Formulation and Evaluation of Pravastatin Sodium-Loaded PLGA Nanoparticles: In vitro-in vivo Studies Assessment.

Purpose: Pravastatin sodium (PVS) is a hypolipidemic drug which suffers from extensive first-pass metabolism and short half-life. Poly(d,l-lactide-co-glycolide) (PLGA) is considered a promising carrier to improve its hypolipidemic and hepatoprotective activities. Methods: PVS-loaded PLGA nanoparticles (PVS-PLGA-NPs) were prepared by double emulsion method using a full 32 factorial design. The in vitro release and the physical stability studies of the optimized PVS-PLGA-NPs (F5) were performed. Finally, both hypolipidemic and hepatoprotective activities of the optimized F5 NPs were studied and compared to PVS solution. Results: All the studied physical parameters of the prepared NPs were found in the accepted range. The particle size (PS) ranged from 90 ± 0.125 nm to 179.33 ± 4.509 nm, the poly dispersity index (PDI) ranged from 0.121 ± 0.018 to 0.158 ± 0.014. The optimized NPs (F5) have the highest entrapment efficiency (EE%) (51.7 ± 5%), reasonable PS (168.4 ± 2.506 nm) as well as reasonable zeta potential (ZP) (-28.3 ± 1.18mv). Solid-state characterization indicated that PVS is well entrapped into NPs. All NPs have distinct spherical shape with smooth surface. The prepared NPs showed a controlled release profile. F5 showed good stability at 4 ± 2°C during the whole storage period of 3 months. In vivo study and histopathological examination indicated that F5 NPs showed significant increase in PVS hypolipidemic as well as hepatoprotective activity compared to PVS solution. Conclusion: The PVS-PLGA-NPs could be considered a promising model to evade the first-pass effect and showed improvement in the hypolipidemic and hepatoprotective activities compared to PVS solution.

Development and optimization of transethosomal gel of apigenin for topical delivery: In-vitro, ex-vivo and cell line assessment.

The main aim of this study was to optimize the transethosomes of apigenin formulated by the thin film hydration method using surfactant Span 80. Response surface Box-Behnken design with three levels of three factors was used to design and optimize the formulations. The prepared transethosomal formulations were characterized for entrapment efficiency, vesicle size, and flux to obtain the optimized formulation batch. The optimized batch was further incorporated into the gel and characterized for the in-vitro, ex-vivo, and cytotoxic studies. The result showed the optimized transethosomes were smooth, nanosized, unilamellar, and spherical with an entrapment efficiency of 78.75 ± 3.14 %, a vesicle size of 108.75 ± 2.31 nm, and a flux of 4.10 ± 0.63 µg/cm2/h. In-vitro cumulative drug release of transethosomal gel of apigenin (TEL gel) and the conventional gel was 92.25 ± 3.5 % and 53.40 ± 3.10 %, respectively, after 24 h study. Ex-vivo permeation of TEL gel and conventional gel showed 86.20 ± 3.60 % and 51.20 ± 3.20 % permeation of apigenin at 24 h, respectively. A cytotoxic study confirmed that TEL gel significantly reduces cell viability compared to conventional gel. The results suggested that topical application of apigenin transethosomal gel may be a better treatment strategy for skin cancer because of the prolonged sustained release of the drug and the better permeability of apigenin through the skin.

Dermal Delivery of Lipid Nanoparticles: Effects on Skin and Assessment of Absorption and Safety.

During the recent decades, dermal delivery has achieved visible popularity mainly due to the increase of chronic skin diseases and the demand for targeted delivery and patient compliance. Dermal delivery provides an attractive alternative to oral drug delivery, promoting the drug application directly at the site of action, resulting in higher localized drug concentration with reduced systemic drug exposure. Among several types of drug delivery systems used in dermal delivery are the lipid nanoparticles, which include solid lipid nanoparticles (SLNs) and nanostructured lipid carriers (NLCs). These lipid nanocarriers have attracted great interest and have been intensively studied for their use in dermal applications. Lipid nanoparticles increase the transport of active compounds through the skin by improving drug solubilization in the formulation, drug partitioning into the skin, and fluidizing skin lipids. Moreover, these nanocarriers are composed of biologically active and biodegradable lipids that show less toxicity and offer many favorable attributes such as adhesiveness, occlusion, skin hydration, lubrication, smoothness, skin penetration enhancement, modified release, improvement of formulation appearance providing a whitening effect, and offering protection of actives against degradation.This chapter focuses on the effects of lipid nanoparticles in dermal delivery, on the types of active compounds that are used in their formulation and application, some aspects related to their possible toxicity, and a description of the most commonly used techniques for the evaluation of drug absorption on the skin.

Infrared and Raman spectroscopy for purity assessment of extracellular vesicles.

Extracellular vesicles (EVs) are a complex and heterogeneous population of nanoparticles involved in cell-to-cell communication. Recently, numerous studies have indicated the potential of EVs as therapeutic agents, drug carriers and diagnostic tools. However, the results of these studies are often difficult to evaluate, since different characterization methods are used to assess the purity, physical and biochemical characteristics of the EV samples. In this study, we compared four methods for the EV sample characterization and purity assessment: i) the particle-to-protein ratio based on particle analyses with nanoparticle tracking and protein concentration by bicinchoninic acid assay, ii) Western Blot analysis for specific EV biomarkers, iii) two spectroscopic lipid-to-protein ratios by either the attenuated total reflection Fourier transform infrared (ATR-FTIR) or Raman spectroscopy. The results confirm the value of Raman and ATR-FTIR spectroscopy as robust, fast and operator independent tools that require only a few microliters of EV sample. We propose that the spectroscopic lipid-to-protein (Li/Pr) ratios are reliable parameters for the purity assessment of EV preparations. Moreover, apart from determining protein concentrations, we show that ATR-FTIR spectroscopy can also be used for indirect measurements of EV concentrations. Nevertheless, the Li/Pr ratios do not represent full characterization of the EV preparations. For a complete characterization of selected EV preparations, we recommend also additional use of particle size distribution and EV biomarker analysis.

Assessment of a Novel Vitamin D3 Formulation with Nanostructured Lipid Carriers for Transdermal Delivery.

OBJECTIVE: Develop and assess a transdermal emulsion loaded with nanostructured lipid carriers for vitamin D3 supplementation. METHODS: Vitamin D3 loaded nanostructured lipid carriers, produced via high shear homogenization and ultrasonication, were assessed for their particle size, distribution, morphology, zeta potential, entrapment efficiency, and cytotoxicity. They were incorporated into a transdermal vehicle, and the stability and ex vivo permeation were evaluated. RESULTS: Spherical nanoparticles were developed with a particle size of 192.5 nm, a polydispersity index of 0.13, a zeta potential of -29.0 mV, and an entrapment efficiency of 99.75%. They were stable (particle size and distribution) for 15 days when stored in a refrigerator, and for 30 days at room temperature and 32°C. The nanoparticles decreased the drug cytotoxicity against fibroblasts, as shown by IC50 (nanoparticle: 32.48 µg mL-1 vitamin D3: 16.73 µg mL-1). The emulsion loaded with nanoparticles minimized the degradation of vitamin D3 when compared with the nanoparticle dispersion. Additionally, the emulsion provided the skin permeation of vitamin D3 following the recommended daily allowance. CONCLUSION: To the best of our knowledge, this is the first study to use nanostructured lipid carriers for transdermal delivery of vitamin D. The developed formulation is a promising strategy to overcome the vitamin D3 variable oral bioavailability. It also represents a comfortable route of administration; thus it could be beneficial for patients and clinicians. However, further studies are needed to allow the permeation of larger amounts of vitamin D3, and the combination of these nanoparticles with microneedles would be interesting.

Topical Delivery of Rapamycin by Means of Microenvironment-Sensitive Core-Multi-Shell Nanocarriers: Assessment of Anti-Inflammatory Activity in an ex vivo Skin/T Cell Co-Culture Model.

INTRODUCTION: Rapamycin (Rapa) is an immunosuppressive macrolide that inhibits the mechanistic target of rapamycin (mTOR) activity. Thanks to its anti-proliferative effects towards different cell types, including keratinocytes and T cells, Rapa shows promise in the treatment of skin diseases characterized by cell hyperproliferation. However, Rapa skin penetration is limited due to its lipophilic nature (log P = 4.3) and high molecular weight (MW = 914 g/mol). In previous studies, new microenvironment-sensitive core multishell (CMS) nanocarriers capable of sensing the redox state of inflamed skin were developed as more efficient and selective vehicles for macrolide delivery to inflamed skin. METHODS: In this study, we tested such redox-sensitive CMS nanocarriers using an inflammatory skin model based on human skin explants co-cultured with Jurkat T cells. Serine protease (SP) was applied on skin surface to induce skin barrier impairment and oxidative stress, whereas phytohaemagglutinin (PHA), IL-17A, and IL-22 were used to activate Jurkat cells. Activation markers, such as CD45 and CD69, phosphorylated ribosomal protein S6 (pRP-S6), and IL-2 release were monitored in activated T cells, whereas pro-inflammatory cytokines were measured in skin extracts and culture medium. RESULTS: We found that alteration of skin barrier proteins corneodesmosin (CDSN), occludin (Occl), and zonula occludens-1 (ZO-1) as well as oxidation-induced decrease of free thiol groups occurred upon SP-treatment. All Rapa formulations exerted inhibitory effects on T cells after penetration across ex vivo skin. No effects on skin inflammatory markers were detected. The superiority of the oxidative-sensitive CMS nanocarriers over the other formulations was observed with regard to drug delivery as well as downregulation of IL-2 release. CONCLUSION: Overall, our results demonstrate that nanocarriers addressing features of diseased skin are promising approaches to improve the topical delivery of macrolide drugs.

Formulation development and in vitro-in vivo assessment of protransfersomal gel of anti-resorptive drug in osteoporosis treatment.

Osteoporosis is a major cause of morbidity, mortality, and economic burden worldwide. Despite being an effective in combating the bone-deteriorating disorders, bisphosphonates have several shortcomings including poor and variable bioavailability, low permeability, high toxicity, etc. In this study, we developed and optimized protransfersome formulation for the drug risedronate sodium (RIS-Na) with the goal of enhancing its bioavailability and hence patient compliance. Phase separation coacervation technique was utilized for development of optimized formulation. Optimization was achieved by using three-factor, three-level Box-Behnken design combined with Response Surface Methodology (RSM). This enabled us to decipher the effect of 3 independent variables (Phospholipid, Tween-80 and Sodium Deoxycholate) on three dependent parameters (entrapment efficiency, vesicle size and transdermal flux). Optimized formulation was further evaluated for pharmacokinetic and pharmacodynamic parameters. Smooth, spherical protransfersomes with a size of 260 ± 18 nm, having entrapment efficiency and flux of 80.4 ± 4.90% and 8.41 ± 0.148 µg/cm2/h, respectively were prepared. Ex vivo studies revealed a shorter lag time of 1.21 ± 0.18 h and higher flux associated with transdermal formulation. CLSM analysis further revealed better drug penetration (220 µm) through the skin in case of protransfersomes as compared to drug solution (72 µm). Additionally, biomechanical, biochemical, and histo-pathological studies further validated the results. Thus, it was concluded that protransfersome formulation has a great potential in providing better therapeutic efficacy of risedronate than its conventional counterpart.

Pharmaceutical Technology Innovation Strategy Based on the Function of Blood Transport Proteins as DDS Carriers for the Treatment of Intractable Disorders and Cancer.

Blood transport proteins are biogenic molecules with unique and interesting inherent characteristics that make up living organisms. As the utilization of their inherent characteristics can be a groundbreaking strategy to resolve and improve several clinical problems, attempts have been made to develop pharmaceutical and biomedical preparations based on blood transport proteins for the treatment and diagnosis of disorders. Among various blood transport proteins, we focus on the immense potential of hemoglobin and albumin to serve as carriers of biomedical gases (oxygen and carbon monoxide) and anticancer agents (low-molecular compounds and antisense oligodeoxynucleotides), respectively, for the development of innovative drug delivery systems (DDS) to treat intractable disorders and solid cancers. In this review, I introduce the pharmaceutical technology, strategies, and application of DDS carriers that have been designed on the basis of the structure and function of hemoglobin and albumin. In addition, the prospect of using hemoglobin and albumin as materials for DDS carriers is discussed.

Comparative assessment of in vitro/in vivo performances of orodispersible electrospun and casting films containing rizatriptan benzoate.

The electrospinning process is a promising approach to produce various drug-loaded orodispersible films (ODFs) with a rapid onset of their actions. However, there is only limited number of studies comparing the pharmacological performances of electrospun ODFs (eODFs) with traditional casting films (CFs). In this study, rizatriptan benzoate (RB), a pain relieving agent was formulated with PVP and PVA into ODFs using electrospinning and casting methods. The ODFs were subsequently characterized with respect to their morphology, solid state properties and mechanical characteristics. The uniformity of the dosage units, disintegration behavior and dissolution patterns of the ODFs were also evaluated prior to the pharmacokinetic study. The obtained CFs and eODFs were semitransparent and white in appearance, respectively. The scanning electron microscopy revealed that the eODFs contained nanoporous structure, while the CFs showed no observable pores. RB was amorphously dispersed in both these films without drug-polymer interactions. The uniformity of dosage units for both eODFs and CFs was complied with European Pharmacopeia. As compared to the CFs, the eODFs were more flexible and lesser rigid in nature and showed faster disintegration and dissolution rates. In addition, the eODFs exhibited a higher bioavailability with a shorter Tmax relative to the CFs and commercial RB tablets. This study demonstrated that eODFs were superior to CFs with respect to in vivo pharmacological effects, which could be attributed to the submicron structure of eODFs obtained through the electrospinning process.

Oxiconazole nitrate solid lipid nanoparticles: formulation, in-vitro characterization and clinical assessment of an analogous loaded carbopol gel.

The aim of the present work was to develop a promising drug delivery system of oxiconazole nitrate-loaded solid lipid nanoparticles (SLNs) topical gel to enhance the drug effectiveness for the treatment of Tinea infection. SLNs were prepared by emulsification-solvent evaporation method. Particle size and entrapment efficiency of the prepared SLNs were investigated. An appropriate formulation was selected and examined for morphology and physicochemical characterization adopting Scanning electron microscope and Differential scanning colorimetry. In-vitro drug release was also investigated. The selected SLNs were loaded into 1% Carbopol 934 gel that was investigated for homogeneity, pH, grittiness, spreadability, viscosity and in vitro drug release. Clinical study for the developed gel system compared to the corresponding marketed product was conducted on 28 patients. The results revealed that the prepared oxiconazole nitrate SLNs had drug entrapment efficiency ranging from 41.34% to 75.07% and zeta potential lying between -13 and -50. Physicochemical characterization revealed a decrease in the drug crystallinity in the prepared SLNs. The gel formulation showed appropriate physical characteristics and sustained in-vitro drug release. Clinical study for the prepared oxiconazole nitrate SLNs gel showed significantly less side effects, better patient satisfaction and superior clinical improvement compared with the corresponding marketed product.

Fabrication of rosuvastatin-loaded polymeric nanocapsules: a promising modality for treating hepatic cancer delineated by apoptotic and cell cycle arrest assessment.

Nanotechnology has provided several advantages for the treatment of cancer. Polymeric nanocapsules (PNCs) were proven promising in the treatment of different cancer types, such as hepatic cancer. Meanwhile, the exploration of novel indications of old molecules with the purpose of cancer treatment has been widely reported. Among the promising therapeutic moieties, rosuvastatin (RV) was delineated as a potential anticancer drug. Hence, the target of the presented manuscript was to develop PNCs loaded with RV to overcome its delivery challenges and augment its anticancer activity. RV PNCs were fabricated by the nanoprecipitation method using poly-lactide-co-glycolide (PLGA) polymer, and were characterized for the size, polydispersity index (PDI), charge, entrapment efficiency EE%, in vitro release, stability, and morphology. Furthermore, their anticancer activity was tested on HepG2 cells using MTT assay, followed by elucidating the cytotoxic activity using flow cytometry. Results showed that RV PNCs displayed particle size ranging from 186 to 239 nm, average PDI, and negative zeta potential with sufficient stability for 3 months. PNCs were able to load RV at high EE% reaching 82.6% and sustain its release for eight hours. RV PNCs were superior in their anticancer activity on HepG2 cells, as delineated from the viability study and further elucidated by enhanced apoptosis in addition to cell cycle arrest at G2/M phase, suggesting their promise in treatment of hepatic cancer.

In vitro and in vivo assessment of delivery of hydrophobic molecules and plasmid DNAs with PEO-PPO-PEO polymeric micelles on cornea.

The stability and bio-distribution of genes or drug complexes with poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO, Pluronic F-68) polymeric micelles (PM) are essential for an effective nanosized PM delivery system. We used Förster resonance energy transfer (FRET) pairs with PM and measured the FRET ratio to assess the stability of PM in vitro and in vivo on the cornea. The FRET ratio reached a plateau at 0.8 with 3% PM. Differential scanning calorimetry measurement confirmed the complex formation of FRET pairs with PM. Confocal imaging with the fluorophores fluorescein isothiocyanate isomer I (FITC) and rhodamine B base (RhB) also showed the occurrence of FRET pairs in vitro. The fluorophores were mixed with 3% PM solution or the FITC-labeled PEO-PPO-PEO polymers (FITC-P) were mixed with RhB-labeled plasmids (RhB-DNA). In addition, the in vitro corneal permeation of FRET pair complexes with PM reached a 0.8 FRET ratio. One hour after eye drop administration, FRET pairs colocalized in the cytoplasm, and surrounded and entered the nuclei of cells in the cornea, and the polymers were located in the corneal epithelial layers, as detected through anti-PEG immunohistochemistry. Furthermore, fluorescence colocalization in the cytoplasm and cell nucleus of the corneal epithelium was confirmed in tissues where RhB or RhB-DNA complexed with FITC-P was found to accumulate. We demonstrate that at a concentration of 3%, PM can encapsulate FRET pairs or RhB-DNA and retain their integrity within the cornea 1 h after administration, suggesting the feasibility and stability of PEO-PPO-PEO polymers as a vehicle for drug delivery.

Formulation and in vivo assessment of terconazole-loaded polymeric mixed micelles enriched with Cremophor EL as dual functioning mediator for augmenting physical stability and skin delivery.

The aim of the current study was to formulate terconazole (TCZ) loaded polymeric mixed micelles (PMMs) incorporating Cremophor EL as a stabilizer and a penetration enhancer. A 23 full factorial design was performed using Design-Expert® software for the optimization of the PMMs which were formulated using Pluronic P123 and Pluronic F127 together with Cremophor EL. To confirm the role of Cremophor EL, PMMs formulation lacking Cremophor EL was prepared for the purpose of comparison. Results showed that the optimal PMMs formulation (F7, where the ratio of total Pluronics to drug was 40:1, the weight ratio of Pluronic P123 to Pluronic F127 was 4:1, and the percentage of Cremophor EL in aqueous phase was 5%) had a high micellar incorporation efficiency (92.98 ± 0.40%) and a very small micellar size (33.23 ± 8.00 nm). Transmission electron microscopy revealed that PMMs possess spherical shape and good dispersibility. The optimal PMMs exhibited superior physical stability when compared with the PMMs formulation of the same composition but lacking Cremophor EL. Ex vivo studies demonstrated that the optimal PMMs formula markedly improved the dermal TCZ delivery compared to PMMs lacking Cremophor EL and TCZ suspension. In addition, it was found that the optimal PMMs exhibited a greater extent of TCZ deposition in the rat dorsal skin relative to TCZ suspension. Moreover, histopathological studies revealed the safety of the optimal PMMs upon topical application to rats. Consequently, PMMs enriched with Cremophor EL, as a stable nano-system, could be promising for the skin delivery of TCZ.

Effect of amphiphilic graft co-polymer-carrier on physical stability of bosentan nanocomposite: Assessment of solubility, dissolution and bioavailability.

Bosentan is a dual endothelin receptor antagonist used in the treatment of pulmonary arterial hypertension (PAH). But the solubility and bioavailability of this drug are poor, which has restricted the design and development of dosage forms for efficient and successful therapy. The present study was carried out to develop nanocomposites using an amphiphilic graft co-polymer (Soluplus®) as a carrier to enhance the solubility and bioavailability of bosentan. The graft co-polymer-based nanocomposite formulation was prepared using the single-emulsion technique. The nanocomposite was characterised in terms of particle size analysis, solubility, percentage entrapment efficiency, drug-loading capacity, surface morphology, drug content, in vitro dissolution, stability and bioavailability. FT-IR study revealed that there was no interaction between the drug and Soluplus®. DSC analysis of the nanocomposite formulation confirmed that the bosentan was completely encapsulated within a Soluplus®. XRD analysis showed that the drug was converted to an amorphous form irreversibly. SEM images showed that the particles were of size 96-129µm and had slightly smooth to rough textured surface. TEM analysis indicated that the diameters of the prepared bosentan nanocomposite after dispersion in distilled water were 13.69-96.78nm. Statistically significant increases in the solubility, dissolution and bioavailability of the drug were observed. It was confirmed that the use of a graft co-polymer carrier-based nanocomposite formulation is a good approach for efficient delivery of bosentan, the solubility and bioavailability being increased manifold.

Preparation and evaluation of novel chitosan: gelrite ocular system containing besifloxacin for topical treatment of bacterial conjunctivitis: scintigraphy, ocular irritation and retention assessment.

This study was aimed to prepare, characterize and evaluate in situ gel formulations based on a blend of chitosan (CS), polyvinyl alcohol (PVA) and gellan gum (Gelrite™) for a sustained ocular delivery of besifloxacin (BSF). The developed formulations were evaluated for physicochemical properties, gelation time (Tsol-gel), rheological behaviour, antimicrobial efficacy, pharmacokinetic assessment, gamma scintigraphy study and ocular irritation. The results showed BSF sol-gel system were found to be sensitive enough which underwent instantaneous phase transition upon getting physiological stimulation. The ex vivo permeation experiments indicated that the developed formulation was able to enhance the retention of BSF at corneal surface. The HET-CAM confirmed the non-irritancy of developed formulation and also demonstrated the ability of ocular protection against strongly irritant substances. The results of gamma scintigraphy study revealed the higher concentration of drug retains at the corneal surface. In addition, optimized BSF sol-gel system showed enhanced anti-bacterial activity compared to BSF suspension.

Lipid drug conjugate nanoparticle as a potential nanocarrier for the oral delivery of pemetrexed diacid: Formulation design, characterization, ex vivo, and in vivo assessment.

The present work was to develop lipid drug conjugated (LDC) nanoparticles for the potential oral delivery of pemetrexed diacid (PTX) and evaluation of its in vitro, ex vivo and in vivo potentials. The LDC was prepared by salt formation of PTX with stearic acid and followed by cold homogenization technique to produce the LDC nanoparticles. FTIR analysis of LDC proved the presence of amide bond in LDC powder indicating the conjugation between drug and lipid. LDC nanoparticles was found to have particle size 121.9±1.85nm and zeta potential -51.6mV±1.23 and entrapment efficiency 81.0±0.89%. TEM images revealed spherical morphology and were in corroboration with particle size measurements. Ex vivo gut permeation studies revealed a very good enhancement in permeation of drug present in the LDC as compared to plain drug solution and were confirmed by CLSM. MTT assay conformed significant% toxicity at the end of 24h and 48h. Furthermore, the AUC0-24 of PTX from the optimized LDC nanoparticels was found to be 4.22 folds higher than that from PTX suspension on oral administration. Thus, LDC has high potential for the oral delivery of PTX in cancer therapy and future prospects for the industrial purpose.

Transdermal anti-inflammatory activity of bilayer film containing olive compound hydroxytyrosol: physical assessment, in vivo dermal safety and efficacy study in Freund's adjuvant-induced arthritic rat model.

Previous studies have shown that hydroxytyrosol (HT) can be a potential alternative therapeutic agent for the treatment of rheumatoid arthritis (RA). However, HT is extensively metabolized following oral administration, which leads to formulating HT in a topical vehicle to prolong drug action as well as to provide a localized effect. Hidrox-6 is a freeze-dried powder derived from fresh olives and contains a high amount of HT (∼3%) and other polyphenols. Alginate bilayer films containing 5% and 10% Hidrox-6 were formulated. The films were characterized with respect to their physical, morphology, rheological properties; drug content uniformity; and in vitro drug release. Acute dermal irritancy tests and a skin sensitization study were carried out in rats. An efficacy study of the bilayer films for RA was conducted using Freund's adjuvant-induced polyarthritis rats. Animal data showed that the bilayer film formulations did not cause skin irritancy. The efficacy in vivo results showed that the Hidrox-6 bilayer films lowered the arthritic scores, paw and ankle circumference, serum IL-6 level and cumulative histological scores compared with those measured for controls. The topical Hidrox-6 bilayer films improve synovitis and inflammatory symptoms in RA and can be a potential alternative to oral RA therapy.

Effect of variation in hemorheology between human and animal blood on the binding efficacy of vascular-targeted carriers.

Animal models are extensively used to evaluate the in vivo functionality of novel drug delivery systems (DDS). However, many variations likely exist in vivo between the animals and human physiological environment that significantly alter results obtained with animal models relative to human system. To date, it is not clear if the variation in hemorheology and hemodynamics between common animal and human models affect the functionality of DDS. This study investigates the role of hemorheology of humans and various animal models in dictating the binding efficiency of model vascular-targeted carriers (VTCs) to the wall in physiological blood flows. Specifically, the adhesion of sLe(A)-coated nano- and micro-spheres to inflamed endothelial cells monolayers were conducted via a parallel plate flow chamber assay with steady and disturbed red blood cells (RBCs)-in-buffer and whole blood flows of common animal models. Our results suggest that the ratio of carrier size to RBC size dictate particle binding in blood flow. Additionally, the presence of white blood cells affects the trend of particle adhesion depending on the animal species. Overall, this work sheds light on some deviation in VTC vascular wall interaction results obtained with in vivo animal experimentation from expected outcome and efficiency in vivo in human.

Optimized luciferase assay for cell-penetrating peptide-mediated delivery of short oligonucleotides.

An improved assay for screening for the intracellular delivery efficacy of short oligonucleotides using cell-penetrating peptides is suggested. This assay is an improvement over previous assays that use luciferase reporters for cell-penetrating peptides because it has been scaled up from a 24-well format to a 96-well format and no longer relies on a luciferin reagent that has been commercially sourced. In addition, the homemade luciferin reagent is useful in multiple cell lines and in different assays that rely on altering the expression of luciferase. To establish a new protocol, the composition of the luciferin reagent was optimized for both signal strength and longevity by multiple two-factorial experiments varying the concentrations of adenosine triphosphate, luciferin, coenzyme A, and dithiothreitol. In addition, the optimal conditions with respect to cell number and time of transfection for both short interfering RNA (siRNA) and splice-correcting oligonucleotides (SCOs) are established. Optimal transfection of siRNA and SCOs was achieved using the reverse transfection method where the oligonucleotide complexes are already present in the wells before the cells are plated. Z' scores were 0.73 for the siRNA assay and 0.71 for the SCO assay, indicating that both assays are suitable for high-throughput screening.

Box-Behnken study design for optimization of bicalutamide-loaded nanostructured lipid carrier: stability assessment.

Bicalutamide (BCM) is an anti-androgen drug used to treat prostate cancer. In this study, nanostructured lipid carriers (NLCs) were chosen as a carrier for delivery of BCM using Box-Behnken (BB) design for optimizing various quality attributes such as particle size and entrapment efficiency which is very critical for efficient drug delivery and high therapeutic efficacy. Stability of formulated NLCs was assessed with respect to storage stability, pH stability, hemolysis, protein stability, serum protein stability and accelerated stability. Hot high-pressure homogenizer was utilized for formulation of BCM-loaded NLCs. In BB response surface methodology, total lipid, % liquid lipid and % soya lecithin was selected as independent variable and particle size and %EE as dependent variables. Scanning electron microscopy (SEM) was done for morphological study of NLCs. Differential scanning calorimeter and X-ray diffraction study were used to study crystalline and amorphous behavior. Analysis of design space showed that process was robust with the particle size less than 200 nm and EE up to 78%. Results of stability studies showed stability of carrier in various storage conditions and in different pH condition. From all the above study, it can be concluded that NLCs may be suitable carrier for the delivery of BCM with respect to stability and quality attributes.