Pharmaceutical Research (v.28, #2)

Nanomedicine for Cancer Therapy by Mansoor M. Amiji (181-186).
RPh, Ph.D., received his undergraduate degree in pharmacy from Northeastern University in 1988 and his Ph.D. in pharmaceutics from Purdue University in 1992. His areas of specialization include polymeric biomaterials, advanced drug delivery systems, and nanomedical technologies.Dr. Amiji’s research interests include synthesis of novel polymeric materials for medical and pharmaceutical applications; surface modification of cationic polymers by the complexation-interpenetration method to develop biocompatible materials; preparation and characterization of polymeric membranes and microcapsules with controlled permeability properties for medical and pharmaceutical applications; target-specific drug and vaccine delivery systems for gastrointestinal tract infections; localized delivery of cytotoxic and anti-angiogenic drugs for solid tumors in novel biodegradable polymeric nanoparticles; intracellular delivery systems for drugs and genes using target-specific, long-circulating, biodegradable polymeric nanoparticles; gold and iron-gold core-shell nanoparticles for biosensing, imaging, and delivery applications. His research has received sustained funding from the National Institutes of Health (NIH), National Science Foundation (NSF), other foundations, and local industries.Dr. Amiji is currently Distinguished Professor and Chairman of the Pharmaceutical Sciences Department and Co-director of Northeastern University Nanomedicine Education and Research Consortium (NERC). NERC oversees a doctoral training grant in Nanomedicine Science and Technology that was co-funded by the NIH and NSF. He has three published books: Applied Physical Pharmacy (McGraw-Hill, 2003), Polymeric Gene Delivery: Principles and Applications (CRC Press, 2005), and Nanotechnology for Cancer Therapy (CRC Press, 2007), along with over 150 book chapters, peer-reviewed manuscripts, and abstract publications. Dr. Amiji has received a number of awards, including the 2006 NSTI Award for Outstanding Contributions towards the Advancement of Nanotechnology, Microtechnology, and Biotechnology, and the 2007 American Association of Pharmaceutical Scientist’s Meritorious Manuscript Award.Dr. Amiji has supervised research efforts of over 80 post-doctoral associates, doctoral and master’s level graduate students, and undergraduate honors students over the last 17 years. His teaching responsibilities are in Doctor of Pharmacy (Pharm.D.) program and graduate programs (M.S. and Ph.D.) in Pharmaceutical Sciences, Biotechnology, and Nanomedicine.

Clinical Developments in Nanotechnology for Cancer Therapy by Jeremy D. Heidel; Mark E. Davis (187-199).
Nanoparticle approaches to drug delivery for cancer offer exciting and potentially “game-changing” ways to improve patient care and quality of life in numerous ways, such as reducing off-target toxicities by more selectively directing drug molecules to intracellular targets of cancer cells. Here, we focus on technologies being investigated clinically and discuss numerous types of therapeutic molecules that have been incorporated within nanostructured entities such as nanoparticles. The impacts of nanostructured therapeutics on efficacy and safety, including parameters like pharmacokinetics and biodistribution, are described for several drug molecules. Additionally, we discuss recent advances in the understanding of ligand-based targeting of nanoparticles, such as on receptor avidity and selectivity.
Keywords: clinical; nanoparticle; oncology; pharmacokinetics; targeting

Cancer Immunotherapy and Nanomedicine by Wei-Yun Sheng; Leaf Huang (200-214).
The immune system has the ability to recognize and kill pre-cancer and cancer cells. However, with the immune system’s surveillance, the survival tumor cells learn how to escape the immune system after immunoselection. Cancer immunotherapy develops strategies to overcome these problems. Nanomedicine applications in cancer immunotherapy include the nanodiagnostics and nanobiopharmaceuticals. In cancer nanodiagnostics, it looks for specific “molecular signatures” in cancer cells or their microenvironment by using genomics and proteomics. Nanobiopharmaceuticals is the field that studies nanotechnology-based therapeutic agents and drug carriers. DNA, RNA, peptides, proteins and small molecules can all be used as cancer therapies when formulated in nanocarriers. Currently, cancer vaccines are applied in treatments with existing cancer or to prevent the development of cancer in certain high risk individuals. Most of the non-specific immune activation agents include adjuvants which enhance immunogenicity and accelerate and prolong the response of cancer vaccines. The carriers of vaccines, such as viruses and nanoparticles, have also been in clinical studies for many years. This review will discuss the relationships between the tumor and the immune system, and also will include topics covering the strategies used in eliminating tumors by using nanomedicine.
Keywords: adjuvant; cancer immunotherapy; cancer vaccine; nano; tumor immunoediting

Nanoparticle Delivery Systems in Cancer Vaccines by Yogita Krishnamachari; Sean M. Geary; Caitlin D. Lemke; Aliasger K. Salem (215-236).
Therapeutic strategies that involve the manipulation of the host’s immune system are gaining momentum in cancer research. Antigen-loaded nanocarriers are capable of being actively taken up by antigen-presenting cells (APCs) and have shown promising potential in cancer immunotherapy by initiating a strong immunostimulatory cascade that results in potent antigen-specific immune responses against the cancer. Such carrier systems offer versatility in that they can simultaneously co-deliver adjuvants with the antigens to enhance APC activation and maturation. Furthermore, modifying the surface properties of these nanocarriers affords active targeting properties to APCs and/or enhanced accumulation in solid tumors. Here, we review some recent advances in these colloidal and particulate nanoscale systems designed for cancer immunotherapy and the potential for these systems to translate into clinical cancer vaccines.
Keywords: cancer immunotherapy; colloidal nanocarriers; liposomes; polymeric nanoparticles; tumor targeting

Inorganic Nanoparticles in Cancer Therapy by Sanjib Bhattacharyya; Rachel A. Kudgus; Resham Bhattacharya; Priyabrata Mukherjee (237-259).
Nanotechnology is an evolving field with enormous potential for biomedical applications. The growing interest to use inorganic nanoparticles in medicine is due to the unique size- and shape-dependent optoelectronic properties. Herein, we will focus on gold, silver and platinum nanoparticles, discussing recent developments for therapeutic applications with regard to cancer in terms of nanoparticles being used as a delivery vehicle as well as therapeutic agents. We will also discuss some of the key challenges to be addressed in future studies.
Keywords: cancer; drug delivery; gold; inorganic; nanoparticle; therapeutics

Nanoparticles: A Promising Modality in the Treatment of Sarcomas by Michiro Susa; Lara Milane; Mansoor M. Amiji; Francis J. Hornicek; Zhenfeng Duan (260-272).
Improvements in surgical technique, chemotherapy, and radiotherapy have enhanced the prognosis of sarcoma patients, but have since reached a plateau in recent years. Novel approaches have been sought but with limited results. Nanomedicine offers solutions in diverse areas of sarcoma therapy including diagnosis and treatment. Several varieties of nanoparticles, including multifunctional nanoparticles, are available that localize the biodistribution of conventional chemotherapeutics to the tumor site. Also, nanoparticles loaded with chemotherapeutic drugs have the ability to overcome drug resistance which is a major obstacle impeding the progress of the treatment. Multifunctional nanoparticles, which have the potential to further augment the bioavailability of drugs, are being actively investigated. In this review, we will discuss the application of nanoparticles for improving the treatment of sarcoma patients.
Keywords: nanoparticle; sarcoma; tyrosine kinase; drug resistance; cancer-initiating cell

The new generation of nanotechnology-based drug formulations is challenging the accepted ways of cancer treatment. Multi-functional nanomaterial constructs have the capability to be delivered directly to the tumor site and eradicate cancer cells selectively, while sparing healthy cells. Tailoring of the nano-construct design can result in enhanced drug efficacy at lower doses as compared to free drug treatment, wider therapeutic window, and lower side effects. Nanoparticle carriers can also address several drug delivery problems which could not be effectively solved in the past and include reduction of multi-drug resistance effects, delivery of siRNA, and penetration of the blood-brain-barrier. Although challenges in understanding toxicity, biodistribution, and paving an effective regulatory path must be met, nanoscale devices carry a formidable promise to change ways cancer is diagnosed and treated. This article summarizes current developments in nanotechnology-based drug delivery and discusses path forward in this field. The discussion is done in context of research and development occurring within the NCI Alliance for Nanotechnology in Cancer program.
Keywords: drug delivery; multi-functional nanoparticle carriers; nanoparticle; toxicity

An Effective Strategy for the Synthesis of Biocompatible Gold Nanoparticles Using Cinnamon Phytochemicals for Phantom CT Imaging and Photoacoustic Detection of Cancerous Cells by Nripen Chanda; Ravi Shukla; Ajit Zambre; Swapna Mekapothula; Rajesh R. Kulkarni; Kavita Katti; Kiran Bhattacharyya; Genevieve M. Fent; Stan W. Casteel; Evan J. Boote; John A. Viator; Anandhi Upendran; Raghuraman Kannan; Kattesh V. Katti (279-291).
The purpose of the present study was to explore the utilization of cinnamon-coated gold nanoparticles (Cin-AuNPs) as CT/optical contrast-enhancement agents for detection of cancer cells.Cin-AuNPs were synthesized by a “green” procedure, and the detailed characterization was performed by physico-chemical analysis. Cytotoxicity and cellular uptake studies were carried out in normal human fibroblast and cancerous (PC-3 and MCF-7) cells, respectively. The efficacy of detecting cancerous cells was monitored using a photoacoustic technique. In vivo biodistribution was studied after IV injection of Cin-AuNPs in mice, and also a CT phantom model was generated.Biocompatible Cin-AuNPs were synthesized with high purity. Significant uptake of these gold nanoparticles was observed in PC-3 and MCF-7 cells. Cin-AuNPs internalized in cancerous cells facilitated detectable photoacoustic signals. In vivo biodistribution in normal mice showed steady accumulation of gold nanoparticles in lungs and rapid clearance from blood. Quantitative analysis of CT values in phantom model revealed that the cinnamon-phytochemical-coated AuNPs have reasonable attenuation efficiency.The results indicate that these non-toxic Cin-AuNPs can serve as excellent CT/ photoacoustic contrast-enhancement agents and may provide a novel approach toward tumor detection through nanopharmaceuticals.
Keywords: cancer cells; cellular internalization; cinnamon-stabilized gold nanoparticles; in vivo biodistribution; phantom CT imaging; photoacoustic detection

Nanochannel Technology for Constant Delivery of Chemotherapeutics: Beyond Metronomic Administration by Alessandro Grattoni; Haifa Shen; Daniel Fine; Arturas Ziemys; Jaskaran S. Gill; Lee Hudson; Sharath Hosali; Randy Goodall; Xuewu Liu; Mauro Ferrari (292-300).
The purpose of this study is to demonstrate the long-term, controlled, zero-order release of low- and high-molecular weight chemotherapeutics through nanochannel membranes by exploiting the molecule-to-surface interactions presented by nanoconfinement.Silicon membranes were produced with nanochannels of 5, 13 and 20 nm using standardized industrial microfabrication techniques. The study of the diffusion kinetics of interferonα-2b and leuprolide was performed by employing UV diffusion chambers. The released amount in the sink reservoir was monitored by UV absorbance.Continuous zero-order release was demonstrated for interferonα-2b and leuprolide at release rates of 20 and 100 μg/day, respectively. The release rates exhibited by these membranes were verified to be in ranges suitable for human therapeutic applications.Our membranes potentially represent a viable nanotechnological approach for the controlled administration of chemotherapeutics intended to improve the therapeutic efficacy of treatment and reduce many of the side effects associated with conventional drug administration.
Keywords: chemotherapy; controlled release; drug delivery; implants; nanochannel

Palmitoyl Ascorbate-Loaded Polymeric Micelles: Cancer Cell Targeting and Cytotoxicity by Rupa R. Sawant; Onkar Vaze; Gerard G. M. D’Souza; Karen Rockwell; Vladimir P. Torchilin (301-308).
To evaluate the potential of palmitoyl ascorbate (PA)-loaded micelles for ascorbate-mediated cancer cell targeting and cytotoxicity.PA was incorporated in polyethylene glycol-phosphatidyl ethanolamine micelles at varying concentrations. The formulations were evaluated for PA content by RP-HPLC. A stable formulation was selected based on size and zeta potential measurements. A co-culture of cancer cells and GFP-expressing non-cancer cells was used to determine the specificity of PA micelle binding. In vitro cytotoxicity of the micellar formulations towards various cancer cell lines was investigated using a cell viability assay. To elucidate the mechanism of action of cell death in vitro, the effect of various H2O2 scavengers and metal chelators on PA-mediated cytotoxicity was studied. The in vivo anti-cancer activity of PA micelles was studied in female Balb/c mice bearing a murine mammary carcinoma (4T1 cells).PA micelles associated preferentially with various cancer cells compared to non-cancer cells in co-culture. PA micelles exhibited anti-cancer activity in cancer cell lines both in vitro and in vivo. The mechanism of cell death was due primarily to generation of reactive oxygen species (ROS).The anti-cancer activity of PA micelles associated with its enhanced cancer cell binding and subsequent generation of ROS.
Keywords: cancer; micelles; nanocarriers; palmitoyl ascorbate; targeting

High Loading of Gentamicin in Bioadhesive PVM/MA Nanostructured Microparticles Using Compressed Carbon-Dioxide by Elisa Elizondo; Santiago Sala; Edurne Imbuluzqueta; David González; María J. Blanco-Prieto; Carlos Gamazo; Nora Ventosa; Jaume Veciana (309-321).
To investigate, for the first time, the viability of compressed antisolvent methodologies for the preparation of drug-loaded particles of the biodegradable and bioadhesive polymer poly (methyl vinyl ether-co-maleic anhydride) (PVM/MA), utilizing gentamicin (Gm) as a model drug.Precipitation with a Compressed Antisolvent (PCA) method was used for the preparation of PVM/MA particles loaded with gentamicin. Before encapsulation, gentamicin was modified into a hydrophobic complex, GmAOT, by exchanging its sulphate ions with an anionic surfactant. GmAOT:PVM/MA composites were fully characterized in terms of size, morphology, composition, drug distribution, phase composition, in vitro activity and drug release.Homogeneous nanostructured microparticles of PVM/MA loaded with high and uniformly distributed quantities of GmAOT were obtained by PCA. The drug loading factors could be tuned at will, improving up to ten times the loadings obtained by other precipitation techniques. Gentamicin retained its bioactivity after being processed, and, according to its release profiles, after an initial burst it experienced a sustained release over 30 days.Compressed antisolvent methods are suitable technologies for the one-step preparation of highly loaded nanostructured PVM/MA matrices with promising application in the delivery of low bioavailable drugs.
Keywords: Gantrez AN; gentamicin; microparticles; PVM/MA; supercritical fluids

Chemical Inhibitors of the Calcium Entry Channel TRPV6 by Christopher P. Landowski; Katrin A. Bolanz; Yoshiro Suzuki; Matthias A. Hediger (322-330).
Calcium entry channels in the plasma membrane are thought to play a major role in maintaining cellular Ca2+ levels, crucial for growth and survival of normal and cancer cells. The calcium-selective channel TRPV6 is expressed in prostate, breast, and other cancer cells. Its expression coincides with cancer progression, suggesting that it drives cancer cell growth. However, no specific inhibitors for TRPV6 have been identified thus far.To develop specific TRPV6 inhibitors, we synthesized molecules based on the lead compound TH-1177, reported to inhibit calcium entry channels in prostate cancer cells in vitro and in vivo.We found that one of our compounds (#03) selectively inhibited TRPV6 over five times better than TRPV5, whereas TH-1177 and the other synthesized compounds preferentially inhibited TRPV5. The IC50 value for growth inhibition by blocking endogenous Ca2+ entry channels in the LNCaP human prostate cancer cell line was 0.44 ± 0.07 μM compared to TH-1177 (50 ± 0.4 μM).These results suggest that compound #03 is a relatively selective and potent inhibitor for TRPV6 and that it is an interesting lead compound for the treatment of prostate cancer and other cancers of epithelial origin.
Keywords: breast cancer; calcium channel; chemical inhibitors; prostate cancer; TRPV6

Evidence of Deep Percutaneous Penetration Associated with Anti-Inflammatory Activity of Topically Applied Helicteres gardneriana Extract: A Photoacoustic Spectroscopy Study by Juliana Oliveira de Melo; Franciana Pedrochi; Mauro L. Baesso; Luzmarina Hernandes; Maria Conceição T. Truiti; Silmara Baroni; Ciomar A. Bersani-Amado (331-336).
To apply the photoacoustic spectroscopy technique to investigate the penetration rate of topically applied Helicteres gardneriana extract used as anti-inflammatory agent.Experiments were performed ex vivo in a well-controlled group of mice. The crude extract was obtained from leaves of the plant Helicteres gardneriana. Croton oil was applied into the ventral surface of the mouse’s right and left auricles in order to induce an inflammatory response. The left auricle was treated with crude extract, while the right one served as the control. After 6 h, the auricles were sectioned for measurements of edema intensity, myeloperoxidase activity and the formulation penetration rate.Croton oil induced inflammatory response in both auricles. The application of Helicteres gardneriana extract reduced significantly the edema of the auricle and inhibited the activity of the myeloperoxidase enzyme. The photoacoustic data showed that the propagation of the formulation was efficient to reach the deep region of the auricle, crossing the cartilage. The strong anti-inflammatory effect was associated with the observed deep penetration of the formulation.This pre-clinical study showed the anti-inflammatory effect of Helicteres gardneriana extract. The photoacoustic technique was useful to demonstrate that this anti-inflammatory activity was associated with deep percutaneous penetration.
Keywords: anti-inflammatory agent; Helicteres gardneriana extract; percutaneous penetration; photoacoustic spectroscopy

To mimic the physicochemical selectivity of the blood-brain barrier (BBB) and to predict its passive permeability using a PAMPA model based on porcine brain lipid extract (PBLE 10%w/v in alkane).Three PAMPA (BD pre-coated and PBLE with 2 different lipid volumes) models were tested with 108 drugs. Abraham solvation descriptors were used to interpret the in vitro-in vivo correlation with 282 in situ brain perfusion measurements, spanning over 5 orders of magnitude. An in combo PAMPA model was developed from combining measured PAMPA permeability with one H-bond descriptor.The in combo PAMPA predicted 93% of the variance of 197 largely efflux-inhibited in situ permeability training set. The model was cross-validated by the “leave-many-out” procedure, with q2 = 0.92 ± 0.03. The PAMPA models indicated the presence of paramembrane water channels. Only the PBLE-based PAMPA-BBB model with sufficient lipid to fill all the internal pore space of the filter showed a wide dynamic range window, selectivity coefficient near 1, and was suitable for predicting BBB permeability.BBB permeability can be predicted by in combo PAMPA. Its speed and substantially lower cost, compared to in vivo measurements, make it an attractive first-pass screening method for BBB passive permeability.
Keywords: blood-brain barrier; brain permeability-surface area (PS); in combo PAMPA-BBB; P-glycoprotein; rodent in situ brain perfusion

Influence of Temperature on Solvent-Mediated Anhydrate-to-Hydrate Transformation Kinetics by Haiyan Qu; Tommy Munk; Claus Cornett; Jian X. Wu; Johan P. Bøtker; Lars P. Christensen; Jukka Rantanen; Fang Tian (364-373).
To achieve an in-depth understanding of the underlying mechanism of the acceleration or deceleration effect of temperature on solvent-mediated anhydrate-to-hydrate phase transformation.The effect of temperature on the phase transformation rate and onset time of two model compounds was investigated using in situ Raman spectroscopy. The thermodynamic driving force of the phase transformation (e.g. supersaturation) at different temperatures was determined by measuring the solubility of the anhydrate and the hydrate.Both acceleration and deceleration effects of temperature on the phase transformation were observed. The mechanism of these temperature effects was studied by exploring the influence of temperature on supersaturation level and crystallization kinetics. Increasing temperature usually leads to accelerated phase transformation kinetics, but it simultaneously decreases supersaturation, which has the opposite effect on the kinetics of the phase transformation. The overall effect of temperature on the phase transformation is therefore determined by the combined effects of supersaturation and temperature on the nucleation and crystal growth kinetics of the hydrate.By differentiating and comparing the effects of temperature and supersaturation on the anhydrate-to-hydrate phase transformation, a deeper understanding of the underlying principle of the acceleration and deceleration effects of temperature on the phase transformation has been achieved.
Keywords: anhydrate-to-hydrate; crystallization; phase transformation; supersaturation

To effectively inhibit succinate buffer crystallization and the consequent pH changes in frozen solutions.Using differential scanning calorimetry (DSC) and X-ray diffractometry (XRD), the crystallization behavior of succinate buffer in the presence of either (i) a crystallizing (glycine, mannitol, trehalose) or (ii) a non-crystallizing cosolute (sucrose) was evaluated. Aqueous succinate buffer solutions, 50 or 200 mM, at pH values 4.0 or 6.0 were cooled from room temperature to −25°C at 0.5°C/min. The pH of the solution was measured as a function of temperature using a probe designed to function at low temperatures. The final lyophiles prepared from these solutions were characterized using synchrotron radiation.When the succinic acid solution buffered to pH 4.0, in the absence of a cosolute, was cooled, there was a pronounced shift in the freeze-concentrate pH. Glycine and mannitol, which have a tendency to crystallize in frozen solutions, remained amorphous when the initial pH was 6.0. Under this condition, they also inhibited buffer crystallization and prevented pH change. At pH 4.0 (50 mM initial concentration), glycine and mannitol crystallized and did not prevent pH change in frozen solutions. While sucrose, a non-crystallizing cosolute, did not completely prevent buffer crystallization, the extent of crystallization was reduced. Sucrose decomposition, based on XRD peaks attributable to β-D-glucose, was observed in frozen buffer solutions with an initial pH of 4.0. Trehalose completely inhibited crystallization of the buffer components when the initial pH was 6.0 but not at pH 4.0. At the lower pH, the crystallization of both trehalose dihydrate and buffer components was evident.When retained amorphous, sucrose and trehalose effectively inhibited succinate buffer component crystallization and the consequent pH shift. However, when trehalose crystallized or sucrose degraded to yield a crystalline decomposition product, crystallization of buffer was observed. Similarly, glycine and mannitol, two widely used bulking agents, inhibited buffer component crystallization only when retained amorphous. In addition to stabilizing the active pharmaceutical ingredient, lyoprotectants may prevent solution pH shift by inhibiting buffer crystallization.
Keywords: buffer crystallization; cosolute; frozen solution; pH shift

Quality of Original and Biosimilar Epoetin Products by Vera Brinks; Andrea Hawe; Abdul H. H. Basmeleh; Liliana Joachin-Rodriguez; Rob Haselberg; Govert W. Somsen; Wim Jiskoot; Huub Schellekens (386-393).
To compare the quality of therapeutic erythropoietin (EPO) products, including two biosimilars, with respect to content, aggregation, isoform profile and potency.Two original products, Eprex (epoetin alfa) and Dynepo (epoetin delta), and two biosimilar products, Binocrit (epoetin alfa) and Retacrit (epoetin zeta), were compared using (1) high performance size exclusion chromatography, (2) ELISA, (3) SDS-PAGE, (4) capillary zone electrophoresis and (5) in-vivo potency.Tested EPO products differed in content, isoform composition, and potency.Of the tested products, the biosimilars have the same or even better quality as the originals. Especially, the potency of originals may significantly differ from the value on the label.
Keywords: biosimilar; immunogenicity; protein characterization; recombinant human erythropoietin

A Semi-mechanistic Gastric Emptying Model for the Population Pharmacokinetic Analysis of Orally Administered Acetaminophen in Critically Ill Patients by Kayode Ogungbenro; Lakshmi Vasist; Robert Maclaren; George Dukes; Malcolm Young; Leon Aarons (394-404).
To develop a semi-mechanistic population pharmacokinetic model based on gastric emptying function for acetaminophen plasma concentration in critically ill patients tolerant and intolerant to enteral nutrition before and after prokinetic therapy.Acetaminophen plasma concentrations were available from a study with 10 tolerant and 20 intolerant patients before and after prokinetic therapy with either erythromycin or metoclopramide. Population pharmacokinetic modelling was carried out in a nonlinear mixed effects analysis software, NONMEM.A four-compartment semi-mechanistic model for stomach, intestine, central and peripheral compartments was described. The rate of emptying of the stomach was described by a first-order rate parameter. The final model has two gastric emptying rate constant parameters: kg1 (1.30 h−1, RSE = 53.84%, T1/2 = 0.53 h) for the intolerant group before prokinetic therapy and kg2 (27.8 h−1, RSE = 59.35%, T1/2 = 0.025 h) for both the intolerant group after prokinetic therapy and the tolerant group. Other parameters and estimates (RSE) in the model were ka = 5.12 h−1 (28.13%), CL = 13.0 L/h (19.62%), CLD = 22.6 L/h (19.78%), V1 = 63.8 L (12.79%) and V2 = 69 L (38.70%).The four-compartment semi-mechanistic population pharmacokinetic model adequately described the data. The gastric emptying half-time is improved by a factor of about 20 in the patients that are intolerant to enteral nutrition after treatment with prokinetic agents.
Keywords: gastric emptying; mixed effects modelling; paracetamol; population pharmacokinetics; prokinetic agents

Human NPC1L1 Expression is Positively Regulated by PPARα by Yuki Iwayanagi; Tappei Takada; Fumiya Tomura; Yoshihide Yamanashi; Tomohiro Terada; Ken-ichi Inui; Hiroshi Suzuki (405-412).
Niemann-Pick C1-like 1 (NPC1L1), a pharmacological target of ezetimibe, is responsible for cholesterol absorption in enterocytes and hepatocytes. In the present study, the involvement of peroxisome proliferator-activated receptor α (PPARα) and its cofactor, PPARγ coactivator 1α (PGC1α) in the transcriptional regulation of human NPC1L1 was analyzed.Reporter gene assays and electrophoretic mobility shift assays (EMSAs) were performed with the 5′-flanking region of the human NPC1L1 gene and the effect of siPPARα was examined.PPARα-mediated transactivation was observed with human NPC1L1 promoter constructs. Detailed analyses using deletion- and mutated-promoter constructs revealed the presence of a functional PPARα-response element (PPRE) upstream of the human NPC1L1 gene (−846/−834), a direct binding of PPARα and RXRα to which was confirmed by EMSAs. Moreover, PPARα-specific knockdown resulted in a significant decrease in the endogenous expression of NPC1L1 mRNA and protein in human-derived HepG2 cells. Furthermore, cotransfection of PGC1α stimulated the SREBP2/HNF4α- and PPARα/RXRα-mediated activation of the human NPC1L1 promoter.We found that PPARα positively regulates human NPC1L1 transcription via direct binding to a PPRE. Additionally, PGC1α stimulates the SREBP2/HNF4α- and PPARα/RXRα-mediated transactivation of human NPC1L1. These findings may provide new insights into the close relationship of glucose, fatty acids and cholesterol homeostasis.
Keywords: cholesterol; ezetimibe; fenofibrate; fibrates; transporter

Application of Alpha7 Nicotinic Acetylcholine Receptor Agonists in Inflammatory Diseases: An Overview by Mario B. Marrero; Merouane Bencherif; Patrick M. Lippiello; Rudolf Lucas (413-416).
Inflammatory disorders are characterized by the influx of immune cells into the vascular wall of veins and/or arteries in response to stimuli such as oxidized-LDL and various pathogens. These factors stimulate the local production of pro-inflammatory cytokines by macrophages and other cells that promote various inflammatory diseases such as atherosclerosis, Crohn’s, Alzheimer’s and diabetes. Numerous cytokines play a significant role in this process, though tumor necrosis factor (TNF) and various interleukins are thought to be among the most important regulators. These proinflammatory cytokines promote the above-described diseases by inducing endothelial cell dysfunction. In this brief commentary we will discuss some of the latest advances and discoveries in the treatment of these inflammatory diseases, making use of alpha7 nicotinic acetylcholine receptor (alpha7 nAChR) agonists.
Keywords: inflammation; alpha7 nicotinic acetylcholine receptor agonists; cholinergic anti-inflammatory pathway; Jak/STAT pathway

AAPS Connection (417-419).