Pharmaceutical Research (v.27, #8)

Orphan Nuclear Receptors as Targets for Drug Development by Subhajit Mukherjee; Sridhar Mani (1439-1468).
Orphan nuclear receptors regulate diverse biological processes. These important molecules are ligand-activated transcription factors that act as natural sensors for a wide range of steroid hormones and xenobiotic ligands. Because of their importance in regulating various novel signaling pathways, recent research has focused on identifying xenobiotics targeting these receptors for the treatment of multiple human diseases. In this review, we will highlight these receptors in several physiologic and pathophysiologic actions and demonstrate how their functions can be exploited for the successful development of newer drugs.
Keywords: agonist; antagonist; ligand binding domain; orphan nuclear receptor

Phospholipids and Lipid-Based Formulations in Oral Drug Delivery by Gert Fricker; Torsten Kromp; Armin Wendel; Alfred Blume; Jürgen Zirkel; Herbert Rebmann; Constanze Setzer; Ralf-Olaf Quinkert; Frank Martin; Christel Müller-Goymann (1469-1486).
Phospholipids become increasingly important as formulation excipients and as active ingredients per se. The present article summarizes particular features of commonly used phospholipids and their application spectrum within oral drug formulation and elucidates current strategies to improve bioavailability and disposition of orally administered drugs. Advantages of phospholipids formulations not only comprise enhanced bioavailability of drugs with low aqueous solubility or low membrane penetration potential, but also improvement or alteration of uptake and release of drugs, protection of sensitive active agents from degradation in the gastrointestinal tract, reduction of gastrointestinal side effects of non-steroidal anti-inflammatory drugs and even masking of bitter taste of orally applied drugs. Technological strategies to achieve these effects are highly diverse and offer various possibilities of liquid, semi-liquid and solid lipid-based formulations for drug delivery optimization.
Keywords: emulsion; liposome; phospholipid; self-emulsifying drug delivery systems; solid lipid nanoparticles

Gene Therapy: A Pharmacokinetic/Pharmacodynamic Modelling Overview by Zinnia P. Parra-Guillén; Gloria González-Aseguinolaza; Pedro Berraondo; Iñaki F. Trocóniz (1487-1497).
Since gene therapy started over 20 years ago, more than one-thousand clinical trials have been carried out. Nonviral vectors present interesting properties for their clinical application, but their efficiency in vivo is relatively low, and further improvements in these vectors are needed. Elucidating how nonviral vectors behave at the intracellular level is enlightening for vector improvement and optimization. Model-based approach is a powerful tool to understand and describe the different processes that gene transfer systems should overcome inside the body. Model-based approach allows for proposing and predicting the effect of parameter changes on the overall gene therapy response, as well as the known application of the pharmacokinetic/pharmacodynamic modelling in conventional therapies. The objective of this paper is to critically review the works in which the time-course of naked or formulated DNA have been quantitatively studied or modelled.
Keywords: computational modelling; gene therapy; pharmacokinetic/pharmacodynamic modelling

Novel Mixed Polymeric Micelles for Enhancing Delivery of Anticancer Drug and Overcoming Multidrug Resistance in Tumor Cell Lines Simultaneously by Xinru Li; Pingzhu Li; Yanhui Zhang; Yanxia Zhou; Xingwei Chen; Yanqing Huang; Yan Liu (1498-1511).
To evaluate novel mixed polymeric micelles based on monomethoxy poly(ethylene glycol)-poly(D,L-lactic acid) (mPEG-PLA) and Pluronic L61 for delivery of paclitaxel (PTX) to circumvent unfavorable effects resulting from Cremophore EL in Cremophore EL-based PTX formulation and overcoming multidrug resistance (MDR) in tumor cells at the same time.PTX-loaded plain micelles and mixed micelles were prepared and characterized by determining PTX release in vitro, MDR reversal effect in human breast cancer MDR MCF-7/ADR cell sublines and pharmacokinetics in vivo.Both PTX-loaded plain micelles and mixed micelles had similar in vitro release profile. Mixed micellar PTX significantly reduced IC50 of PTX in MCF-7/ADR cells compared to free PTX and plain micellar PTX, and mixed micelles substantially enhanced cellular accumulation of R 123 in MCF-7/ADR cells compared to free R123 and plain micelles. PTX-loaded mixed micelles with lower content of L61 exhibited comparable cytotoxicity to that observed with Cremophore EL-based PTX formulation in inhibiting the growth of MCF-7/ADR cells. Moreover, plain micelles and mixed micelles retained the pharmacokinetic characteristics of PTX in rats compared with Cremophore EL-based PTX formulation.This study suggested that the mixed micelles could enhance delivery of PTX and cell-killing effect for MDR MCF-7/ADR cells.
Keywords: methoxy poly(ethylene glycol)-poly(D,L-lactic acid) (mPEG-PLA); mixed polymeric micelles; multidrug resistance (MDR); paclitaxel; pluronic L61

To analyze contribution of short aggregation-prone regions (APRs), which may self-associate via cross-β motif and were earlier identified in therapeutic mAbs, towards antigen recognition via structural analyses of antibody-antigen complexes.A dataset of 29 publically available high-resolution crystal structures of Fab-antigen complexes was collected. Contribution of APRs towards the surface areas of the Fabs buried by the cognate antigens was computed. Propensities of amino acids to occur in APRs and to be involved in antigen binding were compared. Coincidence between APRs and individual CDR loops was examined.All Fabs in the dataset contain at least one APR in CDR loops and adjacent framework β-strands. The average contribution of APRs towards buried surface area of Fabs is 16.0 ± 10.7%. Aggregation and antigen recognition may be coupled via aromatic residues (Tyr, Trp), which occur with high propensities in both APRs and antigen binding sites. APRs are infrequent in the heavy chain CDR 3 (H3) loops (7%), but are frequent in H2 loops (45%).Co-incidence of APRs with antigen recognition sites can potentially lead to the loss of function upon aggregation. Rational structure-based design or selection strategies are suggested for biotherapeutics with improved druggability while maintaining potency.
Keywords: biotherapeutics; drug development; fab-antigen interface; monoclonal antibody; structure

A computational fluid dynamics (CFD) study examined the impact of particle size on dissolution rate and residence of intravitreal suspension depots of Triamcinolone Acetonide (TAC).A model for the rabbit eye was constructed using insights from high-resolution NMR imaging studies (Sawada 2002). The current model was compared to other published simulations in its ability to predict clearance of various intravitreally injected materials. Suspension depots were constructed explicitly rendering individual particles in various configurations: 4 or 16 mg drug confined to a 100 μL spherical depot, or 4 mg exploded to fill the entire vitreous. Particle size was reduced systematically in each configuration. The convective diffusion/dissolution process was simulated using a multiphase model.Release rate became independent of particle diameter below a certain value. The size-independent limits occurred for particle diameters ranging from 77 to 428 μM depending upon the depot configuration. Residence time predicted for the spherical depots in the size-independent limit was comparable to that observed in vivo.Since the size-independent limit was several-fold greater than the particle size of commercially available pharmaceutical TAC suspensions, differences in particle size amongst such products are predicted to be immaterial to their duration or performance.
Keywords: computational fluid dynamics (CFD); convective diffusion; dissolution; drug clearance; intravitreal injection; volume of fluid (VOF)

This study investigates the mechanisms of G3.5 poly (amido amine) dendrimer cellular uptake, intracellular trafficking, transepithelial transport and tight junction modulation in Caco-2 cells in the context of oral drug delivery.Chemical inhibitors blocking clathrin-, caveolin- and dynamin-dependent endocytosis pathways were used to investigate the mechanisms of dendrimer cellular uptake and transport across Caco-2 cells using flow cytometry and confocal microscopy.Dendrimer cellular uptake was found to be dynamin-dependent and was reduced by both clathrin and caveolin endocytosis inhibitors, while transepithelial transport was only dependent on dynamin- and clathrin-mediated endocytosis. Dendrimers were quickly trafficked to the lysosomes after 15 min of incubation and showed increased endosomal accumulation at later time points, suggesting saturation of this pathway. Dendrimers were unable to open tight junctions in cell monolayers treated with dynasore, a selective inhibitor of dynamin, confirming that dendrimer internalization promotes tight junction modulation.G3.5 PAMAM dendrimers take advantage of several receptor-mediated endocytosis pathways for cellular entry in Caco-2 cells. Dendrimer internalization by dynamin-dependent mechanisms promotes tight junction opening, suggesting that dendrimers act on intracellular cytoskeletal proteins to modulate tight junctions, thus catalyzing their own transport via the paracellular route.
Keywords: endocytosis; oral drug delivery; PAMAM dendrimers; tight junctions; transport

Fast Surface Crystallization of Amorphous Griseofulvin Below T g by Lei Zhu; Janan Jona; Karthik Nagapudi; Tian Wu (1558-1567).
To study crystal growth rates of amorphous griseofulvin (GSF) below its glass transition temperature (T g) and the effect of surface crystallization on the overall crystallization kinetics of amorphous GSF.Amorphous GSF was generated by melt quenching. Surface and bulk crystal growth rates were determined using polarized light microscope. X-ray powder diffraction (XRPD) and Raman microscopy were used to identify the polymorph of the crystals. Crystallization kinetics of amorphous GSF powder stored at 40°C (T g−48°C) and room temperature (T g−66°C) was monitored using XRPD.Crystal growth at the surface of amorphous GSF is 10- to 100-fold faster than that in the bulk. The surface crystal growth can be suppressed by an ultrathin gold coating. Below T g, the crystallization of amorphous GSF powder was biphasic with a rapid initial crystallization stage dominated by the surface crystallization and a slow or suspended late stage controlled by the bulk crystallization.GSF exhibits the fastest surface crystallization kinetics among the known amorphous pharmaceutical solids. Well below T g, surface crystallization dominated the overall crystallization kinetics of amorphous GSF powder. Thus, surface crystallization should be distinguished from bulk crystallization in studying, modeling and controlling the crystallization of amorphous solids.
Keywords: amorphous solids; coating; crystallization kinetic; griseofulvin; surface-enhanced crystallization

Glucuronidation via UDP-glucuronosyltransferases (or UGTs) is a major metabolic pathway. The purposes of this study are to determine the UGT-isoform-specific metabolic fingerprint (or GSMF) of wogonin and oroxylin A, and to use isoform-specific metabolism rates and kinetics to determine and describe their glucuronidation behaviors in tissue microsomes. In vitro glucuronidation rates and profiles were measured using expressed UGTs and human intestinal and liver microsomes.GSMF experiments indicated that both flavonoids were metabolized mainly by UGT1As, with major contributions from UGT1A3 and UGT1A7-1A10. Isoform-specific metabolism showed that kinetic profiles obtained using expressed UGT1A3 and UGT1A7-1A10 could fit to known kinetic models. Glucuronidation of both flavonoids in human intestinal and liver microsomes followed simple Michaelis-Menten kinetics. A comparison of the kinetic parameters and profiles suggests that UGT1A9 is likely the main isoform responsible for liver metabolism. In contrast, a combination of UGT1As with a major contribution from UGT1A10 contributed to their intestinal metabolism. Correlation studies clearly showed that UGT isoform-specific metabolism could describe their metabolism rates and profiles in human liver and intestinal microsomes.GSMF and isoform-specific metabolism profiles can determine and describe glucuronidation rates and profiles in human tissue microsomes.
Keywords: flavonoids; intestine; liver; UGT; wogonin

Mannan-Modified Solid Lipid Nanoparticles for Targeted Gene Delivery to Alveolar Macrophages by Wangyang Yu; Chunxi Liu; Yu Liu; Na Zhang; Wenfang Xu (1584-1596).
Cationic solid lipid nanoparticles (SLN) have established themselves during the past decades. They can efficiently bind DNA directly via ionic interaction and mediate gene transfection. One major problem with SLN is the lack of cell-targeting ability. In the present study, a mannan-based PE-grafted ligand was synthesized and used for the surface modification of DNA-loaded cationic SLN to prepare Man-SLN-DNA.For in vitro test, the cytotoxicity and transfection investigation was carried out on murine macrophage cell line RAW 264.7. For in vivo evaluation, Man-SLN-DNA was delivered into the lung of the rats, and the alveolar macrophages (AM) were isolated for the fluorescence determination of transfection efficiency.When compared with non-modified SLN-DNA and Lipofectamine 2000-DNA, Man-SLN-DNA produced the highest gene expressions, especially in vivo.These results demonstrated the active targeting ability of this kind of mannan-modified DNA-loaded vehicles, which may have great potential for targeted gene delivery.
Keywords: active targeting; cationic solid lipid nanoparticles; gene delivery; mannan-based PE-grafted ligand; surface modification

To investigate the effect of the antibiotic release profiles of levofloxacin-loaded polymeric nanoparticles on their antibacterial efficacy against E. coli biofilm cells.Three distinct antibiotic release profiles are produced by encapsulating levofloxacin in PCL and PLGA nanoparticles by nanoprecipitation and emulsification-solvent-evaporation methods. The antibacterial efficacy is examined over six days by time-dependent biofilm susceptibility testing that takes into account the effects of the biofilm age, antibiotic exposure history, and simulated drug removal.Biofilm cells that survive the initial antibiotic exposure exhibit a higher antibiotic tolerance than fresh biofilm cells, where the lower the initial exposure, the higher the tolerance of the surviving biofilm cells. The lower antibiotic susceptibility of the surviving biofilm cells is transferred to their planktonic cell progeny, which can subsequently form new biofilm colonies having a higher antibiotic tolerance, hence exacerbating the infections. A biphasic extended release profile at an appropriate dose can inhibit the biofilm growth for four days, therefore reducing the dosing frequency. The importance of a high initial antibiotic exposure renders a slow release profile ineffective despite the same dosing amount.The antibiotic release profile has an equally significant influence on the biofilm eradication rate as the antibiotic dose.
Keywords: antibiotics aerosols; biofilm; bronchiectasis; cystic fibrosis; persister cells

High-Throughput Screening System for Identifying Phototoxic Potential of Drug Candidates Based on Derivatives of Reactive Oxygen Metabolites by Satomi Onoue; Masanori Ochi; Graham Gandy; Yoshiki Seto; Naoko Igarashi; Yukinori Yamauchi; Shizuo Yamada (1610-1619).
The present study aimed to develop a high-throughput screening strategy for predicting the phototoxic potential of pharmaceutical substances, using a derivatives-of-reactive-oxygen-metabolites (D-ROM) assay.The assay conditions of the D-ROM assay were optimized with a focus on screening run time, sensitivity, solvent system, and reproducibility. The phototoxic potentials of 25 model compounds were assessed by the D-ROM assay, as well as by other screening systems for comparison, including the reactive oxygen species (ROS) assay, the DNA-photocleavage assay, and the 3T3 neutral red uptake phototoxicity test (3T3 NRU PT).Some phototoxic drugs tended to yield D-ROM when exposed to simulated sunlight (250 W/m2), whereas D-ROM generation was negligible for non-phototoxic chemicals. Compared with the ROS assay, the assay procedure for the D-ROM assay was highly simplified with a marked reduction in screening run time. Comparative experiments also demonstrated that D-ROM data were related to the outcomes of the DNA-photocleavage assay and the 3T3 NRU PT, with prediction accuracies of 76 and 72%, respectively.The D-ROM assay has potential for identifying the phototoxic potential of a large number of new drugs as a 1st screening system in the early stages of drug discovery.
Keywords: 3T3 neutral red uptake phototoxicity test; derivatives of reactive oxygen metabolites; photogenotoxicity; phototoxicity; reactive oxygen species

Characterization of the Ascending Colon Fluids in Ulcerative Colitis by Maria Vertzoni; Konstantinos Goumas; Erik Söderlind; Bertil Abrahamsson; Jennifer B. Dressman; Androniki Poulou; Christos Reppas (1620-1626).
To characterize the fluid composition in ascending colon of fasted adults with ulcerative colitis in relapse and in remission with a view to predicting variations on dosage form performance in the lower inflamed gut.Twelve patients participated in a two-phase, crossover study. Enrolment to the relapse phase (Phase A) and designation of the remission state for the second colonoscopy (Phase B) were based on Clinical Rachmilewicz Index values. Samples were analyzed for pH and buffer capacity immediately upon collection. After ultracentrifugation, osmolality, surface tension, soluble protein, soluble carbohydrates, and the levels of ten bile acids, seven short-chain fatty acids (SCFAs), three long-chain fatty acids, triglycerides, diglycerides, monoglycerides, phosphatidylcholine, and cholesterol were measured.Total SCFAs are significantly decreased in relapse, but pH remains unaffected. Regardless of remission/relapse status, pH and isobutyric acid levels are lower than in healthy adults. Buffer capacity, osmolality, and soluble protein are higher than in healthy adults. Treatment with prednisolone increases the volume of intracolonic contents.Variations in fluid composition of the ascending colon with activity and severity of ulcerative colitis may have an impact on the performance of orally administered products that are targeted to release the therapeutic agent in the colon.
Keywords: ascending colon; colonic targeting; lumenal composition; ulcerative colitis

Thermosensitive Micelles from PEG-Based Ether-anhydride Triblock Copolymers by Aijun Zhao; Shaobing Zhou; Qi Zhou; Tao Chen (1627-1643).
The thermosensitive micelles based on the poly(PEG:CPP:SA) terpolymer composed of poly(ethylene glycol) (PEG), 1,3-bis(carboxyphenoxy) propane (CPP) and sebacic acid (SA) were fabricated for application as a promising drug carrier.The terpolymer can self-assemble into micelles in water by a precipitation technology. The sol–gel transition behaviors were investigated by the tube-tilting method and dynamic rheology. The drug release behaviors were investigated in phosphate-buffered solution (PBS) at 25, 37 and 45°C, respectively, and the tumor cell growth inhibition assays were also evaluated.The diameters of these micelles increased as the environmental temperature, and the length of CPP and SA chains increased. The micelles with a low concentration underwent sol-to-nanogel transition as temperature increased from the room temperature to the body temperature, while the polymer solutions with a high concentration underwent sol-to-gel transition as the temperature increased from 20 to 70°C. In vitro release profiles consisted of a burst release followed a sustained release. The cytotoxicity results showed that the terpolymer micelles were biocompatible, and the encapsulated doxorubicin. HCl maintained its potent anti-tumor effect.These micelles may bring the ether-anhydride family of polymers great potential as a novel carrier in nanomedicine.
Keywords: biodegradable; ether-anhydride; polymeric micelles; self-assemble; thermo-responsive

To investigate the permeation of two ionisable drug molecules, warfarin and verapamil, across artificial membranes. For the first time since the introduction of the parallel artificial membrane permeation assay (PAMPA) in 1998, in situ permeation-time profiles of drug molecules are studied.The method employs a rotating-diffusion cell where the donor and acceptor compartments are separated by a lipid-impregnated artificial membrane. The permeation of the solute is investigated under well-defined hydrodynamic conditions with control over the unstirred water layer. The flux of the permeating molecule is analysed in situ using UV spectrophotometry. In situ permeation-time profiles are obtained under hydrodynamic control and used to determine permeability coefficients. An advanced analytical transport model is derived to account for the membrane retention, two-way flux and pH gradient between the two compartments. Moreover, a numerical permeation model was developed to rationalise the time-dependent permeation profiles. The membrane permeability, intrinsic permeability and unstirred water permeability coefficients of two drug molecules are obtained from two independent methods, hydrodynamic extrapolation and pH profiling, and the results are compared.Both warfarin and verapamil exhibit high permeability values, which is consistent with the high fraction absorbed in human. Our results demonstrate that a considerable lag-time, varying with the solute lipophilicity and stirring rate, exists in membrane permeation and leads to incorrect compound ranking if it is not treated properly. Comparison of the permeability data as a function of pH and stirring rate suggests that some transport of the ionized molecules occurs, most likely via ion-pairing.
Keywords: hydrodynamic control; in situ permeation; PAMPA; permeability; unstirred water layer

Development of New Localized Drug Delivery System Based on Ceftriaxone-Sulbactam Composite Drug Impregnated Porous Hydroxyapatite: A Systematic Approach for In Vitro and In Vivo Animal Trial by Biswanath Kundu; Chidambaram Soundrapandian; Samit K. Nandi; Prasenjit Mukherjee; Nandadulal Dandapat; Subhasis Roy; Bakul K. Datta; Tapan K. Mandal; Debabrata Basu; Rupnarayan N. Bhattacharya (1659-1676).
Present investigation deals with an extensive approach incorporating in vitro and in vivo experimentation to treat chronic osteomyelitis, using hydroxyapatite porous scaffolds.Hydroxyapatite was synthesized in the laboratory by wet chemical method, different porous scaffolds have been fabricated. In vitro studies include variation of porosity with interconnectivity, pore-drug interfacial studies by SEM-EDAX and drug elution studies (by HPLC) both in contact with PBS and SBF at ~37°C. In vivo trials were based on experimental osteomyelitis in rabbit model induced in tibia by Staphylococcus aureus. Characterizations included observation of histopathology, radiology and estimation of drug in both bone and serum for 42 days by HPLC method and subsequent bone-biomaterial interface by SEM.It was established that lower pore percentage with a distribution of mainly micro-pores were found to be superior over the higher pore percentage both in vitro and in vivo. The criteria was matched with the 50N50H samples which had 50–55% porosity with an average pore size ~110 μm, having higher interconnectivity (10–100 μm), moderately high adsorption efficiency (~50%) when loaded with CFS (drug combinations consisting of irreversible b-lactamase inhibitor and b-lactam antibiotic). CFS release from HAp implants were faster in PBS than SBF. Further, both the results of in vitro and in vivo drug elution after 42 days showed release higher than minimum inhibitory concentration of CFS against Staphylococcus aureus. In vivo studies also proved the superiority of CFS loaded HAp implants than parenteral group based on eradication of infection and new bone formation.HAp based porous scaffold loaded with CFS and designed porosity (in terms of micro- and macro-porosity, interconnectivity) was found to be an ideal delivery system which could locally, sustainably release the composite antibiotic in reliable manner both in terms of in vitro drug elution behaviour in contact with SBF and in vivo animal trial.
Keywords: ceftriaxone-sulbactam composite; in vivo animal trial; osteomyelitis and new bone formation; porous hydroxyapatite; SBF

Bioadhesive Films Containing Benzocaine: Correlation Between In Vitro Permeation and In Vivo Local Anesthetic Effect by Daniele Ribeiro de Araujo; Cristina Padula; Cíntia Maria Saia Cereda; Giovana Radomille Tófoli; Rui Barbosa Brito Jr.; Eneida de Paula; Sara Nicoli; Patrizia Santi (1677-1686).
The aim of this work was to develop anesthetic bioadhesive films containing benzocaine and study their in vitro skin permeation and in vivo performance, in comparison with commercial formulations.Films containing 3% and 5% w/w of benzocaine were prepared and characterized by weight, drug content, thickness and morphology. In vitro permeation assays were performed in vertical diffusion cells using full-thickness pig ear skin as barrier. Intensity and duration of analgesia were evaluated in rats by tail-flick test, and skin histological analysis was carried out.Tail-flick test showed that the duration of benzocaine-induced analgesia was significantly prolonged with the films compared to commercial creams, in agreement with the higher in vitro permeation. Histological analysis of the rat tail skin did not reveal morphological tissue changes nor cell infiltration signs after application of the commercial creams or films.Results from our study indicate that the films developed in this work can be considered as innovative dermal/transdermal therapeutic systems for benzocaine local delivery.
Keywords: benzocaine; in vitro–in vivo correlation; local anestetic; patch; transdermal

Evaluation of Docetaxel-Loaded Intravenous Lipid Emulsion: Pharmacokinetics, Tissue Distribution, Antitumor Activity, Safety and Toxicity by Mingming Zhao; Min Su; Xia Lin; Yanfei Luo; Haibing He; Cuifang Cai; Xing Tang (1687-1702).
The purpose of this study was to carry out a detailed evaluation of an intravenous lipid emulsion for docetaxel (DLE) without Tween 80 before clinical administration.The pharmacokinetics in rats and beagle dogs, tissue distribution, antitumor activity, safety test and toxicity of DLE have been investigated systematically to evaluate the formulation and compared with Taxotere® (DS).The pharmacokinetic study in rats revealed that DLE exhibited higher plasma concentrations and AUC than DS, and a good correlation was observed between AUC and dose, while, in beagle dogs, the DLE was bioequivalent to DS. The tissue distribution study showed that the profiles of the two formulations were similar, indicating the DLE did not change the distribution of docetaxel in vivo. Furthermore, DLE was as safe as DS in the safety investigation and displayed significant antitumor activities against the A549, BEL7402 and BCAP-37 cell lines in nude mice, similar to DS. The corresponding results of the long-term toxic study demonstrated the DLE was less toxic than DS, and the toxic effects could be reversed.The DLE investigated in this paper was found to be an attractive new formulation and an appropriate choice for the clinical administration of docetaxel.
Keywords: docetaxel; evaluation; lipid emulsion

Effects of Commonly Used Excipients on the Expression of CYP3A4 in Colon and Liver Cells by Leslie Tompkins; Caitlin Lynch; Sam Haidar; James Polli; Hongbing Wang (1703-1712).
The objective of this investigation was to assess whether common pharmaceutical excipients regulate the expression of drug-metabolizing enzymes in human colon and liver cells.Nineteen commonly used excipients were evaluated using a panel of experiments including cell-based human PXR activation assays, real-time RT-PCR assays for CYP3A4 mRNA expression, and immunoblot analysis of CYP3A4 protein expression in immortalized human liver cells (HepG2 and Fa2N4), human primary hepatocytes, and the intestinal LS174T cell models.No excipient activated human PXR or practically induced CYP3A4. However, three excipients (polysorbate 80, pregelatinized starch, and hydroxypropyl methylcellulose) tended to decrease mRNA and protein expression across experimental models.This study represents the first investigation of the potential role of excipients in the expression of drug-metabolizing enzymes. Findings imply that some excipients may hold potential for excipient-drug interactions by repression of CYP3A4 expression.
Keywords: excipients; CYP3A4; PXR; induction; repression

Novel Cyclopeptides for the Design of MMP Directed Delivery Devices: A Novel Smart Delivery Paradigm by El-Farouck Moustoifa; Mohamed-Anis Alouini; Arnaud Salaün; Thomas Berthelot; Aghleb Bartegi; Sandra Albenque-Rubio; Gérard Déléris (1713-1721).
Matrix metalloproteinases (MMP) are a family of proteolytic enzymes, the expression of which in a key step of tumor progression has been better defined recently. The studies highlighted the ongoing need for very specific inhibitors, substrates or release devices designed to be selective for one or at least very few MMPs.This report deals with the design, synthesis and in vitro evaluation of linear and especially novel cyclic peptidic moieties, embodying MMP cleavable sequences designed to answer these questions. FRET (fluorescence resonance energy transfer) labelling via chromophore-modified amino-acids was used to give access to enzyme kinetics.Evaluation of these peptides showed that cyclisation gives rise to high specificity for certain MMP, suggesting that this approach could provide very specific MMP substrate. Moreover, cyclic structures present a very good plasma stability.These original derivatives could allow the design of MMP-controlled delivery devices, the specificity of which will be retained in complex biological media and in vivo.
Keywords: cancer; cyclic peptide; delivery device; fluorescent substrate; MMP

Deposition of Albuterol Aerosol Generated by Pneumatic Nebulizer in the Sophia Anatomical Infant Nose-Throat (SAINT) Model by Beth L. Laube; Gail Sharpless; Charles Shermer; Omer Nasir; Vincent Sullivan; Kenneth Powell (1722-1729).
To quantify distribution of albuterol aerosol generated by a pneumatic nebulizer within the nose and lungs of a model of a 9-month-old child (SAINT) and aerosol loss to the environment, during simulated breathing at increasing tidal volumes (TVs). 99mtechnetium-labeled albuterol aerosol was generated by an IPI nebulizer with face-mask. Deposition was quantified as a percentage of emitted dose using gamma scintigraphy.Lung deposition was similar for all TVs, averaging 7.17 ± 0.01%, 9.34 ± 0.01% and 9.41 ± 0.02% at 50, 100 and 200 mL TV, respectively. In contrast, nose deposition increased significantly with TV, averaging 4.40 ± 0.02%, 11.39 ± 0.02% and 22.12 ± 0.02% at 50 mL, 100 mL and 200 mL TV, respectively (all p < 0.0167). Aerosol loss to the environment was significantly lower at 200 mL TV (53.81 ± 0.04%), compared to 50 mL (71.99 ± 0.02%) (p < 0.0167).Our results suggest that nasal deposition of albuterol aerosol generated by a pneumatic nebulizer in 9-month-old infants may be significantly affected by changes in TV, ranging between 50 to 200 mL, whereas total lung deposition may not be affected. These results also predict that environmental losses would be highest when administering to a child breathing at 50 mL TV. These data should be useful to companies who are working to improve aerosol delivery systems to treat infants.
Keywords: albuterol; pneumatic nebulizer; aerosol deposition; scintigraphy; modeling

In VitroIn Vivo Correlations of Scalable PLGA-Risperidone Implants for the Treatment of Schizophrenia by Laura C. Amann; Michael J. Gandal; Robert Lin; Yuling Liang; Steven J. Siegel (1730-1737).
Nonadherence to antipsychotic medications is a major obstacle preventing optimal outcomes for patients with schizophrenia. Extended release systems exist in the form of depot injections, but these formulations exhibit several disadvantages. To address these concerns, we previously demonstrated proof of concept for an antipsychotic implant containing risperidone and the biodegradable polymer poly(lactic-co-glycolic) acid (PLGA).We build upon recently published data by utilizing a scalable single-screw extrusion system for the production of PLGA-risperidone implants. Implants were composed of 40% risperidone and 60% PLGA, with varying ratios of lactide to glycolide (50:50, 65:35, 75:25 or 85:15). Risperidone release was assessed in vitro and in vivo in rats, and Level A, B and C correlations (IVIVCs) attempted for all. Bioavailability was verified with locomotor testingLevel B analysis yielded the greatest correlation between in vitro and in vivo data (R 2 = 0.9425), while Level C yielded the lowest (R 2 = 0.8336). Although, based on qualitative results, a Level A correlation was not achieved, it did produce an R 2 of 0.9261. Locomotor testing demonstrated that peak serum concentrations coincide with significant reductions in activity.Data demonstrate the applicability of our modeling system and advance long-term, implantable antipsychotics toward clinical application.
Keywords: implant; in vitroin vivo correlation; PLGA; risperidone; schizophrenia

Chitosan Film Containing Poly(D,L-Lactic-Co-Glycolic Acid) Nanoparticles: A Platform for Localized Dual-Drug Release by Dayane B. Tada; Surinder Singh; Dattatri Nagesha; Evan Jost; Craig O. Levy; Evin Gultepe; Robert Cormack; G. Mike Makrigiorgos; Srinivas Sridhar (1738-1745).
To characterize and evaluate chitosan film containing PLGA nanoparticles (NPs) as a platform for localized dual-drug release.Fluorescent Paclitaxel (FPTX), a hydrophobic drug, was incorporated into PLGA NPs. FPTX-loaded PLGA NPs and Carboxyfluorescein (CF), a hydrophilic model drug, were embedded into chitosan films. Release of CF and NPs from chitosan and release of FPTX from PLGA NPs were monitored by fluorescence. The stability of the platform was observed through SEM and dynamic light scattering (DLS).Chitosan films containing CF and FPTX-loaded PLGA NPs showed a biphasic release profile. In the first phase, 78% of CF and 34% of NPs were released within few days. In the second phase, the release was slower, showing an additional release of 22% of CF and 18% of NPs after 3 weeks. SEM images and DLS measurements showed that NP release depends on film degradation rate. FPTX-loaded PLGA NPs showed the release of 19.8% of total drug in 2 days, and no additional release was detected in the next 26 days.The ability of chitosan film containing PLGA NPs to coat gold surface and to incorporate and release two different drugs of different hydrophilicity make it a promising platform for localized dual-drug release.
Keywords: chitosan; PLGA; paclitaxel; drug release; implants coating

Application of Nanotechnology in Cosmetics by Li Mu; Robert L. Sprando (1746-1749).

Hyperthermia and Thermosensitive Liposomes for Improved Delivery of Chemotherapeutic Drugs to Solid Tumors by Gerben A. Koning; Alexander M. M. Eggermont; Lars H. Lindner; Timo L. M. ten Hagen (1750-1754).
Lipid-based nanocarriers or liposomes have been proven successful in the delivery of chemotherapeutic agents and are currently applied clinically in the treatment of various types of cancer. Liposomes offer the advantage of a high drug payload, decreased drug toxicity and enhanced drug accumulation at tumor sites. Increased accumulation is due to the relatively leaky tumor vasculature that allows liposome extravasation. Between different types of tumors and even within one tumor, vascular permeability and thus liposome extravasation may differ greatly. Furthermore, upon accumulation of liposomes in the tumor area, drug bioavailability is not guaranteed. At present, these are the major issues for clinically used liposomal drugs.Mild hyperthermia (HT), the heating of tumor tissue to temperatures of up to 43°C, has been developed in the past decades as an established and efficacious treatment modality in combination with chemo- and radiotherapy. HT can be used to further improve liposomal chemotherapy in two ways: HT is known to increase vascular permeability in solid tumors and may therefore increase levels of liposome accumulation, and thermosensitive liposomes have been developed that can be triggered to release their contents upon hyperthermia. By applying these two strategies, drug delivery to tumors can be strongly enhanced.
Keywords: cancer; chemotherapy; hyperthermia; nanomedicine; thermosensitive liposomes; triggered release

AAPS Connection (1755-1756).