Nanotopography applications in drug delivery

Refinement of micro- and nanofabrication in the semiconductor field has led to innovations in biomedical technologies. Nanotopography, in particular, shows great potential in facilitating drug delivery. The flexibility of fabrication techniques has created a diverse array of topographies that have been developed for drug delivery applications. Nanowires and nanostraws deliver drug cytosolically for in vitro and ex vivo applications. In vivo drug delivery is limited by the barrier function of the epithelium. Nanowires on microspheres increase adhesion and residence time for oral drug delivery, while also increasing permeability of the epithelium. Low aspect ratio nanocolumns increase paracellular permeability, and in conjunction with microneedles increase transdermal drug delivery of biologics in vivo. In summary, nanotopography is a versatile tool for drug delivery. It can deliver directly to cells or be used for in vivo delivery across epithelial barriers. This editorial highlights the application of nanotopography in the field of drug delivery.     

In Vivo and in Vitro Assessment of Baseline Blood-Brain Barrier Parameters in the Presence of Novel Nanoparticles

Purpose. Nanoparticles have advantage as CNS drug delivery vehicles given they disguise drug permeation limiting characteristics. Conflicting toxicological data, however, is published with regard to blood-brain barrier integrity and gross mortality. Methods. To address this issue two novel nanoparticle types: emulsifying wax/Brij 78and Brij 72/Tween 80 nanoparticles were evaluated in vivo for effect on cerebral perfusion flow, barrier integrity, and permeability using the in situ brain perfusion technique. Additional evaluation was completed in vitro using bovine brain microvessel endothelial cells for effect on integrity, permeability, cationic transport interactions, and tight junction protein expression. Results. In the presence of either nanoparticle formulation, no overall significant differences were observed for cerebral perfusion flow in vivo. Furthermore, observed in vitro and in vivo data showed no statistical changes in barrier integrity, membrane permeability, or facilitated choline transport. Western blot analyses of occludin and claudin-1 confirmed no protein expression changes with incubation of either nanoparticle. Conclusions. The nanoparticle formulations appear to have no effect on primary BBB parameters in established in vitro and in vivo blood-brain barrier models.     

皮肤给药的经皮吸收与测量技术研究进展

经皮给药系统（TDDS）可避免首关效应、胃肠道破坏，为新型皮肤给药系统，可通过控制释放而延长治疗效果，成为药物制剂开发研究的热点之一。但是，药物的理化性质以及皮肤屏障影响药物的经皮吸收。综述了TDDS常用的促渗透技术，包括化学、物理、纳米、天然促渗透技术；介绍了促渗透能力的测定方法，包括体外、离体和体内评估皮肤渗透性的方法。通过对经皮药物递送系统和经皮吸收能力测定方法的归纳与总结，以期为TDDS的合理使用和快速发展提供参考。     

Prediction of Drug Transport Through the Blood-Brain Barrier in Vivo: A Comparison Between Two in Vitro Cell Models

Purpose. Studies were conducted to evaluate whether the use of an in vitro model of the blood-brain barrier (BBB) resulted in more accurate predictions of the in vivo transport of compounds compared to the use of a human intestinal cell line (Caco-2). Methods. The in vitro BBB model employs bovine brain capillary endothelial cells co-cultured with primary rat astrocytes. The Caco-2 cells originate from a human colorectal carcinoma. The rat was used as experimental animal for the in vivo studies. Results. Strong correlations (r = 0.93-0.95) were found between the results generated by the in vitro model of the BBB and two different methodologies to measure the permeability across the BBB in vivo. In contrast, a poor correlation (r = 0.68) was obtained between Caco-2 cell data and in vivo BBB transport. A relatively poor correlation (r = 0.74) was also found between the two in vitro models. Conclusion. The present study illustrates the limitations of the Caco-2 model to predict BBB permeability of compounds in vivo. The results emphasize the fact that the BBB and the intestinal mucosa are two fundamentally different biologic barriers, and to be able to make accurate predictions about the in vivo CNS penetration of potential drug candidates, it is important that the in vitro model possesses the main characteristics of the in vivo BBB.     

Antitumor activity of electrospun polylactide nanofibers loaded with 5-fluorouracil and oxaliplatin against colorectal cancer

The purpose of this study was to evaluate both in vitro and in vivo anticancer activities against colorectal cancer (CRC) of electrospun polylactide (PLA) nanofibers loaded with 5-fluorouracil (5-Flu) and oxaliplatin. For in vitro evaluation, human CRC HCT8 cells were directly exposed to the drug-loaded fiber mats, followed with MTT and flow cytometry (FCM) assay. For in vivo evaluation, the drug-loaded fiber mats were locally implanted into mouse colorectal CT26 tumor-bearing mice, followed with histological analysis and detection of survival rate. The results showed that the drug-loaded fiber mats was similar to that of the combination of free 5-Flu and oxaliplatin in vitro cytotoxicity but was much superior to intravenous injection of free drug in vivo anticancer activities, presenting with suppressed tumor growth rate and prolonged survival time of mice. In conclusion, anticancer activities of 5-Flu and oxaliplatin against CRC can be significantly improved by using PLA electrospun nanofibers as local drug delivery system.     

Enhanced Intracellular Uptake of Sterically Stabilized Liposomal Doxorubicin <Emphasis Type="Italic">in Vitro</Emphasis> Resulting in Improved Antitumor Activity <Emphasis Type="Italic">in Vivo</Emphasis>

Purpose To investigate the correlation between the in vitro intracellular uptake and the in vivo antitumor activity of anticancer drugs delivered by sterically stabilized liposomes (SSL).Methods Arginine-glycine-aspartic acid (RGD) peptide or RGD mimetic (RGDm) was coupled onto the surface of SSL to obtain the cell-binding carrier to facilitate the intracellular delivery of the encapsulated drugs. DOX-loaded SSL (SSL-DOX), DOX-loaded RGD-modified SSL (RGD-SSL-DOX) and DOX-loaded RGDm-modified SSL (RGDm-SSL-DOX) were prepared by lipid film dispersion followed by remote loading of DOX. The intracellular uptake of DOX from the various liposomal formulations was evaluated in vitro with melanoma B16 cells, and the pharmacokinetics, biodistribution, and antitumor activity were compared in C57BL/6 mice carrying melanoma B16 tumors.Results In vitro intracellular uptake of DOX by B16 cells and in vivo antitumor activity in terms of tumor growth inhibition and mice survival time prolongation for various liposomal DOX were in the following order: RGD-SSL-DOX > RGDm-SSL-DOX > SSL-DOX. The mean survival time of the mice treated with RGD-SSL-DOX, RGDm-SSL-DOX, and SSL-DOX was 55, 49, and 44 days, respectively. The three liposomal DOX formulations produced very close DOX accumulation in tumor, which is significantly higher than that of free DOX. RGD- or RGDm-SSL-DOX demonstrated prolonged circulation time similar to that of SSL-DOX, whereas they showed significantly lower DOX level in blood and remarkably higher uptake by spleen than SSL-DOX.Conclusions Enhanced intracellular uptake of DOX encapsulated in SSL could produce an improved therapeutic effect for the melanoma B16 tumors. Enhancing intracellular delivery of the anticancer drugs encapsulated in SSL may be a promising strategy to improve their therapeutic efficacy for solid tumors.     

Assessment of Subconjunctival Delivery with Model Ionic Permeants and Magnetic Resonance Imaging

No HeadingPurpose. The objective was to assess the permeation and clearance of model ionic permeants after subconjunctival injection with nuclear magnetic resonance imaging (MRI).Methods. New Zealand white rabbit was the animal model and manganese ion (Mn²⁺) and manganese ethylenediaminetetraacetic acid complex (MnEDTA^2–) were the model permeants. The current study was divided into three parts: in vitro, postmortem, and in vivo. Transscleral passive permeation experiments were conducted with excised sclera in side-by-side diffusion cells in vitro. Subconjunctival delivery experiments were conducted with rabbits postmortem and in vivo. The distribution and elimination of the probe permeants from the subconjunctival space after subconjunctival injections were determined by MRI.Results. The data of excised sclera in vitro suggest large effective pore size for transscleral transport and negligible pore charge effects upon the permeation of the ionic permeants. The permeability coefficients of Mn²⁺ and MnEDTA^2- across the sclera in vitro were 3.6 × 10^-5 cm/s and 2.4 × 10^-5 cm/s, respectively. Although relatively high sclera permeability was observed in vitro, subconjunctival injections in vivo did not provide significant penetration of Mn²⁺ and MnEDTA^2- into the globe; permeant concentrations in the eye were below the detection limit, which corresponds to less than 0.05% of the concentration of the injection solution (e.g., less than 0.02 mM when 40 mM injection solution was used). The volume of the subconjunctival pocket and the concentration of the permeants in the pocket were observed to decrease with time after the injection, and this could contribute to the lower than expected subconjunctival absorption in vivo. Different from the results in vivo, experiments with rabbits postmortem show significant penetration of Mn²⁺ and MnEDTA^2- into the globe with the permeants primarily delivered into the anterior segment of the eye. This difference suggests blood vasculature clearance as a main barrier for passive transscleral transport. The data also show that the pars plicata/pars plana is the least resistance pathway for passive transscleral drug delivery of the polar permeants, and there are indications of the presence of another barrier, possibly the retinal epithelium and/or Bruchs membrane, at the back of the eye.Conclusions. Subconjunctival delivery of the ionic permeants in vivo cannot be quantitatively predicted by the in vitro results. MRI is a noninvasive complementary technique to traditional pharmacokinetic methods. It can provide insights into ocular pharmacokinetics without permeant redistribution that can occur in surgical procedure postmortem in traditional pharmacokinetic studies when the blood vasculature barrier is absent.     

应用于药物评价的血脑屏障模型研究进展

介绍了几种常见的血脑屏障体内外模型及一种脑渗透性分类方案。其中体外模型主要有溶剂水/分配模型、平行人工膜渗透模型、Transwell细胞模型、微流控芯片血脑屏障模型、永生化内皮细胞系建立的血脑屏障模型、三维血脑屏障模型等。体内模型主要有脑/血浆比率测定法、小鼠脑摄取量分析法、啮齿类动物原位脑灌注法、脑微透析法等。随着血脑屏障模型的不断成熟完善,将有助于筛选中枢神经系统药物,为神经系统疾病的治疗提供更多的理论依据。通过分析和评价各种不同的血脑屏障模型,以期为血脑屏障模型研究及中枢神经系统药物研发提供新的思路。     

In vitro and ex vivo experimental models for evaluation of intranasal systemic drug delivery as well as direct nose-to-brain drug delivery

The intranasal route of administration provides a noninvasive method to deliver drugs into the systemic circulation and/or directly into the brain. Direct nose-to-brain drug delivery offers the possibility to treat central nervous system diseases more effectively, as it can evade the blood–brain barrier. In vitro and ex vivo intranasal models provide a means to investigate physiological and pharmaceutical factors that could play a role in drug delivery across the nasal epithelium as well as to determine the mechanisms involved in drug absorption from the nose. The development and implementation of cost-effective pharmacokinetic models for intranasal drug delivery with good in vitro-in vivo correlation can accelerate pharmaceutical drug product development and improve economic and ecological aspects by reducing the time and costs spent on animal studies. Special considerations should be made with regard to the purpose of the in vitro/ex vivo study, namely, whether it is intended to predict systemic or brain delivery, source and site of tissue or cell sampling, viability window of selected model, and the experimental setup of diffusion chambers. The type of model implemented should suit the relevant needs and requirements of the project, researcher, and interlaboratory. This review aims to provide an overview of in vitro and ex vivo models that have been developed to study intranasal and direct nose-to-brain drug delivery.     

Transferrin receptor-targeted vitamin E TPGS micelles for brain cancer therapy: preparation,characterization and brain distribution in rats

Abstract

The effective treatment of brain cancer is hindered by the poor transport across the blood–brain barrier (BBB) and the low penetration across the blood–tumor barrier (BTB). The objective of this work was to formulate transferrin-conjugated docetaxel (DTX)-loaded d-alpha-tocopheryl polyethylene glycol 1000 succinate (vitamin E TPGS or TPGS) micelles for targeted brain cancer therapy. The micelles with and without transferrin conjugation were prepared by the solvent casting method and characterized for their particle size, polydispersity, drug encapsulation efficiency, drug loading, in vitro release study and brain distribution study. Particle sizes of prepared micelles were determined at 25?°C by dynamic light scattering technique. The external surface morphology was determined by transmission electron microscopy analysis and atomic force microscopy. The encapsulation efficiency was determined by spectrophotometery. In vitro release studies of micelles and control formulations were carried out by dialysis bag diffusion method. The particle sizes of the non-targeted and targeted micelles were <20?nm. About 85% of drug encapsulation efficiency was achieved with micelles. The drug release from transferrin-conjugated micelles was sustained for >24?h with 50% of drug release. The in vivo results indicated that transferrin-targeted TPGS micelles could be a promising carrier for brain targeting due to nano-sized drug delivery, solubility enhancement and permeability which provided an improved and prolonged brain targeting of DTX in comparison to the non-targeted micelles and marketed formulation.     

Recent advances in three-dimensional cell culturing to assess liver function and dysfunction: from a drug biotransformation and toxicity perspective

The liver is a vital organ fulfilling a central role in over 500 major metabolic functions, including serving as the most essential site for drug biotransformation. Dysfunction of the drug biotransformation processes may result in the exposure of the liver (and other organs) to hepatotoxins, potentially interacting with cellular constituents and causing toxicity and various lesions. Hepatotoxicity can be investigated on a tissue, cellular and molecular level by employing various in vivo and in vitro techniques, including novel three-dimensional (3?D) cell culturing methods. This paper reflects on the liver and its myriad of functions and the influence of drug biotransformation on liver dysfunction. Current in vivo and in vitro models used to study liver function and dysfunction is outlined, emphasizing their advantages and disadvantages. The advantages of novel in vitro 3?D cell culture models are discussed and the possibility of novel models to bridge the gap between in vitro and in vivo models is explained. Progression made in the field of cell culturing methods such as 3?D cell culturing techniques over the last decade promises to reduce the use of in vivo animal models in biotransformation and toxicological studies of the liver.     

Tumor-preferential sustained drug release enhances antitumor activity of block copolymer micelles

Abstract

Nanoparticles are widely used as drug carriers for controlled, tumor-targeted delivery of various anticancer agents that have biopharmaceutical limitations such as water solubility and tissue permeability. Growing evidence suggests that nanoparticles not only reduce toxic side effects of anticancer drugs but also improve the therapeutic efficacy as a function of their drug-release profile. The purpose of this study is to confirm such hypothetical effects of tunable drug release on improving antitumor activity of nanoparticles in vitro and in vivo, using block copolymer micelles as drug carriers. Micelles were prepared from poly(ethylene glycol)-poly(aspartate) block copolymers modified with hydrazide (HYD), aminobenzoate hydrazide (ABZ) and glycine hydrazide (GLY) linkers to achieve a pH-dependent, tunable release of doxorubicin (DOX), a model anticancer drug. Regardless of the drug-release profile, all three micelles showed similar properties in vitro, such as pH-dependent drug release, intracellular drug delivery and cancer cell growth inhibition. However, micelles releasing DOX slowly in vitro showed that the most effective antitumor activity in vivo, compared to the micelles releasing drugs faster. These results demonstrate that tumor-preferential sustained drug release can enhance the antitumor activity of the micelles.     

A journey from the endothelium to the tumor tissue: distinct behavior between PEO-PCL micelles and polymersomes nanocarriers

Polymeric nanocarriers must overcome several biological barriers to reach the vicinity of solid tumors and deliver their encapsulated drug. This study assessed the in vitro and in vivo passage through the blood vessel wall to tumors of two well-characterized polymeric nanocarriers: poly(ethyleneglycol-b-ε-caprolactone) micelles and polymersomes charged with a fluorescent membrane dye (DiO: 3,3'-dioctadecyloxacarbo-cyanine perchlorate). The internalization and translocation from endothelial (human primary endothelial cells HUVEC) to cancer cells (human tumor cell line HCT-116) was studied in conventional 2D monolayers, 3D tumor spheroids, or in an endothelium model based on transwell assay. Micelles induced a faster DiO internalization compared to polymersomes but the latter crossed the endothelial monolayer more easily. Both translocation rates were enhanced by the addition of a pro-inflammatory factor or in the presence of tumor cells. These results were confirmed by early in vivo experiments. Overall, this study pointed out the room for the improvement of polymeric nanocarriers design to avoid drug losses when crossing the blood vessel walls.     

Prediction of whole-body metabolic clearance of drugs through the combined use of slices from rat liver,lung, kidney,small intestine and colon

1.?The aim was to investigate whether precision-cut rat tissue slices could be used to predict metabolic drug clearance in vivo. To obtain a complete picture, slices not only from liver, but also from lung, kidney, small intestine and colon were included.

2.?The metabolic clearances of 7-ethoxycoumarin, 7-hydroxycoumarin, testosterone, methyltestosterone and warfarin were determined by measuring the disappearance of these compounds during incubation with slices prepared from liver, lung, kidney, small intestine and colon.

3.?The total in vitro metabolic clearance was determined by adding the individual in vitro organ clearances from the slices. Prediction based on the in vitro clearance was within an order of magnitude to the corresponding in vivo values. Interestingly, the relative contribution of extrahepatic metabolic clearance of the studied compounds to total clearance was remarkably high, ranging from 35 to 72% of the total metabolic clearance.

4.?It is concluded that the model of multi-organ precision-cut slices is a useful in vitro tool for prediction of in vivo metabolic clearance. In addition, it provides information about the relative contribution of the liver, lung, kidney, small intestine and colon to the total metabolic clearance.     

Epidermal growth factor receptor-targeted peptide conjugated phospholipid micelles for doxorubicin delivery

Specific targeting of tumor cells to achieve higher drug levels in tumor tissue and to overcome side effects is the major goal in cancer chemotherapy. In this study, we used a tumor targeting peptide, GE11, to conjugate onto the surface of doxorubicin encapsulated phospholipid micelles. The GE11 peptide triggered specific binding to epidermal growth factor receptor (EGFR), leading to enhanced cellular uptake and cytotoxicity in vitro and highly accumulation in the tumors in vivo. The results indicated that GE11 conjugated phospholipid micelles should have potential applications in cancer therapy.     

Analysis of Drug Permeation Across Caco-2 Monolayer: Implication for Predicting In Vivo Drug Absorption

Purpose. The aim of the present work is to characterize in vitro drug permeation processes across Caco-2 monolayer and to identify the advantages of this cultured cell system in predicting in vivo drug absorption after oral administration. Methods. The passive permeability of various drugs through Caco-2 monolayer was measured using Ussing-type chambers and compared with that of the isolated rat jejunum and colon. The in vivo drug permeability to the intestinal membrane was estimated by means of an intestinal perfusion study using the rat jejunum. Results. In Caco-2 monolayer, drug permeability increased with increasing drug lipophilicity and showed a good linear relationship with the in vivo permeability. In contrast, in the isolated jejunum and colon, the permeability of high lipophilic drugs was almost constant and, propranolol, a drug with the highest lipophilicity, hardly passed through the jejunal membrane in vitro. As a result, there was no significant relationship between in vitro and in vivo drug permeability in rat jejunum. However, the amount of drugs accumulated in the jejunal mucosa increased with increasing drug lipophilicity even under the in vitro condition. Conclusions. The permeation and the accumulation studies suggested that the rate-limiting process of in vitro permeation of lipophilic drugs through the intestinal membrane differs from that of in vivo drug absorption. On the other hand, drug permeation through Caco-2 monolayer, which consists of an epithelial cell layer and a supporting filter, is essentially the same process as that of in vivo drug absorption. We concluded that the simple monolayer structure of a cultured cell system provides a distinct advantage in predicting in vivo drug absorption.     

Stimulation of tumor growthin vitro andin vivo by suramin on the VX2 model

Suramin is an antitrypanosomal compound with confirmed efficacy against several human malignancies. It is generally assumed that its mechanism of action includes the interaction with different growth factors, unlike most of the anticancer drugs. Its anticancer activity has not been testedin vivo against squamous cell carcinoma. The purpose of this study was to assess the efficacy and toxicity of suraminin vivo andin vitro on the VX2 tumor model at therapeutic monitored plasma concentrations. We determined the pharmacokinetics of suramin in rabbits, and modelized its administration in order to obtain plasma concentrations between 150 and 300 μg/ml throughout the treatment course of 3 weeks. Under these conditions, antitumor effects of suramin were evaluatedin vivo by comparing liver tumor involvement in suramin-treated and control rabbits. Liver involvement was quantified by image analysis andin vitro effects were also determined at the same concentrations.In vivo, suramin promoted liver tumor growth significantly (p<0.05), compared to untreated controls.In vitro, suramin significantly stimulated tumor cell growth at concentrations above 200 μg/ml (p<0.01). Suramin may have stimulatory effects on tumor growth in squamous cell carcinoma at relevant plasma drug concentrations. Caution should be taken in further trials in patients with squamous cell carcinomas.     

Neutrophil-mediated delivery of pixantrone-loaded liposomes decorated with poly(sialic acid)–octadecylamine conjugate for lung cancer treatment

Poly(sialic acid) (PSA) is a natural hydrophilic biodegradable and non-immunogenic biopolymer, receptors for its monomer are expressed on peripheral blood neutrophils (PBNs), which plays important roles in the progression and invasion of tumors. A poly(sialic acid)–octadecylamine conjugate (PSA–ODA) was synthesized and then anchor it on the surface of liposomal pixantrone (Pix-PSL), to achieve an improved anticancer effect. The liposomes were prepared using a remote loading method via a pH gradient, and then assessed for particle size, zeta potential encapsulation efficiency, in vitro release, and in vitro cytotoxicity. Simultaneously, in vitro and in vivo cellular uptake studies confirmed that PSA-decorated liposomes provided an enhanced accumulation of liposomes in PBNs. An in vivo study presented that the anti-tumor activity of Pix-PSL was superior to that of other Pix formulations, probably due to the efficient targeting of PBNs by Pix-PSL, after which PBN containing Pix-PSL (Pix-PSL/PBNs) in the blood circulation are recruited by the tumor microenvironment. These findings suggest that PSA-decorated liposomal Pix may provide a neutrophil-mediated drug delivery system (DDS) for the eradication of tumors, which represents a promising approach for the tumor targeting of chemotherapeutic treatments.     

Bio-distribution and anti-tumor efficacy of PEG/PLA nano particles loaded doxorubicin

As a more effective in vivo drug delivery system, several methods loading anti-cancer drugs to biodegeradable and biocompatible nano-particles have been explored and developed. Supposedly due to the enhanced permeability and retention (EPR) effect, systemic administration of these nano-particles have been found to result in accumulation of nano-particles into solid tumors. In this study, we prepared nano-particles using polyethylene glycol (PEG)/poly-l-lactide (PLLA) diblock copolymer and loaded doxorubicin into these nano-particles (Nano-dox). The fabricated nano-particles exhibited sustained release kinetics of the drug in vitro. To follow the in vivo biodistribution of 200–350 nm sized nano-dox particles in tumor (syngenic renal cell adenocarcinoma: RENCA) bearing mouse, the carboxylfluorescenin diacetate succinimidyl ester (CFSE) was loaded into the nano-particles. Nano-dox accumulated preferentially in tumors; however, in terms of its anti-tumor efficacy, it did not show any marked benefits, compared to freely-administered doxorubicin. This result suggests the need to re-consider and evalute what type of anti-cancer reagents we to be used in the ongoing efforts of coupling drug delivery system with tumor EPR effects.     

Feasibility study of cavitation-induced liposomal doxorubicin release in an AT2 Dunning rat tumor model

Background: Targeted and triggered release of liposomal drug using heat or ultrasound represents a promising treatment modality able to increase the therapeutic-totoxicity ratio of encapsulated drugs.

Purpose: To study the ability for high-intensity focused ultrasound to induce liposomal drug release mainly by focused inertial cavitation in vitro and in an animal model.

Methods: A 1 MHz ultrasound setup has been developed for in vitro and in vivo drug release from a specific liposomal doxorubicin formulation at a target cavitation dose.

Results: Controlled cavitation at 1 MHz was applied within the tumors 48 hours after liposome injection according to preliminary pharmacokinetic study. A small non-significant therapeutic effect of US-liposomal treatment was observed compared to liposomes alone suggesting no beneficial effect of ultrasound in the current setup.

Conclusion: The in vitro study provided a suitable ultrasound setup for delivering a cavitation dose appropriate for safe liposomal drug release. However, when converting to an in vivo model, no therapeutic benefit was observed. This may be due to a number of reasons, one of which may be the difficulty in converting in vitro findings to an in vivo model. In light of these findings, we discuss important design features for future studies.