International Journal of Pharmaceutics (v.500, #1-2)

Dissolvable microneedle fabrication using piezoelectric dispensing technology by Evin A. Allen; Conor O’Mahony; Michael Cronin; Thomas O’Mahony; Anne C. Moore; Abina M. Crean (1-10).
Display OmittedDissolvable microneedle (DMN) patches are novel dosage forms for the percutaneous delivery of vaccines. DMN are routinely fabricated by dispensing liquid formulations into microneedle-shaped moulds. The liquid formulation within the mould is then dried to create dissolvable vaccine-loaded microneedles. The precision of the dispensing process is critical to the control of formulation volume loaded into each dissolvable microneedle structure. The dispensing process employed must maintain vaccine integrity. Wetting of mould surfaces by the dispensed formulation is also an important consideration for the fabrication of sharp-tipped DMN. Sharp-tipped DMN are essential for ease of percutaneous administration.In this paper, we demonstrate the ability of a piezoelectric dispensing system to dispense picolitre formulation volumes into PDMS moulds enabling the fabrication of bilayer DMN. The influence of formulation components (trehalose and polyvinyl alcohol (PVA) content) and piezoelectric actuation parameters (voltage, frequency and back pressure) on drop formation is described. The biological integrity of a seasonal influenza vaccine following dispensing was investigated and maintained voltage settings of 30 V but undermined at higher settings, 50 and 80 V. The results demonstrate the capability of piezoelectric dispensing technology to precisely fabricate bilayer DMN. They also highlight the importance of identifying formulation and actuation parameters to ensure controlled droplet formulation and vaccine stabilisation.
Keywords: Microneedle; Vaccine delivery; Piezoelectric dispensing; Bilayer; Influenza vaccine; Ohnesorge number;

Poloxamer-hydroxyethyl cellulose-α-cyclodextrin supramolecular gels for sustained release of griseofulvin by Xelhua Marcos; Silvia Pérez-Casas; José Llovo; Angel Concheiro; Carmen Alvarez-Lorenzo (11-19).
Display OmittedSupramolecular gels of poloxamer-hydroxyethyl cellulose (HEC)-α-cyclodextrin (αCD) were developed aiming to obtain synergisms regarding solubilization and sustained release of griseofulvin for topical application. The effects of αCD concentration (0-10%w/w) on the phase behavior of aqueous dispersions of Pluronic® P123 (14%w/w) mixed with HEC (2%w/w) were evaluated at 4, 20 and 37 °C. The cooperative effects of the inclusion complex formation between poly(ethylene oxide) (PEO) blocks and HEC with αCD prevented phase separation and led to supramolecular networks that solubilize the antifungal drug. Rheological and bioadhesive properties of gels with and without griseofulvin could be easily tuned modulating the polymers proportions. Supramolecular gels underwent sol-gel transition at lower temperature than P123 solely dispersions and enabled drug sustained release for at least three weeks. All gels demonstrated good biocompatibility in the HET-CAM test. Furthermore, the drug-loaded gels showed activity against Trichophyton rubrum and Trichophyton mentagrophytes and thus may be useful for the treatment of tinea capitis and other cutaneous fungal infections.
Keywords: Supramolecular gel; Hydroxyethyl cellulose; Pluronic; Antifungal activity; Dermatophytosis; Controlled release;

Rheological and solid-state NMR assessments of copovidone/clotrimazole model solid dispersions by Fengyuan Yang; Yongchao Su; Lei Zhu; Chad D. Brown; Lawrence A. Rosen; Kenneth J. Rosenberg (20-31).
Display OmittedThis study aims to assess several model solid dispersions by using dynamic oscillatory rheology, solid-state NMR and other solid phase characterization techniques, and correlate their viscoelastic responses with processing methods and microstructures. A model active pharmaceutical ingredient (API), clotrimazole, was compounded with copovidone to form solid dispersions via various techniques with different mixing capabilities. Physicochemical characterizations of the resulting solid dispersions demonstrated that simple physical mixing led to a poorly mixed blend manifested by existence of large API crystalline content and heterogeneous distribution. Cryogenic milling significantly improved mixing of two components as a result of reduced particle size and increased contact surface area, but produced limited amorphous content. In contrast, hot melt extrusion (HME) processing resulted in a homogenous amorphous solid dispersion because of its inherent mixing efficiency. Storage modulus and viscosities versus frequency of different solid dispersions indicated that the incorporation of API into the polymer matrix resulted in a plasticizing effect which reduced the viscosity. The crystalline/aggregated forms of API also exhibited more elastic response than its amorphous/dispersed counterpart. Temperature ramps of the physical mixture with high API concentration captured a critical temperature, at which a bump was observed in damping factor. This bump was attributed to the dissolution of crystalline API into the polymer. In addition, heating–cooling cycles of various solid dispersions suggested that cryomilling and HME processing could form a homogeneous solid dispersion at low API content, whereas high drug concentration led to a relatively unstable dispersion due to supersaturation of API in the polymer.
Keywords: Rheology; Solid-state NMR; Hot melt extrusion and amorphous solid dispersion;

Hydrophobically modified inulin as an amphiphilic carbohydrate polymer for micellar delivery of paclitaxel for intravenous route by Pratik Muley; Sunny Kumar; Fadoua El Kourati; Siddharth S. Kesharwani; Hemachand Tummala (32-41).
Display OmittedMicellization offers several advantages for the delivery of water insoluble drugs including a nanoparticulate ‘core–shell’ delivery system for drug targeting. Recently, hydrophobically modified polysaccharides (HMPs) are gaining recognition as micelle forming polymers to encapsulate hydrophobic drugs. In this manuscript, for the first time, we have evaluated the self-assembling properties of a lauryl carbamate derivative of the poly-fructose natural polymer inulin (Inutec SP1® (INT)) to form paclitaxel (PTX) loaded micelles. INT self-assembled into well-defined micellar structures in aqueous environment with a low critical micellar concentration of 27.8 μg/ml. INT micelles exhibited excellent hemocompatibility and low toxicity to cultured cells. PTX loaded INT micelles exhibited a mean size of 256.37 ± 10.45 nm with excellent drug encapsulation efficiency (95.66 ± 2.25%) and loading (8.69 ± 0.22%). PTX loaded micelles also displayed sustained release of PTX and enhanced anti-cancer efficacy in-vitro in mouse melanoma cells (B16F10) compared to Taxol formulation with Cremophor EL as solvent. In addition, PTX loaded INT micelles exhibited comparable in-vivo antitumor activity in B16F10 allograft mouse model at half the dose of Taxol. In conclusion, INT offers safe, inexpensive and natural alternative to widely used PEG-modified polymers for the formulation of micellar delivery systems for paclitaxel.
Keywords: Micelles; Inutec SP1®; Inulin; Paclitaxel; Melanoma; Xenograft; Carbohydrate polymer;

The transport mechanism of integrin αvβ3 receptor targeting nanoparticles in Caco-2 cells by Yining Xu; Juan Xu; Wei Shan; Min Liu; Yi Cui; Lian Li; Chong Liu; Yuan Huang (42-53).
Display OmittedAs for the existence of epithelium barrier, accelerating the transport remains huge challenges for orally delivered protein and peptide drugs into blood circulation. Modifying nanopaticles (NPs) with targeting peptides can enhance the intestinal absorption of loaded macromolecular drugs. However, the transport process, which mainly means how the NPs pass through the apical membrane and the basolateral side and then enter into blood circulation, is needed comprehensive investigation. In this study, we systemically studied the transport mechanisms in Caco-2 cell model of trimethyl chitosan based NPs (TMC NPs) before and after modification of FQS, an integrin αvβ3 receptor targeting peptide. Our results showed FQS peptide mediated multiple endocytosis pathways and could activate integrin αvβ3 receptor by interacting with FAK and Src-family kinases to induce receptor-mediated endocytosis of the NPs. Then, both endocytosed NPs could transport from early endosome to lysososmes via late endosomes/lysosome pathway, as well as to recycling endosomes and Golgi apparatus through early endosome/recycling endosomes and Golgi apparatus/recycling endosomes/plasma membrane pathways, respectively. After FQS peptide modification, the endocytosis subpathways of NPs have been changed, and more pathways are involved in exocytosis process for FQS-modified NPs compared with non-modified NPs. Our study indicated the ligand modification could enhance the uptake and transport by altering some pathways in whole transport process of NPs.
Keywords: Transport mechanism; Nanoparticles; FQSIYPpIK; Oral delivery; Protein and peptide drugs; Epithelium;

Improved antitumor activity of epirubicin-loaded CXCR4-targeted polymeric nanoparticles in liver cancers by Sun Di-Wen; Guo-Zheng Pan; Long Hao; Jian Zhang; Qing-Ze Xue; Peng Wang; Qing-Zhong Yuan (54-61).
Display OmittedA liver-targeted drug delivery system (CX-EPNP) composed of PLGA/TPGS was prepared and characterized. The surface of nanoparticle was conjugated with LFC131 peptide to increase the specific interaction of carrier with CXCR4 overexpressing liver cancers to enhance the Epirubicin (EPI) delivery to tumors. The particles were nanosized with size than 150 nm and portrayed a sustained release kinetics suggesting its suitability for cancer targeting. The in vitro cell uptake results showed that the introduction of LFC131 to the nanoparticles could increase significantly the affinity to human hepatic carcinoma cells (HepG2) with approximately a 3-fold improvement in cellular uptake than non-targeted one. A specific receptor-mediated uptake was observed by confocal laser scanning microscopy. In addition, CX-EPNP showed remarkable cytotoxicity towards HepG2 cells, and could effectively inhibit tumor growth. The more significant EPI accumulation from CX-EPNP in the cancer cells gave rise to the enhanced EPI cytotoxicity and cell apoptosis. The CX-EPNP distributed mostly in the xenograft tumor after intravenous administration to mice and adequately remained in the blood for at least 24 h. It seemed that CX-EPNP upon intravenous injection avoided rapid recognition by Kupffer cells and adequately remained in the blood. These findings suggest that CX-EPNP could effectively inhibit the growth of liver tumors in situ and could potentially reduce the systemic side effects. However, extensive investigation is still needed to assess the possible applications of the CX-EPNP in humans.
Keywords: Liver cancer; Epirubicin; CXCR4; Targeted nanoparticles; Biodistribution;

Design, preparation, and in vitro characterization of a trimodally-targeted nanomagnetic onco-theranostic system for cancer diagnosis and therapy by Abdolhossein Zarrin; Somayeh Sadighian; Kobra Rostamizadeh; Omidreza Firuzi; Mehrdad Hamidi; Soliman Mohammadi-Samani; Ramin Miri (62-76).
Display OmittedIn this study, the aim was to introduce and characterize a new trimodally-targeted nanomagnetic onco-theranostic system for simultaneous early diagnosis and efficient treatment of cancer. The onco-theranostic system was designed as it could target the tumor site through three targeting approach, i.e. magnetic, folic acid receptor, and pH sensitivity, and concurrently, due to the presence of superparamagnetic iron oxide nanoparticles (SPIONs) with super paramagnetic characteristics could be useful as MRI contrast agent for early cancer diagnosis. To achieve this goal, SPIONs were coated with chitosan and folic acid-conjugated chitosan via ionic gelation method in order to obtain non-targeted nanomagnetic onco-diagnostic (NT/NOD) and targeted nanomagnetic onco-diagnostic (T/NOD) systems. Finally, doxorubicin was loaded successfully into NT/NOD and T/NOD in order to obtain nanomagnetic onco-theranostic (NT/NOT) and targeted nanomagnetic onco-theranostic (T/NOT) systems. The entrapment efficiency and drug loading of T/NOT was determined to be 62.33 ± 5.20% and 10.26 ± 1.36%, respectively. MTT assay revealed that all systems were biocompatible within the concentration range investigated. Also, the T/NOT system showed the lowest IC50 comparing with free doxorubicin and NT/NOT system. In addition, uptake studies and competitive inhibition study verified the folate receptor mediated endocytosis of targeted system by MCF-7 as a folate receptor-positive cell line. The finding revealed that the extent of drug release from theranostic systems was pH-sensitive as it was higher at acidic media compared to that of in the neutral condition. Finally, T 2-weighted phantom images, with an acceptable and dose-dependent resolution, proved the potential of T/NOT system as promising T 2 MR contrast agent for diagnostic purpose. These finding proved that the prepared T/NOT system have great potential as a novel tumor-targeting nanotheranostic agent for simultaneous MRI imaging and treatment of folate receptor-positive cancers. Further studies are needed to test their behavior in vivo.
Keywords: Folate-receptor targeting; SPION; Nanomagnetic onco-theranostic system; Nanomagnetic onco-diagnostic system;

Facile large-scale preparation of mesoporous silica microspheres with the assistance of sucrose and their drug loading and releasing properties by Yanping Bi; Chaonan Wu; Ming Xin; Shuyan Bi; Chengxin Yan; Jifu Hao; Fei Li; Shou Li (77-84).
Display OmittedMesoporous silica microspheres (MSMs) with a pore-size larger than 10 nm and a large pore-volume have attracted considerable attention for their application in delivering poorly water-soluble drugs. Here we developed a simple method for large-scale synthesis of MSMs using sodium silicate as silica precursor. The novelty of this approach lies in the use of sucrose solution to achieve large size and volume of nanopores. The highest values of pore size and pore volume are 13.2 nm and 1.97 cm3/g, respectively. Importantly, the method is reliable and easily upscalable. The blank and drug-loaded MSMs were characterized by differential scanning calorimetry (DSC) and X-ray powder diffraction (XRPD). Ibuprofen and resveratrol were successfully loaded into the nanopores of MSMs in amorphous and nanocrystalline form and showed high drug-loadings and enhanced dissolution rates. This kind of MSMs appears to be a promising candidate as a new oral drug delivery vehicle providing a rapid drug release.
Keywords: Mesoporous silica microspheres; Sodium silicate; Nanocrystal; Ibuprofen; Resveratrol;

Drug loading of foldable commercial intraocular lenses using supercritical impregnation by A. Bouledjouidja; Y. Masmoudi; M. Sergent; V. Trivedi; A. Meniai; E. Badens (85-99).
Display OmittedThe drug delivery through intraocular lenses (IOLs) allows the combination of cataract surgery act and postoperative treatment in a single procedure. In order to prepare such systems, “clean” supercritical CO2 processes are studied for loading commercial IOLs with ophthalmic drugs. Ciprofloxacin (CIP, an antibiotic) and dexamethasone 21-phosphate disodium (DXP, an anti-inflammatory drug) were impregnated into foldable IOLs made from poly-2-hydroxyethyl methacrylate (P-HEMA). A first pre-treatment step was conducted in order to remove absorbed conditioning physiological solution. Supercritical impregnations were then performed by varying the experimental conditions. In order to obtain transparent IOLs and avoid the appearance of undesirable foaming, it was necessary to couple slow pressurization and depressurization phases during supercritical treatments. The impregnation yields were determined through drug release studies. For both drugs, release studies showdeep and reproducible impregnation for different diopters.For the system P-HEMA/CIP, a series of impregnations was performed to delimit the experimental range at two pressures (80 and 200 bar) in the presence or absence of ethanol as a co-solvent for two diopters (+5.0 D and +21.0 D). Increase in pressure in the absence of a co-solvent resulted in improved CIP impregnation. The addition of ethanol (5 mol%) produced impregnation yields comparable to those obtained at 200 bar without co-solvent. A response surface methodology based on experimental designs was used to study the influence of operating conditions on impregnation of IOLs (+21.0 D) in the absence of co-solvent. Two input variables with 5 levels each were considered; the pressure (80–200 bar) and the impregnation duration (30–240 min). CIP impregnation yields ranging between 0.92 and 3.83 μgCIP/mgIOL were obtained from these experiments and response surface indicated the pressure as a key factor in the process.The DXP impregnation in P-HEMA was higher than CIP at all the tested conditions (8.50–14.53 μgDXP/mgIOL). Furthermore, unlike CIP, highest DXP impregnation yields were obtained in the presence of ethanol as a co-solvent (5 mol%). NMR spectroscopy was performed to confirm complete removal of ethanol in the co-solvent-treated IOLs.
Keywords: Cataract post-operative treatment; Drug delivery systems; Supercritical impregnation; Foldable P-HEMA intraocular lenses; Response surface methodology; Drug release study;

Display OmittedThe pharmacological target of many anticancer drugs is those molecules associated with genetic information which are localized in nucleus. To efficiently deliver drugs into cancer cell nucleus, in our previous study, a fork-like sub-unit, with one end conjugated with a targeting peptide named R8NLS (CRRRRRRRRPKKKRKV) and the other end conjugated with c-Myc oncogene inhibitor H1-S6A,F8A (H1) peptide, was linked onto the N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer via an enzyme degradable glycylphenylalanylleucylglycine (GFLG) tetrapeptide spacer. Here, an in vitro mechanism investigation of the fork-like sub-unit was studied in detail. We found that the fusion with two complementary R8 and NLS motifs is required to exert the multifunctional targeting capability of the tandem R8NLS peptide in overcoming various intracellular barriers, including enhancing cellular uptake, facilitating endosomal escape and penetrating through the double-layered nuclear membrane. Also required is the tactful detachment of the fork-like sub-unit from the copolymer in response to intracellular stimulus, because a smaller sub-unit not only increases the intracellular trafficking efficiency by reducing the size burden of magical bullet R8NLS, but also guarantees successful entry through the restricted nucleopore. Herein, this study highlights that both nuclear targeting ligand R8NLS and detachable fork-like sub-unit are dispensable for programmed nuclear delivery and may show feasibility on other drug delivery systems, such as nanoparticles and micelles.
Keywords: HPMA copolymers; Nuclear-targeted; Detachable fork-like structure; Cancer therapy;

The research about microscopic structure of emulsion membrane in O/W emulsion by NMR and its influence to emulsion stability by Yiqiao Xie; Jisheng Chen; Shu Zhang; Kaiyan Fan; Gang Chen; Zerong Zhuang; Mingying Zeng; De Chen; Longgui Lu; Linlin Yang; Fan Yang (110-119).
Display OmittedThis paper discussed the influence of microstructure of emulsion membrane on O/W emulsion stability.O/W emulsions were emulsified with equal dosage of egg yolk lecithin and increasing dosage of co-emulsifier (oleic acid or HS15). The average particle size and centrifugal stability constant of emulsion, as well as interfacial tension between oil and water phase were determined. The microstructure of emulsion membrane had been studied by 1H/13C NMR, meanwhile the emulsion droplets were visually presented with TEM and IFM.With increasing dosage of co-emulsifier, emulsions showed two stable states, under which the signal intensity of characteristic group (orient to lipophilic core) of egg yolk lecithin disappeared in NMR of emulsions, but that (orient to aqueous phase) of co-emulsifiers only had some reduction at the second stable state. At the two stable states, the emulsion membranes were neater in TEM and emulsion droplets were rounder in IFM. Furthermore, the average particle size of emulsions at the second stable state was bigger than that at the first stable state.Egg yolk lecithin and co-emulsifier respectively arranged into monolayer and bilayer emulsion membrane at the two stable states. The microstructure of emulsion membrane was related to the stability of emulsion.
Keywords: Emulsion membrane; Microscopic structure; O/W emulsion; Stability; 1H NMR/13C NMR spectroscopy;

Next generation of buccadhesive excipient: Preactivated carboxymethyl cellulose by Flavia Laffleur; Lukas Bacher; Stefan Vanicek; Claudia Menzel; Ijaz Muhammad (120-127).
Display OmittedAssessment of preactivated carboxymethyl cellulose as potential excipient for buccal drug delivery.Firstly, carboxymethyl cellulose (CMC) and cysteine (SH) were covalently coupled via amide bond formation to obtain thiolated carboxymethyl cellulose (CMC–SH). Further, preactivated carboxymethyl cellulose (CMC-S-S-MNA) was obtained by preactivation with 2-mercaptonicotinic acid (MNA). Sulforhodamine 101 (SRH101) was used as a model drug for permeation study through buccal mucosa. CMC-S-S-MNA was evaluated with respect to mucoadhesive and permeation enhancing effect and cytotoxicity.Thiolated carboxymethyl cellulose exhibited a total amount of 112.46 ± 0.46 thiol groups. CMC-S-S-MNA exhibited around 50% of preactivated thiol groups. The preactivated polymer showed no toxic effect. Furthermore, compared to unmodified CMC, CMC-S-S-MNA revealed 3.0-fold improved mucoadhesive properties according to the rotating cylinder method and 8.8-fold enhancement in mucoadhesiveness by tensile assay, respectively.Preactivated carboxymethyl cellulose fulfills the requirements as potential excipient of being mucoadhesive and permeation enhancing for the buccal drug delivery.
Keywords: Buccal; Carboxymethyl cellulose; Mucosa; Preactivation; Thiomers;

Role of scavenger receptors in peptide-based delivery of plasmid DNA across a blood–brain barrier model by Artita Srimanee; Jakob Regberg; Mattias Hällbrink; Opa Vajragupta; Ülo Langel (128-135).
Display OmittedReceptor-mediated transcytosis remains a major route for drug delivery across the blood–brain barrier (BBB). PepFect 32 (PF32), a peptide-based vector modified with targeting ligand (Angiopep-2) binding to low-density lipoprotein receptor-related protein-1 (LRP-1), was previously found to be a promising vector for plasmid delivery across an in vitro model of the BBB. Cellular uptake of PF32/plasmid DNA (pDNA) complexes was speculated the internalization via LRP-1 receptor. In this study, we prove that PF32/pDNA nanocomplexes are not only transported into brain endothelial cells via LRP-1 receptor-mediated endocytosis, but also via scavenger receptor class A and B (SCARA3, SCARA5, and SR-BI)-mediated endocytosis. SCARA3, SCARA5, and SR-BI are found to be expressed in the brain endothelial cells. Inhibition of these receptors leads to a reduction of the transfection. In conclusion, this study shows that scavenger receptors also play an essential role in the cellular uptake of the PF32/pDNA nanocomplexes.
Keywords: Blood–brain barrier; bEnd.3; Plasmid transfection; Scavenger receptors; angiopep-2; LRP-1 receptor; Receptor-mediated endocytosis;

Effect of hydrophobic inclusions on polymer swelling kinetics studied by magnetic resonance imaging by Michaela Gajdošová; Daniel Pěček; Nina Sarvašová; Zdeněk Grof; František Štěpánek (136-143).
Display OmittedThe rate of drug release from polymer matrix-based sustained release formulations is often controlled by the thickness of a gel layer that forms upon contact with dissolution medium. The effect of formulation parameters on the kinetics of elementary rate processes that contribute to gel layer formation, such as water ingress, polymer swelling and erosion, is therefore of interest. In the present work, gel layer formation has been investigated by magnetic resonance imaging (MRI), which is a non-destructive method allowing direct visualization of effective water concentration inside the tablet and its surrounding. Using formulations with Levetiracetam as the active ingredient, HPMC as a hydrophilic matrix former and carnauba wax (CW) as a hydrophobic component in the matrix system, the effect of different ratios of these two ingredients on the kinetics of gel formation (MRI) and drug release (USP 4 like dissolution test) has been investigated and interpreted using a mathematical model.
Keywords: MRI; HPMC; Drug dissolution; Gel layer; Carnauba wax;

Development of TMTP-1 targeted designer biopolymers for gene delivery to prostate cancer by John W. McBride; Ashley S. Massey; J. McCaffrey; Cian M. McCrudden; Jonathan A. Coulter; Nicholas J. Dunne; Tracy Robson; Helen O. McCarthy (144-153).
Display OmittedDesigner biopolymers (DBPs) represent state of the art genetically engineered biomacromolecules designed to condense plasmid DNA, and overcome intra- and extra- cellular barriers to gene delivery. Three DBPs were synthesized, each with the tumor molecular targeting peptide-1 (TMTP-1) motif to specifically target metastases. Each DBP was complexed with a pEGFP-N1 reporter plasmid to permit physiochemical and biological assay analysis. Results indicated that two of the biopolymers (RMHT and RM3GT) effectively condensed pEGFP-N1 into cationic nanoparticles <100 nm and were capable of transfecting PC-3 metastatic prostate cancer cells. Conversely the anionic RMGT DBP nanoparticles could not transfect PC-3 cells. RMHT and RM3GT nanoparticles were stable in the presence of serum and protected the cargo from degradation. Additionally it was concluded that cell viability could recover post-transfection with these DBPs, which were less toxic than the commercially available transfection reagent Lipofectamine® 2000. With both DBPs, a higher transfection efficacy was observed in PC-3 cells than in the moderately metastatic, DU145, and normal, PNT2-C2, cell lines. Blocking of the TMTP-1 receptors inhibited gene transfer indicating internalization via this receptor. In conclusion RMHT and RM3GT are fully functional DBPs that address major obstacles to gene delivery and target metastatic cells expressing the TMTP-1 receptor.
Keywords: Biopolymer; Delivery system; Nanoparticles; Gene Therapy; TMTP-1;

Lipid nanoparticles enhance the absorption of cyclosporine A through the gastrointestinal barrier: In vitro and in vivo studies by Melissa Guada; Beatriz Lasa-Saracíbar; Hugo Lana; Maria del Carmen Dios-Viéitez; Maria J. Blanco-Prieto (154-161).
Display OmittedIn the present work, the feasibility of cyclosporine A lipid nanoparticles (CsA LN) for oral administration was investigated. Three CsA LN formulations were developed using Precirol as lipid matrix, one stabilized with Tween® 80 (Tw) and the other two with mixtures of phosphatidylcholine or Pluronic® F127 with taurocholate (Lec:TC and PL:TC, respectively). The physical characteristics of the LN were studied under gastrointestinal pH and their integrity was found to be dependent on the stabilizers. The in vitro intestinal permeability was assessed with a human colon adenocarcinoma cell model and in vivo pharmacokinetic and biodistribution studies were performed in Balb/c mice using Sandimmune Neoral® as reference. In vitro results showed the highest CsA permeability with the LN containing Lec:TC. In contrast, the best in vivo performance was achieved from the LN containing Tw. The bioavailability of CsA was matched and even enhanced with Precirol nanoparticles. This study suggests the suitability of LN as promising vehicles for CsA oral delivery.
Keywords: Cyclosporine A; Lipid nanoparticles; Gastrointestinal stability; Caco-2 cells; Bioavailability; Tissue distribution; Oral route;

Folate receptor-targeted multimodal polymersomes for delivery of quantum dots and doxorubicin to breast adenocarcinoma: In vitro and in vivo evaluation by Mona Alibolandi; Khalil Abnous; Fatemeh Sadeghi; Hossein Hosseinkhani; Mohammad Ramezani; Farzin Hadizadeh (162-178).
Display OmittedIn this study, we report the design and delivery of tumor-targeted, quantum dot (QD) and doxorubicin (DOX)-encapsulated PEG-PLGA nanopolymersomes (NPs) for the imaging and chemotherapy of breast cancer. To achieve active cancer targeting, QD and DOX-encapsulated NPs were conjugated with folate for folate-binding protein receptor-guided delivery, which overexpressed in many cancer cells. Hydrophobic DOX and hydrophilic MSA-capped QD were encapsulated in the bilayer and core of the PEG-PLGA nanopolymersomes, respectively.The data show that the formulated NPs sustained DOX release for a period of 12 days. Fluorescence microscopy and MTT assay demonstrated that the developed folate-targeted DOX-QD NPs had higher cytotoxicity than non-targeted NPs and the free form of the drug; moreover, they preferentially accumulated in 4T1 and MCF-7 cells in vitro. In vivo experiments including whole organ tissue–homogenate analysis and organ fluorescence microscopy imaging of BALB/c mice bearing 4T1 breast adenocarcinoma showed that the folate receptor-targeted QD encapsulated NPs accumulate at tumor sites 6 h following intravenous injection. Acute toxicity studies of the prepared targeted QD-loaded NPs showed no evidence of long-term harmful histopathological and physiological effects on the treated animals. The in vivo tumor inhibitory effect of folic acid (FA)-QD-DOX NPs demonstrated an augmented therapeutic efficacy of targeted formulation over the non-targeted and free drug. The data obtained illustrate a high potential of the prepared targeted theranostic nanoplatform in the treatment and imaging of breast cancer. This study may open new directions for preparation of QD-based theranostic polymersomes for clinical application.
Keywords: Nanopolymersome; Doxorubicin; Quantum dot; Breast cancer; 4T1; PEG-PLGA;

Effect of poly ethylene glycol on the mechanical and thermal properties of bioactive poly(ε-caprolactone) melt extrudates for pharmaceutical applications by P. Douglas; Ahmad B. Albadarin; M. Sajjia; Chirangano Mangwandi; Manuel Kuhs; Maurice N. Collins; Gavin M. Walker (179-186).
Display OmittedThis paper investigates the effects of polyethylene glycol (PEG), on the mechanical and thermal properties of nalidixic acid/poly ε-caprolactone (NA)/PCL blends prepared by hot melt extrusion. The blends were characterized by tensile and flexural analysis, dynamic mechanical analysis, differential scanning calorimetry, thermogravimetric analysis and X-ray diffraction. Results show that loading PEG in the PCL had a detrimental effect on the tensile strength and toughness of the blends, reducing them by 20–40%. The partial miscibility of the PCL-PEG system, causes an increase in T g. While increases in the crystallinity is attributed to the plasticisation effect of PEG and the nucleation effect of NA. The average crystal size increased by 8% upon PEG addition. Experimental data indicated that the addition of NA caused loss of the tensile strength and toughness of PCL. Thermal analysis of the PCL showed that on addition of the thermally unstable NA, thermal degradation occurred early and was autocatalytic. However, the NA did benefit from the heat shielding provided by the PCL matrix resulting in more thermally stable NA particles.
Keywords: Mechanical and thermal properties; Melt extrusion; Drug formulation; Poly-caprolactone; Nalidixic acid;

Impact of PEG and PEG-b-PAGE modified PLGA on nanoparticle formation, protein loading and release by René Rietscher; Justyna A. Czaplewska; Tobias C. Majdanski; Michael Gottschaldt; Ulrich S. Schubert; Marc Schneider; Claus-Michael Lehr (187-195).
Display OmittedThe effect of modifying the well-established pharmaceutical polymer PLGA by different PEG-containing block-copolymers on the preparation of ovalbumin (OVA) loaded PLGA nanoparticles (NPs) was studied. The used polymers contained poly(d,l-lactic-co-glycolic acid) (PLGA), polyethylene glycol (PEG) and poly(allyl glycidyl ether) (PAGE) as building blocks. The double emulsion technique yielded spherical NPs in the size range from 170 to 220 nm (PDI < 0.15) for all the differently modified polymers, allowing to directly compare protein loading of and release. PEGylation is usually believed to increase the hydrophilic character of produced particles, favoring encapsulation of hydrophilic substances. However, in this study simple PEGylation of PLGA had only a slight effect on protein release. In contrast, incorporating a PAGE block between the PEG and PLGA units, also eventually enabling active targeting introducing a reactive group, led to a significantly higher loading (+25%) and release rate (+100%), compared to PLGA and PEG-b-PLGA NPs.
Keywords: Poly(d,l-lactic-co-glycolic acid); Polyethylene glycol; Poly(allyl glycidyl ether); Ovalbumin; Protein delivery; Drug delivery;

AN in vitro evaluation of a carmustine-loaded Nano-co-Plex for potential magnetic-targeted intranasal delivery to the brain by Olufemi D. Akilo; Yahya E. Choonara; André M. Strydom; Lisa C. du Toit; Pradeep Kumar; Girish Modi; Viness Pillay (196-209).
Display OmittedTargeted delivery of carmustine (BCNU), an efficient brain tumor therapeutic, has been challenged with bioavailability issues due to the Blood Brain Barrier (BBB). The currently effective delivery approach is by implants at the site of the tumor, but this is highly invasive. The intranasal route, which is non-invasive and bypasses the BBB, may be alternative route for delivering BCNU to the brain. In this work, polyvinyl alcohol/polyethyleneimine/fIuorecein isothiocyanate complex (Polyplex) coated iron-oxide nanoparticles (Magnetite) were synthesized employing co-precipitation, epoxidation and EDC/NHS coupling reactions. The Polyplex coated magnetite (Nano-co-Plex) was loaded with BCNU for potential magnetically targeted delivery to the brain following intranasal administration. The Nano-co-Plex was characterized employing Thermogravimetric analysis (TGA), Superconducting Quantum Interference Device (SQUID) magnetometry, Fourier Transform Infrared Spectroscopy (FTIR), Nuclear Magnetic Resonance (NMR), X-ray Diffractometry (XRD), Transmission Electron Microscopy (TEM) and Zetasize analysis. Results revealed superparamagnetic hexagonally shaped “core-shell” nanoparticles with cell labeling attributes, of size ranging between 30–50 nm, and a zeta potential value of +32 ± 2 mV. The Nano-co-Plex synthesized was found to possess high degree of crystallinity with 32% Polyplex coating. The loading and release studies indicated a time-dependent loading with maximum loading capacity of 176.82 μg BCNU/mg of the carrier and maximum release of 75.8% of the loaded BCNU. Cytotoxicity of the BCNU-loaded Nano-co-Plex displayed superiority over the conventional BCNU towards human glioblastoma (HG) cells. Cell studies revealed enhanced uptake and internalization of BCNU-loaded Nano-co-plex in HG cells in the presence of an external magnetic field. These Nano-co-Plexes may be ideal as an intranasal magnetic drug targeting device for BCNU delivery.
Keywords: Carmustine; Iron oxide nanoparticles; Magnetite; Superparamagnetic; Core-shell; Chemotherapeutics; Blood brain barrier; Brain tumor; Polyplex; Nano-co-Plex;

Display OmittedUsing a topical product is part of the overall strategy for skin cancer prevention. The level of protection attainable when using commercial products is indicated by the Sun Protection Factor (SPF) value, in use everywhere. This value reflects the level of protection primarily in the UVB range. However, UVA radiation also has deleterious effects on the skin, and it is essential to prevent it, which is why products must offer a wide spectrum of protection. Tests conducted in vivo, before any marketing, are done by applying the studied product at a rate of 2.0 mg cm−2, while users, in practice, only use 1.0–1.5 mg cm−2. We now know that this reduction in the amount of applied product greatly affects the SPF. To complete the state of knowledge in this area, we sought to evaluate the effect of a decrease in the amount of applied sunscreen product by studying sunscreen creams and oils on the level of protection attainable in the UVA range. We have shown that the PF-UVA is divided by a factor of 2.2, on average, when the amount of applied product is reduced by half, with differences depending on the product type under consideration (cream or oil) and depending on the SPF of the preparation.
Keywords: Sunscreens; Oil; Creams; Protection factor UVA (PF-UVA); Amount of product;

Display OmittedBuccal films were prepared from aqueous and ethanolic Metolose gels using the solvent casting approach (40 °C). The hydration (PBS and simulated saliva), mucoadhesion, physical stability (20 °C, 40 °C), in vitro drug (omeprazole) dissolution (PBS and simulated saliva), ex vivo permeation (pig buccal mucosa) in the presence of simulated saliva, ex vivo bioadhesion and cell viability using MTT of films were investigated. Hydration and mucoadhesion results showed that swelling capacity and adhesion was higher in the presence of PBS than simulated saliva (SS) due to differences in ionic strength. Omeprazole was more stable at 20 °C than 40 °C whilst omeprazole release reached a plateau within 1 h and faster in PBS than in SS. Fitting release data to kinetic models showed that Korsmeyer–Peppas equation best fit the dissolution data. Drug release in PBS was best described by zero order via non-Fickian diffusion but followed super case II transport in SS attributed to drug diffusion and polymer erosion. The amount of omeprazole permeating over 2 h was 275 ug/cm2 whilst the formulations and starting materials showed cell viability values greater than 95%, confirming their safety for potential use in paediatric buccal delivery.
Keywords: Permeation; Cell toxicity; Omeprazole; Metolose; Buccal films; Paediatric;

Biokinetic studies of non-complexed siRNA versus nano-sized PEI F25-LMW/siRNA polyplexes following intratracheal instillation into mice by Jens Lipka; Manuela Semmler-Behnke; Alexander Wenk; Jana Burkhardt; Achim Aigner; Wolfgang Kreyling (227-235).
Display OmittedSuccessful gene therapy requires stability and sufficient bioavailability of the applied drug at the site of action. In the case of RNA interference (RNAi), non-viral vectors play a promising role for delivering intact siRNA molecules. We selected a low molecular weight polyethyleneimine (PEI F25-LMW) and investigated the biokinetics of PEI F25-LMW/siRNA polyplexes in comparison to non-complexed siRNA molecules upon intratracheal application into mice. Additionally, a bronchoalveolar lavage was performed to locate the siRNA within the different lung compartments and to analyse possible inflammatory reactions. Liquid scintillation counting of a 32P-label was used to follow the siRNA within the whole body. During the complete observation time more than 75% of the applied dose was found at the target site. The complexation with PEI F25- LMW prevented the siRNA from being degraded and cleared and prolonged its retention time. A low inflammatory reaction was observed on the basis of cell differentiation. Taken together, PEI F25-LMW meets fundamental requirements on non-viral vectors for local pulmonary siRNA delivery.
Keywords: Biokinetics; Radiolabeled polyethyleneimine (PEI)/siRNA polyplexes; Lung therapy;

Insulin loaded mucus permeating nanoparticles: Addressing the surface characteristics as feature to improve mucus permeation by Irene Pereira de Sousa; Thomas Moser; Corinna Steiner; Barbara Fichtl; Andreas Bernkop-Schnürch (236-244).
Display OmittedIt was the aim of this study to combine two strategies – namely the virus-mimicking strategy and the surface PEGylation strategy – in order to generate highly mucus permeating nanocarriers for oral insulin delivery.Chondroitin sulphate was covalently conjugated with poly(ethylene glycol) 5 kDa at different degree of modification and with the functionalized polymers NPs were formulated via complexation with chitosan. NPs were characterized by particle size, zeta potential, surface hydrophilicity and permeation ability in porcine mucus and on excised mucosa.The NPs presented a size between 510 and 670 nm and a zeta potential between −1 and −5 mV when dispersed in simulated intestinal fluid. The mucus permeation test revealed a correlation between the NPs hydrophilicity and their ability to move through mucus. A 5-fold higher amount of NPs with the higher degree of PEGylation could permeate through fresh mucus compared to non-PEGylated NPs. Moreover, highly PEGylated NPs showed a 3.7-fold greater ability to be retained in intestinal mucosa against buffer flow compared to unmodified NPs. Finally, insulin was incorporated with a payload of 2.18% and the release profile showed a 65% release within 4 h.Results of this study provide strong evidence for the potential of combining different surface modification strategies in order to improve the mucus permeating properties of NPs for oral peptide delivery.
Keywords: Mucus-penetrating particles; Diffusion; Mucus; Insulin; PEGylation;

Display OmittedThis work aimed to develop an optimized ethosomal formulation of glimepiride then loading into transdermal films to offer lower drug side effect, extended release behavior and avoid first pass effect. Four formulation factors were optimized for their effects on vesicle size (Y1), entrapment efficiency (Y2) and vesicle flexibility (Y3). Optimum desirability was identified and, an optimized formulation was prepared, characterized and loaded into transdermal films. Ex-vivo permeation study for the prepared films was conducted and, the permeation parameters and drug permeation mechanism were identified. Penetration through rat skin was studied using confocal laser microscope. In-vivo study was performed following transdermal application on human volunteers. The percent of alcohol was significantly affecting all the studied responses while the other factors and their interaction effects were varied on their effects on each response. The optimized ethosomal formulation showed observed values for Y1, Y2 and Y3 of 61 nm, 97.12% and 54.03, respectively. Ex-vivo permeation of films loaded with optimized ethosomal formulation was superior to that of the corresponding pure drug transdermal films and this finding was also confirmed after confocal laser microscope study. Permeation of glimepiride from the prepared films was in favor of Higushi-diffusion model and exhibited non-Fickian or anomalous release mechanism. In-vivo study revealed extended drug release behavior and lower maximum drug plasma level from transdermal films loaded with drug ethosomal formulation. So, the ethosomal formulation could be considered a suitable drug delivery system especially when loaded into transdermal vehicle with possible reduction in side effects and controlling the drug release.
Keywords: Glimepiride; Ethosomes; Transdermal delivery; Ex-vivo permeation; Human volunteers;

Risk based in vitro performance assessment of extended release abuse deterrent formulations by Xiaoming Xu; Abhay Gupta; Manar Al-Ghabeish; Silvia N. Calderon; Mansoor A. Khan (255-267).
Display OmittedHigh strength extended release opioid products, which are indispensable tools in the management of pain, are associated with serious risks of unintentional and potentially fatal overdose, as well as of misuse and abuse that might lead to addiction. The issue of drug abuse becomes increasingly prominent when the dosage forms can be readily manipulated to release a high amount of opioid or to extract the drug in certain products or solvents. One approach to deter opioid drug abuse is by providing novel abuse deterrent formulations (ADF), with properties that may be viewed as barriers to abuse of the product. However, unlike regular extended release formulations, assessment of ADF technologies are challenging, in part due to the great variety of formulation designs available to achieve deterrence of abuse by oral, parenteral, nasal and respiratory routes. With limited prior history or literature information, and lack of compendial standards, evaluation and regulatory approval of these novel drug products become increasingly difficult. The present article describes a risk-based standardized in-vitro approach that can be utilized in general evaluation of abuse deterrent features for all ADF products.
Keywords: Abuse deterrent formulation; Opioid analgesics; Evaluation matrix; Drug abuse; manipulation; Mode of abuse; Syringeability; Injectability;

An elastic liposomal formulation for RNAi-based topical treatment of skin disorders: Proof-of-concept in the treatment of psoriasis by Eline Desmet; Stefanie Bracke; Katrien Forier; Lien Taevernier; Marc C.A. Stuart; Bart De Spiegeleer; Koen Raemdonck; Mireille Van Gele; Jo Lambert (268-274).
Display OmittedRNA interference (RNAi) is a rapidly emerging approach for targeted gene silencing to alleviate disease pathology. However, lack of efficient carriers for targeted delivery delays the clinical translation of RNAi. An interesting target for local RNAi therapeutics is the skin as it allows direct access to target cells. Still, applications are limited due to the effective skin barrier which hinders penetration. Herein, a description is given of a liposomal carrier, called ‘DDC642’, capable of delivering RNAi molecules to the epidermis of impaired and intact human skin, without targeting the dermis or circulatory system. In a psoriasis tissue model, down-regulation of the psoriasis marker human beta-defensin 2 by DDC642-delivered siRNA was confirmed, providing proof-of-concept. These liposomes thus hold great potential as topical delivery system for RNAi therapeutics in the treatment of numerous skin diseases.
Keywords: Lipid-based nanoparticle; Skin disorder; RNA interference; Liposome; Topical drug delivery; Gene therapy;

Display OmittedThe relationships between data of passage through Caco-2 cultured cell lines (log  P app), taken from the literature, for 38 structurally unrelated compounds and both n-octanol lipophilicity parameters (log  P N and log  D 7.4) and phospholipid affinity indexes were investigated. Phospholipid affinity ( log k W I A M ) was experimentally determined by HPLC on two different phospholipid stationary phases and the polar/electrostatic interaction component drug/phospholipids ( Δ log k W IAM ) was calculated according to a method we previously proposed. Log  P app moderately related to lipophilicity values measured at pH 7.4 (log D7.4), according to a parabolic pattern, but poorly related with log k W IAM . Furthermore, a significant inverse linear relationship with Δ log k W IAM values was only observed for the analytes with m.w. >300 Da, for which paracellular diffusion can be considered a minor transport route in vivo. Indeed, it has been reported that Caco-2 passage data also encode secondary passage mechanisms, which participate in a different extent to the jejunal absorption in vivo and cannot be directly equated to the corresponding human in situ log  P eff values, unless a normalization is performed.In an attempt to elucidate this issue, 47 structurally unrelated compounds whose cultured cell line passage data were corrected for the effects of the aqueous boundary layer and paracellular permeability, so as to express transcellular intrinsic permeability, log  P 0 Caco-2/MDCK, were also considered. Highly significant inverse linear relationships were observed between log  P 0 Caco-2/MDCK and Δ log k W IAM values from both IAM.PC.MG (r 2  = 0.765) and IAM.PC.DD2 (r 2  = 0.806) stationary phases whereas the relationships with either lipophilicity in n-octanol or log k W IAM values were very poor. The results of the present study, in complete agreement with those of our recent study on the relationships between jejunal absorption data measured in situ and Δ log k W IAM values, confirm the soundness of Δ log k W IAM parameters in the prediction of the intestinal absorption of drugs. From a mechanistic point of view, they suggest that the polar/electrostatic forces between drugs and phospholipids play a major role in the passage through biomembranes.
Keywords: IAM-HPLC; Lipophilicity; Caco-2; MDCK; Intestinal absorption; Oral bioavailability; Transcellular passage;

Display OmittedTo avoid the toxicological concerns associated to synthetic surfactants, proteins might be an alternative for the stabilization of pharmaceutical nanoemulsions. The present study investigates the use of β-lactoglobulin (β-lg) to stabilize oil in water biocompatible nanoemulsions intended for a pharmaceutical use and prepared by high pressure homogenization (HPH). The effects of composition (nature and weight fraction of oil, β-lg concentration) and of process parameters (pressure and number of cycles) on the droplet size and on the stability of nanoemulsions were thoroughly assessed. The nanoemulsions prepared with β-lg at 1 wt% and with 5 wt% Miglyol 812 (the oil with the lowest viscosity) displayed a relatively small particle size (about 200 nm) and a low polydispersity when a homogenization pressure of 100 MPa was applied for 4 cycles. These nanoemulsions were the most stable formulations over 30 days at least. Emulsification efficiency of β-lg was reduced at higher homogenization pressures (200 MPa and 300 MPa). The effect of HPH process on the interfacial properties of β-lg was evaluated by drop shape analysis. This treatment had an effect neither on the interfacial tension nor on the interfacial dilatational rheology of β-lg at the Miglyol 812/water interface.
Keywords: Nanoemulsions; β-Lactoglobulin; High pressure homogenization; Stability; Interfacial tension; Interfacial rheology;

Anti-glioma activity and the mechanism of cellular uptake of asiatic acid-loaded solid lipid nanoparticles by Tanem Garanti; Aneta Stasik; Andrea Julie Burrow; Mohamed A. Alhnan; Ka-Wai Wan (305-315).
Display OmittedAsiatic acid (AA), a pentacyclic triterpene found in Centella Asiatica, has shown neuroprotective and anti-cancer activity against glioma. However, owing to its poor aqueous solubility, effective delivery and absorption across biological barriers, in particular the blood brain barrier (BBB), are challenging. Solid lipid nanoparticles (SLNs) have shown a promising potential as a drug delivery system to carry lipophilic drugs across the BBB, a major obstacle in brain cancer therapy. Nevertheless, limited information is available about the cytotoxic mechanisms of nano-lipidic carriers with AA on normal and glioma cells. This study assessed the anti-cancer efficacy of AA-loaded SLNs against glioblastoma and their cellular uptake mechanism in comparison with SVG P12 (human foetal glial) cells. SLNs were systematically investigated for three different solid lipids; glyceryl monostearate (MS), glyceryl distearate (DS) and glyceryl tristearate (TS). The non-drug containing MS-SLNs (E-MS-SLNs) did not show any apparent toxicity towards normal SVG P12 cells, whilst the AA-loaded MS-SLNs (AA-MS-SLNs) displayed a more favourable drug release profile and higher cytotoxicity towards U87 MG cells. Therefore, MS-SLNs were chosen for further in vitro studies. Cytotoxicity studies of SLNs (±AA) were performed using MTT assay where AA-SLNs showed significantly higher cytotoxicity towards U87 MG cells than SVG P12 normal cells, as confirmed by flow cell cytometry. Cellular uptake of SLNs also appeared to be preferentially facilitated by energy-dependent endocytosis as evidenced by fluorescence imaging and flow cell cytometry. Using the Annexin V-PI double staining technique, it was found that these AA-MS-SLNs displayed concentration-dependent apoptotic activity on glioma cells, which further confirms the potential of exploiting these AA-loaded MS-SLNs for brain cancer therapy.
Keywords: Triterpenoid; Centella Asiatica; Tristearin; Asiaticoside; Brain delivery; Glyceride; Apoptosis;

Evaluating the sensitivity, reproducibility and flexibility of a method to test hard shell capsules intended for use in dry powder inhalers by Rosalind H.E. Chong; Brian E. Jones; Fernando Díez; James C. Birchall; Sion A. Coulman (316-325).
Display OmittedPharmaceutical tests for hard shell capsules are designed for orally administered capsules. The use of capsules in dry powder inhalers is widespread and increasing and therefore more appropriate tests are required to ensure quality and determine if these capsules are fit for purpose. This study aims to determine the flexibility, reproducibility and sensitivity of a quantitative method that is designed to evaluate the puncture characteristics of different capsule shell formulations under different climatic conditions. A puncture testing method was used to generate force displacement curves for five capsule formulations that were stored and tested at two different temperatures (5 °C and 19 °C). Force-displacement puncture profiles were reproducible for individual capsule shell formulations. The methodology was able to discriminate between capsules produced using different primary materials i.e. gelatin versus hypromellose, as well as more minor changes to capsule formulation i.e. different material grades and excipients. Reduced temperature increased the forces required for capsule puncture however further work is required to confirm its significance. Results indicate the method provides a reproducible and sensitive means of evaluating capsule puncture. Future studies should validate the methodology at different test sites, using different operators and with different capsule shell formulations.
Keywords: Capsule; Hypromellose; Gelatin; Dry powder inhaler; Quality Assurance; Puncture;

Characterisation and in vitro stability of low-dose, lidocaine-loaded poly(vinyl alcohol)-tetrahydroxyborate hydrogels by D.H. Abdelkader; M.A. Osman; S.A. El-Gizawy; A.M. Faheem; P.A. McCarron (326-335).
Display OmittedPoly(vinyl alcohol) hydrogels cross-linked with the tetrahydroxyborate anion possess textural and rheological properties that can be used as novel drug-loaded vehicles for application to traumatic wounds. However, addition of soluble drug substances causes concentration-dependent phase separation and rheological changes. The aim of this work was to investigate the effect of adding a local anaesthetic, but keeping the concentration low in an attempt to prevent these changes. Cross-linked hydrogels prepared from three grades of poly(vinyl alcohol) were characterised rheologically. Temperature sweep studies showed an elevated complex viscosity upon moving from 25 °C to 80 °C, which remained high for 48 h following completion of the cycle. Adhesion to model dermal surfaces achieved a maximum of 2.62 N cm−2 and were greater than that observed to epidermal substrates, with a strong dependence on the rate of detachment used during testing. An optimised formulation (6% w/w PVA (31–50; 99) and 2% w/w THB) containing lidocaine hydrochloride loaded to an upper maximum concentration of 1.5% w/w was assessed for phase separation and drug crystallisation. After six months, crystallisation was present in formulations containing 0.7% and 1.5% lidocaine HCl. Changes in pH in response to increases in lidocaine loading were low. Drug release was shown to operate via a non-Fickian process for all three concentrations, with 60% occurring after approximately 24 h. It can be concluded that using a low concentration of lidocaine hydrochloride in hydrogels based on poly(vinyl alcohol) will result in crystallisation. Furthermore, these hydrogels are unlikely to induce rapid anaesthesia due to the low loading and slow release kinetics.
Keywords: PVA–THB hydrogels; Viscoelasticity; Texture analysis; Lidocaine hydrochloride; Topical delivery;

Oral drug dosage forms administered to hospitalized children: Analysis of 117,665 oral administrations in a French paediatric hospital over a 1-year period. by A. Lajoinie; E. Henin; K.A. Nguyen; S. Malik; Y. Mimouni; J.M. Sapori; V. Bréant; P. Cochat; B. Kassai (336-344).
Display OmittedSelecting the most appropriate dosage form, that ensures safe administration and adherence of medications, is a major issue for children. Marketed drugs, however, have rarely been tested for their use in children. There is a need for more data on drug formulations administered to children to identify unmet needs, and drive future paediatric research. We observed, over a 12-month follow-up, 117,665 oral drug administrations to 1998 hospitalized children. Nine-tenths belonged to five Anatomical Therapeutic Chemical classes: Alimentary tract & metabolism, Nervous system, Cardiovascular system, Anti-infectives for systemic use and Blood & blood forming organs, one third of drug doses administered to school-age children and adolescents were liquids, and extemporaneous capsules were commonly used in younger children. Our study shows that despite the advantages of solid dosage forms and recent evidence from randomized controlled trials showing their acceptability in infants, they are seldom used in paediatric practice.
Keywords: Oral drug administration; Paediatric; Formulation appropriateness; Age appropriateness; Drug formulation; Administration practices;

PEG-derivatized octacosanol as micellar carrier for paclitaxel delivery by Bingyang Chu; Ying Qu; Yixing Huang; Lan Zhang; Xiaoxin Chen; Chaofeng Long; Yunqi He; Caiwen Ou; Zhiyong Qian (345-359).
Display OmittedIn this study, PEG-derivatized octacosanol copolymer was successfully developed to improve the anti-tumor activity and eliminate toxicity of the commercial formulation of paclitaxel (PTX). MPEG2K-C28, the conjugation of monomethoxy Poly(ethylene glycol) 2000 and octacosanol, was readily soluble in aqueous solution and self-assembled to form micelles with small sizes (< 20 nm) that are efficient in encapsulating PTX with a drug loading of 9.38 ± 0.18% and an encapsulation efficiency of 93.90 ± 2.12%. Meanwhile, octacosanol is very safe for humans and amazingly exhibits antitumor activity through inhibition activity of matrix metalloproteinases (MMPs) and translocation of the transcription factor (nuclear factor-kappa B, NF-κB) to the nucleus, which may be able to promote synergistic effects with PTX. A sustained and slower in vitro release behavior was observed in the (PTX micelles) than that of Taxol. PTX micelles exhibited more potent cytotoxicity than Taxol in the 4T1 breast cancer cell line. More interestingly, MPEG2K-C28 selectively inhibited the growth of 4T1 cells rather than the normal cells (HEK293 and L929 cell lines), indicating the antitumor activity of octacosanol remained after conjugation with MPEG. Acute toxicity evaluations indicated that MPEG2K-C28 was a safe drug carrier. Pharmacokinetic study revealed that PTX micelles improved the T 1/2 and AUC of PTX (compared with Taxol) from 1.910 ± 0.139 h and 13.999 ± 1.109 mg/l × h to 2.876 ± 0.532 h and 76.462 ± 8.619 mg/l × h in vivo, respectively. The maximal tolerated dose (MTD) for PTX micelles (ca. 120 mg PTX/kg) in mice was significantly higher than that for Taxol (ca. 20 mg PTX/kg). PTX micelles exhibited slightly better antitumor activity than Taxol but safer in 4T1 breast cancer model in vivo. The cell apoptosis in the immunofluorescent studies and the cell proliferation in the immunohistochemical studies also proved the results. In conclusion, MPEG2K-C28 is a simple, safe and effective drug delivery carrier for PTX, and has some therapeutic effects in 4T1 cells in vitro. PTX micelles showed significant antitumor activity in vivo with low systemic toxicity in 4T1 breast cancer. MPEG2K-C28 micelles entrapping PTX deserve more studies in the future.
Keywords: Micelles; Octacosanol; Paclitaxel; Drug delivery; Cancer therapy;

Humidity-corrected Arrhenius equation: The reference condition approach by Klemen Naveršnik; Rok Jurečič (360-365).
Display OmittedAccelerated and stress stability data is often used to predict shelf life of pharmaceuticals. Temperature, combined with humidity accelerates chemical decomposition and the Arrhenius equation is used to extrapolate accelerated stability results to long-term stability. Statistical estimation of the humidity-corrected Arrhenius equation is not straightforward due to its non-linearity. A two stage nonlinear fitting approach is used in practice, followed by a prediction stage. We developed a single-stage statistical procedure, called the reference condition approach, which has better statistical properties (less collinearity, direct estimation of uncertainty, narrower prediction interval) and is significantly easier to use, compared to the existing approaches. Our statistical model was populated with data from a 35-day stress stability study on a laboratory batch of vitamin tablets and required mere 30 laboratory assay determinations. The stability prediction agreed well with the actual 24-month long term stability of the product. The approach has high potential to assist product formulation, specification setting and stability statements.
Keywords: Arrhenius equation; Forced degradation; Mathematical model; Shelf-life; Accelerated stability; Chemical stability; Kinetics;

Diclofenac-β-cyclodextrin for colonic drug targeting: In vivo performance in rats by Amélia C.F. Vieira; Arménio C. Serra; Francisco J. Veiga; António M.d’A Rocha Gonsalves; Abdul W. Basit; Sudaxshina Murdan (366-370).
Display OmittedThe aim of this in vivo study was to assess the ability of the prodrug conjugate diclofenac-β-cyclodextrin to release diclofenac in the colon following oral administration, using sulfapyridine (a metabolite of sulfasalazine) as a marker of colonic absorption. Two groups of rats were used; the test rats received a suspension containing the two prodrugs, diclofenac-β-cyclodextrin and sulfasalazine, while the control rats received a suspension containing the corresponding free drugs, sodium diclofenac and sulfapyridine. The rats were fasted overnight with free access to water before and throughout the first 12 h of the study. Blood was collected from the tail vein at pre-determined time points and the plasma analyzed for the concentrations of diclofenac and sulfapyridine. Following the oral administration of the two prodrugs, a more extended absorption profile was observed and C max was achieved 10 h post-dose, in contrast to rapid absorption of the free drugs (t max of diclofenac being 1.3 h, and that of sulfapyridine being 2.1 h). In addition to a later t max, conjugation of diclofenac to β-cyclodextrin also resulted in a reduced C max and a reduced AUC. The same t max for diclofenac-β-cyclodextrin as for sulfasalazine confirms the colonic metabolism of diclofenac-β-cyclodextrin. This study shows the potential of this new cyclodextrin-based prodrug to target and release diclofenac specifically in the colon following oral administration.
Keywords: Large intestine; Colonic targeting; Microbiota; Gut; Conjugate; Cyclodextrin;

Closed-loop glycaemic control using an implantable artificial pancreas in diabetic domestic pig (Sus scrofa domesticus) by M.J. Taylor; R. Gregory; P. Tomlins; D. Jacob; J. Hubble; T.S. Sahota (371-378).
Display OmittedThe performance of a completely implantable peritoneal artificial pancreas (AP) has been demonstrated in principle in a live diabetic domestic pig. The device consists of a smart glucose-sensitive gel that forms a gateway to an insulin reservoir and is designed to both sense glucose and deliver insulin in the peritoneal cavity. It can be refilled with insulin via subcutaneous ports and surgery was developed to insert the AP. Diabetes was induced with streptozotocin (STZ), the device filled with insulin (Humulin® R U-500) in situ and the animal observed for several weeks, during which time there was normal access to food and water and several oral glucose challenges. Blood glucose (BG) levels were brought down from >30 mmol/L (540 mg/dL) to non-fasted values between 7 and 13 mmol/L (126–234 mg/dL) about five days after filling the device. Glucose challenge responses improved ultimately so that, starting at 10 mmol/L (180 mg/dL), the BG peak was 18 mmol/L (324 mg/dL) and fell to 7 mmol/L (126 mg/dL) after 30 min, contrasting with intravenous attempts. The reservoir solution was removed after 8 days of blood glucose levels during which they had been increasingly better controlled. A rapid return to diabetic BG levels (30 mmol/L) occurred only after a further 24 days implying some insulin had remained in the device after removal of the reservoir solution. Thus, the closed loop system appeared to have particular influence on the basal and bolus needs for the 8 days in which the reservoir solution was in place and substantial impact for a further 3 weeks. No additional insulin manual adjustment was given during this period.
Keywords: Diabetes; Insulin; Closed-loop; Smart material; Glucose-sensitive; Artificial pancreas; Implantable;

How do the full-generation poly(amido)amine (PAMAM) dendrimers activate blood platelets? Platelet membrane zeta potential and other membrane-associated phenomena by Cezary Watala; Kamil Karolczak; Hassan Kassassir; Karolina Siewiera; Katarzyna Maczynska; Anna Pieniazek; Magdalena Labieniec-Watala (379-389).
Display OmittedWe explored the hypothesis that zeta potential altered by polycations affects blood platelet activation and reactivity, the phenomena associated with membrane lipid fluidity and platelet mitochondrial bioenergetics.PAMAM dendrimers generation- and dose-dependently enhanced zeta potential of platelets (from −10.7 mV to −4.3 mV). Increased expressions of activation markers, P-selectin and the active complex α IIb β 3, as well as significantly enhanced fibrinogen binding occurred upon the in vitro incubation of blood platelets in the presence of PAMAMs G3 and G4 (resp. 62.1% and 69.4% vs. 1.4% and 2.7% in control for P-selectin, P  < 0.0001). PAMAM dendrimers increased fluidity of platelet membrane lipid bilayer, while they did not affect platelet mitochondria respiration. Increased platelet activation and their responses to agonists in vitro were statistically associated with the revealed alterations in zeta potential.Our results support the hypothesis that polycation-mediated “neutralized” zeta potential may underlie the activating effects of PAMAMs on blood platelets.
Keywords: Platelet activation; Electric charge; Membrane potential; Membrane fluidity; Blood flow; Mitochondrial respiration;