scanning electron microscopy revealed that the nanosuspensions were flaky-shaped. Powder X-ray diffraction and differential scanning calorimetry analysis confirmed that the nanoparticles were in the amorphous state. Fourier transform infrared analysis demonstrated that the reaction between CAR and VES is probably due to hydrogen bonding. The nanosuspension was physically stable at 25°C for 1 week, which allows it to be further processing such as drying. The dissolution rate of the nanosuspensions was markedly enhanced by reducing the size. The in vivo test demonstrated that the C_max and AUC_0–36 values of nanosuspensions were approximately 3.3- and 2.9-fold greater than that of the commercial tablets, respectively.

Pharmaceutical Research: Leveraging Physiological Data from Literature into a Pharmacokinetic Model to Support Informative Clinical Study Design in Pregnant Women

Hasselt; Green; Morrish — 2012-01-13

Type: Original Paper
Purpose

Physiological changes during pregnancy can effect pharmacokinetic (PK) parameters, which may lead to altered dose requirements. We aimed to leverage literature-based physiological changes during pregnancy into a PK model and compare its performance to a published reference model in pregnant women and to use the literature-based model to determine informative PK sampling times for a clinical study that aims to quantify the PK of enoxaparin throughout pregnancy.

Methods

Changes in total body water (BW) and creatinine clearance (CRCL) during pregnancy were described using regression models. BW and CRCL were linked to a PK model of enoxaparin in non-pregnant women. Performance of the literature-based PK model was compared to a previously published empirical reference model. D-optimal sampling times were determined and evaluated for literature-based and reference models.

Results

The literature-based model adequately predicted anti-Xa plasma concentrations when compared to reference model predictions. An informative sampling design was succesfully developed, with parameters expected with good precision (RSE < 36.4%).

Conclusion

A literature-based model describing enoxaparin PK during pregnancy was developed and evaluated. The modelling framework could be used to support development of informative designs in pregnancy when prior models are unavailable.

Pharmaceutical Research: Force Dependent Internalization of Magnetic Nanoparticles Results in Highly Loaded Endothelial Cells for Use as Potential Therapy Delivery Vectors

MacDonald; Barbee; Polyak — 2012-01-11

Type: Original Paper
Purpose

To investigate the kinetics, mechanism and extent of MNP loading into endothelial cells and the effect of this loading on cell function.

Methods

MNP uptake was examined under field on/off conditions, utilizing varying magnetite concentration MNPs. MNP-loaded cell viability and functional integrity was assessed using metabolic respiration, cell proliferation and migration assays.

Results

MNP uptake in endothelial cells significantly increased under the influence of a magnetic field versus non-magnetic conditions. Larger magnetite density of the MNPs led to a higher MNP internalization by cells under application of a magnetic field without compromising cellular respiration activity. Two-dimensional migration assays at no field showed that higher magnetite loading resulted in greater cell migration rates. In a three-dimensional migration assay under magnetic field, the migration rate of MNP-loaded cells was more than twice that of unloaded cells and was comparable to migration stimulated by a serum gradient.

Conclusions

Our results suggest that endothelial cell uptake of MNPs is a force dependent process. The in vitro assays determined that cell health is not adversely affected by high MNP loadings, allowing these highly magnetically responsive cells to be potentially beneficial therapy (gene, drug or cell) delivery systems.

Pharmaceutical Research: Rectal Absorption of Vigabatrin, a Substrate of the Proton Coupled Amino Acid Transporter (PAT1, Slc36a1), in Rats

Holm; Kall; Frølund; Nielsen; Jensen; Broberg; Nielsen — 2012-01-11

Type: Original Paper
Purpose

To investigate the rectal absorption of vigabatrin in rats, based on the hypothesis that PAT1 (Slc36a1) is involved.

Methods

Male Sprague–Dawley rats were dosed rectally with five different gels, varying in buffer capacity, the amount of vigabatrin, and co-administration of proline or tryptophan. Western blotting was used to detect rPAT1 in rat rectal epithelium. X. Laevis oocytes were injected with SLC36A1 cRNA for the expression of hPAT1, prior to two-electrode voltage clamp measurements.

Results

rPAT1 protein was present in rat rectal epithelium. Approximately 7%–9% of a 1 mg/kg vigabatrin dose was absorbed after rectal administration, regardless of the formulation used. Increasing the dose of vigabatrin 10-fold decreased the absolute bioavailability to 4.2%. Co-administration of proline or tryptophan changed the pharmacokinetic profile, indicating a role of PAT1 in the rectal absorption of vigabatrin. Transport of vigabatrin via hPAT1 expressed in X. Laevis oocytes had a K_m of 5.2 ± 0.6 mM and was almost completely inhibited by tryptophan.

Conclusions

Although vigabatrin is a PAT1 substrate and the rPAT1 protein is expressed in the rectum epithelium, vigabatrin has low rectal absorption in rats.

AAPS PharmSciTech: A Novel Synergistic Galactomannan-Based Unit Dosage Form for Sustained Release of Acarbose

Kumar; Sinha — 2012-01-11

Type: Original Paper

In the current study, the potential of a novel combination of a galactomannan with acarbose (100 mg) was evaluated for attaining a desired hypoglycaemic effect over a prolonged period of time. Three major antidiabetic galactomannans viz., fenugreek gum, boswellia gum, and locust bean gum were selected in order to achieve a synergistic effect in the treatment alongwith retardation in drug release. In vitro studies indicated that batches containing various proportions of fenugreek gum (AF40-60) were able to control drug release for a longer duration of approximately 10–12 h. In contrast, the matrices prepared using boswellia and locust bean gum were able to sustain the release for relatively shorter durations. Drug release mainly followed first-order release kinetics owing to the highly soluble nature of the drug. In vivo study depicted a significant reduction (p < 0.001) in the postprandial blood glucose and triglyceride levels in the diabetic rats on treatment with formulation AF40. Thus, the developed system provides a better control of the postprandial glycaemic levels and it also obviates the need of conventional multiple dosing of acarbose. Furthermore, it also reduces the occurrence of side effects like diarrhea and loss of appetite.

AAPS PharmSciTech: Nanostructured Lipid Carriers Improve Skin Permeation and Chemical Stability of Idebenone

Li; Ge — 2012-01-11

Type: Original Paper

Idebenone (IDB) is a synthetic antioxidant and analog of coenzyme Q10. The percutaneous permeation of IDB was investigated in guinea pig skin after application of different formulations. The enhancing effects of various formulations [nanostructured lipid carriers (NLCs), nanoemulsion (NE), or oil solution] on the permeation of IDB were evaluated using ex vivo guinea pig skins. Furthermore, stability of different formulations and in which chemical stability of IDB was determined during storage. Permeation experiments revealed that formulations varied in their ability to enhance the skin permeation of IDB. For NLC formulation, the cumulative amount of IDB in the epidermis, dermis, and acceptor medium of diffusion cells was approximately threefold more than NE or oil solution at the end of 24-h experiment. No significant difference between NE and oil solution was observed in the enhancement of penetration efficacy of IDB. Different formulations resulted in stability with different properties. NLC formulation revealed preferentially more stable than NE. The residual percentage of IDB loaded in NLCs, NE, and oil solution was 90.1%, 65.4%, and 51.3%, respectively, when stored at 40°C under 75% RH and 3,000 lx light conditions for 180 days. The results obtained here demonstrated that the abilities of NLCs to improve the chemical stability of IDB and enhance the skin permeation are much better than NE and oil solution. These suggest that NLCs containing IDB have significant potential use for skin care as an alternative topical formulation.

Pharmaceutical Research: AAPS Connection

2012-01-10

Type: News
No abstract available.

Pharmaceutical Research: Targeted Delivery of Proteins into the Central Nervous System Mediated by Rabies Virus Glycoprotein-Derived Peptide

Fu; Wang; Zhan; Zhou — 2012-01-10

Type: Original Paper
Purpose

Delivery of therapeutic proteins across the blood-brain barrier (BBB) is severely limited by their size and biochemical properties. Here we showed that a 39-amino acid peptide derived from the rabies virus glycoprotein (RDP) was exploited as an efficient protein carrier for brain-targeting delivery.

Methods

Three proteins with different molecular weight and pI, β-galactosidase (β-Gal), luciferase (Luc) and brain-derived neurotrophic factor (BDNF), were fused to RDP and intravenously injected into the mice respectively. The slices of different tissues with X-Gal staining were used to examine whether RDP could deliver β-Gal targeted into the CNS. The time-course relationship of RDP-Luc was studied to confirm the transport efficiency of RDP. The neuroprotective function of RDP-BDNF was examined in mouse experimental stroke to explore the pharmacological effect of RDP fusion protein.

Results

The results showed that the fusion proteins rapidly and specific entered the nerve cells in 15 min, and the t_1/2 was about 1 hr. Furthermore, RDP-BDNF fusion protein showed the neuroprotective properties in mouse experimental stroke including reduction of stroke volume and neural deficit.

Conclusions

RDP provides an effective approach for the targeted delivery of biological active proteins into the central nervous system.

AAPS PharmSciTech: Improvement of Tablet Coating Uniformity Using a Quality by Design Approach

2012-01-10

Type: Original Paper

A combination of analytical and statistical methods is used to improve a tablet coating process guided by quality by design (QbD) principles. A solid dosage form product was found to intermittently exhibit bad taste. A suspected cause was the variability in coating thickness which could lead to the subject tasting the active ingredient in some tablets. A number of samples were analyzed using a laser-induced breakdown spectroscopy (LIBS)-based analytical method, and it was found that the main variability component was the tablet-to-tablet variability within a lot. Hence, it was inferred that the coating process (performed in a perforated rotating pan) required optimization. A set of designed experiments along with response surface modeling and kriging method were used to arrive at an optimal set of operating conditions. Effects of the amount of coating imparted, spray rate, pan rotation speed, and spray temperature were characterized. The results were quantified in terms of the relative standard deviation of tablet-averaged LIBS score and a coating variability index which was the ratio of the standard deviation of the tablet-averaged LIBS score and the weight gain of the tablets. The data-driven models developed based on the designed experiments predicted that the minimum value of this index would be obtained for a 6% weight gain for a pan operating at the highest speed at the maximum fill level while using the lowest spraying rate and temperature from the chosen parametric space. This systematic application of the QbD-based method resulted in an enhanced process understanding and reducing the coating variability by more than half.

AAPS PharmSciTech: The Influence of the Polar Head and the Hydrophobic Chain on the Skin Penetration Enhancement Effect of Poly(Ethylene Glycol) Derivatives

Casiraghi; Grigoli; Cilurzo; Gennari; Rossoni; Minghetti — 2012-01-10

Type: Original Paper

The effect of a homologue series of nonionic surfactants, namely poly(ethylene glycol) (PEG) fatty acid esters, differing in oxyethylene (PEG 8, PEG 12, and PEG 40) and fatty acid (stearate, mono and di-laurate, and mono and di-oleate) chain lengths, on in vitro skin permeability of ketoprofen (KTP) vehicled in plasters was investigated. The drug diffusion through hairless mouse skin as well as the effect of the surfactant type and strength was studied by Franz diffusion cells and ATR-FTIR spectroscopy. The use of PEG stearate series revealed that the surfactant with the largest polar head, namely PEG 40, was ineffective in enhancing the skin permeation of KTP, independently of the plaster concentrations. The effect of the hydrophobic chain was investigated only by using the shortest oxyethylene chains. The experimental results revealed that the oxyethylene chain length of surfactants appeared to be more influent than the alkyl chain. The prediction of the absorption enhancing capability of these PEG derivatives appeared related to the vehicle other than the proper combination of the number of ethylene oxide groups and alkyl groups.

AAPS PharmSciTech: Rice Germ Oil as Multifunctional Excipient in Preparation of Self-Microemulsifying Drug Delivery System (SMEDDS) of Tacrolimus

Pawar; Vavia — 2012-01-10

Type: Original Paper

Surmounting the constraints of limited solubilization efficiency and prime requisite of antioxidant for conventional lipid formulations, the research work explores an edge over formulation utilizing potential applicability of rice germ oil (RGO) as a multifunctional excipient. Self-microemulsifying drug delivery system (SMEDDS) of tacrolimus (TAC) was formulated with RGO, an indigenous source of gamma-oryzanol. Being the same biological source, RGO and rice bran oil (RBO) were compared and it was found that RGO have more solubilization potential for TAC (2.2-fold) as well as higher antioxidant activity (8.06-fold) than the RBO. TAC-SMEDDS was prepared using RGO/Capmul PG8 (2:3) as an oil phase, Cremophore EL as a surfactant, and Transcutol P as a cosurfactant. The approximate particle size of TAC-SMEDDS was found to be 38 nm by dynamic light scattering and 12 nm by small angle neutron scattering. The in vitro dissolution studies showed complete and rapid drug release in 30 min compared to a plain drug (<5%) and marketed capsule (<50%). AUC and C_max were found to be 45.05 ± 15.64 ng h/ml and 3.91 ± 1.2 ng/ml for TAC-SMEDDS, 12.59 ± 5.54 ng h/ml and 0.48 ± 0.12 ng/ml for plain TAC, and 30.23 ± 10.34 ng h/ml and 2.31 ± 0.68 ng/ml for marketed formulation, respectively. The improved pharmacokinetic profile of TAC-SMEDDS is correlating to the dissolution results. Thus, gamma-oryzanol-enriched RGO acts as a potential multifunctional excipient for lipid formulations.

Pharmaceutical Research: Identification of Magnetic Nanoparticles for Combined Positioning and Lentiviral Transduction of Endothelial Cells

2012-01-10

Type: Original Paper
Purpose

The combination of magnetic nanoparticles (MNPs) with a magnetic field is a powerful approach to enable cell positioning and/or local gene therapy. Because physical requirements for MNPs differ between these two applications we have explored whether the use of different MNPs can provide site-specific positioning combined with efficient viral transduction of endothelial cells (ECs).

Methods

A variety of MNPs was screened for magnetic cell labeling and lentivirus binding. Then two different MNPs were chosen and their combined application was evaluated regarding EC magnetization and transduction efficiency.

Results

The combined use of PEI-Mag2 and NDT-Mag1 particles provided both efficient lentiviral transduction and high magnetic responsiveness of ECs that could be even retained within the vascular wall under flow conditions. The use of these MNPs did not affect biological characteristics of ECs like surface marker expression and vascular network formation. Importantly, with this method we could achieve an efficient functional overexpression of endothelial nitric oxide synthase in ECs.

Conclusions

The application of two different MNPs provides optimal results for magnetic labeling of ECs in combination with viral transduction. This novel approach could be very useful for targeted gene therapy ex vivo and site-specific cell replacement in the vascular system.

Pharmaceutical Research: Electrolyte-Stimulated Biphasic Dissolution Profile and Stability Enhancement for Tablets Containing Drug-Polyelectrolyte Complexes

Kindermann; Matthée; Sievert; Breitkreutz — 2012-01-07

Type: Original Paper
Purpose

Recently introduced drug-polyelectrolyte complexes prepared by hot-melt extrusion should be processed to solid dosage forms with tailor-made release properties. Their potential of stability enhancement should be investigated.

Methods

Milled hot-melt extruded naproxen-EUDRAGIT® E PO polyelectrolyte complexes were subsequently processed to double-layer tablets with varying complex loadings on a rotary-die press. Physicochemical interactions were studied under ICH guideline conditions and using the Gordon-Taylor equation. Sorption and desorption were determined to investigate the influence of moisture and temperature on the complex and related to stability tests under accelerated conditions.

Results

Naproxen release from the drug-polyelectrolyte complex is triggered by electrolyte concentration. Depending on the complex loading, phosphate buffer pH 6.8 stimulated a biphasic dissolution profile of the produced double-layer tablets: immediate release from the first layer with 65% loading and prolonged release from the second layer within 24 h (98.5% loading). XRPD patterns proved pseudopolymorphism for tablets containing the pure drug under common storage conditions whereas the drug-complex was stable in the amorphous state.

Conclusions

Drug-polyelectrolyte complexes enable tailor-made dissolution profiles of solid dosage forms by electrolyte stimulation and increase stability under common storage conditions.

The AAPS Journal: Disposition of Human Recombinant Lubricin in Naive Rats and in a Rat Model of Post-traumatic Arthritis After Intra-articular or Intravenous Administration

2012-01-07

Type: Original Paper

We have recently demonstrated that intra-articular (IA) administration of human recombinant lubricin, LUB:1, significantly inhibited cartilage degeneration and pain in the rat meniscal tear model of post-traumatic arthritis. In this report, we show that after a single IA injection to naïve rats and rats that underwent unilateral meniscal tear, [¹²⁵I]LUB:1 had a tri-phasic disposition profile, with the alpha, beta, and gamma half-life estimates of 4.5 h, 1.5 days, and 2.1 weeks, respectively. We hypothesize that the terminal phase kinetics was related to [¹²⁵I]LUB:1 binding to its ligands. [¹²⁵I]LUB:1 was detected on articular cartilage surfaces as long as 28 days after single IA injection. Micro-autoradiography analysis suggested that [¹²⁵I]LUB:1 tended to localize to damaged joint surfaces in rats with meniscal tear. After a single intravenous (IV) dose to rats, [¹²⁵I]LUB:1 was eliminated rapidly from the systemic circulation, with a mean total body clearance of 154 mL/h/kg and a mean elimination half-life (t_1/2) of 6.7 h. Overall, LUB:1 has met a desired disposition profile of a potential therapeutic intended for an IA administration: target tissue (knee) retention and fast elimination from the systemic circulation after a single IA or IV dose.

Pharmaceutical Research: Silica-Iron Oxide Magnetic Nanoparticles Modified for Gene Delivery: A Search for Optimum and Quantitative Criteria

2012-01-06

Type: Original Paper
Purpose

To optimize silica-iron oxide magnetic nanoparticles with surface phosphonate groups decorated with 25-kD branched polyethylenimine (PEI) for gene delivery.

Methods

Surface composition, charge, colloidal stabilities, associations with adenovirus, magneto-tranduction efficiencies, cell internalizations, in vitro toxicities and MRI relaxivities were tested for the particles decorated with varying amounts of PEI.

Results

Moderate PEI-decoration of MNPs results in charge reversal and destabilization. Analysis of space and time resolved concentration changes during centrifugation clearly revealed that at >5% PEI loading flocculation gradually decreases and sufficient stabilization is achieved at >10%. The association with adenovirus occurred efficiently at levels over 5% PEI, resulting in the complexes stable in 50% FCS at a PEI-to-iron w/w ratio of ≥7%; the maximum magneto-transduction efficiency was achieved at 9–12% PEI. Primary silica iron oxide nanoparticles and those with 11.5% PEI demonstrated excellent r₂* relaxivity values (>600 s⁻¹(mM Fe)⁻¹) for the free and cell-internalized particles.

Conclusions

Surface decoration of the silica-iron oxide nanoparticles with a PEI-to-iron w/w ratio of 10-12% yields stable aqueous suspensions, allows for efficient viral gene delivery and labeled cell detection by MRI.

Alert: 2012 AAPS National Biotechnology Conference

2011-10-21

Type: Alert

Alert: Thank You!

2011-01-28

Type: Alert

Alert: 2010 Impact Factors Released for the Official AAPS Journals

2008-10-03

Type: Alert