Research paper
Novel inulin-based mucoadhesive micelles loaded with corticosteroids as potential transcorneal permeation enhancers

https://doi.org/10.1016/j.ejpb.2017.05.005Get rights and content

Abstract

In this work a new copolymer of inulin (INU) derivatized with ethylendiamine (EDA) and retinoic acid (RA), named INU-EDA-RA, was synthetized, characterized and employed to produce micelles as carriers for topical administration of corticosteroids for the potential treatment of diseases of posterior eye segment. Spectroscopic analysis confirmed a molar derivatization degree of 11.30 and 4.30% in EDA and RA, respectively. INU-EDA-RA micelles are capable of strong mucoadhesive interactions which result time-independent and stable over time but concentration depending. Moreover micelles are able to encapsulate efficiently from 3 to 13% (w/w) of lipophilic drugs, as dexamethasone, triamcinolone and triamcinolone acetonide. Drug loaded micelles are stable for three months when stored as freeze-dried powders and able to release high amount of drug when compared to drug dissolution profiles from suspensions. Moreover, drug loaded micelles are compatible with different ocular cell lines that are also able to internalize fluorescent micelles. Finally, drug loaded micelles enhance drug fluxes and permeability coefficients across corneal epithelial cells, thus reducing drug loss due to retention inside the cells.

Introduction

Vascular degenerative disorders of the posterior eye segment, such as age-related macular degeneration (ARMD), diabetic retinopathy (DR) and diabetic macular edema (DME), are the main cause of visual impairment and blindness worldwide [1], [2], [3].

Today, intravitreal injections of vascular endothelial growth factor (VEGF) inhibitors are the standard of care for patients with retinal neovascularization. The “intravitreal operative medication” (IVOM) is the main technique used because it yields effective local drug concentrations while markedly reducing systemic side effects. However, IVOM is a painful and invasive procedure that must be performed under sterile conditions. Moreover, intraocular injections increase the risk of haemorrhages, endophthalmitis, retinal detachment, cataracts and other complications [4]. In addition, anti-VEGF drugs only bind to the synthesized VEGF and inhibit its action, while corticosteroids exert a complex and multi-factorial anti-inflammatory, anti-edematous and anti-angiogenetic mechanism of action. In particular, they decrease VEGF synthesis by destabilizing VEGF mRNA, inhibit cellular proliferation, stabilize the blood-retinal barrier, increase the production of tight junctional proteins, regulate the expression and distribution of ion channels and water channels in retinal glial Müller cells and improve retinal oxygenation. To summarize, corticosteroids improve vision by re-establishing the blood retinal barrier, thereby reducing exudation and lessening interstitial edema [5], [6], [7]. As a consequence, during the past 10 years, corticosteroid therapy emerged as a promising treatment for the main degenerative pathologies of the retina. However, it is necessary to found appropriate ocular delivery systems able to release in efficacious manner corticosteroids after a topical application that is the most widely accepted route for ocular delivery due to its non-invasive nature, easy administration and patient compatibility. Unfortunately, only 2–7% of drugs administered topically result bioavailable due to the innate defensive mechanisms of the eye like naso-lacrimal drainage, absorption into the systemic circulation, very low capacity of human cul-de-sac, winking, basal and reflex tearing, tear dilution and metabolism of drugs by tear enzymes.

The use of nanocarriers could be a useful strategy for topical ocular drug delivery, for their capacity to protect the encapsulated molecules while facilitating their transport to the different compartments of the eye. Furthermore, mucoadhesive polymer based nanosystems could reduce the drainage rate thus improving ocular bioavailability. Thus different ocular drug delivery systems based on nanotechnology could be promising for the treatment of chronic ocular diseases requiring frequent drug administrations [8], [9], [10], [11]. In particular, polymeric micelles are self-assembling colloidal systems obtained by amphiphilic copolymers with distinct hydrophobic and hydrophilic segments. The polymer self-assembles to form micelles in aqueous media, wherein water insoluble segments form the core and hydrophilic segments form the corona. This hydrophobic core allows an effortless encapsulation of several poorly water soluble therapeutic agents, such as corticosteroids. Moreover, polymeric micelles seem to be very promising in ocular drug delivery for many peculiarities, including their ability to act as absorption promoters which might improve drug permeability across ocular epithelia [13], [14].

Thus, the aim of this work was to produce novel polymeric micelles to increase corticosteroid water solubility and retention time on the ocular surface as well as to enhance drug absorption through the corneal layer.

In this paper, inulin, a natural occurring polysaccharide, was chosen as a starting polymer because of its biocompatibility and chemical versatility [15], [16], [17], [18], [19]. Inulin was functionalized with ethylendiamine (EDA) and retinoic acid (RA), to introduce reactive groups able also to interact with mucin and to give an appropriate hydrophobic chain, respectively. This new copolymer, named INU-EDA-RA, was then used to prepare polymeric micelles encapsulating corticosteroids (dexamethasone, triamcinolone and triamcinolone acetonide). Micelles were characterized in order to investigate their potential as effective ocular drug delivery systems administered topically. In particular, mucoadhesive properties, drug loading and drug release as well as stability were evaluated. In addition, in vitro experiments were performed on primary human and murine ocular cells, in order to evaluate their biocompatibility and to study the cellular uptake. Finally, great attention has been given in the assessment of in vitro transcorneal permeation using transwell support systems.

Section snippets

Materials

All solvents and chemicals were of analytical grade and were used without further purification. Inulin (INU), 4-bis-nitrophenyl carbonate (BNPC), Ethylendiamine (EDA), N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDCradical dotHCl), N-hydroxysuccinimide (NHS), retinoic acid (RA), Triethylamine (TEA), Dulbecco’s phosphate buffer saline (DPBS; composition: KCl 0.2 g/l; KH2PO4 0.2 g/l; NaCl 8.2 g/l; Na2HPO4 anhydrous 1.15 g/l), dexamethasone (DEX), triamcinolone (T), triamcinolone acetonide

Results and discussion

In this work, in order to prepare new effective micelles to be administered topically on the ocular surface and useful for retinopathies, inulin (INU) was chosen as a starting natural polysaccharide. It is a linear carbohydrate consisting of glucopyranose end-capped fructose units (β-1,2). It is water soluble and has a lot of reactive hydroxyl groups so that it is a good candidate to produce new derivatives with appropriate and desired characteristics. Firstly, as in a previously published

Conclusion

A novel amphiphilic derivative of inulin (INU), obtained by introducing ethylendiamine (EDA) and retinoic acid (RA) chains on polysaccharide backbone, is able to form stable micelles in aqueous media. INU-EDA-RA micelles, loaded with dexamethasone (DEX), triamcinolone (T) and triamcinolone acetonide (TA), chosen as examples of corticosteroid drugs for the treatment of degenerative pathologies of the retina, are compatible with human and murine cell lines. Their appropriate particle size (lower

Conflict of interest

Authors declare that there is no conflict of interest involving this study.

Acknowledgements

This work has been funded by MIUR by means of the National Program PON R&C 2007-2013, “Piattaforma scientifico-tecnologica mirata allo sviluppo di nuovi approcci terapeutici nel trattamento delle principali patologie degenerative della retina, REACT (REtinopathies Advanced Care Therapies)” (PON01_01434).

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