Formulation Development and Evaluation of Migraine Almotriptan Loaded Ethosomes Using Box Behnken Design

One of the most common nervous system illnesses is headache disorders, which are characterized by chronic headaches. In Present investigation Almotriptan loaded Ethosomes were prepared by water phase addition method. The three independent factors including Phosphotidylcholine: Cholestrol: DSPE (6:3:1molar ratio), Surfactant concentration and sonication time. A factorial design 3*3(3 factor 3 level) was applied to prepare 17 formulation. Optimization of ethosomal preparation was carried out by applying Box Behnken response surface randomized factorial design following quadratic model using Design of Experiment (DOE) software version 11.04.0. The factor Soya PC: Cholesterol: DSPE in molar ration (6:3:1), Concentration of Tween-80 and sonication time were selected as dependable process and formulation factors that can be effect formulation characteristics like entrapment efficiency, average vesicle size, Polydispersity Index (PDI). All other factors like sonication speed and rotation speed was kept constant All the formulation were prepared by simple solvent evaporation thin film formation method and characterized for the drug entrapment, average vesicle size and PDI, shape morphology. Formulations were optimized on the basis of responses such as average vesicle size, PDI, and entrapment efficiency. All the characterized values of the responses were putted in the formulation design table and analyse to best fitted model for the design. It was observed that, quadaratic model is best fitted model for the design. The prepared ethosomes formulation can further incorporated in situ gel for effective treatment of migraine.

Development of Itraconazole Loaded Transethosomes for improved transdermal delivery

Objective: The objective of the present work was to develop, optimize and characterize itraconazole loaded transethosomes for enhanced transdermal delivery. In this study, screening of formulation and process variables was conducted by using Box-Behnken design approach to observe significant and insignificant influence on the transethosomes. Methods: The transethosomes was developed by homogenization technique (hot method). The optimized itraconazole loaded transethosomes were evaluated for its vesicle size, polydispersity index, zeta potential, loading capacity and entrapment efficiency. Characterization was done by P-XRD, DSC and TEM. Further, in-vitro drug release study, stability study and confocal laser scanning microscopy (CLSM) study were also performed. Results: The itraconazole loaded transethosomes are developed by using soya lecithin as phospholipid, oleic acid as edge activator and cholesterol as stabilizer. Developed transethosomes showed acceptable desired vesicle size (207-409 nm), excellent colloidal dispersion characteristics (polydispersity index- 0.131 to 0.312, zeta potential -16.12to -21.96 mV) and high drug entrapment (63.37-73.02%). P-XRD and DSC results suggested that itraconazole encapsulated in amorphous state within transethosomes. In-vitro drug release study show prolonged release of itraconazole for 24 hr and confocal laser scanning microscopy confirmed accumulation of transethosomes in deeper layers of the skin. Results of stability studies showed optimized transethosomes are more stable in refrigerated temperature (4°C) as compared to room temperature (25°C). Conclusion: The results suggested that transethosomes could be better alternative to deliver drugs across the skin and potential carrier for efficient transdermal drug delivery.

EUDRAGIT COATED ALGINATE BEADS BEARING OXALIPLATIN LOADED LIPOSOMES: FORMULATION, OPTIMIZATION AND IN VITRO CHARACTERIZATION

Objective: The present investigation aimed to develop and characterize Eudragit S-100 coated alginate beads bearing oxaliplatin loaded liposomes for colon-specific drug delivery. Methods: Liposomes were formulated by the thin-film hydration method. The process and formulation variables were optimized by Box-Behnken design (BBD) with the help of Design-Expert® Software. Three independent variables taken were HSPC: Chol molar ratio (X1), hydration time (X2), and sonication time (X3). The response variables selected were entrapment efficiency of oxaliplatin, polydispersity index, and vesicle size. Results: The liposomes possessed an average vesicle size of 110.1±2.8 nm, PDI 0.096±0.3, zeta potential of-6.70±1.4 mV, and entrapment efficiency of 27.65%. The beads were characterized for their size, in vitro drug release, and swelling index. The degree of swelling of the beads was found to be 2.3 fold higher at pH 7.4 than at pH 1.2. The in vitro drug release depicted a sustained drug release in 48 h. Conclusion: The outcomes of the study proposed that the developed system can be effectively used for site-specific drug delivery to the colon via the oral route.

A novel method for producing functionalized vesicles that efficiently deliver oligonucleotides in vitro and in vivo in mice

Nano-based delivery systems have greatly enhanced our ability to administer and target drugs and macromolecules to their therapeutic targets. Oligonucleotide drugs have great therapeutic potential but have off-target effects and stability issues, so they are often encapsulated in functionalized vesicles with targeting ligands such as antibodies (Ab). Herein, we describe a novel, scalable and straightforward approach to produce functionalized vesicles called the “Functionalized Lipid Insertion Method.” This method differs significantly from an older approach referred to as the “Detergent-Dialysis Method.” The older method requires excess detergent and extensive dialysis over many hours to produce the functionalized vesicles. With the “Functionalized Lipid Insertion Method,” only the functionalized lipid is detergent-solubilized during the formation of the functionalized vesicle. The approach reduces the dialysis time, keeps the vesicle intact, and orients the functionalized lipid to improve targeting compared to the older method. The dynamic light scattering (DLS) technique demonstrated that vesicle size is sensitive to the initial detergent-solubilized component mixture by the older method. In contrast, functionalized vesicle size increases are consistent with functionalized lipid insertion into the vesicle. In vitro, functionalized vesicles using our approach are able to deliver oligonucleotides selectively and can functionally affect liver cancer HepG2 cells. Functionalized vesicles produced by this method can also achieve targeted delivery of oligonucleotides in mice without inducing a significant immune response through cytokine production or showing physical signs of an immune response. The industrial and therapeutic significance and implications of functionalized vesicles produced by our method are also discussed.

Low-Power Sonication Can Alter Extracellular Vesicle Size and Properties

Low-power sonication is widely used to disaggregate extracellular vesicles (EVs) after isolation, however, the effects of sonication on EV samples beyond dispersion are unclear. The present study analysed the characteristics of EVs collected from mesenchymal stem cells (MSCs) after sonication, using a combination of transmission electron microscopy, direct stochastic optical reconstruction microscopy, and flow cytometry techniques. Results showed that beyond the intended disaggregation effect, sonication using the lowest power setting available was enough to alter the size distribution, membrane integrity, and uptake of EVs in cultured cells. These results point to the need for a more systematic analysis of sonication procedures to improve reproducibility in EV-based cellular experiments.

Enlarged extracellular vesicles are a negative prognostic factor in patients undergoing TACE for primary or secondary liver cancer–a case series

Background Transarterial chemoembolization (TACE) has evolved as a standard treatment option in patients with intermediate stage, unresectable HCC [Barcelona Clinic Liver Cancer (BCLC) stage B] as well as in patients with liver metastases, when surgery or systemic therapy is considered not appropriate. Concentration and sizes of extracellular vesicles (EVs) recently emerged as novel diagnostic and prognostic biomarkers in patients with liver cancer, but no data on its prognostic relevance in the context of TACE exists. Here, we evaluate pre-interventional EVs as a potential biomarker in patients undergoing TACE for primary and secondary hepatic malignancies. Methods Vesicle size distribution and concentration were measured by nanoparticle tracking analysis (NTA) in patient sera before and after TACE in 38 patients. Results Extracellular vesicle size distribution measured before TACE is of prognostic significance with respect to overall survival in patients after TACE. Overall survival is significantly reduced when initial vesicle size (X50) is in the upper quartile (>145.65nm). Median overall survival in patients in the upper quartile was only 314 days, compared to 799 days in patients with vesicle size in the first to third quartile (<145.65nm; p = 0.007). Vesicle size was also shown to be a significant prognostic marker for overall survival in Cox regression analysis [HR 1.089, 95% CI: 1.021–1.162, p = 0.010]. In addition, a significant correlation was observed between initial EVs concentration/BMI (rS = 0.358, p = 0.029), X50/IL-8-concentration (rS = 0.409, p = 0.011) and X50/CRP-concentration (rS = 0.404, p = 0.016). In contrast, with regard to immediate tumor response after TACE, EVs concentration and size did not differ. Summary Sizes (but not concentrations) of EVs represent a novel prognostic marker in patients receiving TACE for primary and secondary hepatic malignancies since patients with enlarged EVs display a significantly impaired prognosis after TACE.

Preparation and Biological Evaluation of Silybin Liposomes for the Treatment of Liver Disorders

Aim: The aim of the present study was to develop silybin liposome by incorporating phosphatidyl choline & cholesterol so as to increase its oral bioavailability and liver targeted enhanced hepatoprotection. Methodology: Thin film hydration technique was used for the development of liposomes by using phosphatidyl choline, cholesterol and drug. Liposomes were evaluated for vesicle size, zeta potential, PDI, encapsulation efficiency, surface morphology and in vitro drug release study. Further the optimized formulation was evaluated for APAP-induced alterations in liver and kidney function tests in rats and histopathological studies. Results: The results were promising with a sustained drug release of 80% within 20hrs, optimized vesicle size of 276nm and 89% encapsulation efficiency. The animal studies demonstrated superior hepatoprotective effect compared to silybin solution. Conclusion: The silybin liposomes showed better in-vitro release & in-vivo hepatoprotection along with better animal activity & improvement in histopathological changes as compared to silybin.

Experimental Design Based Optimization and Ex Vivo Permeation of Desmopressin Acetate Loaded Elastic Liposomes Using Rat Skin

The study aimed to develop elastic-liposome-based transdermal delivery of desmopressin acetate for enhanced permeation to control enuresis, central diabetes insipidus, and traumatic injury. Elastic liposomes (ELs)-loaded desmopressin acetate was prepared, optimized, and evaluated for improved transdermal permeation profiles using rat skin. Full factorial design with independent factors (X1 for lipid and X2 for surfactant) at three levels was used against four responses (Y1, Y2, Y3, and Y4) (dependent variables). Formulations were characterized for vesicle size, polydispersity index (PDI), zeta potential, % entrapment efficiency (% EE), in vitro drug release, in vitro hemolysis potential, ex vivo drug permeation and drug deposition (DD), and ex vivo vesicle–skin interaction using scanning electron microscopy studies. The optimized formulation ODEL1 based on desirability function was found to have vesicle size, % EE, % DR, and permeation flux values of 118.7 nm, 78.9%, 75.1%, and 5.3 µg/h·cm2, respectively, which were close to predicted values. In vitro release profiles indicated slow and sustained delivery. Permeation flux values of ODEL1 and ODEL2 were 5.3 and 3.1 µg/h·cm2, respectively, which are 7.5- and 4.4-fold higher as compared to DS (0.71 µg/h·cm2). The obtained flux was relatively higher than the clinical target value of the drug for therapeutic efficacy. Moreover, the DD value of ODEL1 was significantly higher than ODEL2 and DS. Hemocompatibility study confirmed safety concerns. Finally, vesicle–skin interaction corroborated mechanistic views of permeation through rat skin. Conclusively, the transdermal delivery may be a suitable alternative to oral and nasal delivery to treat nocturnal enuresis, central diabetes insipidus, hemophilia A and von Willebrand’s disease, and any traumatic injuries.

Inhalable Spray Dried Pro-Liposome Powder Containing Budesonide for Pulmonary Delivery

Budesonide is an anti-asthmatic agent which is used to control the symptoms of asthma like bronchospasm, oedema. Drug delivered to lung through inhalation will provide systemic and local drug delivery at lower dose in chronic and acute diseases. Dry powder inhalers are the best choice for targeting the anti-asthmatic drugs through pulmonary route. The objective of the present study is to prepare inhalable lipid coated budesonide microparticles by spray drying method so effective delivery of budesonide to the lungs can be achieved. The microparticles in the form of dry powder were obtained by either spray drying liposomal drug suspension or lipid drug suspension. The liposomes were initially prepared by solvent evaporation method using Hydrogenated Soyabean Phosphatidylcholine and Cholesterol (1:1, 1:2, 2:1) as lipid carrier and then spray dried later with mannitol as bulking agent at different lipid to diluent ratio (1:1.25, 1:2.5 & 1:5). The liposomes and liposomal dry powder were evaluated for vesicle size, % entrapment efficiency, in vitro drug release studies, powder characteristics, aerosol performance and stability studies. The liposomes prepared showed vesicle size (2-8 µm), Entrapment efficiency (92.22%) at lipid: drug ratio of (2.5:1) and observed 80.41 % drug release in 24 hrs. Pro-liposomes prepared by spray drying of liposomal drug suspension (LSD1) showed emitted dose, mean mass aerodynamic diameter, geometric standard deviation and fine particle fraction of 99.01%, 3.12 µm, 1.78 and 43.5% along with good powder properties. The spray dried powder was found to be stable at 4 ± 2 °C & 65% ± 5 % RH. The inhalable microparticles containing Budesonide containing lipid dry powder was successfully prepared by spray drying method that showed good aerodynamic properties and stability with mannitol as diluent. The microparticles produced with this novel approach could deliver drug on target via inhalation route and also ease manufacture process at large scale in fewer production steps.

Development and Optimization of Proniosomal Gel Containing Etodolac: In-vitro, Ex-vivo and In-vivo Evaluation

Introduction: Etodolac is used in the treatment of acute pain and inflammation. It has low solubility because of high hydrophobicity and it is reported that upon oral administration shows gastric disturbances. This encourages the development of topical vesicular formulation. Method: In this work we used coacervation-phase separation method for the development of etodolac loaded vesicular system by using non-ionic surfactants, cholesterol and soya lecithin. Central composite design (rotatble) was used to optimize the concentrations of soy lecithin, surfactant and cholesterol. The prepared formulations were characterized by number of vesicles formed, vesicle size, zeta potential, entrapment efficiency, in-vitro permeation, ex-vivo permeation and anti-inflammatory study. Results: Etodolac was successfully entrapped in all formulations having efficiency in the range of 74.36% to 90.85%, which was more at 4 °C than room temperature. When hydrated with water; niosome in the range of 54 to 141 (per cubic mm) were spontaneously produced. The results of in-vitro diffusion study revealed that etodolac was released in the range of 71.86 to 97.16% over a period of 24 hrs. The average vesicle size of optimized formulation was found 211.9 nm with PDI of 0.5. The observed responses i.e. % encapsulation efficiency and drug release were 74.12 and 95.08 respectively. The zeta potential was -19.4mV revealed the stability of formulation which was further confirmed by no changes in drug content and drug release after stability studies. The % inhibition in paw volume was 40.52% and 43.61% for test and marketed proniosomal gel. Conclusion: Proniosomal gel formulation was stable and could enhance skin delivery of etodolac because of excellent permeation capability of vesicular system.