Drug Discovery Today  Volume 21, Number 6  June 2016
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`REVIEWS
`
`Given that cyclosporine (CsA) is poorly water soluble and currently marketed
`products are not well tolerated, novel approaches for safe and efficient CsA
`delivery to the eye are of great interest.
`
`ReviewsKEYNOTEREVIEW
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`Modern approaches to the ocular
`delivery of cyclosporine A
`
`Priyanka Agarwal and Ilva D. Rupenthal
`
`Buchanan Ocular Therapeutics Unit, Department of Ophthalmology, New Zealand National Eye Centre, Faculty of
`Medical and Health Sciences, University of Auckland, New Zealand
`
`Cyclosporine A (CsA) has long been the mainstay treatment for dry eye
`syndrome (DES), one of the most common disorders of the eye. However,
`the poor water solubility of CsA renders it difficult to formulate it into
`
`topical ocular dosage forms. Restasis1 is currently the only US Food and
`Drug Administration (FDA)-approved CsA formulation, while Ikervis1
` has
`recently been launched in Europe, with both commonly associated with
`severe ocular discomfort. Therefore, several CsA formulations have been
`investigated with the aim to improve bioavailability while reducing
`adverse effects associated with the marketed formulations. In this review,
`we summarize recent advances in ocular CsA delivery that provide safer
`and more effective alternatives for the management of DES and other
`ocular inflammatory conditions.
`
`Introduction
`CsA is a metabolite of the fungi Tolypocladium inflatum and Beauveria nevus that was initially
`suggested for use as an antifungal agent. Its immunosuppressive activity soon became evident
`and, because of the reduced incidence of associated myelotoxicty, CsA eventually became the
`mainstay treatment after organ transplantation [1,2]. Systemic CsA is still used, although to a
`lesser extent, to treat several other autoimmune diseases, including those with eye involvement
`[3]. CsA is the preferred immunomodulatory agent for topical treatment of several immune-
`mediated ocular surface disorders [4], and its prevalence in the treatment of these ocular disorders
`is second only to corticosteroids, whose adverse effects are well documented [5]. CsA therapy has
`been approved for the treatment of keratoconjunctivitis sicca (KCS), more commonly known as
`dry eye syndrome (DES). It is also frequently used off-label to treat several other ophthalmic
`conditions, such as posterior blepharitis [6,7], ocular rosacea [8,9], vernal keratoconjunctivitis
`[10–14], atopic keratoconjunctivitis [15–17], acute corneal graft rejection [18], and conjunctival
`graft versus host disease [19–21].
`DES is one of the most prevalent ocular surface disorders, and is usually characterized by
`increased evaporation or decreased production of tear fluid, resulting in damage to the inter-
`palpebral ocular surface and moderate to severe discomfort [22]. Symptoms of DES have been
`reported in approximately one out of seven individuals above the age of 48, with its prevalence
`nearly doubling after 59 years of age [23–25]. A recent study estimated that nearly 20% of
`
`Priyanka Agarwal is
`currently pursuing a
`doctorate within the
`Buchanan Ocular
`Therapeutics Unit,
`Department of
`Ophthalmology, University
`of Auckland, and is
`investigating novel
`cyclosporine A (CsA) formulations for topical
`administration. She completed her BPharm at the
`University of Mumbai, India, and subsequently
`obtained a Postgraduate Diploma in Health Sciences
`from the School of Pharmacy at the University of
`Auckland. Priyanka has extensive industrial research
`experience in formulation development and
`pharmacokinetics and worked as a Veterinary
`Formulation Development Scientist for several years.
`During her time in industry, she primarily worked on
`the development of new platforms for safe drug
`delivery to animals and she is currently one of the
`inventors on a series of patents filed in this field.
`
`Ilva Rupenthal is a senior
`lecturer in the Department
`of Ophthalmology, New
`Zealand National Eye
`Center, University of
`Auckland, and the
`inaugural Director of the
`Buchanan Ocular
`Therapeutics Unit, which
`aims to translate ocular therapeutic-related scientific
`research into the clinical setting, whether
`pharmaceutical, cell, or technology based. Her
`current research, funded by a Sir Charles Hercus
`Health Research Fellowship from the New Zealand
`Health Research Council, focuses mainly on the
`development of stimuli-response devices, with
`projects investigating ocular implants responsive to
`light or a small electrical current. Moreover, Dr
`Rupenthal’s group is developing tailored controlled
`delivery systems that specifically target the drug to the
`site of action with projects around dry eye, optic
`neuropathy, diabetic retinopathy, and age-related
`macular degeneration treatment.
`
`Corresponding author: Rupenthal, I.D.
`
`(i.rupenthal@auckland.ac.nz)
`
`1359-6446/ß 2016 Elsevier Ltd. All rights reserved.
`
`http://dx.doi.org/10.1016/j.drudis.2016.04.002
`
`www.drugdiscoverytoday.com 977
`
`1
`
`ALL 2011
`MYLAN PHARMACEUTICALS V. ALLERGAN
`IPR2016-01128
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`

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`REVIEWS
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`Drug Discovery Today  Volume 21, Number 6  June 2016
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`N
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`O
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`HN
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`NH
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`O
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`O
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`O
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`N
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`O
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`HN
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`O
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`N
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`O
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`N
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`N
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`Drug Discovery Today
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`FIGURE 1
`Molecular structure of cyclosporine A (CsA), a cyclic undecapeptide with very
`low aqueous solubility.
`
`neutrally charged and hydrophobic molecule (Fig. 1) with an
`aqueous solubility of less than 10 mg/ml (0.001%) at physiological
`temperature [42], formulation of aqueous eye drops at these con-
`centrations is difficult. Attempts have been made to improve the
`aqueous solubility of CsA using surfactants and/or penetration
`enhancers, such as macrogolglycerol ricinoleate (Cremophor
`EL1) and benzalkonium chloride, with the latter also commonly
`used as a preservative in ocular formulations [43]. Although these
`excipients can improve the solubility and penetration of CsA, their
`use is limited by their high irritation potential because these
`molecules typically function by compromising the integrity of
`the ocular tissues [44–46].
`CsA solutions in vegetable oils were considered as the next best
`alternative for topical administration to the eye and concentra-
`tions as high as 2% could be achieved [47,48]. Despite their poor
`absorption, oily CsA solutions have demonstrated success in the
`treatment of DES by improving tear production and inducing
`in a canine model
`regression of corneal neovascularization
`[49–54]. Similar results were also observed in humans after topical
`application of 2% CsA in olive oil [55,56]. However, one of the
`major limitations of vegetable oils in ophthalmic preparations is
`the increased incidence of ocular toxicity after frequent use, with
`blurring of vision also commonly observed because of the high
`viscosity of the carrier oils. BenEzra et al. [38,57,58] showed that
`penetration of CsA from olive oil drops was negligible in normal or
`slightly inflamed eyes, but increased significantly over time be-
`cause the corneal barrier was compromised as a result of toxic
`effects of the oily vehicle. Thus, the use of topical CsA oily drops
`for the management of ophthalmic conditions has largely been
`discontinued.
`CsA ointments [59–61] and oil-in-water (o/w) emulsions [62]
`have also been used to reduce the toxicity associated with oily
`solutions, although they have not been able to eliminate it
`completely. Currently, Restasis1
` (Allergan) is the only CsA for-
`mulation approved by the FDA for DES therapy in humans. It is a
`0.05% CsA emulsion of castor oil in water and is often associated
`with severe adverse effects, such as ocular burning (most com-
`mon), conjunctival hyperemia, discharge, epiphora, eye pain,
`
`hospitalized patients above the age of 50 had DES, with old age and
`illiteracy being major predictors of the disorder [26]. On the
`in
`recommendation of the International Dry Eye Workshop
`2007, DES was classified as a multifactorial disease of the tears
`and ocular surface that can be triggered by a variety of underlying
`causes. However, recently, there has been increasing evidence that
`inflammation has a key role in the manifestation of DES. Ocular
`surface abnormalities, such as the appearance of inflammatory cell
`intermediates in the lacrimal gland and an increase in immune-
`related antigens and cytokines at the conjunctival epithelium,
`are commonly demonstrated by both autoimmune (Sjo¨gren’s
`(non-Sjo¨gren’s syndrome)-
`syndrome) and non-autoimmune
`mediated DES [27–30], making treatment of the underlying cyto-
`inflammatory processes
`the primary
`kine–receptor-mediated
`‘causative therapeutic approach’ [31,32]. A combination of symp-
`tomatic therapy, which includes modification of the ocular envi-
`ronment (by increasing humidity, occlusion of lacrimal canaliculi,
`or simulation of tears), and pathogenic treatments, including the
`use of antibacterial and anti-inflammatory agents (corticosteroids,
`antihistamines, tetracyclines, and CsA), is currently recommended
`for DES therapy [32].
`CsA is the anti-inflammatory agent of choice for the treatment
`of DES, because it can be used long term without adverse effects
`commonly associated with other anti-inflammatory agents, such
`as steroids [5]. Furthermore, unlike corticosteroids, the activity of
`CsA results from specific and reversible action on T cells, making it
`safe for prolonged use. For example, corneal epitheliopathy and
`eyelid maceration associated with long-term CsA therapy were
`found to be reversible with complete cessation on discontinuation
`of the drug [33]. The immunomodulatory activity of CsA helps in
`reducing the inflammation associated with subconjunctival and
`lacrimal glands, resulting in increased goblet cell density and tear
`production [34,35]. CsA tends to bind with specific nuclear pro-
`teins that initiate the activation of T cells, thus preventing T cell
`production of inflammatory cytokines and disrupting the im-
`mune-mediated inflammatory response [31,36]. CsA is a hydro-
`phobic molecule and, therefore, is difficult to formulate into
`conventional topical ocular delivery systems. Significant research
`has been performed over recent years to develop safe and effective
`ocular delivery systems for CsA. In this review, we highlight recent
`efforts to improve the ophthalmic delivery of CsA in terms of its
`bioavailability and ocular tolerability while reducing adverse
`effects.
`
`Conventional CsA formulations
`Systemic CsA is generally not considered for the treatment of
`ocular pathologies because of severe systemic adverse effects, such
`as nephrotoxicity and hypertension [2,37], although significant
`concentrations of CsA have been reported in tears and lacrimal
`glands after oral administration [38]. Thus, the topical route is
`generally preferred because, as well as reducing systemic adverse
`effects, it also helps to achieve improved bioavailability and spe-
`cific targeting to the ocular tissues [39,40].
`Dose-ranging randomized clinical trials have shown that topi-
`cally applied CsA is effective at concentrations between 0.05 and
`0.1% (w/v). No additional benefits were observed at higher con-
`centrations; hence, clinical trials are generally recommended at a
`maximum concentration of 0.1% [4,41]. However, because CsA is a
`
`978 www.drugdiscoverytoday.com
`
`Reviews  KEYNOTE REVIEW
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`Drug Discovery Today  Volume 21, Number 6  June 2016
`
`REVIEWS
`
`*
`
`*
`
`120
`
`100
`
`20
`
`60
`
`40
`
`20
`
`*
`
`Cornea
`
`Conjunctiva
`
`0
`
`Aqueous
`humor
`
`CsA in oil 0.50% formulation
`Prodrug in water 0.50% formulation
`
`ReviewsKEYNOTEREVIEW
`
`Iris-Ciliary
`body
`
`
`*
`
`*
`
`Vitreous
`humor
`
`Retina
`
`Converted CsA
`Unconverted CsA
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`Drug Discovery Today
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`8000
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`7000
`
`6000
`
`5000
`
`4000
`
`3000
`
`2000
`
`1000
`
`0
`
`
`
`Total CsA (ng/g)
`
`FIGURE 2
`Ocular distribution of cyclosporine A (CsA) in oil and CsA prodrug in water.
`Prodrug formulations significantly increased the amount of CsA absorbed by,
`and accumulated in, the cornea and conjunctiva; however, no improvement
`was observed in the penetration of CsA across the cornea.
`Source: Adapted from [75].
`
`a significant
`in DES
`in basal tear production
`improvement
`[71,72]. Aqueous prodrug solutions have also been evaluated for
`their efficacy in the treatment of corneal graft rejection and it was
`found that prodrug eye drops applied five times a day were
`therapeutically equivalent to a 10 mg/kg/day intramuscular injec-
`tion in rats [73]. Recent studies have shown that 2% prodrug
`solutions have a 200–500-fold higher conjunctival permeability
`than the conventional 2% CsA in oil formulation. Accumulation
`of CsA prodrug formulations in the cornea to form large tissue
`deposits that provide a sustained release effect over prolonged
`periods of time was also observed. However, prodrug formulations
`did not show much improvement in the permeability across the
`cornea and into the aqueous humor compared with the conven-
`tional CsA emulsion, probably because of the rapid conversion of
`the prodrug into CsA at the corneal surface (Fig. 2). This depot
`formation could have the added advantage of overall poor system-
`ic absorption of prodrug formulations, reducing the incidence of
`systemic complications and immunosuppression. Prodrug formu-
`lations with CsA concentrations higher than 2% are currently
`under investigation for safety and toxicity [74,75].
`
`Novel application of excipients
`Several new excipients have been introduced to improve CsA
`solubility in ocular formulations and enhance their bioavailability
`(Table 1).
`Cyclodextrins
`The introduction of cyclodextrins (CDs) in ophthalmic drug de-
`livery is relatively recent; however, they have rapidly gained
`popularity because of their pronounced benefits in drug solubili-
`zation and stabilization. CDs are cyclic oligosaccharides of six
`(a-CD), seven (b-CD), or eight (g-CD) glucopyranose units with
`a hydrophobic cavity in the center. In aqueous solutions, water
`from the hydrophobic cavity is displaced by hydrophobic mole-
`cules, resulting in the formation of large water-soluble complexes.
`CDs have been used on several occasions to prepare aqueous CsA
`
`www.drugdiscoverytoday.com 979
`
`foreign body sensation, pruritus, stinging, and visual disturbance
`[63]. Devecı et al. [64] recently showed that ocular irritation
`associated with Restasis usually decreases within 1 week of con-
`tinuous therapy and significant resolution of DES can be observed
`at the 1-month follow-up. A veterinary ointment of 0.2% CsA
`(Optimmune1, Schering-Plough Animal Health) has also been
`approved for DES therapy in dogs and, although it has been shown
`to resolve the symptoms of DES, associated ocular toxicity, proba-
`bly because of the oily base, is significant [65,66]. Ointments are
`generally more viscous and have a cloudy appearance and, thus,
`tend to blur the vision, which, in addition to their tolerability
`issues, reduces patient acceptability. Recently, a cationic nanoe-
`mulsion containing 0.1% CsA was launched in Europe for the
`treatment of severe DES under the brand name Ikervis1
` (Santen).
`Unlike emulsions and ointments, this system does not cloud
`vision because of its low viscosity; however, adverse effects, such
`as stinging and pain, have frequently been reported [67].
`The limitations of currently approved formulations leave tre-
`mendous scope for the development of improved ocular formula-
`tions for the safe and efficient delivery of CsA to the eye. The focus
`of new CsA technologies has predominantly been the improve-
`ment of solubility, transcorneal penetration, and precorneal resi-
`dence time, with a simultaneous reduction in the frequency of
`dosing and irritation potential. Such CsA formulations could
`significantly improve patient comfort and compliance, thus in-
`creasing the quality of life of patients with DES.
`
`Recent approaches in CsA delivery
`Over the past few years, several novel strategies have been sug-
`gested for delivering CsA to the ocular tissues, with topical,
`episcleral, subconjunctival, and intravitreal routes being investi-
`gated. Of these, topical administration remains the most preferred
`route for the treatment of DES, because it is non-invasive, painless,
`and convenient. For the purpose of this review, the key approaches
`to improving ocular drug delivery of CsA have been classified into
`three major areas of research: (i) chemical modification of the
`drug; (ii) novel application of excipients; and (iii) novel ophthal-
`mic dosage forms.
`
`Chemical modification of the drug
`Reversible chemical modification of CsA to obtain prodrugs with
`improved aqueous solubility was first suggested by Hammel et al.
`[68] when they coupled CsA with diketopiperazine to synthesize
`dipeptide esters with improved oral bioavailability. Monomethox-
`y(polyethyleneglycol) derivatives of CsA have also been suggested
`for improving solubility and absorption of CsA in oral prepara-
`tions. However, esterification of the free hydroxyl group of CsA
`with a solubilizing moiety is the generally accepted approach for
`prodrug synthesis for ocular applications. Lallemand et al. [69,70]
`developed a series of amphiphilic acidic prodrug molecules (pat-
`ented by DeBiopharm, Switzerland) having an approximately
`25 000 times higher solubility than CsA in isotonic phosphate
`buffer solution (PBS) at pH 7. These prodrugs are quantitatively
`hydrolyzed in artificial tears to release CsA within 1 min. Prodrug
`conversion into the parent molecule was significantly faster in tear
`fluid than in a buffer at physiological pH, indicating that the
`hydrolysis is enzyme mediated. Aqueous formulations of these
`esterified CsA prodrugs were well tolerated and have shown
`
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`Drug Discovery Today  Volume 21, Number 6  June 2016
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`TABLE 1
`Novel excipients used in ocular CsA delivery systems.
`
`Excipients
`
`Dosage form
`
`Advantages
`
`Limitations
`
`Cyclodextrins
`
`Aqueous eye drops
`
`Solubilize CsA in aqueous formulations; show
`improved bioavailability and low toxicity at
`optimized concentrations
`
`Cannot be used in concentrated
`solutions; ocular toxicity can be
`observed at higher concentrations
`because of complexation of
`cellular components
`
`Refs
`
`[76–79]
`
`Semifluorinated
`alkanes
`
`Eye drops (CyclASolW)
`
`Safe and well tolerated; solubilize hydrophobic
`drugs to form clear eye drops; improved
`penetration; no preservative required;
`lubricating nature
`
`Cationic vectors
`(amines, chitosan,
`poly-L-lysine, Eudragit)
`
`Emulsions, liposomes,
`nanoparticles
`
`Improve precorneal residence time because
`of mucoadhesion
`
`No sustained release effect
`
`[80,81]
`
`Amine vectors can cause stability
`problems; safety and toxicity is a
`concern because ocular irritation
`is usually observed with most
`cationic moieties
`
`[95]
`
`Reviews  KEYNOTE REVIEW
`
`eye drops with significant improvement in transcorneal penetra-
`tion and retention [76–79]. a-CDs have been found to be the most
`efficient in solubilizing CsA, because the cavity size is a good fit for
`the cyclic CsA molecule. When used to prepare concentrations
`below 0.025%, a-CDs improved CsA bioavailability without dem-
`onstrating any toxic effects in vivo [77,79].
`Semi-fluorinated alkanes (SFAs)
`SFAs are a class of amphiphilic fluorinated compounds that are
`physically, chemically, and biologically inert and capable of dis-
`solving several hydrophobic drugs. Although they have been used
`intraocularly as intravitreal tamponades for several years, their use
`in drug delivery to the front of the eye is relatively recent [80].
`Given their established safety profile, excellent spreading proper-
`ties, and ability to solubilize and stabilize several hydrophobic
`drugs, SFAs can be used as suitable carriers for ocular preparations.
`The added advantages of SFAs include their ability to form clear
`solutions that do not cause any blurring of vision or ocular
`discomfort while also providing lubrication to the ocular surface,
`which is particularly useful in DES treatment. Moreover, because
`these solutions are nonaqueous, they do not require any preser-
`vatives, surfactants, or pH modifiers, which are frequently impli-
`cated in ocular toxicity. Recent studies showed that SFA-based
`solutions containing 0.05% CsA were well tolerated and signifi-
`cantly increased CsA penetration across the cornea and into the
`aqueous humor when compared with Restasis, rendering SFA-
`based formulations promising candidates for the treatment of
`ocular inflammatory conditions [81]. This technology is currently
`registered as CyclASol1
` (Novaliq GmbH, Germany) and recently
`conducted Phase I clinical trials have shown promising results
`[82], with the company now recruiting patients for a Phase II
`clinical trial [83].
`Positively charged vectors
`One of the major objectives of topical CsA delivery has been to
`improve ocular bioavailability of the drug by increasing corneal
`penetration and precorneal residence time. Therefore, positively
`charged ocular formulations have received much interest because
`of their higher mucoadhesion and, thus, longer precorneal resi-
`dence time. Studies performed by Daull et al. [84] showed that
`cationic emulsions have significantly greater bioavailability than
`Restasis, which is an anionic o/w emulsion. Safety profiles of
`
`980 www.drugdiscoverytoday.com
`
`cationic and anionic emulsions were further compared and, while
`their tolerability was similar, only the cationic emulsion was able to
`maintain the normal healing rate of the human corneal epithelium
`in vitro and reduce inflammation in vivo [85]. Similar to most epithe-
`lia, corneal and conjunctival cells are negatively charged at physio-
`logical pH and, therefore, cations can adhere to and penetrate them
`more easily [86,87]. Their cell membranes are further coated with a
`layer of mucin containing negatively charged sialic acid groups,
`which develop electrostatic interactions with positively charged
`vectors and thus improve the precorneal residence time [88].
`Stearylamine has been used extensively to impart a positive
`charge to liposomes [89–91] and ocular emulsions [92–94] to
`increase mucoadhesion and, thus, retention. Oleylamine is anoth-
`er cationic lipid that has been used for preparation of cationic
`ophthalmic emulsions [88]. However, a major disadvantage of
`these amines is their poor stability and compatibility with other
`excipients, while ocular tolerability of amines remains a signifi-
`cant concern. Hence, chitosan has emerged as the cationic agent of
`choice in ocular formulations. Chitosan is a biodegradable and
`linear polysaccharide with
`several positively
`biocompatible
`charged free amino groups that interact with mucin. There is
`some evidence that, in addition to the positive charge, the specific
`nature of chitosan might also be responsible for improving the
`uptake of drug molecules. Calvo et al. [95] showed that the
`bioavailability of nanocapsules coated with chitosan was at least
`twofold higher than for poly-L-lysine-coated nanocapsules. How-
`ever, commercialization of chitosan formulations might be diffi-
`cult, because raw materials of natural origin often demonstrate
`high batch-to-batch variability. Recently, several studies also
`reported that, despite its biodegradability, chitosan can display
`some toxicity, especially when administered as nanoparticles
`[96,97], while chitosan hydrogels of a certain molecular weight
`have also shown to initiate an inflammatory response and, thus,
`delay wound closure in a corneal scrape wound model [98]. There-
`fore, Eudragit1, a synthetic polymer frequently used for the prepa-
`ration of enteric oral dosage forms, has been suggested as an
`alternative for ophthalmic formulations [99]. Eudragit is a cationic
`copolymer based on dimethylaminoethyl methacrylate, butyl
`methacrylate, and methyl methacrylate groups. Eudragit-based
`ocular colloidal formulations have shown good tolerability and
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`TABLE 2
`Novel ophthalmic dosage forms for CsA delivery to the front of the eye.
`
`Drug delivery system
`
`Advantages
`
`Limitations
`
`REVIEWS
`
`Refs
`
`[103–106]
`
`Improved uptake into all layers of the cornea;
`sustained release over extended periods
`
`Generally poor shelf life; irritation potential and
`toxicity can be significant
`
`ReviewsKEYNOTEREVIEW
`
`Form depots in cornea for sustained drug
`release over prolonged periods
`Improved stability and simple manufacturing
`procedure
`
`Short half-life at the corneal surface results in
`poor transcorneal penetration
`Safety and irritation potential need to be assessed
`
`[113,114,118]
`
`[119]
`
`Improved retention and uptake into cornea;
`reduced drug toxicity can be observed
`Improved uptake because of transport by
`transcellular pathway
`Improved bioavailability; improved shelf-life
`compared with nanoemulsions
`
`Difficult to scale up; can show some long-term
`toxicity
`Generally poor stability and manufacturability
`
`Generally poor long-term stability and
`manufacturability
`
`[133,134]
`
`[142]
`
`[143–145]
`
`Increased precorneal residence time and
`sustained release
`Sustained release of the drug over prolonged
`periods
`
`High burst release; blurring of vision can reduce
`patient compliance
`Discomfort and foreign body sensation; adverse
`effects because of poor oxygen permeability
`
`[154,157,160]
`
`[164,165,171]
`
`Micelles
`
`Liposomes
`Liposomes
`
`Proliposomes
`
`Nanoparticles
`Nanospheres/nanocapsules
`
`Nanoemulsions
`
`Solid lipid nanoparticles
`
`Other
`In situ gelling systems
`
`Hydrogels
`
`low incidence of ocular irritation [100–102], while decomposition
`of Eudragit is typically slow, providing controlled drug release with a
`low burst effect.
`
`Novel ophthalmic dosage forms
`The field of ocular therapeutics has seen the development of
`several novel delivery systems in the form of micelles, liposomes,
`nanoparticles, in situ gelling systems, and hydrogels, which have
`also been evaluated for the delivery of CsA (Table 2 and Fig. 3).
`
`Micelles
`Micelles are self-assembling spherical colloidal systems that are
`frequently used for the solubilization of hydrophobic molecules.
`Typically, micelles are formed around a hydrophobic drug in
`aqueous solution because of the orientation of surfactant mole-
`cules to form a hydrophobic core enclosing the drug within a
`hydrophilic shell. N-Octyl chitosan has been used as a surfactant
`for preparation of CsA micelles for DES therapy [103]. CsA uptake
`from these micelles was found to be significantly higher than from
`
`In situ gelling systems
`
`+ Improved retention
`+ Sustained release
`
`- Blurring
`- Burst release
`
`Hydrogels
`+ Improved retention
`+ Sustained release
`
`- Patient discomfort
`- Poor oxygen permeability
`
`Micelles
`+ Improved absorption
`+ Deeper penetration
`
`- Surface toxicity
`- Poor stability
`
`Liposomes
`+ Corneal accumulation
`+ Sustained release
`
`- Poor scalability
`- Low entrapment
`
`Nanoparticles
`+ Improved penetration
`+ Improved absorption
`
`- Poor scalability
`- Poor stability
`
`Drug Discovery Today
`
`FIGURE 3
`Schematic representation of novel ophthalmic dosage forms used to improve the bioavailability of topically applied cyclosporine A (CsA).
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`Drug Discovery Today  Volume 21, Number 6  June 2016
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`from a saturable interaction with mucin [116] and, thus, concen-
`trations could be optimized to achieve maximal mucoadhesion
`with minimal toxicity. Li et al. [117] prepared liposomes coated
`with low-molecular-weight chitosan and observed a significant
`improvement in the ocular bioavailability of CsA. The number of
`free positively charged groups available in low-molecular-weight
`chitosan is considerably reduced, resulting in better ocular tolera-
`bility of these formulations.
`Despite their many benefits, the fast degradation on the corneal
`surface, poor entrapment efficacy, and difficulty in manufacturing
`sterile formulations at a large scale are disadvantageous to the use
`of liposomal formulations. To counter these problems, Karn et al.
`[118] developed CsA-loaded multilamellar liposomes from egg
`lecithin and phosphatidyl choline from soybean using supercriti-
`cal fluid of carbon dioxide as the antisolvent. Liposomes, thus
`prepared, showed better stability on the corneal surface, had an
`improved shelf life, and were easy to manufacture on a larger scale.
`Considerable improvements were also seen in the precorneal
`residence time, corneal absorption, and tolerance in comparison
`to Restasis [112]. Recently, the same method was also used to
`prepare proliposomes [119], which are dry free-flowing powders
`that assemble spontaneously in an aqueous medium to form
`liposomes. These proliposomes had excellent stability and were
`easy to scale up. However, some additional studies might be
`required to establish their efficacy in ocular CsA delivery.
`Nanoparticles
`A large body of research has also been dedicated to the develop-
`ment of nanoparticle-based systems for targeted CsA delivery to
`the eye. Nanoparticles are submicron particles ranging from 10 to
`1000 nm in size that can have drug molecules dissolved, adsorbed,
`or entrapped in the nanoparticle matrix. Nanoparticles have been
`shown to accelerate drug penetration and significantly improve
`corneal absorption of drugs, primarily by the transcellular path-
`way [120–123]. They have also been reported to decrease the
`incidence of ocular irritation and toxicity because of their small
`particle size [124,125]. Moreover, because they are similar to
`aqueous eye drops in appearance and viscosity, nanoparticle for-
`mulations are easy to handle and ensure convenient ocular appli-
`cation [126–128]. However, despite their advantages, products
`based on nanotechnology rarely reach the market [129] as a result
`of technical issues involved in scale up and manufacturing, use of
`new excipients or nonpharmacopoeial solvents [130], as well as
`the relatively poor stability of these colloidal systems [131].
`CsA nanosuspensions have been prepared using zirconia beads
`in a water/polyvinyl alcohol system and were found to show
`significantly lower irritation compared with Restasis [132]. How-
`ever, biodegradable polymers, such as poly-D,L-lactide-co-glycolide
`(PLGA) and poly-e-caprolactone-co-lactide (PCL), are usually pre-
`ferred for ophthalmic applications because of their established
`safety profile. PLGA nanoparticles prepared by solvent evaporation
`have shown a 11–16% drug-loading capacity and have been used
`for sustained release of CsA for up to 65 days [133]. It was found
`that coating these nanoparticles with a surface-modifying poly-
`mer, such as PEG, could influence the penetration behavior of CsA
`across the cornea. PCL nanoparticles have also been used for ocular
`CsA delivery in conjugation with penetration-enhancing surfac-
`tants, such as benzalkonium chloride [134]. Although the bio-
`availability from these nanospheres was 10–15 times higher than
`
`conventional CsA oil drops, and the bioavailability could be
`further increased by coating the micelles with carbopol to give
`them mucoadhesive properties. Penetration of micellar formula-
`tions can also be enhanced by reducing the size of the micelles. In a
`recent study using an optimized blend of hydrogenated castor oil-
`40 and octoxynol-40, Cholkar et al. [104] prepared a clear aqueous
`nanomicellar solution containing 0.025–0.1% CsA with an aver-
`age particle size of 22.4 nm. High corneal and conjunctival levels
`were observed with low ocular toxicity. One of the most significant
`findings of this study was that this formulation could also deliver
`CsA to the posterior segment of the eye, with as much as 53.7 ng/g
`of CsA recovered from the retina/choroid in vivo. Polyvinyl capro-
`lactam–polyvinyl acetate–polyethylene glycol nanomicelles have
`also shown improved penetration of CsA along with excellent
`ocular tolerability and safety [105]. CsA uptake from these micelles
`is predicted to be an energy-dependent intracellular pathway,
`leading to improved efficacy in vivo. Recently, in vivo studies
`performed with nano-sized micellar systems based on methoxy
`poly(ethylene) glycol-hexyl substituted poly(lactide) (mPEGhex-
`PLA) copolymers showed increased uptake and prolonged release
`of CsA because of better penetration into, and accumulation in, all
`layers of the cornea [106]. Significant advantages of this mPE-
`GhexPLA nanomicellar system included the absence of ocular
`toxicity and a longer shelf life compared with conventional mi-
`celle systems [107,108]. Further improvement in shelf life by
`lyophilization of mPEGhexPLA micellar systems has also been
`suggested; however, long-term studies still need to be performed
`to verify this [109]. Poor long-term stability is the major drawback
`of most micelle systems. Therefore, in an attempt to overcome this
`limitation, CsA-loaded self-assembling micellar systems were pre-
`pared using two PEG-fatty alcohol ether-type surfactants [110].
`The micellar formulation had good entrapment efficiency and
`showed a threefold increase in corneal CsA concentration com-
`pared with Restasis. However, the toxicity and irritation potential
`of this formulation still needs to be evaluated.
`Liposomes
`Over