Pharmaceutical Research (v.33, #1)

Risk Management for Materials and Components Used in Orally Inhaled and Nasal Drug Products by Cheryl L. M. Stults; Gaby Reckzuegel; Arthur Bailey; Jonathan Petersen; Lisa Dick; Thomas Feinberg; Andrew Feilden; Sara Miller; James Connors; Lee M. Nagao (1-17).
Keywords: container closure systems; nasal products; orally inhaled products; packaging; risk management

Polymeric Micelles: Recent Advancements in the Delivery of Anticancer Drugs by Avinash Gothwal; Iliyas Khan; Umesh Gupta (18-39).
Nanotechnology, in health and medicine, extensively improves the safety and efficacy of different therapeutic agents, particularly the aspects related to drug delivery and targeting. Among various nano-carriers, polymer based macromolecular approaches have resulted in improved drug delivery for the diseases like cancers, diabetes, autoimmune disorders and many more. Polymeric micelles consisting of hydrophilic exterior and hydrophobic core have established a record of anticancer drug delivery from the laboratory to commercial reality. The nanometric size, tailor made functionality, multiple choices of polymeric micelle synthesis and stability are the unique properties, which have attracted scientists and researchers around the world to work upon in this opportunistic drug carrier. The capability of polymeric micelles as nano-carriers are nowhere less significant than nanoparticles, liposomes and other nanocarriers, as per as the commercial feasibility and presence is concerned. In fact polymeric micelles are among the most extensively studied delivery platforms for the effective treatment of different cancers as well as non-cancerous disorders. The present review highlights the sequential and recent developments in the design, synthesis, characterization and evaluation of polymeric micelles to achieve the effective anticancer drug delivery. The future possibilities and clinical outcome have also been discussed, briefly.
Keywords: cancer; hydrophilic corona; hydrophobic core; nanotechnology; polymeric micelles

Assessment of Interspecies Differences in Drug-Induced QTc Interval Prolongation in Cynomolgus Monkeys, Dogs and Humans by V. F. S. Dubois; W. E. A. de Witte; S. A. G. Visser; M. Danhof; O. Della Pasqua (40-51).
The selection of the most suitable animal species and subsequent translation of the concentration-effect relationship to humans are critical steps for accurate assessment of the pro-arrhythmic risk of candidate molecules. The objective of this investigation was to assess quantitatively the differences in the QTc prolonging effects of moxifloxacin between cynomolgus monkeys, dogs and humans. The impact of interspecies differences is also illustrated for a new candidate molecule.Pharmacokinetic data and ECG recordings from pre-clinical protocols in monkeys and dogs and from a phase I trial in healthy subjects were identified for the purpose of this analysis. A previously established Bayesian model describing the combined effect of heart rate, circadian variation and drug effect on the QT interval was used to describe the pharmacokinetic-pharmacodynamic relationships. The probability of a ≥10 ms increase in QT was derived as measure of the pro-arrhythmic effect.For moxifloxacin, the concentrations associated with a 50% probability of QT prolongation ≥10 ms (Cp50) varied from 20.3 to 6.4 and 2.6 μM in dogs, monkeys and humans, respectively. For NCE05, these values were 0.4 μM vs 2.0 μM for monkeys and humans, respectively.Our findings reveal significant interspecies differences in the QT-prolonging effect of moxifloxacin. In addition to the dissimilarity in pharmacokinetics across species, it is likely that differences in pharmacodynamics also play an important role. It appears that, regardless of the animal model used, a translation function is needed to predict concentration-effect relationships in humans.
Keywords: cardiovascular safety; drug development; interspecies differences; PKPD modelling; QT interval prolongation

Assessment of Age-Related Changes in Pediatric Gastrointestinal Solubility by Anil R. Maharaj; Andrea N. Edginton; Nikoletta Fotaki (52-71).
Compound solubility serves as a surrogate indicator of oral biopharmaceutical performance. Between infancy and adulthood, marked compositional changes in gastrointestinal (GI) fluids occur. This study serves to assess how developmental changes in GI fluid composition affects compound solubility.Solubility assessments were conducted in vitro using biorelevant media reflective of age-specific pediatric cohorts (i.e., neonates and infants). Previously published adult media (i.e., FaSSGF, FeSSGF, FaSSIF.v2, and FeSSIF.v2) were employed as references for pediatric media development. Investigations assessing age-specific changes in GI fluid parameters (i.e., pepsin, bile acids, pH, osmolality, etc.) were collected from the literature and served to define the composition of neonatal and infant media. Solubility assessments at 37°C were conducted for seven BCS Class II compounds within the developed pediatric and reference adult media.For six of the seven compounds investigated, solubility fell outside an 80–125% range from adult values in at least one of the developed pediatric media. This result indicates a potential for age-related alterations in oral drug performance, especially for compounds whose absorption is delimited by solubility (i.e., BCS Class II).Developmental changes in GI fluid composition can result in relevant discrepancies in luminal compound solubility between children and adults.
Keywords: biopharmaceutics; biorelevant; pediatric; solubility

A Role for Low Density Lipoprotein Receptor-Related Protein 1 in the Cellular Uptake of Tissue Plasminogen Activator in the Lungs by Swan Lin; Jennifer Racz; Melissa F. Tai; Kristina M. Brooks; Phillip Rzeczycki; Lauren J. Heath; Michael W. Newstead; Theodore J. Standiford; Gus R. Rosania; Kathleen A. Stringer (72-82).
To gain knowledge of lung clearance mechanisms of inhaled tissue plasminogen activator (tPA).Using an in vivo mouse model and ex vivo murine whole organ cell suspensions, we examined the capability of the lungs to utilize LRP1 receptor-mediated endocytosis (RME) for the uptake of exogenous tPA with and without an LRP1 inhibitor, receptor associated protein (RAP), and quantitatively compared it to the liver. We also used a novel imaging technique to assess the amount LRP1 in sections of mouse liver and lung.Following intratracheal administration, tPA concentrations in the bronchoalveolar lavage fluid (BALF) declined over time following two-compartment pharmacokinetics suggestive of a RME clearance mechanism. Ex vivo studies showed that lung and liver cells are similarly capable of tPA uptake via LRP1 RME which was reduced by ~50% by RAP. The comparable lung and liver uptake of tPA is likely due to equivalent amounts of LRP1 of which there was an abundance in the alveolar epithelium.Our findings indicate that LRP1 RME is a candidate clearance mechanism for inhaled tPA which has implications for the development of safe and effective dosing regimens of inhaled tPA for the treatment of plastic bronchitis and other fibrin-inflammatory airway diseases in which inhaled tPA may have utility.
Keywords: plastic bronchitis; receptor mediated endocytosis; therapeutic proteins

In Vitro and In Vivo Performance of a Pre-Filled, Electrochemically-Actuated Infusion System by Assaf Shaked; Ella Tenenbaum-Koren; Jacob Atsmon; Peter Noymer (83-91).
A novel infusion system (PatchPump®) has been developed using an electrochemical actuator and a semi-flexible container to deliver liquid formulations through an infusion set. In vitro studies characterized pump performance, and clinical studies evaluated functionality and pharmacokinetic (PK) performance in humans.Total delivered volume and flow rate accuracy were determined in vitro by real-time gravimetric analysis. Functionality was evaluated in vivo using prototype devices delivering saline for 18 h in a clinical study enrolling 10 healthy volunteers. A second clinical study examined the PK of delivering treprostinil at an average dose of 2.3 ng/kg/min to 5 healthy volunteers for 18 h.Relative to a design target of 0.042 mL/hr, the average flow rate of 23 PatchPumps operating for 48 continuous hours was 0.043 ± 0.007 mL/hr as tested in vitro. In vivo functionality was confirmed by complete infusion of saline for the entire duration with only mild and transient adverse effects. PK results with an infusion rate of 0.045 mL/hr of treprostinil resulted in mean Css of 297 pg/mL and T1/2 of 4.44 h, comparable to prior studies using conventional infusion pumps.The results of these studies demonstrate a successful proof-of-concept for the PatchPump technology.
Keywords: parenteral; pharmacokinetic; pump; treprostinil; wearable

Biological Effects of Simvastatin Formulated as pMDI on Pulmonary Epithelial Cells by Alaa S. Tulbah; Hui Xin Ong; Wing-Hin Lee; Paolo Colombo; Paul M. Young; Daniela Traini (92-101).
The aim of this study is to evaluate the biological effects of Calu-3 epithelial cells in response to the delivery of simvastatin (SV) via solution pressurized metered dose inhaler (pMDI).SV pMDI was aerosolised onto Calu-3 air-interface epithelial cells using a modified glass twin stage impinger. The transport of SV across Calu-3 cells, mucus production, inflammatory cytokines production i.e., interleukin (IL) 6, 8 and tumour necrosis factor alpha (TNF- α) and oxidative stress from Calu-3 cells following treatment with SV pMDI was investigated and compared to untreated cells.It was found that SV had the ability to penetrate into the respiratory epithelium and convert into its active SV hydroxy acid (SVA) metabolite. Furthermore, the amount of mucus produced was significantly reduced when SV was deposited on Calu-3 compared to untreated cells. Additionally, SV delivered by pMDI reduces production of IL-6, 8 and TNF-α from Calu-3 following stimulation with lipopolysaccharide (LPS). SV also showed equivalent antioxidant property to vitamin E.Treatment with SV solution pMDI formulation on Calu-3 cells reduces mucus production, inflammatory cytokines and oxidative stress. This formulation could potentially be used clinically as muco-inhibitory and anti-inflammatory therapy for treatment of chronic lung diseases.
Keywords: anti-inflammatory; Calu-3 epithelial cell; mucus; pMDI; simvastatin

Development of a Sustained Release Solid Dispersion Using Swellable Polymer by Melting Method by Tuong Ngoc-Gia Nguyen; Phuong Ha-Lien Tran; Toi Van Vo; Wei Duan; Thao Truong-Dinh Tran (102-109).
This study is to design a sustained release solid dispersion using swellable polymer by melting method.Polyethylene glycol 6000 (PEG 6000) and hydroxypropyl methylcellulose 4000 (HPMC 4000) were used in solid dispersion for not only enhancing drug dissolution rate but also sustaining drug release. HPMC 4000 is a common swellable polymer in matrix sustained release dosage form, but could not be used in preparation of solid dispersion by melting method. However, the current study utilized the swelling capability of HPMC 4000 accompanied by the common carrier PEG 6000 in solid dispersion to accomplish the goal.While PEG 6000 acted as a releasing stimulant carrier and provided an environment to facilitate the swelling of HPMC 4000, this swellable polymer could act as a rate-controlling agent. This greatly assisted the dissolution enhancement by changing the crystalline structure of drug to more amorphous form and creating a molecular interaction.These results suggested that this useful technique can be applied in designing a sustained release solid dispersion with many advantages.
Keywords: poorly water-soluble drug; solid dispersion; sustained release; swellable polymer

Development of Disposal Systems for Deactivation of Unused/Residual/Expired Medications by Anushree Herwadkar; Neha Singh; Carter Anderson; Andrew Korey; William Fowler; Ajay K. Banga (110-124).
The objective of this work was to identify deactivation agents and develop a disposal system for unused/ residual/ expired medications.Deactivation agents screened included oxidizing agent-sodium percarbonate, hydrolysis agent- sodium carbonate and adsorbants- zeolite and activated carbon. Deactivation studies using these agents were performed on four active pharmaceutical agents (APIs) including ketoprofen, dexamethasone sodium phosphate, metformin hydrochloride and amoxicillin trihydrate. Disposal systems were also designed for deactivation studies on dexamethasone pills, amoxicillin trihydrate capsules and fentanyl transdermal patches (Duragesic®). Briefly, APIs/ dosage forms were allowed to be in close contact with deactivation agents for a specified period of time and percentage decrease in the amount of API from the initial amount was measured.Sodium percarbonate and sodium carbonate were only successful in deactivation of amoxicillin trihydrate API. Adsorption agents resulted in more universal deactivation with activated carbon resulting in efficient deactivation of most APIs and all dosage forms tested. Also adsorption of oral dosage medications on activated carbons was maintained even on dilution and shaking and no desorption was observed.Deactivation systems containing activated carbon are promising for efficient, safe and environment friendly disposal of unused/residual/expired medications.
Keywords: deactivation; disposal systems; unused medications

Sustained release of small molecule hydrophilic drugs at high doses remains difficult to achieve from electrospun fibers and limits their use in clinical applications. Here we investigate tunable release of several water-soluble anti-HIV drugs from electrospun fibers fabricated with blends of two biodegradable polyesters.Drug-loaded fibers were fabricated by electrospinning ratios of PCL and PLGA. Fiber morphology was imaged by SEM, and DSC was used to measure thermal properties. HPLC was used to measure drug loading and release from fibers. Cytotoxicity and antiviral activity of drug-loaded fibers were measured in an in vitro cell culture assay.We show programmable release of hydrophilic antiretroviral drugs loaded up to 40 wt%. Incremental tuning of highly-loaded drug fibers within 24 h or >30 days was achieved by controlling the ratio of PCL and PLGA. Fiber compositions containing higher PCL content yielded greater burst release whereas fibers with higher PLGA content resulted in greater sustained release kinetics. We also demonstrated that our drug-loaded fibers are safe and can sustain inhibition of HIV in vitro.These data suggest that we were able to overcome current limitations associated with sustained release of small molecule hydrophilic drugs at clinically relevant doses. We expect that our system represents an effective strategy to sustain delivery of water-soluble molecules that will benefit a variety of biomedical applications.
Keywords: electrospinning; high loading; HIV; programmable release; tenofovir

Novel Gefitinib Formulation with Improved Oral Bioavailability in Treatment of A431 Skin Carcinoma by Chandraiah Godugu; Ravi Doddapaneni; Apurva R Patel; Rakesh Singh; Roger Mercer; Mandip Singh (137-154).
Oral administration of anticancer agents presents a series of advantages for patients. However, most of the anticancer drugs have poor water solubility leading to low bioavailability.Controlled released spray dried matrix system of Gefitinib with hydroxypropyl β-cyclodextrin, chitosan, hydroxy propyl methyl cellulose, vitamin E TPGS, succinic acid were used for the design of formulations to improve the oral absorption of Gefitinib. Spray drying with a customized spray gun which allows simultaneous/pulsatile flow of two different liquid systems through single nozzle was used to prepare Gefitinib spray dried formulations (Gef-SD). Formulation was characterized by in vitro drug release and Caco-2 permeability studies. Pharmacokinetic studies were performed in Sprague Dawley rats. Efficacy of Gef-SD was carried out in A431 xenografts models in nude mice.In Gef-SD group 9.14-fold increase in the AUC was observed compared to free Gef. Improved pharmacokinetic profile of Gef-SD translated into increase (1.75 fold compared to Gef free drug) in anticancer effects. Animal survival was significantly increased in Gef formulation treated groups, with superior reduction in the tumor size (1.48-fold) and volumes (1.75-fold) and also increase in the anticancer effects (TUNEL positive apoptotic cells) was observed in Gef-SD treated groups. Further, western blot, immunohistochemical and proteomics analysis demonstrated the increased pharmacodynamic effects of Gef-SD formulations in A431 xenograft tumor models.Our studies suggested that Gefitinib can be successfully incorporated into control release microparticles based oral formulation with enhanced pharmacokinetic and pharmacodynamic activity. This study demonstrates the novel application of Gef in A431 tumor models.
Keywords: A431; control release formulations; epidermal growth factor receptor; gefitinib; spray drying

Increased solution viscosity results in difficulties in manufacturing and delivery of therapeutic protein formulations, increasing both the time and production costs, and leading to patient inconvenience. The solution viscosity is affected by the molecular properties of both the solute and the solvent. The purpose of this work was to investigate the effect of size, charge and protein-protein interactions on the viscosity of Dual Variable Domain Immunoglobulin (DVD-IgTM) protein solutions.The effect of size of the protein molecule on solution viscosity was investigated by measuring intrinsic viscosity and excluded volume calculations for monoclonal antibody (mAb) and DVD-IgTM protein solutions. The role of the electrostatic charge resulting in electroviscous effects for DVD-IgTM protein was assessed by measuring zeta potential. Light scattering measurements were performed to detect protein-protein interactions affecting solution viscosity.DVD-IgTM protein exhibited significantly higher viscosity compared to mAb. Intrinsic viscosity and excluded volume calculations indicated that the size of the molecule affects viscosity significantly at higher concentrations, while the effect was minimal at intermediate concentrations. Electroviscous contribution to the viscosity of DVD-IgTM protein varied depending on the presence or absence of ions in the solution. In buffered solutions, negative k D and B 2 values indicated the presence of attractive interactions which resulted in high viscosity for DVD-IgTM protein at certain pH and ionic strength conditions.Results show that more than one factor contributes to the increased viscosity of DVD-IgTM protein and interplay of these factors modulates the overall viscosity behavior of the solution, especially at higher concentrations.
Keywords: bispecific antibody; charge; excluded volume; hydration; monoclonal antibody; protein formulation

The Effect of Excipients on the Permeability of BCS Class III Compounds and Implications for Biowaivers by Alan Parr; Ismael J. Hidalgo; Chris Bode; William Brown; Mehran Yazdanian; Mario A. Gonzalez; Kazuko Sagawa; Kevin Miller; Wenlei Jiang; Erika S. Stippler (167-176).
Currently, the FDA allows biowaivers for Class I (high solubility and high permeability) and Class III (high solubility and low permeability) compounds of the Biopharmaceutics Classification System (BCS). Scientific evidence should be provided to support biowaivers for BCS Class I and Class III (high solubility and low permeability) compounds.Data on the effects of excipients on drug permeability are needed to demonstrate that commonly used excipients do not affect the permeability of BCS Class III compounds, which would support the application of biowaivers to Class III compounds. This study was designed to generate such data by assessing the permeability of four BCS Class III compounds and one Class I compound in the presence and absence of five commonly used excipients.The permeability of each of the compounds was assessed, at three to five concentrations, with each excipient in two different models: Caco-2 cell monolayers, and in situ rat intestinal perfusion. No substantial increases in the permeability of any of the compounds were observed in the presence of any of the tested excipients in either of the models, with the exception of disruption of Caco-2 cell monolayer integrity by sodium lauryl sulfate at 0.1 mg/ml and higher.The results suggest that the absorption of these four BCS Class III compounds would not be greatly affected by the tested excipients. This may have implications in supporting biowaivers for BCS Class III compounds in general.
Keywords: BCS class III; bioavailability; Caco-2; permeability; rat intestinal perfusion model

In vivo Functional Evaluation of Increased Brain Delivery of the Opioid Peptide DAMGO by Glutathione-PEGylated Liposomes by Annika Lindqvist; Jaap Rip; Joan van Kregten; Pieter J Gaillard; Margareta Hammarlund-Udenaes (177-185).
The purpose of this study was to evaluate formulation factors causing improvement in brain delivery of a small peptide after encapsulation into a targeted nanocarrier in vivo.The evaluation was performed in rats using microdialysis, which enabled continuous sampling of the released drug in both the brain (striatum) and blood. Uptake in brain could thereby be studied in terms of therapeutically active, released drug.We found that encapsulation of the peptide DAMGO in fast-releasing polyethylene glycol (PEG)ylated liposomes, either with or without the specific brain targeting ligand glutathione (GSH), doubled the uptake of DAMGO into the rat brain. The increased brain delivery was observed only when the drug was encapsulated into the liposomes, thus excluding any effects of the liposomes themselves on the blood–brain barrier integrity as a possible mechanism. The addition of a GSH coating on the liposomes did not result in an additional increase in DAMGO concentrations in the brain, in contrast to earlier studies on GSH coating. This may be caused by differences in the characteristics of the encapsulated compounds and the composition of the liposome formulations.We were able to show that encapsulation into PEGylated liposomes of a peptide with limited brain delivery could double the drug uptake into the brain without using a specific brain targeting ligand.
Keywords: blood–brain barrier; brain delivery; liposomes; microdialysis; opioid peptide

The Evaluation of the Therapeutic Efficacy and Side Effects of a Macromolecular Dexamethasone Prodrug in the Collagen-Induced Arthritis Mouse Model by Lingdong Quan; Yijia Zhang; Anand Dusad; Ke Ren; P. Edward Purdue; Steven R. Goldring; Dong Wang (186-193).
To investigate the efficacy and safety of N-(2-hydroxypropyl) methacrylamide (HPMA) copolymer-dexamethasone conjugate (P-Dex) in the collagen-induced arthritis (CIA) mouse model.HPMA copolymer labeled with a near infrared fluorescence (NIRF) dye was administered to mice with CIA to validate its passive targeting to inflamed joints and utility as a drug carrier system. The CIA mice were treated with P-Dex, dexamethasone (Dex) or saline and the therapeutic efficacy and skeletal toxicity evaluated using clinical scoring and micro-computed tomography (μ-CT).The NIRF signal of the HPMA copolymer localized to arthritic joints consistent with its passive targeting to sites of inflammation. While the CIA mice responded more rapidly to P-Dex compared to Dex, the final clinical score and endpoint μ-CT analyses of localized bone erosions indicated that both single dose P-Dex and dose equivalent daily Dex led to comparable clinical efficacy after 30 days. μ-CT analysis of the proximal tibial metaphyses showed that P-Dex treatment was associated with significantly higher BMD and BV/TV compared to Dex and the saline control, consistent with reduced glucocorticoid (GC) skeletal toxicity.These results validate the therapeutic efficacy of P-Dex in the CIA mouse model. P-Dex treatment averted the adverse effects of GC’s on systemic bone loss, supporting its utility in clinical development for the management of rheumatoid arthritis.
Keywords: collagen-induced arthritis; ELVIS; glucocorticoid; HPMA copolymer; prodrug

Thermodynamic Proxies to Compensate for Biases in Drug Discovery Methods by Sean Ekins; Nadia K. Litterman; Christopher A. Lipinski; Barry A. Bunin (194-205).
We propose a framework with simple proxies to dissect the relative energy contributions responsible for standard drug discovery binding activity.We explore a rule of thumb using hydrogen-bond donors, hydrogen-bond acceptors and rotatable bonds as relative proxies for the thermodynamic terms. We apply this methodology to several datasets (e.g., multiple small molecules profiled against kinases, Mycobacterium tuberculosis (Mtb) high throughput screening (HTS) and structure based drug design (SBDD) derived compounds, and FDA approved drugs).We found that Mtb active compounds developed through SBDD methods had statistically significantly larger PEnthalpy values than HTS derived compounds, suggesting these compounds had relatively more hydrogen bond donor and hydrogen bond acceptors compared to rotatable bonds. In recent FDA approved medicines we found that compounds identified via target-based approaches had a more balanced enthalpic relationship between these descriptors compared to compounds identified via phenotypic screensAs it is common to experimentally optimize directly for total binding energy, these computational methods provide alternative calculations and approaches useful for compound optimization alongside other common metrics in available software and databases.
Keywords: enthalpy; entropy; high-throughput screening; structure based drug design; tuberculosis

Ceramide and N,N,N-Trimethylphytosphingosine-Iodide (TMP-I)-Based Lipid Nanoparticles for Cancer Therapy by Prabagar Balakrishnan; Chung Kil Song; Alexander Jahn; Hyun-Jong Cho (206-216).
To evaluate the anti-tumor effect of ceramide or trimethylphytosphingosine-iodide (TMP-I) containing solid lipid nanoparticles (SLNs) prepared using trymyristin, phosphatidylcholine (PC), and Pluronic P85 (P85) for intravenous delivery of docetaxel.Docetaxel-loaded SLNs using ceramide or TMP-I at 3.22% (w/w) with a mean diameter of 89–137 nm were successfully prepared by high pressure homogenization. The prepared nanoparticles were characterized by particle size, zeta potential, drug content, and TEM analysis. Cellular uptake and cytotoxicity were studied using adriamycin-resistant breast cancer (MCF-7/ADR) cells. The optimized formulation’s dissolution profile, pharmacokinetics, and antitumor effect in mice tumor model were compared with that of control (Taxotere®).The drug release rate of docetaxel from SLNs was lower than that of control (Taxotere®). The prepared SLNs showed higher cellular uptake of docetaxel compared to that of Taxotere® in MCF-7/ADR cell lines, which was further confirmed by the confocal laser scanning microscopy (CLSM) study using coumarin 6 (C6). Prepared SLNs exhibited significantly increased antitumor efficacy, compared to Taxotere®, in MCF-7/ADR cells. In vivo pharmacokinetic study in rats (at 10 mg/kg dose) showed that the SLNs significantly reduced in vivo clearance of drug than Taxotere®. Interestingly, ceramide and TMP-I SLNs efficiently inhibited the tumor growth compared to Taxotere® in MCF-7/ADR tumor xenografted mouse model.This work showed that TMP-I and ceramide SLNs not only significantly enhanced systemic exposure of drug, but also increased antitumor efficacy compared to Taxotere® and control SLN.
Keywords: ceramide; enhanced antitumor activity; multidrug resistance; reduced in vivo clearance; solid lipid nanoparticles; trimethylphytosphingosine-iodide

A Promising Emodin-Loaded Poly (Lactic-Co-Glycolic Acid)-d-α-Tocopheryl Polyethylene Glycol 1000 Succinate Nanoparticles for Liver Cancer Therapy by Hongyan Liu; Meng Gao; Hong Xu; Xin Guan; Li Lv; Sa Deng; Chenghong Zhang; Yan Tian (217-236).
Emodin (EMO) has multi-targets and multi-way antitumor effect, which was limited by the instability and poor solubility of EMO. The aim of this study was to formulate EMO-loaded poly (lactide-co-glycolide)-d-α-tocopheryl polyethylene glycol 1000 succinate (PLGA-TPGS) nanoparticles (EPTN) to increase the liver targeting of EMO for cancer therapy.EMO/coumarin-6-loaded PLGA-TPGS nanoparticles (ECPTN) and EMO-loaded PLGA nanoparticles (EPN) were also prepared as comparison. The cellular uptake of ECPTN by HepG2 and HCa-F cells was investigated using Confocal laser scanning microscopy. The apoptosis of HepG2 cells handled with EPTN was assayed by flow cytometry. The liver targeting property of ECPTN in mice was evaluated using the drug concentration determined by RP-HPLC and the freezing slices were investigated via fluorescence inversion microscopy. The blood samples were obtained from vein intubation to illustrate the pharmacokinetics process of EPTN. The tumor-bearing mice model was established to elucidate the in vivo therapeutic effect of EPTN.The results demonstrated that ECPTN could be internalized by HepG2 and HCa-F cells respectively. The ratio of apoptosis cells was increased after dealing with EPTN. The detection indexes of drug concentration and fluorescence inversion microscopy images indicated ECPTN had an excellent effect on liver targeting property than EMO solutions (EMS). The pharmacokinetics process of EPTN showed obvious sustained-release effect than EMS. Compared with EPN, the in vivo antitumor activity of EPTN against tumor cells were better.In conclusion, EPTN could be used in the treatment of liver cancer acted as a kind of promising intravenous dosage forms.
Keywords: Emodin; liver cancer; liver targeting property; nanoparticles; PLGA-TPGS

To investigate the destruction of clinically-relevant bacteria within biofilms via the sustained release of the antibiotic tetracycline from zein-based electrospun polymeric fibrous matrices and to demonstrate the compatibility of such wound dressing matrices with human skin cells.Zein/PCL triple layered fibrous dressings with entrapped tetracycline were electrospun. The successful entrapment of tetracycline in these dressings was validated. The successful release of bioactive tetracycline, the destruction of preformed biofilms, and the viability of fibroblast (FEK4) cells were investigated.The sustained release of tetracycline from these matrices led to the efficient destruction of preformed biofilms from Staphylococcus aureus MRSA252 in vitro, and of MRSA252 and ATCC 25923 bacteria in an ex vivo pig skin model using 1 × 1 cm square matrices containing tetracycline (30 μg). Human FEK4 cells grew normally in the presence of these matrices.The ability of the zein-based matrices to destroy bacteria within increasingly complex in vitro biofilm models was clearly established. An ex vivo pig skin assay showed that these matrices, with entrapped tetracycline, efficiently kill bacteria and this, combined with their compatibility with a human skin cell line suggest these matrices are well suited for applications in wound healing and infection control.
Keywords: biofilm; controlled drug delivery; electrospinning; nanofibre; pig skin

Increased Insulin Secretion from Insulin-Secreting Cells by Construction of Mixed Multicellular Spheroids by Kosuke Kusamori; Makiya Nishikawa; Narumi Mizuno; Tomoko Nishikawa; Akira Masuzawa; Yutaro Tanaka; Yuya Mizukami; Kazunori Shimizu; Satoshi Konishi; Yuki Takahashi; Yoshinobu Takakura (247-256).
We previously have shown that multicellular spheroids containing insulin-secreting cells are an effective therapy for diabetic mice. Here we attempted to increase insulin secretion by incorporating other cell types into spheroids.Multicellular spheroids of mouse MIN6 pancreatic β cells were formed in microwells alone and with aortic vascular endothelial MAEC cells or embryo fibroblast NIH3T3 cells. mRNA expression of insulin genes and insulin secretion of MIN6 cells in each spheroid were measured by real-time PCR and an insulin ELIZA kit. Moreover, collagen IV expression in each spheroid was analyzed by western blot.In all cases, uniformly sized (about 300 μm) multicellular spheroids were obtained. MAEC or NIH3T3 cell incorporation into MIN6 spheroids significantly increased mRNA expression of insulin genes and insulin secretion. In addition, collagen IV expression, which was reported to enhance insulin secretion from pancreatic β cells, also increased in their spheroids.The formation of mixed multicellular spheroids containing collagen IV-expressing cells can improve the insulin secretion from insulin-secreting MIN6 cells, and mixed multicellular spheroids can be a potent therapeutic option for patients with type I diabetes mellitus.
Keywords: diabetes; extracellular matrix; insulin; spheroid; β cells

AAPS Connection (257-259).