Local perivascular delivery of anti-restenotic agents from a drug-eluting poly(epsilon-caprolactone) stent cuff

The introduction of drug-eluting stents (DES) to prevent in-stent restenosis is one of the major advances in interventional cardiology. Currently many types of DES are under evaluation for effectiveness and safety, a time-consuming and difficult procedure in humans. An animal model that allows rapid evaluation of the present and upcoming therapeutic approaches to prevent in-stent restenosis is most valuable and still lacking. Here, a perivascular cuff to induce restenosis was constructed of a poly(epsilon-caprolactone) (PCL) formulation suitable for the controlled delivery of drugs. Placing the PCL cuff around the femoral artery, in vivo, resulted in reproducible restenosis-like lesions containing predominantly smooth muscle-actin positive cells. Loading the cuff with the anti-restenotic compounds paclitaxel and rapamycin resulted, in vitro, in a sustained and dose-dependent release for at least 3 weeks. Paclitaxel- and rapamycin-eluting PCL cuffs placed around the femoral artery of mice in vivo significantly reduced intimal thickening by 76 +/- 2% and 75 +/- 6%, respectively, at 21 days. Perivascular sustained release of both anti-restenotic agents is restricted to the cuffed vessel segment with no systemic adverse effects or effect on cuffed contralateral femoral arteries. Drug-eluting PCL cuffs provide an easy and rapid tool to evaluate anti-restenotic agents to be used in combination with the DES strategies.     

Development and in vitro evaluation of a thiomer-based nanoparticulate gene delivery system

Chitosan-thiobutylamidine was developed and evaluated as a novel tool for gene delivery. The conjugate, displaying 299.1+/-11.5 micromol free thiol groups per gram polymer, formed coacervates with pDNA at a mean size of 125 nm and a zeta potential of +9 mV. Thiol groups, being susceptible for oxidation, were immobilised on the polymeric backbone of chitosan in order to introduce the property of extracellular stability and intracellular pDNA release by forming reversible disulfide bonds. The integrity of the new particles was compared to unmodified chitosan under simulated physiological conditions. Within 10h, pDNA was completely released from chitosan-DNA particles while only 12% were released from the thiomer-based particles. At pH 7, the amount of thiol groups significantly (p<0.05) decreased by more than 25% within 6h. In contrast, in a reducing environment as found intracellularly, chitosan-thiobutylamidine-DNA nanoparticles dissociated continuously, liberating approximately 50% of pDNA within 3h. Transfection studies performed in a Caco2 cell culture evinced the highest efficiency for chitosan-thiobutylamidine-DNA nanoparticles in combination with a glycerol shock solution. The combination of improved stability, enhanced pDNA release under reducing conditions, and higher transfection efficiency identifies chitosan-thiobutylamidine as a promising new vector for gene delivery.     

Chitin/PLGA blend microspheres as a biodegradable drug delivery system: a new delivery system for protein

Novel chitin/PLGAs and chitin/PLA based microspheres were developed for the delivery of protein. These biodegradable microspheres were prepared by polymers blending and wet phase-inversion methods. The parameters such as selected non-solvents, temperature of water and ratio of polylactide to polyglycolide were adjusted to improve thermodynamic compatibility of individual polymer (chitin and PLGAs or chitin/PLA), which affects the hydration and degradation properties of the blend microspheres. Triphasic pattern of drug release model is observed from the release of protein from the chitin/PLGAs and chitin/PLA microspheres: the initially fast release (the first phase), the following slow release (the second phase) and the second burst release (the third phase). Formulations of the blends, which are based on the balance among the hydration rate of the chitin phase and degradation of chitin/PLA and PLGA phase, can lead to a controllable release of bovine serum albumin (BSA). In conclusion, such a chitin/PLGA 50/50 microsphere is novel and interesting, and may be used as a protein delivery system.     

Monoolein aqueous dispersions as a delivery system for quercetin

This study describes the preparation, characterization and in vitro release of monoolein aqueous dispersions (MAD) encapsulating quercetin (QT). As emulsifier, sodium cholate was employed at two different concentrations, namely 0.15% and 0.25% with respect to the total weight of the formulation. Cryogenic Transmission electron microscopy and X-ray analysis indicated that MAD015 are a mixture of vesicles and cubic structures, whilst MAD025 are mainly characterized by unilamellar vesicular structures. Photon correlation spectroscopy (PCS) and Sedimentation Field Flow Fractionation (SdFFF) showed a MAD size higher than 300 nm that over 100 days from analysis reduces up to 200 nm. In vitro Franz cell experiments showed that the two systems had a similar behaviour in the release of QT. Experiments on antioxidant activity of MAD containing QT demonstrated that their activity parallel with the content of encapsulated QT within the MAD formulations produced. Taken together these results allow us to conclude that MAD can be potentially proposed for the delivery of QT.     

Development of topical BAPN delivery system for acute spinal cord injury in dogs

Topical sustained release of various medications by a subdurally implantable device at the site of spinal cord injury is considered advantageous in the treatment of early symptoms of tissue damage. A typical case is the interference with collagenous scar by beta-aminopropionitrile, inhibiting collagen polymerization. Four materials, silicone, polyethylene, polytetrafluoroethylene (PTFE), and polyacrylonitrile-based hydrogel were evaluated for biocompatibility in subcutaneous implantations. The hydrogel, the least reactive, was then compared with silicone sheets as subcural implants. The histology favored the hydrogel as the most inert material, which was then used for the construction of soft, pliable pouches, releasing the drug through the hydrated wall at a rate controlled by an osmotic pump.     

Development and in vivo evaluation of an oral drug delivery system for paclitaxel

The aim of the present study was to investigate the effect of poly(acrylic acid)-cysteine (PAA-cysteine) exhibiting a molecular mass of 100 and 250 kDa and reduced glutathione (GSH) on the absorption of the P-glycoprotein (P-gp) and cytochrome P450 (CYP450) substrate paclitaxel in vitro and in vivo. In vitro transport studies were performed with Caco-2 monolayers. Furthermore, the delivery system based on PAA-cysteine, GSH and paclitaxel was evaluated in vivo in rats. In vitro, the formulation comprising 0.5% (m/v) PAA-cysteine (100 kDa)/0.5% (m/v) GSH improved the transport of paclitaxel 6.7-fold (P(app) = 8.7 ± 1.3 × 10(-6) cm/s) in comparison to paclitaxel itself serving as buffer only control (P(app) = 1.3 ± 0.4 × 10(-6) cm/s). Moreover, in the presence of the formulation containing 0.5% (m/v) PAA-cysteine (250 kDa)/0.5% (m/v) GSH paclitaxel absorption was even 7.4-fold (P(app) = 9.7 ± 0.3 × 10(-6) cm/s) improved in comparison to the buffer only control. In vivo, the oral administration of formulations containing 1 mg of paclitaxel, 1 mg of GSH and 8 mg of PAA-cysteine (100 kDa or 250 kDa) resulted in an improved paclitaxel plasma concentration and bioavailability. The area under the plasma concentration-time curve (AUC(0-8)) of paclitaxel was 4.7-fold and 5.7-fold improved in comparison to the oral formulation containing paclitaxel alone, respectively. Moreover, c(max) was improved by 6.3-fold and even 7.3-fold in comparison to the oral formulation containing paclitaxel alone, respectively. Thus, according to the achieved results it is suggested that PAA-cysteine in combination with GSH would be a potentially valuable tool for improving the oral bioavailability of P-gp and CYP450 substrates such as paclitaxel.     

Development and evaluation of a complementation-dependent gene delivery system based on cucumber mosaic virus

To engineer cucumber mosaic virus (CMV-Ix) into a gene vector, genome component RNA 3 of the virus was modified and split into two sub-components, RNA 3A and RNA 3B. In RNA 3A, the open reading frame of the movement protein (MP) was replaced by a reporter gene encoding the green fluorescent protein (GFP), to monitor virus replication and movement. In RNA 3B, the coat protein (CP) gene was eliminated and a multiple cloning site (MCS) was created for foreign gene insertion. Each sub-component alone is defective and relies on its companion sub-component to restore full RNA 3 function. The vector system was evaluated for its ability to deliver and express the bacterial beta-glucuronidase (GUS) gene and a modified bean yellow mosaic virus coat protein (BYMV-CP) gene in Nicotiana benthamiana plants. Results showed that the engineered virus was able to move from cell to cell in the inoculated leaf and enter the minor veins of the inoculated leaf. Foreign gene expression was detected in the inoculated leaves. However, intermolecular recombination between RNA 3A and 3B occurred frequently, preventing efficient systemic expression of the foreign gene(s). Modifications and further evaluations are being undertaken to improve the gene delivery system.     

Design of a continuous-wave Doppler ultrasonic flowmeter for perivascular application

The paper details the specification of an ultrasonic Doppler flowmeter intended for perivascular application and describes the elements of the design of an ultrasonic blood flow transducer. An optimised design is outlined for probes to be used on vessels in the range 5–12 mm. The design is based on the use of continuous-wave Doppler techniques and incorporates a novel polymer ultrasonic couplant. A constrained jet technique is used to test the performance of the probe in vitro and the results indicate that in use velocity profile effects are likely to lead to sensitivity changes of only 3 per cent. A companion paper details the design and performance of the electronic processing system needed for the complete flowmeter.     

Design of a continuous-wave Doppler ultrasonic flowmeter for perivascular application

The paper details the design of the signal processing system of an ultrasonic Doppler flowmeter intended for perivascular application. The system is designed to work in conjunction with a transducer system described in an earlier paper, in a range of vessels varying from 5 mm to 12 mm. The system incorporates conventional quadrature processing for directional discrimination and uses phaselocked loop technology to extract signals corresponding to the instantaneous mean frequency (velocity equivalent). The use of a PLL also permits the extraction of a quasi-spectral analysis signal. The baseline stability of the Doppler system is enhanced by the use of an auto-zeroing system which enables the system to resolve Doppler shifts down to 20Hz. Testing of the complete transducer/signal processor system in a hydraulic rig demonstrates the ability of the flowmeter to accurately measure both steady and pulsatile flow.     

Dose error analysis for a scanned proton beam delivery system

All particle beam scanning systems are subject to dose delivery errors due to errors in position, energy and intensity of the delivered beam. In addition, finite scan speeds, beam spill non-uniformities, and delays in detector, detector electronics and magnet responses will all contribute errors in delivery. In this paper, we present dose errors for an 8 × 10 × 8 cm(3) target of uniform water equivalent density with 8 cm spread out Bragg peak and a prescribed dose of 2 Gy. Lower doses are also analyzed and presented later in the paper. Beam energy errors and errors due to limitations of scanning system hardware have been included in the analysis. By using Gaussian shaped pencil beams derived from measurements in the research room of the James M Slater Proton Treatment and Research Center at Loma Linda, CA and executing treatment simulations multiple times, statistical dose errors have been calculated in each 2.5 mm cubic voxel in the target. These errors were calculated by delivering multiple treatments to the same volume and calculating the rms variation in delivered dose at each voxel in the target. The variations in dose were the result of random beam delivery errors such as proton energy, spot position and intensity fluctuations. The results show that with reasonable assumptions of random beam delivery errors, the spot scanning technique yielded an rms dose error in each voxel less than 2% or 3% of the 2 Gy prescribed dose. These calculated errors are within acceptable clinical limits for radiation therapy.     

Biodegradable microspheres as a vaccine delivery system

The utility of biodegradable and biocompatible microspheres as a vaccine delivery system for the induction of systemic and disseminated mucosal antibody responses was investigated. Intraperitoneal (ip) injection into mice of 1-10 microns microspheres, constructed of the copolymer poly(DL-lactide-coglycolide) (DL-PLG) which contained approximately 1% by weight a formalinized toxoid vaccine of staphylococcal enterotoxin B (SEB), dramatically potentiated the circulating IgG anti-toxin antibody response as compared to the free toxoid. The initiation of vaccine release was delayed in larger microspheres, and a mixture of 1-10 and 20-50 microns microspheres stimulated both a primary and an anamnestic secondary anti-toxin response following a single injection. However, neither free nor microencapsulated SEB toxoid induced a detectable mucosal IgA anti-toxin response following systemic injection. In contrast, three peroral immunizations with toxoid-microspheres stimulated circulating IgM, IgG and IgA anti-toxin antibodies and a concurrent mucosal IgA response in saliva, gut washings and lung washings. Systemic immunization with microencapsulated toxoid primed for the induction of disseminated mucosal IgA responses by subsequent oral or intratracheal (it) boosting in microspheres, while soluble toxoid was ineffective at boosting. These results indicate that biodegradable and biocompatible microspheres represent an adjuvant system with potentially widespread application in the induction of both circulating and mucosal immunity.     

A polymeric micelle system with a hydrolysable segment for drug delivery

A potential anti-cancer drug-delivery polymeric micelle system with an in vitro degradation half-life of about 48 h that releases its drug upon application of ultrasound was synthesized. This vehicle was composed of an amphiphilic co-polymer, poly(ethylene oxide)-b-poly(N-isopropylacrylamide-co-2-hydroxyethyl methacrylate-lactate _n ). The degree of polymerization of the lactate side group, n, was 0, 3 or 5. The molar ratio of NIPAAm to HEMA-lactate _n to PEO in polymerization was optimized to produce an in vitro polymeric micelle half-life of about 48 h at 40°C. 1,6-Diphenyl-1,3,5-hexatriene (DPH) was used as a fluorescent probe to study the hydrophobicity of the cores of the polymeric micelles. The results showed that the cores of the polymeric micelles were hydrophobic enough to sequester DPH and the anti-cancer drug doxorubicin (Dox). Dox was encapsulated into the polymeric micelles having a molar feed ratio of NIPAAm to HEMA-lactate₃ to PEO equal to 20 : 5 : 1; this drug was released upon the application of low-frequency ultrasound. The Dox release was about 2% at room temperature and 4% at body temperature, and the drug returned to the polymeric micelles when insonation ceased.     

A polymeric micelle system with a hydrolysable segment for drug delivery

A potential anti-cancer drug-delivery polymeric micelle system with an in vitro degradation half-life of about 48 h that releases its drug upon application of ultrasound was synthesized. This vehicle was composed of an amphiphilic co-polymer, poly(ethylene oxide)-b-poly(N-isopropylacrylamide-co-2-hydroxyethyl methacrylate-lactate(n)). The degree of polymerization of the lactate side group, n, was 0, 3 or 5. The molar ratio of NIPAAm to HEMA-lactate(n) to PEO in polymerization was optimized to produce an in vitro polymeric micelle half-life of about 48 h at 40 degrees C. 1,6-Diphenyl-1,3,5-hexatriene (DPH) was used as a fluorescent probe to study the hydrophobicity of the cores of the polymeric micelles. The results showed that the cores of the polymeric micelles were hydrophobic enough to sequester DPH and the anti-cancer drug doxorubicin (Dox). Dox was encapsulated into the polymeric micelles having a molar feed ratio of NIPAAm to HEMA-lactate3 to PEO equal to 20:5:1; this drug was released upon the application of low-frequency ultrasound. The Dox release was about 2% at room temperature and 4% at body temperature, and the drug returned to the polymeric micelles when insonation ceased.     

Building a polysaccharide hydrogel capsule delivery system for control release of ibuprofen

Development of a delivery system which can effectively carry hydrophobic drugs and have pH response is becoming necessary. Here we demonstrate that through preparation of β-cyclodextrin polymer (β-CDP), a hydrophobic drug molecule of ibuprofen (IBU) was incorporated into our prepared β-CDP inner cavities, aiming to improve the poor water solubility of IBU. A core-shell capsule structure has been designed for achieving the drug pH targeted and sustained release. This delivery system was built with polysaccharide polymer of Sodium alginate (SA), sodium carboxymethylcellulose (CMC) and hydroxyethyl cellulose (HEC) by physical cross-linking. The drug pH-response control release is this hydrogel system’s chief merit, which has potential value for synthesizing enteric capsule. Besides, due to our simple preparing strategy, optimal conditions can be readily determined and the synthesis process can be accurately controlled, leading to consistent and reproducible hydrogel capsules. In addition, phase-solubility method was used to investigate the solubilization effect of IBU by β-CDP. SEM was used to prove the forming of core and shell structure. FT-IR and ¹H-NMR were also used to perform structural characteristics. By the technique of UV determination, the pH targeted and sustained release study were also performed. The results have proved that our prepared polysaccharide hydrogel capsule delivery system has potential applications as oral drugs delivery in the field of biomedical materials.     

Polymeric micelle-templated synthesis of hydroxyapatite hollow nanoparticles for a drug delivery system

Hydroxyapatite (HA) hollow nanoparticles (HNPs) have great potential in nanoscaled delivery devices due to their small size, excellent biocompatibility and expected high capacity. However, the preparation of HA HNPs for their application in a drug delivery system has rarely been reported because HA has a complicated crystal structure and it is difficult to obtain stable HA HNPs with hollows that are only nanoscaled in size. In the present study, HA HNPs were successfully produced through a novel polymeric micelle-templating method. The micelles were structured with completely insoluble Pluronic P123 molecules at cloud point as the core and Tween-60 molecules as the shell by the hydrophobic interaction of the alkyl chains with the insoluble P123 core. The morphology of the HA HNPs could be transformed from nanospheres to nanotubes by adding citric acid as a cosurfactant. The prepared HA HNPs had a much higher drug payload than traditional nanoparticles, using vancomycin as the model drug. Most importantly, the HA nanotubes were coupled with a layer of citrate molecules on the HA surfaces, which could further improve the drug load efficiency and could form an excellent pH-controlled open/closed gate for drug release with the addition of cationic polyelectrolytes.     

Fluorination of electrospun hydrogel fibers for a controlled release drug delivery system

Electrospinning and fluorination were carried out in order to obtain a controlled release drug delivery system to solve the problem of both an initial burst of the drug and a limited release time. Poly(vinyl alcohol) was electrospun with Procion Blue as a model drug and heat treated in order to obtain cross-linked hydrogel fibers. Two different kinds of electrospun fibers of thin and thick diameters were obtained by controlling the electrospinning conditions. Thin fibers offer more available sites than thick fibers for surface modification during fluorination. Fluorination was conducted to control the release period by introducing hydrophobic functional groups on the surface of fibers. With an increase in the reaction pressure of the fluorine gas hydrophobic C–F and C–F₂ bonds were more effectively introduced. Over-fluorination of the fibers at higher reaction pressures of fluorine gas led to the introduction of C–F₂ bonds, which made the surface of the fibers hydrophobic and resulted in a decrease in their swelling potential. When C–F bonds were generated the initial drug burst decreased dramatically and total release time increased significantly, by a factor of approximately 6.7 times.     

Development of a local vasodilator delivery system using fibrin glue to prevent arterial graft from spasm

The clinical benefits of coronary artery bypass graft operations can be compromised by postoperative vasospasm. Traditionally, local papaverine (PPV) has been employed during the procedure to prevent and counteract vasospasm. But the relatively short action period limited its application. Fibrin glue (FG) might be a potential carrier of PPV for counteracting vasospasm in a longer action period than PPV solution. After FG incorporated with PPV (PPV-FG) was locally administrated in axillary and femoral arteries of dogs, PPV concentrations in artery vessels surrounding the administration sites were compared with the concentrations at the same sites in dogs given PPV solution. The properties of PPV's release in vitro and maintenance in vessel as well as the influence on the mean peripheral blood pressure and drug concentration in peripheral vein after the introduction PPV-FG on the surface of artery in dogs were evaluated. FG was considered to provide a sustained release of PPV and could maintain a high PPV concentration in artery vessel around the administration site. The results suggested that FG was an effective substrate for reserving PPV in the administrated site in a defined period.     

Development of an injectable two-phase drug delivery system for sequential release of antiresorptive and osteogenic drugs

Unlike controlled release systems that deliver a single drug, dual or multidrug delivery systems with distinct release profiles are more likely to promote timely and effective tissue regeneration as they provide both temporally and concentration-dependent release of different molecules to mimic natural biological events. In this study, an injectable and biodegradable delivery system was developed to sequentially release an antiresorptive drug (clodronate) followed by an osteogenic agent (simvastatin) to treat bone disease. The injectable delivery system comprised simvastatin-loaded gelatin microspheres suspended in a viscous solution of carboxymethylcellulose (CMC) containing clodronate. Several factors (CMC concentration, glutaraldehyde concentration, simvastatin loading, and gelatin microsphere processing conditions) were investigated for their effects on drug release. Clodronate release was not affected by CMC concentration, with complete delivery within 12 hr, and simvastatin release could be modulated by cross-linking of the gelatin microspheres, loading, and washing conditions. Burst release of simvastatin was reduced from 70% to 6% in conjunction with sustained release for up to 3 weeks. The combined system showed early release of the antiresorptive clodronate sequentially followed by sustained delivery of the osteogenic simvastatin. This robust and flexible two-phase delivery system may prove useful for applications in which multiple drug delivery is desired.     

A pH-sensitive charge-conversion system for doxorubicin delivery

A novel pH-sensitive charge-conversion shielding system was designed by the electrostatic binding of polyethylenimine (PEI)-poly(l-lysine)-poly(l-glutamic acid) (PELG), PEI, and cis-aconityl-doxorubicin (CAD). Doxorubicin (DOX) was modified by cis-aconityl linkage to form acid-sensitive CAD, which was then adsorbed by the positively charged PEI. The PEI/CAD complexes were subsequently shielded with the pH-responsive charge-conversion PELG. In normal tissues, the PELG/PEI/CAD complexes were negatively charged; in acidic tumor tissues, the shielding PELG was positively charged and detached from the PELG/PEI/CAD complexes. The resulting positively charged PEI/CAD complexes thus became exposed and were endocytosed. CAD was then cleaved in the acidic intracellular environment of endosomes and lysosomes, and converted back into DOX. The charge reversal of the PELG/PEI/CAD complexes was verified by zeta potential analysis at different pH values. Moreover, DOX release increased with decreasing pH. Cell uptake and confocal laser scanning microscopy analyses showed that, at pH 6.8, PELG/PEI/CAD had the highest endocytosis rate and more DOX entered cell nuclei. More importantly, the system showed remarkable cytotoxicity against cancer cells. These results revealed that the combination of pH-sensitive charge-conversion shielding with pH-sensitive drug release is a potential drug delivery system for tumor treatment.