International Journal of Pharmaceutics (v.266, #1-2)

Hans E. Junginger: a life in pharmaceutics by Alexander T. Florence (1-2).

Shifting paradigms: biopharmaceuticals versus low molecular weight drugs by Daan J.A. Crommelin; Gert Storm; Ruud Verrijk; Leo de Leede; Wim Jiskoot; Wim E. Hennink (3-16).
Biopharmaceuticals are pharmaceutical products consisting of (glyco)proteins. Nowadays a substantial part of the FDA-approved drugs belong to this class of drugs. Biopharmaceuticals deserve special attention as they have a number of characteristics that set them aside from low molecular weight drugs.Their activity depends on their complicated shape based on secondary, tertiary and (sometimes) quaternary structures. These structures cannot be fully defined with our present set of analytical techniques and approaches for potency testing. They often are the same as (or closely resemble) endogenous proteins. This means that in safety testing and clinical test programs questions have to be addressed regarding species specific responses, selection of dosing schedules and route of administration, and the possible occurrence of immunogenicity. As the conformational structure of a protein is easily disturbed, formulation and handling of biopharmaceuticals needs special attention in order to optimize the therapeutic effect and minimize adverse reaction, among which immune responses.The issue of biogenerics is gaining more and more interest and different critical elements in the development of biogenerics are touched upon.In conclusion, biopharmaceuticals cannot be characterized fully in terms of their structure like low molecular weight drugs. The performance of biopharmaceuticals relies on strict production protocols and close monitoring of their activity in the clinical situation.
Keywords: Biopharmaceuticals; Biologicals; Biogenerics; Immunogenicity; PK–PD;

Generation of Toxoplasma gondii GRA1 protein and DNA vaccine loaded chitosan particles: preparation, characterization, and preliminary in vivo studies by Maytal Bivas-Benita; Marleen Laloup; Soetkin Versteyhe; Joelle Dewit; Jos De Braekeleer; Erik Jongert; Gerrit Borchard (17-27).
Chitosan microparticles as carriers for GRA-1 protein vaccine were prepared and characterized with respect to loading efficiency and GRA-1 stability after short-term storage. Chitosan nanoparticles as carriers for GRA-1 pDNA vaccine were prepared and characterized with respect to size, zeta potential, and protection of the pDNA vaccine against degradation by DNase I. Both protein and pDNA vaccine preparations were tested with regard to their potential to elicit GRA-1-specific immune response after intragastric administration using different prime/boost regimen. The immune response was measured by determination of IgG2a and IgG1 antibody titers. It was shown that priming with GRA1 protein vaccine loaded chitosan particles and boosting with GRA1 pDNA vaccine resulted in high anti-GRA1 antibodies, characterized by a mixed IgG2a/IgG1 ratio. These results showed that oral delivery of vaccines using chitosan as a carrier material appears to be beneficial for inducing an immune response against Toxoplasma gondii. The type of immune response, however, will largely depend on the prime/boost regimen and the type of vaccine used.
Keywords: Mucosal vaccination; Toxoplasmosis; GRA-1; DNA vaccine; Chitosan;

Novel pH-responsive complexation hydrogels containing pendent glucose (P(MAA-co-MEG)) or grafted PEG chains (P(MAA-g-EG)) were synthesized by photopolymerization. The feasibility of these hydrogels as oral protein delivery carriers was evaluated. The pH-responsive release behavior of insulin was analyzed from both P(MAA-co-MEG) and P(MAA-g-EG) hydrogels. In acidic media (pH 2.2), insulin release from the hydrogels was very slow. However, as the pH of the medium was changed to 6.5, a rapid release of insulin occurred. In both cases, the biological activity of insulin was retained. For P(MAA-co-MEG) hydrogels, the biological activity of insulin decreased when the pendent glucose content increased. In P(MAA-g-EG) hydrogels, when the grafted PEG molecular weight increased, the insulin biological activity decreased. Finally, hydrogels of P(MAA-co-MEG) prepared with an initial ratio of 1:4 MEG:MAA and P(MAA-g-EG) hydrogels containing PEG chains of molecular weights of 200 showed the greatest change in insulin release rate from acidic to basic pH solutions and the greatest protective effect for insulin in simulated GI tract conditions.
Keywords: Hydrogels; pH-responsive; Insulin; Oral drug delivery; Protein-release; Protein-stability;

Hydrolytic degradation of poly(lactide-co-glycolide) films: effect of oligomers on degradation rate and crystallinity by Gesine Schliecker; Carsten Schmidt; Stefan Fuchs; Ralf Wombacher; Thomas Kissel (39-49).
Oligomers are thought to accelerate the hydrolytic degradation of devices prepared from poly(lactide-co-glycolide), PLGA, due to their increased number of carboxylic end groups. To experimentally verify this hypothesis, two d,l-lactic acid oligomers having molecular weights close to their critical limit of solubility were synthesized and incorporated into PLGA films in three concentrations (0, 10, and 30% w/w). All films were translucent, rather flexible and initially amorphous. With increasing oligomer concentration the glass transition temperature (T g) and the molecular weight of films decreased prior to erosion. The degradation studies show that initial mass loss and water absorption are increased in oligomer-containing films as a function of average molecular weight and oligomer concentration. However, the incorporation of oligomers does not accelerate the degradation of films. By contrast, oligomer-containing films show extended lag phase until onset of polymer erosion. This was shown to be related to crystallization. Moreover, it was found that crystallization occurs earlier in oligomer-containing films and that the degree of crystallization is related to the average molecular weight of the oligomer. These findings bring new insight into the role of oligomers in the degradation process and can be used to explain why erosion in massive polymer devices occurs from the center to the surface.
Keywords: d,l-lactic acid oligomers; PLGA; Blends; Degradation; Crystallinity;

Stabilisation by freeze-drying of cationically modified silica nanoparticles for gene delivery by M. Sameti; G. Bohr; M.N.V. Ravi Kumar; C. Kneuer; U. Bakowsky; M. Nacken; H. Schmidt; C.-M. Lehr (51-60).
Core shell silica particles with a hydrodynamic diameter of 28 nm, an IEP of 7.1 and a zeta potential of +35 mV at pH 4.0 were synthesised. The role of freeze-drying for the conservation of zwitterionic nanoparticles and the usefulness of different lyoprotective agents (LPA) for the minimisation of particle aggregation were studied. The activity of the nanoparticles was measured as DNA-binding capacity and transfection efficiency in Cos-1 cells before and after lyophilisation. It was found that massive aggregation occurred in the absence of LPA. Of the various LPAs screened in the present investigations, trehalose and glycerol were found to be well suited for conservation of cationically modified silica nanoparticles with simultaneous preservation of their DNA-binding and transfection activity in Cos-1 cells.
Keywords: Freeze-drying; Gene delivery; Lyoprotective agents; Nanoparticles; Silica;

Iontophoretic R-apomorphine delivery in combination with surfactant pretreatment: in vitro validation studies by Gai Ling Li; Arne Grossklaus; Meindert Danhof; Joke A Bouwstra (61-68).
To validate the efficacy and controllability of a newly developed transdermal delivery system for R-apomorphine in combination with the surfactant pretreatment, iontophoresis was performed in three-chamber continuous-flow-through diffusion cells in vitro. The transdermal iontophoretic transport of R-apomorphine was examined with both human SC and freshly dermatomed human skin, at room temperature and at 32 °C. Furthermore, the relationship between current density and iontophoretic flux was investigated. By increasing the temperature from 22 to 32 °C, the iontophoretic transport rate of R-apomorphine in human SC was increased 1.9-fold. Also the iontophoretic flux increased linearly with the increase in the current density from 100 to 500 μA/cm2. When using dermatomed human skin instead of SC, the iontophoretic flux at a current density of 500 μA/cm2 was decreased from 362 ± 45 to 259 ± 30  nmol/cm2  h, and the corresponding lag time was prolonged from 0.8 to 2.8 h. In conclusion, the combination of non-occlusive pretreatment with the surfactant formulation and iontophoresis has shown to substantially increase the transdermal transport rate of R-apomorphine. A linear relationship between current density and R-apomorphine flux indicates that the iontophoretic delivery combined with surfactant pretreatment allows a controlled and individualised administration of R-apomorphine.
Keywords: Iontophoresis; Surfactant pretreatment; R-apomorphine; Parkinson’s disease;

Post-iontophoresis transport of ibuprofen lysine across rabbit ear skin by Patrizia Santi; Sara Nicoli; Gaia Colombo; Ruggero Bettini; Mariella Artusi; Silvia Rimondi; Cristina Padula; Paola Rizzo; Paolo Colombo (69-75).
The aim of this work was to study in vitro the post-iontophoresis transport of ibuprofen lysine across rabbit ear skin, from ibuprofen lysine water solutions (20–200 mg/ml and pH 6.8–7.8). Current densities of 0.125, 0.25, and 0.5 mA/cm2 were applied either continuously or for 15, 30, and 60 min followed by passive diffusion for up to 5 h. The results showed a significantly higher cathodal transport compared to passive flux. Anodal iontophoresis also increased ibuprofen permeation, even though the drug is negatively charged. The application of an electric current for a limited period of time, followed by passive diffusion from the reservoir in contact with the skin, produced much higher post-iontophoresis fluxes of ibuprofen than passive diffusion. Post-iontophoresis transport of ibuprofen from lysine salt solutions linearly depended on the total amount of current applied during iontophoresis, and in the absence of background ions was independent of donor drug concentration. The reason for this behavior was the creation of a drug reservoir in the skin owing to the short period of current application.
Keywords: Iontophoresis; Post-iontophoresis flux; Rabbit skin; Ibuprofen lysine;

Targeted and sustained drug delivery using PEGylated galactosylated liposomes by Chittima Managit; Shigeru Kawakami; Makiya Nishikawa; Fumiyoshi Yamashita; Mitsuru Hashida (77-84).
To achieve a sustained and targeted delivery of liposomes to liver parenchymal cells (PC), we modified distearoyl-l-phosphatidylcholine (DSPC)/cholesterol (Chol) (60:40) (DSPC/Chol) liposomes with a galactosylated cholesterol derivative (Gal-C4-Chol), and polysorbate (Tween) 20 or 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-polyethylene glycol (PEG x -DSPE). After intravenous injection, DSPC/Chol/Gal-C4-Chol (60:35:5) (Gal) liposomes were rapidly eliminated from the blood circulation and mostly recovered in the liver. The blood elimination of DSPC/Chol/Gal-C4-Chol/Tween 20 (55:35:5:5) (Tween 20-Gal) liposomes was slightly reduced as compared to Gal-liposomes. In contrast, a significant reduction in the blood elimination was observed with DSPC/Chol/Gal-C4-Chol/PEG2000-DSPE (59:35:5:1) (PEG2000-Gal) liposomes. Hepatic uptake of DSPC/Chol/Gal-C4-Chol/PEG350-DSPE (59:35:5:1) (PEG350-Gal) liposomes was intermediate between PEG2000-Gal-liposomes and Tween 20-Gal-liposomes. The uptake of PEG350-Gal-liposomes by liver PC was 7.7-fold higher than that by non-parenchymal cells (NPC). These results suggest that PEG350-DSPE can control the delivery rate of Gal-liposomes to liver PC without losing its targeting capability.
Keywords: Sustained delivery; Targeting; Galactosylated liposomes; PEGylation; Hepatocytes;

Cyclodextrins and emulsions by Dominique Duchêne; Amélie Bochot; Shan-Chen Yu; Céline Pépin; Monique Seiller (85-90).
This paper synthesises the literature on interactions between cyclodextrins (CD) and fatty acids and glycerides, and explains how these interactions allow the use of cyclodextrins to stabilise emulsions. An example of formulation with cyclodextrins is given which discusses the preparation of simple o/w emulsions, the addition of a model active ingredient, and the preparation of multiple emulsions in the absence of preformed surface active agents.
Keywords: Cyclodextrins; Emulsions; Multiple emulsions; Fatty acids; Glycerides;

Microtubules formed by capillary extrusion and fusion of surfactant vesicles by Behrooz Nasseri; Alexander T. Florence (91-98).
Polyhedral non-ionic surfactant vesicles formed from mixtures of polyoxyethylene-5-cetyl ether (C16EO5) or polyoxethylene-5-stearyl ether (C18EO5) with poly-24-oxyethylene cholesteryl ether (Solulan C24) and low amounts of cholesterol, when extruded from microcapillaries under pressure fuse to form multi-lamellar tubules up to about 80 μm in length. The diameter of the extruded tubules depends on the exit diameter of the capillaries used, in this paper generally around 1 μm. Under some circumstances, instead of linear tubules, the tubules form as concentric whorls which can be unraveled into their constituent tubules, indicating the strength of the systems. Vesicles can be formed within these tubular structures to act as a model for vesicular flow in elastic capillaries. Microparticles encapsulated with the primary polyhedral vesicles after extrusion are seen within the tubules, promising the possibility of a model for the study of microparticle flow within vessels. Preliminary studies on the use of the tubules as templates for polymerisation are also presented.
Keywords: Polyhedral niosomes; Microfabrication; Microtubules; Surfactant tubules;