Sanaa A. El-Gizawy

Preparation and in vivo evaluation of insulin-loaded biodegradable nanoparticles prepared from diblock copolymers of PLGA and PEG

The aim of this study was to design a controlled release vehicle for insulin to preserve its stability and biological activity during fabrication and release. A modified, double emulsion, solvent evaporation, technique using homogenisation force optimised entrapment efficiency of insulin into biodegradable nanoparticles (NP) prepared from poly (DL-lactic-co-glycolic acid) (PLGA) and its PEGylated diblock copolymers. Formulation parameters (type of polymer and its concentration, stabiliser concentration and volume of internal aqueous phase) and physicochemical characteristics (size, zeta potential, encapsulation efficiency, in vitro release profiles and in vitro stability) were investigated. In vivo insulin sensitivity was tested by diet-induced type II diabetic mice. Bioactivity of insulin was studied using Swiss TO mice with streptozotocin-induced type I diabetic profile. Insulin-loaded NP were spherical and negatively charged with an average diameter of 200-400 nm. Insulin encapsulation efficiency increased significantly with increasing ratio of co-polymeric PEG. The internal aqueous phase volume had a significant impact on encapsulation efficiency, initial burst release and NP size. Optimised insulin NP formulated from 10% PEG-PLGA retained insulin integrity in vitro, insulin sensitivity in vivo and induced a sustained hypoglycaemic effect from 3h to 6 days in type I diabetic mice.

Nano-encapsulation of a novel anti-Ran-GTPase peptide for blockade of regulator of chromosome condensation 1 (RCC1) function in MDA-MB-231 breast cancer cells

Ran is a small ras-related GTPase and is highly expressed in aggressive breast carcinoma. Overexpression induces malignant transformation and drives metastatic growth. We have designed a novel series of anti-Ran-GTPase peptides, which prevents Ran hydrolysis and activation, and although they display effectiveness in silico, peptide activity is suboptimal in vitro due to reduced bioavailability and poor delivery. To overcome this drawback, we delivered an anti-Ran-GTPase peptide using encapsulation in PLGA-based nanoparticles (NP). Formulation variables within a double emulsion solvent evaporation technique were controlled to optimise physicochemical properties. NP were spherical and negatively charged with a mean diameter of 182-277nm. Peptide integrity and stability were maintained after encapsulation and release kinetics followed a sustained profile. We were interested in the relationship between cellular uptake and poly(ethylene glycol) (PEG) in the NP matrix, with results showing enhanced in vitro uptake with increasing PEG content. Peptide-loaded, pegylated (10% PEG)-PLGA NP induced significant cytotoxic and apoptotic effects in MDA-MB-231 breast cancer cells, with no evidence of similar effects in cells pulsed with free peptide. Western blot analysis showed that encapsulated peptide interfered with the proposed signal transduction pathway of the Ran gene. Our novel blockade peptide prevented Ran activation by blockage of regulator of chromosome condensation 1 (RCC1) following peptide release directly in the cytoplasm once endocytosis of the peptide-loaded nanoparticle has occurred. RCC1 blockage was effective only when a nanoparticulate delivery approach was adopted.

Characterisation and in vitro stability of low-dose, lidocaine-loaded poly(vinyl alcohol)-tetrahydroxyborate hydrogels

Poly(vinyl alcohol) hydrogels cross-linked with the tetrahydroxyborate anion possess textural and rheological properties that can be used as novel drug-loaded vehicles for application to traumatic wounds. However, addition of soluble drug substances causes concentration-dependent phase separation and rheological changes. The aim of this work was to investigate the effect of adding a local anaesthetic, but keeping the concentration low in an attempt to prevent these changes. Cross-linked hydrogels prepared from three grades of poly(vinyl alcohol) were characterised rheologically. Temperature sweep studies showed an elevated complex viscosity upon moving from 25°C to 80°C, which remained high for 48 h following completion of the cycle. Adhesion to model dermal surfaces achieved a maximum of 2.62 N cm(-2) and were greater than that observed to epidermal substrates, with a strong dependence on the rate of detachment used during testing. An optimised formulation (6% w/w PVA (31-50; 99) and 2% w/w THB) containing lidocaine hydrochloride loaded to an upper maximum concentration of 1.5% w/w was assessed for phase separation and drug crystallisation. After six months, crystallisation was present in formulations containing 0.7% and 1.5% lidocaine HCl. Changes in pH in response to increases in lidocaine loading were low. Drug release was shown to operate via a non-Fickian process for all three concentrations, with 60% occurring after approximately 24h. It can be concluded that using a low concentration of lidocaine hydrochloride in hydrogels based on poly(vinyl alcohol) will result in crystallisation. Furthermore, these hydrogels are unlikely to induce rapid anaesthesia due to the low loading and slow release kinetics.

Effect of poly(ethylene glycol) on insulin stability and cutaneous cell proliferation in vitro following cytoplasmic delivery of insulin-loaded nanoparticulate carriers – a potential topical wound management approach

Abstract We describe the development of a nanoparticulate system, with variation of poly(ethylene glycol) (PEG) content, capable of releasing therapeutic levels of bioactive insulin for extended periods of time. Recombinant human insulin was encapsulated in poly(d,l‐lactide‐co‐glycolide) nanoparticles, manufactured with variation in poly(ethylene glycol) content, and shown to be stable for 6 days using SDS‐PAGE, western blot and MALDI MS. To determine if insulin released from this sustained release matrix could stimulate migration of cell types normally active in dermal repair, a model wound was simulated by scratching confluent cultures of human keratinocytes (HaCaT) and fibroblasts (Hs27). Although free insulin was shown to have proliferative effect, closure of in vitro scratch fissures was significantly faster following administration of nano‐encapsulated insulin. This effect was more pronounced in HaCaT cells when compared to Hs27 cells. Variation in PEG content had the greatest effect on NP size, with a lesser influence on scratch closure times. Our work supports a particulate uptake mechanism that provides for intracellular insulin delivery, leading to enhanced cell proliferation. When placed into an appropriate topical delivery vehicle, such as a hydrogel, the extended and sustained topical administration of active insulin delivered from a nanoparticulate vehicle shows promise in promoting tissue healing. Graphical abstract Figure. No caption available.

Effect of poly(ethylene glycol) content and formulation parameters on particulate properties and intraperitoneal delivery of insulin from PLGA nanoparticles prepared using the double-emulsion evaporation procedure

Abstract Context: Size, encapsulation efficiency and stability affect the sustained release from nanoparticles containing protein-type drugs. Objectives: Insulin was used to evaluate effects of formulation parameters on minimizing diameter, maximizing encapsulation efficiency and preserving blood glucose control following intraperitoneal (IP) administration. Methods: Homogenization or sonication was used to incorporate insulin into poly(D,L-lactic-co-glycolic acid) (PLGA) nanoparticles with increasing poly(ethylene glycol) (PEG) content. Effects of polymer type, insulin/polymer loading ratio and stabilizer in the internal aqueous phase on physicochemical characteristics of NP, in vitro release and stability of encapsulated insulin were investigated. Entrapment efficiency and release were assessed by radioimmunoassay and bicinconnic acid protein assay, and stability was evaluated using SDS-PAGE. Bioactivity of insulin was assessed in streptozotocin-induced, insulin-deficient Type I diabetic mice. Results: Increasing polymeric PEG increased encapsulation efficiency, while the absence of internal stabilizer improved encapsulation and minimized burst release kinetics. Homogenization was shown to be superior to sonication, with NP fabricated from 10% PEG–PLGA having higher insulin encapsulation, lower burst release and better stability. Insulin-loaded NP maintained normoglycaemia for 24u2009h in diabetic mice following a single bolus, with no evidence of hypoglycemia. Conclusions: Insulin-loaded NP prepared from 10% PEG–PLGA possessed therapeutically useful encapsulation and release kinetics when delivered by the IP route.

Polymeric nano-encapsulation of 5-fluorouracil enhances anti-cancer activity and ameliorates side effects in solid Ehrlich Carcinoma-bearing mice

Biodegradable PLGA nanoparticles, loaded with 5-fluorouracil (5FU), were prepared using a double emulsion method and characterised in terms of mean diameter, zeta potential, entrapment efficiency and in vitro release. Poly (vinyl alcohol) was used to modify both internal and external aqueous phases and shown have a significant effect on nanoparticulate size, encapsulation efficiency and the initial burst release. Addition of poly (ethylene glycol) to the particle matrix, as part of the polymeric backbone, improved significantly the encapsulation efficiency. 5FU-loaded NPs were spherical in shape and negatively charged with a size range of 185-350u202fnm. Biological evaluation was performed in vivo using a solid Ehrlich carcinoma (SEC) murine model. An optimised 5FU-loaded formulation containing PEG as part of a block copolymer induced a pronounced reduction in tumour volume and tumour weight, together with an improved percentage tumour growth inhibition. Drug-loaded nanoparticles showed no significant toxicity or associated changes on liver and kidney function in tested animals, whereas increased alanine aminotransferase, aspartate aminotransferase and serum creatinine were observed in animals treated with free 5FU. Histopathological examination demonstrated enhanced cytotoxic action of 5FU-loaded nanoparticles when compared to the free drug. Based on these findings, it was concluded that nano-encapsulation of 5FU using PEGylated PLGA improved encapsulation and sustained in vitro release. This leads to increased anti-tumour efficacy against SEC, with a reduction in adverse effects.

Layer-by-layer gelatin/chondroitin quantum dots-based nanotheranostics: combined rapamycin/celecoxib delivery and cancer imaging

AIMnNanotheranostics consisting of highly-fluorescent quantum dots coupled with gelatin/chondroitin layer-by-layer assembled nanocapsules were developed.nnnMATERIALS & METHODSnxa0The hydrophobic drugs celecoxib (CXB) and rapamycin (RAP) were co-loaded into the oily core of nanocapsules (NCs) to enable synergistic growth inhibition of breast cancer cells. To overcome the nonspecific binding of actively targeted CS-NCs with normal cells, a matrix metalloproteinase (MMP-2)-degradable cationic gelatin layer was electrostatically deposited onto the surface of the negatively-charged CS-NCs.nnnRESULTSnThe prepared nanocarriers displayed strong fluorescence which enabled tracing their internalization into cancer cells. An enhanced cytotoxicity of the NCs against breast cancer cells was demonstrated. In vivo, the nanoplatforms displayed superior antitumor efficacy as well as nonimmunogenic response.nnnCONCLUSIONnTherefore, these multifunctional nanoplatforms could be used as potential cancer theranostics.

Lactoferrin-tagged quantum dots-based theranostic nanocapsules for combined COX-2 inhibitor/herbal therapy of breast cancer

AIMnHerein, tumor-targeted quantum dots (QDs)-based theranostic nanocapsules (NCs) coloaded with celecoxib and honokiol were developed. Materialsxa0& methodology: The anionic CD44-targeting chondroitin sulfate and cationic low density lipoprotein (LDL)-targeting lactoferrin (LF) were sequentially assembled onto the surface of the positively charged oily core. As an imaging probe, highly fluorescent mercaptopropionic acid-capped cadmium telluride QDs were coupled to LF.nnnRESULTSnIn vitro, fluorescence of QDs was quenched (OFF state) due to combined electron/energy transfer-mediated processes involving LF. After intracellular uptake of NCs, fluorescence was restored (ON state), thus enabled tracing their internalization. The NCs demonstrated enhanced cytotoxicity against breast cancer cells as well as superior in vivo antitumor efficacy.nnnCONCLUSIONnWe propose these multifunctional nanotheranostics for imaging and targeted therapy of breast cancer.

Regional difference in intestinal drug absorption as a measure for the potential effect of P-glycoprotein efflux transporters

The aim of this research was to assess regional difference in the intestinal absorption of ranitidine HCl as an indicator for the potential effect of P‐glycoprotein (P‐gp) efflux transporters.

Development and optimization of a novel drug free nanolipid vesicular system for treatment of osteoarthritis

Abstract Objective: The goal of this study is to improve the transdermal delivery of phosphatidylcholine (PC) via constructing a novel nanolipid vesicular system (NLVS) with high level of permeability through the stratum corneum (SC). Significance: In our study, a novel drug free NLVS was developed. The system depends on PC boundary cartilage lubrication to relieve osteoarthritic pain without developing gastrointestinal problems associated with anti-inflammatory drug. Materials and methods: A full two-level (23) factorial design is applied to optimize the quality of the prepared NLVS. The selected independent variables are the concentration of PC, the concentration of edge activator (EA), and EA type. The developed NLVS was evaluated for in-vitro, ex-vivo as well as in-vivo efficacy in rat animal model. Results: Based on the factorial design, the selected formulation variables significantly affect the tested responses. The prepared NLV formulations have a particle size (PS)in the range of 10.34 to 496.3u2009nm, polydispersity index (PdI) values less than one, and negative zeta potential (ZP) range of −1.42 to −32.01u2009mV. In-vitro and ex-vivo study results reveal that the designed NLVS is effective in sustaining PC release and enhancing its transdermal permeation over 24u2009h. The optimal permeation flux through ex-vivo study is 0.415u2009mg/cm2/h following zero-order kinetics. Moreover, in-vivo study of the optimized formulations demonstrated remarkable reduction in inflammatory mediators associated with osteoarthritis (OA). Conclusion: The results indicate that the optimized drug free NLVS significantly augment transdermal delivery of PC and have a potential role in treatment of OA without the risk of systemic side effects.