Comparative Studies of the Uptake and Internalization Pathways of Different Lipid Nano-Systems Intended for Brain Delivery.

Lipid nano-systems were prepared and characterized in a series of well-established in vitro tests that could assess their interactions with the hCMEC/D3 and SH-SY5Y cell lines as a model for the blood-brain barrier and neuronal function, accordingly. The prepared formulations of nanoliposomes and nanostructured lipid carriers were characterized by z-average diameters of ~120 nm and ~105 nm, respectively, following a unimodal particle size distribution (PDI < 0.3) and negative Z-potential (-24.30 mV to -31.20 mV). Stability studies implied that the nano-systems were stable in a physiologically relevant medium as well as human plasma, except nanoliposomes containing poloxamer on their surface, where there was an increase in particle size of ~26%. The presence of stealth polymer tends to decrease the amount of adsorbed proteins onto a particle's surface, according to protein adsorption studies. Both formulations of nanoliposomes were characterized by a low cytotoxicity, while their cell viability was reduced when incubated with the highest concentration (100 µg/mL) of nanostructured lipid formulations, which could have been associated with the consumption of cellular energy, thus resulting in a reduction in metabolic active cells. The uptake of all the nano-systems in the hCMEC/D3 and SH-SY5Y cell lines was successful, most likely following ATP-dependent internalization, as well as transport via passive diffusion.

Lipid nano-carriers loaded with Cannabis sativa extract for epilepsy treatment - in vitro characterization and in vivo efficacy studies.

Taking into consideration the latest reported beneficial anticolvusant effects of cannabidiol (CBD) and cannabiodiolic acid (CBDA) for clinical applications and the advantages of lipid nano-systems as carriers for targeted brain delivery, the aim of this study was set in direction of in vitro physico-chemical and biopharmaceutical characterization and in vivo evaluation of nanoliposomes and nanostructured lipid carriers loaded with Cannabis sativa extract intended for safe and efficient transport via blood-brain barrier and treatment of epilepsy. These nanoliposomes and nanostructured lipid formulations were characterized with z-average diameter <200 nm, following unimodal particle size distribution, negative values for Z-potential, high drug encapsulation efficiency and prolonged release during 24h (38.84-60.91 %). Prepared formulations showed statistically significant higher antioxidant capacity compared to the extract. The results from in vivo studies of the anticonvulsant activity demonstrated that all formulations significantly elevated the latencies for myoclonic, clonic and tonic seizures and, therefore, could be used in preventing different types of seizures. A distinction in the potential of the nano-systems was noted, which was probably anticipated by the type and the characteristics of the prepared formulations.

Design and evaluation of nanostructured lipid carriers loaded with Salvia officinalis extract for Alzheimer's disease treatment.

Considering the potential of Salvia officinalis in prevention and treatment of Alzheimer's disease (AD), as well as the ability of nanostructured lipid carriers (NLC) to successfully deliver drug molecules across blood-brain barrier (BBB), the objective of this study was design, development, optimization and characterization of freeze-dried salvia officinalis extract (FSE) loaded NLC intended for intranasal administration. NLC were prepared by solvent evaporation method and the optimization was carried out using central composite design (CCD) of experiments. Further, the optimized formulation (NLCo) was coated either with chitosan (NLCc) or poloxamer (NLCp). Surface characterization of the particles demonstrated a spherical shape with smooth exterior. Particle size of optimal formulations after 0.45 µm pore size filtration ranged from 127 ± 0.68 nm to 140 ± 0.74 nm. The zeta potential was -25.6 ± 0.404 mV; 22.4 ± 1.106 mV and - 6.74 ± 0.609 mV for NLCo, NLCc, and NLCp, respectively. Differential scanning calorimetry (DSC) confirmed the formation of NLC whereas Fourier-transform infrared spectroscopy confirmed the FSE encapsulation into particles. All formulations showcased relatively high drug loading (>86.74 mcg FSE/mg solid lipid) and were characterized by prolonged and controlled release that followed Peppas-Sahlin in vitro release kinetic model. Protein adsorption studies revealed the lowest adsorption of the proteins onto NLCp (43.53 ± 0.07%) and highest protein adsorption onto NLCc (55.97 ± 0.75%) surface. The modified ORAC assay demonstrated higher antioxidative activity for NLCo (95.31 ± 1.86%) and NLCc (97.76 ± 4.00%) as compared to FSE (90.30 ± 1.53%). Results obtained from cell cultures tests pointed to the potential of prepared NLCs for FSE brain targeting and controlled release.

The impact of molecular tumor profiling on the design strategies for targeting myeloid leukemia and EGFR/CD44-positive solid tumors.

Nanomedicine has emerged as a novel cancer treatment and diagnostic modality, whose design constantly evolves towards increasing the safety and efficacy of the chemotherapeutic and diagnostic protocols. Molecular diagnostics, which create a great amount of data related to the unique molecular signatures of each tumor subtype, have emerged as an important tool for detailed profiling of tumors. They provide an opportunity to develop targeting agents for early detection and diagnosis, and to select the most effective combinatorial treatment options. Alongside, the design of the nanoscale carriers needs to cope with novel trends of molecular screening. Also, multiple targeting ligands needed for robust and specific interactions with the targeted cell populations have to be introduced, which should result in substantial improvements in safety and efficacy of the cancer treatment. This article will focus on novel design strategies for nanoscale drug delivery systems, based on the unique molecular signatures of myeloid leukemia and EGFR/CD44-positive solid tumors, and the impact of novel discoveries in molecular tumor profiles on future chemotherapeutic protocols.

Implementation of quality by design principles in the development of microsponges as drug delivery carriers: Identification and optimization of critical factors using multivariate statistical analyses and design of experiments studies.

Microsponges drug delivery system (MDDC) was prepared by double emulsion-solvent-diffusion technique using rotor-stator homogenization. Quality by design (QbD) concept was implemented for the development of MDDC with potential to be incorporated into semisolid dosage form (gel). Quality target product profile (QTPP) and critical quality attributes (CQA) were defined and identified, accordingly. Critical material attributes (CMA) and Critical process parameters (CPP) were identified using quality risk management (QRM) tool, failure mode, effects and criticality analysis (FMECA). CMA and CPP were identified based on results obtained from principal component analysis (PCA-X&Y) and partial least squares (PLS) statistical analysis along with literature data, product and process knowledge and understanding. FMECA identified amount of ethylcellulose, chitosan, acetone, dichloromethane, span 80, tween 80 and water ratio in primary/multiple emulsions as CMA and rotation speed and stirrer type used for organic solvent removal as CPP. The relationship between identified CPP and particle size as CQA was described in the design space using design of experiments - one-factor response surface method. Obtained results from statistically designed experiments enabled establishment of mathematical models and equations that were used for detailed characterization of influence of identified CPP upon MDDC particle size and particle size distribution and their subsequent optimization.

Formulation and characterization of ORMOSIL particles loaded with budesonide for local colonic delivery.

In this study, hybrid silica xerogel particles were developed as carriers of budesonide (BDS) for efficient local treatment of inflammatory bowel diseases (IBD). Organically modified silica particles (ORMOSILs) were prepared by co-condensation of 3-aminopropyltriethoxysilane (APTES) and tetraethyl orthosilicate (TEOS) by an ambient temperature acid catalysed sol-gel process followed by spray-drying. Formulation for preparation of BDS-loaded particles was optimized and their physicochemical parameters and drug release profiles were evaluated in vitro. Optimal formulation had a small particle size (mean diameter of 1.45±0.02µm) with unimodal narrow size distribution and high encapsulation efficiency (98.0 ± 1.85%). Due to the positive surface charge originated from amino group of APTES, ORMOSILs showed excessive mucoadhesiveness in comparison to native TEOS particles. The drug release decreased with increasing pH from 2.0 to 7.4. In order to avoid undesirable erroneous performance in the upper GI tract, particles were additionally coated with Eudragit(®) FS 30D, as a barrier to the drug release at pH range from 2.0 to 7.0. After Eudragit(®) FS 30D coating, the release of BDS in acidic media was sustained, while no significant differences in drug release were observed at pH 7.4. In conclusion, pH-responsive ORMOSILs showed great potential for efficient BDS delivery to the colon region.

Wheat germ agglutinin-functionalised crosslinked polyelectrolyte microparticles for local colon delivery of 5-FU: in vitro efficacy and in vivo gastrointestinal distribution.

We have previously reported the development and characterisation of wheat germ agglutinin (WGA)-functionalised chitosan-Ca-alginate (CTS-Ca-ALG) microparticles (MPs) loaded with acid-resistant particles of 5-fluorouracil (5-FU). In the present work, our goal was to evaluate the potential of these carriers for efficient treatment of colon cancer by studying in vitro permeability and cell association of 5-FU and [methyl-³H]thymidine uptake in Caco-2 cells, as well as in vivo gastrointestinal distribution. The amount of 5-FU permeated through Caco-2 cells was 15.1, 7.7 and 6.5% for 5-FU solution, CTS-Ca-ALG MPs and WGA conjugates. The concentration of 5-FU associated with Caco-2 cells was significantly greater when delivered from MPs. By incorporation of 5-FU into MPs and further decoration with WGA, an increased [methyl-³H]thymidine uptake was observed few hours after continuous drug treatment followed by significantly reduced uptake after 6 h. Gastrointestinal distribution was in favour of increased localisation and concentration of the particles in colon region.

Chitosan coated Ca-alginate microparticles loaded with budesonide for delivery to the inflamed colonic mucosa.

Using a novel one-step spray-drying process uncoated and Eudragit S 100 coated chitosan-Ca-alginate microparticles efficiently loaded with budesonide (BDS), with bioadhesive and controlled release properties in GIT, were prepared. Microparticles were spherical with mean particle size of 4.05-5.36 microm, narrow unimodal distribution and positive surface charge. A greater extent of calcium chloride limited the swelling ratio of beads, while swelling behaviour of coated beads was mainly determined by properties of enteric coating. Comparing the release profiles of formulations, under different pH conditions, influence of polymer properties and concentration of cross-linker on the rate and extent of drug release was evident. Coating has successfully sustained release of BDS in buffers at pH 2.0 and 6.8, while providing potential for efficient release of BDS at pH 7.4. Release data kinetics indicated influence of erosion and biodegradation of polymer matrix on drug release from microparticles. Prepared formulations were stable for 12 months period at controlled ambient conditions. In conclusion coated microparticles prepared by one-step spray-drying procedure could be suitable candidates for oral delivery of BDS with controlled release properties for local treatment of inflammatory bowel diseases.

5-Fluorouracil in topical liposome gels for anticancer treatment--formulation and evaluation.

Liposome gels bearing an antineoplastic agent, 5-fluorouracil, intended for topical application have been prepared and drug release properties in vitro have been evaluated. Different formulations of liposomes were prepared by the film hydration method by varying the lipid phase composition (PL 90H/cholesterol mass ratio) and hydration conditions of dry lipid film (drug/aqueous phase mass ratio). Topical liposome gels were prepared by incorporation of lyophilized liposomes into a structured vehicle (1%, m/m, chitosan gel base). Also, hydrogels containing different concentrations of 5-fluorouracil were prepared and drug release properties were investigated. The rate of drug release from liposome gels was found to be dependent on the bilayer composition and the dry lipid film hydration conditions. Also, liposomes embedded into a structured vehicle of chitosan showed significantly slower release than hydrogels. The drug release obeyed the Higuchi diffusion model, while liposomes acted as reservoir systems for continuous delivery of the encapsulated drug.